Brian A. Baldo Nghia H. Pham Clinical Aspects, Diagnosis, Mechanisms

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Brian A. Baldo Nghia H. Pham Drug Allergy

Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity Relationships

123 Drug Allergy

[email protected] Brian A. Baldo • Nghia H. Pham

Drug Allergy

Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity Relationships

[email protected] Brian A. Baldo Nghia H. Pham Formerly-Molecular Immunology Unit Formerly-Molecular Immunology Unit Kolling Institute of Medical Research Kolling Institute of Medical Research Royal North Shore Hospital of Sydney Royal North Shore Hospital of Sydney Sydney , Australia Sydney , Australia

ISBN 978-1-4614-7260-5 ISBN 978-1-4614-7261-2 (eBook) DOI 10.1007/978-1-4614-7261-2 Springer New York Heidelberg Dordrecht London

Library of Congress Control Number: 2013937589

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[email protected] Dedicated to my mother, Sylvia, and the memory of my father, Disma, for their care, guidance, and enduring support and whose hard work, self-reliance, and plain commonsense instructed by example.

BA B

To my father and memory of my mother with love and gratitude. To my wife, Phong-Thuy, and my daughter, Cecile, for their unconditional love, endless support, and encouragement.

NHP

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[email protected] Preface

With its roots in immunology and pharmacology, advancement in the science of drug allergy and its application to clinical medicine has ultimately always been heavily reliant on application of a broad range of investigative methodologies in humans rather than laboratory animal models. The variety of chemically diverse pharmacological agents administered to patients is large and continues to expand and with every new drug released, there is always potential for adverse reactions, some of them allergic. This diversity in chemical structure and pharmacological action together with the range of observed adverse clinical responses; the need to access suffi cient numbers of ade- quately phenotyped patients to study; the necessity of collaborative inputs from laboratory and clinic; and the variety of chemical, cellular, and clinical methodologies needed ensured that progress in the fi eld has generally fallen short of the hoped-for insights. In fact, post the penicillin era when drug allergy was given a much-needed structural perspective, at both the research level and in terms of patient benefi ts this specialized section of allergic diseases could not be said to have made great advances. There are a number of reasons for this. In the fi rst place, the seemingly perpetual confusion over what constitutes an allergic reaction is something that affects many clinicians as well as the public and the mass media. The term “allergy” continues to be used inappropriately to refer to all sorts of nonimmune-based reactions to a drug and, for many in the medical profession, this refl ects a state of mind that is not conducive to reporting/recording, diagnosing, and seeking to understand the true nature of many drug-induced reactions. To reinforce this point, attention is drawn to the 1970s and 1980s when the value of skin testing— prick, intradermal, and patch—although widely advocated and promoted by a few practitioners and afi cionados, was not widely understood, appreciated, and applied. Examples were the neglect of the skin test in the diagnosis of drug allergies to anesthetic agents, drugs used in surgery, antibiotics, and other antimicrobial drugs where attitudes to the test sometimes ranged from the uninformed to cynicism of its scientifi c and clinical relevance. One only needs to talk to anesthetists from that period who were aware of anaphylaxis to neuromuscular blocking drugs to learn of the skepticism of skin testing by many of their professional colleagues. To help overcome this problem, research fi ndings and leadership and instruction, for example, in the form of issued practice parameters and standard operating procedures by the relevant professional bodies, were needed. In anesthesia, this is, in fact, now being

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[email protected] viii Preface undertaken by Societe Francaises d’Anesthesie et de Reanimation, the Association of Anaesthetists of Great Britain and Ireland, the British Society of Allergy and Clinical Immunology, and the Scandinavian Society of Anaesthesiology and Intensive Care Medicine, all of whom have issued clinical practice guidelines. Over the whole broad area of drug allergy, the European Network for Drug Allergy under the aegis of the European Academy of Allergology and Clinical Immunology has published numerous position papers on allergy practice over the last decade with emphasis, for example, on standardization of methods for the diagnosis of drug allergies. While acknowledging the sometimes under-appreciation of the problem of drug allergy and the inadequacy of its diagnosis, a third inhibitory factor to progress was probably inevitable. This relates to research directed at identifying underlying mechanisms and improving patient outcomes for cell-mediated drug-induced hypersensitivities such as the various cutaneous reactions ranging from mild exanthemas to severe bullous eruptions. Knowledge of the intricate cellular immune processes involved in antigen recognition, lymphocyte receptor repertoires, and the adaptive immune response as well as recognition of the value and application of a pharmacogenetic approach needed to progress to somewhere near their present levels of understanding before signifi cant inroads could begin to occur. In particular, understanding the role of MHC restriction, drug-specifi c T cells, and the availability of improved genotyping technologies should signifi cantly increase the chances of advancing knowledge of the cellular hypersensitivity mechanisms and developing new diagnostic and predictive tests. As advantage is increasingly being taken of the results obtained from the extraordinary investigative activity directed at defi ning cellular and molecular mechanisms of immune processes, chemical approaches, used so effectively in the studies on penicillin and neuromuscular blocking drugs, are being less often utilized as biological and clinical emphases dominate research efforts. The results of this neglect can be seen in the dearth of detail available on the structures recognized by the cellular immune system in delayed hypersensitivity responses. With increasing employment of mass spectrometric characterizations, carefully selected synthetic drug conjugates, and the realization that drugs may be recognized or participate in immune processes in their free state, we can expect that this situation may soon be remedied as investigators seek to expand their current cellular preoccupation, much of it often speculative in nature, with a deeper understanding of the fi ne structural features that determine allergenic recognition in cell-mediated drug reactions. With this background and perspective in mind, we set out here to identify the most important culprit drugs implicated in immediate and delayed drug hypersensitivities and to collate up-to-date information on classifi cations, clinical features, diagnoses, underlying mechanisms, and structure–activity relationships. Chapters dealing with the molecular and cellular mechanisms of drug hypersensitivities, nonimmune-mediated sensitivities, and diagnostic methods are presented as introductory material for in-depth treatises on the β-lactam antibiotics, other antibiotics and antimicrobials, drugs used in anesthesia and surgery, opioid analgesics, corticosteroids, monoclonal antibodies and other biologics, drugs used in chemotherapy, proton pump inhibitors,

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iodinated and gadolinium-based contrast media, and nonsteroidal anti- infl ammatory drugs. For the latter two groups of drugs where only some of the adverse reactions are truly allergic in nature, discussions have been extended to cover the more dominant and more often seen drug-induced sensitivities or intolerances. Readers with a historical perspective may be able to detect in this book the infl uence of two past investigators who made important contributions to hypersensitivity research. Each had widely different professional training, research backgrounds, and clinical involvement, but both were well known for their infectious, unrelenting enthusiasm and the pleasure they derived from pursuing, over many years, original ideas and observations that were very much their own. Time spent by the author in the 1970s in both laboratories left a career-long imprint. In so many ways, the diffi cult Elvin Kabat in the Columbia University College of Physicians and Surgeons, New York Presbyterian Hospital, and the urbane Jack Pepys at the Brompton Hospital, London, could not have been more different but both were undoubtedly exceptional investigators, one in the laboratory relentlessly applying his quantitative approaches and the other in the world of patients, exploiting the diagnostic potential of, and promoting, one of the simplest technical procedures ever employed in clinical work. The quantitative immunochemical methodologies introduced and developed by the Landsteiner-Heidelberger school of immunochemistry and so expertly applied and propagated by Kabat in his classic text Kabat and Mayer ’ s Experimental Immunochemistry (C . C . Thomas , Springfi eld , Il ) infl uenced a generation of immunologists and maintained a direct line back to Landsteiner and the origins of immunochemistry. By the early 1950s in studies backed by the Offi ce of the Surgeon General, U.S. Army, Kabat had demonstrated a relationship between dextran structures and molecular weight and the propensity of the polysaccharides to provoke systemic allergic reactions. This work ultimately led to a dramatic 90-fold reduction in dextran-induced anaphylactic reactions by pre-dosing with a dextran monovalent hapten. Application of this competitive hapten inhibition strategy, straight out of the Landsteiner–Heidelberger–Kabat quantitative immunochemical protocols, made dextrans easily the safest of all the plasma volume expanders in use. Likewise, Pepys’ championing and application of the specifi city, sensitivity, and wide applicability of skin prick and provocation testing, despite their apparent simplicity, aided understanding of some important fungal-induced hypersensitivity diseases of the chest, increased appreciation of the clinical value of the procedures, and emphasized their utility for research, diagnosis, and studies of mechanisms in clinical immunology and allergology. Together with his original contributions over many years in the fi eld of occupational allergic diseases studying hypersensitivity pneumonitis (extrinsic allergic alveolitis), his early contributions to our understanding of the late reaction and the training of a constant stream of visiting clinicians from all over the world, Pepys was also fascinated by what often appeared to be hypersensitive responses to “small” molecules including drugs and in his later years he began studies in this area. This was after his earlier pioneering investigations into the sensitizing and allergenic properties of platinum in refi nery workers. This work, including the detection

[email protected] x Preface of IgE antibodies to platinum salts, was to prove a forerunner of later interest in patient reactions to the important and heavily used platinum chemotherapeutic drugs. The legacies of Elvin Kabat and Jack Pepys remain apparent today in the originality of their scientifi c research and value of their clinical contributions. To that can be added the many practitioners in laboratories and clinics who pass on what they themselves learned from the enthusiastic tute- lage of these too- often forgotten important early contributors to our knowledge of hypersensitivity states. In pursuing the authors’ own interests and research in drug allergies, some of it recounted in this volume, we would like to acknowledge our enduring collaboration with Dr Malcolm M. Fisher who introduced one of us to the then mechanistically poorly understood problem of perioperative anaphylaxis to what, at the time, were called muscle relaxants. The long-standing clinical interest by Dr Fisher provided all the necessary clinical background and patient material for successful investigations of underlying mechanisms, led on to the study of a range of other drug allergies, and ultimately the development of a useful battery of routine in vitro drug allergy tests. In what was a remarkably small manpower input over many years, we are indebted to Gail Knowland in particular for her long-standing, versatile, and always reliable input into all of the projects, to Dr David Harle for his sustained careful investigations and technical expertise, and, in later years, to Dr. Zhenjun Zhao who, like all his fellow investigators, assiduously pursued the laboratory’s quantitative approaches to mechanistic and diagnostic studies on a wide range of poorly understood adverse drug reactions. Dr. John Redmond, Dr. Mary Smal, Dr. Sue Cooney, and Dr. Alistair McCaskill had key roles in the laboratory’s research on PAF mentioned here and the development of a sensitive, high- throughput immunoassay for the mediator. The inclusion in this book of some important photographs and fi gures was greatly assisted by the generosity and cooperation of Professor S. R. Durham, Dr. D. G. Ebo, Dr. J. S. Fok, Dr. D. Gin, Dr. F. Hasdenteufel, Professor R. J. Heddle, Dr. A. Mar, Dr. P. A. J. Russo, Dr. R. Spiewak, Dr. F. C. K. Thien, and Dr. S. Van Nunen. Our intention has been to provide a scientifi cally based textbook with the relevant chemical, immunological, pharmacological, biochemical, and, where appropriate, pharmacogenomic information without losing the clinical perspective that is, in any case, the stimulus and the need for studying drug allergies in the fi rst place. In addition to clinicians, other healthcare professionals, and researchers, the book has been aimed at undergraduate and graduate courses in the biomedical sciences and to serve as a text for students of medicine, pharmacy, nursing, and dentistry. Finally, as with any subject still beset by many questions, alternative interpretations and different priorities, some analyses, arguments, or conclusions expressed here may not fi nd universal acceptance. In such cases, we remain open and ready to consider all comments in an ongoing effort to improve the book and correct any errors.

Sydney, Australia Brian A. Baldo Nghia H. Pham

[email protected] Contents

1 Adverse Reactions to Drugs and Drug Allergy: Scope of This Book ...... 1 1.1 Adverse Drug Reactions ...... 1 1.1.1 Defi nition ...... 1 1.1.2 Terminology: Classifi cation of Adverse Drug Reactions and the terms Hypersensitivity, and Allergy ...... 2 1.1.3 Usage of the Term “Allergy” ...... 5 1.2 Drug Allergy ...... 6 1.2.1 The Early Years ...... 6 1.2.2 Drugs, Haptens, and Prior Exposure ...... 6 1.2.3 Drug Allergies, Hypersensitivities, and Sensitivities (Intolerances) ...... 7 1.2.4 Risk Factors for Drug Allergy ...... 8 1.3 The Promise of Pharmacogenomics for understanding and managing Adverse Drug Reactions Including Drug Allergies ...... 9 1.4 Improvements In Drug Delivery and the Allergenicity of Drugs ...... 11 1.5 Scope of This Book ...... 11 1.5.1 The Place of Drug Allergy in Immunology ...... 11 1.5.2 The Need for a Single Text Book on Drug Allergy ...... 12 Summary ...... 12 Further Reading ...... 13 2 Classifi cation and Descriptions of Allergic Reactions to Drugs ...... 15 2.1 Hypersensitivity: The Early Years—From Koch to Gell and Coombs ...... 16 2.2 Classifi cation of Hypersensitivity Reactions ...... 16 2.2.1 Type I Hypersensitivity ...... 16 2.2.2 Type II Hypersensitivity ...... 23 2.2.3 Type III Hypersensitivity ...... 23 2.2.4 Delayed-Type (Type IV) Hypersensitivity ...... 24 Summary ...... 34 Further Reading ...... 35

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3 Mechanisms of Hypersensitivity ...... 37 3.1 Allergic Sensitization to Drugs and the Dogma of Previous Exposure ...... 38 3.1.1 Immunogenicity of Free and Conjugated Drugs ...... 38 3.1.2 Mediator Release by Free and Conjugated Drugs in Immediate Allergic Reactions ...... 40 3.1.3 Immunological Recognition of Free, Unconjugated Drug Molecules ...... 41 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities ...... 42 3.2.1 Initiating Events in the Production of IgE Antibody ...... 42 3.2.2 Allergic Release of Mediators of Hypersensitivity from Mast Cells ...... 43 3.2.3 Amplifi cation of IgE Antibody Production by Cellular Interaction ...... 45 3.2.4 Low-Affi nity IgE Receptor FcεRII (CD23) ...... 45 3.2.5 Important Mediators of the Type I Immediate Allergic Response ...... 46 3.2.6 Anaphylaxis ...... 58 3.2.7 Other Mechanisms of Anaphylaxis: IgG, PAF, and Nitric Oxide ...... 61 3.2.8 Drug-Induced Urticaria and Angioedema ...... 62 3.3 The Allergen-Induced Late Phase Reaction ...... 68 3.3.1 Early Studies: Implication of IgE Antibodies ...... 69 3.3.2 Cellular Responses in the Late Phase Reaction and Comparison with the Delayed- Type Hypersensitivity Response ...... 69 3.4 Drug-Induced Hypersensitivity and Immune Receptors ...... 70 3.4.1 Background ...... 70 3.4.2 Recognition and the Immune Response to Free, Unconjugated Drug ...... 71 3.4.3 Abacavir and the MHC- Presented Altered Peptide Model of Drug Hypersensitivity...... 72 3.4.4 Carbamazepine and Other HLA-Drug Hypersensitivity Associations ...... 73 3.4.5 The Question of Direct Drug Activation of T Cells Without Involvement of a Specifi c Peptide ...... 74 3.5 Desensitization of Drug- Allergic Patients ...... 75 3.6 Delayed-Type (Type IV) Hypersensitivity Drug Reactions ...... 76 3.6.1 The Cellular Basis of Type IV Hypersensitivity Cutaneous Drug Reactions ...... 76 3.6.2 T Helper Cell Responses and Th17 ...... 78 3.6.3 Delayed Cutaneous Adverse Drug Reactions ...... 79 3.7 Type II Hypersensitivity Drug Reactions ...... 84 3.8 Type III Hypersensitivity Drug Reactions ...... 86 Summary ...... 87 Further Reading ...... 90

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4 Diagnosis of Allergic Reactions to Drugs ...... 91 4.1 Case History ...... 92 4.2 Skin Testing ...... 93 4.2.1 General Aspects of Skin Testing for Drug Hypersensitivity ...... 93 4.2.2 Skin Prick Test Method ...... 94 4.2.3 Intradermal Testing ...... 95 4.2.4 Patch Tests ...... 97 4.3 Serum Immunoglobulin E Antibody Tests ...... 100 4.3.1 In Vitro Detection of Drug- Reactive IgE Antibodies ...... 101 4.3.2 Tests for the Clinic and Research ...... 103 4.3.3 Quantitation, Interpretation and Reporting of Results ...... 104 4.4 Drug Challenge (Provocation) Testing ...... 105 4.5 Detection and Measurement of Released Mediators/Markers of Hypersensitivity ...... 107 4.5.1 Tryptase ...... 107 4.5.2 Histamine ...... 109 4.5.3 Cysteinyl Leukotrienes ...... 111 4.6 Basophil Activation Test...... 113 4.6.1 Basophils and Background to the Basophil Activation Test (BAT) ...... 113 4.6.2 Basophil Activation Markers ...... 114 4.6.3 Some Technical Aspects ...... 115 4.6.4 Controls ...... 116 4.6.5 Evaluation of Results ...... 116 4.6.6 Application to Drug Allergies ...... 117 4.6.7 Analysis by Flow Cytometry of Intracellular Histamine and Its Release by Activated Basophils at the Single Cell Level ...... 117 4.6.8 Future Research and Conclusions ...... 118 4.7 Tests for Delayed Type Drug Hypersensitivity Reactions ..... 118 4.7.1 Lymphocyte Transformation Test ...... 120 4.7.2 The Local Lymph Node Assay ...... 121 4.7.3 Toward Nonproliferation- Based In Vitro Assays: Cell Surface Activation Markers, Cytokines, Chemokines, and Skin-Homing Receptors ...... 121 4.8 And Finally: Is the Patient Allergic to the Drug? ...... 124 Summary ...... 125 Further Reading ...... 127 5 β-Lactam Antibiotics ...... 129 5.1 Penicillins ...... 131 5.1.1 Incidence of Penicillin Hypersensitivity and Clinical Aspects ...... 131 5.1.2 Penicillin Antigens and Allergenic Determinants ...... 132 5.1.3 Heterogeneity of IgE Antibody Responses to Penicillins and the Spectrum of Penicillin Allergenic Determinants ...... 141

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5.1.4 Risk Factors for Immediate (Type I) Reactions to Penicillins ...... 147 5.1.5 Skin Testing Today for Immediate Hypersensitivity to Penicillin ...... 148 5.1.6 In Vitro Tests for Immediate Hypersensitivity to Penicillins ...... 151 5.1.7 Challenge (Provocation) Testing for Penicillin Hypersensitivity ...... 153 5.1.8 Penicillin Desensitization ...... 154 5.1.9 Delayed-Type Hypersensitivity Reactions to Penicillins ...... 156 5.1.10 The Genetic Basis of Penicillin-Induced Liver Injury...... 159 5.2 Cephalosporins ...... 159 5.2.1 Incidence of Cephalosporin Hypersensitivity and Clinical Aspects ...... 160 5.2.2 Clinical Aspects of Cross-Reactivity of Cephalosporins and Penicillins ...... 160 5.2.3 Structures and Classiﬁ cation of Cephalosporin Drugs...... 161 5.2.4 Cephalosporin Antigens and Allergenic Determinants ...... 161 5.2.5 Skin Testing with Cephalosporins ...... 173 5.2.6 Delayed Reactions to Cephalosporins ...... 174 5.3 Monobactams ...... 175 5.4 Carbapenems ...... 176 5.5 Clavams ...... 177 Summary ...... 178 Further Reading ...... 180 6 Other Antimicrobial Drugs ...... 183 6.1 Antibiotics ...... 184 6.1.1 Macrolides ...... 184 6.1.2 Tetracyclines ...... 186 6.1.3 Rifamycins ...... 187 6.1.4 Vancomycin and Teicoplanin ...... 188 6.1.5 Antibiotics Used Topically with Emphasis on Neomycin and Bacitracin ...... 193 6.1.6 Ribostamycin ...... 197 6.1.7 Chloramphenicol ...... 198 6.1.8 Clindamycin ...... 198 6.1.9 Pristinamycin ...... 200 6.1.10 Fosfomycin ...... 200 6.2 Antimicrobials Other than Antibiotics ...... 200 6.2.1 Sulfonamides ...... 200 6.2.2 Trimethoprim ...... 213 6.2.3 Quinolones ...... 218 6.2.4 Chlorhexidine ...... 225 6.2.5 Povidone–Iodine ...... 230 Summary ...... 231 Further Reading ...... 232

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7 Drugs and Other Agents Used in Anesthesia and Surgery ...... 235 7.1 Drug-Induced Reactions During Anesthesia ...... 235 7.2 Incidences of Drug- and Other Agent-Induced Anaphylaxis During Anesthesia ...... 237 7.3 Clinical Features of Anaphylactic and Anaphylactoid Reactions During Anesthesia ...... 238 7.4 Anaphylaxis to Neuromuscular Blocking Drugs ...... 239 7.4.1 Some Epidemiological Background ...... 239 7.4.2 Mechanisms Underlying Anaphylaxis to Neuromuscular Blocking Drugs ...... 240 7.4.3 Diagnosis of Anaphylaxis to Neuromuscular Blocking Drugs ...... 246 7.4.4 Cross-Reactions Between Neuromuscular Blocking Drugs ...... 256 7.4.5 The NMBD–IgE Conundrum: The Origin of IgE Antibodies to Neuromuscular Blocking Drugs ...... 263 7.4.6 Sugammadex and Anaphylaxis to Rocuronium ...... 267 7.4.7 Antigenic Similarity Between α-Bungarotoxin and Neuromuscular Blocking Drugs ...... 272 7.5 Anaphylaxis to Hypnotic Drugs Used in Anesthesia...... 273 7.5.1 Thiopentone ...... 273 7.5.2 Propofol ...... 276 7.6 Anaphylaxis to Colloids ...... 278 7.6.1 Hydroxyethyl Starch...... 278 7.6.2 Gelatin ...... 279 7.6.3 Dextrans...... 279 7.7 Local Anesthetics ...... 281 7.7.1 Chemistry ...... 281 7.7.2 Adverse Reactions to Local Anesthetics ...... 281 7.7.3 Diagnosis of Reactions to Local Anesthetics ...... 284 7.8 Polypeptides ...... 284 7.8.1 Protamine ...... 284 7.8.2 Aprotinin ...... 286 7.8.3 Latex ...... 287 7.9 Heparin ...... 289 7.9.1 Adverse Reactions to Heparin ...... 289 7.9.2 Diagnostic Methods ...... 289 7.9.3 Danaparoid and Hirudins...... 290 7.9.4 Fondaparinux ...... 290 7.10 Patent Blue V, Isosulfan Blue, and Methylene Blue ...... 291 Summary ...... 293 Further Reading ...... 294 8 Opioid Analgesic Drugs ...... 295 8.1 Terminology ...... 296 8.2 Structure–Activity Relationships ...... 296 8.3 Classiﬁ cation of Opioid Drugs ...... 301 8.4 Opioids and Histamine Release ...... 303 8.4.1 Histamine Receptors...... 303

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8.4.2 Early Studies on Opioid Drug-Induced Release of Histamine ...... 304 8.4.3 Summary of Morphine- Induced Hemodynamic and Cutaneous Changes in Humans ...... 305 8.5 Allergenicity of Opioid Analgesic Drugs ...... 308 8.5.1 Naturally Occurring and Semisynthetic Opioid Drugs ...... 308 8.5.2 Synthetic Opioid Drugs ...... 311 8.6 Resolving the Histamine- Releasing and Allergenic Effects in Diagnosing Reactions to Opioid Drugs ...... 313 8.7 Why Are Opioid Analgesic Drugs So Poorly Allergenic? ...... 315 8.8 Some Important Clinical Implications Related to the Use of Opioid Analgesic Drugs...... 316 Summary ...... 317 Further Reading ...... 317 9 Nonsteroidal Anti-Infl ammatory Drugs ...... 319 9.1 Therapeutic Applications of NSAIDs ...... 320 9.2 Patient Usage ...... 320 9.3 Classifi cation of NSAIDs ...... 321 9.3.1 Salicylates ...... 321 9.3.2 Propionic Acid Derivatives ...... 321 9.3.3 Aryl and Heteroaryl Acetic Acids ...... 321 9.3.4 Anthranilates (Fenamic Acid Derivatives) ...... 323 9.3.5 Oxicams (Enolic Acid Derivatives) ...... 323 9.3.6 Phenylpyrazolones...... 323 9.3.7 Anilides ...... 323 9.3.8 COX-2 Selective Inhibitors ...... 323 9.4 Mechanism of Action of NSAIDs ...... 324 9.4.1 Biosynthesis of the Prostanoids ...... 324 9.4.2 The Cyclooxygenase Isoforms COX-1 and COX-2 ...... 324 9.4.3 Classifi cation of NSAIDs by COX Isoenzyme Selectivity...... 326 9.4.4 The So-Called COX-3 Isoform ...... 327 9.5 Sensitivities to NSAIDs ...... 328 9.5.1 Defi nition and Epidemiology ...... 329 9.5.2 Clinical Classifi cation of Sensitivities to NSAIDs ...... 330 9.5.3 IgE Antibodies to Aspirin ...... 339 9.5.4 NSAID Sensitivities and BAT ...... 340 9.5.5 Genetic Mechanisms of Aspirin-Induced Sensitivities ...... 340 Summary ...... 341 Further Reading ...... 342 10 Contrast Media ...... 343 10.1 Iodinated Contrast Media ...... 344 10.2 Usage and Safety of Contrast Media ...... 346 10.3 Adverse Reactions ...... 347 10.3.1 Classifi cation and Symptoms ...... 347

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10.3.2 Incidence of Reactions ...... 349 10.3.3 Risk Factors ...... 350 10.3.4 Biphasic Reactions ...... 350 10.4 Mechanisms of Adverse Reactions to Iodinated Contrast Media ...... 351 10.4.1 Anaphylactoid and Anaphylactic Reactions ...... 352 10.4.2 Delayed Reactions ...... 355 10.5 Tests for the Diagnosis and Study of Adverse Reactions to Contrast Media ...... 356 10.5.1 Skin Tests ...... 356 10.5.2 IgE Antibody Tests ...... 357 10.5.3 Tests to Detect the Release of Mediators ...... 358 10.5.4 Challenge Tests ...... 359 10.6 Premedication for the Prevention of Anaphylactoid/ Anaphylactic Reactions to Iodinated Contrast Media ...... 360 10.7 Gadolinium-Based Contrast Agents ...... 361 10.7.1 Molecular Structures of Gadolinium-Based Contrast Agents ...... 361 10.7.2 Nephrogenic Systemic Fibrosis ...... 361 10.7.3 Other Adverse Reactions ...... 361 Summary ...... 365 Further Reading ...... 366 11 Biologics ...... 369 11.1 Monoclonal Antibodies for Therapy...... 370 11.1.1 Nomenclature ...... 370 11.1.2 Monoclonal Antibodies Approved for Therapy ...... 370 11.1.3 Immune Reactions to Monoclonal Antibodies ...... 371 11.2 Etanercept ...... 379 11.3 Interferons ...... 380 11.4 Interleukin 2...... 380 11.5 Denileukin Diftitox and Aﬂ ibercept ...... 381 11.6 Anakinra ...... 381 11.7 Anti-thymocyte Globulin ...... 382 11.8 Epoetins ...... 382 11.9 Human Insulin ...... 383 Summary ...... 384 Further Reading ...... 385 12 Corticosteroids ...... 387 12.1 Introduction, Incidence of Allergy, and Sensitization ...... 387 12.2 Corticosteroid Haptens: Metabolism and Degradation ...... 388 12.3 Delayed Reactions ...... 390 12.3.1 Clinical Presentation ...... 390 12.3.2 Diagnosis of Corticosteroid Hypersensitivity ...... 390 12.3.3 Structure–Activity Relationships of Corticosteroid Cross-Reactions...... 393 12.4 Immediate Reactions to Corticosteroids...... 394 12.4.1 Incidence, Sensitization, Clinical Presentation, and Risk Factors ...... 394

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12.4.2 Immediate reactions: Important/Interesting Findings and Observations Reported so Far ...... 394 12.4.3 Identifying and Understanding Cross- Reactions: Finding a Safe Alternative Corticosteroid ...... 396 12.4.4 Diagnostic Methods ...... 396 Summary ...... 397 Further Reading ...... 397 13 Drugs Used for Chemotherapy ...... 399 13.1 Taxanes ...... 400 13.1.1 Premedication for Taxanes ...... 404 13.1.2 Desensitization for Hypersensitivity Reactions to Taxanes ...... 405 13.2 Organoplatinum Chemotherapeutic Drugs ...... 405 13.2.1 Symptoms of Hypersensitivity to Platinum Drugs ...... 405 13.2.2 Incidences of, and Risk Factors for, Hypersensitivity Reactions to Platinum Drugs ...... 407 13.2.3 Mechanisms and Diagnosis of Platinum Drug-Induced Hypersensitivity Reactions ...... 407 13.2.4 Desensitization ...... 408 13.3 Tyrosine Kinase Inhibitors ...... 409 13.3.1 The Philadelphia Chromosome and Tyrosine Kinases ...... 409 13.3.2 Imatinib Mesylate ...... 410 13.3.3 Geﬁ tinib and Erlotinib ...... 411 13.4 Proteasome Inhibitors ...... 412 13.4.1 The Proteasome...... 412 13.4.2 Bortezomib ...... 412 13.4.3 Second Generation Proteasome Inhibitors ...... 415 13.5 Cytokine-release and tumor lysis syndromes ...... 415 Summary ...... 415 Further Reading ...... 417 14 Proton Pump Inhibitors ...... 419 14.1 Chemistry ...... 419 14.2 Mechanism of Action ...... 420 14.3 Hypersensitivity Reactions to Proton Pump Inhibitors ...... 420 14.4 Diagnosis of Hypersensitivity to Proton Pump Inhibitors ...... 422 14.5 Proton Pump Inhibitors, Gastroesophageal Reﬂ ux Disease, and Asthma...... 423 14.6 Other Safety Concerns with Proton Pump Inhibitors ...... 423 Summary ...... 423 Further Reading ...... 424

Postface – Concluding Remarks ...... 425

Index ...... 429

[email protected] Adverse Reactions to Drugs and Drug Allergy: Scope 1 of This Book

Abstract In what is essentially a pharmacologically based classiﬁ cation of adverse drug reactions (ADRs), unpredictable and dose-independent drug reactions, designated type B reactions, include hypersensitivity responses while those reactions designated as type A are predictable, dose- dependent, and make up about 80 % of all ADRs. Previous exposure is not always a prerequisite for allergic sensitization, and there are many instances where reactions occur after initial contact with poorly reactive drugs that do not bind to proteins. Risk factors for drug allergy can be divided into those that are patient-related (age, sex, current diseases, previous exposure, genetic factors) and those that are drug-related (nature and cross-reactivity of drug, degree of exposure, route of administration). Genomic studies are already helping to explain some ADRs, for example, the association in Han Chinese of carbamazepine-induced Stevens–Johnson syndrome with HLA-B*15:02 and the association of abacavir hypersensitivity in abacavir hypersensitivity syndrome with HLA-B*57:01. It seems likely that multiple rather than single genes are involved in ADRs. Drug allergy studies promise to provide signiﬁ cant insights into important areas of biomedical investigation including cell recognition and interaction processes, relationships between receptors and effector pathways and mechanisms of mediator actions.

severe and even life-threatening, necessitating 1.1 Adverse Drug Reactions the substitution or discontinuation of preferred medications. An additional unwanted clinical 1.1.1 Deﬁ nition response in a sick patient already under treatment constitutes extra burdens for the patient A major and seemingly ever-present risk of and the managing physician. With approximately pharmacotherapy is adverse drug reactions 5 % of patients developing adverse reactions (ADRs). Drug reactions occur frequently and during drug therapy and up to 10 % said to react may need expert management. Reactions can be in hospitals, this adds up to a signiﬁ cant public

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 1 Relationships, DOI 10.1007/978-1-4614-7261-2_1, © Springer Science+Business Media, LLC 2013

[email protected] 2 1 Adverse Reactions to Drugs and Drug Allergy: Scope of This Book health problem. The often quoted WHO 1.1.2 Terminology: Classifi cation defi nition of an ADR, published 40 years ago, is of Adverse Drug Reactions “a response to a drug that is noxious and unin- and the terms Hypersensitivity, tended and occurs at doses normally used in man and Allergy for the prophylaxis, diagnosis or therapy of disease, or for modifi cation of physiological func- The study of ADRs falls within the discipline of tion.” Questioning this defi nition and in particular pharmacovigilance. In an early pharmacological the inclusiveness of the wording for minor reac- classifi cation, ADRs were distinguished primar- tions, Laurence and Carpenter in A dictionary of ily on the basis of dose-related and non-dose- pharmacology and allied topics: Elsevier, 1998 , related reactions. These two types of reactions suggested: “A harmful or signifi cantly unpleas- were sometimes called types A and B, respec- ant effect caused by a drug at doses intended for tively. Approximately 80 % or more of ADRs are therapeutic effect (or prophylaxis or diagnosis) predictable, can be anticipated from the drug’s which warrants reduction of dose or withdrawal pharmacological actions, are dose-dependent, of the drug and/or foretells hazard from future and resolve when the dose is reduced or with- administration.” I.R. Edwards of the Uppsala drawn. Unpredictable reactions, sometimes Monitoring Centre, a WHO collaborating centre called idiosyncratic drug reactions, are generally for international drug monitoring, and J.K. unrelated to the drug’s pharmacological actions, Aronson of the Radcliffe Infi rmary, Oxford, are independent of dose, and, even though they regard the WHO defi nition as vague, especially usually resolve when treatment is stopped, reac- with regard to the term “noxious,” ask just how tions sometimes progress. From about the early minor can an adverse reaction be, query the nar- 1980s, three further reaction categories were rowness of the term “drug” and disagree with recognized, one related to dose and time and one what they regarded as ambiguities in other pub- classifi ed as delayed but divided into time- lished defi nitions. To cover these perceived defi - related and withdrawal reactions. More recently, ciencies, Edwards and Aronson proposed their a sixth category, “unexpected failure of therapy” own succinct defi nition: “An appreciably harm- has been added. In some classifi cations of ADRs, ful or unpleasant reaction, resulting from an a seventh category G, “genetic reactions,” is intervention related to the use of a medicinal included. This essentially pharmacologically product, which predicts hazard from future based overall classifi cation of ADRs, together administration and warrants prevention or spe- with some distinguishing features and examples cifi c treatment, or alteration of the dosage regi- of drug reactions, are shown in Table 1.1 . men, or withdrawal of the product.” For drug allergy, and for our purposes, an The United States Food and Drug immunological classifi cation is more informative Administration (FDA) states that any serious and useful. The unpredictable and dose- adverse event should be reported to the FDA and independent drug reactions, that is, type B reac- defi nes such an event as “any undesirable experi- tions, include the reactions that are said to be ence associated with the use of a medical prod- hypersensitivity responses but also three catego- uct in a patient.” The event is said to be serious ries of nonimmune drug sensitivities termed when the patient outcome is death, life-threatening, pseudoallergy, idiosyncratic reactions and type B hospitalization (initial or prolonged), disability intolerances. In Fig. 1.1 , the four different cate- or permanent damage, congenital anomaly/birth gories of allergic reactions are referred to as defect, required intervention to prevent perma- hypersensitivity reactions while the nonimmune nent impairment or damage (devices), and other (or nonallergic) type B adverse drug reactions are serious important medical events (e.g., allergic referred to simply as sensitivities. The term bronchospasm, serious blood dyscrasias, or sei- “hypersensitivity” is somewhat problematic since zures or convulsions that do not result in it has a long and well-established usage in hospitalization). immunology and allergy but also a history of

[email protected] 1.1 Adverse Drug Reactions 3

Table 1.1 Classifi cation of adverse drug reactions Reaction type Examples of reaction Main features of reaction A. Augmented – Toxic (intolerant) reactions—e.g., – Majority of ADRs pharmacologic effects a serotonin syndrome to opioids, – Common antidepressants; digoxin toxicity – Predictable – Side effects—e.g., bronchospasm to – Usually dose dependent β-blocker in hypertensive patient; dry – Related to pharmacologic reaction mouth to antidepressants of drug – Low mortality B. Bizarreb (see Fig. 1.1 ) – Immunologic reactions – Relatively uncommon – Idiosyncratic reactions – Unpredictable – Pseudoallergy – Rarely dose dependentc – Intolerance – Unrelated to drug’s pharmacologic action – High mortality C. Chronic (continuous) – Corticosteroid-induced suppression – Uncommon effects of hypothalmic–pituitary–adrenal axis – Cumulative dose and long-term – Renal papillary necrosis caused by exposure required phenacetin D. Delayed effects – Carcinogenesis – Time-related. Apparent some time – Teratogenesis—e.g., vaginal after drug exposure adenocarcinoma induced by – Uncommon diethylstilbestrol – Usually dose-related E. End-of-treatment effects – Opiate withdrawal syndrome – Occurs with little or no delay after (withdrawal effects) – β-Blocker withdrawal withdrawal of drug – Uncommon F. Failure of therapy – Resistance to drug action—e.g., – Common resistant bacteria to antibiotic or – Usually dose-related tumor to chemotherapy – May be caused by drug – Oral contraceptive dose too low interactions G. Genetic reactionsb,d – Abnormal drug metabolism due – Abnormal drug metabolism to inherited factors (alleles appears to be uncommon of P450 (CYP), N -acetyltransferase, – Pharmacogenomic studies still in pseudocholin-esterase) early stages – HLA–drug hypersensitivity – Ethnicity seems to matter for some associations (e.g., abacavir, drugs, e.g., carbamazepine carbamazepine, allopurinol) – Succinylcholine sensitivity – Porphyria a Said to account for ~80 % of ADRs b There is evidence that some type B reactions are under genetic control c Exceptions exist, e.g., with type IV hypersensitivity skin reactions; responses to vaccines; desensitization with increasing dosages of drug d ADRs likely to be multigenetic phenomena meaning different things to different people, and what was described as acceptable nomenclature this confusion is still apparent today. In 2001, the for allergic diseases with the ultimate goal of European Academy of Allergology and Clinical improving communication in the fi eld of allergy. Immunology (EAACI) published a EAACI Acknowledging that the nomenclature proposed Position Statement entitled, in part, “A revised by the EAACI was based on reaction mecha- nomenclature for allergy.” After setting up a nisms causing the signs and symptoms of allergic Nomenclature Review Committee to review the disease and these mechanisms were usually EAACI position statement, the World Allergy infl ammatory, the WAO Nomenclature Review Organization (WAO) set about promoting globally Committee issued a revised nomenclature for

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ADVERSE DRUG REACTIONS TYPE B

ALLERGIC NONIMMUNE DRUG SENSITIVITIES

Type I Type II Type III Type IV Idiosyncratic Pseudoallergy Intolerence hypersensitivity hypersensitivity hypersensitivity hypersensitivity reactions Immediate Immune Delayed Most reactions Malignant Tinnitus IgE-mediated Cytotoxic complex Cell-mediated to NSAIDs hypothermia induced by Direct mast cell Halothane small doses Penicillins Cephalosporins Penicillins NSAIDs degranulation hepatitis aspirin Cephalosporins Penicillins Cephalosporins Anti-microbials (muscle relaxants, Antibacterials Quinine Sulfonamides Anti-convulsants Drug-induced opioids, Neuromuscular Tetracycline (carbamazepine) Glu-6-PO vancomycin, 4 blocking drugs Local anesthetics dehydrogenase contrast media) deficiency anemia (antimalarials, sulfonamides)

Fig. 1.1 Classifi cation of immune (allergic) and nonim- nonallergic sensitivities or intolerances. Some drugs com- mune sensitivities to drugs. The former are referred to monly involved in allergic reactions and a few examples here as hypersensitivities and the latter as nonimmune or of nonimmune sensitivities are shown allergy in 2003. Of its proposed defi nitions and II and immune complex-mediated type III hyper- explanations, those dealing with the terms, hypersensitivities, are included with the type I and type sensitivity, nonallergic hypersensitivity, and ana- IV hypersensitivities in the Gell and Coombs phylaxis can be considered contentious. classifi cation of hypersensitivity reactions (see Anaphylaxis, and the proposed defi nition of it, is Chap. 2). However, while this classifi cation has discussed in Chap. 2 . “Hypersensitivity” was served allergists, clinical immunologists, and defi ned as “objectively reproducible symptoms researchers well for half a century, it is clear that and signs initiated by exposure to a defi ned stim- for some adverse drug reactions there are ulus at a dose tolerated by normal persons.” responses that simply do not fi t into the four Gell “Sensitivity” was said to be an acceptable alter- and Coombs categories. Some reactions to con- native in special circumstances. This defi nition of trast media and nonsteroidal anti-infl ammatory hypersensitivity is inadequate. Apart from the drugs (NSAIDs) are two examples that readily absence of any reference to the adverse nature of spring to mind as well as skin reactions such as a reaction, by omitting any mention of an immu- alopecia, folliculitis, and hyperpigmentation. nologic mechanism, the description fl ies in the With some of these drugs, reactions occur that face of well entrenched, widely accepted, and have an immune basis (viz., type I or type IV), long-term usage. Firmly established, if not that is, they are true hypersensitivities, but in ingrained, use of the labels “immediate hypersen- other responses to drugs no immune mechanism sitivity” for IgE antibody-mediated, type I aller- can be identifi ed. For NSAIDs, the mechanism gic reactions and “delayed hypersensitivity” for underlying most adverse patient responses delayed-type, type IV, or cell-mediated reactions appears to be drug- induced redirection of media- illustrates how the term hypersensitivity has tor synthesis in the arachidonic acid cascade from become synonymous with an immune reaction. the cyclooxygenase to the lipoxygenase pathway This is reinforced by the still accepted and com- with no antibody or immune cell involvement monly used classifi cation where two other immune (see Chap. 9). Showing some features of a hyper- mechanisms, antibody-dependent cytotoxic type sensitivity response and usually presented to an

[email protected] 1.1 Adverse Drug Reactions 5 allergist, clinical immunologist, or dermatologist, to the accepted scheme of classifi cation of hyper- it is not diffi cult to see why such responses are sensitivity states. At present, the common feature commonly viewed as hypersensitive reactions, for grouping many different reactions with a but should the long-standing defi nition of wide range of signs and symptoms into a broad hypersensitivity be changed to accommodate but unifying classifi cation scheme is the immune diverse drugs that exert their effects by a num- basis of each of the responses. Employment of ber of different mechanisms and where no com- the terminology used for these responses should mon humoral or cellular immune basis of action not be stretched to accommodate reactions that exists? proceed by an entirely different mechanism. It In defi ning “allergy,” the WAO describe it as “a seems likely, however, that many currently inad- hypersensitivity reaction initiated by specifi c equately researched and poorly understood drug immunologic mechanisms.” “Drug allergy” should reactions will probably be shown to be allergic. therefore only be used for an ADR where an In the meantime, for reactions to drugs such as immunologic mechanism has been established. It NSAIDs, contrast media and many adverse skin was further stated that: “When other mechanisms responses where it is already clear that immune can be proven, as in hypersensitivity to aspirin, the mechanisms are not involved, or where evidence term nonallergic hypersensitivity should be used.” one way or the other is not yet available, the terms If a contrast medium, a NSAID, or any other agent “sensitivity” or “intolerance” should be used is known not to act via an immune mechanism or instead of “hypersensitivity.” if an immune basis of the reaction cannot be established, the NSAID aspirin can indeed help in providing the appropriate terminology, not by the 1.1.3 Usage of the Term “Allergy” suggested use of “nonallergic hypersensitivity” but by the already commonly employed and clear Derived from the Greek words allos, meaning terms “aspirin sensitivity,” “aspirin-intolerant,” or other and ergon, meaning work, task, purpose “aspirin- induced” (as in asthma). Thus, a patient (or ergein, to work), the term allergy was intro- with asthma induced by (say) celecoxib would duced in 1906 by the Austrian pediatrician simply be described as celecoxib-sensitive, or C.P. von Pirquet who seems to have thought of it intolerant to celecoxib, and the condition referred as a state of changed reactivity by the host, cov- to as celecoxib-induced asthma, celecoxib-intoler- ering both an increase (hypersensitivity) and a ant asthma, or celecoxib-exacerbated asthma. decrease (hyposensitivity) in the allergic A number of late, polymorphic reactions to drugs response. However, the word was not used to with mechanisms still to be fully worked out occur mean hypersensitivity or immunity but as a term several days after administration. Reactions may for the response that could produce protective take the form of maculopapular eruptions, urti- immunity on the one hand or hypersensitivity caria, fi xed drug eruptions, acute generalized with its detrimental effects on the other. This is exanthematic pustulosis, drug reaction (rash) with different to today’s use where the word “allergy” eosinophilia and systemic symptoms (DRESS), is restricted to specifi c hypersensitivity to for- Stevens–Johnson syndrome, toxic epidermal eign antigens, some of which are also toxic in necrolysis, and vasculitis. Temporal relationships, their own right (such as venoms) and some not together with accumulating evidence for activated (such as foods). It is often said that the medical CD4+ and CD8+ activated lymphocytes from profession, as well as the public and the media, lesions and the generation of drug- specifi c T cell uses the term allergy inappropriately, loosely, or clones, suggest that these reactions are allergic too casually, and nothing short of a concentrated rather than due to direct toxicity. educational campaign (which is unlikely to hap- In summary, and simply put, there seems to be pen) will help to overcome this misuse. There is no compelling reason to alter an established and a widespread tendency to consider a large variety widely understood defi nition that is fundamental of drug-induced adverse systemic and local effects

[email protected] 6 1 Adverse Reactions to Drugs and Drug Allergy: Scope of This Book as allergic in nature due to lazy use of terminol- combine covalently with protein and what ogy or lack of medical understanding of what Landsteiner described as a “loose attachment” constitutes a true allergic response. It is not to protein seen with, for example, salts of heavy uncommon, for example, to hear a wide range of metals. Despite the contemporary demonstra- different responses to an administered drug, tion of passive sensitization of normal animals from minor affl ictions such as headache, dry to simple chemicals with serum antibodies, the mouth, nausea, or syncope to cardiovascular and existence of drug-reactive antibodies in subjects CNS reactions, described as an “allergic reac- allergic to drugs was generally discounted. The tion.” This needs to be taken into account in any prevailing view seemed to be that there was no analysis or consideration of drug allergy where a clear division between drug allergy and other wide coverage of many different drugs of differ- immunological manifestations. It took almost a ent pharmacological actions may need to be quarter of a century before the chemist and phy- reviewed. In addition to this pharmacological sician, Bernard Levine and others with their diversity and nonallergic adverse responses, the work on penicillins (see Chap. 5 ), laid solid spectrum of true allergic reactions elicited by foundations substantial enough to ultimately drugs can range from a clear type I immediate support the necessary scientifi c chemical frame- hypersensitivity reactions manifesting as cata- work that was to come. Progress was initially strophic anaphylaxis with all or some of urti- slow but now the “fi nished” edifi ce promises to caria, angioedema, bronchospasm, and be far more complex than Landsteiner and his cardiovascular collapse to mild IgE antibody- contemporaries could have imagined as special- mediated rhinitis or a mild and transient T cell- ist workers with specialized tools of the modern mediated rash. era move in to expand structures and add the necessary, and sometimes surprising, detail.

1.2 Drug Allergy 1.2.2 Drugs, Haptens, and Prior 1.2.1 The Early Years Exposure

For the still incomplete construct of “Drug From the time of immunology’s earliest practitio- Allergy,” activity can be seen to have been initi- ners, how the immune system recognizes and ated in 1907 when Wolff-Eisner surveyed the deals with “small” molecules with molecular site with the prediction that nonantigenic sub- weights less than 1 kDa has intrigued researchers. stances can induce dermatological sensitization Following earlier suggestions and results indicat- after combination with patients’ “self” proteins. ing that some small molecular weight chemicals In the 1920s, Bloch and Steiner-Wourlisch, can sensitize skin after combination with host Wedroff, Mayer, and then Landsteiner and proteins, Landsteiner and Jacobs demonstrated a Jacobs (1935 , 1936 ) subsequently cleared the clear relationship between skin sensitizing capac- ground and put down the footings by demon- ity and the chemical reactivity of haptens to cova- strating sensitization of guinea pigs and humans lently bind to a carrier protein. While with simple chemicals. Although the explana- Landsteiner’s landmark studies on hapten recog- tion for the formation of antigens from simple nition inﬂ uenced generations of subsequent reactive chemicals such as acyl chlorides and investigators, recent progress in elucidating acid anhydrides seemed satisfying, Landsteiner mechanisms, both immune and pharmacological, was aware that for less reactive compounds, for of some different drug intolerances promises to example, quinine, “a chemical interpretation is expand our understanding of the body’s responses not immediately to hand.” Two explanations to small antigens free of a macromolecular car- were advanced—conversion of unreactive rier. Note also the doubtful relevance of delayed chemicals in vivo to reactive compounds able to (contact) hypersensitivity studies in laboratory

[email protected] 1.2 Drug Allergy 7 animals to human allergies, especially immediate and particularly neuromuscular blocking drugs, reactions. In addition, rodents are far from being and sometimes β-lactams produce positive skin an ideal animal model for the human allergic reactions in allergic patients. In fact, the former state since, apart from the guinea pig, they are not drugs are routinely used as free drugs in routine always easy to sensitize and make allergic, the diagnosis and some penicillins and cephalospo- spectra of mediators released and the end organ rins sometimes elicit clearer and more specifi c responses to these mediators often differ from the skin reactions than the corresponding drugs in human responses, and homocytotropic antibodies conjugated form. This subject is considered in may differ and are not always of the IgE class. forthcoming chapters. The conundrum of previous exposure is also still with us, and there are numerous examples of allergic reactions to poorly reactive drugs where 1.2.3 Drug Allergies, no protein binding, either by the parent molecule Hypersensitivities, or any putative metabolite or degradation prod- and Sensitivities (Intolerances) uct, can be demonstrated. Landsteiner believed that previous exposure to an antigen is a prereq- In accordance with the defi nition of allergies pos- uisite for sensitization and an allergic response sessing an immune basis, an allergic reaction to a but, even acknowledging the possibility of cross- drug is mediated by antibody or cells of the reactions, it is clear that this dogma of prior con- immune system. Thus, immediate type I IgE tact does not always hold. Examples of this keep antibody- mediated and delayed-type, type IV T cropping up as subsequent pages in this book cell-mediated reactions to a drug are considered reveal. There seems to be little information on true drug allergies. Type II antibody-mediated what could be termed innate allergic sensitivity, (generally IgG or IgM) cytotoxic reactions with for example, “natural” IgE antibodies to some complement involvement to drugs such as peni- allergens (including drugs) but its involvement in cillins, cephalosporins, quinine, and pyrazolones some cases seems likely. For drug allergy, the and type III reactions mediated by soluble question of whether the allergic sensitivity to a immune complexes with antibodies mostly of the “small” organic molecule (usually prepared by IgG class to drugs such as penicillins, sulfon- total synthesis and therefore not “natural” in any amides, and quinolones are seen as true drug sense) is genetically determined or derived from hypersensitivities. Where reactions to drugs pro- natural exposure, remains of great intellectual ceed with no identifi able immune mediation, and practical medical interest. There are already such as occurs with nonsteroidal anti- infl ammable signs, however, that with application of modern drugs and contrast media and with direct hista- genetic technical approaches and insights, includ- mine releasing agents such as opioids and neuro- ing identifi cation of HLA markers for drug- muscular blockers, responses are viewed here as induced hypersensitivities, progress in this area is sensitivities or intolerances, not as hypersensi- well underway in an atmosphere of signifi cant, tivities and, therefore, not true allergic responses. but seemingly justifi ed, expectation. With true drug allergies defi ned as reactions Just as conjugation of a small nonantigenic involving immune mechanisms, and bearing in molecule such as a drug to a macromolecular car- mind the ADRs covering the wide range of rier is said to be a requirement for allergic sensi- reaction types set out in Table 1.1, it is to be tization and subsequent reaction, multivalency of expected that the two lists of drugs (immune- verse the small molecule or hapten is said to be neces- nonimmune-based) will be signifi cantly different. sary for detecting skin test sensitivity to drugs. In From a recent survey of nearly 3,700 patient practice, this is not always the case—drugs such episodes analyzed for ADRs in hospital inpatients as trimethoprim, sulfamethoxazole, opioids in the UK, the drugs most frequently implicated in (diluted beyond their histamine-liberating con- provoking the reactions were, in order: loop diuret- centrations), some contrast media, thiopentone ics, opioids, systemic corticosteroids, inhaled

[email protected] 8 1 Adverse Reactions to Drugs and Drug Allergy: Scope of This Book

RISK FACTORS FOR DRUG ALLERGY

PATIENT-RELATED DRUG-RELATED

Current Genetic Previous Cross sensiti- Degree of Route of Age Sex Nature of diseases factors exposure drug vity/reactivity exposure administration

Adults ≥ Females Asthma Atopy not a Risk ↑ with Protein- Recognition Intermittent IV, IM, children ≥ males Infection risk previous reactive of structurally exposure topical ↑ with exposure drugs similar drugs → sensi- Herpesvi- Ethnicity BUT tization Oral safer ridea, HIV & genetics − reactions Protein HLA asso- can occur on (insulin, ciations first contact streptokinase etc)

Fig. 1.2 Patient-related and drug-related risk factors for drug allergies beta-agonists, penicillins, oral anticoagulants, seem to indicate that female gender is a risk for cephalosporins, compound analgesics with opioid, adverse drug reactions, especially cutaneous reac- macrolide antibiotics, and low molecular weight tions, but other surveys dispute this. Asthma is heparins. From the above list, a top ten compila- associated with increased risk but the claim that tion of drugs causing true allergic reactions would hypersensitivity is more common in systemic include only the penicillins, cephalosporins, pos- lupus erythematosus has not been confi rmed. sibly heparins, and the compound analgesics with Some drugs, particularly β-lactams like ampicillin opioid if the analgesics were nonsteroidal anti- and amoxicillin and particularly in children, show infl ammatory drugs, and perhaps, but not likely, a temporal association between drug exposure macrolide antibiotics and opioids. and rashes. It seems likely, however, that in most cases the reaction is produced by the infectious agent (e.g., in infectious mononucleosis) or by 1.2.4 Risk Factors for Drug Allergy interaction between the drug and the infectious agent. Such responses do not appear to be immu- Risk factors for most patients and for most drugs nologically mediated. An increased risk of reac- are not always identifi ed or, at best, only vaguely tions is seen in patients infected with herpesviridae defi ned, and there is usually little certainty in try- and HIV viruses, good examples being reactions ing to anticipate a drug reaction. In recent years, to cotrimoxazole, abacavir, or nevirapine in signifi cant progress has been made with a very patients with HIV infection. Atopic patients do small number of drugs (see, for example, abaca- not show a higher rate of sensitization to drugs vir, Sects. 1.3 and 3.4.2), but these remain the but, once a reaction occurs, they are at increased exception. In most cases, some broad risk factors risk of it being serious, and this is also true for related to the patient and the drug (Fig. 1.2 ) are uncontrolled asthmatics and patients with food considered, but this highlights our ignorance of the allergies. A few studies indicate that genetics and subject and represents an early stage in the under- ethnicity can be important in certain drug aller- standing of the subject of drugs and patient risk. gies but the situation is certain to be complex with Hypersensitivity reactions appear to be less fre- multiple genes as well as environmental factors quent in children than adults and, in some surveys, involved. The search for associations between less frequent in the elderly. Epidemiological data drug allergies and human leukocyte antigens

[email protected] 1.3 The Promise of Pharmacogenomics for understanding and managing Adverse Drug Reactions... 9

(HLA) of the major histocompatibility complex agents. This is particularly important with, for (MHC) on chromosome 6 remains an active area example, the neuromuscular blocking drugs, the of investigation, and one might expect further dis- β-lactams, sulfonamides, and pyrazolones where coveries that help to distinguish at risk patients clearly similar or identical structural groupings before administration of a potential harmful drug have been implicated as allergenic determinants (see example of abacavir below). As so often seen in allergic recognition. Cross-sensitization may in immune responses, previous exposure to drugs also be important not only from an immuno- can also induce increased sensitivity and here the chemical basis but also from the pharmacological question of cross-reactivity needs to be kept in action of drugs, for example, NSAIDs where mind (see below). Note, however, as already structurally different drugs such as aspirin and pointed out, there are many examples of reactions oxicams provoke sensitivity to COX-1 inhibitors occurring on fi rst exposure as witnessed by reac- even though their chemical structures show tions to cotrimoxazole, quinolones, muscle relax- marked differences. ants, contrast media, and so on. When examining drug-related risks, the nature of the drug itself is the fi rst and most obvious 1.3 The Promise of consideration. A drug’s chemical properties, in Pharmacogenomics for particular its molecular weight, structural com- understanding and managing plexity, and chemical reactivity are relevant. Adverse Drug Reactions Protein reactivity of a drug is a special consider- Including Drug Allergies ation, and drugs that are themselves proteins such as insulin, vaccines, biologics such as monoclo- The application of genomics, the study of the nal antibodies, interferons, etanercept, afl iber- entire genome of an organism, together with cept, anakinra, streptokinase and the anticancer molecular genetics, promises to reduce some agent L-asparaginase are already potential immu- adverse drug reactions in the future, improve nogens and allergens. The dose of drug and the therapeutic outcomes, and help tailor individual duration and frequency of treatment with the therapies. The variable clinical response to a drug drug can affect the risk of reaction. High dosage by different individuals is a major problem in and prolonged administration may lead to higher medicine since it can lead to both clinical failure risk but again, this is not a hard and fast rule. If and adverse effects in individuals and subpopula- anything, intermittent dosage seems more likely tions of patients. The study of pharmacogenom- to lead to sensitization and hence increased risk. ics is concerned with the relationship between The route of administration of drug can have a patients’ genes and their responses to drugs. marked effect exemplifi ed by the higher inci- Pharmacogenomics offers one avenue toward dence of anaphylaxis to a particular drug when personalized medicine aimed at prescribing the given intravenously. Intramuscular administra- optimum drug at the optimum dose for each tion also carries a higher risk than subcutaneous patient. Pharmacogenetics had its initial impact injection with the oral route being the safest. on drug therapy when the prolonged muscle However, sensitization and severe reactions can relaxation of succinylcholine experienced by also follow oral dosage. Topical application is some patients was explained by an inherited associated with a high incidence of sensitization defi ciency of a serum cholinesterase, and anti- and should be completely avoided with some malarial-induced hemolysis was shown to be due drugs such as the antibiotics chloramphenicol, to inherited variants of glucose-6-phosphate penicillins, and neomycin. The same is true for dehydrogenase (Fig. 1.1 ). More recently, the dis- sulfonamides. Lastly, cross-reactivity generally covery of genetic polymorphisms of the drug- has a structural basis and it is unwise to prescribe metabolizing enzyme cytochrome P450 2D6 or drugs or change medications without at least a CYP2D6 has led to the discovery of numerous basic knowledge of the chemical structures of the variant alleles at the CYP2D6 locus, many of

[email protected] 10 1 Adverse Reactions to Drugs and Drug Allergy: Scope of This Book which are associated with marked differences in An explanation of why abacavir is tolerated by enzyme function. Almost 25 % of drugs are 45 % of patients positive for HLA-B*57:01 is still metabolized by CYP2D6 and differences in to be found but, in the meantime, a genetic screen- CYP2D6 activity and drug clearance of up to ing test has been implemented globally as part of 40-fold can lead to severe adverse effects and primary HIV clinical practice. Basic studies in the patients who do not respond to their drug laboratory have revealed that AHS is specifi cally therapy. restricted by HLA-B*57:01 and mediated by Some examples of how pharmacogenomic CD8+ lymphocytes. This classic example of what studies can provide invaluable insights into ADRs has become a successful translation of pharma- are provided by the discoveries of the strong cogenomics into the clinic under the direction of association in Han Chinese of carbamazepine- lead investigator Simon Mallal in Western induced Stevens–Johnson syndrome and the Australia serves as a prototype study for other HLA-B*15:02 allele and myelotoxicity seen in drugs where genetic testing might be utilized for thiopurine methyltransferase-defi cient patients on the prevention of drug reactions. The recently elu- azothioprine. Such discoveries have led to the US cidated molecular basis of AHS is covered in FDA recommending genetic testing of patients Chap. 3 , Sect. 3.4.3 . prescribed certain drugs and the employment of In an interesting evaluation of the potential labels containing pharmacogenomic information. role of pharmacogenomics in reducing the inci- Abacavir hypersensitivity syndrome (AHS) pro- dence of ADRs, 22 variant allele articles were vides another fascinating example of the contribu- reviewed and matched with 27 drugs frequently tion of genetic factors to the predisposition of implicated in ADRs from 18 ADR studies. Fifty some individuals to drug-induced hypersensitiv- nine percent of the drugs are known to be metab- ity reactions. Abacavir, a reverse transcriptase olized by at least one enzyme with a variant allele inhibitor administered to patients infected with that causes poor metabolism whereas only human immunodefi ciency virus-1 (HIV-1), can 7–22 % of drugs randomly selected are metabo- provoke a potentially fatal multiorgan response, lized by a variant allele. The authors concluded AHS, involving fever, skin rash, and gastrointesti- that drug therapy suited to an individual’s genetic nal and respiratory symptoms in about 2–8 % of makeup might lead to signifi cant reductions in patients. Investigations revealed a strong associa- ADRs. As a logical extension of such fi ndings, a tion between abacavir hypersensitivity and the proposal to collect and utilize pharmacogenomic well-defi ned 57.1 MHC ancestral haplotype information after regulatory approval of a drug which comprises HLA-B*57:01, HLA-DR7, and has been put forward. As part of surveillance HLA-DQ3 with an odds ratio greater than 100. after a drug’s launch, patients taking the drug Although early results indicated that HLA- would lodge a biological specimen (e.g., a blood B*57:01 held promise as a screening test to pre- spot) and undergo a genetic scan. DNA compari- vent AHS, some cases of AHS proved negative for sons between patients with and without ADRs the HLA allele and low sensitivities of it for AHS would then identify genetic markers that could were seen in nonwhites. However, screening stud- be used to identify patients at risk of an ADR to ies suggested that HLA-B*57:01 screening can the drug. The pharmacogenomic approach to largely eliminate AHS, and patch testing was ADRs is now extending to some of the most found to distinguish patients with true immuno- important medications with, for example, new logically mediated AHS from false positives. It is information on the frequently prescribed statin now known that HLA-B*57:01 has a negative drugs and warfarin, the anticoagulant widely predictive value of 100 % for AHS confi rmed by used to prevent thrombosis and thromboembolism. patch testing across both white and black popula- The SLCO1B1 gene on chromosome 12 encodes tions, and HIV treatment guidelines recommend a transporting polypeptide that regulates hepatic HLA-B*57:01 screening as part of the routine uptake of statins. Recently, common variants in care of patients before being prescribed abacavir. SLCO1B1 that are strongly associated with an

[email protected] 1.5 Scope of This Book 11 increased risk of statin-myopathy have been not known to provoke allergic reactions needs to identifi ed and an algorithm for estimating the be kept in mind in the planning of preclinical appropriate warfarin dose on both clinical and drug safety assessments of those drugs modifi ed genetic data has been developed. by efforts to improve drug delivery. Note, however, that it now seems more likely that multiple rather than single genes are involved in ADRs, and this may add greatly to the diffi cul- 1.5 Scope of This Book ties of ultimately successfully applying these approaches to the everyday clinical situation. 1.5.1 The Place of Drug Allergy in Immunology

1.4 Improvements In Drug Drug allergy has never been a topic given much Delivery and the Allergenicity coverage by textbooks of immunology and the of Drugs subject, if considered at all by general allergy texts, is often restricted to penicillin with no, or With increasing usage of different carrier sys- scant, details provided on other drug allergies. In tems to improve drug solubility, delivery, or sta- two of the currently most widely studied and bility, selected drug modifi cations may give rise respected immunology textbooks used at under- to some unanticipated and unwanted conse- graduate and graduate level and by medical stu- quences as well as the hoped-for improvement(s). dents, coverage in one is restricted to three As more drug carriers such as cyclodextrins, sentences on anaphylaxis to penicillin and, in the dendrimers, vesicles like liposomes and nio- other, to a short paragraph on the involvement of somes, liquid crystals, soluble polymers, and penicillin in IgE-mediated reactions and type II micelles formed by self-assembly of amphiphi- hypersensitivity responses. In the latter case, the lic block copolymers, cell ghosts, and hydrogels name of the only other drug mentioned is found are employed for a wider variety of drugs, in the sentence: “Another drug that is known to carrier-induced changes including chemical provoke anaphylaxis is the anesthetic hexame- modifi cations, exposure, masking of structural thonium.” To some extent, this situation can be groupings, and altered orientation of groups may understood since many insights into mechanisms occur. This will carry with it the possibility of of the allergic responses to a whole range of dif- antigenic and allergenic changes as well as the ferent drugs have occurred only in recent years. possibility of an additional allergenic contribu- Many of the well-established advances, even tion by the carrier molecule. Widely and heavily simply the names of important culprit allergenic used drugs such as β-lactams, neuromuscular drugs (e.g., cephalosporins, anesthetic agents, blockers, and many other commonly used oral, and chimeric monoclonal antibodies), let alone injected, and topical medicaments are already details of their allergenic properties, have yet to being formulated and/or marketed as drug-carfi nd their way into the general immunological rier complexes. With the cyclodextrins alone, and even some of the allergological literature. there were more than 45 drug-cyclodextrin Ultimately, improvements in diagnosis and treat- inclusion complexes approved in 2010. ment should result from increased understanding Allergenic modifi cation of the neuromuscular of the mechanisms underlying immediate and blocker rocuronium by a chemically modifi ed delayed reactions and some nonimmune drug γ-cyclodextrin used to sequester the drug from intolerances. In the process, fundamental insights the neuromuscular junction to reverse neuro- into the recognition and handling by the immune muscular blockade is discussed in Sect. 7.4.6 . system of small (<1 kDa) usually synthesized, The possibility of change in the allergic recogni- nonprotein, non-carbohydrate molecules are tion of a known allergenic drug as well as appear- likely to be gained and to enter the mainstream ance of allergenic activity in drugs previously immunological literature.

[email protected] 12 1 Adverse Reactions to Drugs and Drug Allergy: Scope of This Book

1.5.2 The Need for a Single Text chlorhexidine; anesthetic agents and drugs Book on Drug Allergy used in surgery; NSAIDs; opioid analgesics; iodinated and gadolinium-based contrast At the research level, drug allergy is a mixture of media; therapeutic monoclonal antibodies and chemistry, immunology, pharmacology, and bio- some recombinant biologics used for therapy; chemistry with increasing inputs in the labora- corticosteroids; a wide variety of drugs with tory from immunochemistry, cell biology, anti-neoplastic properties used in cancer ther- molecular biology, and molecular genetics. apy; and proton pump inhibitors. Although in the early development of research on • Where appropriate, concentrations of drugs the immune response to chemicals, animal mod- for skin and challenge testing, photographs of els were instrumental in the discovery and subse- examples of important drug-induced cutane- quent study of contact sensitivity and delayed ous and cutaneous/systemic reactions, struc- type hypersensitivity, allergy research has never tural information, and relevant metabolic been far removed from human patients and the pathways are presented. Treatments of drug- clinical application of its fi ndings. Reports of induced reactions are beyond the scope of this advances in the understanding of diagnosis and book and are not considered. treatment of allergic reactions to drugs are likely • At the conclusion of each chapter a “Further to be found in journals in any of the above- reading” list of seminal/authoritative/innova- mentioned disciplines as well as journals devoted tive publications is provided. In some cases, to allergy and the general medical literature. With an early seminal work, a particularly informa- signifi cant recent advances in our understanding tive and/or comprehensive review, or an of the mechanisms underlying the induction, advanced treatment of a subject is included. development, and manifestations of the various allergic and other drug sensitivities, now seems to be an appropriate time to review, in one vol- Summary ume, the research and clinical developments made since the fi rst relevant publications • With approximately 5 % of patients develop- appeared just over a century ago. In other words, ing ADRs during drug therapy and up to 10 % there is a need for an integrated text that treats the reacting to drugs in hospitals, ADRs are a sig- subject in its own right. In doing this, we have nifi cant health problem. pursued a straightforward approach that can be • In what is essentially a pharmacologically summarized as follows: based classifi cation of ADRs, six or seven differ- • Classifi cation and description of the various ent categories are recognized. Approximately drug-induced hypersensitivity and sensitivity 80 % of reactions are predictable, dose depen- responses. dent, and resolve upon withdrawal of drug. • Presentation of current knowledge of the Unpredictable reactions are generally unre- mechanisms underlying the various systemic lated to the drug’s pharmacological action, and cutaneous drug reactions including mech- are independent of dose, and usually resolve anisms of immediate, late, and delayed reac- when treatment stops. tions, type II and type III hypersensitivities, • Unpredicted and dose-independent drug reac- and some important drug-induced sensitivities tions, known as type B reactions, include lacking an immune basis. hypersensitivity responses. • A comprehensive review of diagnostic • “Allergy” is a much misused term. The well methods. entrenched and widely accepted term “hyper- • In depth discussions of those groups of drugs sensitivity” is reserved here for immune-based most frequently implicated in reactions—the reactions. Some nonimmune reactions such as β-lactams; other antibacterials including adverse reactions to NSAIDs and contrast media antibiotics, sulfonamides, trimethoprim, and are referred to as sensitivities or intolerances.

[email protected] Further Reading 13

• The foundations of modern allergy research signaling pathways, mechanisms of action of and practice were laid at the end of the nine- a variety of soluble mediators, and the genetic teenth and fi rst quarter of the twentieth centu- bases of many adverse reactions to drug ries and extended by the work of Landsteiner molecules and collaborators with their demonstration of • Drug allergy has been neglected in text books the sensitization of laboratory animals and of immunology and the time is right for the humans with simple reactive chemicals. subject to be presented in detail and with wide • Previous exposure is not always a precondi- coverage of its many aspects. tion for allergic sensitization. There are numerous examples of reactions to poorly reactive drugs where no protein binding to the Further Reading parent drug, metabolites, or degradation products can be demonstrated. Baldo BA, McDonnell NJ, Pham NH. Drug-specifi c cyclodextrans with emphasis on sugammadex, the • Type II antibody-mediated cytotoxic reactions neuromuscular blocker rocuronium and perioperative and type III reactions mediated by soluble anaphylaxis: implications for drug allergy. Clin Exp immune complexes are seen as true hypersen- Allergy. 2011;41:1663–78. sitivity reactions. Barone C, Mousa SS, Mousa SA. Pharmacogenomics in cardiovascular disorders: steps in approaching per- • Risk factors for drug allergies can be divided sonalized medicine in cardiovascular medicine. into those that are patient-related or drug- Pharmgenomics Pers Med. 2009;2:59–67. related. The former group covers the infl uences Bharadwaj M, Illing P, Theodossis E, et al. Drug hyper- of age, sex, current diseases, previous exposure, sensitivity and human leukocyte antigens of the major histocompatibility complex. Annu Rev Pharmacol and genetic factors; the latter, cross-sensitivity/ Toxicol. 2012;52:401–31. reactions of drugs, nature of the drug, the degree Daly AK. Using genome-wide association studies to iden- of exposure, and route of administration. tify genes important in serious adverse drug reactions. • The application of genomics promises to bet- Annu Rev Pharmacol Toxicol. 2012;52:21–35. Edwards IR, Aronson JK. Adverse drug reactions: defi ni- ter explain and ultimately reduce some ADRs. tions, diagnosis, and management. Lancet. 2000;356: Results so far have demonstrated the associa- 1255–9. tion in Han Chinese of carbamazepine- Landsteiner K. The specifi city of serological reactions. induced Stevens–Johnson syndrome with the New York: Dover; 1962. Landsteiner K, Jacobs J. Studies on the sensitization of HLA-B*15:02 allele and the association of animals with simple chemical compounds. J Exp Med. abacavir hypersensitivity in AHS with 1935;61:643–56. HLA-B*57:01. Landsteiner K, Jacobs J. Studies on the sensitization of • As a result of the latter fi ndings, genetic animals with simple chemical compounds II. J Exp Med. 1936;64:625–39. screening is now part of primary HIV clinical Phillips E, Mallal S. Successful translation of pharmaco- practice. genetics into the clinic: the abacavir example. Mol • It seems likely that multiple rather than single Diagn Ther. 2009;13:1–9. genes are involved in ADRs. Phillips KA, Veenstra DL, Oren E, et al. Potential role of pharmacogenomics in reducing adverse drug reac- • Efforts to improve drug delivery may lead to tions. A systematic review. JAMA. 2001;286:2270–9. loss, or enhancement, of a drug’s former aller- Rawlins MD, Thomas SHL. Mechanisms of adverse drug genicity, or the appearance of allergenicity in reactions. In: Davies DM, Ferner ER, de Glanville H, a drug previously not implicated in allergic editors. Davies’ textbook of adverse drug reactions. 5th ed. London: Chapman and Hall Medical; 1998. reactions. This will need to be recognized in p. 40–64. preclinical assessments of drug safety. Talbot J, Aronson JK, editors. Stephen’s detection and • Studies of drug allergies have the potential of evaluation of adverse drug reactions. 6th ed. Chichester: providing insights into some important areas Wiley-Blackwell; 2012. Wilke RA, Lin DW, Roden DM, et al. Identifying genetic of biomedical investigation including cell risk factors for serious adverse drug reactions: current recognition and interactive processes, rela- progress and challenges. Nat Rev Drug Discov. tionships between receptors and effector and 2007;6:904–16.

[email protected] Classiﬁ cation and Descriptions of Allergic Reactions to Drugs 2

Abstract Four types of hypersensitivities may be distinguished. Type I, or immediate hypersensitivity, occurs within about 30 min, is IgE antibody-mediated, and the allergic signs and symptoms are triggered by cross-linking of mast cell-bound IgE which leads to mast cell degranulation and release of inﬂ ammatory mediators. Drugs well known to cause type I reactions include β-lactams, neuromuscular blockers, and some NSAIDs. Anaphylactoid reactions may mimic the signs and symptoms of anaphylaxis but, unlike the latter reactions, anaphylactoid reactions are not immune-mediated. Clinical manifestations of anaphylaxis include erythema, urticaria, angioedema, bronchospasm, and cardiovascular collapse. Urticaria is often associated with angioedema and anaphylaxis. ACE inhibitors are responsible for one in six hospital admissions for angioedema. Types II and III hypersensitivities are known as antibody-dependent cytotoxic and immune complex-mediated hypersensitivities, respectively. Examples of drug-induced type II reactions are hemolytic anemia, thrombocytopenia, and granulocytopenia. A serum sickness-like reaction is the prototype type III drug hypersensitivity. Type IV drug hypersensitivities are mediated by antigen-speciﬁ c T cells. Reactions occur 48–72 h after antigen exposure and are therefore referred to as delayed. Examples of delayed cutaneous reactions include allergic contact dermatitis, psoriasis, FDE, AGEP, DRESS, SJS, and TEN.

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 15 Relationships, DOI 10.1007/978-1-4614-7261-2_2, © Springer Science+Business Media, LLC 2013

[email protected] 16 2 Classiﬁ cation and Descriptions of Allergic Reactions to Drugs

their classifi cation of hypersensitivity reactions 2.1 Hypersensitivity: The Early based on the immune mechanisms underlying Years—From Koch to Gell the different reactions, that is, the latency of and Coombs reactions, humoral or cellular involvement, complement involvement, and pathophysiological The phenomena that constitute the basis of consequences for the host. hypersensitivity reactions were discovered, and studies initiated, in the approximately 20-year period between Robert Koch’s demonstration of 2.2 Classifi cation a delayed hypersensitivity reaction to the tubercle of Hypersensitivity Reactions bacillus in 1882 and the demonstrations of anaphylactic, Arthus, and serum sickness reactions A number of arguments have been voiced over the in the fi rst decade of the twentieth century. years against the Gell and Coombs classifi cation Having discovered the tubercle bacillus, Koch and published criticisms include its alleged too showed that the delayed infl ammatory response narrow focus on the deleterious consequences to to an intradermal injection of the organism could the host, that some antigens such as drugs do not indicate previous exposure to Mycobacterium fi t well into the categories, and that the classifi ca- tuberculosis in an asymptomatic person. This tion scheme should be simplifi ed to pseudo- response became known as the tuberculin reac- allergic, antibody-mediated, and cell-mediated tion and, as the Mantoux test, is recognized today reactions. More recently, subdivisions have been as the classical and best known example of a suggested for type IV reactions. We believe that delayed hypersensitivity reaction. Continuing the existing classifi cation has defi ciencies but, studies initiated with P. Portier in 1902, Charles with mechanism-based categories, it remains, Richet by 1907 had shown that numerous pro- overall, the simplest, most valid, and most logical teins in small doses could provoke not “phylaxis” way of distinguishing the host’s immune sensitivi- or protection, but “anti-protection” or what he ties. Four types of hypersensitivity reactions, des- termed “anaphylaxis,” and this state could be ignated types I, II, III, and IV, are distinguished. passively transferred to a normal animal by serum. In the fi rst demonstration of the reaction which came to bear his name, N.M. Arthus in 2.2.1 Type I Hypersensitivity 1903 produced erythematous and edematous reactions in rabbits after repeated injections of Type I hypersensitivity is also known as immedi- horse serum, and in 1906, von Pirquet and Bėla ate, or sometimes anaphylactic, hypersensitivity. Schick demonstrated the immune complex- Responses usually occur within 30–60 min but mediated serum sickness reaction (see Sects. 2.2.3 can be extremely quick (within minutes) and dra- and 3.8 ) so named because it sometimes occurred matic as in anaphylaxis. In some cases a late following the administration of antisera prepared onset reaction may occur about 3 or 4 h after in horses (e.g., anti-pneumococcal antisera). allergen exposure and the immediate reaction. Despite Landsteiner’s initial conviction that This late phase reaction generally peaks at about cell- bound antibody was the cause of contact 6–12 h and subsides at about 24 h (see Sect. 3.3 ). sensitivity, Chase’s experiments in 1941–1942 Immediate reactions can affect a single organ transferring “sticky,” unclarifi ed peritoneal exudate such as the skin (urticaria, eczema), eyes (con- from guinea pigs sensitized by hapten–stromata junctivitis), nasopharynx (allergic rhinitis), conjugate with Freund’s complete adjuvant led to mucosa of mouth/throat/tongue (angioedema), the discovery that viable cells from the perito- bronchopulmonary tissue (asthma), and gastroin- neum, lymph nodes, and spleen-mediated contact testinal tract (gastroenteritis) or multiple organs sensitivity and tuberculin reactivity. It was then a (anaphylaxis), causing symptoms ranging from further 21 years before Gell and Coombs produced minor itching and infl ammation to death.

[email protected] 2.2 Classiﬁ cation of Hypersensitivity Reactions 17

Immediate hypersensitivity is mediated by IgE involving the release of potent allergic and antibodies interacting with mast cells and baso- infl ammatory mediators from mast cells and phils (Sect. 3.2) and with eosinophils, platelets, basophils and producing at least some of the and neutrophils amplifying the response. IgE signs and symptoms of severe immediate reac- antibodies bind to their complementary FcεRI tions such as cardiovascular symptoms (tachy- receptors on mast cells via their Cε3 region leav- cardia, hypotension, cardiovascular collapse), ing both antibody combining sites free to interact respiratory involvement (dyspnea, broncho- with the complementary allergenic determinants. spasm, wheeze), gastrointestinal symptoms (nau- This interaction causes cross-linking of the cell- sea, vomiting, abdominal pain), and cutaneous bound antibodies leading to degranulation of the manifestations (urticaria, angioedema, erythema, anchoring mast cells and release of the variety of pruritus). With the knowledge that some agents preformed, and then newly formed, mediators of induce anaphylaxis via an IgG antibody-mediated infl ammation and hypersensitivity. mechanism, for example, as occurs in some Drugs well known for causing type I allergic responses to dextrans (and an IgG-mediated reactions include penicillins, cephalosporins, qui- pathway to anaphylaxis is known in some labora- nolones, chlorhexidine, neuromuscular blocking tory animals such as mice), the wording “IgE drugs, some nonsteroidal anti-infl ammatory drugs antibody-mediated” in the defi nition above is (NSAIDs) such as pyrazolones, trimethoprim, more correctly replaced by, simply, “antibody- sulfamethoxazole, proton pump inhibitors, heparin, mediated.” The term anaphylactoid is used for insulin, L-asparaginase, etanercept, and chimeric reactions that may show clinically similar or even human–animal monoclonal antibodies used for identical signs and symptoms but where no therapy, but it is uncommon to fi nd a drug at least immune-mediated mechanism can be shown, for moderately used that has not provoked an anaphy- example, reactions caused by direct mast cell lactic reaction in at least one rare individual. degranulation induced by drugs such as vancomycin, opioids, or contrast media. 2.2.1.1 Anaphylaxis to Drugs In 1998 in the USA, a Joint Task Force on It seems that every new drug administered to Practice Parameters for Allergy and Immunology humans has the potential to provoke an anaphy- defi ned anaphylaxis as an “immediate systemic lactic response, and the likelihood of such a reaction caused by rapid, IgE-mediated immune response therefore increases with increased release of potent mediators from tissue mast cells administration. Unfortunately, while the signs and peripheral basophils.” Anaphylactoid reac- and symptoms of what is commonly termed tions were seen as mechanistically different and “anaphylactic shock” seem clear enough, inci- said to “mimic signs and symptoms of anaphy- dences of anaphylaxis in different countries and laxis, but are caused by non-IgE-mediated release to different drugs, as well as the terminology of potent mediators from mast cells and baso- used for reactions is often far from consistent. phils.” By 2004 when the National Institute of Allergy and Infectious Diseases (NIAID) and the 2.2.1.1.1 Terminology Food Allergy and Anaphylaxis Network (FAAN) For many years, two terms, “anaphylaxis” and cosponsored a symposium on the defi nition and “anaphylactoid,” have been used to describe rela- management of anaphylaxis, the existing mecha- tively rare reactions that have features commonly nistic defi nition of anaphylaxis was judged to be associated with severe immediate, often life- “of marginal utility” to the physician and other threatening, allergic reactions. These two terms health care personnel when faced with “the vari- are distinguished by the underlying mechanisms able constellation of signs and symptoms of this of the reactions. The term anaphylaxis is used by disorder.” In a second symposium in 2005 the many for an immune IgE antibody-mediated, NIAID and FAAN brought together allergists, systemic immediate type I hypersensitivity reac- immunologists, pediatricians, emergency and intention, often occurring within seconds or minutes, sive care physicians, internists, and pathologists

[email protected] 18 2 Classifi cation and Descriptions of Allergic Reactions to Drugs together with representatives from the profes- think more mechanistically of their art. If ana- sional bodies in Canada, Europe, and Australia to phylaxis is defi ned in terms of being a systemic, (among other aims) work toward a universally rapidly proceeding, immune-based immediate accepted defi nition of anaphylaxis and establish reaction involving potent mediators released clinical criteria to accurately identify cases of from mast cells and basophils and producing a anaphylaxis. The defi nition of anaphylaxis that range of clearly defi ned profound clinical effects, emerged from this gathering was: “Anaphylaxis the addition of descriptors such as “allergic,” is a severe, potentially fatal, systemic allergic “IgE-mediated allergic,” and “nonallergic” to the reaction that occurs suddenly after contact with word anaphylaxis becomes completely unneces- an allergy-causing substance.” For a defi nition sary. Anaphylaxis has, over many years, come to that would be useful to both the medical and lay be known as an allergic reaction mediated by communities, anaphylaxis was defi ned as “a seri- IgE. Hence, the terms “allergic anaphylaxis,” ous allergic reaction that is rapid in onset and “IgE- mediated allergic anaphylaxis” and “nonal- may cause death.” For the researcher, and those lergic anaphylaxis” each have added words that practicing clinical medicine but perhaps not for are redundant or are a contradiction in terms. It the layman, it is hard to see how these defi nitions then follows that “anaphylactoid” is a suitable are an improvement or more useful than the for- and convenient term to cover those immediate mer ones. systemic reactions that mimic anaphylaxis but As mentioned in Chap. 1, in 2003, the are not immune-mediated and, with the stipula- Nomenclature Review Committee of the World tions and distinctions outlined above in mind, Allergy Organization (WAO) defi ned anaphy- reactions will be referred to either as anaphylac- laxis (which they referred to as an “umbrella tic or anaphylactoid throughout this book. term”) as “a severe, life-threatening generalized or systemic hypersensitivity reaction.” It was fur- 2.2.1.1.2 Incidence of Anaphylaxis ther stated that “the term allergic anaphylaxis The incidence of all causes of anaphylaxis in should be used when the reaction is mediated by Western countries is estimated to be from about an immunological mechanism, e.g., IgE, IgG, 8 to 50 per 100,000 persons per year with 3–4 % and immune complex complement related” and, hospitalized and a lifetime prevalence of “an anaphylactic reaction mediated by IgE anti- 0.05–2 %. In one retrospective Danish study of bodies, such as peanut-induced food anaphylaxis, anaphylaxis over a 13-year period outside hospi- may be referred to as IgE-mediated allergic ana- tal in a catchment area of 48,000 subjects, an phylaxis.” Lastly, “anaphylaxis from whatever incidence of 3.2 cases per 100,000 persons per nonimmunologic cause should be referred to as year was found. Of the 20 cases of anaphylaxis nonallergic anaphylaxis.” As with the WAO sug- identifi ed, seven were provoked by oral penicil- gested changes to the defi nition of hypersensitiv- lin and three by oral aspirin, indicating that ana- ity and introduction of the term “non-allergic phylaxis to penicillin in the non-hospital hypersensitivity,” (see Chap. 1, Sect. 1.1.2 ), the environment was more common than thought. proposed defi nition of anaphylaxis is not only A retrospective population-based cohort study of unnecessary but inadequate and the distinction of 1,255 US residents in one county over a 5-year anaphylaxis into “IgE- mediated allergic” and period revealed an incidence of anaphylaxis of “nonallergic” is needlessly complicating, cum- 30 per 100,000 persons per year with an average bersome, and redundant. To move away from a annual incidence rate of 21 per 100,000 person- mechanistic defi nition in defi ning anaphylaxis years. Drugs, along with foods and insect stings, for professionally trained workers in medicine were the main causes. In a prospective study of and science in an age when scientifi c progress is 432 Australian patients, medication was the infl uencing clinical medicine to an unprecedented cause in 8.3 % of cases. Minimum occurrence extent, seems to be a step backward. There seems and incidence of new cases were estimated to be to be much to gain by encouraging physicians to 12.6 and 9.9 episodes per 100,000 patient-years,

[email protected] 2.2 Classifi cation of Hypersensitivity Reactions 19 respectively. An analysis of anaphylaxis admis- induced anaphylaxis are the fi gures for drugs sions to UK critical care units between 2005 and used perioperatively. Published incidences of 2009 revealed 1,269 adult and 81 pediatric anaphylactic and anaphylactoid reactions include admissions representing 0.3 % of admissions to 1 in 5,000–13,000 for Australia, 1 in 1,250–5,000 adult units and 0.1 % of admissions to pediatric for New Zealand, 1 in 3,500 for England, 1 in units. The data showed that hospital admission 5,000 in Thailand, and 1 in 4,600 in France. By rates for anaphylaxis have increased sevenfold in far the biggest contributor to anaphylaxis alone the last two decades and the absolute numbers of (that is, with anaphylactoid reactions not both adults and children rose year-on-year. The involved) during anesthesia is the group of neuro- authors remarked that drug-triggered reactions muscular blocking drugs. Reported incidences to are more common in older people and concluded these drugs are 1 in 10,000–20,000 Australia, 1 that each UK critical care unit is likely to see at in 5,500 France, 1 in 5,200 Norway, 1 in 10,263 least one anaphylaxis case per year. Spain, 1 in 5,500 Thailand and approximately A large proportion of anaphylactic reactions is 500 cases per year in the UK. Of the drugs impli- due to drugs. In hospitalized patients the preva- cated in anaphylaxis, the most complete and reli- lence is said to be 3 in 10,000 with deaths able data on incidences of reactions are those occurring in 3–9 % of patients. The overall inci- obtained from studies of perioperative anaphy- dence and the mortality of anaphylaxis induced laxis in Australia and France. A breakdown of the by drugs are not known but fi gures are available main drugs responsible for anaphylaxis during for some individual drugs or groups of drugs in anesthesia in Australia and France shows the fol- some localities. Most data over the years for inci- lowing incidences of reactions (as percentages, dences of anaphylaxis to a single drug (or group Australian fi gures fi rst): neuromuscular blockers of drugs) have been for penicillins with published 61.9, 58.1; induction agents 10.4, 2.3; antibiotics estimates of approximately one reaction for every 8.6, 12.9; colloids 4.6, 3.4; opioids 2.6, 1.7 (see 10,000 prescriptions and 15–40 reactions per also Table 7.1 ). 100,000 persons. During the 1960s and 1970s penicillins were often claimed to be the most 2.2.1.1.3 Clinical Features of Drug- common cause of drug-induced anaphylaxis in Induced Anaphylactic Reactions the US and that may still be the case today. In the Dutch study discussed above in which 43 Reactions to contrast media and blood-volume different drugs were implicated in causing at replacement fl uids have been reported in one of least one anaphylactic reaction, the main symp- every 600 and 400 persons, respectively, receiv- toms seen in the patients admitted to hospitals ing the drugs. More detail of incidences of ana- were (in decreasing order of occurrence) ery- phylaxis to other drugs are to be found in the thema, angioedema, hypotension, bronchospasm, chapters on the different drug groups. In a pruritus, urticaria, nausea/vomiting, tachycardia, population-based case–cohort study in The loss of consciousness, diarrhea, upper airways Netherlands, a drug was found to be the causative symptoms, conjunctivitis, laryngeal edema, agent in 107 of 252 cases of suspected anaphy- abdominal pain, and bradycardia. Erythema was laxis classifi ed as “causal relationship certain” or seen in 57 % of patients, angioedema in 51 %, “causal relationship probable.” Of the 107 cases, hypotension in 36 %, bronchospasm and pruritus 19 % were caused by the NSAID glafenine, 11 % in 35 %, urticaria in 31 %, tachycardia in 19 %, by amoxicillin, 7 % for each of diclofenac and and bradycardia in 4 %. For drug-induced ana- acetaminophen and 6 % for propyphenazone. In phylactic and anaphylactoid reactions, however, fact, at least 42.6 % of the 107 cases of possible/ it is the perioperative data accumulated over probable cases of anaphylaxis were due to a many years by Fisher and Baldo in Australia and NSAID (27 miscellaneous drugs each causing by the Laxenaire and Mertes groups in France one reaction were not named). Perhaps the most (see Fig. 7.2 ) that is the most informative. As reliable data available on incidences of drug- emphasized by the former workers, the list of

[email protected] 20 2 Classiﬁ cation and Descriptions of Allergic Reactions to Drugs

Table 2.1 Clinical features in 315 patients with life-threatening anaphylaxis to anesthetic drugs Total Worst feature Sole feature Clinical features Number % Number % Number % Cardiovascular collapse 283 89.8 251 79.7 33 10.5 Bronchospasm Severe 52 16.5 52 16.5 10 3.2 Transient 50 15.9 Angioedema 73 23.2 4 1.3 4 1.3 Urticaria 43 13.7 Rash 42 13.3 Erythema 151 47.9 Gastrointestinal symptoms 30 9.5 Pulmonary edema 11 3.5 3 1.0 1 0.3 Generalized edema 15 4.8 Other major common features: vomiting, diarrhea, laryngeal edema Uncommon major features: cardiac failure, disseminated intravascular coagulation, hemoptysis, melena Minor features: rash, fl ush, rhinitis, cough, lacrimation, urticaria, pruritus, aura, conjunctivitis Late features: headache, thromboembolism, edema, wound hematoma, vaginal discharge Data adapted from Fisher M, Baldo BA. Eur J Anaesthesiol 1994;11:263 and Med J Aust 1988;149:43 complete signs and symptoms (Table 2.1 ) does least 12 h. Gastrointestinal symptoms include not appear in every patient; cardiovascular col- severe abdominal pain, vomiting, diarrhea, lapse is the most common symptom and usually hematemesis, and melena and last up to 6 h. Non- the worst; in addition to cardiovascular collapse cardiogenic pulmonary edema, which occasionally there is usually at least one other sign; asthmatic occurs as a single clinical feature of anaphylaxis patients who experience anaphylaxis usually get and is a common postmortem fi nding in resultant bronchospasm; and a transient bronchospasm or deaths, has been reported as a sole delayed reaction diffi culty in infl ating the lungs is often seen as the to protamine after cardiac bypass surgery. fi rst sign of a reaction. Note that when the problem of lung infl ation persists, it is often the most 2.2.1.1.4 Clinical Grading System diffi cult feature to reverse. Cardiovascular col- for Anaphylaxis lapse, the most common life-threatening feature, Defi nitions of anaphylaxis vary, sometimes mak- is due to vasodilation and the pooling of periph- ing it diffi cult to interpret and compare clinical eral blood which reduces the venous return and fi ndings. Classifi cation of anaphylactic reactions the cardiac output. Whether the heart is a target on the bases of clinical features observed and their organ in anaphylaxis in humans as it is in some severity is obviously needed, but there has been other animals is still not clear. Cardiac failure no uniformity or agreement on the relevance and occurs in anaphylactic patients with cardiac importance of the parameters to be included in disease but rarely in patients with normal cardiac any grading system. Some grading systems are function. Table 2.1 indicates that when broncho- simple descriptions of common and/or important spasm is severe, it is usually the worst feature and symptoms while others are based on statistical the sole feature in about 20 % of the cases. analyses of individual reaction features, sums of Bronchospasm may be critical since the high allotted scores, the appearance of “two or more” pressures needed for infl ation reduce venous clinical features, or cardiovascular compromise. return and may increase ventricular compliance. From case records of over 1,100 acute generalized Angioedema, which involves the head, neck, and hypersensitivity reactions, logistic regression upper airways, usually progresses slowly and it is analyses of associations between reaction features therefore prudent to observe the patient for at and hypotension and hypoxia were used to con-

[email protected] 2.2 Classiﬁ cation of Hypersensitivity Reactions 21

Table 2.2 Grading system for hypersensitivity reactions including anaphylaxis Grade Broad clinical features Deﬁ ning symptoms and signs 1 Cutaneous and subcutaneous only Generalized erythema, periorbital edema, urticaria, MILD or angioedema 2 Cardiovascular, respiratory, or Dyspnea, stridor, wheeze, nausea, vomiting, dizziness, MODERATE gastrointestinal involvement diaphoresis, chest or throat tightness, or abdominal pain

3 Hypoxia, hypotension, or Cyanosis or SpO2 ≤ 92 % at any stage, hypotension SEVERE neurologic compromise (systolic BP < 90 mmHg in adults), confusion, collapse, loss of consciousness, or incontinence Adapted from Brown SGA. Clinical features and severity grading of anaphylaxis. J Allergy Clin Immunol 2004;114:317 with kind permission from Elsevier Limited struct a grading system suitable for defi ning reaction severity in clinical and research settings. Three grades that correlated with epinephrine usage, mild, moderate, and severe (Table 2.2 ), were discerned from the clinical features of confusion, collapse, unconsciousness, incontinence, diaphoresis, vomiting, presyncope, dyspnea, stridor, wheeze, and nausea, all of which were associated with documented hypotension and hypoxia. The clinical features in the moderate and severe grades fi t in with a defi nition of anaphylaxis. A major difference between this and other grading systems is the identifi ed importance of gastrointestinal symptoms. A possible limitation of this study and its conclusions is that clinical assessments were undertaken by emergency medicine Fig. 2.1 A case of severe generalized chronic urticaria clinicians rather than allergists, so confi rmatory and nonlife-threatening angioedema unresponsive to anti- skin and IgE antibody tests are lacking. histamines. From Fox R, Lieberman P, Blaiss M. Centralized urticarial and angioedema and angioedema of the face. Atlas of allergic diseases; 2002;IS:08. With kind 2.2.1.2 Urticaria and Angioedema permission from Springer Science+Business Media B.V Urticaria or hives, the second most common cutaneous reaction induced by drugs after exanthem- ger than 24 h and which are painful, burning, or atous reactions, occurs often in association with leave bruising and/or pigmentation changes may angioedema, in cases of anaphylaxis and in serum indicate urticarial vasculitis. Acute urticaria that sickness. Virtually any drug can cause urticaria. is more common in children, may appear early Hives are generally raised, circumscribed, ery- after exposure (perhaps minutes) and can last thematous papules and plaques (Fig. 2.1 ) with a several weeks; chronic urticaria, more common central area of pallor, often round in shape and of in adults, occurs in episodes that last longer than variable size. Erythematous areas may be 6 weeks. Drugs are only infrequently implicated smooth surfaced, patchy, or confl uent and gener- in cases of chronic urticaria and, in fact, no exter- alized (Fig. 2.2 ). Outbreaks that may occur any- nal agent or disease state has been implicated in where on the skin are extremely pruritic and 80–90 % of patients with chronic urticaria. The transient, usually appearing within 36 h of drug incidence of chronic urticaria is said to be as exposure and resolving without sequelae within high as 1 % in the USA and several other coun- 24 h. On rechallenge with drug, lesions may tries, but the fi gure is likely to be closer to about appear within minutes. Lesions that persist lon- 0.1–0.3 %. The mechanisms involved in the

[email protected] 22 2 Classiﬁ cation and Descriptions of Allergic Reactions to Drugs

Fig. 2.2 An example of generalized urticaria (hives) showing smooth, erythematous, and pruritic conﬂ uent papules and plaques. Aspirin and other nonsteroidal anti- inﬂ ammatory drugs are a frequent cause of hives but virtually any drug can precipitate the condition. From Anderson J, Lieberman P, Blaiss M. Allergic drug reactions. Atlas of allergic diseases; 2002;IS;26. With kind permission from Springer Science+Business Media B.V

spectrum of urticarial reactions are reviewed in Sect. 3.2.8 . Drugs implicated in reactions include those that provoke type I IgE-mediated reactions β (e.g., -lactams, other antibiotics, antimicrobials Fig. 2.3 Angioedema of the face showing non-pruritic such as sulfonamides and trimethoprim, neuro- swelling of the cutaneous tissues with some erythema. muscular blockers, pyrazolones); direct mast cell Angioedema persists longer than urticaria due to the accu- degranulation (e.g., opioids, contrast media, van- mulated fl uid in the tissues. From Fox R, Lieberman P, Blaiss M. Centralized urticarial and angioedema and comycin, quinine, pentamidine, atropine); drugs angioedema of the face. Atlas of allergic diseases; that promote or exacerbate urticaria due to their 2002;IS:08. With kind permission from Springer pharmacological effects on metabolic pathways Science+Business Media B.V (angiotensin-converting enzyme [ACE] inhibitors, e.g., captopril, enalopril, lisinopril; NSAIDs, e.g., leakage of fl uid which produces edema of the sub- aspirin, indomethacin, ibuprofen); drugs involved mucosal tissues, deep dermis, and subcutaneous in type III immune complex formation as in serum tissues. As a result of compression of nerves, sickness (e.g., amoxicillin, cefaclor, ciprofl oxacin, patients may experience decreased sensation in monoclonal antibodies); excipients, preservatives, the affected areas. In severe reactions involving coloring agents, antioxidants (e.g., benzoic acid, the airways, stridor, gasping, and wheezing may sulfi tes, tartrazine, butylated hydroxytoluene). indicate the need for tracheal intubation. As with Note that evidence for adverse effects, including urticaria, angioedema is often an IgE the induction of urticaria, is often lacking for antibody-mediated reaction to a drug and one of implicated agents in the latter group. its main causes is the ACE inhibitor group of Angioedema (Quincke’s edema), which is seen drugs which induce reactions with an incidence of less often than urticaria but often accompanies it, 0.1–0.5 %. For black Americans and Afro- is a vascular reaction resulting in swelling of the Caribbeans this incidence is three times higher. face (Fig. 2.3) around the mouth, in the mucosa of ACE inhibitors are the responsible agents for the mouth, throat and tongue, eyelids, genitalia, approximately one in six patients admitted to hos- and occasionally involving the hands and else- pital for treatment of angioedema. It is thought the where (Fig. 2.1 ). Swelling, which can be itchy and drugs increase bradykinin levels in peripheral tis- painful, is the result of increased permeability and sues, and this leads to the rapid fl uid accumulation

[email protected] 2.2 Classifi cation of Hypersensitivity Reactions 23 seen in angioedema (Sect. 3.2.8.5.2 ). Angioedema ifi city) appears to be involved. Drug- induced to ACE inhibitors usually occurs within the fi rst thrombocytopenia with quinine, quinidine, pro- week of treatment and the drug should be with- pylthiouracil, gold salts, acetaminophen, vanco- drawn immediately in the patients who experience mycin, and sulfonamides in immune complexes a reaction. Aspirin and other NSAIDs are other adsorbed onto platelet membranes is well docu- common causes of angioedema generally involv- mented. Other mechanisms are also operative in ing the face. Responses to the NSAIDs may be immune-mediated thrombocytopenia. Cytotoxic complex with mixed cutaneous and respiratory antibodies to pyrazolone drugs, thiouracil, sulfon- symptoms (Sect. 9.5.2 ). amides, anticonvulsives, and phenothiazines may Unlike the other types of hypersensitivity produce granulocytopenia by the destruction of caused by antibodies, namely types II and III, peripheral neutrophils. For more detail, including only a proportion of the population, the so-called mechanisms, of each of these reactions, the reader atopics, have a predisposition to developing a is referred to Chap. 3 , Sect. 3.7 . type I hypersensitivity reaction when exposed to the allergen in question. Atopy may have a genetic component but type II and III reactions 2.2.3 Type III Hypersensitivity like, for example, penicillin-induced hemolytic anemia and serum sickness, may occur in all indi- Type III hypersensitivity, also called immune viduals, and this may result without a prior sensi- complex hypersensitivity, is mediated by soluble tization phase. immune complexes of antigen with antibodies mostly of the IgG class but sometimes IgM. Deposition of immune complexes in tissues 2.2.2 Type II Hypersensitivity results in a tissue reaction initiated by complement activation that may lead to mast cell degran- Type II hypersensitivity, also known as cytotoxic ulation, leukocyte chemotaxis, and infl ammation hypersensitivity and antibody-dependent cytotoxi- induced by the cell infl ux. After exposure, reac- city, causes reactions that are serious and poten- tions may develop over a period of about 3–10 h tially life-threatening. A number of different against antigens that can be endogenous as in organs and tissues may be affected with the DNA/anti-DNA/complement deposits in the kidneys involvement of multiple underlying mechanisms. of patients with systemic lupus erythematosus or, The antigens involved are often endogenous but more often, exogenous as in the Arthus reaction in drugs can attach to cell membranes provoking rabbits to intradermal injection of soluble antigen drug-induced immune hemolytic anemia, throm- or intrapulmonary Arthus-like reactions in humans bocytopenia, and granulocytopenia, all examples to inhaled antigen associated with farmer’s lung of type II reactions. Reaction times can range and extrinsic allergic alveolitis. Other exogenous from minutes to hours; IgM or IgG antibodies antigens eliciting type III responses include those mediate the reactions and in antibody-dependent from organisms such as fi larial worms, dengue cellular cytotoxicity, target cells coated with anti- virus, and microbial antigens abruptly released body are killed by a non-phagocytic process following chemotherapy in patients with high involving Fc receptor-bound leukocytes (NK antibody levels. More importantly for our pur- cells, monocytes, neutrophils, and eosinophils). poses are type III reactions with drug involvement Drug-induced hemolytic anemias with IgG anti- where examples include erythema nodosum lep- bodies and complement-mediated cytotoxicity (or rosum in the skin of leprosy patients treated with an autoantibody) implicated may occur after treat- dapsone, the Jarisch–Herxheimer reaction in ment with penicillins, quinidine, α-methyldopa, syphilitic patients treated with penicillins, and some cephalosporins. In the penicillin- and serum sickness-like reactions caused by a induced condition, an atypical anti-penicillin number of different drugs including penicillins, antibody (perhaps with drug-surface protein spec- cephalosporins, sulfonamides, ciprofl oxacin,

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tetracycline, lincomycin, NSAIDs, carbamazepine, are said to make up 4 % of all adverse drug phenytoin, allopurinol, thiouracil, propanolol, reactions to amoxicillin. Hypersensitivity vascu- griseofulvin, captopril, gold salts, barbiturates, litis is another example of a type III hypersensi- and monoclonal antibodies. tivity response that may be induced by drugs. For a description of the immunological events Drugs most commonly implicated include central to the mechanism of serum sickness, see β-lactams, cotrimoxazole, NSAIDs and some Sect. 3.8 . As well as the most common cause of monoclonal antibodies (see Chap. 11 , Fig. 11.1 ). classical serum sickness reactions, namely equine antisera given as an antitoxin, other foreign proteins such as vaccines, anti-lymphocyte 2.2.4 Delayed-Type (Type IV) globulins, streptokinase, and hymenoptera ven- Hypersensitivity oms have also been implicated in reactions. Drugs can cause a reaction that is clinically sim- Unlike types I, II, and III hypersensitivities that ilar to the protein-induced condition although proceed with the involvement of antibodies, nonprotein antigens generally do not induce the type IV hypersensitivity reactions are cell-medi- response. Symptoms typically show up 6–21 ated, in particular, by antigen-specifi c effector T days after drug administration and are cells. The term “delayed” refers to the cellular similar to those seen in the classical reaction. response that generally becomes apparent Lymphadenopathy is common and fever, one of 48–72 h after antigen exposure and distin- the earliest signs of serum sickness, tends to per- guishes the response from type I or immediate sist throughout the illness. Note, however, that reactions that often appear within minutes and lymphadenopathy has not been reported in peni- peak in a matter of minutes or just a few hours. cillin-induced serum sickness. Cutaneous symp- Type IV hypersensitivity is not represented by a toms occur in up to 95 % of patients with single reaction. Rather, it is a number of related urticarial and morbilliform eruptions being the responses seen in a variety of reactions that may most common and erythema and petechiae some- have benefi cial or undesirable consequences for times appearing. Angioedema may be seen and the host and which, at fi rst sight, do not seem to arthritis or arthralgia and gastrointestinal symp- have a lot in common except for their cellular toms of cramping, nausea, vomiting, or diarrhea immune base. Apart from the prototypic tuber- occur in up to 67 % of patients. Joints (knee, culin test, these different reactions include cel- ankle, shoulder, elbow, wrist, spine, jaw) may be lular responses to intracellular pathogens such severely affected; respiratory symptoms and as mycobacteria, fungi, and parasites; graft splenomegaly occurs; and hepatomegaly, periph- rejection; graft verse host reactions; granuloma- eral neuropathies, encephalomyelitis, and peri- tous infl ammation as occurs in Crohn’s disease carditis have been reported. With few if any and sarcoidosis; tumor immunity; some autoim- laboratory-detected changes to help with the mune reactions; and in contact allergy and aller- diagnosis, serum sickness-like reactions, as with gic contact dermatitis. Other infectious diseases classic serum sickness, tend to be diagnosed in which type IV reactions play a part include clinically. Although the drug-induced reaction leprosy, histoplasmosis, toxoplasmosis, blasto- can be severe, most are mild and usually resolve mycosis, and leishmaniasis. A well-known spontaneously within a few days or weeks after example of allergic contact dermatitis is the discontinuing the drug. True incidences of the reaction provoked by the lipid-soluble chemi- reaction to various drugs are not known although cals, mixed pentadecacatechols in urushiol oil amoxicillin and amoxicillin–clavulanic acid were present in plants of the Rhus genus, namely, poi- involved in 18 % of serum sickness-like reactions son ivy, poison oak, and poison sumac. After examined in a pediatric emergency department, crossing the cell membrane, the catechol deriva- and there are numerous reports implicating cefa- tives interact with intracellular proteins before clor in children. Serum sickness-like reactions binding with MHC (major histocompatibility

[email protected] 2.2 Classifi cation of Hypersensitivity Reactions 25 complex) class I molecules. These modifi ed Table 2.3 presents a side-by-side summary of peptides are recognized by CD8+ T cells which all four hypersensitivities according to the Gell respond with the secretion of cytokines such as and Coombs classifi cation viewed from the per- IFN-γ and cell destruction. spective of the responses to drugs in humans. In some classifi cations, type IV reactions are Delayed cutaneous hypersensitivity reactions subdivided into three categories based on the to drugs generally begin from about 7 to 21 days time of onset, the clinical manifestations, and the after contact with the drug. Subsequent reactions cells involved. The main features of the different may appear only 1 or 2 days after reexposure. categories in one such classifi cation include: Identifi cation and specifi city of the drug is estab- 1. Tuberculin type reaction (seen as local indura- lished from oral challenge studies, patch tests, tion): antigen presented intradermally; reac- and intradermal tests read after a delay of at least tion time 48–72 h; cells involved, lymphocytes, 48 h. Different T cell subsets with their individ- monocytes, and macrophages ual profi les of cytokines and chemokines are 2. Contact type reaction (seen as eczema): cuta- associated with different skin hypersensitivity neous contact (e.g., with chemicals, poison reactions although there is often overlapping ivy, nickel); reaction time 48–72 h; cells cytokine involvement. involved, lymphocytes, macrophages Summarized descriptions of the most impor- 3. Granulomatous type reaction (as in leprosy): tant immune-mediated delayed cutaneous antigen persists in the host; reaction time adverse drug reactions follow. Readers should 21–28 days; cells involved, macrophages, epi- refer to Sect. 3.6.3 for details of the mechanisms theloid, and giant cells involved in these reactions. Another subdivision of type IV hypersensitivity reactions has categories distinguished by the 2.2.4.1 Allergic Contact effector cells and mediators involved together Dermatitis with the resulting associated cutaneous reactions. Contact hypersensitivity may result from sensiti- Four subdivisions are defi ned: type IVa, medi- zation to chemicals such as chromates and picryl ated by monocytes with IFN-γ as dominant chloride in industrial settings; from chemicals cytokine; type IVb, mediated by eosinophils such as p -phenylenediamine in hair dyes; nickel with IL-5 and IL-4 involvement; type IVc, in jewelry, glasses, and devices (Fig. 2.4 ); urishiol mediated by T cells with perforin and granzyme in Rhus plants; and drugs such as neomycin (Sect. B as important effector molecules; and type 6.1.5.1 ; Fig. 6.7 ) and bacitracin (Sect. 6.1.5.2 ) IVd, mediated by neutrophils with IL-8 involve- used in topical applications. Contact dermatitis is ment. Representative skin reactions for the so- the most common occupational disease although called types IVa, b, c, and d are, respectively, it should be remembered that not all contact der- maculopapular rash, maculopapular rash with matitis has an immune basis; some irritants such eosinophilia, Stevens–Johnson syndrome (SJS), as detergents, solvents, acids, and alkalis may toxic epidermal necrolysis (TEN), and acute provoke irritant contact dermatitis. Allergic con- generalized exanthematous pustulosis (AGEP). tact dermatitis may appear at any age. The time It should be remembered that for delayed, between the initial exposure to the offending type IV cutaneous allergic drug reactions (and agent and the development of skin sensitivity sometimes for type I reactions as well), the aller- may only be 2–3 days while the interval between gic response is generally heterogeneous with exposure and the fi rst symptoms may be as early overlapping reactions and with the involvement as 12 h but is usually closer to 48 h. The reaction of different effector cells including various T is generally confi ned to the contact site but gener- cells with separate functions. Naturally, this het- alized reactions may occur. When the face is erogeneity affects the clinical picture and adds to involved, swelling of the eyelids is common. Other the diffi culties of the clinician in coming to a skin sites may become involved by the patient confi dent diagnostic conclusion. touching other areas of skin after touching the

[email protected] 26 2 Classiﬁ cation and Descriptions of Allergic Reactions to Drugs

Table 2.3 Hypersensitivities to drugs according to the Gell and Coombs classifi cationa Type of hypersensitivity I II III IV Other designations Immediate; anaphylactic Cytotoxic Immune complex Delayed; cell- mediated; T cell mediated Time for reaction Seconds to 30 minb Hours (~1 day) 3–10 h 24–72 h to reach maximum Immune reactant(s) IgE antibody IgG (and IgM) IgG antibody Th1, Th2 and/ antibodies (and/or IgM) or Th17 cells Cytotoxic lymphocytes Effector mechanism Mast cell and basophil Complement fi xation Complement Macrophage activation activation Phagocytes, NK cells Phagocytes Cytotoxic lymphocytes (Fc receptor cells) Eosinophil activation Intradermal Wheal and fl are Lysis and necrosis Erythema and edema Erythema and response to antigen induration Histology Degranulated mast cells; Immunofl uorescence Acute infl ammatory Perivascular Cellular infi ltrates shows antibody, reaction; infl ammation; including neutrophils b complement, Mainly neutrophils Mainly mononuclear neutrophils cells Sensitivity Serum IgE antibody Serum antibody Serum antibody Lymphoid cells transferred by Examples Erythema; urticaria; Drug-induced Serum sickness; Allergic contact of disease states angioedema; hemolytic anemia, Drug-induced dermatitis; Psoriasis; respiratory symptoms; thrombocytopenia, vasculitis Maculopapular GI symptoms; agranulocytosis exanthema; AGEP; anaphylaxis (immune form) FDE; DRESS; SJS; TEN; EM Drugs implicated β -Lactams; other β-Lactams; quinine; β-Lactams; cipro- NSAIDs; β-lactams; antibacterials; NMBDs; quinidine; sulfon- fl oxacin; sulfon- other antibiotics; some NSAIDs; amides; NSAIDs; amides; lincomycin; anti-convulsants; quinolones; mAbs; proton procainamide; tetracycline; NSAIDs; antimalarials; local pump inhib’s gold; carbamazepine; carbamazepine; anesthetics; barbitu- propylthiouracil; allopurinol; gold; rates; quinolones; ticlopidine methyldopa; mAbs dapsone a Coombs RRA, Gell PGH. In: Gell PGH et al. (eds.) Clinical Aspects of Immunology. Oxford: Blackwells. 1975, p. 761–81 b Late reaction may occur ~3–4 h after immediate reaction, peak at ~12 h and subside by ~24 h AGEP acute generalized exanthematous pustulosis, DRESS drug reaction with eosinophilia and systemic symptoms, EM erythema multiforme, FDE fi xed drug eruption, mAbs monoclonal antibodies, NMBDs neuromuscular blocking drugs, NSAIDs nonsteroidal anti-infl ammatory drugs, SJS Stevens–Johnson syndrome, TEN toxic epidermal necrolysis affected area. The skin may be red, swollen, and a genetic predisposition to psoriasis illustrated by show blistering or be dry and bumpy but in the a concordance rate of 70 % in monozygotic twins active stage, the skin usually shows redness, with and prevalences of 50 % and 16 % respectively raised areas and blisters (Fig. 2.5 ). when both parents or only one has psoriasis. Several genetic susceptibility loci have been 2.2.4.2 Psoriasis reported, particularly the so-called psoriasis sus- Psoriasis is a chronic, immune-mediated infl am- ceptibility 1 (PSORS1) locus on chromosome 6 matory skin disease with a prevalence generally which appears to be associated with up to 50 % of stated to be around 2 % but, depending on the cases of psoriasis. As well as drugs, psoriasis may population, this fi gure can range from zero in be triggered by smoking, alcohol, and withdrawal Samoa to as high as 4.8 % in the USA (US range of systemic or topical corticosteroids. One study 0.6–4.8 %). Ethnicity appears to be involved, for showed 23 % of patients were taking more example, American blacks show a prevalence of than three medications and 11 % of these were only 0.45–0.7 % and, likewise, there appears to be taking more than ten medications. The clinical

[email protected] 2.2 Classiﬁ cation of Hypersensitivity Reactions 27

presentation of the disease is, typically, sharply demarcated erythematous papules and rounded plaques covered by silvery micaceous scales most commonly on the scalp, elbows, umbilicus, lum- bar region, and knees (Fig. 2.6 ). Lesions of psoriasis vulgaris may show small pustules but various forms of pustular psoriasis including generalized and localized variants have been described. Both the more common vulgar form and the pustular form may progress to psoriatic erythroderma affecting the whole body. Fingernails and toenails can also be affected in all types of psoriasis. A relationship between psoriasis and certain drugs is well recognized. The authors of one study, for example, suggested that acute generalized exanthematous pustulosis (AGEP) is a cutaneous reaction pattern that might be favored by a “psoriatic background,” and medications could be responsible for psoriasis in up to 83 % of cases. Drugs can affect psoriasis in a number of ways—they may induce the disease, cause skin Fig. 2.4 Allergic nickel contact dermatitis caused by (a ) eruptions, induce lesions in previously unaffected reading glasses and ( b) a multifunction key on a cell skin in patients with psoriasis, and induce a form phone. From Veien NK, in: Johansen JD, Frosch PJ, Lepoittevin J-P, editors. Contact Dermatitis. 5th ed. of psoriasis that is resistant to treatment. Drugs Berlin: Springer-Verlag; 2011. With kind permission from that provoke psoriasis can be divided into two Springer Science+Business Media categories—drugs that induce psoriasis but withdrawal of the drug stops further progression of the disease and drugs that aggravate psoriasis but the disease still progresses even after drug withdrawal. Lithium, β-blockers, and synthetic antimalarials are the drugs most commonly mentioned in triggering or worsening psoriasis, but there are many other drugs that have been implicated either as inducers of the disease or for provoking eruptions. In the former case, the list includes acetazolamide, aminoglutethimide, amiodarone, amoxicillin, ampicillin, aspirin, chloroquine, cimetidine, corticosteroids, cyclosporin, diclofenac, diltiazem, hydroxychloroquine, indomethacin, lithium, methicillin, propranolol, and terbinaﬁ ne. According to Litt ( 2006 ), there are at least 125 different drugs known to be responsible Fig. 2.5 Acute allergic contact dermatitis to the topical for the eruption or induction of psoriasis. Psoriatic antiviral tromantadine hydrochloride showing blistering. eruptions occur in 3.4–45 % of patients treated From Brandão FM, in: Johansen JD, Frosch PJ, with lithium. The mechanism is currently Lepoittevin J-P, editors. Contact Dermatitis. 5th ed. Berlin: Springer-Verlag; 2011. With kind permission from believed to be by inhibition of the intracellular Springer Science+Business Media release of calcium as a result of lithium-induced

[email protected] 28 2 Classiﬁ cation and Descriptions of Allergic Reactions to Drugs

Fig. 2.6 Psoriasis of the elbows showing irregular red neum (image by courtesy of the Center for Disease patches covered by dry, scaly hyperkeratotic stratum cor- Control and Richard S. Hibbets)

depletion of inositol monophosphatase. of which are available over the counter, are fre- Supporting this is the benefi cial effect of inositol quently used by patients with psoriasis or psori- supplementation in lithium-provoked psoriasis. atic arthritis, but it has been claimed that the The administration of the antimalarials chloro- NSAIDs are the most common cause of both the quine and hydroxychloroquine to patients with induction and exacerbation of psoriasis. Naproxen psoriasis is considered to be contraindicated by in particular has been implicated. NSAIDs some. Exacerbation of psoriatic lesions and the exhibit a short latency period of only about 1.6 induction of the disease have been found after weeks. Leukotrienes, which increase due to use of these drugs and chloroquine has been NSAID-induced redirection of the arachidonic implicated in resistance to treatment when given acid prostanoid pathway to the lipoxygenase as an antimalarial. Clinical improvement is seen pathway (see Chap. 9 . Sects. 9.4.1 and 9.5.2.1.2 ), after withdrawal of β-blockers in cases where the are thought to aggravate psoriasis. Other drugs drugs have induced or exacerbated psoriasis. The for which there is enough data to be sure that they reactions to β-blockers usually manifest 1–18 are a signifi cant problem for psoriasis include months after initiation of therapy although, ACE inhibitors, amiodarone, benzodiazepines, because of some histological fi ndings and clini- cimetidine, clonidine, digoxin, fl uoxetine, gemfi - cal features (such as absence of elbow and knee brozil, gold, interferons, quinidine, and tumor involvement and lesions that are less thick and necrosis factor. ACE inhibitors appear to be a scaly), some believe that the condition induced problem in patients over 50 years of age. by these drugs is not true psoriasis. The mecha- Although the treatment and management of nism underlying the reactions induced by drug reactions are beyond the scope of this book, β-blockers is thought to involve blockade of epi- it is worth noting that a number of biological dermal β 2 receptors, which leads to decreased agents are currently being used either in the clinic cAMP levels in the epidermis and ultimately or experimentally to treat psoriasis. These include keratinocyte hyper-proliferation. NSAIDs, many etanercept, a fusion protein inhibitor of tumor

[email protected] 2.2 Classiﬁ cation of Hypersensitivity Reactions 29 necrosis factor (see Sect. 11.2 ), and the chimeric and human monoclonal antibodies adalimumab, brodalumab, inﬂ iximab, ixekizumab, and ustekinumab (Sect. 11.1 ).

2.2.4.3 Maculopapular Exanthema Maculopapular exanthema, also called morbilliform drug reaction, morbilliform exanthem, and maculopapular drug eruption, is the most frequently seen pattern of drug-induced skin eruption. Clinical presentation is diverse and can vary from an erythematous rash mimicking a viral or bacterial exanthem to generalized symmetric eruptions of both isolated and confl uent erythematous plaques. These often start on the trunk and then spread to the extremities and neck without involvement of the mucosa. The pink to red macules or papules tend to blanch when pressed and purple, non-blanchable spots (purpura) may appear on the lower legs. Reactions appear 7–14 days after exposure to drug but after only 1 or 2 days in already sensitized patients. Reactions usually tend to progress over a few days before regressing over a 2-week period often with accompanying desquamation. This may be the course of events even if the drug has been withdrawn. Reactions may be associated with a mild fever and itch. Histologically, lymphocytes are seen in papillary dermis and at the junction of the dermis and epidermis while degenerated, necrotic and some dyskeratotic keratinocytes and spongiosis are visible in early lesions. Maculopapular exanthema can be caused by a wide variety of drugs. Those commonly implicated are chiefl y antibiotics, especially aminopenicillins (Fig. 2.7 ) and cephalosporins, but also sulfonamides, anticonvulsants Fig. 2.7 Generalized maculopapular exanthema follow- like carbamazepine, allopurinol, and NSAIDs. ing the introduction of amoxicillin therapy showing lesions on the trunk ( a) and targeted lesions on the hands and forearms ( b). The patient had positive patch tests to 2.2.4.4 Acute Generalized amoxicillin and ampicillin and negative tests to benzyl- Exanthematous Pustulosis penicillin, dicloxacillin, and a number of cephalosporins. AGEP, also termed toxic pustuloderma and pustu- From Gonçalo M, in: Johansen JD, Frosch PJ, Lepoittevin J-P, editors. Contact Dermatitis. 5th ed. Berlin: Springer- lar drug eruption, is induced in 90 % of cases by Verlag; 2011. With kind permission from Springer drugs and characterized by fever and acute non- Science+Business Media follicular pustular eruptions that overlay erythrodermic skin. The incidence of the disease is said to be about three to fi ve cases per million per year and groin a few days after administration of the with a mortality rate of about 5 %. Reactions gen- offending drug before becoming widespread. erally begin with a rash on the face or in armpits Time of onset after drug administration can be

[email protected] 30 2 Classiﬁ cation and Descriptions of Allergic Reactions to Drugs

Fig. 2.8 A case of acute generalized erythematous pustulosis (AGEP) to hydroxychloroquine sulfate (photograph courtesy of Dr. Adrian Mar)

Fig. 2.9 A patient with drug reaction (or rash) with remarkably short—even as little as 24 h in some eosinophilia and systemic symptoms (DRESS), also cases. The rash lasts about 2 weeks, and the reac- referred to as hypersensitivity syndrome or drug-induced tion regresses with skin desquamation as it hypersensitivity syndrome. The patient experienced sys- resolves 5–10 days after drug withdrawal, often temic symptoms, skin reactions with nonspecifi c maculopapular rash, and exfoliative dermatitis with facial edema with rapid spontaneous healing. AGEP is charac- (photograph courtesy of Dr. Adrian Mar) terized by symmetrical widespread edematous erythema with small non-follicular sterile pustules (Fig. 2.8 ) due to neutrophil accumulation on the drug. DRESS involves systemic symptoms predominating in body folds. Recruitment of neu- and skin reactions with nonspecifi c maculopapular trophils occurs in the late phase of development rash or a more generalized exfoliative dermatitis of lesions. Histopathology shows spongiosis, sub- and facial edema (Fig. 2.9 ). Symptoms consist of corneal pustules, leukocytoclastic vasculitis, dis- fever, malaise, arthralgia, enlarged lymph nodes, crete vacuolar keratinocyte degeneration, and hepatitis, renal impairment, pneumonitis, and dermal and intradermal infi ltration of lympho- hematological abnormalities with atypical acti- cytes. AGEP is also characterized by the great vated lymphocytes and elevated levels of eosino- predominance (over 80 % of cases) of antibiotics phils that infi ltrate the skin and other organs and (including amoxicillin, ampicillin, tetracycline, are thought to cause tissue damage. Visceral spiramycin, pristinamycin, and metronidazole) as involvement differentiates DRESS from other causative agents. Other implicated drugs include exanthematous serious acute drug reactions. hydroxychloroquine, cotrimoxazole, diltiazem, Another distinguishing feature is the extended terbinafi ne, and carbamazepine. interval between drug exposure and the onset of symptoms, which is usually about 2–8 weeks. 2.2.4.5 Drug Reaction (Rash) with DRESS also tends to regress more slowly and the Eosinophilia and Systemic disease may sometimes reactivate after exposure to Symptoms an unrelated drug or even without drug exposure. Drug reaction (or rash) with eosinophilia and sys- Viruses may have a role in the development of the temic symptoms (DRESS), also referred to as symptoms of DRESS. This is suggested by reacti- hypersensitivity syndrome or drug-induced hyper- vation of the Herpesviridae viruses, cytomegalovi- sensitivity syndrome, is a severe life-threatening rus, EBV, and human herpes virus 6 (HHV-6), the adverse drug reaction with an incidence reported to latter evidenced by the rise of anti-HHV-6 IgG be between 1 in 1,000 and 1 in 10,000 depending titers and the presence of HHV-6 DNA 2–3 weeks

[email protected] 2.2 Classifi cation of Hypersensitivity Reactions 31 after the onset of rash. The main drugs involved in eliciting reactions are the anticonvulsants carbamazepine, lamotrigine and phenytoin, pheno- barbitone, dapsone, sulfonamides, allopurinol, minocycline, and mexiletine. In a retrospective analysis of 1,544 DRESS cases reported to the United States Food and Drug Administration between 2004 and 2010, 137 cases (8.9 %) had a fatal outcome; the sex ratio was fi ve females to four males and the most frequently involved age range was the 60–69 group. Those older than 70 had a higher incidence of fatalities. Approximately 60 % of cases developed DRESS within 4 weeks but some late-onset cases developed after 6 months Fig. 2.10 A fi xed drug eruption showing the characteristic, often-seen circular shape. Lesions often resolve with post- exposure to the drug. The top 20 drugs most fre- infl ammatory pigmentation (photograph courtesy of quently implicated in provoking DRESS, in order Dr. Adrian Mar) of highest to lowest frequency, were: carbamazepine, sulfasalazine, allopurinol, vancomycin, amoxicillin, lamotrigine, phenytoin, minocycline, zonisamide, abacavir, ciprofl oxacin, piperacillin, rifampicin, ibuprofen, diclofenac, valproate, acetaminophen, phenobarbital, lamivudine, omeprazole. Increased reporting of DRESS is apparent, although this increase is not related to newly marketed drugs but rather the already well-known causes. Only 12 % of the 1,544 cases reported to the FDA were from the USA whereas 527 cases (34 %) were reported from France, 286 (19 %) from Japan, 79 (5 %) from the UK, and 135 cases (9 %) unknown. It is clear that there is underre- porting to the extent, as some have claimed, that Fig. 2.11 An example of a well-circumscribed bullous the reports received by the FDA are only 1–10 % fi xed drug eruption. The reaction was induced by carbam- of the real number. Finally, there is some disagree- azepine, a drug implicated in some severe drug-induced ment over the variability of the clinical pattern of delayed hypersensitivity responses (photograph courtesy of Dr. Adrian Mar) cutaneous symptoms and the classifi cation of some patients as having DRESS. Some claim that use of the term DRESS in not consistent; cutane- symptoms are usually absent. The period ous and systemic signs vary and eosinophilia is required for sensitization ranges from weeks to not a constant fi nding. With this in mind, the des- years and the time between drug administration ignation DIHS/DRESS to include DRESS and and eruption can be anything from a day or two drug- induced hypersensitivity syndrome (DIHS) to a few weeks. Initially, one or a small number has been suggested. of round or oval pruritic, well circumscribed, erythematous macules appear (Fig. 2.10 ). These 2.2.4.6 Fixed Drug Eruption may progress to edematous plaques or bulla Fixed drug eruption (FDE) is due to drug hyper- (Fig. 2.11 ) and may regress over a period of sensitivity in more than 95 % of cases. Patients 10–15 days. In a few cases, lesions can be so may complain of burning in the affected area widespread that it is diffi cult to distinguish FDE before the appearance of lesions but systemic from TEN. FDE is so named because the site of

[email protected] 32 2 Classifi cation and Descriptions of Allergic Reactions to Drugs the eruption is fi xed, that is, it occurs in exactly the same place when the same drug is again encountered. Upon discontinuation of the drug, lesions resolve leaving hyperpigmentation although pigmentation is often absent when the skin is fair. A FDE can occur anywhere on the skin or on mucous membranes, but the most commonly affected sites are the hands, feet, penis, groin and perianal areas, and the lips. Drugs implicated in causing FDE include allopurinol, cotrimoxazole, antibiotics (tetracycline, doxycycline, amoxicillin, ampicillin, cephalosporins, and clindamycin), NSAIDs (aspirin, ibuprofen, naproxen, and diclofenac), acetamin- Fig. 2.12 Erythema multiforme with circumscribed macular and papular lesions progressing to plaques with dark and ophen, carbamazepine, dapsone, quinine, phe- purpuric centers (photograph courtesy of Dr. Adrian Mar) nolphthalein, and benzodiazepines. To confi rm diagnosis of FDE, oral challenge with a single dose of the suspected drug at one-tenth the normal therapeutic dose is accepted as safe. Interestingly, desensitization has been successful for allopurinol-induced FDE but allopurinol appears to remain the only drug where desensitization for FDE has worked.

2.2.4.7 Erythema Multiforme Erythema multiforme is a self-limiting cutaneous hypersensitivity reaction to infection (mostly) or drugs occurring mainly in adults 20–40 years of age although it can occur in patients at any age. Prodromal symptoms are either lacking or mild Fig. 2.13 Erythema multiforme involving the hands (itch, burning) and the condition usually resolves (photograph courtesy of Dr. Adrian Mar) spontaneously in 3–5 weeks without sequelae. Although considered to be closely related to SJS and TEN, and with the designation erythema involved, it is usually limited to the oral cavity. multiforme major sometimes interchanged with Patients may experience multiple outbreaks in a the former, erythema multiforme is now accepted year. More than half the cases of erythema multi- as a distinct condition. Major reasons for this are forme are due to Herpes simplex (HSV), and its lesser severity, minimal involvement with recurrent cases may be secondary to reactivation mucous membranes, and the extent of epidermal of HSV-1 and HSV-2. Acyclovir, famciclovir, or detachment which is usually less than 10 %. valacyclovir have been used to treat recurrent Lesions appear as circumscribed pink-red mac- outbreaks. Mycoplasma pneumoniae and fungal ules before progressing to papules which may infection are also commonly involved with reac- enlarge into plaques, the centers of which become tions. Drugs most commonly associated with darker or even purpuric (Fig. 2.12). Crusting or erythema multiforme are anticonvulsants, barbi- blistering may occur. Several days after onset, turates, ciproﬂ oxacin, NSAIDs, penicillins, phe- lesions of various morphology are visible, hence nothiazines, sulfonamides, and tetracyclines. the name “multiforme.” Palms (Fig. 2.13 ) and Some drugs released in more recent years are soles may be affected and if the mucosa is now being implicated, for example, the monoclo-

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Fig. 2.14 Toxic epidermal necrolysis in an adult patient tions: emergency approach to non-burn epidermolytic covering more than 30 % of the total body surface area. syndromes. Intensive Care Medicine 2010;36:22. With From Struck MF et al. Severe cutaneous adverse reac- kind permission from Springer Science+Business Media nal antibody adalimumab, bupropion, candesartan cilexetil, and metformin and reactions to some vaccines against bacteria (diphtheria–tetanus) and viruses (hepatitis B, hepatitis C, cyto- megalovirus, and HIV) are seen.

2.2.4.8 Stevens–Johnson Syndrome and Toxic Epidermal Necrolysis SJS and TEN are potentially fatal, severe, rare, adverse cutaneous drug reactions involving both the skin and mucous membranes. Beginning with relatively unremarkable signs like fever, malaise, and perhaps eye discomfort, macular Fig. 2.15 Lips and facial involvement in a child with developing drug-induced Stevens–Johnson syndrome lesions become symmetrically distributed on the (photograph courtesy of Dr. Adrian Mar) trunk, face, palms, and soles. Lesions develop a central bulla and coalesce into large sheets of necrotic tissue covering at least 30 % of the body cal, and genital mucosa involvement. Figure 2.15 in the case of TEN (Fig. 2.14 ). For SJS, skin shows lip and facial involvement in a child with denudation/detachment is generally less than developing drug-induced SJS. Respiratory and 10 % of body surface area, and this extent of skin gastrointestinal tracts can also be affected. involvement is the main distinguishing feature Ocular effects can include acute conjunctivitis between the two diseases. Skin erythema and with discharge, eyelid edema, erythema, and erosions progress to the extremities with more corneal erosion or ulceration. The collective than 90 % of patients experiencing ocular, buc- clinical features sometimes do not ﬁ t neatly into

[email protected] 34 2 Classifi cation and Descriptions of Allergic Reactions to Drugs either a clear SJS or TEN diagnosis and may be are trimethoprim–sulfamethoxazole, other sul- classifi ed as overlapping SJS or TEN. In the fonamides, aminopenicillins, cephalosporins, SJS–TEN overlap group, primary lesions tend to quinolones, and chlormezanone. In the long be dusky red or purpuric macules or fl at atypical period group: carbamazepine, phenytoin, pheno- targets widely distributed as isolated lesions but barbitone, and valproic acid, NSAIDs of the oxi- with quite marked confl uence on the face and cam type, allopurinol, and corticosteroids. upper trunk. As with TEN, but not necessarily Allopurinol is the drug most commonly impli- with SJS, systemic symptoms are always pres- cated in causing SJS/TEN in Europe and Israel. ent. Skin detachment occurs on 10–30 % of the Some newer drugs identifi ed as being of increased body surface area. comparative risk include nevirapine, lamotrigine, In the second phase of development of the dis- and sertraline. Because of the fear of provoking a eases, large areas of epidermis become detached. second episode or aggravating an existing case of Sequelae and late effects are common in SJS and SJS/TEN, skin testing in all its forms and provo- particularly TEN. These can include changes in cation tests are not to be considered although skin pigmentation, nail dystrophies, and ocular there are at least two reports of intradermal test- problems. Long-term sequelae in surviving TEN ing that did not trigger another episode. Patch patients can include eye affl ictions like entropion testing and the lymphocyte transformation test and symblepharon, on-going mucous membrane (Sect. 4.7.1 ) have been used but proved to be of erosion, and cutaneous scarring. Dry mouth and low sensitivity. Clinical features and histology dry eyes resembling Sjögren syndrome may be a therefore remain the mainstay of diagnosing long-term complication. SJS/TEN. Although SJS and TEN are rare with incidences of usually less than two per million per year (e.g., 1.89 Western Germany, 1.9 USA), Summary some infectious diseases can have a marked effect in the incidence. This is seen with HIV • In the Gell and Coombs classifi cation of aller- where the annual incidence is approximately gic reactions, four types of hypersensitivities 1,000 times higher than in the general popula- designated types I, II, III, and IV are tion. Infections M. pneumoniae and H. simplex distinguished. are known to cause SJS and TEN without drug • Type I, also called immediate or anaphylactic involvement. A strong association shown hypersensitivity, occurs within about 30 min between HLA-B*15:02, SJS, and carbamazepine but reactions can be dramatic and appear in Han Chinese and confi rmed in a Thai popula- within seconds or minutes as in anaphylaxis. tion is a clear demonstration of a relationship • Type I reactions are IgE antibody-mediated. between ethnicity and drug hypersensitivity. This Receptor-bound drug-reactive IgE on the sur- fi nding has stimulated interest in the investiga- face of mast cells is cross-linked by completion of genetic factors in drug hypersensitivity mentary drug determinants causing cell (see Sect. 3.4); one recent demonstration being degranulation and the release of infl ammatory the large collaborative European RegiSCAR mediators. project which showed that HLA-B*15:02 is not a • Drugs well known to cause type I reactions genetic marker for carbamazepine, sulfamethox- include penicillins, cephalosporins, neuro- azole, lamotrigine, or NSAIDs of NSAID muscular blocking drugs, some NSAIDs, oxicam- type-induced SJS/TEN in Europe. The monoclonal antibodies, quinolones, and prodrugs most commonly responsible for SJS/TEN ton pump inhibitors. have been divided by Roujeau into those that pro- • The term “anaphylactic” is used to describe an voke the disease after only a short period of immune-mediated immediate systemic reac- administration and those that do so after being tion involving the release of potent mediators taken for longer periods. In the short period group from mast cells and basophils that produce

[email protected] Further Reading 35

a range of possible symptoms including Further Reading erythema, urticaria, angioedema, bronchospasm, gastrointestinal symptoms, and cardio- Aster RH, Bougie DW. Drug-induced immune thrombo- vascular collapse. The term “anaphylactoid” cytopenia. N Engl J Med. 2007;357:580–7. is used for reactions that show clinically simi- Berliner N, Horwitz M, Loughran TP Jr. Congenital and lar signs and symptoms but where no immune- acquired neutropenia. Hematology Am Soc Hematol mediated mechanism can be demonstrated. Educ Program. 2004;63–79. Bock G, Goode J, editors. Anaphylaxis. Novartis founda- • It is important to have a suitable clinical grad- tion symposium 257. Chichester: Wiley; 2004. ing system for anaphylaxis. Castells MC, editor. Anaphylaxis and hypersensitivity • Urticaria is the second most common cutane- reactions. New York, NY: Humana; 2011. ous reaction induced by drugs, often in associa- Grattan CE, Charlesworth EN. Urticaria. In: Holgate ST, Church M, Lichtenstein LM, editors. Allergy. 3rd ed. tion with angioedema and anaphylaxis. Many Philadelphia, PA: Mosby; 2006. p. 95–106. drugs are implicated including β-lactams, Harr T, French LE. Toxic epidermal necrolysis and NSAIDs, sulfonamides, vancomycin, and con- Stevens-Johnson syndrome. Orphanet J Rare Dis. trast media. ACE inhibitors are responsible for 2010;5:39–49. Johansen JD, Frosch PJ, Lepoittevin J-P, editors. Contact approximately one in six patients admitted to dermatitis. Heidelberg: Springer; 2011. hospital with angioedema. Joint Task Force on Practice Parameters. The diagnosis • Type II hypersensitivity is also known as and management of urticaria: a practice parameter. antibody- dependent cytotoxic hypersensitiv- Part I. Acute urticaria/angioedema. Part II. Chronic urticaria/angioedema. Ann Allergy Asthma Immunol. ity. Drugs can attach to cell membranes 2000;85:521–44. producing drug-induced hemolytic anemia, Lieberman P. Defi nition and criteria for the diagnoses of thrombocytopenia, and granulocytopenia. anaphylaxis. In: Castells MC, editor. Anaphylaxis and Drugs implicated: hemolytic anemia—peni- hypersensitivity reactions. New York, NY: Humana; 2011. p. 1–12. cillins, quinidine, methyldopa; thrombocyto- Litt JZ. Drug eruption reference manual. 12th ed. London: penia—quinine, quinidine, propyl thiouracil, Taylor and Francis; 2006. vancomycin, sulfonamides; granulocytope- Mockenhaupt M, Viboud C, Dunant A, et al. Stevens- nia—pyrazolones, thiouracil, anticonvulsants, Johnson syndrome and toxic epidermal necrolysis: assessment of medication risks with emphasis on and sulfonamides. recently marketed drugs. The EuroSCAR-study. J • Type III hypersensitivity is mediated by solu- Invest Dermatol. 2008;128:35–44. ble immune complexes mostly involving IgG Radic M, Martinovic Kaliterna D, Radic J. Drug-induced antibodies. Drug-induced serum sickness-like vasculitis: a clinical and pathological review. Neth J Med. 2012;70:12–7. reaction is the prototype example of type III Salama A, Schutz B, Kiefel V, et al. Immune-mediated drug hypersensitivity. Hypersensitivity vascu- agranulocytosis related to drugs and their metabolites: litis is another example of a type III hypersen- mode of sensitization and heterogeneity of antibodies. sitivity response induced by drugs. Br J Haematol. 1989;72:127–32. Sampson HA, Munoz-Furlong A, Bock SA, et al. • Type IV hypersensitivity reactions are medi- Symposium on the defi nition and management of ana- ated by antigen-specifi c effector T cells. phylaxis: summary report. J Allergy Clin Immunol. Reactions generally occur 48–72 h after anti- 2005;115:584–91. gen exposure and are therefore referred to as Schön MP, Boehncke W-H. Psoriasis. N Engl J Med. 2005;352:1899–912. delayed reactions. Shiohara T. Fixed drug eruption: pathogenesis and diag- • Important delayed cutaneous reactions include nostic tests. Curr Opin Allergy Clin Immunol. maculopapular exanthema; allergic contact 2009;9:316–21. dermatitis; psoriasis; acute generalized exan- Shiohara T, Inaoka M, Kano Y. Drug-induced hypersensitivity syndrome (DIHS): a reaction induced by a com- thematous pustulosis; drug reaction with plex interplay among Herpes viruses and antiviral and eosinophilia and systemic symptoms; fi xed antidrug immune responses. Allergol Int. drug eruption; erythema multiforme; Stevens– 2006;55:1–8. Johnson syndrome; and toxic epidermal Sidoroff A, Halevy S, Bavinck JNB, et al. Acute generalized exanthematous pustulosis (AGEP) – a clinical necrolysis. reaction pattern. J Cutan Pathol. 2001;28:113–9.

[email protected] Mechanisms of Hypersensitivity 3

Abstract Allergic reactions to drugs are not always the result of the drug’s protein- binding capacity, biotransformation, or degradation. Mediator release may occur via cross-linking of cell-bound IgE by di-(multi-) valent free drug. Physiological and pharmacological effects of histamine are

mediated through four receptors, H1 , H2 , H3 , and H4. The H3 receptor has a regulatory role in the release of neurotransmitters such as serotonin and

dopamine; the H4 receptor exerts a chemotactic effect on several cell types associated with allergy and asthma. Cysteinyl leukotrienes and PAF are powerful mediators of anaphylaxis, asthma, and shock. Sphingosine-1- phosphate, elevated in the lungs of asthmatics, regulates pulmonary epithelium permeability and contributes to the pathogenesis of anaphylaxis. Urticaria is a heterogeneous disease with many subtypes. Both ACE inhibitors and angiotensin II receptor blockers may cause angioedema. Abacavir changes the shape of the HLA antigen-binding cleft producing an alteration in the repertoire of self-peptides that bind HLA-B*57:01 and a T cell response to self-proteins. Drug-induced delayed-type cutaneous hypersensitivity reactions are mediated by CD4+ and CD8+ CD3+ T cells in the dermis and epidermis. Granulysin appears to be a key molecule for keratinocyte killing in TEN/SJS. Drugs provide good examples of types II (immune hemolytic anemia, drug-induced thrombocytopenia) and III (serum sickness-like) hypersensitivities.

In this chapter, emphasis has been placed on the sensitivities including, for example, responses to core mechanisms underlying the broad categories reactive metabolites from chemically “inert” par- of hypersensitivity responses distinguished on ent drugs such as sulfamethoxazole; relationships the basis of the Gell and Coombs classiﬁ cation between chemical structures and immune and based on differences in the immune reactants responses seen with, for example, anaphylactic (antibodies or cells), the form of the presented reactions to neuromuscular blocking drugs during antigen, and the effector mechanisms involved. anesthesia; hypersensitivities and other Mechanisms involved in individual drug hyper- intolerances to nonsteroidal anti-inﬂ ammatory

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 37 Relationships, DOI 10.1007/978-1-4614-7261-2_3, © Springer Science+Business Media, LLC 2013

[email protected] 38 3 Mechanisms of Hypersensitivity drugs (NSAIDs); and mechanisms underlying the soluble or cell-bound protein, by biotransforma- killing of malignant cells by some drugs used in tion of the drug to form a reactive metabolite able chemotherapy are not confi ned to this chapter but to bind to a carrier protein, or by degradative presented in the relevant chapters dealing with changes to the parent molecule forming reactive pharmacologically different groups of drugs. groupings (Fig. 3.1 ). In practice, however, it is Most hypersensitivities to drugs manifest as type often not possible to show protein binding by a I or type IV reactions. Type II and type III drug drug or to even offer a satisfying explanation of hypersensitive reactions are far less often seen how such binding might occur given the known and are considered after the discussions of the chemical properties of the drug and the metabolic type I and IV responses. Mechanisms, to the processes to which it is exposed. While a number extent that they are currently understood, of other of allergenic drugs such as the β-lactams undergo types of “hypersensitivity” reactions or intoler- well-known ring-opening and subsequent ances, some mediated by antibodies other than protein-binding reactions (Chap. 5), for many IgE, and others by cells, are also discussed. We drugs chemical reactivity, protein binding, bio- begin by examining the mechanisms underlying transformation, or the involvement of degrada- type I drug-induced IgE antibody allergic sensiti- tive products and /or reactive impurities has not zation, regulation and production, and the effec- been demonstrated. This raises the question of tor mechanisms operative in IgE-mediated another possible mechanism(s) to explain the allergic reactions. immune recognition of “small” drug molecules and the subsequent immunological steps involved in the drug-induced hypersensitivity responses 3.1 Allergic Sensitization to Drugs (see Sects. 3.1.2 , 3.1.3 , 3.4 , 6.2.3.3 , 7.4.2.3 , and and the Dogma of Previous 7.4.5.3 ). Over the last few decades, a large num- Exposure ber of drugs have been implicated as provoking agents for a range of hypersensitivity states, and 3.1.1 Immunogenicity of Free although the list of identifi ed drug allergenic and Conjugated Drugs determinants has expanded, this aspect of hypersensitivity research is still in its infancy. As well as the chemical nature of a drug, its size Structures as diverse as substituted ammonium and complexity infl uence its antigenicity. ions, simple disaccharides, small side chain Chemicals of molecular mass less than 5 kDa and groups and ring structures on β-lactams, haloge- sometimes up to about 10 kDa are often poorly or nated isoxazolyl groups, and whole molecules as non-antigenic. From the time of the early immu- seen with trimethoprim and chlorhexidine are nochemical studies on antigenicity and haptens, known to be recognized as allergenic determi- organic chemicals of small molecular mass have nants in some drug-allergic patients. Of the pres- been assumed to be antigenic and capable of ently known drug allergenic determinants, most stimulating an immune response only as a com- have been reliably identifi ed in IgE antibody plex with a macromolecular carrier, usually pro- recognition studies using quantitative immuno- tein. By coupling a wide range of different chemical hapten inhibition techniques. The chemicals that are not antigenic in their free state, approaches and methods already applied so for example, steroids, sugars, purines, pyrimi- successfully to a range of drugs (see Chaps. 5 – 8 ) dines, nucleosides, and aromatic ring compounds need to be expanded to cover other yet-to-be- such as phenols, etc., Landsteiner and other early defi ned IgE antibody-binding determinants and investigators demonstrated clear and specifi c extended to T cell-mediated drug hypersensitiv- antibody responses in laboratory animals. ity reactions where progress has been slow. Chemicals such as drugs may form hapten–carrier Results from the IgE studies have demonstrated complexes in vivo in three different ways—by that more than one allergenic determinant gener- direct chemical covalent interaction with a ally occurs on drugs and such antigenic

[email protected] 3.1 Allergic Sensitization to Drugs and the Dogma of Previous Exposure 39

metabolite + drug soluble or cell-bound protein in the body

degradation product hapten-protein conjugates

non-allergenic non-allergenic potentially allergenic

Fig. 3.1 Diagrammatic representation of possible, BA & Pham NH. Structure–activity studies on drug- and potentially allergenic, hapten–protein complexes induced anaphylactic reactions. Chem Res Toxicol 1994 ; that may form in vivo from a drug and/or its 7: 703. Reproduced with permission from American metabolite(s) and degradative product(s). From Baldo Chemical Society

heterogeneity is refl ected in patients’ IgE anti- responses to allergens, parasites, and fungi, some body recognition responses. As more drug aller- of the antibodies are “natural,” that is, antibodies gies and more allergic individuals are studied, the formed without exposure to foreign antigens via extent of this heterogeneity will emerge and with infection or passive or active immunization. it the possibility of gaining greater insights into Examples of such antibodies appear to be those the structural basis of drug allergenicity. that are complementary to various cross-reactive For an allergic reaction to a given drug, immu- carbohydrate determinants (the so-called CCDs), nological dogma requires that the response and to phosphorylcholine connected by phospho- occurs on reexposure to the drug after the initial diester linkages in some N -linked proteoglycans sensitizing exposure to that drug. However, this and glycolipids and found in pneumococcal tei- seemingly obvious requirement may not always choic acid (“C substance”) and other “C sub- hold true or appear to hold true. Some allergic stances” in bacteria, fungi, arthropods, responses, sometimes even life-threatening as helminthes, protozoa, and plants. The curious with anaphylaxis, occur on fi rst exposure to connection between IgE natural antibodies to the a drug. Such reactions to the neuromuscular D-galactose disaccharide found on cetuximab, a blocking drugs are well known and there are chimeric mouse–human IgG1 monoclonal anti- numerous other investigations and case studies body used for cancer treatment, and anaphylaxis involving a variety of pharmacologically differ- in some treated patients (see Sect. 11.1.3.2 ) and ent drugs including trimethoprim, iodinated con- the possible cross-reaction of natural anti- trast media, opioids, and some antibiotics that phosphorylcholine IgE antibodies with ammo- report the same phenomenon. In some cases, this nium groups on neuromuscular blocking drugs might be explained by previous exposure to a (Sect. 7.4.5.3 ) are indicators of the likely exis- structurally similar drug or to a structurally simi- tence of other natural IgE antibodies with poten- lar compound that may not even be administered tially cross-reactive specifi cities. Although some as a drug. An example of the former case is a of these antibodies may appear to have no con- reaction to a cephalosporin in a patient previ- nection whatsoever with a particular drug, struc- ously given a penicillin while a reaction to a drug tural features recognized by the antibody may also result from previous exposure to the combining site may resemble structures on the drug (e.g., an antibiotic in meat) or an antigeni- drug molecule resulting in allergenic cross- cally cross-reactive chemical in some foods or in reactivity. It should also be kept in mind, how- the environment. Although IgE antibodies are ever, that not all exposures to a potentially almost invariably thought of as induced humoral sensitizing drug will result in a patient becoming

[email protected] 40 3 Mechanisms of Hypersensitivity sensitized and every sensitized patient will not Bridging of adjacent cell-bound IgE molecules necessarily respond with allergic symptoms by at least bivalent allergenic determinants react- following reexposure to the sensitizing drug. ing with their complementary antibody combin- Findings so far on immune recognition of ing sites (Fig. 3.2a–d ) triggers cell degranulation drugs, especially recognition by IgE antibodies in and release of a variety of mediators that cause cases of type I immediate hypersensitivity and to the signs and symptoms of a type I hypersensi- a much lesser extent for drug recognition by spe- tivity reaction. In the case of drug–carrier conju- cifi c T cells, have shown that small parts of drug gates, cross-linking of IgE antibodies is readily molecules, sometimes only one or a few chemi- explained by the presence on the conjugates of cal groups that form part of the molecule, consti- multiple reactive drug determinant sites, but for tute the allergenic determinant structures that are free, uncomplexed drug molecules both the size complementary to the immunoglobulin E com- and number of reactive determinants would bining sites and T cell receptors. In addition to appear to be too small for cross-linkage of anti- allergic cross-sensitivity to drugs in the same body combining sites to occur. Drugs with a family, for example, between different β-lactams, single IgE-binding determinant cannot, of recognition of widely distributed structures com- course, cross-link adjacent cell-bound antibody monly occurring in many different drugs and molecules (Fig. 3.2e ), but even if two or more chemicals also represents potentially immuno- determinants are present, they must be separated logically cross-reactive determinants. Substituted by a suitable distance and/or be suitably spatially ammonium ions identifi ed as the most important arranged if cross-linking via adjacent comple- IgE antibody-binding structures in neuromuscu- mentary antibody combining sites is to occur lar blocking drugs (Sect. 7.4.2.1 ) occur in many (Fig. 3.2f, g ). Despite this problem of explaining drugs and chemicals frequently encountered by the mechanism of apparently monovalent drug- humans. Bioisosteres, that is, groups with similar induced allergic mediator release, there is at physical and/or chemical properties that impart least one group of drugs, the neuromuscular similar biological properties to a drug, should blockers (and probably more to be identifi ed) also be kept in mind when prior allergic sensiti- that can specifi cally elicit antibody-induced mast zation is suspected in patients who are fi rst time cell activation and release without fi rst undergo- reactors to a drug. In this era of so-called rational ing coupling to a macromolecular carrier. For drug design, bioisosteres are commonly seen for these drugs, di- or multi-valency is an inherent example, in the replacement of a six-membered part of the molecular structure and, even in the phenyl ring with a fi ve-membered thiophene ring absence of protein binding, cross-linking of cell- in many synthesized drugs. The importance of bound antibodies can be effected (Fig. 3.2a). Of bioisosterism in the identifi cation of allergenic the polymethylene bismethonium compounds, structures and allergic cross-reactivity is dis- the 2 nm molecular length of the C-10 congener cussed further in Chap. 5 . decamethonium is optimal for neuromuscular block, that is, it best fi ts the distance between the receptive sites on the muscle nicotinic acetyl- 3.1.2 Mediator Release by Free and choline receptor. Interonium distances, however, Conjugated Drugs in are less, for example, 1.4 nm for decametho- Immediate Allergic Reactions nium, 1.08 nm for d-tubocurarine (molecular length 1.8 nm), and 1.14 nm for pancuronium In allergic subjects, IgE antibodies, as well as (molecular length 1.9 nm). These distances being free in serum, are fi xed to the extracellular appear to be suitable for the neuromuscular D1 distil and D2 proximal domains of the FcεRI blockers to bridge and thus activate adjacent IgE receptor on mast cells and basophils via the Cε2 molecules on the mast cell surface (see also and Cε3 domains of the antibody Fc region. Sects. 7.4.2.1 and 7.4.2.3 ).

[email protected] 3.1 Allergic Sensitization to Drugs and the Dogma of Previous Exposure 41

ef g

Fig. 3.2 Different ways in which a free drug (shown in containing two (or more) different determinants that elicit bold in a , b , e , f , and g ) and a drug–protein conjugate ( c , d ) an IgE response. ( c ) and (d ) Bridging via conjugated drug may cross-link or bridge adjacent cell-bound IgE molecules molecules with cross-linking effected by the same, or dif- which triggers release of the mediators of immediate hyper- ferent, determinants, respectively. Failure to bridge adja- sensitivity. (a ) Bridging via an allergenically divalent cent cell-bound IgE molecules because: (e ) drug is unconjugated drug molecule with the same or closely allergenically monovalent; (f ) and (g ) drug determinants related allergenic determinants. This is the mechanism are not positioned to effect cross-linkage. From Baldo BA thought to occur in patients who experience anaphylaxis & Pham NH. Structure–activity studies on drug-induced following administration of a neuromuscular blocking anaphylactic reactions. Chem Res Toxicol 1994 ; 7: 703. drug. ( b) Bridging via a free, unconjugated drug molecule Adapted with permission from American Chemical Society

3.1.3 Immunological Recognition the drug is said to bind directly to self-peptides in of Free, Unconjugated Drug the antigen-binding cleft of the major histocam- Molecules patibility complex (MHC). In another possible alternative, the drug may couple directly to the The generally accepted explanation for the rec- MHC itself on regions involved in binding to the ognition of drugs causing an immune-mediated T cell receptor. In drug interaction with the MHC, hypersensitivity reaction is based on the binding recognition may be restricted to a limited number of drug to a protein carrier molecule, immune of peptides or it may be promiscuous, that is, recognition and processing of the drug–protein independent of peptide. For some drugs at least, complex, presentation of drug–peptide conju- direct stimulation of T cells via the T cell recep- gates to the T cells, and recognition and reaction tor in an MHC-dependent way has been sug- of the T cell with the drug antigen. However, gested. With sulfamethoxazole for example, a although there is no evidence that many drugs, drug known to be metabolized to its reactive either as the parent compound or as a metabolite, nitroso derivative, only a minority of T cell clones bind to a suitable carrier, there is evidence that T reactive with this metabolite were isolated from cells recognize metal ions such as Ni2+ and some sulfamethoxazole-allergic patients. The short drugs like sodium aurothiomalate that do not time period for T cell activation to occur with require antigen processing. In one explanation, some free, unmetabolized drugs, T cell clone

[email protected] 42 3 Mechanisms of Hypersensitivity reactivity with glutaraldehyde-fi xed antigen- Resting or naïve B cells are also nondividing, presenting cells, and removal of free drug by and to undergo clonal expansion and differentia- washing all suggests a drug–T cell receptor inter- tion to effector B cells, that is, to produce aller- action that is independent of metabolism and pro- gen-reactive IgE antibodies, B cells also require cessing. Further consideration of the recognition the participation of a specifi c receptor, the B cell and the immune response to free, unconjugated receptor or BCR, and co-stimulation from T drugs is set out in Sect. 3.4 below. helper cells. The BCR has immunoglobulin anchored in the cell membrane, and, in concert with the B cell co-receptor complex, it is the 3.2 IgE Antibodies interaction between this surface immunoglobu- and IgE- Mediated Drug lin and its complementary antigen that initiates Hypersensitivities B cell activation. Upon binding to the antigen, the BCR–antigen complex is internalized within The central importance of IgE antibodies in both an endosome, processing follows, and the pro- the immediate and late phases of an allergic cessed antigen is presented back on the surface response involving infl ammatory reactions is well by MHC type II molecules. If maturation of the established. IgE mediates the allergic infl amma- B cell to a plasma cell or a memory cell is to tory response by binding to both its high-affi nity continue, interaction with, and co- stimulation receptor FcεRI on mast cells and basophils and its by, an activated T helper cell is required. low-affi nity IgE receptor FcεRII (CD23) (on a Interaction between the B cell and an activated number of different hematopoietic cells including Th2 cell with the appropriate TCR involves rec- B cells) to augment humoral and cellular responses. ognition of the MHC-processed antigen by the TCR and co-stimulatory CD80/CD86 (B7) signals. Co-stimulation of the B cell that eventually 3.2.1 Initiating Events in the leads to clonal expansion and isotype switching Production of IgE Antibody is also enabled through upregulation of the CD40 ligand (CD40L) (Fig. 3.3 ). If CD40L-CD40 IgE is produced by plasma cells at the site of an receptor interaction and co- stimulation do not allergic reaction generally in mucosal, cutane- eventuate, B cells undergo apoptosis and are ous, and gut lymphoid tissue. IgE antibody pro- eliminated. Cell proliferation and isotype switch- duction begins with the interaction between ing for the synthesis of IgE are aided by the cyto- antigen-bearing antigen-presenting cells (APC) kines Il-4 and IL-13 generated by Th2 cells. and lymphocytes. APCs can be dendritic cells, These two cytokines initiate transcription of the most important cell in initiating the adaptive germ-line mRNA for IgE antibodies and are immune response, macrophages, and B cells. regarded as the fi rst of two signals necessary for Naïve T lymphocytes not only need to have anti- class switching from IgM- to IgE-bearing cells. gen presented in a special way, they also require The second signal is delivered by the interaction precise signals to become activated. Both of of CD40L on the T cell surface with its receptor these requirements are fulfi lled by the APC CD40 on the B cell. This interaction results in all fi rstly via the membrane-associated MHC that of the elements necessary for the ε-heavy chain interacts with the T cell receptor (TCR) (activa- being brought into close proximity. tion signal 1) and secondly by the provision of IgE levels infl uence IgE receptor density on co-stimulatory signals in the form of the mem- cells. High levels of antibody increase both the brane protein ligand CD80 (B7-1) working in number of FcεRI receptors and the degranulation tandem with another membrane ligand CD86 of mast cells and basophils. Along with degranu- (B7-2) (activation signal 2). These ligands inter- lation, increased release of cytokines such as IL-4 act with their complementary receptor CD28 occurs and these in turn stimulate increased IgE constitutively expressed on naïve T cells to allow levels and receptor density. A reduction of IgE the cells to undergo clonal expansion (Fig. 3.3 ). results in a reduction of receptor levels on mast

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 43

T cell Dendritic cell B cell Co-stimulatory B7 signals Allergen

Allergen captured B cell receptor & internalized CD28 Stimulation B7 (CD80/CD86)

TCR Activation of B cell MHC+ processed peptide CD40 Activation of T cell by dendritic cell MHC

B cell stimulation of T cell Co-stimulatory B7 signals

Fully activated B cell CD28 Stimulation B7 undergoes TCR MHC+peptide clonal expansion CD40L and class switching Upregulates CD40 CD40

T cell Cytokines B cell eg. IL-4, IL-13

Fig. 3.3 Cellular events in the production of IgE anti- Antigen presentation to activated T cells by activated B bodies. Presentation of antigen (usually in peptide form) cells ultimately results in co-stimulation of the B cells, to T cells via MHC molecules on dendritic cells. This class switching, clonal expansion, and differentiation to results in the T cells undergoing clonal expansion. effector cells cells and decreased degranulation. IgE is also differentiate into IgE- secreting plasma cells and capable of upregulating the FcεRII receptor (see Th2 cells and their functions in the allergic below). From the therapeutic aspect then, inhibi- response are inhibited. Modulation of cytokines tion of IgE is desirable since it leads to a decrease involved in the production of IgE is yet another in the release of mediators from mast cells and therapeutic strategy. For example, IL-12 and basophils. This is, in fact, the rationale for the use IFN-γ inhibit cytokine production by Th2 cells of omalizumab, a recombinant humanized IgG1k so interference with the expression of these monoclonal antibody (see Sect. 11.1.3.1 ) used for cytokines suppresses IgE synthesis. For further patients with difﬁ cult-to- manage severe persistent biologic strategies in directing therapies for hyper- allergic asthma. Omalizumab binds to the Cε3 sensitivities, see Chap. 11 . region of circulating human IgE antibodies inhibiting their binding to the FcεRI and FcεRII receptors and thus ultimately suppressing IgE-mediated 3.2.2 Allergic Release of Mediators mast cell activation and the allergic inﬂ ammatory of Hypersensitivity from response. It does not target receptor-bound IgE on Mast Cells mast cells and thus does not trigger mast cell degranulation. Another potential therapeutic The critical role of IgE in both the immediate approach to treat allergic disorders is the interfer- and late phases of the allergic response is well ence with the interaction between IL-4 and its established and, together with the mast cell, the receptor. Without this interaction, B cells do not resultant humoral and cellular interactions produce

[email protected] 44 3 Mechanisms of Hypersensitivity

Antigen cross-links IgE antibody electron adjacent antibodies dense FceRI granules

granules (less electron dense)- contents released & open to exterior

Activated mast cell

Resting mast cell containing granules and inflammatory mediators Mediators of hypersensitivity and inflammation

Contracts Dilates and increases Stimulates Attracts Activates airways permeability of blood secretions of eosinophils platelets smooth muscle vessels mucous glands

Fig. 3.4 Diagrammatic representation of antigen- cells by antigen-effected cross-linking of adjacent cell- induced degranulation of, and mediator release from, mast bound complementary IgE antibodies

the infl ammatory mediators and symptoms surfaces (Fig. 3.4 ). The high affi nity of the recep- −10 characteristic of allergic reactions. On the basis tor (~ Ka 10 M) means that a high proportion of of the type of proteases and proteoglycans in IgE is bound even in situations where there are their granules, human mast cells can be divided low levels of circulating IgE antibodies. The into three populations: tryptase-only positive FcεRI complex is a receptor in tetramer form mast cells in the lungs and intestinal mucosa; made up of a ligand-binding α chain structurally tryptase, chymase, and carboxypeptidase positive related to the α chains of FcγR, a tetraspan β mast cells in the skin, connective tissues, and chain, and the FcγR γ chain dimer. The α chain intestinal mucosa; and a smaller population of has two protruding Ig type domains that bind the chymase-only positive cells in the nasal and Cε3 region of IgE and in the presence of the anti- intestinal mucosae. For more details of tryptase body the receptor is upregulated while the Fc and its importance as a diagnostic marker for receptor for IgG is downregulated. The β and γ anaphylaxis, the reader is referred to Sect. 4.5.1 . chains each contain an ITAM (Immunoreceptor The initial event in the activation of mast cells for Tyrosine-based Activation Motif) that interact mediator release is the binding of IgE antibodies with the Lyn, Syk, and Fyn protein tyrosine to the high-affi nity FcεRI IgE receptor abun- kinases. The critical event and signal for mediator dantly expressed on the mast cell and basophil release, as occurs in anaphylaxis, is the cross-linking

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 45 of receptor-bound IgE antibodies by allergen basophils. Cross-linkage of antibodies by antigen molecules reacting with the bivalent antibody that reacts with the combining sites of adjacent combining sites. The IgE–FcεRI complex is receptor-bound IgE molecules activates the long-lasting and dissociates exceptionally slowly. receptors and triggers the cells to express CD40 Cross-linking of receptors causes their aggrega- ligand (CD40L) and secrete IL-4. These mole- tion, rapid migration to lipid rafts, activation of cules react with their complementary receptors the Lyn and Fyn protein tyrosine kinases, and expressed on the B cell surface, and hence, like ultimately transphosphorylation of the β and γ Th2 cells, mast cells and basophils can induce chains and involvement of the Syk kinase. Mast class switching and increase the production of cell degranulation (Fig. 3.4 ), which can occur IgE antibody. within seconds, follows a series of activation steps induced by phosphorylation reactions discussed in more detail in Sect. 3.2.6 . Some of the 3.2.4 Low-Affi nity IgE Receptor released mediators of infl ammation and anaphy- FcεRII (CD23) laxis stored in the cytoplasmic granules including histamine (see below, Sect. 3.2.5.1 ), heparin, A second receptor for IgE, the low-affi nity recep- platelet-activating factor (PAF) (Sect. 3.2.5.3 ), tor FcεRII also known as CD23, is expressed on serotonin, the enzymes tryptase, chymase, and airways smooth muscle cells and several types of carboxypeptidase, and eosinophil, neutrophil, hematopoietic cells including mature B lympho- and monocyte chemotactic factors are preformed cytes, macrophages, monocytes, dendritic cells, while others are newly synthesized. The pre- and eosinophils. The designation “low” affi nity is formed mediators are responsible for the imme- derived from the receptor’s lower affi nity −7 −8 diate signs and symptoms of vasodilation, edema, ( KD = ~10 –10 ) than the affi nity of the FcεRI −10 −11 bronchoconstriction, and itching. The newly syn- receptor (K D = ~10 –10 ). CD23 has multiple thesized group of released mediators includes functions by virtue of its capacity to bind a range prostaglandin D2 (PGD2) , thromboxanes, and leu- of different ligands. As well as binding IgE in kotrienes LTB4 , LTC4 , and LTD4 (Sect. 3.2.5.2 ). both its secreted and B cell-bound form, CD23 A host of cytokines (pro- and anti-infl ammatory), binds CD21 (also known as complement receptor chemokines, and chemotactic, stimulating, and 2), CD18/CD11b (complement receptor 3), growth factors including interleukins -1, -3, -4, CD18/11c (complement receptor 4), and α v β3 , the -5, -6, -8, -9, -10, -11, and -13, tumor necrosis vitronectin receptor. CD23 is a 45 kD type II factor (TNF), granulocyte–macrophage colony- membrane protein with homology to calcium- stimulating factor (GM-CSF), monocyte chemo- dependent (C-type) lectins. It is involved in both tactic protein-1 (MCP-1), regulated upon the up- and downregulation of IgE synthesis by B activation normal T cell expressed and secreted cells, augmentation of humoral and cellular (RANTES; CCL5), and eotaxin (CCL-11) are responses, and facilitation of the phagocytosis of also released. IgE opsonized antigens. Upon antigen-mediated cross-linking of bound IgE, the low-affi nity receptor on B cells downregulates IgE synthesis. 3.2.3 Amplifi cation of IgE Antibody Augmentation of IgE-mediated responses can be Production by Cellular demonstrated in vivo by the prevention of an Interaction immunogen and antigen-specifi c IgE-induced increase in serum IgE titers following pretreat- Mast cells, basophils, and even dendritic cells ment with anti-CD23 antibodies. As well as its can accentuate B cell production of IgE antibod- effects on the FcεRI receptor, IgE can also upreg- ies by direct interaction (Fig. 3.5 ). IgE antibodies ulate CD23 resulting in an increased allergic newly synthesized by plasma cells bind to the response in the bronchial mucosa. This is thought FcεRI receptors on the surfaces of mast cells and to occur via enhancement of allergen uptake and

[email protected] 46 3 Mechanisms of Hypersensitivity

IgE secreted by plasma cell

IgE antibody Antigen

IL-4 IL-4R

FceRI

CD40L CD40 Bound antigen cross-links Stimulated IgE molecules IgE production Mast cell B cell

Fig. 3.5 Amplification of IgE antibody production their complementary receptors on the B cell surface by B cells by direct interaction of mast cells express- inducing class switching and the production of IgE ing CD40L and secreting IL-4. These interact with antibodies

presentation. Considering the effects of IgE on controlled by sCD23, and further, sCD23 binds the high and low IgE receptors, inhibition of the to cells co-expressing IgE and membrane CD21. antibody leads to downregulation of both recep- These results have been interpreted as membrane- tors and ultimately decreased mediator release bound IgE and CD21 having a role in the sCD23- from mast cells and basophils. mediated positive regulation of IgE synthesis Important ﬁ ndings on CD23 control of IgE with feedback occurring when the concentration antibody synthesis and homeostasis in human B of IgE becomes great enough to allow binding to cells have recently been forthcoming. The endog- mCD23, thus preventing further release of its enous metalloprotease, a disintegrin and metal- soluble form. loproteinase domain-containing protein 10 (ADAM10), demonstrated the existence of two forms of CD23 by releasing its soluble form 3.2.5 Important Mediators of the (sCD23) from membrane CD23 (mCD23). Type I Immediate Allergic Upregulation of mCD23 in tonsil B cells follow- Response ing treatment with Il-4 and anti-CD40 led to accumulation of sCD23 in the medium prior to 3.2.5.1 Histamine class switching to IgE synthesis. Inhibition of The reader is also referred to Sect. 4.5.2 for a mCD23 cleavage by an inhibitor of ADAM10 or consideration of the place of histamine in the small interfering RNA inhibition of CD23 syn- diagnosis of drug allergies and to Sect. 8.4.1 for a thesis suppressed IL-4- and anti-CD40-induced summary of histamine receptors and their rele- IgE synthesis, but addition of recombinant vance to opioid analgesics. sCD23 enhanced IgE synthesis. Since this Histamine (2-(imidazol-4-yl)ethylamine) is occurred even when mCD23 is protected from one of the most intensely studied molecules in cleavage, it seems that IgE synthesis is positively all biological systems. This fact, and its appar-

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 47 ent myriad physiological and pathological This is achieved by methylation and oxida- effects, is behind a seemingly ever-expanding tion. In mammals, histamine is inactivated in literature on an extraordinarily broad spread of two main ways—methylation of the imidazole activities including its role in infl ammatory and ring effected by histamine N-methyltransferase allergic reactions; many aspects of the immune (HMT) and oxidative deamination of the pri- response; differentiation; cell proliferation; mary amino group catalyzed by diamine oxidase hematopoiesis; neurotransmission; regulation of (DAO)—to form N -methylhistamine and circulatory functions, vasodilation, and blood imidazole-4-acetaldehyde, respectively (Fig. 3.6 ). pressure; wound healing; gastrointestinal func- HMT, which is specifi c for histamine, is present tion; and, no doubt, numerous others yet to be in most tissues and responsible for the inactiva- elucidated. In peripheral tissues, more than tion of intracellular histamine. The enzyme cata- 90 % of body stores of histamine are found in lyzes the transfer of a methyl group from mast cells and basophils, although there are two S -adenosyl-L -methionine to the secondary amino other main sources in humans—enterochromaf- group of the imidazole ring. DAO is stored in fi n-like cells of the gut and histaminergic nerves secretory vesicles and expressed mainly in intes- in the brain. In mast cells and basophils, hista- tinal and kidney epithelial cells. Its release is mine is stored in granules in association with stimulated by heparin which is liberated together different anionic proteoglycans—heparin in with histamine by activated mast cells. Heparin mast cells and condroitin-4-sulfate in basophils. terminates the action of histamine by inactivat- Upon degranulation elicited by specifi c IgE ing it locally. DOA is also active in the gut where antibodies, cytokines, or histamine releasers it catabolizes histamine present in some foods, like compound 48/80, calcium ionophore, thus preventing it from entering the circulation. N-formyl-met-leu-phe, phorbol 12-myristate The products of histamine inactivation by the 13-acetate, and some drugs such as opioid anal- two different routes are further metabolized gesics and neuromuscular blockers, histamine is (Fig. 3.6). N -methylhistamine is converted to released from the granules in large amounts N- methylimidazole-4-acetaldehyde by mitochon- with the associated proteoglycan. drial monoamine oxidases and this aldehyde, in turn, is catalyzed by aldehyde dehyrogenases to 3.2.5.1.1 Histamine Biosynthesis and N -methylimidazole-4-acetic acid. In the DAO Metabolism pathway, the fi rst product from the breakdown of Histamine is synthesized from L -histidine exclu- histamine, imidazole-4-acetaldehyde, is also cata- sively by the inducible enzyme L -histidine lyzed to the acetic acid derivative by aldehyde decarboxylase located in the cytosol and widely dehydogenase before its subsequent ribosylation expressed in the body in various cells including for transport and excretion. mast cells, basophils, parietal cells, gastric mucosa, neurons, and cells of the central nervous 3.2.5.1.2 Histamine Receptors system. The mammalian enzyme requires The physiological and pharmacological effects of pyridoxine-5-phosphate as an active site cofac- histamine are mediated through four different tor (Fig. 3.6 ). Once synthesized, histamine is receptors H 1 , H 2 , H3 , and H 4 , all members of the transported from the cytosol to the secretory 7-transmembrane g protein-coupled receptor granules by vesicular monoamine transporter 2 (GPCR) family with amino terminal glycosylation (VMAT2). L-Histidine decarboxylase is detect- sites and phosphorylation sites for protein kinases able only in cells producing histamine since it is A and C. The receptors are widely expressed on synthesized only when the mediator is required different tissues that are responsive to histamine. and degraded as soon as synthesis is terminated. For the H 1 receptor these tissues include smooth Given histamine’s pronounced physiological muscle cells of the airways and vasculature, the gas- actions, its inactivation to metabolites that do not trointestinal tract, cardiovascular system, neutrophils, interact with histamine receptors is a requirement. endothelial cells, T and B cells, hepatocytes, nerve

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O N OH

NH HN 2 L-histidine

pyridoxine-5-PO4 L-histidine decarboxylase

N NH2

HN histamine

histamine N-methyltransferase (HMT) diamine oxidase (DAO)

NH N 2 N H

O N HN

H3C N-methylhistamine imidazole-4-acetaldehyde

monoamine oxidase (MAO) aldehyde dehydrogenase

N H N OH

O O N HN

H3C N-methylimidazole-4-acetaldehyde imidazole-4-acetic acid aldehyde dehydrogenase ribosyl transferase

N OH

N OH O N O N O OH HO H3C OH N-methylimidazole-4-acetic acid N-ribosylimidazole-4-acetic acid

Fig. 3.6 Biosynthesis of histamine from L -histidine by the and its metabolism by methylation (via histamine widely expressed enzyme L -histidine decarboxylase N-methyltransferase) and oxidation (via diamine oxidase)

cells, and cells of the genitourinary system sug- systems while H 4 receptors are largely expressed gesting an important role for the autacoid in the in hemopoietic cells where they modulate eosino- modulation of immune, inﬂ ammatory, and allergic phil migration and selective recruitment of mast processes. The H2 receptor is expressed in gastric cells. For signal transduction, the H1 and H2 recep- parietal cells, the central nervous system, vascular tors activate Gq and G s -coupled proteins respec- smooth muscle, heart, neutrophils, and uterus. H3 tively while both H3 and H4 are coupled to, and receptors appear to be less widely distributed activate, Gi/o proteins. occurring in the central and peripheral nervous

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 49

Pathophysiological effects resulting from The H 3 receptor regulates the synthesis and stimulation of the H 1 receptor include those release of histamine and also has a regulatory responses seen in immediate allergic reactions, role in the release of neurotransmitters such as viz, redness, itch, swelling, asthma, anaphy- serotonin, dopamine, and norepinephrine. The laxis, bronchoconstriction, and vascular permea- receptor is expressed in those regions of the cen- bility. The primary activation of the H 1 receptor, tral nervous system associated with cognition, in a Gα q/11 -coupled protein, proceeds through particular, the hippocampus, basal ganglia, and phospholipase C which catalyzes the formation cortical areas, and in the peripheral nervous sys- of inositol-1,4,5-triphosphate (IP3 ) and 1,2- tem, namely, the cardiovascular system, gastroin- diacylglycerol (DAG) from phosphatidylinositol testinal tract, and airways. The H 3 receptor

4,5-biphosphate. IP3, released into the cytosol, signals through G i/o proteins and alternative sig- binds to its receptor in the endoplastic reticulum naling pathways appear to be activated by these causing an increase in cytosolic Ca2+ levels. proteins. Stimulation of the receptor results in DAG, acting as a second messenger, activates adenyl cyclase inhibition and lower levels of protein kinase C (PKC). This pathway is acti- cAMP and PKA. Alternative signaling pathways vated and proceeds in the brain, airways, and may be activated including activation of phos- intestinal and vascular smooth muscle. H 1 recep- pholipase A2 (PLA 2), stimulation of mitogen- tor activation in some other tissues can stimulate activated protein kinase (MAPK), the inhibition adenyl cyclase and cAMP formation. The signal- of Na+ /H+ exchange, and K + -induced Ca2+ mobi- ing pathways are not yet fully understood, par- lization. A study of H3 receptor-mediated attenu- ticularly details of the involvement of Ca 2+ . Some ation of norepinephrine exocytosis in cardiac of the resultant responses in vascular endothelial sympathetic nerves identiﬁ ed a novel pathway in cells after stimulation of the H 1 receptor and ele- which stimulation of the receptors on nerve end- vated intracellular Ca2+ levels are permeability ings produces intraneuronal activation of the changes, synthesis of prostacyclin and platelet- MAPK cascade. PLA2 , phosphorylated by activating factor (PAF), and release of Von MAPK, translocates to the cell membrane where Willebrand factor and nitric oxide (NO). it acts on membrane phospholipids producing

Whereas H1 receptors are involved with posi- arachidonic acid, the substrate for cyclooxygen- tive effects, H 2 receptors appear to mainly medi- ase and the production of prostaglandin E 2 ate suppressive activities of histamine including (PGE2 ). PGE2 activates prostaglandin E receptor gastric acid secretion, heart contraction, cell pro- 3 (EP3 R) on the cell membrane where the Gβγ i 2+ liferation, differentiation, and some effects on the subunit of EP3 R inhibits Ca entry resulting in immune response. H2 receptors are coupled to the attenuation of norepinephrine exocytosis. It is adenylate cyclase as well as the phosphoinositide apparent that with the H3 receptor, different sig- second messenger systems via separate GTP- naling can be employed in different cell systems. dependent mechanisms, but H2 -dependent A further illustration of this is the demonstration effects, particularly those of the central nervous of H 3 receptor-mediated activation in the inhibi- system, are predominantly mediated through tion of the growth of cholangiocarcinoma in vitro cAMP. It has been shown that receptor binding and in vivo. Activation of H3 receptors by a high- stimulates activation of c-Fos, c-Jun, PKC, and afﬁ nity H 3 agonist decreased cholangiocarci-

P70S6 kinase. Alternative signaling pathways noma growth by increasing levels of IP 3 , 2+ have been reported (Fig. 3.7 ). These include a translocation of PKCα, and IP 3 /Ca -dependent receptor-mediated increase in intracellular Ca2+ dephosphorylation of the extracellular signal- and/or IP 3 levels in HL-60 human promyelocytic regulated kinases ERK 1/2. leukemia cells and an increase in cAMP and inhi- A new signaling pathway of the H3 receptor bition of release of arachidonic acid in Chinese involving receptor modulation of the activity of hamster ovary (CHO) cells transfected with rat the serine/threonine-speciﬁ c protein kinase Akt cDNA and induced by calcium ionophore. (protein kinase B, PKB)/GSK-3β (glycogen

[email protected] 50 3 Mechanisms of Hypersensitivity

HISTAMINE N G protein-coupled receptor

α C β γ

RECEPTORS H1 H2 H3 H4 α α α α GTP q GTP s GTP i/o GTP i/o

Main signaling PLC-β IP3 +DAG AC cAMP AC cAMP AC cAMP pathway 2+ Ca PKC PKA PKA PKA

Intracellular MAPK PLA2 AA PLCIP3 DAG Alternative 2+ 2+ AC cAMP Ca & IP PGE EP3R Ca pathways 3 2 Ca2+ in HL-60 cells in guinea-pig cardiac synaptosomes & heart in mouse mast cells cAMP & AA High affinity agonist Agonist-biased signaling: , 2+ b in CHO cells IP3 PKCα IP3/Ca - recruitment of -arrestin dephosphylation of ERK1/2 independent of G protein in human cholangiocarcinoma activation in human osteosarcoma cell line U20S Modulation of Akt/GSK-3β axis in human neuroblastoma

Fig. 3.7 Summarized comparisons of G protein coupling and main and alternative signaling pathways for the four histamine receptors H1, H2, H3 , and H4. (see also Table 3.1 )

synthase kinase 3β) axis was recently demon- Following the realization that not all of the strated in SK-N-MC cells from a neuroepitheli- biological effects of histamine could be attributed oma cell line. Receptor stimulation with an H3 to histamine receptors H1 , H2 , and H3, a fourth agonist induced the phosphorylation of Ser473 receptor was postulated and histamine receptor and Thr308 on Akt, a kinase important for neuro- H 4 was subsequently cloned in 2000–2001. nal development and function. Studies suggested Receptor H4 shows a 35 % amino acid sequence that the Akt activation occurs via a Gi/o -mediated homology with the H 3 receptor and the two are activation of PI3K (see Sect. 3.2.6.1 ). H3 receptor similar in gene structure. The receptor, essentially activation also resulted in phosphorylation of Ser confi ned to hemopoietic cells, exerts a chemotac- 9 on GSK-3β, a ser/thr kinase which acts down- tic effect on several cell types associated with stream of Akt. This kinase is important in brain immune and infl ammatory responses such as function and this newly identifi ed signaling path- allergy, asthma, rheumatoid arthritis, and infl am- way adds important knowledge to our under- matory bowel disease and this has led to interest standing of the role of H3 receptor-controlled in the development of new agents targeting these histamine in brain function. The three above- diseases. H4 receptors are functionally expressed outlined alternative pathways are summarized in on mast cells, eosinophils, monocytes, dendritic Fig. 3.7 . cells, and CD8+ T cells. Although the presence

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 51

Table 3.1 Summarized comparison of function, G protein coupling and signaling pathways of histamine H 1 , H 2 , H 3 , a and H 4 receptors

Receptor H 1 H 2 H3 H4 Best characterized function Acute allergic Gastric acid secretion Modulation of Immuno- reaction neurotransmitters modulation Indications for antagonists Allergy, Gastroesophageal reﬂ ux Sleep and cognition Pruritus, pruritus b disease, peptic ulcerb disordersc asthmac

G protein coupling Gα q/11 G αs Gα i/o Gα i/o Main signaling pathway Ca2+ ↑ cAMP ↑ Inhibition of cAMP Ca2+ ↑ a See also Fig. 3.7 b Approved indications c Potential indications

of large amounts of histamine in mast cells and result seen in mice deﬁ cient in the H4 receptor. the cell’s histamine-releasing properties are well Interestingly, expression of the H4 receptor on known, expression of histamine receptors on mast cells or any other cell was not required for mast cells had not been convincingly demon- the pruritic activity, leading to the speculation strated and there has been little information on that H4 receptor-mediated pruritus may result the effect of histamine on the cell. It is now from actions on peripheral neurons. While the known that mast cells express the H4 but not the relevance to pruritus in humans of the results

H3 receptor, but exposure to histamine, or hista- with animal models is uncertain, there is opti- mine in combination with antigen–IgE antibody mism that antihistamines specifi cally targeting complexes, does not lead to degranulation of the H4 receptor may lead to more effective treat- mast cells. The H4 receptor has, however, been ment of pruritic conditions in humans. clearly implicated in infl ammation and pruritus The H4 receptor is mainly coupled to G i/o pro- in animal models. In a rat model of carrageenan- teins and, in common with the H3 receptor, this induced acute infl ammation, antagonists of the leads to inhibition of adenyl cyclase and receptor inhibited edema formation and reversed decreased production of cAMP and downstream the thermal hyperalgesia. In a histamine-induced effects on cAMP response element-binding itch model in mice, H4 antagonists inhibited but (CREB) gene transcription. As with the other his- did not abolish scratching and itch was reduced tamine receptors, other signaling pathways have in H4 -defi cient mice. Centrally acting H1 receptor been reported (Fig. 3.7 ). From a study of the sig- antagonists produced a partial reduction and naling pathways of the endogenous mouse H4 combined treatment with both antagonists com- receptor of bone marrow-derived mast cells, his- pletely eliminated itch. Further evidence for the tamine activation of the receptor was shown to involvement of both H4 and H1 receptors in induce chemotaxis without affecting degranula- histamine-induced itch was the production of itch tion of the mast cells. The following interpreta- following administration of agonists of both tions and sequence of events were suggested. receptors. There are many mediators of itch and Binding of histamine to the receptor on mast cells mechanisms are complex. With the belief that the and eosinophils activates the pertussis toxin- mechanisms underlying itch in chronic condi- sensitive Gαi/o proteins triggering PLC possibly tions such as atopic dermatitis are more likely via the G protein βγ subunits dissociated from the those associated with mast cell degranulation, a Gα i/o proteins. PLC hydrolyzes phosphatidylino- mouse model of itch was set up by injecting anti- sitol 4,5-biphosphate to IP3 and DAG. IP3 dif- gen-specifi c IgE intradermally and challenging fuses into the cytosol and binds to its receptor on with antigen 24 h later. H 4 receptor antagonists the endoplastic reticulum where it activates a signifi cantly reduced itch and this was also the Ca 2+ channel causing the release of intracellular

[email protected] 52 3 Mechanisms of Hypersensitivity

Ca2+ . The increased Ca2+ triggers mast cell che- pounds are synthesized by leukocytes, they motaxis toward histamine by pathways yet to be contain three conjugated double bonds or alkenes, worked out. It has been suggested that this mech- and four members of the group, LTC4, LTD4 , anism might be responsible for mast cell accu- LTE 4 , and LTF4 , contain the amino acid cysteine. mulation in allergic tissues. Although the leukotrienes were originally identi- There is mounting evidence that when the ﬁ ed by their contractile effect on smooth muscle, same receptor can activate more than one path- they are now recognized as potent inﬂ ammatory way, some agonists can activate one pathway in mediators with a range of other biologic effects. preference to another. The need to consider more In particular, LTC4 and LTD4 are powerful medi- than one downstream signaling pathway in ators of asthma, airway hypersensitivity, and histamine- GPCR studies was again reinforced by allergies, inducing bronchoconstriction, increas- a recent investigation of signaling at the H4 recep- ing vascular permeability, and promoting mucous tor using the selective antagonist for G secretion. Upon inhalation, both mediators are up protein- dependent signaling JNJ7777120 to 1,000 times as potent as histamine whereas

(1-[(chloro-1-H -indol-2-yl)carbonyl]-4-methyl- LTE4 is only 39 times as potent as histamine in piperazine). Downstream signaling measure- reducing maximum expiratory ﬂ ow at 30 % of ments of G protein activation and β-arrestin vital capacity. LTE4 , the most stable of the three recruitment demonstrated that the antagonist is cysteinyl leukotrienes, is present in greatest what has been described as a biased agonist, act- amount in vivo where it induces bronchial eosin- ing as an agonist in a non-G protein-dependent ophilia and airway hyperresponsiveness. Unlike manner to recruit β-arrestin to the receptor. LTC4 and LTD 4 , LTE 4 persists longer in serum, β-Arrestin is part of the mechanism for regulating urine, and bronchoalveolar lavage ﬂ uid of asth- the activity of GPCRs. In stabilizing an alterna- matics. Urinary excretion of LTE4 is therefore tive active conformation of the H4 receptor that sometimes used as an indicator of asthma. The initiates β-arrestin recruitment but not G protein bronchoconstriction provoked by LTE 4 is strong activation, that is, agonist-biased signaling, in patients with aspirin-sensitive asthma but

JNJ7777120 may be exhibiting the capacity to much weaker in other asthmatics, whereas LTD4 exist in multiple active conformations. This may is much more pronounced in asthmatic patients result in an agonist stabilizing a slightly different not sensitive to aspirin (see Chap. 9 ). Another state that preferentially couples to one pathway difference between the two mediators in their and not another. effects on asthmatics is the recruitment into spu- Summarized comparisons of functions, indi- tum of basophils, mast cells, and eosoniphils by cations for antagonists, G protein coupling, and LTE 4 but not by LTD 4 . LTD4 aids the adhesion signaling pathways for the four histamine recep- and migration of some cancer cells and increases tors are shown in Table 3.1 and Fig. 3.7 . proliferation of mast cells. All three cysteinyl leukotrienes produce an equiactive wheal and 3.2.5.2 Cysteinyl Leukotrienes ﬂ are reaction characteristic of an allergic response For a discussion of cysteinyl leukotrienes in rela- when injected intradermally at a concentration of tion to diagnostic investigations of suspected 1 nmol per site. drug allergies, see Sect. 4.5.3 . Originally isolated after stimulation of lung 3.2.5.2.1 Biosynthesis tissue by histamine and snake venom and named As part of the response to leukocyte cell activa- over 70 years ago as “slow reacting substance of tion, cysteinyl leukotrienes are generated de novo anaphylaxis,” (SRS-A), leukotrienes are a family from arachidonic acid liberated from cell mem- of bioactive peptide-conjugated eicosanoid lipids brane phospholipid by cytosolic phospholipase produced by mast cells, basophils, eosinophils, A2 (Fig. 3.8 ). In concert with 5-lipoxygenase- and macrophages. The name “cysteinyl leukotri- activating protein (FLAP), the enzyme enes” is derived from the facts that the com- 5-lipoxygenase (5-LO) converts arachidonic

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 53

Cellular Membrane cPLA2

COOH

Arachidonic acid (5,8,11,14-Eicosatetraenoic acid) 5-lipoxygenase- 5-lipoxygenase (5-LO) activating protein (FLAP)

OOH OH COOH COOH

5-HPETE 5-HETE (5-Hydroxyperoxyeicosatetraenoic acid) (5-Hydroxyeicosatetraenoic acid)

5-LO (LTA4 synthase) OH OH O COOH COOH

LTA4 hydrolase synthaseS LTA4 (LTA4H) LTB4 4 4 Glutathione LTC(LTC ) Glutamic acid Glycine OH OH OH COOH COOH COOH

SCysGly γ-Glutamyl S Cys Gly Dipeptidase SCys LTC Glu transpepti- LTD LTE 4 dase (γ-GT) 4 4 or γ-Glutamyl leukotrienase (γ -GL)

Fig. 3.8 Biosynthesis of cysteinyl leukotrienes from arachidonic acid showing the pathways to the formation of LTA4, LTB 4 , LTC4 , LTD4 , and LTE4

acid to 5-hydroxyperoxyeicosatetraenoic acid express LTC4 synthase, LTA4 is conjugated to (5-HPETE) which spontaneously reduces to reduced tripeptide glutathione to form the cyste-

5-hydroxyeicosatetraenoic acid (5-HETE). 5-LO inyl leukotriene LTC 4. After transportation to the converts 5-HPETE to leukotriene A4, an unstable cell surface in an energy-dependent step with the peroxide. Note that the enzyme involved in this assistance of multidrug resistance-associated step is sometimes referred to as LTA4 synthase. protein 1 (MRP-1), LTC 4 is converted extracel-

LTA 4 synthase activity co-puriﬁ es with 5-LO and lularly to LTD4 by a γ-glutamyl transpeptidase the same cytosolic and membrane-bound active (γ-GT) or γ-glutamyl leukotrienase (γ-GL). In proteins are required for reactions catalyzed by the ﬁ nal step in the pathway, a dipeptidase

5-LO and the so-called LTA4 synthase in crude removes glycine from LTD4 producing LTE 4 human leukocyte homogenates leading to the which is excreted unchanged in the urine. LTF4 conclusion that a single enzyme is responsible which has an S-glutamylcysteinyl group has for the production of 5-HPETE from arachidonic been prepared in vitro from LTE 4 with glutathi- acid and for its subsequent conversion to LTA4 . In one and γ-glutamyltranspeptidase but, as yet, it neutrophils and monocytes which have the has not been found in vivo. In comparison to the enzyme LTA4 hydrolase, LTA4 is converted to the other cysteinyl leukotrienes, LTF 4 contracts vas- dihydroxyacid leukotriene LTB4 , a chemoattrac- cular smooth muscle poorly—the rank order of tant for neutrophils, whereas in mast cells, baso- potency being LTD4 > LTC4 > LTE4 >> LTF4 . phils, eosinophils, and macrophages, all of which Although leukotriene synthesis generally proceeds

[email protected] 54 3 Mechanisms of Hypersensitivity via the 5-LO pathway, a second family of tially studied with prototypes of the later-to-be-

leukotrienes (eoxins, given the preﬁ x EX) can be developed “lukast” antagonists. CysLT1 R, which generated from the action of 15-LO (and 12-LO) bound LTD4 much more strongly than LTC 4 , was ﬁ rst on arachidonic acid and then, for the competitively blocked by the antagonists while

15- lipoxygenase compounds, 15-HPETE to CysLT2 R bound the two cysteinyl leukotrienes form the 15-epoxytriene 15-LTA 4 (EXA4 ) fol- with equal affi nity and bound LTD4 at one-tenth lowed by the pro-infl ammatory cysteinyl 15-leu- the affi nity shown by CysLT1 R. LTE4 , however, kotrienes 15-LTC4 (EXC4 ), 15-LTD4 (EXD4 ), did not display any appreciable binding to either and 15-LTE4 (EXE4 ) in eosinophils, mast cells, receptor. Despite this, some early recognized and nasal polyps of allergic subject (see also pharmacologic properties of LTE4 , viz, its supe- Sects. 9.4.1 , 9.4.3 and Fig. 9.3 ). IL-4-primed rior potency to its related compounds in contract- human mast cells incubated with arachidonic ing guinea pig tracheal smooth muscle and acid synthesize and release EXC 4 and possess enhancement of this effect produced by hista- the capacity to produce EXD4 cells while nasal mine, its known peripheral and central airway polyps spontaneously release EXC4. Eoxins effects in guinea pigs, and its capacity to increase modulate and enhance vascular permeability, permeability in guinea pig and human skin, all being 100 times more potent in this respect than suggested a distinct pathobiologic role and the histamine and almost as potent as LTC4 and existence of a distinct receptor for LTE4. Studies

LTD 4. Two types of the 15-LO enzyme are by K. Frank Austen’s group of cysteinyl leukotri- known, 15-LO-1 (which also has about 10 % ene-dependent swelling of ear tissue in mice 12-lipoxygenating activity) and 15-LO-2, both lacking both CysLT receptors proved the exis- of which produce 15(S)-HETE from arachidonic tence of a distinct LTE4 -reactive cutaneous recep- acid but 15-LO-1 oxygenates arachidonic acid at tor. Ear swelling, a measure of LTE 4 - mediated carbons 15 and 12 while 15-LO-2 adds oxygen vascular leakage, was inhibited by pretreatment only at carbon 15. Human eosinophils and air- with pertussis toxin or a Rho kinase inhibitor, ways epithelial cells contain high amounts of indicating the involvement of a human GPCR to

15-LO-1 as do some subsets of human mast Gα i proteins and Rho kinase. Until this cutaneous cells, macrophages, and dendritic cells. receptor is cloned, it has been designated

Expression of 15-LO-2 appears to be restricted CysLTE R. Further studies of single receptor- to lung, skin, prostate, and cornea. deﬁ cient strains of mice compared to wild-type mice showed that the permeability response to −/− 3.2.5.2.2 Cysteinyl Leukotriene Receptors LTC 4 or LTD4 was reduced by half in Cslt1r Two human cysteinyl leukotriene receptors mice but was normal in magnitude and delayed in −/− CysLT 1R and CysLT2 R, cloned at the turn of the Cslt2r mice. These results suggested that century, do not bind the three cysteinyl leukotri- CysLT 1R is the major signaling receptor for LTC4 ene ligands equally. The rank order of binding for and LTD4 while CysLT2 negatively regulates

CysLT1 R is LTD 4 > LTC4 = LTE4 and for CysLT 2 R, CysLT1 R. Vascular leakage was not reduced by −/− LTC 4 = LTD4 > LTE4 . The receptors are expressed LTE 4 in Cslt1r mice but again sustained and on a wide range of organ tissues and cell types— delayed in the Cslt2r −/− strain, indicating that

CysLT1 R on spleen, lung, small intestine, pla- CysLTE R is the dominant receptor for LTE4 and centa, bronchial smooth muscle, mast cells, that CysLT 2 R once again acts as a negative neutrophils, eosinophils, macrophages, mono- regulator. cytes, and hemopoietic progenitor cells and Studies on expression of cysteinyl leukotriene

CysLT 2 R on lung, heart, lymph node, spleen, receptors by human mast cells unexpectedly brain, bronchial and coronary smooth muscle, revealed that LTE4 helps to induce greater num- adrenal medulla, mast cells, eosinophils, macro- bers of mast cells from cord blood progenitor phages, and monocytes. The receptors were ini- cells cultured together with IL-6 and IL-10 than

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both LTC 4 and LTD4 and it is also more potent for 3.2.5.3.1 Chemistry and Structure– the production of the infl ammatory chemokine Activity Relationships macrophage infl ammatory protein-1β (MIP-1β) PAF, 1-O -alkyl-2-acetyl-sn -glycero-3- phospho- and for the expression of COX-2 and prostaglan- choline, a phospholipid of relatively simple but din D2 . Sequence homologies of the classical unique structure, belongs to a relatively minor type 1 and 2 cysteinyl leukotriene receptors and class of lipids, the ether-linked phospholipids. The the P2Y receptor family together with computer distinguishing feature of its unique structure is an modeling studies indicated that LTE4 might be a acetyl group at position 2 of the glycerol backbone surrogate ligand for a previously unrecognized (Fig. 3.9). Removal of the acetyl group produces receptor on mast cells. Human mast cells express lyso-PAF which is devoid of biological activity. the P2Y12 receptor, a Gαi-linked receptor for ade- When produced and liberated naturally in the cel- nosine diphosphate. Subsequent investigations lular environment, PAF is made up of a mixture of by Austen’s group using ovalbumin-sensitized homologs differing in the number of carbons and and Cyslt1r/Clt2r −/− mice, expression of IL-13, the degree of unsaturation of the alkyl chain at and the P2Y12 receptor-selective antagonist clopi- position 1 of the glycerol backbone. The main dogrel suggested that LTE4 acted as an agonist homologs usually present are the C16:0 , C18:0 , and for platelet activation in the pulmonary vascula- C18:1 structures. The structures for maximum activ- ture. It seems, therefore, that P2Y12 is the receptor ity are a 16 carbon chain, the 1-O -alkyl ether link- for LTE4 -mediated amplifi cation of allergic pul- age, the acetyl group at position 2, the sn monary infi ltration and proliferation of mast cells confi guration, and the phosphate group at carbon 3 and this receptor is separate and distinct from the (Fig. 3.9 ). Activity decreases progressively as the

CysLTE R in the skin. C chain backbone is shortened or lengthened; replacement of the ether linkage leads to no or lit- 3.2.5.3 Platelet-Activating Factor tle biological activity; the unnatural enantiomer Platelet-activating factor (PAF), a preformed with the (S) -confi guration is inactive; for any bio- mediator of anaphylaxis released by degranulat- logical activity the only substituents tolerated at ing mast cells, is one of the most powerful auta- carbon 2 are propionyl and N -methyl carbamoyl coids yet discovered. The PAF story began in the groups (the 2-ethoxy analog has only 10 % of the early 1970s when Benveniste, Henson, and activity of PAF); and fairly major modifi cations of Cochrane demonstrated the release of a substance the substituents on the nitrogen diminish activity. with both powerful anaphylactic and platelet These specifi c structural requirements suggest that aggregating properties from allergically sensi- PAF exerts its biological effects by binding to spe- tized rabbit leukocytes. Although fi rst investi- cifi c receptors and this is in fact so. gated in relation to anaphylaxis and other allergic manifestations, later studies revealed a wide 3.2.5.3.2 Biosynthesis and Cellular diversity of other biological actions and involve- Sources of PAF ment in diseases such as asthma, some delayed Because PAF is such a potent mediator of a range hypersensitivity reactions, septic shock, adult of biological effects, its concentration in body respiratory distress syndrome, rheumatoid arthri- fl uids and tissues needs to be restricted to avoid tis, necrotic bowel disease, and a wide range of adverse or even lethal consequences. This is other infl ammatory conditions. This diversity of achieved intracellularly and extracellularly by a biological actions and pathogenic involvements specifi c acetylhydrolase and by regulation of the is due to the mediator’s activation of other cells conversion of precursor molecules. The activity besides platelets, in particular, eosinophils, neu- of PAF acetylhydrolase for its substrate is trophils, fi brocytes, neurocytes, and endothelial, extremely high ensuring that the half-life of the vascular, cardiac, smooth muscle, pancreatic, and mediator in blood is of the order of only a few secretory cells. minutes. PAF is synthesized by two metabolic

[email protected] 56 3 Mechanisms of Hypersensitivity

a small acyl group ether linkage required required O H2C OCH2 (CH2)nCH3 various chain lengths tolerated H CCO H 3 O + natural configuration (R) H2C OPOCH2CH2N (CH3)3 required choline - O only minor changes tolerated phosphate PAF b

c Excellent recognition

Poor recognition Good recognition

Fig. 3.9 ( a) Two-dimensional structure of PAF high- cule devoid of biological activity. (c ) Outline of PAF lighting the important structural features necessary for model indicating regions of excellent, good, and poor maximum biological activity and recognition by anti- recognition by anti-PAF antibodies. The antibody recog- PAF antibody combing sites. (b ) Three-dimensional nition pattern is very similar to that of the PAF receptor space-ﬁ lling CPK model of PAF with the acetyl group (see Smal MA, Baldo BA and Harle DG. J Mol Recogn circled. Removal of this group alone produces a mole- 1990; 3: 169–73) pathways—the de novo and remodeling path- on the cytoplasmic surface of the endoplasmic ways. In the de novo pathway (Fig. 3.10a ), the reticulum, catalyzes the reaction between speciﬁ c enzyme alkylacetylglycerol choline- 1- O -alkyl-2-acetyl-sn -glycerols and cytidinedi- phosphotransferase, widely distributed in tissues phosphocholine (CDP-choline) in the presence of

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 57

O CH2 OR O CH2 OR H C C O CH H3C C O CH 3 O + CH2 OH H2C O P O CH2CH2N (CH3)3 - O 1-O-alkyl-2-acetyl-sn-glycerol (AAG) PAF Mg 2+, specific AAG CDP-Choline PAF CMP + cholinephosphotransferase +

b Acetate Acyl-CoA

OR OR OR AcO HO Acyl O OPC OPC OPC PAF Lyso-PAF Acyl-PAF

Acetyl-CoA Fatty acid

Fig. 3.10 The two biosynthetic pathways for the synthesis of PAF. (a ) The de novo pathway; (b ) the remodeling pathway

Mg2+ generating PAF and cytidinemonophosphate. cytopenia, neutropenia, and eventually death. This synthetic pathway appears to maintain PAF Infusion of PAF into the heart decreases myocar- levels for normal physiological processes. The dial contractility and coronary fl ow, effects remodeling pathway (Fig. 3.10b ) both activates resembling cardiac anaphylaxis. Intradermal and deactivates PAF via the calcium-dependent injection produces a biphasic infl ammatory enzymes phospholipase A2 and acetyltransfer- response similar to the response of allergic sub- ase, the latter being the rate-limiting enzyme. jects to allergen. PAF has a profound effect on the These enzymes are found particularly in cells of lung producing bronchoconstriction, edema, and the immune system such as basophils, eosino- hyperresponsiveness. PAF is also one of the most phils, platelets, polymorphonuclear cells, macro- powerful ulcerogenic agents known, provoking phages, and endothelial cells and can be hemorrhage and vascular congestion in both the stimulated by a variety of agents including stomach and small intestine. immune complexes, thrombin and histamine. PAF has been implicated in many disease states but since it is often only one of a range of 3.2.5.3.3 Biological Actions of PAF and other mediators present, any preeminent role is Its Role in Health and Disease understandably often diffi cult to establish. For PAF is a hydrophobic molecule and for crossing example, it is frequently present along with hista- cell membranes and transportation to its various mine, numerous metabolites of the cyclooxygen- sites of action, serum albumin serves a carrier ase and lipoxygenase pathways, and a range of function. When injected into mammals, PAF pro- chemokines and cytokines including TNF. As duces both the signs and symptoms of anaphy- well as its undoubted role in anaphylaxis and laxis with hypotension, increased vascular some other allergic reactions, PAF is an impor- permeability and hemoconcentration, thrombo- tant mediator in the asthmatic response.

[email protected] 58 3 Mechanisms of Hypersensitivity

Administration of PAF into the lungs produces 3.2.5.3.5 Measurement of PAF in the severe bronchoconstriction, mucous secretion, Laboratory infl ammation, and long-lasting airway hyperre- Accurate measurement of the very small amounts activity. The latter two effects may be contributed of PAF in fl uids and extracts is a prerequisite for to by PAF-induced recruitment and activation of studying the role of the mediator in health and infl ammatory cells such as macrophages and disease. The most widely used method relies on eosinophils. Recent fi ndings, particularly in stud- the interaction of PAF with platelets, but the pro- ies in the mouse, have identifi ed a second path- cedures are not strictly quantitative, lack specifi c- way of anaphylaxis involving the IgG receptor ity, and are diffi cult to standardize and reproduce; FcγRIII and the release of PAF as the major fresh platelet suspensions are required; and mediator (see Sect. 3.2.7 ). Although long sus- throughput capacity is poor. Mass spectrometric pected, a central role for PAF in anaphylaxis is techniques are sensitive and specifi c but the spe- confi rmed and explained by this alternative path- cialized nature of the equipment, absence of easy way. In a model of peanut allergy for example, access for many laboratories, and diffi culty of although both histamine and PAF are involved in assessing large numbers of samples side by side the response, PAF is more important in shock make this method problematic for routine use by pathogenesis. Along with anaphylaxis and many researchers. Other methods such as mea- asthma, septic shock is a disease in which PAF is surement of 3 H-serotonin after PAF-induced suspected of having a leading role. PAF induces platelet degranulation and radioreceptor assays systemic responses similar to those provoked by are specialized procedures in some laboratories bacterial endotoxin and is found in the spleen and but can be diffi cult to standardize, require cell peritoneum of rats with endotoxic shock. Some labeling, or membrane preparations and may PAF antagonists protect animals against septic show high nonspecifi c binding. Perhaps the best shock caused by infection with gram-negative all-round, high-throughput procedure for quanti- organisms or injection of endotoxin. Because of tating PAF levels in research and test samples is a its potent effect on platelets, PAF is thought to be specifi c immunoassay, available since the fi rst involved in some thrombotic diseases including such assay was introduced in the authors’ labora- stroke. Other suspected roles are in acute graft tory in 1989. This method is highly specifi c, sen- rejection and immune complex deposition in, for sitive in the range 10–1,000 pg (0.02–2 pmoles), example, systemic lupus erythematosis, psoria- has a high capacity, and is not affected by inhibi- sis, and other allergic conditions. tors of platelet aggregation.

3.2.5.3.4 The PAF Receptor The PAF receptor is a MW 48 kD, G-protein- 3.2.6 Anaphylaxis coupled single 342 amino acid protein that shows structural characteristics of the rhodopsin gene Anaphylaxis is a sudden, systemic reaction involv- family. The human, guinea pig, and rat receptors ing a number of different organs of the body that have been cloned and characterized as a seven- may be severe enough to cause death. It is usually transmembrane receptor that induces phos- provoked by exposure to allergens with drugs and phoinositol turnover. The receptor shows wide foods being the most common causes. For the clini- tissue distribution being expressed in lung, kid- cal features of anaphylaxis the reader is referred to ney, liver, spleen, small intestine, and brain. In Chap. 2. Progress continues in identifying key leukocytes, it is expressed on platelets, neutro- intermediates and elucidating mechanisms of regu- phils, monocytes, and B cells but not on resting T latory systems and signaling pathways during mast cells and natural killer cell lines. Human mono- cell activation and degranulation and some impres- cytes treated with IFN-γ show a two- to sixfold sive advances in our understanding of the path- increase in PAF receptor expression compared to ways, the mediators involved, and their contribution untreated cells. to the pathobiology of anaphylaxis are under way.

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 59

3.2.6.1 Mechanisms of FcεRI- the pathophysiology of anaphylaxis. Cross- Mediated Mast Cell Activation linking initiates the signaling cascade that ulti- in Anaphylaxis mately results in anaphylaxis. FcεRI receptor Understanding anaphylaxis involves study of the aggregation causes Lyn, the tyrosine kinase cellular events leading to the release of media- associated with the β and γ subunits of the receptors of infl ammation and hypersensitivity with tor, to phosphorylate the tyrosines of the ITAMs emphasis on the mechanisms, in particular the of these two subunits. The phosphorylated signaling processes, of mast cell and basophil ITAMS, mainly on the γ subunit, then act as activation and degranulation. Upon activation of scaffolds for binding the cytoplasmic tyrosine mast cells and basophils following cross-linking kinase Syk. As outlined above in Sect. 3.2.2 , by allergen of receptor-bound IgE and aggrega- recruitment of the Syk kinase and subsequent tion of the high-affi nity IgE FcεRI receptors, the phosphorylation activation steps involving Lyn cells quickly release preformed mediators from lead to mast cell activation demonstrating the the secretory granules. These mediators, includ- importance of protein tyrosine kinases in the ing histamine, leukotrienes, PGD 2 , PAF, and pathways that result in allergic infl ammation and TNF, causing vasodilation, increased vascular anaphylaxis. These pathways involved in mast permeability and heart rate, bronchoconstriction, cell triggering are summarized in a simplifi ed airway remodeling, pulmonary and coronary form in Fig. 3.11 . Activated Syk is involved in vasoconstriction, and a host of other detrimental the phosphorylation of the transmembrane adap- effects, including cell recruitment with cytokine tor linker for activation of T cells (LAT) as well and chemokine production, are responsible for as the SH2 domain-containing leukocyte-specifi c

Antigen

IgE

γ β α Lipid raft LAT LAT

PIP3

PIP2 Btk PLC-γ PI3K Ca2+ release activated Fyn GAB2 Lyn 2+ Ca channel Grb2/Shc/Sos DAG Hck IP3 Syk Ras internal Vav Rac 2+ stores Ca IP3R in PKC ERK ER SLP-76 JNK

Transcription cPLA2 HISTAMINE RELEASE factors ARACHIDONIC CYTOKINES ACID

Fig. 3.11 Simpliﬁ ed summary of FcεRI-mediated signaling pathways in the mast cell leading to allergic inﬂ ammation and anaphylaxis

[email protected] 60 3 Mechanisms of Hypersensitivity protein of MW 76 kD (lymphocyte cytosolic (PLA-A2) to release arachidonic acid with the protein 2 LCP2 or SLP-76), the guanine nucleo- production of lipid mediators (Fig. 3.11). tide exchange factor Vav, phospholipase C-γ1 Further research, much of it in mice, has con- (PLC-γ1), and PLC-γ2. After involvement of the tinued to highlight the key role of tyrosine kinases enzyme proto-oncogene tyrosine-protein kinase in FcεRI activation and the subsequent signaling (Fyn), tyrosine phosphorylated GAB2 (GRB2 events, although other involved tyrosine kinases [growth factor receptor-bound protein 2]-associ- such as Hck have been identifi ed. The intensity of ated-binding protein 2) binds a subunit (p85) of stimulation of the FcεRI receptor has been shown phosphatidylinositol 3-kinase (PI3K). In the to be important. Low-intensity stimulation by membrane, PI3K catalyzes the conversion of IgE with low antigen concentration or by anti- phosphatidylinositol- 4,5- diphosphate (PIP2 ) to IgE positively regulates mast cell degranulation phosphatidyl-3,4,5-triphosphate (PIP3 ). This and the production of cytokines by inhibiting Lyn attracts a number of proteins containing the activity. High-intensity stimulation with high IgE pleckstrin homology (PH) domain, a 120 amino and high antigen concentrations negatively regu- acid domain occurring in a variety of proteins lates mast cells by enhancing Lyn activity and involved in intracellular signaling. The attracted increased Syk activation. Genetic variation proteins include Bruton’s tyrosine kinase (Btk) appears to infl uence the role of tyrosine kinases. and the PLCs γ1 and γ2 which in tyrosine- For example, an epilepsy- and anaphylaxis- prone phosphorylated form catalyze the hydrolysis of strain of mice was found to be defi cient in the

PIP 2 to inositol-1,4,5-triphosphate (IP 3) and expression of Lyn while a related epilepsy- prone 1,2-diacylglycerol (DAG). Both compounds act variant proved anaphylaxis resistant. Bone mar- as second messengers, the former releasing Ca 2+ row-derived mast cells (BMMCs) from the ana- resulting in a depletion of Ca2+ stores and entry phylaxis-sensitive mice had reduced Lyn and Syk of Ca2+ from the extracellular medium and the activities and showed degranulation typical of latter activating protein kinase C (PKC). These BMMCs of phenotype Lyn(−/−) whereas the events lead to mast cell degranulation. This phenotype of the anaphylaxis-resistant mice was activity takes place in two regions on the inner similar to wild-type animals. side of the plasma membrane. Electron micros- copy has revealed a primary region of activity 3.2.6.2 Sphingosine-1-Phosphate, near the FcεRI receptor involving Gab2, the p85 an Emerging Mediator subunit of PI3K, and PLC-γ2 and a second of Anaphylaxis region near LAT involving PLC-γ1 and the p85 Activated Fyn, involved in a second tyrosine subunit. Tyrosine phosphorylation and activation kinase pathway, has been shown to be required of other enzymes and adaptors, including Vav, for cytokine production as well as degranulation Grb2, the SHC-adaptor protein (Shc) involved in and to have a role in generating sphingosine-1- signaling, and Son of sevenless (Sos) protein (a phosphate (S1P) (Fig. 3.12 ) a blood-borne bioac- guanine nucleotide exchange factor), stimulate tive lipid mediator that is a major regulator of the the small GTPases Ras, Rho, and Rac. These vascular system and B and T cell trafﬁ cking. S1P reactions lead to activation of the extracellular- from mast cells and circulating S1P from macro- signal-regulated kinase ERK, Jun amino-termi- phages, platelets activated by the release of PAF, nal kinase JNK, the p38 mitogen-activated endothelial cells, and many other nonimmune protein (MAP) kinase cascade, and histamine cells are elevated in the lungs of asthmatics where release. Phosphorylation of the transcription fac- they regulate pulmonary epithelium permeability tors activating protein-1 (AP- 1), nuclear factor and are thought to contribute to the pathogenesis of activated T cells (NFAT), and nuclear factor of asthma and diseases such as rheumatoid arthri- kappa-light-chain-enhancer of activated B cells tis. The detection of elevated S1P levels in bron- (NF-κB) induces the synthesis of cytokines and chial alveolar lavage ﬂ uid of challenged the activation of cytoplasmic phospholipase A2 asthmatics and demonstration that it is necessary

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 61

+ NH3 O- O P OH

O OH Sphingosine-1-phosphate

Fig. 3.12 Structure of the bioactive signaling phospholipid sphingosine-1-phosphate, a regulator of immune and vascular systems

for sustained mast cell degranulation through the expression of mouse and human mast cells, the

S1P2 receptor were indications that this lyso- roles of SphK1 and SphK2 may prove to be sig- sphingolipid has a previously unrecognized rela- nifi cantly different in the two species. tionship with anaphylaxis. It is produced by sphingosine kinase (Sphk1 and Sphk2)-catalyzed addition of phosphate to sphingosine, broken 3.2.7 Other Mechanisms down by a S1P lysate and converted back to of Anaphylaxis: IgG, PAF, sphingosine by S1P phosphatase. Recent research and Nitric Oxide has shown that susceptibility to anaphylaxis appears to be due to S1P generated within the Mechanisms of anaphylaxis independent of IgE mast cell and by free, circulating S1P from non- have been suggested, for example, anaphylatox- mast cell sources. The gene SphK2 regulates the ins produced during complement activation, gen- infl ux of Ca2+ into mast cells and the responses to eration of immune complexes, the involvement it, making it a determinant of intrinsic mast cell of T cell activation and cytotoxicity, release of function whereas SphK1 appears to act extrinsi- neuropeptides, and a number of different mecha- cally affecting mast cell responsiveness by regu- nisms acting coincidently without the involve- lating levels of circulating S1P. The surprising ment of allergen-specifi c IgE. Intriguingly, demonstration of a relationship between circulat- anaphylaxis can occur in the mouse via the clas- ing levels of S1P and anaphylaxis is made more sic pathway involving allergen-induced cross- intriguing by the demonstration that reduced S1P linking of mast cell FcεRI receptor-bound IgE levels due to a defi ciency of SphK1 are associ- antibodies with release of histamine (and other ated with resistance to anaphylaxis. While it is mediators) but also by an IgG pathway in which well known that only a small number of individu- allergen–antibody complexes activate macro- als from a large group with similar circulating phages by cross-linking FcγRIII receptors and levels of allergen-specifi c IgE antibodies will with PAF as the main mediator of anaphylaxis. experience anaphylactic shock when challenged Although there is, as yet, no compelling evidence with the allergen, the amount of circulating S1P for an IgG-mediated mechanism in humans, what might help to provide the explanation. Finally, as appears to be anaphylaxis has been described in a exciting as these developments in our under- few cases where there is an apparent absence of standing of the underlying mechanisms of ana- mast cell degranulation, that is, with no increase phylaxis are, it should be remembered that many in serum tryptase. Certainly there are many simi- of the fi ndings result from research on mice not larities between the immune systems of mice and man and that is also true for most of the S1P stud- men; PAF is produced by macrophages of both ies where mice with SphK1 and 2 genes, indi- species, it has the same affect on vascular perme- vidually or jointly deleted, were used. Given the ability, and consequently allergen–IgG complexes diversity of mast cells and differences in gene may have an important role in anaphylaxis in

[email protected] 62 3 Mechanisms of Hypersensitivity humans as well as the mouse. However, human ana- • Relationships, if any, between the risk of anaphylaxis tends to result from low-dose exposure phylaxis and levels of allergen-specifi c IgE whereas mouse IgG-mediated anaphylactic reac- and the affi nities of IgE antibodies. tions may occur in response to relatively larger • The relationship between mediator activity antigen doses and/or adjuvants that elicit IgG as and their turnover, for example, PAF acetylhy- well as IgE antibodies. What may be evidence in drolase may be less active in some individuals favor of anaphylaxis in humans independent of allowing PAF to remain active for longer. IgE are the responses seen in patients after receiv- • Further studies of the relationship of ing the chimeric mouse-human anti-TNF mono- sphingosine- 1-phosphate with anaphylaxis. clonal antibody infl iximab (see also Sect. • Continuing searches for more mediators and 11.1.3.3 ). None of the subjects appeared to have markers of anaphylaxis, especially more sen- complementary IgE antibodies, all had IgG to the sitive ones. mouse immunoglobulin determinants, and there • Is there an IgG-dependent pathway for ana- was no increase (at only 20 min) in serum trypt- phylaxis in humans? If so, what is the ase. From insights gained from his extensive mechanism? studies of mechanisms of anaphylaxis in mouse • Identifi cation of signaling pathways that stim- models, F. D. Finkelman has suggested that large ulate or inhibit anaphylaxis and how these doses of antigen might be used in humans to look pathways can be manipulated. for evidence of anaphylaxis accompanied by • Further studies on the roles and importance of macrophage activation and PAF secretion. NO, eNOS, and iNOS. PAF contributes to hypotension and cardiac • The role of the heart in anaphylaxis, in partic- dysfunction during shock and stimulates, via its ular, the heart mast cells in cardiovascular receptor, a number of signaling pathways includ- collapse. ing those that activate PLA 2 and PI3K. Studies of • Estimation of levels and searches for poly- PAF and anaphylactic shock in mice have shown morphisms of relevant cytokines and cytokine that PAF-induced shock depends on PI3K receptors such as IL-4, IL-13, and TNF. signaling and on NO produced by constitutive • The role, if any, of IgG blocking antibodies. nitric oxide synthase (eNOS) not the inducible This list is far from exhaustive. form of the enzyme (iNOS). Mouse models showed that inhibition of NOS, PI3K, or Akt, or defi ciency of eNOS, gave complete protection 3.2.8 Drug-Induced Urticaria against anaphylaxis. These fi ndings appear to and Angioedema support the belief that eNOS has a detrimental role in vascular function during shock and in For the clinical manifestations of urticaria, see regulating infl ammation. Further, if eNOS- Sect. 2.2.1.2 . derived NO is the principal vasodilator in ana- Of the drugs implicated in provoking urticaria phylactic shock, eNOS and/or PI3K or Akt and angioedema, the NSAIDs are perhaps the might prove to be important targets for treating most important. What is currently understood of anaphylaxis. their proposed mechanisms of action together Clearly, there is much to learn and understand with a review of the arachidonic acid cascade is about anaphylaxis and the list of interesting ques- considered in Chap. 9. Formation of the cysteinyl tions that remain unanswered is disconcertingly leukotrienes is detailed in Sect. 3.2.5.2 (above) extensive. The following topics are suggested as and is also referred to in Chap. 9 . important and potentially productive areas of Urticaria may be classifi ed as acute or chronic. investigations that could be near the top of any The acute form appears early after exposure, current research agenda for anaphylaxis: perhaps within minutes, and can last from hours

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 63 to several weeks whereas the chronic form per- symptoms. An immediate (within 20 min) sists for about six weeks or more. Urticaria is positive skin test is usually presumed to result often an isolated event but drug-induced urti- from an IgE antibody-mediated mechanism or a caria, regarded as one of the most common cuta- direct histamine-releasing effect but one cannot neous drug reactions, can be seen in association necessarily presume that these are the only mech- with anaphylaxis, angioedema, and serum sick- anisms operative in all cases of drug-induced ness. Urticaria is a heterogeneous disease with acute urticaria. Symptoms of urticaria are caused many subtypes caused by a range of agents and by the mediators histamine, leukotrienes, prosta- stimuli. Some infections (e.g., Helicobacter glandin D2, bradykinin, and other vasoactive pylori), intolerance to foods, and autoantibodies substances released from mast cells and baso- to the high-affi nity IgE receptor FcεRI have been phils into the skin. Cases of acute urticaria may implicated, but, apart from the NSAIDs and be immune or nonimmune mediated. Drug- angiotensin-converting enzyme (ACE) inhibi- induced immune-mediated reactions can be elic- tors, there is a dearth of information on mecha- ited by cross-linking of high-affi nity nisms underlying drug-induced urticaria and (FcεRI)-bound IgE antibodies on mast cells and angioedema. basophils by free drug or drug–carrier complex molecules reacting with the bivalent antibody 3.2.8.1 Genetic Mechanisms combining sites via their complementary aller- Knowledge of drug-induced mechanisms of urti- genic determinants. This results in degranulation caria and angioedema is limited and this is even of the cells and histamine release. more apparent when considering current progress Other hypersensitivity responses may lead to on the molecular genetic mechanisms involved. urticarial reactions. A rare cause of the acute Information on genetic polymorphisms of rele- form is a type II hypersensitivity cytotoxic reac- vant genes together with supporting functional tion mediated by cytotoxic antibodies and com- studies is needed to help elucidate molecular plement activation. An example of this type of mechanisms and identify genetic markers. Some reaction occurs in transfusion reactions when progress made in identifying HLA alleles and IgG and IgM antibodies activate complement and promoter polymorphism genetic markers for lyse transfused incompatible red cells. Urticaria aspirin-induced urticaria/angioedema is covered may also result from a type III antigen–antibody in Chap. 9 , Sect. 9.5.5 . complex-mediated hypersensitivity reaction, in particular, serum sickness lasting for several 3.2.8.2 Urticaria Due to Immune weeks and presenting with fever, arthralgias, and Mechanisms glomerulonephritis as well as urticaria. Note that Urticaria following drug administration may there is a drug-induced serum sickness-like reac- occur without previous exposure to the drug or tion that is not associated with circulating after previously tolerated exposures. Drugs immune complexes. Drugs implicated in these appear to cause only a minority of cases of reactions include penicillins, cephalosporins, tet- chronic urticaria and while they are often racyclines, quinolones, sulfonamides, NSAIDs, assumed to be the cause of a high proportion of carbamazepine, thiouracil, allopurinol, and bar- cases of acute urticaria during drug treatment, biturates. Other drug-induced type III hypersen- some results cast doubt on this. An examination, sitivity reactions involving skin infl ammation including skin testing, of 350 patients with sus- include erythema nodosum leprosum induced by pected drug-induced reactions made up of 343 dapsone and the Jarisch–Herxheimer reaction with urticaria/angioedema and seven with ana- following treatment of some microorganisms phylaxis revealed that only 22 (6.3 %) were aller- (e.g., in syphilis) with antimicrobials such as gic and had a positive skin test to the suspected penicillins and tetracyclines. The infl ammatory drug. The positive reactors proved to be the cytokines TNF, IL-6, and IL-8 appear to be patients who presented with the most severe released in these reactions. Urticarial vasculitis is

[email protected] 64 3 Mechanisms of Hypersensitivity another type III hypersensitivity skin eruption strengthening the evidence for an immune basis that can resemble urticaria and which is some- of the disease and supporting histological dem- times drug induced. Antigen–antibody com- onstrations of predominant CD4+ T cell infi l- plexes formed in the vascular lumina lead to trates in biopsies of chronic urticaria lesions. complement activation, chemotaxis of neutro- The observed cytokine profi le of Il-4, IL-5, and phils, and the release of proteolytic enzymes that IFN-γ does not refl ect a predominance of Th1 or damage the vascular lumina. Drugs implicated Th2 cells and cellular infi ltrates indicate a Th0 include ACE inhibitors, penicillins, sulfonamides, profi le or a mixture of activated Th1 and Th2 cells. thiazides, and the antidepressant fl uoxetine. Omalizumab, a recombinant humanized Urticarial reactions are also sometimes seen along monoclonal antibody that inhibits the binding of with other skin manifestations during some drug- human IgE to its high-affi nity receptor FcεRI by induced type IV hypersensitivity responses, but selectively binding the immunoglobulin in solu- the presence of other skin manifestations, fre- tion, has been used as a successful treatment of quently more severe, makes it diffi cult to distin- intractable allergic asthma. The effi cacy of the guish and study the specifi c mechanisms. monoclonal antibody treatment was therefore investigated in patients with chronic autoimmune 3.2.8.3 Urticaria with an Autoimmune urticaria who remained symptomatic on antihis- Basis tamine therapy. Of the 12 patients treated, seven A signifi cant proportion of cases of chronic urti- showed complete resolution of the urticaria, four caria demonstrate no connection with drugs. responded with a decrease in the urticaria activity Observations during the 1980s of the associa- score but the urticaria persisted, and one patient tion of chronic urticaria and angioedema with showed no improvement. thyroid autoimmunity and on the prevalence of anti-IgE autoantibodies in urticarial syndromes 3.2.8.4 Basophils in Chronic Urticaria suggested that autoimmunity might have a role CD203c is a basophil activation marker that is in some cases of chronic urticaria. These fi nd- upregulated by cross-linking the FcεRI recep- ings led to the occasional demonstration of the tors on mast cells and basophils. Incubation of presence of IgG anti-immunogloulin E autoanti- basophils with sera from patients with chronic bodies and functional autoantibodies against the idiopathic urticaria and a positive autologous alpha subunit of the high-affi nity IgE receptor serum skin test (ASST; an intradermal test with (i.e., FcεRIα) in at least one-third of patients the patient’s own serum) demonstrated signifi - with chronic urticaria. These autoantibodies cant upregulation of CD203c and this upregula- activate normal cell function by cross-linking tion correlated with basophil histamine release the receptors on cutaneous mast cells and blood and the ASST. Basophils from chronic urticaria basophils, thus releasing histamine and other patients are less responsive to anti-IgE and C5a mediators responsible for the urticaria and but highly responsive when incubated with angioedema. Activity of the autoantibodies was sera, even normal sera. The stimulatory factor(s) later shown to be augmented by complement in serum has not been identifi ed and the activation with a critical role for component increased response of the cells is not yet under- C5a. Chronic urticaria is now divided into auto- stood. In a fl ow cytometric evaluation of the immune and idiopathic subgroups since in about expression of basophil cell surface markers 55–60 % of patients the etiology remains CD203c, CD63, CD123, and the receptor obscure. As well as releasing histamine and leu- FcεRIα, both CD203c and CD63 were upregu- kotrienes from basophils, sera with the autoim- lated on basophils from patients with chronic mune antibodies also release IL-4. A study of idiopathic urticaria regardless of their ASST lymphocytes from patients with chronic urti- response. High expression of IL-3 receptor on caria showed that activated CD4+ T cells pro- basophils and activated T cells was detected duced high amounts of IL-4 and IFN-γ, only in ASST-positive patients.

[email protected] 3.2 IgE Antibodies and IgE- Mediated Drug Hypersensitivities 65

3.2.8.5 Nonimmune-Mediated angiotensin II and degrades bradykinin, a potent Urticaria and Angioedema vasodilator. The vasoconstrictor action of angio- Angioedema does not always have an allergic tensin II may lead to increased blood pressure basis. Nonallergic angioedema not involving IgE and hypertension, the effect that led to the devel- antibodies and unassociated with urticaria can opment and application of the ACE inhibitor occur. The prototype example is hereditary drugs. ACE inhibitors increase bradykinin levels angioedema arising from a defi ciency of the and prolong its action and decrease angiotensin II inhibitor for C1 esterase that results in the main- levels (and therefore a decrease in aldosterone tenance of undegraded bradykinin. Acquired secretion from the adrenal cortex) leading to dila- angioedema may be due to the accelerated con- tion of blood vessels and a coincident decrease in sumption of C1 esterase inhibitor or, with an arterial blood pressure. ACE inhibitors, now immune component, to autoantibody production. widely used to treat hypertension, congestive During acute attacks of hereditary and acquired heart failure and diabetic nephropathy include angioedemas, plasma bradykinin has been shown the drugs benazapril, captopril, enalapril, fosino- to rise to up to 12 times the normal level. pril, lisinopril, perindopril, quinapril, ramipril, and trandolopril. The combination of actions of 3.2.8.5.1 Drugs that Directly Trigger decreasing angiotensin II and aldosterone levels Mast Cell Release and increasing and maintaining bradykinin levels These reactions are sometimes referred to as pseu- may lead to fl uid extravasation into subcutaneous doallergic responses since their clinical course and tissue ultimately producing angioedema. The presentation are similar to allergic urticaria and increased levels of bradykinin are thought to be angioedema. Drugs including the antimicrobial related to the high incidence of cough in patients vancomycin (Chap. 6), neuromuscular blockers on ACE inhibitors and elevated bradykinin levels used in anesthesia (Chap. 7), opioid analgesics in the peripheral tissues, resulting in rapid fl uid such as morphine (Chap. 8 ), NSAIDs (Chap. 9 ), accumulation, are suspected of playing a key role radiocontrast media (Chap. 10 ), and a wide range in angioedema seen in a small number of patients of other less often used medications can trigger taking ACE inhibitors. The association between urticaria by directly stimulating mast cell degranu- ACE inhibitors and angioedema, fi rst reported in lation and histamine release. The mechanism of the early 1980s, is now well recognized as a mediator release by the NSAIDs is particularly potentially serious but rare side effect of the interesting. The drugs inhibit cyclooxygenase drugs. Reactions occur with an incidence of which in turn leads to overproduction of the vaso- about 0.1–0.5 % but the incidence in blacks active and pro- infl ammatory leukotrienes (black Americans and Afro-Caribbeans) is about (Sect. 3.2.5.2 above). This subject is discussed in three times higher than in white populations. more detail in Chap. 9 . This, and the decreased antihypertensive response to ACE inhibitions in blacks, is thought to be due 3.2.8.5.2 Angiotensin-Converting to decreased production of bradykinin and/or Enzyme (ACE) Inhibitors decreased vasodilation in response to the peptide and Angioedema vasodilator. In terms of the number and severity, ACE occurs as somatic and germinal isozymes. ACE inhibitor-induced angioedema is said to The somatic enzyme, expressed in the lungs and account for 17 % of patients admitted for the in vascular endothelial, kidney, and testicular treatment of angioedema and from 13 to 22 % of Leydig cells, is part of the renin–angiotensin– patients with this form of angioedema require aldosterone system, one of the body’s mecha- airway intervention. In a 2008 study in Boston, nisms for maintaining blood pressure. ACE has USA, records of 220 patients who presented to two actions—it catalyzes the conversion of the fi ve hospital emergency departments were ten amino acid peptide angiotensin I to the reviewed. The frequency of ACE inhibitor- induced potent vasoconstrictor eight amino acid peptide angioedema in all patients who presented with

[email protected] 66 3 Mechanisms of Hypersensitivity angioedema was 30 %. The annual rate of visits It can be seen therefore, that the kallikrein–kinin for the drug-induced reaction was 0.7 per 10,000 and renin–angiotensin systems interact and are emergency department visits. Eleven percent of linked in a mutually dependent way. the patients were admitted to intensive care and Although it is beyond our requirements here, it 18 % admitted to hospital for observation for a should be pointed out that a homolog of ACE, 24 h period. This study confi rmed past experi- angiotensin-converting enzyme 2 (ACE2), has ence and surveys concluding that ACE inhibitor- recently been recognized. The two enzymes show induced angioedema remains a rare condition, it different recognition of bradykinin. ACE2, a car- represents a signifi cant proportion of angioedema boxypeptidase found mainly in the heart, kidney, patients, and a subgroup of these patients require and testis, does not degrade bradykinin but degrades hospitalization for management of upper airway des-Arg(9)-bradykinin at its carboxy-terminal angioedema. amino acid and, unlike ACE which degrades angio- Figure 3.13 summarizes the individual reac- tensin I by cleaving at the penultimate phenylala- tions and the interactions and relationship nine to produce angiotensin II [angiotensin(1–8)], between the renin–angiotensin system and the ACE2 removes the carboxy- terminal leucine to plasma kallikrein–kinin system. Activation by form angiotensin(1–9). This peptide has been the enzyme prolylcarboxypeptidase (lysosomal reported to enhance arachidonic acid release. carboxypeptidase) of the prekallikrein–high molecular weight kininogen complex on endo- 3.2.8.5.3 Angioedema Following thelial cells produces kallikrein which cleaves Administration of Angiotensin high (sometimes low) molecular weight kinino- II Receptor-Binding Inhibitors gen liberating bradykinin. Bradykinin stimulates Angiotensin II receptor blockers (ARBs) are pri- vasodilation and leads to the formation of nitric marily prescribed for high blood pressure but oxide (NO), superoxide and prostacyclin and the may also be used to treat heart attack, stroke, and liberation of tissue plasminogen activator. congestive heart failure. Unlike ACE inhibitors, Kallikrein in plasma and tissues also activates ARBs are not associated with cough. When fi rst prorenin to renin, an aspartyl protease, which in approved for the treatment of hypertension in turn activates angiotensinogen to angiotensin I. 1995, ARBs were considered safe from the risk ACE converts the inactive decapeptide angioten- of edema and they are generally a safe alternative sin I to the biologically active octapeptide to ACE inhibitors, blocking the renin–angioten- angiotensin II which, like bradykinin, stimulates sin system more effectively than the latter drugs. NO and superoxide formation as well as contrib- By binding selectively to the angiotensin 1 recep- uting to the elevation of blood pressure and local tors (AT1), ARBs do not affect ACE and there- vasoconstriction and stimulating the release of fore should not affect bradykinin levels but plasminogen activator inhibitor I. ACE is also a angioedema to ARBs does occur with an inci- major degrading enzyme for bradykinin (in fact, dence ranging from about 0.1 to 0.4 %. From lim- bradykinin is its preferred substrate over angio- ited numbers examined, the risk of patients with tensin I) producing the breakdown pentapeptide angioedema to ACE inhibitors developing angio- bradykinin(1–5) and in addition to its role in the edema to an ARB is said to be from 2 to 17 % and formation of kallikrein, prolylcarboxypeptidase for developing confi rmed angioedema, 0–9.2 %. (with other enzymes) degrades angiotensin II to A review of 19 cases of ARB-induced angio- form angiotensin(1–7) which has vasodilatory edema found that 13 (68 %) had never received and blood pressure-lowering activities. Overall an ACE inhibitor. Angioedema has been reported then, stimulation of the bradykinin and angioten- after administration of losartan, candesartan, sin II receptors results in vasodilation and the eprosartan, irbesartan, olmesartan medoxomil, production of NO and prostacyclin. Stimulation and telmisartan. Cross-reactivity between ACE of the angiotensin I receptor leads to vasocon- inhibitor- and ARB inhibitor-induced angio- striction and the elevation of blood pressure. edema has been estimated to be from 3 to 8 %.

[email protected] 3.2 IgE Antibodies and IgE-Mediated Drug Hypersensitivities 67

High MW Kininogen-Prekallikrein prolylcarboxypeptidase KALLIKREIN

RENIN-ANGIOTENSIN SYSTEM

Extracellular Kallikrein activates Prorenin

Renin

Angiotensinogen

ACE2 Angiotensin I Angiotensin (1-9) [angiotensin (1-10)]

ACE

Angiotensin II ACE2 Angiotensin (1-7) [angiotensin (1-8)] prolylcarboxypeptidase

Vasoconstriction & BP ↑ Vasodilation & BP ↓

tissue plasminogen NO, superoxide NO, superoxide, plasminogen activator activator (tPA) PGI2 inhibitor 1 (PAI-1)

Intracellular Vasodilation

ACE Bradykinin Bradykinin (1-5) ACE2 Kallikrein-cleaved HMW kininogen

Kininogen

Extracellular Kallikrein

KALLIKREIN-KININ SYSTEM

Fig. 3.13 Summary of the individual reactions involved verting enzyme, BP blood pressure, MW molecular in, and relationships between, the renin–angiotensin and weight, NO nitric oxide plasma kallikrein–kinin systems. ACE angiotensin-con-

[email protected] 68 3 Mechanisms of Hypersensitivity

Since angioedema to ACE inhibitors occurs as a fl ow rate (PEFR) following inhalation challenge result of increased bradykinin levels and ARBs with allergen. Figure 3.14 shows that the imme- are not known to affect these levels, the mecha- diate response bottoms at about 30 min after nism of ARB-induced angioedema is not under- allergen challenge before beginning to recover stood. One suggested explanation is that and climbing back over the next 30 min toward, unblocked angiotensin II receptors (AT2) are but not reaching, the pre-challenged PEFR fi g- subjected to secondary stimulation by high levels ure. Three to four hours after the initial allergen of angiotensin II producing an increase in tissue challenge there is a late phase response refl ected bradykinin and hence angioedema. Another sug- in a pronounced fall in PEFR which reaches its gestion is an abnormality in the degradation of maximum at 5–10 h. Thereafter there is a steady the active metabolite of bradykinin, climb back to normal levels. The immediate des-Arg(9)-bradykinin. response is caused by the release of histamine and some other preformed mediators from mast cells that have direct effects on blood vessels and smooth muscle. The initial release of the pre- 3.3 The Allergen-Induced Late formed mediators is supplemented over time by Phase Reaction other powerful infl ammatory agents including vasoactive agents that dilate blood vessels and Exposure to allergen in the skin, lung, nose, or produce edema, swelling, and pain. Figure 3.15 eye of atopic patients provokes an immediate or shows good examples of immediate and late early response that is maximal at 20–30 min, phase cutaneous reactions. An immediate wheal resolves within about an hour, and is often fol- and fl are reaction and a late phase edematous lowed 3–4 h after allergen challenge by a delayed response are seen 15 min and 6 h, respectively, reaction peaking at 6–12 h and subsiding by 24 h. following intradermal injection of antigen. The two reaction phases are well illustrated by an asthmatic response in the lungs of an allergic patient measured as falls in the peak expiratory

Allergen challenge 600

500

400 Immediate response Late phase 300 response

15 mins

0306090251020 Minutes Hours

Fig. 3.14 A typical lung function result as measured by tion at about 30 min is followed by a late phase response peak expiratory ﬂ ow rates (PEFR) in an allergic patient which reaches a maximum 5–10 h after allergen following challenge with allergen. An immediate reac- challenge

[email protected] 3.3 The Allergen-Induced Late Phase Reaction 69

the reactions by a variety of methods including direct demonstration by induction of immediate and late responses with affi nity-purifi ed allergen- specifi c IgE antibodies followed by allergenic challenge. Another important fi nding was the observation that lymphocytes were the predominant cell in the cellular infi ltrates together with a signifi cant number of eosinophils and basophils. It should be remembered, however, that the investigations implicating IgE antibodies in late reactions to Bacillus subtilis enzyme, ragweed pollen, and other inhalant allergens do not necessarily refute the conclusion of a type III Arthus reaction to Aspergillus species and other allergens responsible for hypersensitivity pneumonitis conditions such as bagassosis and farmer’s, bird-fancier’s, coffee worker’s, malt worker’s, and mushroom worker’s lung. These are very different conditions to hypersensitivities to allergen sources such as ragweed pollen and dust mites and are character- Fig. 3.15 An immediate wheal and fl are cutaneous reaction in an allergic patient 15 min after intradermal injec- ized by different antigenic stimuli, symptoms of tion of antigen shown alongside a late phase edematous cough, dyspnea, pleurisy, fatigue, anorexia, and response 6 h post injection (Photograph kindly provided weight loss with interstitial granulomas and by Professor S. R. Durham) mononuclear and giant cells in the lungs.

3.3.1 Early Studies: Implication of IgE Antibodies 3.3.2 Cellular Responses in the Late Phase Reaction and Late reactions have been known for many years Comparison with the with initial published reports dating back nearly Delayed-Type 100 years, but investigations of the underlying Hypersensitivity Response cellular events and mechanisms involved were not pursued in any systematic way until the late From undergraduates to clinicians and research- 1960s when Pepys and colleagues studied ers, there has long been confusion over use of the patients with allergic bronchopulmonary asper- terms “late” and “delayed” with the late phase of gillosis and extrinsic allergic alveoltis, also the immediate wheal and fl are reaction some- known as hypersensitivity pneumonitis. They times being labeled and referred to as a delayed- found edema, perivascular cellular infi ltration, type hypersensitivity reaction, a type IV reaction, deposition of complement and serum immuno- or simply “DTH.” There was therefore a need to globulin precipitins to Aspergillus fumigatus , and research, compare, and contrast these reactions a variety of other extracts from organisms and and this was done in an important study in which agents that cause allergic alveolitis and con- both responses were provoked in the same indi- cluded that the late reactions were the result of an viduals and studied with the same panel of cell Arthus or type III reaction. Soon after, other marker monoclonal antibodies together with investigators came to a different conclusion fail- immunohistologic methods. Skin biopsies from ing to consistently fi nd precipitins and comple- atopic individuals with late phase allergic skin ment but strongly implicating IgE antibodies in reactions to intradermal challenge with grass

[email protected] 70 3 Mechanisms of Hypersensitivity

pollen or house dust mite were sectioned and profi le expressing mRNA for the cytokine gene examined for evidence of infi ltration and activa- cluster IL-3, IL-4, IL-5, and granulocyte macro- tion of T cells and eosinophils. A substantial phage colony-stimulating factor (GM-CSF). number of CD3+ and CD4+ cells but far fewer Cells from tuberculin biopsies on the other hand CD8+ cells were observed together with clearly preferentially expressed mRNAs encoding for different CD4+: CD8+ ratios in the sampled tis- IL-2 and IFN-γ, that is, cells preferentially sue and the peripheral blood. Infi ltrated cells expressing a Th1 cytokine profi le. Comparisons bearing receptors for IL-2 and evidence for IFN-γ of the accumulation of infl ammatory cells and secretion suggested that T cells had become acti- cells expressing mRNA for different cytokines in vated. Activated eosinophils were also detected late phase and delayed-type responses in the and there was a strong correlation between these same subjects showed a relatively rapid (1–6 h) cells and the numbers of CD4+ cells 24 h after accumulation of T cells and granulocytes in the the allergen challenge, suggesting that T cells former case and a much longer accumulation participate in the late phase infl ammatory reac- time (24–48 h) for T cells, macrophages, and tion. In fact, about 50 % of cells infi ltrating a late other cells expressing Th1-type cytokines in phase reaction site are T lymphocytes. It is there- delayed hypersensitivity responses. At 48–96 h fore of interest to compare the late phase response in the late phase response, some cells increas- with the delayed-type hypersensitivity reaction ingly expressed Th1-type cytokines. This may be since it seems that T lymphocytes are important an indication of a classic delayed hypersensitiv- in both responses. In the comparison, grass pol- ity response earlier masked by the IgE antibody- len and house dust mite extracts were used to mediated reaction. Again, with the delayed-type elicit late phase reactions while tuberculin chal- response the distinction from the late phase lenge was used for delayed hypersensitivity response was not totally clear since a small num- responses. Both responses showed accumulation ber of cells in some individuals expressed mRNA of CD4+ T cells, but overall the cells were more encoding IL-4 and IL-5. dispersed in denser accumulations and cells were still being recruited at 48 h in the delayed reactions. This is in contrast to the situation in late 3.4 Drug-Induced phase reactions where cell numbers usually pla- Hypersensitivity and Immune teau between 24 and 48 h. Other differences Receptors found were greater activation of eosinophils in late phase reactions, the detection of small num- 3.4.1 Background bers of these cells in atopics and non-atopics at 24 h but not at 48 h in delayed-type hypersensi- An antibody response to a chemically reactive tivity, and greater T cell activation (demonstrated drug or hapten is said to occur after the drug–pro- by expression of IL-2R) in the latter response. tein complex is recognized, processed, and pre- The release of infl ammatory cytokines in both sented as a drug–peptide conjugate to T cells that reactions was indicated by endothelial expression recognize the drug-modifi ed peptide. A low of HLA-DR. The allergen-induced late phase molecular weight free, unconjugated drug is reaction then has features of a cell-mediated thought to remain unrecognized and not equipped hypersensitivity response, but it shows some sig- to elicit an immune response. For drugs, how- nifi cant differences from the classical delayed ever, immunological dogma is often found want- hypersensitivity response in atopic subjects. The ing on at least two counts. Firstly, despite the difference is perhaps best illustrated by the dif- requirement that “small” molecular weight com- ferent cytokine profi les. Employment of labeled pounds or haptens (generally less than 1,000 kDa) RNA probes for some cytokines showed that need attachment to a macromolecular carrier to infi ltrating cells from allergen-induced late phase become immunogenic, many haptens or drugs cutaneous reactions have a Th2-like cytokine that remain uncomplexed and apparently too

[email protected] 3.4 Drug-Induced Hypersensitivity and Immune Receptors 71 small do in fact elicit a clear immune response. understood that Landsteiner’s studies on the Secondly, despite the pioneering fi ndings of sensitizing properties of some chemicals in the Landsteiner and other early immunochemists, form of “small” molecules linked to a protein car- and the conclusion that previous exposure to an rier constituted the initial investigations of contact allergen is a prerequisite for allergic sensitization hypersensitivity and the fi ndings and interpreta- and reactivity, the dogma of prior exposure does tions from studies on protein conjugated chemi- not always hold. Previous contact with a drug is cals should not automatically be used to explain not necessarily a prerequisite for a drug-induced all delayed reactions and certainly not IgE anti- immune hypersensitivity response. Although body-mediated type I reactions. An early clue to these inconsistencies were emphasized by the specifi c immune recognition of “small,” unbound author and some other investigators over 20 years chemicals and hence drugs was the demonstration ago (see monograph on drug allergy, Further by Sinigaglia and his group of selective interac- Reading), general acceptance of exceptions to the tion of nickel (Ni) ions with an MHC-II-bound dogma has only recently been forthcoming. peptide. Ni-specifi c T lymphocyte clones from a patient with contact dermatitis to Ni responded to the metal ions when Ni salts were presented by 3.4.2 Recognition and the Immune APC in association with DRw11.1(5) molecules. Response to Free, Direct evidence that Ni was bound to the MHC- Unconjugated Drug associated peptide was provided by NMR spectroscopy. These results, the fi rst direct evidence of The implications for drug allergy from the basic interaction between hapten and a MHC-bound and applied research on T cell recognition of hap- peptide, not only demonstrated a model for Ni tens initiated just over 20 years ago, were perhaps recognition by T cells from patients with Ni best summed up by Weltzien who, in commenting hypersensitivity but also indicated that a variety on the advances, declared simply that the work of chemically reactive groups, not only reactive may “contribute to a better understanding of what metal ions, might attach to MHC-bound mole- defi nes an antigen as an allergy-inducing aller- cules to induce MHC-restricted responses to the gen.” Perhaps this will eventuate but since the conjugates. Further work with Ni hypersensitivity fl edgling discipline of hypersensitivity research and the occupational lung disease berylliosis moved beyond the embryonic stage in the early established that these conditions were MHC-II- half of the twentieth century and matured over a linked CD4+ delayed-type hypersensitivity 60-year period to provide impressive insights into responses and that the high frequency of the effector processes in the immediate allergic Ni-reactive T cells occurs by formation of revers- response, in particular the roles of IgE antibodies, ible coordination complexes in which Ni interacts mast cells, infl ammatory mediators, and their with the MHC and TCR via His81 of the HLA-DR receptor- controlled end organ responses, the long- α-chain and Tyr29 and Tyr94 of the CDR1α standing question of what makes an allergen an region of the TCR. This coordination complex of allergen has remained obscure. Whether Ni ions directly linking the MHC peptide and approaches utilizing T cell recognition of antigens TCR is similar to the action of a weak superanti- in eliciting delayed hypersensitivity responses gen. In extending the studies on Ni to investiga- can soon provide the experimental and clinical tions on the T cell recognition of haptens, Weltzien opportunities to obtain the necessary insights and and others have shown, somewhat surprisingly to answers remains to be seen. some, that MHC-restricted hapten-specifi c T cell How then do small molecular weight, nonreac- receptors react to hapten–peptide conjugates tive chemicals such as many drugs stimulate IgE within the MHC peptide-binding groove. This antibody production and provoke immune hyper- opened up a new approach for studying the sensitivity reactions ranging from mild rashes to molecular mechanisms underlying hapten recog- severe, life-threatening anaphylaxis? It must be nition by T cells.

[email protected] 72 3 Mechanisms of Hypersensitivity

3.4.3 Abacavir and the MHC- Presented Altered Peptide NH O Model of Drug N N Hypersensitivity N N

N N H2N N N More recently, some interesting HLA associa- H tions in drug hypersensitivities have been O reported. A strong association of hypersensitivity to the guanosine-related pro-drug and Abacavir Didanosine reverse- transcriptase inhibitor abacavir was OH OH found with the well-defi ned 57.1 MHC haplotype encoding the MHC class I allotype HLA- O O B*57:01 (see Sect. 1.3 ). Multi-organ reactions to N N abacavir, termed abacavir hypersensitivity syn- HN HN drome or AHS, manifests as fever, rash, malaise, N N H2N N H2N N nausea, and diarrhea. It occurs in approximately OH 2–8 % of patients with human immunodefi ciency O virus-1 (HIV-1) infection and can be severe OH enough to cause death in some rechallenged Carbovir Guanosine patients. Abacavir-specifi c CD8+ T cells secrete OH OH TNF and IFNγ and are cytotoxic to abacavir- APCs. In a 2008 study, implication of the fi ne- Fig. 3.16 Structures of abacavir and three purine structural specifi city of the 6-cyclopropylamino analogs didanosine, carbovir, and guanosine. The 6- cyclopropylamino group of abacavir is highlighted group of abacavir as a possible reactive site in the HLA-restricted CD8+ T cell response was demonstrated by lack of recognition of the abacavir structural analogs carbovir, didanosine, tion of having profound implications for under- and guanosine by abacavir- reactive T cells standing the origins and general molecular (Fig. 3.16 ). Specifi city of the interaction was fur- processes of autoimmunity. AHS is mediated by ther mapped to the F pocket (one of six, termed abacavir-specifi c activation of cytotoxic CD8+ T A–F), of the MHC-1 antigen- binding cleft where cells that require HLA-B*57:01 antigen- it was thought that abacavir, or a metabolite, presenting cells, but abacavir-specifi c T cells are binds to one or more self- peptides. At that stage, not activated by cells expressing the closely whether the binding was covalent or not had yet related allotypes HLA-B*57:02, HLA-B*57:03, to be determined. It was predicted that the dem- and HLA-B*58:01, each of which is insensitive onstration that AHS is an MHC-I-restricted cel- to abacavir and not linked to AHS. Two amino lular hypersensitivity response mediated by acid residues, Asp114 and Ser116, distinguish CD8+ T cells might prove to be a forerunner for HLA-B*57:01 from the abacavir-insensitive our better understanding of the basis of immune alleles and abacavir reacts with these two amino receptor recognition in drug hypersensitivities acid residues. In extending the fi nding that the and, more specifi cally, for elucidating the abacavir-HLA-B*57:01 association results from pathogenesis of some of the life-threatening specifi c binding of the drug to the HLA F pocket, drug-induced systemic reactions such as toxic amino acid sequences of a large number of HLA- epidermal necrolysis (TEN) and Stevens– B*57:01- and HLA-B*57:03-bound peptides Johnson syndrome (SJS). from untreated and abacavir-treated cell lines were Right at the time of the completion of this determined. Abacavir-treated HLA- B*57:01 monograph, newly published results explaining cells, but not treated HLA-B*57:03 cells, con- the molecular basis of AHS give every indica- tained unmodifi ed drug but no metabolites

[email protected] 3.4 Drug-Induced Hypersensitivity and Immune Receptors 73 indicating that abacavir bound non-covalently antigen- presenting molecules may reveal further and specifi cally with HLA-B*57:01. Up to 25 % fascinating insights into some poorly under- of the peptides bound to HLA-B*57:01 follow- stood, unpredictable, and potentially life-threat- ing treatment with abacavir proved to be differ- ening adverse drug reactions and ultimately lead ent to those before treatment but a change was to a better understanding of the immunopatho- not seen with peptides bound to HLA-B*57:03 genesis of autoimmunity, infectious diseases, or HLA-B*58:03. These results again suggested and cancer. that abacavir binds specifi cally to the antigen- binding cleft of HLA-B*57:01 and this alters the repertoire of self-peptides bound by HLA- 3.4.4 Carbamazepine and Other B*57:01 but not the repertoire bound by the HLA-Drug Hypersensitivity other HLA alleles. Sequences of peptides that Associations bind HLA-B*57:01 contained Trp → Phe at the C terminus (PΩ) but for HLA-B*57:03 PΩ was In addition to the associations of HLA-B* reversed, i.e., Phe → Trp. After abacavir treat- 57:01 with abacavir hypersensitivity and ment, an increased number of peptides with Ile fl ucloxacillin- induced liver injury (Sect. 5.1.10 ), or Leu at PΩ bound HLA-B*57:01. In another HLA-DRB1*15:01 with lumiracoxib-induced recent study, peptides eluted from an HLA- hepatotoxicity, and HLA-B*58:01 with B*57:01 single allele-transfected cell line treated allopurinol-induced SJS (see below), HLA- or not treated with abacavir were analyzed. A B*15:02 is strongly associated with signifi cant number of peptides with Val at the carbamazepine-induced SJS and TEN. As men- C-terminus were identifi ed in the presence of tioned in Sect. 3.4.3 above, the generality of the abacavir but no peptides with Val at the abacavir–HLA binding results was tested in a C-terminus were found in untreated cells. preliminary way in an examination of the well- Signifi cant numbers of peptides with terminal Ile established strong association between HLA- and fewer peptides with Trp and Phe also B*15:02 and carbamazepine-induced SJS/TEN occurred in the presence of abacavir. Taken in Asian populations. A non-covalent associa- together, the results of the abacavir–HLA bind- tion between carbamazepine and HLA-B*15:02 ing studies indicate that the drug positions itself was established by purifying HLA-B*15:02– at the bottom of the antigen-binding cleft extend- peptide complexes and sequencing of bound ing, via the cyclopropyl moiety (Fig. 3.16 ), into peptides. This revealed a preference for smaller the F pocket and changing the shape of the cleft. amino acids at key positions and signifi cant This results in preferred binding of smaller increases in the presence of some hydrophobic amino acids, an alteration in the repertoire of residues. Comparisons with HLA-B*15:01 self- peptides that bind HLA-B*57:01, and a T show that this allele is not associated with cell response to self-proteins presented only in carbamazepine- induced SJS and an important the presence of abacavir. Extension of this inves- difference between HLA-B*1502 and HLA- tigative approach to the antiepileptic carbamaze- B*15:01 is at position 156 (Leu for the former, pine, a drug strongly associated with Trp for the latter) near where the drug is thought HLA-B*15:02 (see below), showed that the drug to bind in HLA-B*15:02. binds to this allotype and, again, an altered rep- The carbamazepine–HLA-B*15:02 interac- ertoire of presented self-peptides results. This tion has also recently been used by S-I Hung and raises the possibility that antigen-presenting collaborators in Taiwan as a model for the study molecules may be susceptible to modulation by of the pathologic role of HLA in delayed-type drugs (and perhaps even toxins, environmental drug hypersensitivity. No intracellular metabo- chemicals, etc.) causing altered T cell immunity. lism or antigen processing was detected in If this is a general mechanism, investigations of the interaction between carbamazepine and associations of other drug hypersensitivities with HLA- B*15:02 in patients with the bullous skin

[email protected] 74 3 Mechanisms of Hypersensitivity

conditions and surface plasmon resonance assays Other drug hypersensitivities or intolerances showed that HLA-B*15:02, but not other HLA-B thought or claimed to be associated with HLA recombinant proteins, directly binds carbamaze- class I and/or class II alleles include trimethoprim- pine and the structurally related carbamazepine sulfamethoxazole with HLA-A30, aspirin with a 10,11-epoxide. For drug presentation and activa- number of different haplotypes (see Sect. 9.5.5 ), tion of cytotoxic T lymphocytes, endogenous hydralazine-induced systemic lupus erythemato- peptides in the antigen-binding groove were sus with HLA-DRw4, and nevirapine hypersensi- shown to be necessary. This is in contrast to aba- tivities with a surprising and confusing array of cavir which binds to HLA-B*57:01 without pep- associations. Again, these fi ndings highlight the tide loading. Modeling suggested that the Arg62 need for extensive phenotyping studies and inves- side chain, located in the B pocket of the HLA- tigations in much larger populations. B*15:02 protein, was the most likely binding site for carbamazepine by forming a hydrogen bond with the ketone of its 5-carboxamide group on 3.4.5 The Question of Direct Drug the tricyclic ring. Specifi c recognition of this Activation of T Cells Without group was supported by results obtained with Involvement of a Specifi c selected structural analogs. Peptide Allopurinol is an important treatment for hyperuricemia-related diseases, being used to Drawing on earlier speculations on the absence lower uric acid in gout, kidney stones, and Lesch– of prior sensitization in many drug reactions and Nyhan syndrome. Unfortunately, the drug is also the seminal studies of MHC-restricted metal ion a frequent cause of adverse drug reactions, and drug hypersensitivities mediated by T cell accounting, it is said, for up to 5 % of severe cuta- activation, others have suggested some modifi ca- neous adverse reactions. Reactions include drug tions to the possible cellular and drug interac- hypersensitivity syndrome, SJS, and TEN. In a tions involved in drug-specifi c recognition by Taiwanese study designed to identify genetic cells of the immune system. A number of obser- markers for allopurinol cutaneous reactions, the vations including the prevention of T cell activa- HLA-B*58:01 allele was identifi ed in 100 % of tion after removal of drug by washing, rapid 51 patients with allopurinol-induced serious calcium infl ux into T cells after exposure to drug, reactions but in only 15 % (20) of 135 tolerant and the fact that glutaraldehyde-fi xed APC can patients. These results indicate that in Han still present drug have led to the suggestion that T Chinese, allopurinol is strongly associated with cells rather than APCs recognize free, unpro- HLA-B*58:01 and this allele is an important cessed parent drug in allergic individuals. genetic risk factor for the serious cutaneous reac- Proponents of the so-called p–i concept (derived tions with systemic symptoms. from the proposed direct pharmacological inter- The NSAID and phenylbutazone derivative, action of drugs with immune receptors) state that feprazone, was found to be associated with HLA- “drugs bind specifi cally and reversibly to some of B22 in a Scandinavian study—93 % of patients the highly variable antigen-specifi c TCR in a with a fi xed drug eruption caused by the drug direct way, instead of covalently modifying the were HLA-B22 positive but no patients with MHC-peptide complex.” Direct and in-depth fi xed drug eruptions to other drugs were positive experimental fi ndings of the sort presented in the to HLA-B22 and this allele was found in 4 % of Ni, abacavir, and carbamazepine/HLA-B*15:02 healthy controls. However, a number of factors, investigations to support this hypothesis are lack- including the absence of HLA-B22 in 7 % of the ing, and with, for example, abacavir bound at the Scandinavian patients with feprazone-induced bottom of the HLA-binding groove, it is diffi cult fi xed drug eruption, need further scrutiny before to see how the drug can directly contact, or by feprazone-HLA-B22 can be taken as a diagnostic itself directly infl uence, the T cell receptor. marker. At the least, more extensive population Recently the proposed model restricted to T cell studies are needed. binding appears to have been modifi ed to

[email protected] 3.5 Desensitization of Drug-Allergic Patients 75 acknowledge and to try and accommodate the tigation of possible mechanisms. One current association and presentation of some drugs, or investigative approach involves the delivery of drugs with peptide, in the MHC peptide-binding increasing quantities of antigen at fi xed time groove. Given the apparent unequivocal defi ni- intervals to mouse bone marrow mast cells in tion of direct drug binding to the TCR without vitro together with the monitoring of granule modifi cation of the MHC–peptide complex, it is release by detection of β-hexosaminidase and the diffi cult to see how this accommodation can be metabolism of prostaglandins and leukotrienes. achieved. Both of these indicators were inhibited by desensitization and this was achieved by incremental increases in dosage. Importantly, the presence of 3.5 Desensitization of Drug- antigen was necessary for desensitization—as Allergic Patients long as antigen was maintained, and desensitization was maintained. Mast cells desensitized to Only immediate type hypersensitivity drug reac- dinitrophenol did not release preformed and de tions involving IgE antibodies and/or a mast cell- novo synthesized mediators such as TNF and mediated mechanism are considered eligible for IL-6. This may help to explain why desensitized desensitization. patients are not at risk of a delayed reaction. An adverse reaction to a drug can be major Experiments in which mast cells were sensitized problem in efforts to achieve successful treat- to dinitrophenol and ovalbumin showed that oval- ment for common and important diseases includ- bumin-desensitized cells responded fully to dini- ing infections, arthritides, allergies, and trophenol, proving antigen specifi city and that malignancies. Adverse drug reactions occur fre- signaling transduction pathways have not been quently, and as the number, chemical nature, and impaired during desensitization. Furthermore, novel pharmacological actions of registered FcεRI-bound antigen-specifi c IgE molecules did drugs continue to increase, such reactions can not disappear from the cell surface during desen- seriously interrupt therapy and leave patients sitization after becoming bound to small doses of with less than optimal treatments. Rapid drug antigen. These results are reassuring in that they desensitization (RDD) can provide an effective support both the proposed inhibition of the mast and safe means to continue vital therapies while cell response and the basis for the RDD proce- minimizing or avoiding the previously disruptive dures currently used. Over many years, a number impediment. The aim of desensitization is to of other mechanisms have been proposed to administer increasing amounts of drug in an explain the state of clinical tolerance resulting incremental and stepwise manner while at the from the practice of desensitization. The list same time avoiding or minimizing life- includes the formation of IgG blocking antibod- threatening, or even lesser adverse, symptoms. ies, consumption or blocking of the drug-reactive When successful, the procedure induces tempo- IgE antibodies by the gradually increasing quanti- rary tolerance to the drug allowing treatment to ties of administered drug, tachyphylaxis or deple- continue with optimal dosage. tion of the released mediators, hapten inhibition by In considering possible mechanisms leading to monovalent penicillin–protein conjugates, and rapid desensitization to a drug, the mast cell and desensitization of mast cells and basophils by possibly the basophil appear to be the cells most gradually increasing quantities of multivalent likely to be involved. In drug reactions involving drug–carrier complex. It must be concluded, how- complementary IgE antibodies, RDD appears to ever, that few truly revealing insights into the result in the mast cells becoming temporarily tol- mechanisms underlying mast cell tolerance or erant to the drug. A convincing explanation of hypo-responsiveness have been obtained so far. how RDD tolerizes mast cells or interferes with Note that desensitization to a drug does not their activation is lacking and the subject is inad- result in long-term tolerance to the adverse equately understood and in need of further inves- effects of the drug and this therefore means that

[email protected] 76 3 Mechanisms of Hypersensitivity patients need the desensitization procedure to be processed antigen is presented to CD4+ T cells repeated each time they are again exposed to the via the MHC class II molecules on antigen-pre- drug. However, if the medication is maintained, senting cells. The cells involved in many type IV for example, by daily dosage with pharmacologi- responses such as contact hypersensitivity and cally active levels, the desensitization state can psoriasis are Th1 and CD8+ cytotoxic T cells be maintained. while in a condition such as allergic contact dermatitis, CD4+ or CD8+ T cells can be activated depending on the antigen processing pathway. In 3.6 Delayed-Type (Type IV) general, CD4+ T cell activation seems to mediate Hypersensitivity Drug maculopapular and eczematous drug hypersensi- Reactions tivities while CD8+ T cell activation produces the more severe skin reactions involving bullous Unlike types I, II, and III hypersensitivities, manifestation. which are mediated by antibodies, delayed or In a hypersensitivity reaction in the skin such cell-mediated hypersensitivity, classifi ed as type as allergic contact dermatitis, there are two IV, is mediated by antigen-specifi c effector T phases of the hypersensitivity response, sensiti- cells and this means that the hypersensitivity zation (or initiation or induction) and elicitation. response can be transferred by purifi ed T cells or In the sensitization phase following drug admin- a cloned T cell line. Again in contrast to an istration, free drug or drug bound to a protein immediate reaction, a delayed-type hypersensi- carrier reaches the skin where it encounters kera- tivity reaction develops over a period of 24–72 h. tinocytes, present in great numbers and thought Delayed hypersensitivity responses have been to play a major role in the initiation of skin sen- used for many years to assess patients’ cell- sitization. As well as facilitating the formation of mediated immunity by the induction of indura- biologically active haptens and hapten binding tion and erythema 48–72 h after intradermal to protein, keratinocytes release chemotactic injection of so-called “recall” antigens from factors CXCL8, CXCL9, CXCL10, and CXCL11 Mycobacterium tuberculosis , Candida and and adhesion molecules (e.g., ICAM-1) on Trichophyton species, and tetanus toxoid. exposure to sensitizing agents. These chemotactic factors attract more cells to the active skin sites, thus increasing local immune activity. 3.6.1 The Cellular Basis of Type IV Sensitization proceeds with drug–carrier com- Hypersensitivity Cutaneous plex being taken up by immature Langerhans’ Drug Reactions and dendritic cells. These migrate to the draining lymph node and, with the stimulus provided by Delayed-type hypersensitivity reactions in the co-stimulatory molecules, become T cell- skin provoked by systemic drug administration activating cells. Processed antigen is expressed usually occur 7–10 days after the commencement as a drug–peptide complex in association with of therapy. Drug-induced skin reactions manifest MHC class I and II molecules on the surface of mainly as exanthemas, mediated by CD4+ and the mature antigen-presenting cells for presenta- CD8+ CD3+ T cells in the dermis and epidermis. tion to CD8+ and CD4+ T cells, respectively. Antimicrobial drugs, NSAIDs, and some analge- Dendritic cells, Langerhans’ cells, and skin mac- sic drugs are the biggest causes of drug-induced rophages express both MHC (class I and II) mol- adverse cutaneous reactions but a variety of other ecules and can activate CD4+ as well as CD8+ T drugs including anticonvulsants (e.g., carbam- cells. Keratinocytes are also important in the azepine), local anesthetics (lidocaine), cardiovas- elicitation phase and can present antigen via cular drugs (procainamide), and antipsychotics MHC I and MHC II molecules. T cells are (clozapine) are well known to cause reactions. activated, undergo clonal expansion, and give For most proteins and hapten–protein conjugates, rise to cells with different memory and effector

[email protected] 3.6 Delayed-Type (Type IV) Hypersensitivity Drug Reactions 77 functions, that is, Th1, Th2, or Th17 CD4+ or damage in the skin. Activation and proliferation CD8+ T cells. By the time of the elicitation of memory T cells in the dermis release chemo- phase, T cells have gained access to the skin and kines and infl ammatory cytokines such as IFN-γ following reexposure to the drug, skin symptoms and tumor necrosis factor-α/β (hereafter referred occur within about 48 h. Hapten-specifi c T cells to as TNF) that recruit macrophages to the site. recognize the hapten-peptide presented by den- Presentation of antigen by the newly arrived dritic cells and keratinocytes and the resultant macrophages has the affect of further amplifying activated T cells produce Th1 and Th17 cyto- the response. The released chemokines and cytokines such as IFN-γ, IL-12, IL-17, and IL-23. kines increase the permeability of blood vessels Note that although Langerhans’ cells have long leading to local swelling and induce the expres- been considered to be the classical cell net to sion of vascular adhesion molecules. IFN- γ is trap, process, transport, and present antigen to T the key cytokine and it plays a dominant part in lymphocytes, evidence, including mice lacking delayed hypersensitivity, upregulating T cell Langerhans’ cells, has shown that dendritic cells activation markers and MHC molecules, and aid- can act in their place if Langerhans’ cells are ing Th1 while suppressing Th2 cell differentia- absent or functionally affected. The nature of the tion. TNF also has a central role in delayed antigen, that is, the sensitizing drug or drug con- hypersensitivity, inducing chemokine produc- jugate, seems to determine which MHC mole- tion, upregulating expression of adhesion molecule is involved in antigen presentation. cules, and promoting the infl ux of infl ammatory Extracellular antigens (for example, contact cells. CD4+ and CD8+ T cell- mediated cytotox- allergens) are generally presented via MHC class icity of skin cells presenting drug can result from II molecules and intracellular antigens (for interaction with Fas/FasL, release of the cyto- example, drug–protein conjugates formed intra- lytic protein perforin and the serine protease cellularly) via MHC class I. Presentation, for granzyme B from cytotoxic T lymphocytes, and example, of the contact sensitizing agent nitro- release of granulysin from cytotoxic CD8+ T benzene sulfonic acid, appears to be by MHC II cells. In response to infl ammatory agents molecules. released by T cells, skin cells can in turn contrib- With the involvement of the co-stimulatory ute to infl ammation by releasing their own spec- B7-CD28 interaction, T cells are activated and trum of cytokines and chemokines that stimulates memory T cells can be found in the dermis. further leukocyte recruitment into the skin. As Other co-stimulatory molecules have also been understanding of the complexities of the mecha- identifi ed including OX-40-OX-40L, PD-1- nisms of the many processes that make up PD-L1 and PD-L2, RANK-RANKL, and CD40- delayed-type hypersensitivity responses CD40L (CD154). The receptor OX-40, also increases, two other agents, Il-12 and osteopon- known as CD134, and its ligand OX-40L are tin, are attracting the interest of researchers. seen as secondary co-stimulatory molecules IL-12, produced mainly by antigen-presenting expressed after T cell activation and important in cells, aids the proliferation and differentiation of maintaining T cell memory. RANKL, involved Th1 cells, augments IFN-γ production by these with dendritic cell maturation, belongs to the cells, and enhances NK and CD8+ T cell cyto- TNF cytokine family while PD-1 and its ligands, toxicity. Osteopontin (also known as ETA, early belonging to the B7 family, negatively regulate T T lymphocyte activation-1), a phosphoglycopro- cell responses. Binding of the co-stimulatory tein with cytokine and chemotactic functions, molecule CD-40 on antigen-presenting cells to has a Th2 suppressive effect augmenting Th1- its ligand CD40L activates these cells. During mediated allergy such as allergic contact derma- subsequent exposure of the memory T cells to titis and supporting dendritic cell migration and the sensitizing antigen, clonal expansion of the Il-12 expression and secretion. Discussion of activated T cells occurs and this ultimately osteopontin’s role in allergic contact dermatitis results in T cell-mediated infl ammation and cell is continued briefl y below in Sect. 3.6.3.1 .

[email protected] 78 3 Mechanisms of Hypersensitivity

Naive T cells

IL-12 IL-6 IFN-γ IL-21 IL-4 TGF-β TGF-β IL-6 IL-23

Th1 Th2 Th17 Tfh Treg T-bet GATA-3 RORγT Bcl-6 FoxP3 STAT4 STAT6 STAT3 STAT3

IFN-γ IL-4 IL-17A-F TGF-β IL-21 IL-2 IL-5 IL-6 IL-10 ICOS TNF IL-13 TNF IL-17

Fig. 3.17 Naive T cells, under the infl uence of cyto- immune responses to parasites; Th17 cells promote kines produced by other immune cells, undergo activa- infl ammation by helping to recruit neutrophils and Treg tion and polarization to distinct Th subsets. Each subset cells exert a number of inhibitory actions via cell con- displays a distinct cytokine secretion profi le resulting in tact. A more recently identifi ed CD4 T cell subset, different effector functions, e.g., Th1 cells activate mac- termed follicular helper cells (Tfh), provide a helper rophages; Th2 cells promote allergic responses and function to B cells

3.6.2 T Helper Cell Responses by IL-23, a cytokine produced by keratinocytes, and Th17 Langerhans’ cells, dendritic cells, and macrophages. Th17 cells are characterized by expres- As discussed, naive T cells differentiate into Th1 sion of distinct transcription factors RORγT, or Th2 cells during activation induced by interac- STAT3, and IRF-4 and the production of pro- tion with dendritic cells with toll-like pattern- infl ammatory molecules of the IL-17 family recognition receptors that detect the nature of the comprising IL-17A, B, C, D, E, and F (Fig. 3.17 ). antigen. This results in IL-12 production, the IL-17A gives rise to tissue infl ammation by pro- involvement of transcription factors T-bet, ducing pro-infl ammatory cytokines IL-6 and STAT4, or STAT1 within the T cell, the induction TNF and chemokines CCL2 (monocyte chemo- of Th1 differentiation, and production of IFN-γ. tactic protein-1 or MCP-1), CXCL1, and CXCL2 Th2 differentiation is the result of IL-4 cytokine that activate macrophages and granulocytes. IL-25 and GATA-3 and STAT6 transcription factor and IL-27 negatively regulate Th17 cells while involvement that drives production of IL-4, IL-5, Th17 polarization is inhibited by IL-2, IL-4 and IL-13. TGF-β and the transcription factor (induces Th2), and IFN-γ (induces Th1).

FoxP3 results in the Treg T cell subset that secretes Another more recently identiﬁ ed CD4 T cell TGF-β (Fig. 3.17 ). After deﬁ nition of the Th1 subset, termed follicular helper cells (Tfh), pro- and Th2 subsets more than 20 years ago, rela- vides a helper function to B cells. Tfh cells are dis- tively recent research has revealed a new class of tinguished from Th1 and Th2 cells by expression T effector cells Th17, induced from naive T cells of the chemokine CXCR5, their association with by the cytokines TGF-β and IL-6 and enhanced B cell follicles, and their B cell helper function.

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They produce ICOS (inducible T cell co- 3.6.3.1 Allergic Contact Dermatitis stimulator) and IL-21, a cytokine that stimulates B For a description of allergic contact dermatitis, cells to differentiate into antibody-forming cells. see Sect. 2.2.4.1 and Figs 2.4 and 2.5. Not all This cytokine is particularly interesting for those contact dermatitis has an immune basis; some concerned with understanding allergic mecha- irritants such as organic solvents, highly alkaline nisms and the treatment of immediate-type aller- drain cleaners, and sodium lauryl sulfate and gies. IL-21 knockout mice express higher levels of some phototoxins like the psoralens, paradoxi- IgE than normal mice and, in fact, IL-21 has cally used for the treatment of psoriasis, eczema, already been used to attenuate allergic responses and vitiligo, may also provoke reactions. by reducing both IgE and infl ammatory cytokine Common causes of allergic contact dermatitis production in mouse models for rhinitis and pea- include Ni metal, chromium, balsam of Peru, and nut allergy. Toxicodendron plants, for example, poison ivy, poison oak, and poison sumac. Causative agents tend to be reactive small molecules or haptens of 3.6.3 Delayed Cutaneous Adverse less than 1,000 Da that can easily penetrate the Drug Reactions skin barrier and form covalent adducts with cutaneous proteins. Allergic contact dermatitis is For the fi rst episode, these reactions generally regarded as a Th1 and CD8+ T cell-mediated dis- begin 7–21 days after contact with drug. ease and Ni allergy (see also Sect. 3.4.2 ), which Subsequent reactions begin 1 or 2 days after involves activation of HLA-restricted, skin- reexposure. Specifi city is usually demonstrated homing Ni-specifi c T cells by antigen-presenting by oral challenge with small doses of the culprit cells, is perhaps its best-known commonly occur- drug, a positive patch test or intradermal test gen- ring form. Both sensitization and skin reactions erally read after a delay of at least 48 h, and per- to Ni are thought to be mediated by CD4+ and haps a positive in vitro lymphocyte proliferation CD8+ effector T cells producing IFN-γ. During assay. Activated T cells are found in the skin and sensitization when no clinical symptoms are in some cases T cell lines and clones can be iso- apparent, mature Langerhans’ cells originating lated from blood and/or skin sites. Distinct sub- from skin sub-layers present Ni-peptide-MHC sets of T cells with their accompanying profi le of complex to T cells in local lymph nodes. Upon cytokines and chemokines promote the infl am- rechallenge with Ni, the effector phase of allergic matory and cytotoxic responses seen in the dif- contact dermatitis is activated to produce cutane- ferent clinical patterns characteristic of the ous infi ltration of Ni-specifi c and CCR4-positive various drug-induced adverse cutaneous hyper- T cells. Ni-specifi c cytotoxic CD8+ T cells sensitivities. Individual hypersensitivity erup- release infl ammatory cytokines that produce the tions are essentially the result of overlapping characteristic skin lesions at the site of Ni sensiti- cytokine actions with one or a few such actions zation (Chap. 2, Fig. 2.4 ). The T cell cytokine dominant and characteristic of the delayed drug IL-17 can be found in the skin of patients with hypersensitivity phenotypic pattern. This, and the allergic contact dermatitis. Some Ni-specifi c lack of histological and immunocytochemical CD4+ T cell clones isolated from the blood of criteria, has consequences for the diagnosis of allergic contact dermatitis patients express this drug-induced skin reactions where considerable cytokine which regulates the expression of adhe- effort is needed in the development of reliable sion molecules by keratinocytes and the synthe- and specifi c tests that can be easily undertaken sis and release of the chemokines IL-8 and (see Chap. 4 ). Although the mechanisms under- RANTES. IL-17 has been shown to be locally lying the different drug-related skin eruptions released by Ni-specifi c Th0, Th1, and Th2 lym- with an immunological pathogenesis are still far phocytes in the skin of patients with allergic from precisely defi ned, summaries of the prog- contact dermatitis where it amplifi es reactions ress are set out below. and modulates the pro-infl ammatory action of

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keratinocytes by acting together with IFN-γ IL-22 in the serum of patients whose psoriasis and IL-4. There still seems much to learn about had been cleared by treatment with the TNF the role of IL-17 in allergic contact dermatitis inhibitor etanercept; enhanced expression of but already the importance of this cytokine in IL-23 in patients with psoriatic lesions; and the the pathomechanism underlying the condition demonstration of IL-17 mRNA in psoriatic is apparent. lesions. Analysis of psoriatic skin lesions and The phosphoglycoprotein osteopontin is peripheral blood for the presence of IL-17- expressed by a number of different immune cells producing cells revealed Th17 cells localized in including effector T cells and keratinocytes in the lesions and the dermis. In addition, IL-17 allergic contact dermatitis. The molecule is mRNA expression returned to normal with expressed in secreted and intracellular form, it cyclosporin therapy and IL-22 mRNA expres- enhances Th1 and Th17 immunity, and protects sion moved in parallel with IL-17 changes, sug- against apoptosis. Experiments in mice have gesting that both Th1 and Th17 cells are active in shown that T cell clones secreting low levels of the infl ammatory stages of psoriasis. Following IFN-γ may compensate by secreting high levels the demonstration that in addition to Th1 cells of osteopontin which, in turn, down-modulates T producing IFN-γ, CD4+ T lymphocytes produc- cell IL-4 expression. In allergic contact dermati- ing IL-17 were also important in the pathogene- tis, secretion of IFN-γ by effector T cells induces sis of psoriasis, attention turned to the possible osteopontin in keratinocytes which ultimately importance of IL-17-producing CD8+ cells results in the attraction of infl ammatory cells. known to be present in psoriatic plaque. The demonstrations that osteopontin-null mice Investigations showed that CD8+ IL-17+ cells display a reduced infl ammatory response in con- produced the Th1-related cytokines IFN-γ and tact hypersensitivity and anti-osteopontin anti- TNF as well as the Th17 cytokines IL-17, IL-21, bodies partly suppresses established chronic IL-22, and upregulation of the transcription fac- contact sensitivity suggest that osteopontin may tor RORC. These results showing some common be a promising therapeutic target in allergic con- properties between CD8+ IL-17+ T cells and tact dermatitis. Th17 cells and the intriguing fi nding that CD8+ cells, unlike Th17 cells, can also make IFN-γ 3.6.3.2 Psoriasis and TNF may prove signifi cant in fully elucidat- Clinical aspects of psoriasis are presented in ing the pathogenesis of psoriasis. Sect. 2.2.4.2 and Fig. 2.6 . The classifi cation of T Currently, the broad understanding of the cells into Th1 and Th2 cells, essentially on the events and mechanisms leading to psoriasis is basis of their defi ning cytokines IFN-γ and IL-4, as follows. Antigen-presenting cells, probably respectively, and the fairly recent identifi cation Langerhans’ cells, in the skin migrate to regional of a new type of T cell, Th17, together with the lymph nodes where they interact with T cells. The realization of its importance in infl ammation, nature of the presented antigen is not known but has led to the reexamination of many diseases co-stimulatory factors from the antigen-presenting previously considered to be solely Th1 or Th2 cell are believed to be intercellular adhesion mol- mediated. So far, in murine models at least, some ecule 1 (ICAM-1, CD54) and lymphocyte func- diseases, previously thought to be Th1-mediated tion-associated antigen 3 (LFA-3, CD58). These responses, have been found to involve both Th1 molecules interact with their complementary and Th17 cells. Th17 cells produce IL-17, TNF, receptors on the T cell, lymphocyte function-asso- IL-6, IL-21, and IL-22 which are upregulated ciated antigen 1 (LFA-1, integrin), and LFA-2 during infl ammatory disorders and which pro- (CD2), respectively. Activated T cells return to the duce thickening of mouse epidermis suggesting skin where local effects in the dermis and epider- a role in psoriatic infl ammation. Other fi ndings mis of released pro-infl ammatory cytokines such suggestive of a role for Th17 cells in psoriasis as TNF produce the infl ammation and epidermal include the reduction of levels of IL-17 and hyper-proliferation seen in psoriasis.

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3.6.3.3 Maculopapular Exanthema kine CCR6 and aid infi ltration and survival of A case of maculopapular exanthema induced by neutrophils leading to the sterile pustular erup- amoxicillin with lesions on the trunk and hands is tions found in AGEP patients (Fig. 2.8 ). shown in Fig. 2.7 together with a clinical description in Sect. 2.2.4.3. Lymphocytes (CLA+, 3.6.3.5 Drug Reaction (Rash) with CD3+, DR+, CD25+) expressing adhesion mol- Eosinophilia and Systemic ecules are attracted from the blood by adhesion Symptoms molecules expressed by endothelial cells and The pathophysiology of drug reaction (some- keratinocytes and by chemokines such as CCL27 times designated rash) with eosinophilia and (also called cutaneous T cell-attracting chemo- systemic symptoms (DRESS), also called drug- kine CTACK). induced hypersensitivity syndrome (DIHS), is Both CD4+ and CD8+ T cells are found in the still being worked out (see also Sect. 2.2.4.5 ). skin and blood of patients with maculopapular Activated CD4+ and CD8+ T cells expressing exanthema, but fi ndings on the relative impor- CCR10 and producing type 1 cytokines, chiefl y tance of these cells differ with some authors stat- IFN-γ, are found in the blood of DRESS patients ing that CD4+ cells predominate and infl ict cell in the acute phase and these cells increase in damage by expressing high levels of perforin and proportion to the severity of the skin reaction granzyme B while CD8+ cells are found mainly (Fig. 2.9 ). Interestingly, T cell clones from car- in the epidermis. Other results have shown that bamazepine- and lamotrigine-sensitive patients CD8+ cells predominate in acute lesions of the react specifi cally with antigen-presenting cells epidermis and are the major drug-specifi c cyto- apparently without the formation of reactive toxic cell found in the blood of most patients with metabolites and processing, much like the situa- penicillin-induced maculopapular exanthema. tion with a superantigen. The T cell clones pro- Examination of cellular infi ltration in the skin of duce perforin and secrete IL-5 as well as IFN-γ, patients during patch testing demonstrated rapid the former accounting for the eosinophilia asso- recruitment of CD8+ cells after skin contact with ciated with the syndrome. Many investigators drug and before appearance of other cells particu- believe that a concomitant human herpes virus 6 larly CD4+ T cells. Both type 1 and type 2 cyto- (HHV-6) reactivation with hypogammaglobu- kines are produced; IFN-γ (type 1) activates linemia caused by the drug is associated with dendritic cells and keratinocytes; IL-5 (type 2) the hypersensitivity syndrome. This remains to together with eotaxin (CCL11) recruits and acti- be established. vates eosinophils. Other chemokines including While mentioning DRESS, it is opportune to CCL20, CXCL9, and CXCL10 appear to be comment on drug-induced allergic hepatitis. As involved in skin homing. During the acute phase in DRESS, this condition is associated with fever, CD4+ cells express perforin. rash, eosinophilia, and liver infi ltrates and the reaction is generally a type IV hypersensitivity 3.6.3.4 Acute Generalized response involving CD4+ cells, CD8+ cytotoxic Exanthematous Pustulosis lymphocytes, NK, Kupffer and dendritic cells. Activated drug-specifi c CD4+ and CD8+ T cells Type II hypersensitivities may also sometimes producing the neutrophil-attracting chemokine occur. There are two main hypotheses for the CXCL8 (IL-8), infi ltrate the skin of patients mechanism of drug-induced liver injury (DILI) with acute generalized exanthematous pustulo- caused by immune processes. First, the drug or sis (AGEP) (Sect. 2.2.4.4 ) and can be detected active metabolite(s) acts as a hapten and binds to in peripheral blood, in positive patch test biop- endogenous proteins forming conjugates that sies, and in T cell lines and clones. CXCL8- induce antibody- and/or T cell-mediated injury. producing effector memory T cells express Proponents of the second hypothesis suggest that mainly IFN-γ, GM-CSF, TNF, and sometimes most individuals are tolerant to immune- mediated IL-4 and IL-5. These cells express the chemo- DILI and reactions occur only when tolerance is

[email protected] 82 3 Mechanisms of Hypersensitivity overcome. Although the cellular events remain perforin, TWEAK (TNF-like weak inducer of poorly defi ned, knowledge of underlying mecha- apoptosis), and TRAIL (TNF-related apoptosis- nisms of idiosyncratic DILI is even more inducing ligand) pathways. These cytotoxic fragmentary. mediators are found in the serum as well as in blister fl uid where levels are high and where they 3.6.3.6 Fixed Drug Eruption occur with other cytokines including IFN-γ, Mediated by activated CD8+ T cells, fi xed drug IL-10, and IL-18. Several studies suggest that eruption (FDE) is a disease instigated by drugs in TNF has an important role in TEN and this more than 95 % of cases. In regression, large appears to be supported by the success of the numbers of CD8+ effector memory T cells of TNF-targeted monoclonal antibody infl iximab phenotype CD3+, CD45RA+, CD11b+, and (see chapter 11, Sect. 11.1.3.3 ) in promoting the CD27− are found in lesions in the epidermis. resolution of lesions in a number of patients. Reexposure to the culprit drug rapidly leads to a These outlined fi ndings are the conclusions conversion of this benign state to one of aggres- assembled from a number of different investiga- sive cell damage. T cells secrete IFN-γ in high tors, but the explanations leave signifi cant doubts amount as well as TNF, perforin, granzyme B, since some key points remain unexplained. In and Fas ligand (FasL) which initiates cell killing particular, the number of infi ltrating infl amma- by binding to its receptor FasR on keratinocytes. tory cells in the skin lesions is claimed to be too The presence of the “dormant” CD8+ T cells in few to explain the widespread killing of keratino- “resting” lesions explains why patch testing is cytes. In the fi rst place, both of the two favored negative on normal skin but reactivation occurs pathways to cell death, viz., granzyme B- and when patches are applied to areas of residual perforin-mediated exocytosis and Fas-FasL kill- lesions. For a clinical description of FDE see ing, are not restricted to TEN and SJS—both Sect. 2.2.4.6 and Figs 2.10 and 2.11 . pathways are upregulated in some other adverse cutaneous reactions such as maculopapular ery- 3.6.3.7 Toxic Epidermal Necrolysis thema where widespread cell destruction does and Stevens–Johnson not occur. The second inadequacy of the dual Syndrome pathway explanation is the need for cell-to-cell These diseases (Sect. 2.2.4.8 ) are provoked by contact for killing when there seems to be not drugs in more than 90 % of cases with sulfon- enough infl ammatory cells for this to occur. amides, anticonvulsants, some NSAIDs, and These doubts have been expressed by Chung and allopurinol most frequently involved. It is not yet coworkers in Taiwan whose investigations understood why and how a cutaneous adverse recently provided a better understanding of the drug reaction very occasionally progresses to the immune mechanisms and biomarkers of TEN life-threatening TEN or SJS. Clinical features of and SJS and promise new approaches for the both syndromes are similar with the extent of management of these diseases. Gene expression necrotic epidermis/skin detachment greater in profi ling, PCR, and immunohistochemical meth- TEN (>30 %; Fig. 2.14) than in SJS (<10 %) and ods identifi ed granulysin rather than Fas, FasL, the predominance of lesions around mucosal ori- soluble FasL, granzyme B, or perforin as the fi ces in SJS (Fig. 2.15). In fact, the two disorders major cytotoxic molecule responsible for kerati- are considered by many to be variants of the nocyte necrosis in TEN/SJS. Granulysin, a mem- same disease with different severity. In TEN, ber of the saposin-like family of membrane- blister fl uid contains many activated HLA class disrupting proteins, is a cationic cytolytic and I-restricted, drug-specifi c CD8+ CD56+ cyto- pro-infl ammatory protein contained in the cytotoxic T cells with natural killer (NK) cell fea- lytic granules of cytotoxic T lymphocytes and tures. These kill lymphocytes and particularly NK cells. Chung et al. showed that granulysin keratinocytes via, according to different from blister fl uid, in the 15 kD secretory form researchers, several mechanisms including the (a precursor of the 9 kD form), was present in a Fas/FasL (CD95/CD95L), TNF, granzyme B, concentration two to four times higher that soluble

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FasL, granzyme B, and perforin. Depletion of ticing clinician, especially those without spe- granulysin reduced cytotoxicity and when it was cialty knowledge of immunology and injected into mouse skin it produced TEN- and dermatology, the fi eld of drug-provoked cutane- SJS-like skin necrosis. In addition to its cytotoxic ous reactions is an area of great diffi culty starting effects, granulysin is a chemoattractant for other with the requirement of identifying the culprit infl ammatory cells and aids the expression of drug, often amongst multiple drugs being taken. some chemokines and cytokines including There then remains the need to undertake or order RANTES (CCL5), MIP-1α (macrophage infl am- appropriate tests without further aggravating the matory protein-1α, CCL3), MCP-1 (monocyte condition; institute appropriate management chemotactic protein-1, CCL2), MCP-3 (mono- measures; identify other drugs that may be a risk; cyte chemotactic protein-3, CCL7), IL-1, IL-6, and to take measures, including instruction of the IL-10, and IFN-α. patient, to avoid further reactions. A fairly recent In summary, the demonstrations of a pathoge- interesting area of investigation that is particu- netic mechanism for TEN/SJS and that secretory larly promising has emerged from demonstrated granulysin is a key toxic molecule responsible for associations between HLA alleles, certain drugs, disseminated keratinocyte killing open the way diseases such as TEN/SJS, and different human for the development of new diagnostic and thera- populations. Apart from the presentation of drug peutic targets for the diseases. However, impor- or drug metabolite to T cells, HLA alleles may tant questions concerning operative mechanisms also be responsible for genetic susceptibilities for in TEN and SJS remain. For example, what are drug-induced cutaneous reactions. As pointed the precise molecular mechanisms involved in out by Chung et al., “Understanding the molecu- the interactions between the offending drugs, lar mechanism of the interaction of HLA, offend- HLA, and the T cell receptor? What are the precise ing drugs and TCR, as well as CTLs/NK cells steps between taking the drug and the secretion activation, would facilitate the development of of granulysin? How is secretion of the cytolytic new approaches for the management of SJS/ protein regulated? What are the identities of TEN.” With relevance to pathomechanisms and the determinants recognized in the immune pro- regard to classifi cation of reactions, attention has cesses? The benefi cial effect of infl iximab when been drawn to the particular cell type(s) recruited used for selective TNF blockade in some cases of during the so-called second step of drug-induced TEN, and the importance of TNF in causing skin infl ammation following the involvement of direct cytotoxicity and apoptosis, must also be drug-specifi c T cells in the fi rst step. The impor- considered and somehow incorporated into a sat- tant involvements of eosinophils with DRESS isfying explanation of the pathogenesis of this and neutrophils with maculopapular exanthema intriguing toxidermia. and AGEP illustrate the point. As discussed in Chap. 4 , the demonstration or 3.6.3.8 Delayed Cutaneous Drug detection of individual or patterns of cytokines Hypersensitivity Reactions. and chemokines is a promising approach for Conclusions improving the reliability and specifi city of diag- In reviewing what is currently known about the nosing some drug-induced cutaneous hypersensi- pathophysiology and mechanisms underlying the tivity reactions. Surprisingly, this diagnostic T cell-mediated delayed allergic drug reactions it strategy still seems to be underutilized but signifi - is apparent that knowledge of the different cuta- cant advancements probably depend on fi rst reli- neous reactions is still widely incomplete and ably implicating a suitable disease-specifi c agreement, even on some basic processes, is marker or spectrum of markers. Finally, the aller- often inconsistent or lacking. Absence of agree- genic determinants recognized in the cellular ment on the identity of the often-bewildering immune processes remain largely unexplored and number and nature of cytokines and chemokines undefi ned. Identifi cation of the structures of said to be involved is particularly apparent for drug–peptide complexes presented by the MHC some of the drug-induced reactions. For the prac- and fi ne-structural detail of drug determinants

[email protected] 84 3 Mechanisms of Hypersensitivity recognized by the T cell receptor remain areas IgG but may be IgM), and subsequent sorely in need of both investigation and secure Fc-mediated cell destruction and clearance by fi ndings. Progress on these points is needed to macrophages. Activation of complement may reliably identify potentially cross-reacting drugs occur leading to intravascular lysis and renal fail- for patients and offers the possibility of selecting ure. Understanding the mechanisms involved in or tailor-making interfering inhibitory or compet- the action of drug-dependent antibodies is coming molecules to mitigate drug-specifi c reactions. plicated by the observation that drugs that cause some of the worst reactions including hemolysis, renal failure, disseminated intravascular coagula- 3.7 Type II Hypersensitivity tion, and death appear to proceed via a different Drug Reactions mechanism, often involving complement-activating antibodies. The so- called “unifying The classical drug-induced type II hypersensitiv- hypothesis” has been advanced to explain all ity is the well-known immune cytotoxic reaction three types of antibodies implicated in DIIHA. to high doses of penicillin that results from bind- This hypothesis is based on known fi ndings dating of the drug to red cells. This causes the red ing back to Landsteiner of the generation of three cells to be recognized as foreign, resulting in IgM populations of antibodies to an injected hapten– and IgG antibodies reacting with the drug–cell protein complex—one population to the hapten, membrane protein complex. The antibody–anti- one to hapten plus carrier determinants, and one gen complexes so formed activate the classical to the protein carrier. A further, and more recent, complement pathway causing cell lysis and death proposed mechanism for DIIHA is based on non- and the antibody-coated red cells can interact immune adsorption of the drug onto the red cell with macrophages leading to Fc-mediated cell membrane. Cefotetan, often used prophylacti- destruction by the reticuloendothelial system. cally in some surgical procedures, is the most Another example of a type II cytotoxic antibody- common cause of DIIHA with another cephalo- mediated drug reaction when the drug appears to sporin, ceftriaxone, the second most common form an antigenic complex with the red cell sur- cause. For the period 1985–1997, the FDA face is drug-induced immune hemolytic anemia reported 85 cases of cefotetan-induced hemolytic (DIIHA). The drugs most frequently associated anemia with 15 (18 %) fatalities. It is therefore with DIIHA are some cephalosporins (especially usually recommended that all cephalosporins cefotetan and ceftriaxone) and penicillins should be avoided in patients where DIIHA is (especially piperacillin). DIIHA can also be asso- thought to be a possibility but in vitro hapten ciated with red cell autoantibodies induced by the inhibition experiments have shown that only drug affecting the immune system without cefotetan and cephalothin inhibited anti-cefotetan becoming bound to the red cell surface, that is, IgG and IgM antibodies. It would be prudent to the drug does not participate in the antigen–anti- remember, however, that these were in vitro fi nd- body reaction. Such antibodies are referred to as ings and the in vivo consequences may be quite drug-independent. Prototype drugs involved in different especially if allergic reactions (and IgE drug-independent autoantibody formation are antibodies) are involved. With piperacillin, the methyldopa and the chemotherapy drug, fl udara- third most common drug causing DIIHA, bine. In this form of DIIHA the clinical and labo- immune complexes seem to be involved. ratory fi ndings are identical to autoimmune As well as erythrocytes, other cells including hemolytic anemia. It is not known why drugs platelets (thrombocytes) and some hematopoietic sometimes induce drug-independent autoanti- precursor cells can be affected by drug-induced bodies to red cells or what mechanism is involved. type II hypersensitivity reactions. Drug-induced The mechanism of DIIHA when the drug partici- thrombocytopenia for example is increasing as pates as the antigen is thought to proceed by more drugs are released and used. A number of attachment of the drug to the red cell in vivo, different mechanisms appear to be involved. interaction with drug-reactive antibodies (usually Drugs may bind covalently to the platelet mem-

[email protected] 3.7 Type II Hypersensitivity Drug Reactions 85 brane producing a hapten–glycoprotein conju- drug is withdrawn. The prototype drugs in this gate with an antigenic determinant(s) that is category are gold and procainamide. Finally, recognized by antibody. Drugs implicated in this heparin and heparin-like drugs can induce throm- form of thrombocytopenia include penicillins bosis by binding to surface-bound soluble plate- and cephalosporins in particular. Quinine, quini- let factor 4 (PF4), a small chemokine CXCL4 dine, sulfonamides, and NSAIDs may interact that promotes coagulation and is released from non-covalently with platelet membrane glyco- the alpha granules of activated platelets during proteins, including the von Willebrand factor platelet aggregation. Antibodies to the heparin– receptor GPIb-IX-V (GP for glycoprotein) and PF4 complex bind to receptors on the platelet activated integrins GPIIb/IIIa, forming drug–gly- surface via their Fc pieces producing platelet coprotein non-covalently linked complexes. For activation. This mechanism is basically different antibody binding to occur, the presence of the from the other fi ve described mechanisms in that drug is essential—in the absence of the drug, activation and aggregation of platelets is the antibodies do not bind to the platelet surface and result rather than cell lysis and hemorrhage mak- thrombocytopenia does not occur. It remains ing the reaction more like a type III than a type II uncertain whether the antibodies are directed to hypersensitivity response. the drug alone or to the complex of drug and Acute agranulocytosis is rare but when it does platelet glycoprotein. A third mechanism of occur, drugs are responsible in more than 70 % of drug-induced thrombocytopenia is seen with the cases. In its immune form, antibodies are pro- antiplatelet GPIIb/IIIa inhibitor drugs lotrafi ban, duced to circulating neutrophils and/or myeloid tirofi ban, and eptifi batide, the novel cyclic hepta- precursor cells. Immune-mediated agranulocyto- peptide from the venom of the southeastern sis is rapid in onset with symptoms generally pygmy rattlesnake. By binding to the glycopro- occurring within a few days. Drugs commonly tein complex, these drugs induce a conforma- associated with the condition include quinine, tional change and a new determinant to which quinidine, β-lactams, pyrazolones, propylthio- antibodies bind and cause cell destruction. The uracil, clozapine, ticlopidine, carbamazepine, drug does not physically form part of the deter- chlorpromazine, and some sulfonamides. minant. Another inhibitor of platelet activation Numerous other drugs have been implicated in sometimes administered is abciximab, the Fab one or only a few cases. Several mechanisms fragment of a chimeric human–mouse monoclonal have been advanced although detailed and antibody that binds to the platelet glycoprotein convincing evidence is not always offered. Some receptor GPIIb/IIIa. Some patients, even without of the implicated drugs such as penicillins and prior exposure to the monoclonal agent, react to aminopyrine are thought to act as haptens that the mouse component of the hybrid, supporting elicit antibody formation against neutrophils and the belief that natural antibodies may be involved their subsequent destruction. In the case of in the recognition. Such recognition of murine aminopyrine-induced agranulocytosis, antibod- antigens on a chimeric human–mouse antibody ies are directed to neutrophil cell membrane anti- fragment is similar to the recognition by natural gens modifi ed by a reactive metabolite of the antibodies of the chimeric monoclonal antibody drug. Antibody recognition of metabolites was cetuximab (Sect. 3.1.1 ; Sect. 11.1.3.2 ). This also demonstrated for metamizole and diclofenac humoral form of immune-mediated drug-induced in cases of agranulocytosis induced by these thrombocytopenia is regarded as drug-specifi c drugs. In addition to drug-dependent antibodies since the antibodies are formed against the drug of the IgG and/or IgM class, autoantibodies were itself and platelets are destroyed in the process. In found in 13 cases of drug-related agranulocytosis a fi fth mechanism, drug induces the formation of due to penicillins, dimethylaminophenazone, autoantibodies to glycoproteins on the platelet propyphenazone, metamizole, and diclofenac. In surface. The antibodies bind to the platelet anti- the case of clozapine-induced agranulocytosis, gens without participation of the drug and the the drug is converted to the reactive nitrenium ion resultant thrombocytopenia can persist when the which binds to cellular proteins and accelerates

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Antigen Antibody

100

Antigen clearance in absence of antibody

Antigen Antigen-Antibody complexes & serum sickness reaction

0.1 2 468 10 12 14 16 18 20 22 24 26 28 30 Days

Fig. 3.18 Relationship between antigen introduction, subsequent immunological events, and time in serum sickness apoptosis of neutrophils. Propylthiouracil was hypersensitivity reactions. Thus, it can be said that shown to lyse neutrophils via a complement- penicillins can cause all four types of hypersensi- dependent mechanism. An immune mechanism tivity reactions. Other drugs that produce similar does not seem to be involved with drugs such as serum sickness-like reactions include cephalospo- ticlopidine, busulfan, chlorpromazine, and meth- rins, sulfonamides, ciproﬂ oxacin, tetracycline, lin- amizole, each of which has a direct toxic effect comycin, NSAIDs, carbamazepine, allopurinol, on myeloid precursors. thiouracil, propranolol, griseofulvin, metronidazole, furoxone, captopril, gold salts, methyldopa, halothane, ﬂ uoxetine, barbiturates, and monoclo- 3.8 Type III Hypersensitivity Drug nal antibodies. β-Lactam drugs are considered the Reactions most common cause of serum sickness elicited by nonproteins but drugs by themselves are thought Serum sickness (see Sect. 2.2.3) can occur in to be poor antigens for the production of the good response to foreign proteins such as streptokinase antibody responses necessary to induce serum and to antitoxins, antivenins, and vaccines. As sickness. Circulating antigen–antibody complexes mentioned above in Sect. 3.2.8.2 , type III drug- are formed after drugs become protein bound in induced hypersensitivities, that is, antigen–antibody vivo and stimulate IgG and/or IgM antibodies. The complex-mediated reactions, occur in some cases liberation of vasoactive amines is thought to play a that closely resemble classical serum sickness. part in tissue deposition. Antigen also interacts Penicillin has long been known to become with complementary IgE antibodies on mast cells antigenic by conjugating to proteins in vivo to pro- and basophils leading to the release of PAF and duce drug–protein complexes that mediate type III other mediators, platelet aggregation, and further

[email protected] Summary 87 release of histamine and serotonin. The resulting involve drug-specifi c antibodies or, more usually, increase in vascular permeability facilitates the drug-specifi c T cells. Eosinophilic pneumonia deposition of immune complexes which, in turn, can be caused by almost any medication while produces complement activation, the formation of reports of drug-induced hypersensitivity pneu- C3a and C5a, an infl ux of infl ammatory cells to monitis, a combined type III and IV reaction in a the sites of immune complex deposition, and Th1/Th17 response, are increasing, particularly release of further infl ammatory mediators. Drug to antineoplastic drugs. immune complexes are normally rapidly cleared via the antibody Fc piece or complement binding to cells of the reticuloendothelial cells but if this Summary does not occur, for example, because of the high concentrations of immune complexes, deposition • For many drugs it has not been possible to of complexes in glomeruli, arteries, endocardium, explain allergic reactions on the basis of their spleen, and other organs and infl ux of infl amma- chemical reactivity, protein-binding capacity, tory cells may result. In a graph that relates the their biotransformed or degradative products, time of occurrence of tissue lesions to the clear- or the presence of a reactive impurity. ance of antigen and developing antibody produc- • Some allergic responses, sometimes even life- tion, Fig. 3.18 summarizes the immunologic threatening as with anaphylaxis, occur on fi rst events in the patient after antigen exposure. Serum exposure to a drug. concentration of protein-bound drug initially • There is at least one group of drugs, the neuro- decreases sharply as a result of intravascular and muscular blockers (and probably more to be extravascular equilibration and levels continue to identifi ed), that can specifi cally elicit decrease normally as the protein is catabolized antibody- induced mast cell activation and until antibody levels increase, causing rapid release without fi rst undergoing coupling to a immune elimination. The dashed red line in macromolecular carrier. For these drugs, the Fig. 3.18 at about day 6 represents the course of di- or multi-valency which is an inherent part antigen decline in the absence of antibody- of the molecular structure, initiates mediator mediated antigen elimination. From about day 14, release by cross-linking cell-bound antibodies. soluble circulating complexes of antigen with IgG • The initial event in the activation of mast cells or IgM form and may begin to deposit in a number for mediator release is the binding of IgE anti- −10 of tissue sites leading to the clinical manifestations bodies to the high-affi nity (K a 10 M) FcεRI and pathologic fi ndings of serum sickness. IgE receptor abundantly expressed on the Hypersensitivity vasculitis induced by drugs is mast cell and basophil surfaces. another manifestation of a type III response. Drugs • Released preformed mediators of infl amma- involved include some β-lactams, particularly, tion and anaphylaxis stored in the cytoplasmic amoxicillin and cephalexin, cotrimoxazole, granules of mast cells include histamine, hep- NSAIDs, monoclonal antibodies, and chemothera- arin, platelet-activating factor (PAF), sero- peutic drugs such as tamoxifen and erlotinib. A tonin, the enzymes tryptase, chymase, and proportion of small-vessel vasculitis patients have carboxypeptidase, and eosinophil, neutrophil, anti-neutrophil cytoplasmic antibodies. Although and monocyte chemotactic factors. Newly there is evidence of a pathogenic role for these anti- synthesized released mediators include pros- bodies and they are used as a diagnostic marker, taglandin D2 , thromboxanes, and leukotrienes operative mechanisms underlying this hypersensi- LTB 4 , LTC4 , and LTD4. A host of cytokines tivity state are still far from established. (pro- and anti-infl ammatory), chemokines, Hypersensitivity reactions are one of a num- and chemotactic, stimulating, and growth fac- ber of different mechanisms producing drug- tors are also released. induced lung disease. These reactions result from • A second receptor for IgE, the low-affi nity interaction of drug with the immune system and receptor FcεRII also known as CD23, is

[email protected] 88 3 Mechanisms of Hypersensitivity

expressed on airways smooth muscle cells and asthma but much weaker in other asthmatics.

several types of hematopoietic cells including LTD4 is much more pronounced in asthmatic mature B lymphocytes, macrophages, mono- patients not sensitive to aspirin. cytes, dendritic cells, and eosinophils. • Cysteinyl leukotrienes are generated de • Histamine is synthesized from L -histidine novo from arachidonic acid by phospholi- by the inducible enzyme L -histidine decar- pase A2 with the initial participation of boxylase and inactivated by histamine 5- lipoxygenase-activating protein and the N-methyltransferase-catalyzed methylation of enzyme 5-lipoxygenase. the imidazole ring and oxidative deamination • The two human cysteinyl leukotriene recep-

of the primary amino group catalyzed by tors CysLT1 R and CysLT2 R do not bind the diamine oxidase. three cysteinyl leukotriene ligands equally: for

• The physiological and pharmacological CysLT 1 R, LTD4 > LTC4 = LTE4; for CysLT 2 R,

effects of histamine are mediated through four LTC 4 = LTD4 > LTE4 .

different receptors H1 , H2 , H3 , and H4 , all • PAF, 1-O -alkyl-2-acetyl-sn -glycero-3-phospho- members of the 7-transmembrane g protein- choline, a phospholipid of relatively simple coupled receptor (GPCR) family with amino but unique structure, produces both the signs terminal glycosylation sites and phosphoryla- and symptoms of anaphylaxis. PAF is also an tion sites for protein kinases A and C. important mediator in asthma and septic • Pathophysiological effects resulting from stimu- shock. Recent ﬁ ndings in the mouse identiﬁ ed

lation of the H 1 receptor include those responses a second pathway of anaphylaxis involving seen in immediate allergic reactions, viz, red- the IgG receptor FcγRIII and the release of ness, itch, swelling, asthma, anaphylaxis, bron- PAF as the major mediator. choconstriction, and vascular permeability. • Recruitment of the Syk kinase and subsequent

• H2 receptors appear to mainly mediate sup- phosphorylation activation steps involving pressive activities of histamine including gas- Lyn lead to mast cell activation demonstrating tric acid secretion, heart contraction, cell the importance of protein tyrosine kinases in proliferation, differentiation, and some effects the pathways that result in allergic inﬂ amma- on the immune response. tion and anaphylaxis.

• The H3 receptor regulates the synthesis and • Sphingosine-1-phosphate, a major regulator of release of histamine and also has a regulatory the vascular system and B and T cell trafﬁ ck- role in the release of neurotransmitters such as ing, is elevated in the lungs of asthmatics where serotonin, dopamine, and norepinephrine. it regulates pulmonary epithelium permeability

• The H4 receptor is functionally expressed on and is thought to contribute to the pathogenesis mast cells, eosinophils, monocytes, dendritic of anaphylaxis and rheumatoid arthritis. cells, and CD8+ T cells. The receptor exerts a • Mechanisms of anaphylaxis independent of chemotactic effect on several cell types asso- IgE and including PAF-induced shock have ciated with immune and inﬂ ammatory been suggested. responses such as allergy, asthma, rheumatoid • Urticaria is a heterogeneous disease with arthritis, and inﬂ ammatory bowel disease. many subtypes caused by a range of agents

• LTC4 and LTD4 are powerful mediators of and stimuli. Subtypes include urticaria due to asthma, airway hypersensitivity, and allergies genetic or immune mechanisms, urticaria with inducing bronchoconstriction, increasing vas- an autoimmune basis and nonimmune- cular permeability, and promoting mucous mediated urticaria, and angioedema.

secretion. LTE4 is present in greatest amount • The combination of actions of ACE inhibitors in vivo where it induces bronchial eosino- of decreasing angiotensin II and aldosterone philia and airway hyperresponsiveness. The and increasing and maintaining bradykinin lev-

bronchoconstriction provoked by LTE 4 is els may lead to ﬂ uid extravasation into subcuta- strong in patients with aspirin-sensitive neous tissue ultimately producing angioedema.

[email protected] Summary 89

• Angioedema may also occur following admin- The reaction is generally a type IV hypersen- istration of angiotensin II receptor-binding sitivity response involving CD4+ cells, CD8+ inhibitors such as losartan. cytotoxic lymphocytes and NK, Kupffer, and • The allergen-induced late phase reaction has fea- dendritic cells. Type II hypersensitivities may tures of a cell-mediated hypersensitivity response also sometimes occur. Knowledge of mecha- but shows some signifi cant differences best illus- nisms underlying idiosyncratic drug-induced trated by the different cytokine profi les. liver injury is limited. • An early clue to specifi c immune recognition • Granulysin appears to be a key toxic molecule of “small,” unbound chemicals and hence responsible for disseminated keratinocyte kill- drugs was the demonstration of selective ing in TEN/SJS. interaction of nickel ions with an MHC-II- • Examples of type II cytotoxic antibody- bound peptide. mediated drug reactions include drug-induced • Abacavir–HLA binding studies indicate that immune hemolytic anemia, drug-induced the drug changes the shape of the antigen- thrombocytopenia where a number of differ- binding cleft. This results in preferred binding ent mechanisms are involved and acute agran- of smaller amino acids, an alteration in the ulocytosis in which more than 70 % of cases repertoire of self-peptides that bind HLA- are caused by drugs B*57:01, and a T cell response to self-proteins • Type III drug-induced hypersensitivities, that presented only in the presence of abacavir. is, antigen–antibody complex-mediated reac- • Carbamazepine, a drug strongly associated tions, occur in some cases that closely resem- with HLA-B*15:02, binds to this allotype and ble classical serum sickness. Drugs implicated alters the repertoire of presented self-peptides. include β-lactams, sulfonamides, ciprofl oxa- The most likely binding site on the carbam- cin, tetracycline, lincomycin, NSAIDs, carba- azepine molecule is the ketone of its mazepine, allopurinol, thiouracil, propranolol, 5- carboxamide group on the tricyclic ring. griseofulvin, metronidazole, furoxone, capto- • The mast cell and possibly the basophil appear pril, gold salts, methyldopa, halothane, fl uox- to be the cells most likely involved in the etine, barbiturates, and monoclonal antibodies. desensitization of patients to drug allergies. Circulating antigen–antibody complexes are • Drug-induced delayed-type cutaneous hyper- formed after drugs become protein bound sensitivity reactions manifest mainly as exan- in vivo and stimulate IgG and/or IgM antibod- themas, mediated by CD4+ and CD8+ CD3+ ies. The liberation of vasoactive amines is T cells in the dermis and epidermis. There are thought to play a part in tissue deposition. two phases of the hypersensitivity response, • Hypersensitivity vasculitis induced by drugs is sensitization (or initiation or induction) another manifestation of a type III response. involving keratinocytes, Langerhans’, and Drugs involved include some β-lactams, partic- dendritic cells and elicitation via T cells. ularly, amoxicillin and cephalexin, cotrimoxa- • Some progress has been made in identifying zole, monoclonal antibodies, and NSAIDs. mechanisms underlying the different drug- • Hypersensitivity reactions are one of a related skin eruptions with an immunological number of different mechanisms producing pathogenesis but more precise defi nitions are drug-induced lung disease. Eosinophilic needed. Individual important drug-induced pneumonia can be caused by almost any delayed reactions include allergic contact der- medication while reports of drug-induced matitis, psoriasis, maculopapular exanthema, hypersensitivity pneumonitis, a combined AGEP, DRESS, FDE, TEN, and SJS. type III and IV reaction in a Th1/Th17 • Drug-induced allergic hepatitis, as in DRESS, response, are increasing, particularly to is associated with fever, rash, and eosinophilia. antineoplastic drugs.

[email protected] 90 3 Mechanisms of Hypersensitivity

matotoxicology. 7th ed. Boca Raton, FL: CRC Press; Further Reading 2008. p. 259–68. Gould HJ, Sutton BJ, Beavil AJ, et al. The biology of IgE Adams DH, Ju C, Ramaiah SK, Uetrecht J, Jaeschke H. and the basis of allergic disease. Annu Rev Immunol. Mechanisms of immune-mediated liver injury. Toxicol 2003;21:579–628. Sci. 2010;115:307–21. Greaves MW, Tan KT. Chronic urticaria: recent advances. Austen KF, Maekawa A, Kanaoka Y, et al. The leukotriene Clin Rev Allergy Immunol. 2007;33:134–43.

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[email protected] Diagnosis of Allergic Reactions to Drugs 4

Abstract In diagnosing drug allergies, history, skin testing, some in vitro laboratory tests and the challenge test are the backbone of the investigation. If skin prick testing elicits no reaction, intradermal testing is usually employed. The latter test is more sensitive but produces more false positives. The COADEX classifi cation should be used to assess clinical relevance of positive patch tests. Assays for drug-specifi c serum IgE antibodies are useful in cases of skin test-negative or equivocal reactors or when skin tests are unreliable/unavailable. In interpreting results of IgE antibody tests, receiver-operating characteristic (ROC) curves provide more information to aid discrimination between positive and negative results. Drug challenge is the best way to confi rm an allergic reaction, and it is considered to be the “gold standard” in the diagnosis of drug hypersensitivities. In anaphylaxis, the ratio of total to mature tryptase is typically less than 10. Given the technical improvements made with BAT and the test’s validation for a number of drugs, it continues to be applied to many drug reactions. Nonproliferation-based in vitro assays of cell surface activation markers, cytokines, chemokines, and skin-homing receptors will be increasingly applied to diagnosis. ELISPOT assays (e.g., for IFN-γ and granzyme B) show potential for diagnosis and the chemokine CCL27 and CLA are promising markers for aiding efforts to understand the relationship between T cells, drugs, and adverse delayed skin reactions.

In diagnosing drug allergies, the patient’s history, sometimes from inadequate descriptions and skin testing, some in vitro laboratory tests and the recall by patients taking different drugs simulta- challenge test are the backbone of the investiga- neously; agents used for skin testing are not tive procedures. A successful diagnosis of drug always ideal and are usually unstandardized; suit- allergy can be particularly difﬁ cult since each of able laboratory tests are not always available and these investigations has limitations and draw- sensitive enough for testing reactions with a backs. The history is usually pieced together humoral or cell-mediated immune basis; and drug

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 91 Relationships, DOI 10.1007/978-1-4614-7261-2_4, © Springer Science+Business Media, LLC 2013

[email protected] 92 4 Diagnosis of Allergic Reactions to Drugs challenge tests are involved, possibly harmful for the selection of measures for immediate treat- the patient and not always sensitive enough. Even ment if needed, and the formulation of a future so, by application of more than one of these four avoidance strategy. basic investigations and sometimes supplement- 1. A list of all the medications the patient is, or ing the diagnostic process with some more spe- has been, taking including over-the-counter cialized tests, an accurate diagnosis can usually preparations. be achieved. The four basic diagnostic investiga- 2. How much was (is being or has been) taken tions will be reviewed together with other tests and for how long? some of which are essentially still research tools 3. Which drug is the prime suspect of causing with yet to be established clinical diagnostic reli- the reaction and why? ability. Some of these more “specifi c” tests (that 4. When did the reaction occur and how long is “specifi c” in terms of the restricted or specifi ed did it last? nature of what they measure rather than the abso- 5. What was the temporal sequence of events lute preciseness of the measurement) may even- between the initiation of therapy and the tually occupy a regular place in the evaluation onset of symptoms? and management of patients with drug allergies. 6. Did the reaction occur on fi rst exposure to a drug? 7. What were the manifestations of the reac- 4.1 Case History tion? For example, if there was a skin reaction, describe it. Did any swelling, choking, In assessing a case of drug allergy, the patient’s shortness of breath, or itching result? Get a clinical history is the most important component list of all symptoms. of the diagnostic process. It should be self- 8. Has the patient recently been subjected to any evident that diagnosis is not just the selection, medical or dental procedures such as major ordering, and subsequent assessment of tests or minor surgery, radiographic investigation, some of which might not be needed if the physi- immunization, or tooth fi lling or extraction? cian spends the necessary time on a little forensic 9. Has the patient ever had a previous reaction questioning and analysis needed to assemble an to the suspected drug or to any other drug adequate case history. As part of medical training and is there a history of drug allergy? and from the experience of practice, clinicians in 10. Is the patient atopic and is there a family his- all disciplines of medicine learn the importance tory of drug allergy or allergy in general? of the medical history, or anamnesis, of a patient. 11. Does the patient have a viral infection or has The symptoms reported by the patient together he/she had one recently? with the clinical signs ascertained by direct phys- 12. Does the patient have any other disease, in par- ical examination, sometimes confi rmed by clini- ticular, asthma, cystic fi brosis, diabetes, etc.? cal and/or laboratory tests, enables the clinician 13. Questions on home environment, pets, hob- to make a diagnosis which is essentially based on bies etc. pattern recognition, context, and probability. In Answers to questions 3, 4, 5, and 7 can go a addition to the obligatory and standard patient long way toward helping to establish a fi rm diag- information that needs to be gathered such as nosis. In relation to points 4 and 5, information age, weight, height, past medical history, family on the temporal sequence of events can provide history, home environment, work, diet, medica- essential information needed to help determine tion, allergies, habits, smoker or not, alcohol con- the mechanism of the reaction. Immediate, IgE- sumption, and so on, assessment of a case of antibody- mediated reactions that can range from possible drug allergy must include a series of rel- a simple rash to full-blown anaphylaxis generally evant and specifi c drug-related questions. occur from only a few minutes to 1 h after drug Obtaining the following information is aimed at administration. Delayed or late reactions may providing answers that will help the diagnosis, occur from more than 1 h up to several days after

[email protected] 4.2 Skin Testing 93 administration. These reactions that may present as maculopapular rashes, fi xed drug eruptions, 4.2 Skin Testing and different exanthems suggest a drug-specifi c T cell-mediated mechanism. In response to ques- 4.2.1 General Aspects of Skin tion 7, a good description, or better, a direct view Testing for Drug or photograph (e.g., of skin reactions) of the clin- Hypersensitivity ical manifestations of the reaction, can be very informative. Symptoms that result from the acti- The amount of information gained from a vation of mast cells such as anaphylaxis, bron- patient’s history that is potentially useful in a chospasm, angioedema, and urticaria indicate an diagnostic investigation of a suspected drug immune mechanism mediated by drug-reactive allergy will vary with different patients. After IgE antibodies. Note, however, that some drugs, gathering and recording the most comprehensive for example, vancomycin and contrast media can and detailed history that is obtainable from a have a direct, nonimmune effect on mast cells. patient, the usual next step in the diagnostic pro- Cutaneous reactions of the type mentioned above cess is to make a clinical assessment and decide generally indicate responses mediated by T cells. whether the available information suggests that a Some patients are taking multiple drugs so it is hypersensitivity reaction is a possibility or if the often diffi cult to identify a culprit drug. This is a probability of such a reaction is so low that an particular problem in surgery, when many drugs allergic reaction is considered to be most unlikely. are often administered in a short time. Reactions In the latter case, a drug challenge test might be that occur during anesthesia usually cannot be carried out to eliminate any suspicion of an aller- determined reliably without detailed investigations. gic drug sensitivity while in the former case some Diffi culties in identifying the culprit drug and the testing to investigate the possible presence of a sequence of events before and after the reaction hypersensitivity should be pursued. The fi rst test may also occur in patients who are infants or young to consider in this situation, and usually the fi rst children, aphonic, dyspneic, or unconscious, where choice, is skin testing. Skin tests, in prick or skin pigment masks some cutaneous reactions and intradermal form, are an appropriate diagnostic in specifi c clinical situations, for example, during tool for hypersensitivity reactions of the immedi- childbirth and hemodialysis. In all of these cases it ate type, for example, in cases of suspected ana- is the physician or anesthetist who is in the position phylaxis, angioedema, bronchospasm, urticaria, and has the responsibility of identifying the pro- conjunctivitis, and allergic rhinitis. The risk of voking agent and ensuring that this sometimes systemic reactions is lower with skin prick and potentially vital information is recorded for future patch testing than with intradermal testing. access. This may take the form, for example, of a Patients with a history of previous anaphylactic letter given to the patient or the patient being reactions or uncontrolled asthma, pregnant advised to wear a warning chain or bracelet. women, and small children should be considered In conclusion, although taking a detailed drug at higher risk of systemic and anaphylactic reac- history is an integral part of assembling a patient’s tions but the risk of fatality due to prick and patch medical history, in cases of suspected drug reac- tests is remote and anaphylactic reactions are tions it is a task that is all too often inadequately rare. The patch test and, sometimes after very performed by busy clinicians recording incom- careful consideration of the risks, late readings of plete information in a perfunctory way. It should the intradermal test are appropriate for investigat- be remembered that a possible consequence of ing drug reactions such as contact dermatitis, failing to document and subsequently consult a erythema multiforme, exanthematous drug history of a drug reaction(s) before prescribing eruptions, fi xed drug eruptions, leucocytoclastic medications could be the basis of a successful vasculitis, Stevens–Johnson syndrome (SJS), and malpractice suit. toxic epidermal necrolysis (TEN). Note that for

[email protected] 94 4 Diagnosis of Allergic Reactions to Drugs these latter high-risk patients, a careful risk to Before undergoing skin testing, the patient benefi t analysis must be undertaken and, if it is should discontinue taking antihistamines at least decided it is in the patient’s best interest to proceed 5 days before testing commences, and if the with testing, test solutions should start at very patient is pregnant the testing physician should high dilutions and all safety precautions, includ- be informed. Other drugs that must be discontin- ing hospitalisation of the patient (see Sects. 4.2.2 ued prior to skin testing include β-adrenergic and 4.4 ), should be taken. In cases of severe late blocking agents, corticosteroids, including prep- and delayed reactions, the time intervals between arations for topical application, tricyclic antide- tests may be extended or patch testing alone may pressants like amitryptyline and histamine be employed. H2 -receptor antagonists. There appears to be no Many of the drugs that will need to be tested universal agreement that the latter two groups of are available only in tablet or capsule form. In drugs interfere. A consent form setting out the these cases, the tablet, pill, or capsule contents reasons for the test and its procedures, benefi ts, should be accurately weighed before grinding to and risks should be read and signed. Once these a fi ne powder in a mortar. Sugar or other protec- necessary preparations have been completed, the tive coating on pills should be removed fi rst. patient’s blood pressure, pulse, peak expiratory Since most solid dosage forms are formulated fl ow, and oxygen saturation levels are measured with other substances, and bearing in mind the and recorded. These measurements may be specifi ed dose per tablet/capsule of drug, the repeated during and at the conclusion of the test. quantity of the powdered material suffi cient to contain the desired amount of drug needed to prepare the skin test solution or patch test is obtained 4.2.2 Skin Prick Test Method by weighing out the appropriate portion of the original weight of the tablet/capsule. A prick test Skin prick testing is performed on the volar solution is prepared by dissolving or dispersing aspect of the forearm or on the back. A solution the desired concentration of powdered material in of the drug is placed on the skin and a new lancet sterile physiological saline or a 1:1 solution of or fi ne-gauge needle is passed through the drop glycerine:saline. For some drugs, adjustment of pricking the top layer of skin without drawing the pH or use of other solvents may be needed to blood. This allows the test solution to gain access aid solubility. This was done, for example, to to cells of the dermis. The excess solution remain- increase the solubility of trimethoprim and sulfa- ing on the skin is wiped away. If a number of methoxazole where 0.1 M sodium hydroxide and solutions are to be tested, a testing grid can be benzyl alcohol were added (see Sects. 6.2.1.2.1 placed or drawn on the skin and the drops are and 6.2.2.2). Lack of water solubility may also be placed in the center of each grid square. As a overcome by dissolving the drug in dimethyl sulf- positive control, histamine 10 mg/ml (1 % w/v) oxide (DMSO) and further diluting with sterile or codeine phosphate 90 mg/ml (9 % w/v) is physiological saline to the desired concentration. included while the vehicle used for the test solu- In all such cases of where special diluents are tions (often physiological saline or glycerin- used, the diluent itself must be used as a control. saline) is used as a negative control. Results are Prior to proceeding with skin test studies on read and recorded after 15–20 min in the case of any drug, the optimal test concentration for that immediate reactions and after 24–72 h (and drug must be determined. This is the highest con- sometimes longer) for late and delayed reactions. centration that produces no skin reactions in a A longer time interval before reading is often group of control subjects who have never been necessary for aminoglycoside antibiotics like exposed to the drug, and in a group of nonallergic neomycin (see Sect. 6.1.5.1). The size of the patients who have been exposed to the drug, but wheal and erythema (fl are) reaction can be which will elicit a positive response in patients recorded in different ways and different scoring allergic to the drug. systems are employed. For documenting the

[email protected] 4.2 Skin Testing 95 reaction, the outer margins of the wheal and fl are test is more sensitive than the prick test but pro- can be traced on the skin using ink that is trans- duces more false positives, that is, it is less spe- ferable to adhesive translucent cellophane tape. cifi c. For some practitioners of skin testing, the Alternatively, the reactions can be traced directly extra sensitivity provided by the intradermal test onto translucent tape placed over the reaction and the tendency of the prick test to produce site. The recorded area of the reaction on the more false negatives are more than compensated tape can be quantifi ed by weighing, planimetry, for by the greater specifi city of the prick test. or by computerized scanning. Calculation of the This superior specifi city, it is argued, correlates so-called mean diameter is one of the most fre- better with clinical allergy and the sensitivity is quently used procedures to compare and record said to be adequate when suffi ciently potent skin test reactions. This involves the measure- extracts are used. Additional advantages of prick ment of the largest diameter ( D) of the wheal and testing are claimed to be the presence of glycerin a diameter perpendicular to this (d ). The mean (usually at 50 % concentration) that is thought to diameter is then obtained from (D + d)/2. For provide better stability for extracts, superior inhalant and many other allergens, a 3 mm or patient comfort and safety, and an economy of greater wheal mean diameter in the prick test is time in the test’s application. generally considered a positive reaction. The number of drugs employed in skin testing, although growing, is still too few to confi dently 4.2.3 Intradermal Testing assign an overall fi gure for positivity (e.g., wheals resulting from most anesthetic agents are Intradermal tests need to be performed only when usually smaller than those resulting from other the prick test gives negative results with testing agents causing anaphylaxis), and because peni- beginning after a 15–20 min break. Solutions cillins are the drugs most studied in skin testing, for testing are prepared under a laminar fl ow criteria for positivity with these drugs tend to be hood in sterile physiological saline or sterile applied when reading the results of skin tests saline with 0.5 % phenol no longer than 2 h with other drugs. In practice, this means that a before administration. Intradermal testing is wheal diameter at least 3 mm greater than the normally contraindicated in patients who have negative control is considered a positive result in developed SJS, TEN, erythema multiforme, or most tests on drugs. For some drugs, a wheal leucocytoclastic vasculitis although testing such diameter at least half the diameter resulting from high-risk patients may be judged to be necessary the positive control is taken as a positive reaction in special circumstances and with all safety by some investigators (see for example Sect. precautions in place (see Sects. 4.2.1 and 4.4 ). So 7.4.3.3.1 ). Reactions read at later time intervals far, there appears to be no report of skin testing to detect late-phase reactions and delayed reac- provoking or causing the reoccurrence of toxic tions involve the documentation of induration in epidermal necrolysis. particular but also any erythema, papulation, and Using a 25-gauge needle with the bevel upper- vesicles. Any erythema with infi ltration is con- most at an angle of 15–20° to the skin surface, sidered to be a positive reaction. Late-phase 0.02–0.05 ml of the test solution is injected intra- reactions are IgE antibody-dependent but differ dermally (Fig. 4.1 ) on the forearm or back to pro- from immediate type I reactions by the involve- duce a small blister or bleb (Fig. 4.2 ). Depending ment of neutrophils, eosinophils, and mononuclear on the drug and the severity of the patient’s drug cells (see Sect. 3.3 ). Whether or not the investi- reaction, the initial injection may range from a gation is aimed at detecting late or delayed small dilution of 1:10 or 1:100 of the prick test reactions, skin prick test sites should also be read concentration to more extreme dilutions of up to 1 day after the tests. 1:100,000. If no reaction is seen, the concentration If skin prick testing elicits no reaction, intra- is increased in logarithmic steps until the fi nal dermal testing is usually employed. The latter concentration is reached and this maximum

[email protected] 96 4 Diagnosis of Allergic Reactions to Drugs

25 gauge needle epidermis

dermis ~ 15o angle of entry bevel up

subcutaneous tissue

muscle

Fig. 4.1 Diagrammatic representation of an intradermal skin test

Fig. 4.2 An intradermal skin test being performed. Note the small blister or bleb formed from the solution injected into the dermis (Photograph courtesy of Dr. Paul A.J. Russo, Department of Clinical Immunology and Allergy, Royal Adelaide Hospital)

concentration should not be exceeded. Histamine base 0.01-0.1 mg/ml (0.001-0.01% w/v) and the Fig. 4.3 Clear positive wheal and ﬂ are reactions to solvent for the drug are included as positive and amoxicillin (A) and histamine (H) control solution negative controls respectively. Some regard a posi- (10 mg/ml) following intradermal testing (Photograph tive test as the appearance of an erythematous courtesy of Dr. Paul A.J. Russo, Department of Clinical Immunology and Allergy, Royal Adelaide Hospital) wheal after 20 min with a diameter at least twice that of the initial bleb. Probably a more rigorous and widely accepted positive threshold is an 5 mm. Tests should be read after 15–20 min for increase in diameter of more than 3 mm over the immediate reactions and after 48 and 72 h for initial 20 μl injection bleb (usually ~ 2 mm) accom- delayed reactions. With some drugs even later panied by erythema (Fig. 4.3 ). This threshold readings may be needed. The positive predictive caters for different sizes of the bleb formed from value of a skin test is, in general, high so a positive the injected solution and generally means that a result can be taken as diagnostic but a single positive reaction has a diameter of more than negative result cannot necessarily rule out drug

[email protected] 4.2 Skin Testing 97 allergy. After a careful analysis of the risks and baboon syndrome, contact dermatitis, fi xed drug benefi ts, consideration should be given to pro- eruption, lichenoid rash, and acute generalized ceeding to a second test using the next step-up exanthematous pustulosis, and it may be of value concentration of drug. If the intradermal test for drug reaction (rash) with eosinophilia and remains negative, the patient should be contacted systemic symptoms (DRESS). It is thought to be 1 week later and asked whether or not the test site of less use in investigating urticaria, SJS and is still negative, or the patient should be instructed TEN. A signifi cantly higher number of positive to return for the site to be inspected. reactions appear to occur with drug-induced maculopapular rashes than with urticarial or erythrodermic reactions. Positive reactions are often 4.2.4 Patch Tests observed with some drugs, for example, β-lactams particularly amoxicillin, cotrimoxazole, pristina- 4.2.4.1 General Aspects of Patch mycin, hydroxyzine, pseudoephedrine, carbam- Testing azepine, heparinoids, diltiazem, diazepam, and Adverse drug reactions affecting the skin are a tetrazepam. The skin site tested can also be frequently seen condition and, as for other mani- important. In fi xed drug eruptions, patch tests festations of adverse drug reactions, for patients should be performed on both normal skin and on receiving multiple medications it is often diffi cult residual pigmented skin sites of the eruption. In to identify the culprit drug from history alone. the case of toxic necrolysis, a positive patch test Patch testing with drugs, in both pure and com- with cotrimoxazole was obtained on skin previ- mercial form, is valuable in helping to determine ously affected by necrolysis but not at other less, the cause of drug-induced cutaneous drug reac- or unaffected, sites. tions and for studying the pathophysiological mechanisms underlying the reactions. 4.2.4.2 Concentrations of Drug A strong positive feature of the patch test is that it An important advantage of patch tests is their is both a screening test for hypersensitivity and a capacity to utilize virtually any form of a com- provocation test in the target organ, the skin, mercialized drug. When used with a pure drug, a where it can be seen as reproducing the disease. 10 % solution in water or alcohol or a 10 % dis- Unlike patch testing that requires no hospital sur- persion in petrolatum is employed. Some drugs veillance during testing, rarely provokes anything require a specifi c vehicle—for example, alcohol, more than a mild reaction and can be used with not water, should be used for estrogen and pro- commercial forms of drugs, intradermal tests gesterone, and β-lactams tend to give false nega- with drugs carry a greater risk and can only be tive results in an aqueous vehicle and should performed with pure dissolved material in free, therefore be tested at 5–10 % in petrolatum. In sterile solution. On the other hand, patch tests are some cases olive oil, rape oil, or acetone may be less sensitive than intradermal tests. The specifi c- used. Test materials prepared from tablets and ity and sensitivity of patch tests is said to be in the other formulations always contain additives range 70–80 % depending on the test agent but in (diluents, binders, pigments, sweeteners, lubri- four separate studies where a single drug was cants, disintegrants, granulating agents), so a implicated with high imputability, drug patch higher quantity, usually 30 %, of powdered tab- tests proved positive in 31.7–50 % of patients let, pill, or capsule contents is mixed with the with a cutaneous adverse drug reaction. Results diluent for patch application (see Sect. 4.2.1 ). of patch tests depend on the clinical features of Whenever possible these excipient substances the cutaneous drug reaction, the drug tested and should also be tested. If pure drug is used for test- its concentration, the vehicle used, and some- ing, concentrations should begin at about 0.1 % times on the skin test site. It has been claimed that and progress to 1–10 % if results are negative. the test is valuable for investigating generalized If DRESS, SJS, or TEN patients must be tested, eczema, maculopapular rash, photosensitivity, with, for example, aciclovir, carbamazepine, or

[email protected] 98 4 Diagnosis of Allergic Reactions to Drugs

Fig. 4.4 Patch tests showing (a ) patches in place during 4 days further development (d ), both the nickel (+++) and development of reactions and (b ) immediately after their p-phenylenediamine (+) reactions have intensiﬁ ed. For fur- removal, 2 days after being applied. A strong positive reac- ther details of notation of reactions see Fig. 4.5 and tion (++) to nickel (N) and an extreme positive reaction Table 4.1 . From Spiewak R. Patch testing for contact allergy (+++) to fragrance mix (F) are visible in (b ). An equivocal and allergic contact dermatitis. The Open Allergy Journal reaction (?+) to p-phenylenediamine (P) is seen in ( c). After 2008;1:42. Reproduced with permission of the author pseudoephedrine, testing should start with much ties in mind of substituting another drug and lower concentrations to avoid any relapse of a gaining an understanding of cross-reactions, cutaneous adverse drug reaction. related drugs with similar pharmacological action and/or chemical structure should be tested along 4.2.4.3 Method, Materials, with the suspected culprit drug. Patch tests should and Reading be applied to the upper back usually at a distance After obtaining informed consent from the of about 2–4 cm from the centerline using patient, patch testing should begin 6 weeks to 6 Finn Chambers on Scanpor® (a hypoallergenic months after healing of any cutaneous adverse tape) (Epitest Ltd Oy, Tuusula, Finland); Van der drug reaction and 4 weeks after discontinuing Bend Square Chambers (Brielle, The immunomodulating drugs such as glucocorti- Netherlands); IQ Chambers™ (Chemotechnique, coids or cyclosporin and topical corticosteroids. Vellinge, Sweden); or T.R.U.E. Test® (Thin-layer Due to hormonal effects on test results, patch test- Rapid Use Epicutaneous Test, SmartPractice, ing should not be performed during pregnancy Denmark ApS). Figure 4.4a shows patch tests in nor should it be done after a patient has experi- place on the back of a patient, clear positive reac- enced strong ultraviolet exposure, for example, tions (++ and +++) to two contact allergens, after a seaside holiday. Otherwise, patients should nickel and a fragrance mix, after removal of the be in good health, free from virus and other infec- patches (b, c, d) and an equivocal positive reactions, fever, and inﬂ ammation. With the possibili- tion (?+) (c) and weak positive reaction (+) (d) to

[email protected] 4.2 Skin Testing 99

Table 4.1 Scoring of patch test reactions due to drug concentrations that are too low, Score Clinical picture Interpretation insuffi cient occlusion, an inappropriate vehicle, NT reading at too early a time, absence of a drug IR Different types of reactions Irritant reaction metabolite, or reduced or impaired immunoreac- (e.g., vesicles, blister, tivity of the patient. Any repeat test should be necrosis) carried out after a delay of 2 months. Nonspecifi c − No reaction Negative reaction irritant reactions can be induced by some drugs ? or ?+ Faint erythema only—no Doubtful or infi ltration equivocal reaction including colchicine, misoprostol, used for the + Erythema, infi ltration, Weak positive prevention of nonsteroidal anti-infl ammatory possibly discrete papules reaction drug (NSAID)-induced gastric ulcers and by ++ Erythema, infi ltration, Strong positive sodium lauryl sulfate which is included in some papules, vesicles reaction commercial drug formulations. +++ Erythema, infi ltration, Extreme positive confl uent vesicles reaction 4.2.4.4 Clinical Relevance Typical scoring and notation system used when reading of a Positive Patch Test patch test results. See for example, Wilkinson DS, et al. Acta Derm Venereol. 1970;50:287; Spiewak R. The Open Allergy A positive patch test (indicating contact allergy) J. 2008;1:42; Lachapelle J-M, Maibach HI. Patch testing is not necessarily a positive diagnosis of allergic and prick testing. A practical guide offi cial publication of contact dermatitis as indicated by the fact that the ICDRG, 3rd edn. Berlin: Springer-Verlag;2012 some patients with a positive patch result never Note that follicular reactions (usually denoted by F and not shown here) can be categorized as doubtful reactions experience clinical symptoms after exposure to NT not tested the test agent. The so-called COADEX classifi cation is useful in the attempt to assess relevance of positive patch tests. COADEX stands for: • C (Current)—Current relevance: patient p- phenylenediamine . Some practitioners state exposed to drug or test agent prior to current that in order to avoid immediate reactions, patch episode of dermatitis and improves when test reactions should be read at 20 min while exposure ceases. others believe that such reactions should be • O (Old)—Past or old relevance of dermatitis tested and ruled out before patches are applied. to the test agent. Reactions are read after 48 and 72 or 96 h. In • A (Active)—Patient actively sensitized. some cases a reaction occurs in less than 2 days Presents with a late reaction. (e.g., abacavir) while with some other drugs such • D (Doubtful)—Relevance diffi cult to assess. as corticosteroids, aminoglycoside antibiotics, Not known if exposure is current or not. and phenylephrine, reactions may occur after 6 or • E (Exposed)—History of exposure but no der- 7 days. If the result is negative on day 4, a further matitis; no history of exposure but a positive reading should be carried out on day 7. Reactions patch test. based on morphology are scored as shown in • X (Cross-reaction)—Positive test due to cross- Table 4.1 with reactions rated as +, ++, or +++ reaction with another agent. interpreted as increasingly positive (Fig. 4.5 ). Positive reactions to structurally related com- 4.2.4.5 Photopatch Testing pounds may refl ect cross-reactions but “polysen- A photopatch test is used to investigate a drug sitization” or reactions to a number (e.g., at least reaction when a phototoxic or photoallergic fi ve or six) unrelated compounds may indicate a reaction is suspected. A drug patch is applied, highly developed drug sensitivity or the so-called removed after 1 day (or on day 2 if necessary), and angry back or excited skin syndrome refl ecting the skin is irradiated with 5 J/cm2 UVA. The test false positives. A complete absence of positive is read after 2, 3, or 4 days. In performing the test, reactions despite a history highly suggestive of the test agent is applied to two sites, with only drug sensitivity may be a false negative response one being irradiated with UV light. A positive

[email protected] 100 4 Diagnosis of Allergic Reactions to Drugs

Fig. 4.5 Examples of patch test reactions showing (left be viewed with reference to Table 4.1 . From Spiewak R. to right) an equivocal positive reaction (?+); weak posi- Patch testing for contact allergy and allergic contact der- tive reaction (+); strong positive reaction (++); extreme matitis. The Open Allergy Journal 2008;1:42. Reproduced reaction (+++); irritant reaction (IR). This fi gure should with permission of the author result at the irradiated site with a negative result when skin tests are unreliable or unavailable. at the nonirradiated site suggests photoallergy A good example of their value is seen in cases whereas equal positive responses at both sites where patients have a convincing history of suggest contact allergy. immediate allergy to a β-lactam but a negative skin test, a discrepancy that is occasionally seen. 4.2.4.6 Control Subjects They are also valuable for patients on certain For control subjects, healthy volunteers with or medications that must be curtailed for skin test- without exposure to the drug can be recruited as ing, in patients with widespread skin affl ictions negative controls although some investigators such as eczema or psoriasis and when applied to prefer to include dermatitis patients who proved sera taken at the time of a reaction (e.g., an ana- positive to a drug but negative to the drug being phylactic reaction during anesthesia), before sur- investigated. Such control subjects should only gery, and serum taken before or after death. Until be included 6 weeks to 6 months after taking the the introduction of the radioallergosorbent test drugs, and they should enter the study with ethics (RAST) for the detection of allergen-reactive IgE approval and signed informed consent. antibodies in the early 1970s, skin testing and occasionally the Prausnitz-Kűstner test were the only ways of confi rming a diagnosis of type I 4.3 Serum Immunoglobulin E IgE-antibody-mediated disease. In its earliest Antibody Tests form, the test was a solid phase radioimmunoassay utilizing allergen preparations attached to While the patient history and skin tests form the paper discs and a labeled second antibody to core of the investigative procedures for accu- detect IgE antibodies bound to the immobilized rately diagnosing hypersensitivities to drugs, allergens. Despite the introduction of some varia- tests for the detection of IgE antibodies specifi c tions such as liquid phase systems, the solid for individual drugs are a valuable diagnostic aid phase technology has persisted due to improve- to supplement and confi rm skin test fi ndings, and ments in the types of solid phases with enhanced in some cases they offer some advantages. Serum allergen binding capacity, the use of monoclonal IgE antibody determinations are useful in cases antibodies, enhanced sensitivity and accuracy, of skin test-negative or equivocal reactors or and the introduction of calibration methods for

[email protected] 4.3 Serum Immunoglobulin E Antibody Tests 101

drug-protein drug-protein solid conjugate support complex chemical (or (or + coupling antibodies in solid patient’s serum support

some antibodies react drug-with suitable drug-solid phase with drug-protein or functional group) complex) drug-solid phase complex

wash & add labelled second antiboby

detect label in tube− gives a measure of wash the amount of drug- specific antibodies in patient’s serum

Fig. 4.6 Diagrammatic representation of the solid phase Structure–activity studies on drug-induced anaphylactic immunoassay procedure for the detection of drug-reactive reactions. Chem Res Toxicol 1994;7:703. Reproduced IgE antibodies in serum. From Baldo BA and Pham NH. with permission from American Chemical Society the quantitative expression of results. At the same complex is incubated with patient’s serum, time, automation has led to greater precision and washed, and specifi cally bound IgE antibodies are reduced turnaround times, and the addition of detected with an enzyme-, fl uorescent-, fl uoroen- carefully selected nonisotopic labels and sub- zyme-, chemiluminescent-, or radiolabeled sec- strates has improved sensitivity and accuracy and ond antibody (Fig. 4.6 ). The biotin-avidin (or reduced nonspecifi c binding. Automation and streptavidin) reaction utilizing labeled biotin can widely adopted calibration methods have also also be employed as a highly sensitive detection made interlaboratory standardization possible. procedure. In preparing the drug solid phase, the drug must be immobilized but unaltered antigenically for recognition by its complementary anti- 4.3.1 In Vitro Detection bodies. The chemical procedure selected to couple of Drug-Reactive IgE a drug either directly to a solid support or fi rst to a Antibodies carrier macromolecule (usually protein) and then to the support depends on the functional groups Specifi c immunoassays to detect IgE antibodies in that are available on the drug or can be added to the sera of drug-allergic patients are being increas- the drug. Nucleophilic addition reactions employ- ingly used to supplement patient’s histories and ing, for example, bis-oxirane (1,4-butanediol skin tests and increasingly valued as the range and diglycidyl) or divinyl sulfone have proved to be sensitivities of tests increase. In its simplest form, widely applicable, generally not chemically protein-coupled or free drug is covalently linked destructive for the drug to be coupled and easy to to a solid phase (sometimes via a spacer arm); the carry out. Coupling employing carbodiimides is

[email protected] 102 4 Diagnosis of Allergic Reactions to Drugs

Insoluble Drug containing: carbo- - Nucleophile hydrate (-OH, -NH , -SH) Drug-Solid phase complex or Coupling agent 2 + + or or Soluble Drug-Protein conjugate protein - Carboxyl group carrier (-COOH)

(a) Bis-oxirane

OH OCH2CHOHCH2O(CH2)4OCH2CHOHCH2 Nuc Drug NaOH CH CHCH O(CH ) OCH CH 2 2 2 4 2 CH2 Nuc Drug or or + + pH11-12 O O

Protein NH2 Protein NH CH2CHOHCH2O(CH2)4OCH2CHOHCH2 Nuc Drug

(b) Divinylsulfone (DVS)

OH OCH2CH2SO2CH2CH2 Nuc Drug NaOH or CH =CHSO CH=CH Nuc Drug or + 2 2 2 + pH9-12 Drug Protein NH2 Protein NH CH2CH2SO2CH2CH2 Nuc

Protein NH R N=C=N R'HO C Drug Protein NH C Drug 2 + + pH5.5 O O

Fig. 4.7 Some widely applicable chemical strategies drug-induced anaphylactic reactions. Chem Res Toxicol for the preparation of drug-carrier complexes. From 1994;7:703. Reproduced with permission from American Baldo BA and Pham NH. Structure–activity studies on Chemical Society also a relatively mild procedure applicable to a identify the precise structures features that variety of different drugs. Carbodiimides can be constitute the drug allergenic determinants used to form peptide bonds at room temperature (Fig. 4.8 ). Inhibition assays are indispensable in by linking free carboxyl groups on drugs to pro- the study of immediate allergic reactions to drugs tein amino groups (Fig. 4.7 ). Chemical methods not only to check specifi city but also for cross- can usually be applied to conjugate most drugs reactivity and allergen structure investigations but sometimes it is chemically diffi cult and time and to help bridge the clinic-laboratory divide by consuming and may even involve the employment relating and correlating, on a quantitative basis, of complex synthetic steps. Sometimes a simple allergic recognition in the test tube and in the and convenient alternative to a complex synthesis patient. Although this correlation has not always is the substitution of suitable structural analogs been as good and as informative as one would that contain the identical or closely related anti- hope, improvements in assay specifi cities and genic determinant structures. This was the strat- sensitivities and insights gained from increased egy used to prepare specifi c, IgE-reactive solid knowledge of precise allergenic structures and their phases antigenically similar if not identical to the recognition promise to signifi cantly increase muscle relaxants succinylcholine and gallamine the value and utility of drug-IgE antibody both of which lack suitable functional groups for measurements. So far in clinical allergy practice easy coupling to protein or insoluble carbohydrate and research, quantitative immunochemical supports (see Sect. 7.4.3.4 ). approaches have been largely ignored and inter- By using the binding assay in an inhibition pretations of allergic recognition and sensitivity format, the investigator can confi rm specifi city have been based on undoubtedly clinically rele- of the antibody-drug reaction, compare quan- vant skin tests and other tests that are not strictly titatively the recognition of similar drugs, and quantitative.

[email protected] 4.3 Serum Immunoglobulin E Antibody Tests 103

incubate + RT, 1h (or )

antibodies in free drug some antibodies add solid support some free antibodies react patient's serum react with drug with drug-protein or drug-solid phase complex

wash & add labelled second antiboby

detect label in tube − gives a measure of the amount of drug-specific antibodies bound to the solid phase; wash compared with control (no inhibitor) to obtain percent inhibition

Fig. 4.8 Diagrammatic representation of the solid phase allergenic determinants. From Baldo BA and Pham NH. immunoassay inhibition procedure used in quantitative Structure–activity studies on drug-induced anaphylactic hapten inhibition studies for the establishment of speciﬁ c- reactions. Chem Res Toxicol 1994;7:703. Reproduced ity of antibody binding and for the identiﬁ cation of drug with permission from American Chemical Society

4.3.2 Tests for the Clinic and clinical utility making this allergy test system and Research the best described and most studied one available but, even so, the range of available tests for indi- For routine application of immunoassays to vidual drugs remains inadequate. In fact, at the detect drug-reactive IgE antibodies, the current research level, a relatively wide range of drug situation is characterized by the restricted avail- solid phases have been prepared and successfully ability of specifi c drug tests. Perhaps the only used to identify drug- reactive IgE antibodies in suitable standardized testing agents that can be allergic patient’s sera and, in some cases, to iden- accessed widely are the Phadia, now Thermo tify allergenic structural determinants. Results Scientifi c, ImmunoCAP® small range of drug with some of these “in-house” tests have been at solid phases comprising penicilloyl G, penicil- least as good and sometimes superior to the cor- loyl V, amoxicilloyl and ampicilloyl determinants, responding commercial assays. Details for the cefaclor, chlorhexidine, chymopapain, gelatin preparation of drug solid phases and procedures (bovine), insulin (human, bovine and porcine), for the assays utilizing them have been published pholcodine, morphine, and succinylcholine. For for the following drugs: research purposes only, ACTH, protamine, and Penicillins (penicilloyl and penicillanyl determi- tetanus toxoid are offered. Each ImmunoCAP® is nants of benzylpenicillin, phenoxymethylpenicillin, described as a capsule enclosing a cellulose ampicillin, amoxicillin, ticarcillin, fl ucloxacillin, derivative with a high binding allergen capacity and cloxacillin) (Chap. 5 ); cephalosporins (cefaclor, per mg of cellulose. The ImmunoCAP® testing cephalothin, cefalexin, ceftazidime, ceftriaxone, agents and format are now backed by a large cefuroxime, cefotaxime, cefadroxil) (Chap. 5 ); number of publications focusing on performance tetracycline and doxycycline; sulfamethoxazole;

[email protected] 104 4 Diagnosis of Allergic Reactions to Drugs trimethoprim; chlorhexidine; quinolones (cipro- Table 4.2 Necessary procedures and criteria to be satis- fl oxacin, levofl oxacin, moxifl oxacin, ofl oxacin, fi ed for the detection of drug-specifi c IgE antibodies pipemidic acid, rufl oxacin) (Chap. 6 ); neuromus- A. Requirements for assay cular blocking drugs (morphine for the detection of 1. Serum from allergic subjects members of the group, alcuronium, gallamine, 2. Drug (or close structural analog) covalently rocuronium, succinylcholine, d -tubocurarine, linked to suitable solid phase vecuronium); thiopentone (Chap. 7 ); local anes- 3. Monospecifi c, affi nity-purifi ed anti-human IgE antibodies tagged with suitable reporter group thetics (mepivacaine, procaine) (Chap. 7 ); opioids (e.g., radioisotope, enzyme, fl uorescent label, (morphine, codeine) (Chap. 8 ); aspirin, propyphen- colloidal metal particles, etc.) azone, and other pyrazolones (Chap. 9 ); ioxaglic 4. Detector—spectrometers, spectrophotometer, etc. acid (Chap. 10); a number of monoclonal antibod- B. Controls ies (Chap. 11); methylprednisolone succinate ester 1. Free solid phase (Chap. 12 ); L-asparaginase (Chap. 13 ). 2. Solid phase covalently linked to Many of these assays essentially remain as (a) Structurally related compounds unstandardized research tools but each one has (b) Structurally unrelated compounds produced results that suggest further investiga- 3. Sera from nonallergic (“normal”) subjects 4. Serum from cord blood (IgE-“free” control) tion, refi nement and steps to achieve validation 5. Sera from subjects allergic to common allergens; are worth considering. A major obstacle in the include sera with high total IgE levels introduction and development of any new test for C. Demonstration of specifi city the detection of drug- reactive IgE antibodies is 1. Inhibition of binding of IgE antibodies in subject’s the availability of a suffi cient supply of reactive serum to drug solid phase by preincubation with sera from allergic patients. This problem is par- free drug ticularly acute when the drug is an infrequent 2. Inhibition with structurally related compounds cause of type I allergy. In cases where the devel- 3. No inhibition with structurally unrelated compounds opment of a new test is considered desirable, 4. Binding of IgE antibodies to solid phase covalently when suffi cient sera from allergic patients is linked to structurally related compounds available and the necessary chemical procedures have been devised, a series of investigative steps to determine specifi city of the assay are obliga- reactions, the classes are often wide, use different tory. These essential controls and procedures are calibrations, and differ between different test sys- set out in Table 4.2 . tems. The traditional performance characteristics of sensitivity and specifi city for tests measuring specifi c IgE antibodies simply divide the results 4.3.3 Quantitation, Interpretation into positive and negative with a cutoff that is and Reporting of Results often diffi cult to interpret clinically. This is most easily seen when low levels of IgE occur with It is obviously desirable to be able to detect IgE unclear or vague clinical symptoms. Receiver- antibodies in very sensitive, specifi c, and accurate operating characteristic (ROC) curves provide assays and to report the results in quantitative more information to aid discrimination between terms. Over the last two or three decades, a vari- positive and negative results but decision thresh- ety of methods for quantifi cation have been devel- olds are not displayed and they are diffi cult to oped. Many laboratories have reported results apply to small samples. The concentration of IgE using a class or scale system ranging from class 0, antibodies assessed by immunoassay is related to no reaction to class one, a low level of specifi c the presence of allergic symptoms indicating that IgE and so on to class six, an extremely high a quantitative measurement of antibodies will level. Although such a semiquantitative scale yield more informative results than a simple posi- helps to interpret, more fi nely discriminate, and tive or negative answer. This relationship between sort the relative strengths of the different clinician-diagnosed positive and negative fi ndings

[email protected] 4.4 Drug Challenge (Provocation) Testing 105 and the quantitative levels of specifi c IgE was analyzed using a logistic regression model. 4.4 Drug Challenge (Provocation) Without a fi xed cutoff, the logistic model showed Testing better agreement between IgE antibody levels and clinical disease than could be obtained with In the diagnosis of drug hypersensitivity reac- the conventional sensitivity and specifi city tions, a drug challenge, or provocation test, is the approach. This quantifi cation demonstrating a controlled step-wise administration of a drug in a link between specifi c IgE antibodies and allergic supervised hospital environment in order to reactions helps to achieve greater diagnostic determine if the drug was the causative agent in a accuracy. patient’s allergic reaction. The challenge test is The best calibration system for specifi c IgE also used to determine if a particular drug can be antibodies currently in use is based on the World safely administered to a patient. In the ideal diag- Health Organization 75/502 IgE standard using a nostic test for a drug-induced hypersensitivity multipoint calibration curve. A number of clini- reaction, challenge with the suspected drug cal IgE assays, including the ImmunoCAP ® reproduces the identical clinical symptoms and (Thermo Scientifi c) assays, have a working range signs of the original or so-called index reaction. from 0.1 to 100 kU A/l, where A represents the For this reason, the test is the best way to confi rm amount of allergen-specifi c IgE antibody in the an allergic reaction, and it is considered to be the sample and 1 kUA /l = 1 KU/l = 2.42 μg/l. Note that “gold standard” in the diagnosis of drug hyper- although different assay systems may present sensitivity reactions. In fact, a positive challenge their results in the same units, viz. kUA /l, this test will not only produce the symptoms of does not guarantee that the results are correct and hypersensitivity but also those of other adverse interchangeable. This is probably because most responses regardless of the mechanism. The ben- allergen preparations contain a mixture of differ- efi t of the challenge test is obvious when one ent individual allergens with different allergenic considers that a positive result makes the need potencies and the compositions of the allergen for allergen avoidance clear and unequivocal preparations used may vary between manufactur- while a negative result gets rid of the incorrect ers. One might predict that the situation with and unnecessary classifi cation of the patient as drugs should be simpler and more predictable hypersensitive to the drug. If a reaction does since the allergenic material employed in the occur upon challenge, it is likely to be milder assay is a pure, standardized drug that can be because of the slow and incremental dose escala- coupled to the solid support using well-defi ned tions. Drug challenge tests are particularly chemical procedures that can be followed by all important when other usually employed tests, manufacturers to produce a standardized test particularly skin tests, are not available or possi- solid phase. ble, for example, with histamine-releasing drugs Special care should be exercised when results such as codeine; where sensitivity is lacking with for drug-specifi c IgE antibodies fall below the non-β-lactam antibiotics; with glucocorticoids limit of quantitation and are undetectable. and heparins; and with drugs such as local anes- Although this may indicate a nonsensitized thetics (see Chap. 7 ), NSAIDs in patients with individual, the fi nding might be due to the pres- the cross-reactive pattern (Chap. 9), and contrast ence of a hypersensitivity state without IgE media (Chap. 10) that may produce unreliable involvement or the blood sample might have results. A challenge test is also a reliable way to been taken too long or too soon after the adverse check on a previous test result such as a skin test reaction. In the former case, antibody levels or serum IgE antibody fi nding. If these tests do may have decreased over time while in the latter not lead to a conclusive result, the challenge test case, drug-reactive antibodies may have been may be the only way to achieve a diagnosis. largely consumed with no time for their Before undergoing challenge, patients should replacement. be presented with a consent form to read and sign

[email protected] 106 4 Diagnosis of Allergic Reactions to Drugs if they agree to go ahead with the procedure. The Table 4.3 Doses of some commonly used drugs form should state the purpose of the test, set out employed in provocation testing the procedures involved, and summarize the ben- Dosage amounts Usual daily efi ts and risks. Some practitioners believe that Drug (mg) and sequencea,b adult dosec (mg) drug challenge tests should be performed at least β-Lactamsd 4 weeks after the drug reaction, but there is no Cefaclor 1, 5, 25, 125, 500 750 e general agreement on this or on any upper time Cefazolin 1, 5, 25, 100, 500, 1, 500–3,000 2,000 limit. Antihistamines should be discontinued at Cefuroxime 1, 5, 20, 80, 400 500 least 5 days before the scheduled appointment Macrolides and the administering clinician should be advised Azithromycin 1, 5, 25, 57, 125, 250 500 if beta-blockers or ACE-inhibitors are being Erythromycin 1, 5, 25, 100, 500, 2,000–3,000 taken or if the patient is pregnant. The patient’s 1,500 health should be good on the day of testing with Quinolones no sign of allergy or virus infection. Checks on Ciprofl oxacin 1, 5, 25, 100, 500 500–1,500 blood pressure, pulse, peak expiratory fl ow, and NSAIDs f,g oxygen saturation levels may be done before, Ibuprofen 1, 5, 20, 80, 150, 300 200–1,200 during, and at the conclusion of the test. Challenge Diclofenac 1, 5, 20, 80 100–150 can be oral, parenteral (subcutaneous, intramus- Piroxicam 1, 3, 6, 10 20 cular, or intravenous), bronchial, nasal, cutane- Acetaminophen 1, 10, 50, 250, 500, 500–4,000 1,000 ous, or conjunctival depending on the specifi c Steroids reaction and drug. For drugs that are ingested or Prednisolone 2, 10, 20, 40 20–80 injected, the oral route is preferred. Provocation Betamethasone 0.2, 1, 2, 4 3–12 is commenced with a small dose of the drug, and Proton pump inhibitors this is gradually increased at 30-min intervals Omeprazole 1, 5, 10, 20 20–40 provided no adverse reaction occurs after the pre- Local anesthetics vious dose. This procedure is continued until the Lidocaine h 2, 20, 40 20–60 desired dose (usually the dose that would be pre- Doses administered orally unless indicated scribed or the total daily dose) is reached. Doses taken from Messaad D et al. Ann Intern Med Table 4.3 lists a range of commonly used drugs, 2004;140:1,001 a For anaphylactic shock patients start with 1/10th of the their usual daily dosages, and increasing chal- dose shown here lenge doses administered during provocation b Doses administered at 30 min intervals testing. For details of oral and parenteral provo- c Recommendations of the French Agency on Drug Safety cation testing for penicillins, see Chap. 5 , ( http://www.AFSSAPS.sante.fr ) d For penicillin provocation testing see Table 5.3 Table 5.3 . If any adverse reaction that looks like e Doses administered IV an allergic response occurs, for example, wheez- f NSAIDs—nonsteroidal anti-infl ammatory drugs ing, swelling of the throat, a drop in blood pres- g For aspirin provocation testing see Sect. 9.5.2.1.3.1 h sure, or rash, the challenges are discontinued and Administered subcutaneously using solution 20 mg/ml the test interpreted as positive, in other words, the patient is judged allergic to the drug. Minor patient is given the drug and a placebo without symptoms like itching and some redness might knowing which is which for each dose or double not be considered a suffi cient reason to curtail blind when both the patient and the clinician are the test but, if this is done, oral antihistamines unaware whether and when the drug or placebo is are usually enough to control the reactions. Severe given. The time taken for drug challenge tests can reactions are treated promptly with epinephrine vary widely depending on the drug, the possible and/or other medications such as steroids, bron- severity of any allergic reaction it might elicit, chodilators, antihistamines, and, if necessary, and the slow or rapid graded nature of dosage. intravenous fl uids. Placebo-controlled drug chal- Challenges may be as short as 2 or 3 h or extended lenges may be either single blind, when the over many hours or even days. After completion

[email protected] 4.5 Detection and Measurement of Released Mediators/Markers of Hypersensitivity 107 of the test, patients who did not develop a reac- benefi ts warrant proceeding, challenge can be tion are kept under observation for 1 h and undertaken in an intensive care unit for safety rea- patients with minor reaction are observed for 2 h. sons. For patients with a history of anaphylactic Overnight stay in hospital should follow a serious shock, intravenous catheters should be in place allergic reaction. for the duration of the test. After discharge, there Some variations in procedures are needed for is the possibility of a delayed allergic reaction as some drugs. With aspirin, (acetylsalicylic acid) part of a biphasic response. This can occasionally for example, the interval between doses is usually be lethal so at the time of discharge the patient longer and of the order of 60–120 min rather should be provided with adequate emergency than 30 min (see Sect. 9.5.2.1.3) and a starting treatment of antihistamines, steroids, or inhalers. dose of 40–60 mg is proposed for patients with aspirin-exacerbated respiratory disease if they are given a leukotriene-modifying drug. In order to 4.5 Detection and Measurement confi rm the absence of allergy (particularly of the of Released Mediators/ delayed type) to some drugs, for example, antibi- Markers of Hypersensitivity otics, patients who tolerated the test are sometimes sent home with a 5–7 day course of the A wide and chemically varied range of biologi- drug. With the β-lactams, 4–9 % of cases con- cally active agents may be released during drug- fi rmed by challenge testing have a delayed reac- induced adverse reactions in humans. Mast cells tion demonstrated by a positive intradermal or and basophils are the main cells activated during a patch test. With noncooperative young children, type I IgE antibody-mediated reaction. Preformed drug challenge tests with β-lactams have proved mediators of allergy including histamine, the to be well tolerated. enzymes tryptase, carboxypeptidase, chymase In general, drug challenge tests should not be and cathepsin G, serotonin, platelet activating fac- performed on pregnant women, on patients with tor, and eosinophil and neutrophil chemotactic uncontrolled asthma, acute infections, and some factors are released from the granules of mast diseases involving the major organs like the heart, cells while potent lipid mediators such as leukot- lung, liver, and kidneys. It should never be per- rienes LTB4 , LTC4 , and LTD4 ; prostaglandin formed on patients who have experienced SJS, PGD2 ; and thromboxanes are newly synthesized. TEN, DRESS, exfoliative dermatitis, vasculitic Cytokines, including IL-4 and IL-13 that stimu- syndromes, and life-threatening immunocyto- late Th2-cell responses, IL-3, IL-5, granulocyte- toxic reactions. macrophage colony-stimulating factor (GM-CSF), Apart from the obvious diagnostic value and tumor necrosis factor (TNF), and chemokines benefi ts for the patient that a drug challenge can such as CCL3 and CCL5 (RANTES) are also bring, it remains a serious and potentially danger- released. Of this large, assorted group of liberated ous procedure to be used with great caution. It mediators promoting such a diverse range of should not be used without careful analysis of the physiological and pharmacological effects, only possible benefi ts for the patient, the particular histamine, the enzyme tryptase, and to a lesser drug hypersensitivity situation, and the patient’s extent the leukotrienes have so far found any diag- state of health. Essentially, provocation with a nostic application to drug allergies. suspected drug can be undertaken when it is judged that the risk of a serious allergic reaction is low while the value and quality of information 4.5.1 Tryptase gained to aid the patient is high. In other words, performance of this test, more than the other tests 4.5.1.1 Tryptase Genes discussed here, comes down to a decision after Although many, if not most, of the autacoids, weighing up the risk-to-benefi t ratio. If for some enzymes, biologically active lipids, and various reason a high risk remains but it is judged that the other factors involved in the infl ammatory cascade

[email protected] 108 4 Diagnosis of Allergic Reactions to Drugs following mast cell activation contribute to the tryptase levels in blood therefore refl ect, and are clinical manifestations of an allergic reaction, a measure of, mast cell activation. there is no evidence that tryptase has such a role. Tryptases are neutral proteases belonging to a 4.5.1.3 Tests for Mature and Total subgroup of the trypsin family serine peptidases. Tryptase Almost all human mast cells make and store The immunoassay result for total tryptase refl ects tryptases which constitute the major proteins in the mast cell number and measures pro and mature the secretory granules. Basophils, which seem to forms of α and β tryptases. Protryptases account have a lesser role in anaphylaxis than mast cells, for nearly all of the tryptase in normal sera which make far smaller amounts of tryptase. Four gene show a normal level of approximately 5 ng/ml and loci on chromosome 16 encode the human mast a range of 1–15 ng/ml. Results of a study of 126 cell tryptases, and additional diversity is added apparently healthy individuals 12–61 years old by α, β, γ, and δ allelic variations. The gene sites with the Phadia ImmunoCAP Tryptase (Thermo and alleles they harbor are termed TPSAB1 (α Scientifi c) commercially available fl uoroimmu- and β1 alleles), TPSB2 (β2 and β3 alleles), noenzymatic assay showed a geometric mean of TPSD1 (δ alleles), and TPSG1 (γ alleles). 3.8 ng/ml and a 95th percentile of 11.4 ng/ml. δ-Tryptase and γ-tryptase are not found in serum; From the pioneering studies of L. B. Schwartz and the biological role of δ-tryptase is not known coworkers, blood samples (to obtain serum after but γ-tryptase is an active peptidase that stimu- clotting) should be taken 15 min to 3 h after the lates IL-13 production and induces bronchial suspected reaction, although signifi cantly raised hyperresponsiveness in mouse airways. Only α tryptase levels can usually be detected up to 6 h and β tryptases make up the circulating tryptase (and sometimes more) after the reaction. Normal and only these two forms show a relationship levels return 12–14 h after the initial release. For to anaphylaxis. Considering only the α and β shipping, serum samples can be kept at room tem- alleles, three genotypes are possible αα:ββ, perature for 2 days otherwise samples are stored αβ:ββ, and ββ:ββ. at 4 °C for 5 days or −70 °C for longer periods. Total tryptase levels greater than 20 ng/ml are 4.5.1.2 α and β Tryptases generally found in patients with systemic masto- Circulating tryptase levels are now known to con- cytosis. For postmortem examination of tryptase sist mainly of inactive β-protryptase with perhaps levels, blood can be taken up to 24 h after death a small amount of α-protryptase. In fact, total although results from at least one investigation tryptase levels in serum are not signifi cantly dif- suggest that blood samples taken after a signifi - ferent in subjects with and without the gene for cantly longer time interval can still be success- α-tryptase. α-Tryptase is not stored in granules fully examined to yield important diagnostic but is instead secreted as an inactive proenzyme. information. In a study of two fatal cases of peri- This means that upon degranulation, α-tryptase is operative anaphylaxis employing immunoassays not part of any increase in serum tryptase levels for neuromuscular blocking drugs together with and is therefore not a useful marker of mast cell the tryptase assay, highly elevated tryptase levels activation and anaphylaxis. Tryptase, stored in (compared to levels in preoperative blood sam- secretory granules and released during activation ples) found in blood taken 48 and 72 h after death of mast cells, unlike protryptases, is not sponta- were matched by the clear-cut detection of IgE neously secreted by resting mast cells and is antibodies to thiopentone and succinylcholine. referred to as mature tryptase. In skin and lung The assay for mature tryptase is a capture mast cells most, if not all, of the mature tryptase assay using a monoclonal antibody that recog- is β-tryptase. Thus, α and β protryptases are nizes mature α and β tryptases although β-tryptase spontaneously secreted by resting mast cells constitutes the major, if not exclusive, form of while mature β-tryptase is stored and released mature tryptase in blood. Normal levels of mature upon degranulation of the mast cell. Mature tryptase in serum are less than 1 ng/ml whereas

[email protected] 4.5 Detection and Measurement of Released Mediators/Markers of Hypersensitivity 109

Table 4.4 Total and mature tryptase levels in serum of nor- ority. Early studies with the assay showed that mal subjects and patients with anaphylaxis or systemic increases in mast cell tryptase concentrations in mastocytosis serum seemed to occur only in immunological Mature Ratio of total Total tryptase tryptase to mature reactions but this proved to be not invariable— Subject level (ng/ml) level (ng/ml) levels increased tryptase concentrations may occur with Normal 1–15 <1 – direct histamine release. A good reported exam- Acute systemic >Baseline >1 <10 ple of this is an extremely high serum tryptase anaphylaxis level in a patient who died after receiving a bolus a Nonacute >20 <1 >20 of vancomycin. In cases of life-threatening drug systemic mastocytosis reactions during anesthesia where neuromuscular Data from Schwartz LB. Immunol Allergy Clin N Am blocking drugs are known to be the biggest cause 2006 ;26:451 of anaphylaxis and where research is more a Sometimes small elevations advanced than in many other areas of drug allergies, the tryptase test has become part of the levels equal to or above 1 ng/ml indicate mast cell established and standard protocol along with skin activation. Three or four hours after the onset of tests and tests for serum IgE antibodies for diag- anaphylaxis, levels generally revert to less than nosing and establishing the mechanism of drug- 1 ng/ml. In anaphylaxis, the ratio of total to induced reactions. It is concluded that increased mature tryptase is typically less than 10 while in mast cell tryptase concentrations are a valuable systemic mastocytosis the ratio is generally more indicator of an anaphylactic reaction during anes- than 20. Mature tryptase levels greater than 10 ng/ thesia and although elevated levels favor an IgE- ml in postmortem samples suggest that systemic antibody- mediated cause, they do not always anaphylaxis might have occurred (Table 4.4 ). The distinguish between an anaphylactic and an ana- mature tryptase assay is only available in the lab- phylactoid reaction. oratory of Dr Lawrence Schwartz, Medical College of Virginia, Richmond, VA, USA. 4.5.2 Histamine 4.5.1.4 Tryptase and Drug Allergy From the time of its general introduction in the The presence and biological role of histamine in early 1990s, the tryptase assay has gone a long immediate and some other hypersensitivity reac- way in helping to answer the question “is this tions is well established. Histamine release from drug reaction anaphylaxis?” From the beginning, human blood leukocytes after challenge with the test proved valuable for selecting an homog- drug in vitro is occasionally employed but in gen- enous population to evaluate tests for dug allergy; eral, for diagnostic purposes, histamine concen- initial screening with the tryptase test sometimes trations in biological fl uids have rarely been eliminating a signifi cant number of patients from routinely measured. The reasons for this in the subsequent skin and IgE antibody testing. Drug- diagnosis of drug allergy are not hard to fi nd: the reactive IgE antibody tests on sera taken at the half-life of histamine in plasma is short (approxi- time of the reaction may identify an immune mately 1–2 min) due to its rapid methylation by basis for the reaction but such tests are not always histamine methyltransferase and oxidation by available and some assays in their current form diamine oxidase; blood sampling after a reaction, may give false negatives to some drugs. At fi rst especially in an emergency situation like anaphy- site, the determination of plasma histamine con- laxis, is diffi cult and has severe time constraints; centrations can be valuable in demonstrating an false positive results are often assumed to be anaphylactoid reaction, but these tests can be likely following disruption of cells, in particular technically and logistically diffi cult with samples basophils, during blood sampling and handling; needing to be obtained preferably within 10 min and assays have commonly had technical and of the reaction, a time when resuscitation is a pri- practical shortcomings.

[email protected] 110 4 Diagnosis of Allergic Reactions to Drugs

4.5.2.1 Histamine Concentrations are challenged with drug in vitro at 37 °C, super- in Blood natants are frozen, and the histamine content is The threshold for pathological levels of hista- measured after cell lysis. Anti-IgE can be added mine in plasma is said to be 1 ng/ml or 9 nmol/l to assess releasibility of histamine from baso- while levels greater than 10 ng/ml (90 nmol/l) phils, and the total histamine concentration of the cause serious cardiovascular sequelae. Plasma cells is measured after cell lysis. The fi gure for histamine levels in normal subjects are generally spontaneous histamine release is subtracted from less than 9 nmol/l when measured by the fl uori- the drug-induced release and total histamine metric method. Isotope dilution mass fragmen- results. Spontaneous release should normally be tography gave a range of 0.8–3.6 nmol/l, and the less than 5 % of the total histamine content of monoclonal anti-acylated histamine-based radio- blood (20–200 ng/ml). Histamine release greater immunoassay employed in a number of allergy- than 5 % of the total histamine content (after sub- related studies has shown concentrations of tracting the spontaneous release fi gure) is 3.33 nmol/l (n = 14), 0.193 (1.7 nmol/l) ± 0.08 ng/ regarded as a positive drug-specifi c result. In an ml ( n = 40) and 0.8 ± 0.4 nmol/l ( n = 13) in normal effort to investigate the potential of measuring subjects and 1.63 ± 0.61 nmol/l (n = 35) in anes- released plasma histamine for the diagnosis of thetized patients. Histamine was measured 15 drug allergies and the possible problems associ- min after induction of the latter patients none of ated with the necessary measurement procedures, whom had any adverse reactions. The normal a radioimmunoassay utilizing a monoclonal antihistamine concentration represents less than body specifi c for succinyl glycinamide deriva- 0.5 % of the histamine concentration in blood. tized histamine was utilized. This work is Histamine in blood and in plasma is generally mentioned since it covers most of the important said to be unstable but precise fi gures on its half- technical points relevant to measuring histamine life under normal and varied conditions are hard in plasma and highlights the all-important ques- to fi nd. Studies on six normal volunteers showed tion of the mediator’s stability. The assay has the half-life of infused histamine to be 102 s. A cross-reactivity ratios for N -methylhistamine and comparison of the metabolism of infused hista- histidine of 1/14,500 and 1/250,000, respectively, mine in urticarial patients, normal subjects and and a claimed sensitivity of 0.2 nmol/l, although atopics revealed half-lives of 6.2 ± 1.3 min, 4 ± 0.7 it proved to be 0.5 nmol/l with a variation coeffi - min, and 3 ± 1.2 min, respectively. cient of 10 % in this study. A range of seemingly small but essential investigations of aspects of the 4.5.2.2 Measurement of Histamine methodology of sample collection, handling, and At least until the late 1980s, assays for histamine storage as well as stability studies were under- were often diffi cult to undertake and lacking sen- taken, sometimes with surprising results. Many sitivity and specifi city. Some methods for mea- of the precautions thought to be necessary, and suring histamine, for example, the automated taken, when using the fl uorimetric assay were fl uorimetric assay developed by Siraganian, show found to be unnecessary for the radioimmunoas- good sensitivity and specifi city, but the method is say. This was demonstrated in experiments on the somewhat complicated and technically demand- sampling procedure, leakage of histamine from ing and has therefore not been employed in a basophils at room temperature and 4 °C, and large number of laboratories worldwide. freeze-thawing of samples. Of most interest, were Immunoassays for histamine in RIA or ELISA the fi ndings on the stability of histamine in blood format are available commercially from a number and serum samples. The in vivo half-life of of different companies. They are more accessible infused histamine in plasma has been estimated and easy to use than many of the older assays and to be between 1 and 2 min but in 12 patients with some with good performance characteristics have increased plasma histamine concentrations dur- been adopted as a diagnostic tool. For drug ing anaphylaxis, all had increased plasma hista- allergy studies, aliquots of diluted whole blood mine concentrations in the fi rst sample taken

[email protected] 4.5 Detection and Measurement of Released Mediators/Markers of Hypersensitivity 111

5–60 min after the onset of the reaction. The allergy (55 ng/ml). Overall, the sensitivity mean histamine concentration in the second sam- (51.4 %), the specifi city (63 %), and the positive ple taken between 30 and 200 min after the fi rst predictive value (29.3 %) were poor, but the neg- sample was signifi cantly less but also still ele- ative predictive value (81.1 %) was judged to be vated. All but one of the 12 patients still had of value for ruling out some reactions that increased plasma histamine concentrations at appeared to be allergic. On the basis of these pathological levels 60 min after the onset of the results, one could not advocate histamine release reaction. In the anaphylactic patients, these as a routine test for the diagnosis of drug allergies results point to a half-life of histamine signifi - but it seems premature to accept this conclusion cantly higher than 1–2 min—in fact, a fi gure as fi nal and generally applicable to drug allergy closer to 20 min is likely. This prolonged half-life for a number of reasons. More extensive studies may be a refl ection of the extended release of his- are needed with homogenous populations of tamine or saturation of the enzymes involved in patients particularly with regard to the type of its metabolism. In vitro, histamine concentrations hypersensitivity, the drugs involved, and the time did not change signifi cantly in whole blood left at interval between the reaction and testing. Patients room temperature for 2 h or overnight at 4 °C or with any one of the four commonly recognized in separated plasma after 48 h at room tempera- types of hypersensitivity may be classifi ed as ture or 72 h at 4 °C. In previous in vitro studies, drug allergic and testing may compare time inter- histamine has been reported to be stable for 30 vals of as little as a month to a number of years. min at room temperature and at 4 °C when added There are also indications that histamine release to normal plasma. The stability of histamine in tests might be of more value for some classes of patients with normal metabolism was in contrast drugs. With neuromuscular blocking drugs, diag- to results obtained with plasma from heparinized nostic applications of histamine release tests have patients and from pregnant women, the latter yielded encouraging results. One study, for apparently due to increased diamine oxidase example, involving 40 patients allergic to muscle activity during pregnancy. Of interest was the relaxants and 44 controls, showed a sensitivity of fi nding that histamine disappeared from plasma 65 % and specifi city of 100 %. Some encourag- at a slower than expected rate during anaphylaxis ing results have also been obtained with indicating that there is still likely to be time for β-lactams, especially in the case of rapid onset blood sampling after emergency treatment. This reactions. It, therefore, seems too early to leave has led some investigators to conclude that hista- histamine release tests out of any serious consid- mine levels in plasma should be determined in eration of drug allergy diagnosis. Too few well- cases of drug-induced anaphylaxis and to go so designed and executed investigations have been far as to state that there is no justifi cation, apart undertaken so far and it seems true to say that from cost, for not doing so. some of the expected diffi culties of working with histamine are more assumed than real. 4.5.2.3 Histamine in Drug Allergy Diagnosis It is safe to say that the above belief is not widely 4.5.3 Cysteinyl Leukotrienes held and the conclusion not widely supported. An investigation of the predictive capacity of hista- For the structures of the cysteinyl leukotrienes mine release for the diagnosis of drug allergy and a discussion of their biosynthesis and role as using the sensitive anti-acylated histamine immu- infl ammatory mediators, see Sect. 3.2.5.2 . noassay found that net histamine release was The cysteinyl leukotrienes LTC4 , LTD 4 , and positive in only 18 of 35 drug-allergic patients LTE4 , sometimes called sulfi doleukotrienes, are a (median total histamine 61 ng/ml), 12 of 33 aller- family of potent bioactive peptide-conjugated gic patients with no drug allergies (50 ng/ml), lipids formed by mast cells, basophils, eosino- and 15 of 40 controls with no history of drug phils, neutrophils, macrophages, dendritic cells,

[email protected] 112 4 Diagnosis of Allergic Reactions to Drugs and T lymphocytes. Basophils produce more than cysteinyl leukotriene release induced by aller- 100 times the amount produced by eosinophils. gens although some investigators believe that Following allergen-induced cross-linking of IgE IL-3 may stimulate basophils causing false posi- antibody receptors on mast cells, cysteinyl leu- tives, high backgrounds, and nonspecifi c reac- kotrienes are released newly synthesized within tions. In testing for drug hypersensitivity, the minutes. Well recognized for their powerful addition of IL-3 is said by some to be essential to bronchoconstricting effects and exacerbating obtain sensitivity. C5a , a nonspecifi c activator of asthma, these lipid mediators are found in bron- basophils and thought to enhance allergen stimu- choalveolar lavage fl uid and nasal secretions lation, has been added in the CAST® assay but its from atopic human subjects after allergen expo- use seems to be not widely supported. In the sure. As an indication of potency, LTE 4 , for absence of fresh cells from patients, successful example, is considerably more powerful than his- passive sensitization with serum would be an tamine in decreasing airfl ow. important added feature of the CAST® assay. However, investigations so far have shown that it 4.5.3.1 Tests for Allergen-Induced is less sensitive than when untreated fresh cells Release of Cysteinyl are used. In CAST®-ELISA assays, up to 6–8 % Leukotrienes of cells have been reported to be nonresponders. The release of cysteinyl leukotrienes from iso- This has been said to be due to reconstitution of lated peripheral blood leukocytes following aller- the lyophilized anti-IgE reagent in solutions with gen challenge in vitro has been utilized as a test a suboptimal Ca2+ concentration. for immediate hypersensitivity in the form of the The determination of cysteinyl leukotrienes Bühlmann CAST® (Cellular Allergy Stimulation by ELISA in cell supernatants has been com- Test) assays offered commercially as CAST® bined with fl ow cytometric examination (see ELISA, a microtiter plate ELISA immunoassay, Sect. 4.6 ) of CD63 expression on basophils in the and as a fl ow cytometric assay Flow CAST ® Flow CAST® assay. Gating of basophils is (Bühlmann Laboratories AG, Schönenbuch, achieved with the aid of the constitutively Switzerland). Before the CAST ® ELISA test is expressed eotaxin receptor CCR3 detected with a carried out, patients should discontinue treatment phycoerythrin fl uorescence-labeled monoclonal with antihistamines, corticosteroids, or chromo- antibody. CD63, detected with a fl uorescein iso- glycic acid 3–7 days prior to sample collection. thiocyanate (FITC) fl uorescence-labeled mono- Blood is collected in the presence of EDTA and clonal antibody, is utilized as the cell surface stored at 2–8 °C for up to 24 h if necessary before activation marker expressed on sensitized degran- 200 μl is taken for leukocyte isolation by dextran ulating basophils. Positive controls include a sedimentation of red cells. After resuspension of monoclonal antibody to cross-link the FcεRIα leukocytes, aliquots in stimulation buffer are receptor and the tripeptide chemoattractant taken for challenge with allergen and to serve as N-formyl-methionine-leucine-phenylalanine positive control (antibody to high affi nity IgE (fMLP) that activates basophils nonimmunologi- receptor FcεRIα) and background spontaneous cally. The basophil surface expressed antigen release tubes. After incubation for 40 min at CD203c detected with a different fl uorescence- 37 °C and centrifugation, cell-free supernatants labeled antibody can also be used as an extra acti- along with standards (range 50–3,200 pg/ml) and vation marker. Both markers are used in the Flow controls are added to pre-coated microtiter wells CAST® highsens commercially available kit. for the determination of de novo synthesized leu- There are reports of 8–10 % of nonresponders in ® kotrienes (LTC4 , LTD 4 , and LTE 4 ) in a competi- some Flow-CAST studies. tive immunoassay at room temperature. The When considering the cellular antigen stimu- ELISA utilizes alkaline phosphatase for color lation and basophil activation tests, an interesting development. The whole procedure is said to point to bear in mind is the sensitivity of CD63 take5½ h and an analytical sensitivity of 19 pg/ml expression to external Ca2+ concentrations is claimed. Addition of IL-3 is said to increase compared to the calcium requirement for LTC 4

[email protected] 4.6 Basophil Activation Test 113 production. In addition, calcium sensitivity may 34 cases. Sensitivities (percents) for the CAST® vary from one individual to another. The conse- and histamine tests, respectively, were: for quence of this can be a positive CAST® result and β-lactams 43 and 53; for aspirin 21 and 53; for a negative basophil activation test (BAT) result. acetaminophen 33 and 33. Corresponding specifi cities (percents) were 79 and 55; 88 and 35; 100 4.5.3.2 Correlations of CAST® with and 43. The effi ciency of both tests was low and the Clinical Situation and the conclusion reached was that the test for Other Diagnostic Tests released cysteinyl leukotrienes showed little or These are not always clear-cut and uniformly no diagnostic utility and was not any better than good. Good correlation has been reported with the histamine release assay when applied to the the severity of allergic rhinitis reactions but not three drugs. Results from a European multicenter for asthma severity. For β-lactam drugs, CAST® study that evaluated the effi ciencies of skin tests, was shown to be positive only in cases of anaphy- serum IgE antibody tests, and CAST® assays in laxis but another group reported positive fi ndings the diagnosis of β-lactam allergy have been used in cases of β-lactam induced generalized urti- to claim that the CAST® assays “are a decisive caria. When CAST® and histamine release assays and powerful in vitro tool for clear diagnosis of were applied to the investigation of hypersensitiv- β-lactam allergies ahead of sIgE.” Sensitivities ity to NSAIDs, histamine release assays were said and specifi cities of the three different tests were to be less sensitive and correlations poor. With (percentages): Skin tests, 70 and 100; serum IgE regard to skin tests, positive CAST® results have antibodies, 30 and 86; CAST ® ELISA/Flow been detected in some skin test negative cases and CAST ® , 41 and 86. Attention is drawn here to the where no skin tests exist, for example, hypersen- relatively low fi gures for IgE antibody tests, sitivity to NSAIDs and non-IgE-mediated drug especially the low sensitivity result. It can be allergies, CAST® has been advanced as a valuable argued that the currently employed assays for in vitro alternative approach to diagnosis. individual penicillins and cephalosporins are When CAST® results are compared to allergen- inadequate to detect the range of allergenic speci- specifi c serum IgE antibody results, correlations fi cities found in the sera of β-lactam-allergic are found but they are generally not high, for patients. This is especially true for the minor example, for protein allergens involved in inhal- determinants of the penicillins and R1 and R2 ant allergies. However, with allergy to β-lactam side chain determinants of cephalosporins. This antibiotics, the determination of drug-specifi c IgE subject is discussed in detail in Chap. 5 . is claimed to be less sensitive than the CAST®. In summing up, it seems true to say that over- In one early study on 25 patients with hyper- all, the effi ciency of the CAST® tests still falls sensitivity to NSAIDs, the CAST® assay was short for the confi dent diagnosis of drug allergies. positive in 5 of 8 aspirin-intolerant and 8 of 12 Further application of the tests to drugs apart from diclofenac-intolerant patients. Sensitivity of the the β-lactams and NSAIDs will reveal whether or test was in the range 62.5–80 % and specifi city not the tests become part of the standard diagnos- 70–100 % leading the investigators to conclude tic protocol for investigating drug allergies. that the CAST® assay might be a useful in vitro test to reliably and safely screen for hypersensitive reactions to NSAIDs. A comparative evalua- 4.6 Basophil Activation Test tion of the CAST® and histamine release assays on 55 patients with immediate reactions to drugs 4.6.1 Basophils and Background (30 to β-lactams, 19 to aspirin, and 6 to acetamin- to the Basophil Activation ophen) and 64 nonallergic but drug-exposed con- Test (BAT) trols showed that the CAST® gave slightly better results with only 19 of 55 (13 for β-lactams, 4 for Basophils are granulocytes that develop from aspirin, 2 for acetaminophen) and 9 of 64 positive CD34+ progenitor stem cells in the bone marrow. while histamine release exceeded 5 % in 28 and They comprise less than 1 % of nucleated blood

[email protected] 114 4 Diagnosis of Allergic Reactions to Drugs cells in humans and remain one of the least involves employment of a fl uorescence-labeled understood leukocytes. Together with mast cells, monoclonal antibody to the eotaxin receptor they express the high affi nity immunoglobulin E CCR3, cysteine–cysteine chemokine receptor 3, receptor FcεRI and have cytoplasmic granules a receptor closely associated with asthma and containing preformed histamine. Basophils also allergy. CCR3 also occurs on eosinophils but express a variety of other receptors including they can be distinguished from basophils by the those for complement, interleukins, chemokines, former’s increased side scatter. Basophils can and prostaglandins. Intermittent efforts to utilize also be identifi ed as anti-IgE and CD203c (see the basophil for the development of an in vitro below) positive cells. An important advancement allergy diagnostic test have occurred over many was the observation that basophil degranulation years. Approaches have usually centered on chal- was accompanied by the upregulation of lenge with allergen followed by the attempted lysosomal- associated membrane glycoprotein-3 measurement of basophil mediators such as his- (LAMP-3; also known as granulophysin), tamine and cysteinyl leukotrienes or direct micro- belonging to the tetraspanin (TM4SF) family. scopic observation of basophil degranulation. For Now generally known as CD63, this protein is a number of reasons including the small number expressed on the surface of degranulated baso- of basophils in blood, diffi culties involved in phils and is the best-validated activation marker handling them, and the laborious and time- used to quantify basophil activation. When FcεRI consuming nature of the required microscopic receptors on basophils are indirectly cross-linked counting, these approaches never became widely during allergen interaction with receptor-bound accepted and used. The development of fl ow IgE molecules, mediators of hypersensitivity cytometry offered the prospect of working with including histamine are released and activation larger numbers of cells but, initially, advances markers such as CD63 are expressed on the cell were limited by the inability to specifi cally iden- surface. In resting basophils of both normal and tify basophils among the whole leukocyte popu- allergic subjects, CD63 is located in the intracel- lation. With improvements in fl ow cytometric lular granules with little surface expression but technology, the utilization of reliable membrane upon upregulation during exocytosis involving markers for basophil identifi cation and activation fusion between granules and membrane, CD63 is and the availability of monoclonal antibodies expressed on the membrane surface in high specifi c for the range of important cell receptors, density. Expression of CD63 on basophils has the monitoring of basophil activation upon produced convincing and specifi c results with allergen challenge became a standardized tool for some common inhalant and venom allergens but in vitro diagnosis of immediate hypersensitivity with respect to drugs, some early studies reported and for recognition of some pseudo-allergies sensitivities of only 50–64 %, that is, not suffi - without IgE involvement. cient to be clinically useful. It was suggested that a contributing factor to this poor sensitivity may be the expression of CD63 on other activated leu- 4.6.2 Basophil Activation Markers kocytes, including platelets, and the subsequent adhesion of these other cells to basophils. A more The identifi cation of basophil cells was initially specifi c and sensitive activation marker therefore based on the presence of the high affi nity recep- seemed desirable. In 1999 the monoclonal anti- tor for immunoglobulin E and expression of body 97A6 defi ned a novel surface antigen CD45 or protein tyrosine phosphatase receptor belonging to the type II transmembrane protein type C (PTPRC) also known as leukocyte com- family on human basophils. The antigen, ecto- mon antigen, a PTP signaling molecule found on nucleotide pyrophosphatase phosphodiesterase 3 all leukocytes. To distinguish basophils from (E-NPPS3), referred to as CD203c, catalyzes the other leukocyte populations, a number of selec- cleavage of oligonucleotides, nucleoside phos- tion strategies are possible. One procedure phates, and NAD. CD203c is constitutively

[email protected] 4.6 Basophil Activation Test 115 expressed on basophils and has the desirable fea- used basophil activation monitored by expression ture of being expressed apparently on that cell of CD63 is a validated test while the more alone. After stimulation with allergen, CD203c is recently introduced marker requires more exten- rapidly upregulated making it a valuable marker sive study and validation for different clinical for basophil activation and hence allergy diagno- conditions. Some have claimed that CD203c pro- sis. Interestingly, it was thought that the release duces slightly improved sensitivity if not by itself of histamine is not directly associated with then together with CD63. The use of both mark- expression of CD63 and CD203c but recently, ers has been advocated, and the practice of using CD63 expression has been shown to result from dual markers now seems to be common. More only the anaphylactic degranulation form of his- recently identifi ed basophil activation markers tamine release. like CD13, CD107a, and CD164 may be the fore- In a comparison of the performances of CD63 runners of a second generation of BATs. and CD203c in the diagnosis of latex allergy, the Recently, it has been shown that phosphoryla- sensitivities of the two markers were 50 and tion of p38 MAPK accompanies upregulation of 75 %, respectively. Following allergen challenge, CD63 expression offering the prospect that mea- levels of expressed CD203c were increased up to surement of phosphorylation of these mitogen- 350 % above control values whereas the increase activated protein kinases might be another way of for CD63 was less than 100 %. Expressed as a measuring basophil activation when applying the percentage of basophils that were marker- test to the diagnosis of allergy. positive, the result for CD203c was 48 % and for CD63 below 20 %. This led to a clear distinction between resting and activated basophils. Another 4.6.3 Some Technical Aspects stated advantage of CD203c was said to be a three- to eightfold higher fl uorescence signal Ideally, cells should be used in the test within 3 h than CD63 but others have reported the opposite of blood sampling. Heparin, EDTA, or acid- fi nding. Occasional weak spontaneous expres- citrate-dextrose can be used to prevent clotting sion of CD203c on resting basophils can make and blood in the latter two media but not in hepa- cell identifi cation diffi cult but rapid expression rin can be stored for 24 h at 4 °C if necessary. At of the marker following allergen challenge may room temperature, desorption of IgE is thought to allow for single color testing without additional occur. Use of whole, heparinized blood is often staining. A small number of comparative studies preferred since it is simple and practical, involv- have revealed some other clear differences in the ing fewer preparative steps while at the same time activation of CD63 and CD203c. Upregulation preserving the basophils’ natural environment. of CD203c seems to occur in all or most baso- Isolated (or more accurately enriched) leukocytes phils while upregulation of CD63 produces one can be prepared as a buffy coat fraction (centrifu- population of basophils expressing the marker gation at 500 g for 10 min) or as a leukocyte frac- with high intensity and another population with tion prepared on a simple density gradient. lower CD63 expression. Expression of CD203c Isolated leukocytes are more diffi cult to standard- is infl uenced by some differences in enzymic ize than whole blood samples, and the conditions regulation, activation by prostaglandin D2 or of their isolation may affect their reactivity for IL-3 are different and CD203c is more easily example, when used with NSAIDs. Some reports activated nonspecifi cally by, for example, han- show that isolated leukocytes are less sensitive to dling of blood and experimental manipulations anti-IgE and allergen-induced CD63 activation, as well as clinical conditions such as atopic der- and although this certainly appears to be the case matitis and food allergy. for most drugs, some results with the neuromus- Overall, the claimed superior performance of cular blocker rocuronium revealed a sensitivity of CD203c has been questioned in more than one 92 %. For drug-induced IgE antibody-mediated study with comments that the presently widely activation of basophils, parenteral preparations of

[email protected] 116 4 Diagnosis of Allergic Reactions to Drugs drugs are preferred for challenge. Prewarming of can be used as an alternative and more sensitive blood/cells and reagents at 37 °C is recom- positive control. This increases activation and the mended. Activation usually begins within 3 min number of responders. A proportion of individuals and reaches a peak within 15–20 min, but this (about 5–10 %) have cells that are so-called non- may vary with the marker. Expression of CD203c responders, that is, upregulation of activation peaks within a few minutes and begins to decline markers does not follow IgE cross-linkage or after 15–20 min. CD203c and CD63 both disap- FcεRI activation. Apart from nonresponders, false pear after 4–5 h. IL-3 has been reported to negatives may arise because of technical manipu- increase CD63 expression and thus sensitivity. lation problems, poor storage, or too long an inter- Since drugs are sometimes poor stimulators of val between the patient’s allergic experience and expression, addition of IL-3 has been suggested testing (recommended interval 6–12 months). To but the enhancing effect of IL-3 does not seem to be able to interpret the BAT, both negative and be consistent. positive controls need to conform to normal, In theory, utilization of passively sensitization expected patterns—high backgrounds and false basophils should be possible in basophil activa- negative responders preclude normal interpretation analyses. This involves stripping of bound tion and any meaningful fi ndings. IgE antibodies from their receptors on the surface of basophils with the aid of acidic buffers (often a lactic acid buffer, pH 3.9), incubation of these 4.6.5 Evaluation of Results stripped cells with patient’s serum (containing IgE antibodies to the allergen being investigated) In evaluating results of BATs, one needs to take for 1 h at 37 °C and then challenging the pas- account of the absolute number of basophils eval- sively sensitized basophils with the allergen at uated (more than 150 is desirable) and the per- the fi rst stage of the BAT. Donor cells for sensi- centage of activated basophils. Due to the tizing should be from a healthy subject whose differences in the upregulation of CD63 and basophils are known to be good responders. Both CD203c (see above), results for CD63 expression unstripped and stripped donor basophils should are generally expressed as a percentage of CD63 be included in the controls. This procedure, apart basophils whereas results for CD203c tend to be from being laborious with a number of extra expressed as stimulation indices (SI) of the mean steps, carries the risk of nonspecifi c stimulation fl uorescence intensity. Because of the usually or damage to basophils and is diffi cult to stan- small number of patients, arbitrary cutoff points dardize. Results so far indicate rather poor sensi- are usually selected as thresholds for positivity. tivity, but some researchers who use the method Cutoffs for some drugs that provide the highest claim good results. sensitivity and specifi city determined by ROC curves are: β-Lactams > 5 % with a SI > 2; aspirin and NSAIDs > 5 % and SI > 2; metamizol > 5 % 4.6.4 Controls and SI > 5. However, for drugs, where smaller numbers of basophils are usually activated, the Incubation of cells with the stimulation buffer pro- calculation of drug-specifi c thresholds is an abso- vides the all-important negative control, that is, lute requirement and this demands the study of the spontaneous expression of the activation large groups of well-defi ned patients. For the marker. In general, negative controls remain below determination of correct sensitivity, specifi city, 5 % in 80 % of cases. For the positive control, anti- and predictive values, the adoption of optimal IgE, either as a monoclonal or polyclonal antidrug-specifi c positivity thresholds to replace body, is employed although the latter is generally arbitrarily chosen decision thresholds is advo- superior since monoclonal anti-IgE antibodies are cated. In establishing the cutoff points, ROC often poor activators of basophils. A monoclonal curves are necessary to establish optimal sensi- antibody to the high affi nity IgE receptor FcεRI tivity and specifi city.

[email protected] 4.6 Basophil Activation Test 117

4.6.6 Application to Drug Allergies degranulation. It has been proposed that expression of the basophil activation marker CD63 is BAT is still mainly employed in research settings, associated with the anaphylactic degranulation but clinical applications are steadily growing. In form of histamine release whereas CD203c cases where the skin test is negative and alterna- expression is known not to refl ect histamine tive tests are either not applicable (e.g., a serum release. Recently, in an attempt to develop a fl ow IgE test for the solvent Cremophor) or unavail- cytometric technique to analyze histamine and its able, and/or when the clinician is faced with reli- release at the single cell level, D. G. Ebo and col- ance on a potentially dangerous provocation test, leagues in Antwerp studied the expression of the the BAT is increasingly being used in the diagno- activation markers CD63 and CD203c and sis of drug hypersensitivities and to assess safe employed the histaminase, diamine oxidase (with alternative treatment regimens. Given the techni- a fl uorochrome label), after application of aller- cal improvements and advances made with the gen, anti-IgE, fMLP, phorbol 12-myristate test, the increasing acquisition of fl ow cytometric 13-acetate (PMA), ionomycin, and IL-3 to obtain quantifi cation equipment by laboratories, the different degranulation profi les. Nineteen birch test’s validation for a number of drugs, and, in pollen-allergic patients, fi ve healthy controls, and some cases, the potential of the test to detect non- the recombinant birch pollen allergen Bet v 1 IgE- mediated basophil activation, it can readily were used in the study. Upon stimulation with be appreciated why the method continues to be allergen, anti-IgE, or fMLP, basophils that upreg- applied to an increasingly wide range of drugs ulated CD203c generally exhibited a bimodal dis- and other therapeutic agents. This list includes tribution of CD63 expression, but individual cell the β-lactam antibiotics penicillins, cephalospo- responses of CD63 expression proved heteroge- rins and clavulanic acid, quinolones, neuromus- neous with the detection of low and high CD63 cular blocking drugs, NSAIDs, radiocontrast expressing basophilic subpopulations following media, chlorhexidine, omeprazole, methylpred- IgE-mediated stimulation. When activation mark- nisolone, valacyclovir, some plasma expanders, ers and histamine release at the single basophil starch colloids, carboxymethylcellulose, some level were analyzed, like CD63 expression, heparins, patent blue, platinum salts, hyaluroni- diamine oxidase labeling was found to be bimodal. dase, and recombinant hepatitis B vaccine. CD203c (bright) CD63 (bright) cells demon- For discussions of the application of BAT to strated clear histamine release while CD203c individual drugs or groups of drugs, the reader is (bright) CD63 (dim) showed less histamine referred to the following sections: Penicillins, release. Diamine oxidase labeling of Bet v Sect. 5.1.6.3 ; clavulanic acid, Sect. 5.5 ; quino- 1-stimulated cells that did not degranulate was lones, Sect. 6.2.3.6 ; neuromuscular blocking more intense than the labeling seen with cells drugs, Sect. 7.4.3.5; hydroxyethyl starch, Sect. exposed only to buffer. Expression of CD63 7.6.1 ; gelatin, Sect. 7.6.2 ; NSAIDs, Sects. following PMA treatment was poor, supporting an 9.5.2.1.3 , 9.5.2.2.3 , and Sect. 9.5.4 ; contrast earlier conclusion that phorbol esters do not induce media, Sect. 10.5.2.3 . signifi cant anaphylactic degranulation but this did occur following addition of ionomycin to PMA. IL-3 produced an increase of diamine oxidase 4.6.7 Analysis by Flow Cytometry labeling of primed basophils from some patients. of Intracellular Histamine From the study, one can conclude overall that and Its Release by Activated fl ow cytometry can be used to examine histamine Basophils at the Single Cell and its release along with the simultaneous quan- Level tifi cation of basophil activation markers. This methodology has been termed “HistaFlow” by The release of histamine is thought to initially the authors. Results demonstrated that the appear- proceed by piecemeal degranulation before con- ance of CD63 indicates anaphylactic degranula- version to what has been termed anaphylactic tion and signifi cant histamine release whereas

[email protected] 118 4 Diagnosis of Allergic Reactions to Drugs expression of CD203c seems to be associated but research progress on extra- and intracellular with fusion of small vesicles, piecemeal degranu- activation markers and a better understanding of lation, and slow liberation of histamine from acti- relationships between the expression of these vated cells. However, upregulation of CD203c markers and the release of mediators promises to does not per se indicate histamine release, and signifi cantly improve diagnostic performance. In release can occur without expression of CD63, particular, application of the HistaFlow method- that is, the expression of both markers can disso- ology may lead to an understanding of the mech- ciate from histamine release. anisms underlying intracellular signaling and The potential of HistaFlow for application to drug- and other agent-induced degranulation of both research and the clinic is apparent. In the basophils. An area where BAT will continue to be former case, the methodology promises to con- applied is in the examination of allergens, both tribute to our understanding of intracellular sig- pure and in crude form, allergoids, vaccines, naling and degranulation of basophils while fl ow newly introduced drugs, additives in many forms cytometry’s and BAT’s already signifi cant contri- of medication, recombinant preparations, and butions to allergy diagnosis are likely to be fur- drugs and agents where other tests are either not ther advanced. In relation to drug allergy, this is suitable or not available. Passive sensitization of already being demonstrated. Figure 4.9 summa- basophils and the subsequent use of the sensi- rizes fi ndings when fl ow cytometry and the tized cells in BAT examinations is one area where expression of basophil activation markers were greater research effort seems necessary. Being utilized for the analysis of histamine release by able to routinely employ serum samples, some- individual basophils from a patient who experi- times taken years before, together with cells enced profound hypotension and severe broncho- known to be good responders and consistently spasm almost immediately after intravenous obtain reliable results would be a signifi cant administration of the cephalosporin cephazolin. advance. Finally, as more knowledge is accumu- Results in this study clearly demonstrated drug- lated of basophil markers and other routes of acti- specifi c IgE antibody-mediated activation of the vation, BAT might prove to be an appropriate and patient’s basophils together with the visually valuable procedure for the study and diagnosis of clear-cut demonstration of histamine release by some IgE-independent drug reactions. the basophils.

4.7 Tests for Delayed Type Drug 4.6.8 Future Research Hypersensitivity Reactions and Conclusions The mechanisms underlying most delayed or Although the BAT remains largely research non-immediate hypersensitivity drug reactions based, awareness of its relevance to the clinic is appear to be heterogeneous and not yet fully growing, and this is refl ected in the allergy litera- understood. The involvement of T cells in delayed ture where diagnostic applications are steadily type drug hypersensitivity reactions is well estab- increasing. An attraction of BAT comes from its lished with different subsets of cells implicated. obvious features that form part of the in vivo Type IV cell-mediated mechanisms seem to be response to allergenic challenge and the resultant involved in reactions such as maculopapular release of mediators. Basophils do not need to be rashes and a number of other skin manifestations separated or purifi ed in potentially cell-damaging including bullous and pustular exanthemas and manipulations and can be examined in their nor- eczema and for the fi rst three of these conditions mal milieu together with other cells to achieve T cells have been implicated. For the diagnosis of reliable results. At present, however, the proce- delayed reactions to drugs, predominately cuta- dures shortcomings, particularly its sensitivity neous reactions, delayed reading of intradermal and diagnostic accuracy, need to be kept in mind testing and patch testing are sometimes used, but

[email protected] Fig. 4.9 Analysis by fl ow cytometry of intracellular hista- blue ), and CD203cbright+ CD63bright+ (gate R5, red ). Gate R0 mine and its release by activated basophils at the single cell denotes non-activated CD203dim+ basophils (b ). Histamine- level (Histafl ow). Upregulation of basophil activation containing basophils are defi ned as DAO+ cells (c –f ). markers CD63 and CD203c combined with demonstration Stimulation with buffer only (negative control) is shown in of intracellular histamine after 20-min activation with cep- (c ). With the positive control stimulated with anti-IgE ( d ), hazolin sodium (Cefacidal® ) in a patient with profound 24 % of the basophils showed histamine release (DAO− hypotension and severe bronchospasm almost immediately cells—see lower-right quadrant). Upon stimulation with after IV administration of cephazolin. Basophils are char- cephazolin (e ), 30 % of the basophils demonstrated hista- acterized using side scatter (SSC) and CD203cdim+ (purple mine release, (DAO− cells – see lower right quadrant). ( f). population in R2) ( a). Activated basophils are defi ned as Shows the DAO histogram (y -axis in (e )) (Kindly provided anti-IgE and upregulated CD203cbright+ positive cells. by DG Ebo, University Hospital Antwerp. See Ebo DG Co-expression of CD203cbright+ and CD63 reveals three sub- et al. Analyzing histamine release by fl ow cytometry populations of distinct CD63 expression; i.e., CD203cbright+ (HistaFlow): a novel instrument to study the degranulation CD63− (gate R3, green ), CD203cbright+ CD63dim+ (gate R4, patterns of basophils. J Immunol Methods. 2012;375:30)

[email protected] 120 4 Diagnosis of Allergic Reactions to Drugs these skin tests often give negative results even in therapy may interfere with the test; and the test patients with a clear history of delayed hypersen- itself is cumbersome and technically demanding. sitivity. The lymphocyte transformation test is Perhaps most importantly of all, a positive lym- often claimed to be the only test to detect drug- phocyte transformation test does not necessarily sensitized T cells. It can sometimes identify the refl ect the exclusive involvement of T cells. For culprit drug and it begins our list of tests for example, B cells present in PBMC may also pro- delayed drug reactions. liferate in response to drug challenge. Nevertheless, the test has some features in its favor. These include: The procedures are carried 4.7.1 Lymphocyte out in vitro so the test is not harmful to the patient Transformation Test nor is there a risk of the patient developing additional drug allergies; new test reagents are not The lymphocyte transformation test is an in vitro needed for each of the different drugs tested; procedure that measures the antigen-induced pro- simultaneous assessments of T cell responses to liferation of drug-specifi c T cells. Briefl y, periph- multiple drugs can be undertaken; positive reac- eral blood mononuclear cells are isolated with a tions can be detected to drugs with different Ficoll gradient and cultured with the addition of pathomechanisms; and the test is claimed to be autologous plasma in triplicate for 5–7 days in more sensitive than other tests for drug hypersen- the presence of different concentrations of the sitivities. The lymphocyte transfer test measures drug being investigated. As positive and negative a memory T cell response and while it can remain controls, cells in triplicate are incubated with and positive for as long as 10–20 years after a drug without phytohemagglutinin 5 μg/ml respec- reaction, other patients are found to test negative tively. Twenty-four hours before harvesting, only 5–8 weeks after onset of a reaction. Ongoing 3 H-thymidine 1 μC is added. Proliferation of cells investigations show that the lymphocyte transfor- refl ected in the incorporation of radioactive mation test is a promising method to identify a nucleoside into DNA is measured in a liquid causative drug in cases of drug eruptions, but it is scintillation counter and the results expressed as crucial to perform the test at the right time and a stimulation index SI where SI equals the counts that time depends on the type of drug reaction. per minute (cpm) with drug divided by cpm Findings with maculopapular drug eruptions, without drug. Some investigators on the basis SJS, TEN, and DRESS illustrate this. For DRESS, of their experience with the method regard a SI patients should be tested 5–8 weeks after the of 1.8 as positive while a cutoff of 2 is also onset of reactions while for the other three condi- often used with a value between two and three tions, testing should take place within 1 week of regarded as a weak positive. Instead of using a skin rashes. The lymphocyte transformation test radiolabeled nucleoside for incorporation, fl uoro- is claimed to have a sensitivity of 60–70 %. chrome-labeled derivatives of deoxyuridine Specifi city is said to be 100 % for carbamazepine (with or without nucleoside-specifi c antibodies) and lamotrigine hypersensitivities and 93 % for are sometimes used. β-lactam hypersensitivity giving an overall speci- There are a number of practical issues that act fi city of at least 85 %. It will be interesting to see against the routine and widespread application of if these levels of sensitivity are achieved and the the lymphocyte activation test for the diagnosis high fi gures for specifi city are maintained as of drug hypersensitivities. In the fi rst instance, more drugs are examined with the test. The most the test does not easily transfer from the labora- studied drugs in the lymphocyte transformation tory to the clinical situation; the time from setting test are the β-lactam antibiotics and anti-epilep- up the test to recording results is at least 5–7 tics, particularly carbamazepine. Diseases in days; it involves sterile cell culture; the optimum which the test has been found to be frequently times for testing different drug-induced reactions positive include maculopapular exanthema, bul- are not always known; the patient’s existing drug lous exanthema, acute generalized exanthematous

[email protected] 4.7 Tests for Delayed Type Drug Hypersensitivity Reactions 121 pustulosis, DRESS, and severe anaphylaxis. It is response provoked by contact sensitizing agents. occasionally positive in cases of urticaria, angio- Confi rmation of the results of the assay and exten- edema, and some drug-induced hepatitis and sion of their value can be obtained by carrying out nephritis reactions and rarely positive in fi xed concurrent cytokine release measurements. drug eruption, vasculitis, and TEN. Positive lymphocyte transformation tests have been found with quite a big range of drugs including 4.7.3 Toward Nonproliferation- β-lactams, macrolides, tetracyclines, sulfon- Based In Vitro Assays: Cell amides, quinolones, antiepileptics, opioids, ACE- Surface Activation Markers, inhibitors, anti-tuberculosis drugs, NSAIDs, Cytokines, Chemokines, local anesthetics, pyrazolones, contrast media, and Skin-Homing Receptors neuromuscular blocking drugs, vitamins, and contact allergens such as p -phenylenediamine. For the detection of delayed type drug hypersen- Considering the magnitude of the problem of sitivity reactions, the lymphocyte transformation delayed hypersensitivity reactions to drugs and test may be the only readily available and well the diffi culties associated with the lymphocyte investigated ex vivo methodology with a suffi - transformation test, there is a need to develop ciently long-standing pedigree to be employed sensitive and specifi c tests that are more easily with any confi dence. However, as outlined above, and quickly carried out, widely applicable to the the test has some major limitations including its many forms of delayed drug reactions and valu- practicability, lack of specifi city to T cells, and able for use in the clinic as well as the research the time involved in its execution. Although the laboratory. BAT more closely mirrors the in vivo pathways leading to allergic manifestations, it cannot be used to detect non-IgE-mediated allergic reac- 4.7.2 The Local Lymph Node Assay tions. With these considerations in mind and with the steady accumulation of insights into cellular This formally validated test, based on measuring and molecular immune processes underlying the the proliferative activity of draining lymph node secretory and effector functions of antigen-spe- cells from mice following epicutaneous applica- cifi c T lymphocytes, proliferation-based assays tion of the test agent, is now preferred to the are beginning to be supplemented by some novel guinea pig sensitization test by the FDA, EPA, in vitro tests to detect and measure cell activation and OECD as the accepted method for assessing markers, signaling molecules, chemoattractants, the skin sensitizing potential of chemicals, i.e., transcription factors, and cytolytic molecules for identifying contact allergens. Originally based released by some lymphocytes. In the main, these on measuring the incorporation of radiolabeled investigations are still essentially research based thymidine into the DNA of lymph node cells, and before any new assay’s fi ndings and method- more recent protocols substitute 5-bromo-2- ologies can be seriously considered for the diag- deoxyuridine in an ELISA or fl ow cytometric nosis of drug allergies, studies will have to procedure. Although the local lymph node assay confi rm their usefulness in large numbers of clin- is generally applied to assessing the response to ically well-defi ned patients. sensitizing chemicals used industrially and/or contacted in the environment (e.g., dinitrochloro- 4.7.3.1 Cell Surface Activation benzene, picryl chloride), the method has been Markers applied successfully to detect sensitization by Activation markers such as CD69, CD25, CD71, drugs, for example, benzocaine and benzylpeni- and the MHC class II cell surface receptor cillin. The method can also be adapted for use as HLA-DR are expressed and may be upregulated an immune function assay by examining the on the T cell surface. CD69, widely used in vitro effect of orally administered drugs on the T cell and in vivo as a marker of T cell activation for

[email protected] 122 4 Diagnosis of Allergic Reactions to Drugs more than 15 years, was recently exploited in secretion were observed for IL-2, IL-5, IL-13, fl ow cytometry studies for the detection of drug- and IFN-γ in response to sulfonamide and reactive T cells from patients with delayed type β-lactam drugs in the PBMC from patients aller- drug hypersensitivities. Freshly isolated PBMC gic to these drugs but no differences were seen in were cultured in the presence of drug or IL-2 for the cytokine secretion patterns of sulfonamide- 48 h before examining CD69 expression on and β-lactam-reactive PBMC. For PBMC from CD4+/CD8+ T cells by fl ow cytometry with healthy subjects, sulfonamide and β-lactam drugs fl uorescent-labeled monoclonal antibodies to stimulated statistically signifi cant increases in CD69, CD3, CD4, and CD8. Cells from lympho- IL-1β and IL-6 indicating that before measurement cyte transformation test-positive patients with of cytokine/chemokine release can be considered delayed hypersensitivity showed a signifi cantly as a likely diagnostic tool, many more patients increased expression of CD69 following expo- will need to be examined and more information sure to the culprit drug. Upregulation of the acti- will be needed on the background spectrum of vation marker occurred in 0.5–3 % of T cells with cytokines secreted in response to different drugs. only a minority of the reactive cell population In another approach to monitor cytokine being drug-reactive T cells secreting cytokines. release in patients with delayed drug hypersensi- There were a higher number of bystander T cells tivities, fl ow cytometry and an ELISA assay were activated by IL-2 and possibly other cytokines. used to measure cytokine secretion by PBMC. Although it was concluded that upregulation of Drug-induced production of IL-5, a stimulant for CD69 was a promising tool to identify drug- B cells and immunoglobulin secretion, the anti- reactive T cells from patients with drug hypersen- infl ammatory cytokine IL-10, and IFN-γ was sitivities, developments in this area and progress demonstrated by fl ow cytometry, and secretion of with this approach have been less than expected. the T cell growth factor IL-2 was detected in the ELISA assay. Cytokines were not detected in less 4.7.3.2 Monitoring of Cytokines from than a 5-day incubation period. Flow cytometry T Cells and the ELISPOT Assay and ELISA detected drug-specifi c cytokine pro- One relatively new approach is based on the duction in 75 % and 79 % of patients, respectively. detection of drug-specifi c cytokine production by Combining both procedures increased the sensi- cells from patients with a history of drug hyper- tivity to 100 %. In another study designed to uti- sensitivity following stimulation in vitro with lize the drug-induced release of cytokines as a drugs well known to provoke delayed type reac- sensitive assay for the diagnosis of cutaneous tions. During the last decade, work has shown adverse reactions to drugs, detection of IFN-γ was that measurement of secreted cytokines IL-5 and undertaken to investigate T cell involvement in IFN-γ can be useful for diagnosis of drug hyper- patients with maculopapular exanthema caused sensitivity. More recently, the secretion of a range by amoxicillin (see below). IFN-γ was selected of cytokines and chemokines has been examined since it is thought to be important in the patho- in attempts to identify promising “markers” for physiology of maculopapular exanthema, and the the in vitro detection of T cells sensitized to expression of this type 1 cytokine is restricted to drugs. As a fi rst approach, PBMC from patients activated T cells. with delayed type drug hypersensitivity were The ELISPOT (enzyme-linked immunospot) stimulated in vitro with drug for extended periods assay is based on classical immunoassay princi- (generally 72 h), and liberated cytokines were ples and the detection procedures employed in measured in the supernatants using immunoas- ELISA assays. Its sensitivity, ease of use, employ- says for 17 different cytokines/chemokines viz., ment of highly reactive, standardized monoclonal IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8 (CXCL8), antibodies, commercial availability, and ready IL-10, IL-12, IL-13, IL-17, G-CSF, GM-CSF, application to studies on individual cell types IFN-γ, TNF-α, CCL2, and CCL4. Increases in makes it a good choice for research and diagnostic

[email protected] 4.7 Tests for Delayed Type Drug Hypersensitivity Reactions 123 investigations of cellular effector–target inter- T lymphocytes and natural killer (NK) cells. actions. The methodology involves specifi c Granzyme B is constitutively expressed by mem- capture and immobilization of the target mole- ory but not by naïve cytotoxic T lymphocytes cule (e.g., a cytokine) with a complementary making the protease a likely candidate to be monoclonal (usually) antibody and visualization utilized for the assessment of cell-mediated cyto- of the reaction by addition of a detecting (second) toxicity. A highly sensitive ELISPOT assay has antibody tagged with a highly sensitive enzyme been developed for granzyme B but, unlike the label, for example, an anti-cytokine labeled with IFN-γ ELISPOT assay, it directly measures the streptavidin-alkaline phosphatase. The ELISPOT release of a cytolytic protein making it a more assay was employed to monitor the appearance of direct measure of antigen-specifi c cytotoxic T IFN-γ following amoxicillin and ceftriaxone lymphocyte lytic activity. An additional advan- challenge of the PBMC of 22 patients with a tage of monitoring granzyme B secretion is its well-documented history of delayed hypersensi- early release following effector–target contact. tivity to β-lactams manifesting as maculopapular The enzyme is detectable as early as 10 min after exanthema. The assay detected IFN-γ, and hence interaction, signifi cant amounts are measurable drug-specifi c T cells, in 20 of the 22 patients after 30 min, and maximum levels are obtained examined providing evidence that maculopapular after 4 h. Measurable amounts of INF-γ are seen exanthema is mediated by IFN-γ-producing only after 1 h. An ELISPOT assay for granzyme T cells. Results showed that the ELISPOT assay B and surface expression of CD107a were can detect amoxicillin-specifi c T cell precursors recently utilized for the detection of cytotoxic as low as 1:30,000 blood leukocytes and with a and NK cells in peripheral blood of patients with frequency of 30–125 per 106 PBMC; that the test various drug-induced skin diseases. CD107a, can distinguish patients with immediate and also known as LAMP-1 (lysosomal-associated delayed reactions; T cells that cross-react with membrane protein 1), is a membrane glycopro- other β-lactams can be detected; and the tein degranulation marker on CD8+ lymphocytes ELISPOT assay is more sensitive than the lym- and NK cells. The assay proved highly specifi c phocyte transformation test for the diagnosis of for detecting drug-reactive cytotoxic cells in delayed type hypersensitivity to β-lactams. The peripheral blood of drug-allergic patients but no sensitivity and specifi city of the assay for the strict correlation between the granzyme B assay diagnosis of delayed type hypersensitivity to and the lymphocyte transformation test was β-lactams were 91 % and 95 % respectively. found. Both CD4+ and CD8+ T cells expressed From the study, the investigators concluded that CD107a supporting belief both of their involve- other Th1 and Th2 cytokine-producing cells ment in drug hypersensitivities and the contribu- should be examined in sensitive assays with the tion of NK cells to the observed drug-induced view to detecting specifi c T cells and improving degranulation. the diagnosis of many drug-induced delayed reactions and it was suggested that the ELISPOT 4.7.3.4 Chemokines and Skin-Homing assay might represent an important advance in of T Cells the quest for improved tests for in vitro ex vivo The skin-associated chemokine CCL27, also diagnosis of such reactions. called Skinkine, Eskine, and CTACK (cutaneous T cell-attracting chemokine) and its receptor 4.7.3.3 Granzyme B ELISPOT CCR10 are associated with skin-homing of T Assay for the Detection lymphocytes and are implicated in T cell- of Drug-Reactive T Cells mediated infl ammation of the skin. Most skin- An interesting and highly promising candidate infi ltrating lymphocytes in patients suffering marker protein for drug-specifi c T cells is the ser- from contact dermatitis and psoriasis express ine protease granzyme B expressed by cytotoxic CCR10 and CCL27–CCR10 interactions appear

[email protected] 124 4 Diagnosis of Allergic Reactions to Drugs to play an important role in T cell-mediated skin the disease and they may be involved in the infl ammation. CCL27 can be induced by TNF underlying pathophysiologic mechanisms. and IL-1β and suppressed by glucocorticoste- Patient numbers were small, and although CLA roids. Intracutaneous injection of CCL27 attracts must be regarded as a promising marker for aid- lymphocytes and neutralization of the interaction ing efforts to understand the relationship between of the chemokine with its receptor impairs lym- T cells, drugs, and adverse delayed skin reac- phocyte recruitment to the skin and suppresses tions, research in this area must be seen as being allergen-induced infl ammation. Utilization of still in its infancy. this knowledge of the CCL27–CCR10 interaction might provide the basis of investigations that lead to a better understanding of T cell-mediated 4.8 And Finally: Is the Patient skin infl ammation in different delayed type drug Allergic to the Drug? hypersensitivity reactions and stimulation of further investigations of other chemokine–receptor The aim must be to establish or disprove a interactions of skin-infi ltrating lymphocytes. causal relationship between the drug and the Some encouraging in vitro test results with the patient’s reaction and, if there is such a relation- CCL27-CCR10 interaction are beginning to ship, the reaction type and mechanisms should appear, for example, results demonstrating that be determined if possible. The approach should levels of expressed CCL27 in skin biopsies from be both methodical and meticulous. For a reac- two patients with bullous skin reactions were tion that is severe or life-threatening, the drug higher than those found in healthy subjects and should not be readministered. If the reaction is a other drug-induced exanthemas, and resolution type A adverse drug reaction due to the drug’s was associated with return to normal expression pharmacological effects (see Sect. 1.1.2 ), low- levels of both CCL27 and its receptor. These ering the dose may be all that is necessary to fi ndings indicate that the CCL27–CCR10 inter- avoid, for example, a toxic reaction or known action may be involved in the selective recruit- side effect and for medication to safely con- ment to the skin of certain cytotoxic lymphocytes tinue. For a reaction that is not severe, a chal- in SJS and TEN and this and other chemokine-T lenge test to confi rm or eliminate suspicion can cell receptor interactions may be involved in be undertaken. For type B immunologic reac- other drug-induced cutaneous reactions. tions it is necessary to establish the underlying mechanism. This can be done by employing 4.7.3.5 Cutaneous Lymphocyte- confi rmatory tests if they are available. In many Associated Antigen cases, if not most, such tests are not available, Cutaneous lymphocyte-associated antigen (CLA) and then it may be necessary to avoid the drug is a l -fucose-containing carbohydrate epitope on as a precaution and prescribe an alternative drug the P-selectin glycoprotein ligand-1, a surface if one is suitable and available. Otherwise, a glycoprotein expressed on the majority of periph- graded challenge with the implicated drug can eral blood leukocytes. Most T cells that infi ltrate be carried out, but this should be done only if the skin express CLA, and CLA positive cells the reaction was not life-threatening and clearly have been implicated in contact dermatitis to not an IgE-antibody-mediated reaction. nickel in some patients with delayed cutaneous Nevertheless, if the medication is essential or allergic reactions. Studies on the expression of highly desirable, appropriate desensitization CLA by T cells from patients with exanthema- (Sect. 3.5 ) should be considered. The overall tous reactions induced by a range of drugs includ- strategy in looking for a causal relationship and ing β-lactams have so far shown that CLA the methodical approach pursued is summarized positive cells appear to parallel the evolution of in Fig. 4.10 .

[email protected] Summary 125

Compatible with an allergic reaction?

Yes No No Skin test / in vitro test available? Drug important and provocation possible?

Yes No Yes

Severe reaction? Yes Special precautions for Drug provocation test patients with higher risk No

Immediate reaction? Drug hypersensitivity confirmed?

Yes No Yes No

Skin prick test Patch test Intradermal test Intradermal test

++- -

Drug allergy confirmed Drug avoidance No drug avoidance Patient education Documentation

Fig. 4.10 Algorithm for the use of skin testing (and considerations for skin test procedures in the diagnosis of suitable available in vitro tests) in the diagnosis of drug hypersensitivity. Allergy 2002;57:45. Reproduced drug hypersensitivities. From Brockow K et al. General with permission from John Wiley and Sons

• Depending on the drug and the severity of the Summary patient’s drug reaction, the initial intradermal test injection may range from a small dilution • In diagnosing drug allergies, the patient’s his- of 1:10 or 1:100 of the prick test concentra- tory, skin testing, some in vitro laboratory tion to more extreme dilutions of up to tests, and the challenge test are the backbone 1:100,000. If no reaction is seen, the concen- of the investigative procedures. tration is increased in logarithmic steps until • A detailed and thorough clinical history of the the ﬁ nal concentration is reached, and this patient is the most important component of the maximum concentration should not be diagnostic process. exceeded. A positive reaction is an increase in • If skin prick testing elicits no reaction, intra- diameter of more than 3mm over the initial dermal testing is usually employed. The latter 20 ul injection bleb (usually ~2 mm) accom- test is more sensitive but produces more false panied by erythema. positives, that is, it is less speciﬁ c. • Intradermal testing is normally contraindi- • Depending on the drug, a prick test wheal cated in patients who have developed SJS, diameter at least 3 mm greater than the nega- TEN, and erythema multiforme. tive (saline) control or a wheal at least half the • Skin tests should be read at 15–20 min for diameter of the positive (histamine) control is immediate reactions and 48–72 h (or some- considered a positive result. times later) for delayed reactions.

[email protected] 126 4 Diagnosis of Allergic Reactions to Drugs

• The patch test is both a screening test for ratio of total to mature tryptase is typically hypersensitivity and a provocation test in the less than 10. target organ, the skin. • Histamine release from human blood leuko- • If pure drug is used for patch testing, concen- cytes after challenge with drug in vitro is trations should begin at about 0.1 % and prog- occasionally employed but in general, for ress to 1–10 % if results are negative. For diagnostic purposes, histamine concentrations DRESS, SJS and TEN (if testing must be in biological fl uids have rarely been routinely done) and for some drugs, for example, aciclo- measured; the half-life of histamine in plasma vir, carbamazepine, and pseudoephedrine, is short—approximately 1–2 min. testing should start with lower concentrations. • The release of cysteinyl leukotrienes from iso- • Patch test reactions are read after 48 and 72 or lated peripheral blood leukocytes following 96 h. In some cases a reaction occurs in less allergen challenge in vitro has been utilized as than 2 days (e.g., abacavir) while with some a test for immediate hypersensitivity in the other drugs such as corticosteroids, aminogly- form of the Bühlmann CAST® (Cellular coside antibiotics, and phenylephrine, reac- Allergy Stimulation Test) assays offered com- tions may occur after 6 or 7 days. mercially as CAST ® ELISA a microtiter plate • The COADEX classifi cation should be used to ELIZA immunoassay and as a fl ow cytometric assess clinical relevance of positive patch tests. assay Flow CAST® . Although showing some • Assays for drug-specifi c serum IgE antibodies promising results with β-lactams and NSAIDs, are useful in cases of skin test-negative or wider assessment of the methods in more lab- equivocal reactors or when skin tests are unre- oratories is needed. liable or unavailable. • Given the technical improvements and • In interpreting results of IgE antibody tests, advances made with BAT, the increasing ROC curves provide more information to aid acquisition of fl ow cytometric quantifi cation discrimination between positive and negative equipment by laboratories, the test’s validation results. The concentration of IgE antibodies for a number of drugs, and, in some cases, the assessed by immunoassay is related to the potential of the test to detect non-IgE-mediated presence of allergic symptoms. basophil activation, it can be appreciated why • A drug challenge, or provocation test, is the the method continues to be applied to an controlled step-wise administration of a drug increasingly wide range of drugs and other in a supervised hospital environment in order therapeutic agents. In a new application of the to determine if the drug was the causative agent methodology termed “Histafl ow,” fl ow cytom- in a patient’s allergic reaction. The test is the etry has been used to examine histamine and best way to confi rm an allergic reaction, and it its release along with the simultaneous quanti- is considered to be the “gold standard” in the fi cation of basophil activation markers. diagnosis of drug hypersensitivity reactions. • In vitro tests for delayed drug reactions include • Drug challenge tests are particularly important the lymphocyte transformation test and local when other usually employed tests, particu- lymph node assay but nonproliferation-based larly skin tests, are not available or possible. in vitro assays of cell surface activation mark- • Increased mast cell tryptase concentrations ers, cytokines, chemokines, and skin-homing are a valuable indicator of an anaphylactic receptors will be increasingly applied. reaction especially during anesthesia and • The ELISPOT assay shows potential for use in although elevated levels favor an IgE- drug allergy diagnosis. It can detect amoxicillin- antibody-mediated cause, they do not always specifi c T cell precursors as low as 1 : 30,000 distinguish between an anaphylactic and an blood leukocytes and can distinguish patients anaphylactoid reaction. Normal levels of with immediate and delayed reactions. The mature tryptase in serum are less than 1 ng/ml assay is more sensitive than the lymphocyte whereas levels equal to or above 1 ng/ml indi- transformation test for the diagnosis of delayed cate mast cell activation. In anaphylaxis, the type hypersensitivity to β-lactams.

[email protected] Further Reading 127

• A highly sensitive ELISPOT assay has been Lachapelle JM, Maibach HI. Patch testing and prick test- developed for granzyme B but, unlike the ing. A practical guide. Berlin: Springer; 2003. γ Laroche D, Dubois F, Gérard J-L, et al. Radioimmunoassay IFN- ELISPOT assay, it directly measures for plasma histamine: a study of false positive the release of a cytolytic protein making it a and false negative values. Br J Anaesth. 1995;74: more direct measure of antigen-specifi c cyto- 430–7. toxic T lymphocyte lytic activity. Matsson PNJ, Hamilton RG, Esch RE, et al. Analytical performance characteristics and clinical utility of • Studies on chemokines such as CCL27 associ- immunological assays for human immunoglobulin E ated with skin-homing of T cells, and cutane- (IgE) antibodies and defi ned allergen specifi cities; ous lymphocyte-associated antigen CLA, are approved guidelines—second edition. CLSI document promising markers for aiding efforts to under- I/LA20-A2. Wayne, PA: Clinical and Laboratory Standards Institute; 2009. stand the relationship between T cells, drugs, NCCLS. (2004). Protocols for determination of limits and adverse delayed skin reactions. of detection and limits of quantitation: approved guideline. NCCLS document EP17-A. NCCLS, Wayne, PA Santamaria Babi LF, Picker LJ, Perez Soler MT, et al. Further Reading Circulating allergen-reactive T cells from patients with atopic dermatitis and allergic contact dermatitis Aberer W, Kränke B. Provocation tests in drug hypersen- express the skin-selective homing receptor, the cutane- sitivity. Immunol Allergy Clin North Am. 2009;29: ous lymphocyte-associated antigen. J Exp Med. 567–84. 1995;181:1935–40. Barbaud A. Drug patch testing in systemic cutaneous drug Schwartz LB. Diagnostic value of tryptase in anaphylaxis allergy. Toxicology. 2005;209:209–16. and mastocytosis. Immunol Allergy Clin North Am. Barbaud A. Skin testing in delayed reactions to drugs. 2006;26:451–63. Immunol Allergy Clin North Am. 2009;29:517–35. Schwartz LB, Min H-K, Ren S, et al. Tryptase precursors Barbaud A, Gonçalo M, Bruynzeel D, et al. Guidelines for are preferentially and spontaneously released, whereas performing skin tests with drugs in the investigation of mature tryptase is retained by HMC-1 cells, mono- cutaneous adverse drug reactions. Contact Dermatitis. Mac- 6 cells, and human skin-derived mast cells. 2001;45:321–8. J Immunol. 2003;170:5667–73. Bernstein IL, Li JT, Bernstein DI, et al. Allergy diagnostic Söderstrom L, Kober A, Ahlstedt S, et al. A further evalu- testing: an updated practice parameter. Ann Allergy ation of the clinical use of specifi c IgE antibody test- Asthma Immunol. 2008;100 Suppl 3:S66–S121. ing in allergic diseases. Allergy. 2003;58:921–8. Brockow K, Garvey LH, Aberer W, et al. Skin test concen- Steiner M, Harrer A, Lang R, et al. Basophil activation test trations for systemically administered drugs – an for investigation of IgE-mediated mechanisms in drug ENDA/EAACI Drug Allergy Interest Group position hypersensitivity. J Vis Exp. 2011;55:e3263. paper. Allergy. 2013; doi: 10.1111/all.12142 . doi: 10.3791/3263 . Kalyuzhny AE, editor. Handbook of ELISPOT. Methods Zweig MH, Campbell G. Receiver-operating characteris- and protocols. Methods in molecular biology, vol. 792. tic (ROC) plots: a fundamental evaluation tool in clini- New York: Human; 2012. cal medicine. Clin Chem. 1993;39:561–77.

[email protected] β-Lactam Antibiotics 5

Abstract The β-lactam antibiotics comprise four main classes of drugs; penams (penicillins), cephems (cephalosporins), monobactams, and carbapenems. Penicillins can cause all four types of hypersensitivity responses. IgE antibodies in patients’ sera detect a spectrum of antigenic speciﬁ cities, show heterogeneous recognition and cross-reactive responses, and may distinguish ﬁ ne structural features, e.g., amoxicilloyl and amoxicillanyl determinants. With a negative history of penicillin allergy, the incidence of positive skin tests is 2–7 %. For skin test-positive patients the risk of an acute allergic reaction ranges from 10 % (negative history) to 50–70 % (positive history). IDTs with delayed reading and patch tests are used to diagnose delayed reactions. Aminolysis of cephalosporins produces unstable intermediates that decompose to penaldate and penamaldate structures resulting in only the R1 side chain remaining from the original molecule. With some allergic patients the R2 side chain and/or the whole cephalosporin molecule are also recognized by IgE antibodies. Testing with penicillins does not reliably predict cephalosporin allergy unless the side chains of the penicillin and the culprit cephalosporin are similar. Aztreonam shows little, if any, cross-reaction with penicillins and cephalosporins. The practice of avoiding imipenem and meropenem therapies in penicillin- allergic patients should be reconsidered. There has been an increase in cases of immediate hypersensitivity to clavulanic acid.

So named because of the presence of a four- pyrrole ring in the carbapenems. Monobactams membered β-lactam ring in the molecules, the consist of a β-lactam ring free of any other ring β-lactam antibiotics comprise four main classes attachment. Other classes of β-lactam antibacte- of drugs that possess antibacterial action, viz., rials, each with a small number of less often used penams (penicillins), cephems (cephalosporins), drugs, are the penems, clavams, carbacephems, monobactams, and carbapenems. The β-lactam oxacephems, and cephamycins (Fig. 5.1 ). ring is fused to a thiazolidine ring in penams, a Henceforth here, penams and cephems are dihydrothiazine ring in cephems, and a dihydro- referred to by the more commonly used names,

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 129 Relationships, DOI 10.1007/978-1-4614-7261-2_5, © Springer Science+Business Media, LLC 2013 [email protected] MONOCYCLIC -LACTAM MONOBACTAMS H H

R-CONH CH3

N O SO3H e.g.Aztreonam BICYCLIC PENICILLINS (PENAMS) CARBAPENEMS R1 H H H H S CH R-CONH CH3 CH3-CHOH S-R2 N CH3 N O O H COOH COOH e.g. e.g. Penicillin G Ampicillin Doripenem Imipenem Carbenicillin Meropenem Penicillin V Amoxicillin Temocillin Ertapenem Phenethicillin Bacampicillin Ticarcillin Propicillin Epicillin CLAVAMS (OXAPENAMS) Hetacillin Azlocillin H H Oxacillin Pivampocillin O Cloxacillin Mezlocillin H Piperacillin Dicloxacillin Methicillin CH-CH2-R Flucloxacillin Nafcillin N O H COOH e.g. Clavulanic acid

CARBACEPHEMS H H H H S CH2 R1-CONH R1-CONH N N O R2 O R2 COOH e.g. Loracarbef COOH e.g. Cephacetrile Cefaclor Cefdinir Cefadroxil Cefamandole Cefetamet Cephalexin Cefonicid Cefixime Cephaloglycin Cefmenoxime Cephaloridine Cefotiam Cephalothin Cefprozil Cefodizime Cephapirin Cefuroxime Cefoperazone Cefatrizine Cefotaxime Cefopime Cefazedone Cefpiramide Cefozopran Cefazolin Cefpodoxime Cephradine Cefpirome Cefroxadine Ceftazidime Ceftezole Ceftaroline Ceftibuten fosamil Ceftizoxime Ceftobiprole Ceftriaxone CEPHAMYCINS OXACEPHEMS (1-oxocephalosporins) H CO H H CO H 3 S 3 O R1-CONH R1-CONH N N O R2 O R2 COOH COOH e.g. Cefoxitin Cefmetazole Cefotetan e.g. Moxalactam

Fig. 5.1 Structures of the four main classes of β-lactam used β-lactam antibacterials include the carbacephems, antibiotics, penams (penicillins), cephems (cephalospo- oxacephems, cephamycins, clavams (see Sect. 5.5 and rins), carbapenems, and monobactams. Less frequently Fig. 5.18d ), and penems (not shown)

[email protected] 5.1 Penicillins 131 penicillins and cephalosporins, respectively. Table 5.1 Clinical adverse reactions to penicillin and These two drug classes of antibacterials are associated immune mechanisms highly effective in treating infections and gener- Immediate IgE-mediated reactions (Type I) ally not toxic and, even after extensive use over Urticaria many decades and the generation of resistant Angioedema organisms, penicillins and cephalosporins remain Asthma the most frequently prescribed antibiotics. Anaphylaxis However, soon after the introduction of the ﬁ rst Urticaria penicillin, the propensity of this group of drugs to Angioedema Laryngeal edema cause allergic reactions ranging from simple Flushing rashes to life- threatening anaphylaxis was Pruritus recognized. Penicillins, together with cephalo- Nausea, vomiting, diarrhea, headache sporins, are probably still the most common Bronchospasm cause of drug allergy and this continues to be a Tachycardia, arrhythmias problem in antibiotic selection and risk avoid- Cardiovascular collapse ance today. Non-immediate reactions, not IgE-mediated In this chapter, the four main classes of Antibody-mediated (Type II, cytotoxic) β-lactams will be dealt with separately with great- Hemolytic anemia est emphasis on the heavily used penicillins and Thrombocytopenia cephalosporins and with reference to the other Immune complex-mediated (Type III) classes when appropriate. Despite the impressive Vasculitis immunochemical insights from the pioneering Serum sickness drug allergy studies that began over 50 years ago T cell-mediated (Type IV) with the early investigations of Levine, Parker, De Contact dermatitis Weck, and Dewdney, clinically relevant molecu- Drug-induced skin eruptions lar aspects of β-lactam drugs have been largely ignored in recent years with steps necessary to identify and deﬁ ne allergenic structures and or believe, that they are allergic to the medication improve diagnostic agents neglected. While a full and 80–90 % of patients who claim to be allergic and reliable case history, skin tests, challenge to penicillins are not. It has been claimed that vir- tests, and serum IgE antibody investigations still tually all patients with a negative skin test to the form the basis of an accurate diagnosis, a good drug(s) can take penicillins without serious knowledge of different allergenic structures and sequelae. Penicillins have long been known to be an improved range of test materials for the recog- the most common cause of both drug-induced nition of individual sensitivities would assist cli- anaphylaxis and drug-induced allergic reactions nicians in achieving accurate and precise causing an estimated 75 % (500–1,000) of deaths diagnoses, identifying likely cross-reactive drugs each year in the USA and 26 % of fatal drug- and selecting safe alternative drugs. induced anaphylaxis in the UK. Penicillins can cause all four types of hypersensitivity responses provoking type I IgE- 5.1 Penicillins mediated reactions such as urticaria, angioedema, asthma, and anaphylaxis; type II 5.1.1 Incidence of Penicillin antibody-mediated hemolytic anemia and Hypersensitivity and Clinical thrombocytopenia; type III immune complex- Aspects mediated serum sickness-like reactions and vasculitis; and type IV T cell-mediated contact The incidence of hypersensitivity reactions to dermatitis, rashes, and other skin eruptions penicillins is about 1–2 % but, importantly, up to (refer to Chaps. 2 and 3 ). Table 5.1 lists clinical about 10 % of patients taking a penicillin report, adverse reactions, together with their immune

[email protected] 132 5 β-Lactam Antibiotics

mechanisms, that may occur following the administration of a penicillin. As well as a classiﬁ cation based on immune mechanisms, reactions are often grouped as immediate, accelerated, or late onset, with immediate reactions occurring within 1 h, accelerated reactions occurring between 1 and 72 h, and late reactions after 72 h. Clinical manifestations can occur singly or in combination—for immediate responses the most common reactions are urticaria and angioedema; urticaria and maculopapular rashes occur commonly in accelerated reactions; and erythema multiforme, skin eruptions, hemolytic anemia, and a serum sickness- like reaction are seen as delayed reactions. Delayed onset urticarial or maculopapular rashes are frequently seen especially in children and many are labeled allergic without evidence or even testing. Allergy is overdiagnosed in these patients since skin rashes are rarely reproducible by challenge testing (Fig. 5.2a, b ) and viral infections are suspected in many of the penicillin-induced rashes.

5.1.2 Penicillin Antigens and Fig. 5.2 Erythematous rash on a patient’s neck (a), arms, and hands (b) following oral challenge with amoxicillin. Allergenic Determinants The patient showed a negative intradermal test to the drug and responded on the last oral challenge dose of amoxicil- All penicillins contain a β-lactam ring fused to a lin. (Photographs courtesy of Dr P. A. J. Russo and Dr J. thiazolidine ring and individual penicillins are S. Fok, Department of Clinical Immunology and Allergy, Royal Adelaide Hospital, Adelaide) distinguished by the nature of the side chain R group (Fig. 5.3 ). The structures of the most clinically important and frequently used penicillins 5.1.2.1 The Penicilloyl Determinant are shown in Fig. 5.4 . and Benzylpenicillenic Acid Extensive investigations over a period of more Of all the penicillin breakdown products and pro- than 30 years on the formation of antigenic and tein conjugates studied, most is known about the allergenic determinants of benzylpenicillin (peni- penicilloyl determinant. The major populations cillin G) led to the unraveling of complex path- of antibodies in sera from experimental animals ways and steps in the formation of a number of immunized with benzylpenicillin, and from ultimately proven, and some putative, allergenic humans following penicillin therapy, were found determinants. At the time of that research, the to be complementary to this determinant leading accepted view was that low molecular weight to its designation as the major penicillin antigen. drugs and other chemicals must ﬁ rst combine irre- It was estimated that of all the penicillin mole- versibly with a macromolecular carrier, usually cules that became covalently bound to protein protein, to produce hapten–carrier complexes that under physiological conditions, 95 % form peni- stimulate a speciﬁ c antibody response. Although cilloyl groups and it was this quantitative pre- this view is still largely accepted, some exceptions dominance rather than allergenic potency or may occur (see in particular Chaps. 3 and 7 ). clinical or immunological importance that the

[email protected] 5.1 Penicillins 133

HH on protein amino groups, ﬁ rst demonstrated by S R CONH Bernard Levine at high pH (Fig. 5.5 ). Later stud- CH3 ies showed that the reaction also proceeds at neu-

N CH3 tral pH. Benzylpenicillin also readily rearranges O to form an isomer, D -benzylpenicillenic acid, a H COOH highly reactive compound which, like benzylpenicillin, binds selectively to lysine residues of Side-chain b-lactam Thiazolidine human serum albumin (HSA) forming penicil- ring ring R-CONH- loyl–lysine adducts (Fig. 5.5 ). The rearrange- ment to penicillenic acid occurs in vitro and in vivo where it is not dependent on enzymatic involvement. A direct demonstration of the presence of penicilloylated protein conjugates in vivo was fi rst achieved by Levine in inhibition experiments with sera from patients treated with high doses of penicillin. Levine argued that formation of the penicilloyl specifi city more likely pro- ceeded via the penicillenic intermediate because anti-benzylpenicillin antibodies were specifi c for a diastereoisomeric mixture of benzylpenicillin, whereas benzylpenicilloyl hapten, formed from benzylpenicillin by direct aminolysis, would produce only the D - α-diastereoisomer. It was later pointed out, however, that epimerization by the direct route is also possible. Recently, the question of diastereoisomeric benzylpenicilloyl antigen formation from benzylpenicillin and benzylpenicillenic acid was investigated in a mass spectrometric and molecular modeling e.g. Benzylpenicillin study. Both benzylpenicillin and benzylpenicillenic acid were shown to covalently bind to lysine R = CH2 residues in HSA to form penicilloyl adducts in vitro, but the two compounds showed differences Fig. 5.3 Side-by-side two- and three-dimensional peni- in their binding targets. Whereas the parent drug β cillin structure highlighting the side chain (R), -lactam, showed marked preferential binding to Lys199, and thiazolide ring structures. From Baldo BA. Diagnosis of allergy to penicillins and cephalosporins. Allergy Clin benzylpenicillenic acid bound to this residue and Immunol Int. 2000;12:206. Reprinted with permission to Lys525 as well. Characterization of the iso- from © 2000 Hogrefe & Huber Publishers (now Hogrefe meric adducts, formed when benzylpenicillin and Publishing. http://www.hogrefe.com ) benzylpenicillenic acid were incubated with albumin in vitro, revealed two isomers for both term “major” was applied to. Another potential compounds at each of the modifi ed lysines, confusion with the term is the possibility that it although the 5R,6R diastereomer predominated may be thought to refer to the major degradative for the parent drug (pathway 1, Fig. 5.5 ) and the product of the penicillins, viz., penicilloic acid. 5R,6S diastereomer predominated for the acid There is more than one pathway for the formation (pathway 2, Fig. 5.5 ). Prolonged incubation of of the penicilloyl determinant. One is by direct benzylpenicillenic acid with HSA produced an reaction of benzylpenicillin involving the open- increase in the relative amount of the 5R,6S dia- ing of the β-lactam ring and nucleophilic attack stereomer. Investigations showed that epimerization

[email protected] H H S R-CONH CH3

N CH3 O H COOH

R- R-

Benzylpenicillin CH Ampicillin CH (Penicillion G) 2 NH2

Phenoxymethylpenicillin OCH Amoxicillin HO CH (Penicillion V) 2 NH2

Epicillin CH

NH2

OCH Carboxypenicillins 3 CH Ticarcillin Methicillin COOH S

OCH3 Carbenicillin CH COOH Nafcillin Acylaminopenicillins (Ureidopenicillins) CH OC H 2 5 HN Azlocillin O Isoxazolyl Penicillins N O Oxacillin N H N CH3 O CH Cl HN Cloxacillin Mezlocillin O N N CH O 3 O Cl N

SO2CH3 Dicloxacillin

N CH3 CH Cl O HN Cl Piperacillin O O N Flucloxacillin

N CH3 F O O N

CH2CH3

Fig. 5.4 Structures of the different side chain (R) groups lins, and ureidopenicillins) and biological properties on individual penicillins grouped according to their simi- (penicillinase resistance, extended spectrum) larities of structure (aminopenicillins, isoxazolylpenicil-

[email protected] 5.1 Penicillins 135

H H H N S

O N O COOH Pathway 1 Benzylpenicillin Pathway 2 NH2

R NH H H H 2 N S N HS R O HN O HN O NH O COOH COOH Benzylpenicilloyl R Benzylpenicillenic acid determinant−(5R,6R) adduct

epimerization NH2

R H H H N S

O HN O NH COOH

Benzylpenicilloyl R determinant−(5R,6S) adduct

Fig. 5.5 Pathways for the formation of the penicilloyl Maggs, John Farrell, Catherine S Lane, Andrew V determinant. Pathway 1: Opening of the β-lactam ring of Stachulski, Neil S French, Dean J Naisbitt, Munir benzylpenicillin and nucleophilic attack on protein amino Pirmohamed, B. Kevin Park. Direct evidence for the for- groups, specifi cally ε-amino groups of lysine residues mation of diastereoisomeric benzylpenicilloyl haptens Lys199. Pathway 2: Benzylpenicillin rearranges to its iso- from benzylpenicillin and benzylpenicillenic acid in mer benzylpenicillenic acid which binds selectively via patients. J Pharmacol Exp Ther. 2011; 338: 841–9. nucleophilic attack to Lys199 and Lys525 of human serum Reprinted with permission from American Society for albumin to form benzylpenicilloyl-lysine adducts. From Pharmacology and Experimental Therapeutics Xiaoli Meng, Rosalind E Jenkins, Neil G Berry, James L of the 5R,6R diastereomer to the 5R,6S diastereo- of 59 lysine residues were modifi ed. Modifi ed mer does not occur after the drug becomes lysines at positions 541, 432, 195, and 190 were covalently bound. This indicated that the latter detected in the plasma of piperacillin-exposed diastereomer can only be formed by rearrange- patients with cystic fi brosis. ment of benzylpenicillin to benzylpenicillenic Structures of the degradation products of the acid followed by covalent reaction of the acid penicilloyl determinant, viz., penamaldyl and with the lysine residues of the protein (pathway 2, penaldyl determinants and penicillamine, are Fig. 5.5 ) rather than via pathway 1 followed by shown in Fig. 5.6 . epimerization to form the 5R,6S diastereomer. Mass spectrometric methods were also employed 5.1.2.2 The Penicillenate, Penicilloic to detect and characterize antigens derived from Acid, Penicillamine, and the reaction of piperacillin with human serum Penamaldate Determinants albumin in phosphate buffer at pH 7.4. Two differ- Benzylpenicillenic acid, which forms readily from ent haptens were detected: one hypothesized to benzylpenicillin in aqueous solution, is unstable result from hydrolysis of the 2,3-dioxypiperazine and is thought to be allergenic, particularly in con- ring on the piperacillin side chain (Fig. 5.4) and tact skin allergy, after direct reaction with disul- the other by addition to the β-lactam ring. At low fi des and cysteine sulfhydryl groups (Fig. 5.6 ). drug concentrations, modifi cation of Lys541 Immunization of laboratory animals with penicil- occurred while at higher concentrations, up to 13 lenate–protein and penicilloyl–protein conjugates

[email protected] 136 5 β-Lactam Antibiotics

S S NH CH C-(CH ) CO NH 2 3 2 CH2 CH C-(CH3)2

O CNHCHCOOH O CNCH CO HN P HN P Penicoyl determinant Benzylpenicillanyl determinant

Protein NHSucc Protein DCC

S S NH CH C-(CH ) CO NH CH C-(CH ) 2 3 2 CH2 3 2

O CNCHCOOH O C NCHCOOH 6-Aminopenicillanic acid Benzylpenicillin S CH CO NH CH C-(CH ) isomerization 2 3 2 P O CNHCHCOOH S S hydrolysis SH NH N C CH C-(CH ) N C CH C-(CH ) 3 2 3 2 P CH CNHCH COOH CH C NH CH COOH 2 2 Benzylpenicilloyl determinant O O O O Benzylpenicillenate Benzylpenicillenic acid (benzyl penicillenic acid SH CH CO NH C CH C-(CH ) mixed disulfide) 2 3 2 O CNHCH COOH S decarboxylation S NH CO NH CH CO NH CH2 CH2 C-(CH3)2 2 CH C-(CH3)2 P HN CH COOH HOOC HN CH COOH Benzylpenamaldyl Benzylpenilloic acid Benzylpenicilloic acid determinant

SH CH CO NH 2 CH CHO C-(CH3)2 SH O CNH+ CH COOH NH 2 CH2 CO CH CH C(CH3)2 NH HOOC N CH COOH P Benzylpenamaldic acid Benzylpenaldyl Penicillamine determinant endogenous protein with cysteine -S-S- P P S S S S CH CO NH CH CO NH C(CH ) 2 CH CH C(CH3)2 2 CH2 CHO 3 2

HOOC N CH COOH + NH2 CH COOH Benzylpenamaldic acid− Benzylpenillo- Benzylpenamaldic protein conjugate aldehyde acid−protein (benzylpenamaldic acid conjugate cysteine mixed disulfide) (penicillamine cysteine mixed disulfide)

Fig. 5.6 Pathways for the formation of penicilloyl, penicillenate, penicilloic acid, penamaldate, penicillamine, and other determinants of benzylpenicillin showed that the penicillenate and penicilloyl Benzylpenicilloic acid, the main hydrolysis haptens were recognized as distinct determinants. product of benzylpenicillin, elicits wheal and Added evidence of the allergenicity of the peni- ﬂ are skin reactions in some patients and was con- cillenate determinant was the detection of com- sidered to be one of the so-called minor (in a plementary IgE antibodies, although this quantitative sense) determinants by Levine. determinant does not seem to be a clinically Decarboxylation of penicilloic acid gives rise to important allergen. penilloic acid, another of the minor determinants

[email protected] 5.1 Penicillins 137

(see Sect. 5.1.2.4.2 ). It has been suggested that in 5.1.2.4.1 History vivo, penicilloic acid reacts with cystine disulfi de The importance of these so-called minor linkages via its penamaldic acid intermediate to determinants was fi rst demonstrated by Levine and form benzylpenamaldic acid cysteine mixed coworkers who observed their marked association disulfi de and, then, via a penamaldate rearrange- with what was described at the time as “skin- ment, to penicillamine cysteine mixed disulfi de sensitizing” antibodies in penicillin-allergic and benzylpenilloaldehyde (Fig. 5.6). It is possi- patients. Levine concluded that “immediate allergic ble that these degradation products of penicilloic reactions to penicillin are most often mediated by acid can be formed in vivo. They are chemically skin-sensitizing antibodies of minor determinant equipped to react with protein carriers and, theo- specifi cities,” while penicilloyl- specifi c skin-sensi- retically at least, can function as antigens and tizing antibodies were “invariably associated with allergens. Some evidence from skin tests studies accelerated and late urticarial reactions and proba- with benzylpenicilloic acid suggests that positive bly mediate these reactions.” The penicilloyl-spe- skin reactions may be a response to penicillamine cifi c antibodies were thought to be mainly IgG and and/or penamaldate specifi cities, and skin tests IgM and it was suggested that these acted as block- with D -penicillamine-HSA and polylysine conju- ing antibodies preventing penicilloyl-mediated gates revealed a fairly high proportion of positive immediate reactions. Two skin test solutions were responses (13–41 %) in penicillin-sensitive originally recommended for diagnosis of penicillin patients. However, evidence that this group of allergy—benzylpenicilloyl–polylysine conjugate at penicillin determinants is allergenically signifi - a concentration of 10−6 M and a minor determinant cant remains insubstantial. mixture consisting of potassium benzylpenicillin, sodium benzylpenicilloate, and sodium benzylpe- 5.1.2.3 6-Aminopenicillanic Acid nilloate all at a concentration of 10−2 M. The penicil- and the Penicoyl Determinant loyl–polylysine conjugate contained 20 lysine 6-Aminopenicillanic acid, sometimes used as a residues with 13 of them coupled to the penicilloyl starting material for the synthesis of semisyn- hapten and the remaining lysines succinylated. In thetic penicillins, is weakly immunogenic in comparative skin tests on penicillin-allergic patients, laboratory animals, acts as a hapten inhibitor for it became clear that patient responses were hetero- reaction with the penicilloyl specifi city, and geneous—some reacted only to benzylpenicillin, does not appear to be an important penicillin only to penicilloate, or only to benzylpenicilloyl- allergenic structure. Any such importance polylysine or any combination of two or more of the 6-aminopenicillanic acid has is probably due to test reagents. In one of the original clinical studies the penicoyl derivative formed when it reacts on the minor determinants, 26 patients selected for with protein amino groups (Fig. 5.6 ). Early a positive skin test reaction to one or more of potas- reports of the allergenic activity of sium benzylpenicillin, sodium benzylpenicilloate, 6-aminopenicillanic acid were probably due to and sodium benzylpenilloate (all at 10-2 M) contamination by the penicilloyl specifi city so were skin tested with the major determinant ben- its clinical signifi cance and the allergenicity of zylpenicilloyl-polylysine and with benzylpenicillin, the more immunogenic and antigenic penicoyl benzylpenicilloate, benzylpenilloate and benzyl- determinant need to be fully evaluated. penicilloyl-amine to compare the allergenic activity of a range of minor determinants. The major 5.1.2.4 The “Minor” Determinants determinant was positive in 46 % of patients and of Penicillin benzylpenicillin in 62 % while the penicilloate and Metabolites other than the penicilloyl moiety are penilloate preparations were positive in 85 and believed to constitute about 5 % or less of admin- 73 % of patients, respectively, with the latter deter- istered penicillin and, together with penicillin G, minant not detecting any positive reactions missed are often referred to as minor determinants. by penicilloate. Although this seemed to indicate

[email protected] 138 5 β-Lactam Antibiotics that the presence of the penilloate specifi city ably be reused for a limited time since ~85 % of was redundant and could be left out of the mixture, the original activity is retained 3½ h after prepa- it produced more intense skin reactions than ration. Clearly, the solution to the stability prob- the penicilloate determinant in a few patients. lem with the minor determinants is the Penicilloyl-amine, prepared by reacting potassium preparation of freeze-dried compounds that can benzylpenicillin with ammonia and used at 10−2 M, be stored dried in the form of single-dose was positive in 77 % of patients, but other minor ampules and opened and used only when determinants, D -penicillamine, oxazolone (2-ben- needed. Progress in achieving this has been zyl-4-sodium hydroxymethylene-(5)-oxazolone), made (see Sect. 5.1.5.1 ). and benzylpenilloaldehyde, were either negative in The apparent general acceptance of testing all patients or reacted with only about 10 % or less. with penicilloyl–polylysine conjugate and a It was concluded that to avoid missing penicillin- minor determinant mix has obscured some allergic individuals, penicillin, penicilloate, aspects of the recognition of penicillin aller- penilloate, penicilloyl-amine, and benzylpenicil- genic determinants that are still poorly under- lin- polylysine must be used for skin testing. stood and defi ned. Over 40 years ago Levine believed that “the haptenic determinant specifi c- 5.1.2.4.2 Selection and Stability of ity of skin reactivity to penicillin is not known” Penicillin Minor Determinants and that the “specifi city appears to be toward a Although there seems to be general agreement hapten other than the BPO-[benzylpenicilloyl-] that minor determinants should be included in group, which is formed from penicillin but skin testing for penicillin-allergic sensitivity, which is not formed from the penicilloate–penil- uncertainty remains over which of the individ- loate group of compounds. Its identity has not ual compounds should constitute the “ideal” yet been determined.” He was also aware that, minor determinant mix. Results have shown that with more research, the need for additional about 7–14 % of penicillin skin test-positive minor determinants may become evident and, patients react only to the penicilloate specifi city unlike the major determinant, none of the minor and not to other penicillin determinants, but determinants had been covalently linked to a some investigators believe that conclusive dem- carrier to produce more effective skin test onstrations of the importance of minor determi- reagents by forming multivalent hapten conju- nants are lacking and testing should be gates. This remains the case today. undertaken only with penicilloyl- polylysine and Attempts to identify penicillin metabolites penicillin G. Most importantly, however, the have continued over the years by employing a chief problem preventing widespread routine range of techniques including thin layer chroma- evaluation and everyday diagnostic application tography, HPLC coupled with UV detection, of minor determinant mix preparations has been NMR, and mass spectrometry (MS), but the need the highly labile nature of the reagents, particu- for large amounts of sample, lack of sensitivity in larly the penicilloate and penilloate compo- detecting trace amounts of metabolites, and poor nents. For benzylpenicilloic acid, epimerization information of fragmentation for analysis has not at C-5 for (5R,6S)-, (5S,6R)-, and (5R,6R)- always led to the hoped-for progress. Application benzyl-D -penicilloic acids is well known and of newer MS-based approaches, however, such as likewise (5R,6R)-benzyl-d - penicilloate and the recent application of data-dependent liquid (5R)-benzyl-D -penilloate were found to be chromatography/multiple stage tandem MS, labile in aqueous solution, giving rise to a mix- revealed seven minor metabolites of penicillin G ture of diastereoisomers. In fact, aqueous minor in human serum. As well as the already known determinant mix solutions are too labile at room penicilloate and penilloate structures, other com- temperature to use other than immediately after pounds identifi ed were hydroxypenicilloate and preparation. Solutions stored frozen are stable glucuronide conjugates of penicilloate and three for at least 9 days while those at 4 °C can prob- other reactive metabolites. Such methods may

[email protected] 5.1 Penicillins 139 help to provide a full defi nition of penicillin Table 5.2 Results of tests a for the detection of serum breakdown products and ultimately aid the IgE antibodies to benzylpenicillin and amoxicillin. Comparison of results obtained with penicilloyl and selection of an optimal group of minor determi- penicillanyl solid phases nants for diagnostic application. Ratio Test result N Percent (%) positive A/O 5.1.2.5 The Penicillanyl Determinant Sera positive to both 95 7.4 − Reaction with the C-3 carboxyl group of penicil- benzypenicillin and lins produces penicillanyl derivatives, but amoxicillin following administration of the drug, the penicil- Sera positive to 22 1.7 − lanyl determinant formed by covalent interaction benzypenicillin and negative to amoxicillin with a protein carrier (Fig. 5.6 ) is unlikely to Sera negative to 44 3.4 – occur under physiological conditions. This benzypenicillin and almost certainly accounts for the lack of interest positive to amoxicillin shown in the penicillanyl determinant despite a Sera positive to BPO 3 0.2 – small number of promising claims made for its and negative to BPA diagnostic potential. Early immunization studies Sera negative to BPO 55 4.3 55/3 = 18.3 and positive to BPA with laboratory animals revealed that the resul- Sera positive to AmoxO 29 2.2 – tant anti-penicillanyl antibodies did not cross- and negative to AmoxA react with the penicilloyl determinant, but the Sera negative to AmoxO 68 5.3 68/29 = 2.3 antibodies did recognize penicilloyl protein con- and positive to AmoxA jugates when the acyl side chains of the penicil- From Baldo BA. Diagnosis of allergy to penicillins and lanyl immunogen and penicilloyl conjugate were cephalosporins. Structural and immunochemical consid- similar. Hapten inhibition results, including some erations. Allergy Clin Immunol Int. 2000;12:206. Reprinted with permission from © 2000 Hogrefe & Huber with the cephalosporin cephalothin, confi rmed Publishers (now Hogrefe Publishing). http://www. the importance of the side chain structure in hogrefe.com determining specifi city of the antigen, and this BPO benzylpenicilloyl determinant, BPA benzylpenicil- fi nding, together with results showing some anti- lanyl determinant, AmoxO amoxicilloyl determinant, β AmoxA amoxicillanyl determinant body recognition of the -lactam and thiazolidine a 1,290 subjects rings, proved a forerunner for later immunochemical fi ndings with sera from penicillin- allergic patients discussed below in Sect. 5.1.3 . clear indication that the β-lactam ring was pres- Rabbit antibodies to the penicillanyl determinant ent and intact. Recognition by IgE antibodies was bind the parent drug strongly and the determinant signifi cantly greater for the “-anyl” determinants in solid phase form is effective for the detection of benzylpenicillin and amoxicillin than for the of penicillin-reactive IgE antibodies in patients’ “-oyl” determinants with ratios of 18.3 and 2.3 sera (see Sect. 5.1.6.1). In our laboratory, we for benzylpenicillin and amoxicillin, respectively compared penicilloyl and penicillanyl poly-L - (Table 5.2 ). A similar comparison of skin tests lysine and HSA conjugates for this purpose. does not appear to have been done. From results Benzylpenicilloyl and amoxicilloyl conjugates obtained in some of the earliest studies employ- with poly-L -lysine and HSA were prepared by the ing different penicillin determinants in skin tests addition of 0.5 M potassium carbonate at pH 11, on penicillin-allergic patients and reinforced in passage through Sephadex G-25, and dialysis quantitative immunochemical and cellular inves- followed by characterization by the penamaldate tigations, it is clear that the immune response to assay and 1 H NMR spectroscopy. The “-anyl” deter- penicillins is heterogeneous. The penicillanyl minants were prepared with N -hydroxysuccinamide determinant is stable, easy to prepare and charac- and N , N ′-dicyclohexycarbodiimide. To avoid self- terize, and retains the thiazolidine ring, β-lactam condensation, the amino group of amoxicillin was ring, and acyl side chain intact. It thus provides a protected (Boc protection). 1 H NMR spectra gave potentially valuable antigen for studies of the

[email protected] 140 5 β-Lactam Antibiotics allergenic recognition of penicillins at both the humoral and cellular levels. 80

5.1.2.6 Recognition of Penicillin Acyl 70 Side Chains as Allergenic Determinants 60 The side chains (R substituents) of the most important penicillins are shown in Fig. 5.4 . Early 50 immunization studies with laboratory animals showed that the acyl side chain of penicillins 40 elicits the production of complementary antibod- Inhibition (%) ies and there is an extensive literature on the pro- 30 duction of such antibodies and subsequent specifi city investigations undertaken. For our 20 purposes, we are interested in the allergenic contribution, if any, of side chain structures. As long 10 ago as the early 1960s it was appreciated that the side chain of penicillins plays a large part in 0 the specifi city of immunological reactions to the 200 2000 drugs. Side chain antigenicity is easily seen at the Compound (nmol) clinical level in the marked increase in recent years of patients allergic to amoxicillin and/or Fig. 5.7 Example of preferential recognition of a penicil- ampicillin but tolerant of the parent drug. In the lin side chain (R) structure, in this case the 3-thiophene early 1980s it was shown that the addition of moiety of ticarcillin, and of cross-reactivity with other some semisynthetic penicillins such as ampicil- penicillins, by serum IgE antibodies in the serum of a penicillin-allergic patient. Quantitative hapten inhibition lin, ticarcillin, methicillin, and piperacillin to the by β-lactam drugs of IgE binding to a ticarcillin- Sepharose battery of skin testing reagents increased the rate solid phase: ( open inverted triangle ) Ticarcillin; ( fi lled of positive skin tests and, importantly, detected circle) ampicillin; ( fi lled square) phenethicillin; ( open positive reactions to the semisynthetics in some square ) amoxicillin; (open circle ) benzylpenicillin; ( fi lled triangle) piperacillin; ( fi lled diamond) cephalothin. From patients who were skin test negative to benzyl- Harle DG, Baldo BA. Identifi cation of penicillin aller- penicillin. With the marked increase in adminis- genic determinants that bind IgE antibodies in the sera of trations of ampicillin and amoxicillin, subjects with penicillin allergy. Mol Immunol 1990 ; 27: immunologic and provocational evaluations 1063. Reprinted with kind permission from Elsevier Limited revealed increasing numbers of patients responsive only to the semisynthetic penicillins. Although not necessarily refl ective of clinical evidence that side chain groups are the dominant relevance, clear IgE immunologic recognition of allergenic determinant in some immediate aller- some different penicillin side chain substituents gic reactions to penicillins was obtained from was clearly demonstrated in quantitative immu- investigations on a number of patients who nochemical direct binding and inhibition immu- reacted to penicillins with a phenylisoxazolyl R noassays with penicillin-solid phase complexes. substituent. For example, in two patients who For example, sera from some patients showed experienced fl ucloxacillin-induced anaphylaxis preferential recognition of ticarcillin even though confi rmed by obvious clinical features of the other regions of the penicillin structure also reactions, history, skin testing (Fig. 5.8 ), and bound IgE antibodies (Fig. 5.7 ). These results detection of drug-reactive IgE antibodies, quanti- could only be explained by recognition of the tative hapten inhibitions revealed potent IgE anti- ticarcillin R substituent by a population of peni- body reactivity with fl ucloxacillin as well as cillin-reactive IgE antibodies. Further clear-cut pronounced reactivity with three structurally

[email protected] 5.1 Penicillins 141

than 13,200 times as potent an inhibitor as benzylpenicillin, clearly showing overwhelming recognition of the side chain with little or no recognition of the rest of the penicillin molecule. Allergic recognition by some patients of side chain determinants highlights the importance of including different individual penicillins in the battery of penicillin skin test reagents.

5.1.3 Heterogeneity of IgE Antibody Responses to Penicillins and the Spectrum of Penicillin Allergenic Determinants

It was over 40 years ago that Levine pointed out the need for the identifi cation of haptenic determinants of allergenic drugs and although he and numerous other investigators since have helped to place the penicillins near the top, if not at the head, of a list of the best defi ned allergenic determinants on drugs, information on the fi ne Fig. 5.8 Skin test results showing wheals following prick structural detail of allergic recognition of penicil- testing with fl ucloxacillin of a patient who experienced lins remains defi cient. In considering the number anaphylaxis after ingestion of one 500 mg capsule of the penicillin. Positive responses to the drug were obtained in and importance of penicillin allergenic determi- the range 0.125–250 mg/ml and to histamine (H), 10 mg/ nants, two points recognized in the early years of ml. No wheals resulted following prick testing with Pre- research on the drug and its breakdown products Pen (penicilloyl-polylysine, Kremers-Urban, 6 × 10 −5 M are highly relevant. The fi rst is the persisting penicilloyl) and benzylpenicillin at 0.3, 3, 30, and 600 mg/ ml. From Baldo BA et al. Detection and side chain speci- belief that the determinant(s) responsible for the fi city of IgE antibodies to fl ucloxacillin in allergic sub- skin-sensitizing capacity of penicillins is not jects. J Mol Recogn 1995 ; 8: 171. Reprinted with mainly due to the penicilloyl group and the sec- permission from John Wiley and Sons ond is the heterogeneity of the allergic response to penicillins. Early research on the allergenic related penicillins containing a phenylisoxazolyl properties of penicillin, its metabolites, and deg- side chain, viz., oxacillin, cloxacillin, and diclox- radative products was hampered by lack of acillin (Fig. 5.9 ). Analysis of the inhibition knowledge of both the so-called reagins mediat- results showed recognition of the 3-(2-chloro- 6- ing skin and other reactions and their hapten fl uorophenyl)-5-methyl-4-isoxazolyl group of specifi cities. The research effort was conse- fl ucloxacillin by some IgE antibodies and that the quently primarily directed toward the in vitro 5-methyl-3-phenyl-4-isoxazolyl group, with or identifi cation of the main penicillin metabolites without halogen substituents, accounted for the and breakdown products and even when the reactivity of other antibodies and for the strong identifi cation of determinants was pursued via cross-reactions seen with dicloxacillin, cloxacil- antibody recognition studies, it was generally lin, and oxacillin (Figs. 5.9 and 5.10 ). On a molar done with heterologous antisera prepared in labo- basis, and depending on the individual patient, ratory animals. Such antisera almost always dem- the di-halogenated compounds, fl ucloxacillin onstrate heterogeneity of the humoral immune and dicloxacillin, were from about 800 to more response, a potential problem that can often be

[email protected] 142 5 β-Lactam Antibiotics

a 100

40 Inhibition (%)

0 .1 1 10 100 1000 10000 Compound (nmol) b 100

40 Inhibition (%)

0 .1 1 10 100 1000 10000 Compound (nmol)

Fig. 5.9 Demonstration of preferential recognition of patient’s serum. (open circle) Dicloxacillin; (fi lled circle ) side chain (R) groups on isoxazolyl penicillins by IgE fl ucloxacillin; (open square) cloxacillin; ( fi lled square ) antibodies in the sera of two patients who experienced oxacillin; ( open triangle ) benzylpenicillin. From Baldo anaphylaxis to fl ucloxacillin. Quantitative hapten inhibi- BA et al. Detection and side chain specifi city of IgE anti- tion by some β-lactams of the binding of IgE to a fl uclox- bodies to fl ucloxacillin in allergic subjects. J Mol Recogn acillin-solid phase covalent complex: (a ) patient with skin 1995 ; 8: 171. Reprinted with permission from John Wiley test results shown in Fig. 5.8 . ( b) Results with second and Sons overcome by the production of a spectrum of to penicillin determinants have been undertaken monoclonal antibodies. Application of this strat- with the aim of identifying the most important egy to benzylpenicilloyl–protein conjugate delin- allergenic structural features, and for delayed eated three major determinants—the side chain reactions, such studies have been even rarer (as is structure, a compound determinant made up of the case for most drug allergens). It can be argued the amide group on the penicillin molecule that an approach directed at identifying the struc- connected to amino acid residues of the carrier tures recognized by the antibodies mediating the protein, and the thiazolidine ring. immediate allergic reactions is a more direct and In relation to immediate allergic reactions, few clinically relevant one than the potentially more studies employing human sera with IgE antibodies hit-or-miss strategy of fi rst identifying a break-

[email protected] 5.1 Penicillins 143

Space filling model

a Oxacillin

b Cloxacillin

c Dicloxacillin

d Flucloxacillin

Fig. 5.10 Space-fi lling CPK three-dimensional molecu- isoxazolyl penicillins oxacillin, cloxacillin, dicloxacillin, lar models showing the structures and IgE antibody-bind- and fl ucloxacillin (see Fig. 5.9 ). Chlorine atom is green , ing regions (colored blue , green , and orange ) on the fl uorine , orange down product and then accumulating enough of it patients are involved and it is possible to build up to use in tests on allergic patients. An additional a full picture of the spectrum of allergenically risk with the latter approach arises if an allergeni- important structural features recognized in patient cally important metabolite present in only trace responses to the drug. The same general strategy amount remains unidentifi ed. By identifying drug of identifying the determinants via the comple- allergenic structures complementary to combin- mentary immune receptors on cells can be ing sites of IgE antibodies, only the structures rel- employed in cell-mediated responses to drugs. evant to the stimulated allergic responses in Examples of this approach in the investigation of

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T cell- mediated delayed-type hypersensitivity the methodology sometimes detects antibodies with associated HLA alleles, e.g., with abacavir with the capacity to distinguish fi ne structural fea- and carbamazepine, are beginning to accumulate tures on different β-lactam drugs. Good examples (see Sect. 3.4 ). of this are the demonstration of IgE to benzylpeni- When the specifi cities of anti-penicillin IgE cillin that cross-reacted with the cephalosporin antibodies from patients allergic to β-lactam drugs cephalothin (see Sect. 5.2.4.2.2 ) and the detection were fi rst studied, one of the most obvious of serum IgE antibodies in the sera of allergic recorded fi ndings was the marked heterogeneity of patients that distinguished amoxicilloyl and amox- the immune response, a feature often pointed out icillanyl determinants. In the latter study, antibod- by early investigators but seemingly little appreci- ies from a patient who experienced anaphylaxis ated in recent years when we have seen a heavy following an oral dose of amoxicillin reacted only emphasis on clinical aspects and skin testing with with the amoxicilloyl determinant while IgE from the available reagents. IgE antibodies in the sera of a patient with possible penicillin allergy involving patients allergic to β-lactam antibiotics detect a urticaria and angioedema showed multiple reac- spectrum of antigenic specifi cities and IgE in the tivities with penicilloyl and penicillanyl determi- sera of different allergic patients show heteroge- nants of different penicillins but not with the neous recognition and cross-reactive responses. It amoxicilloyl determinant. The explanation for the has been known for many years that some allergic recognition differences shown by the two sera lies patients have more than one population of in the different possible confi gurations of the β-lactam-reactive antibodies in their serum. In amoxicilloyl- and amoxicillanyl-polylysine conju- 1968, evidence was presented for up to eight dif- gates employed as drug-solid phases. Reaction of ferent populations of skin-sensitizing anti-penicil- antibodies with the amoxicilloyl but not the amox- lin antibodies with different binding specifi cities. icillanyl conjugate refl ected antibody recognition Quantitative hapten inhibition investigations of both ends of the amoxicilloyl molecule, that is, employing sera from penicillin-allergic patients in with the aminobenzyl portion of the side chain radioimmunoassay experiments with semisyn- (and perhaps with little or no recognition of the thetic penicillins, the parent molecule, and a range attached ring hydroxyl) and the thiazolide ring. of carefully selected structural analogs often reveal These antibodies could not be detected with the antibody cross- reactivity and recognition of more amoxicillanyl conjugate formed by coupling than one structural domain on penicillin mole- through the thiazoline ring carboxyl group cules. Results obtained with the semisynthetic (Fig. 5.12 ). Reaction of the antibody from the sec- ampicillin illustrate the point. Some antibodies ond patient with the amoxicillanyl but not the recognized discrete regions of the ampicillin mol- amoxicilloyl conjugate refl ected clear and strong ecule such as the side chain only or the thiazoli- antibody specifi city for the aminohydroxybenzyl dine ring only while others were shown to have side chain, and especially for the 4-hydroxy sub- combining sites complementary to compound stituent, which is accessible for binding in the structures made up of the side chain with the “-anyl” but not the “-oyl” conjugate form. With the β-lactam ring, the combination of the β-lactam and amoxicilloyl conjugate where linkage of the drug thiazolidine rings, or the whole molecule is through the open β-lactam ring, rotation and (Fig. 5.11 ). As well as identifying a spectrum of fl exibility around C-6 and C-7 allow the possibil- complementary antibody combining sites recog- ity of close steric association between the side nizing “broad” combinations of groups of atoms chain and the peptide carrier (Fig. 5.12 ). Such such as ring structures or even the entire molecule, close association creates the possibility for

Fig. 5.11 (continued) allergic sera recognize the whole sporins. Allergy Clin Immunol Int 2000; 12: 206. Reprinted ampicillin molecule rather than parts of the structure. From with permission from © 2000 Hogrefe & Huber Publishers Baldo BA. Diagnosis of allergy to penicillins and cephalo- (now Hogrefe Publishing. http://www.hogrefe.com )

[email protected] HH S CH CONH CH3

NH2 N CH3 O H COOH Ampicillin

HH S CH CONH CH3

NH2 N CH3 O H COOH Aminobenzyl side chain HH S CH CONH CH3 NH 2 N CH3 O H COOH Side chain and β -lactam ring

H H S CH CONH CH3

NH2 N CH3 O H COOH Thiazolidine ring

HH S CH CONH CH3

NH2 N CH3 O H COOH β-lactam and thiazolidine rings

HH S CH CONH CH3

NH2 N CH3 O H COOH Whole molecule

Fig. 5.11 Two-dimensional structures and three- molecule complementary to combining sites of ampicillin- dimensional CPK models showing the spectrum of aller- reactive IgE antibodies in the sera of patients allergic to the genic determinants on ampicillin. Regions on the ampicillin aminopenicillin are highlighted . Antibodies in some

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Amoxicillanyl-Lysine conjugate Side-chain Side-chain

-OH

Lysine Lysine

Amoxicilloyl-Lysine conjugate Side-chain Side-chain

-OH

C-6

C-7

Lysine Lysine

Side-chain Side-chain

-OH C-6

C-7

Lysine Lysine

Fig. 5.12 Models (ball-and-stick, left-hand side , CPK β-lactam ring confers rigidity on the molecule with the space-ﬁ lling, right-hand side) showing some possible linked peptide at C-2 (shown here attached to a sin- conﬁ gurations of amoxicillanyl- and amoxicilloyl-lysine gle lysine) below the plane (P), and the C–NH bond at conjugates. In the amoxicillanyl form, the still intact C-6 above the plane, making close association of the

[email protected] 5.1 Penicillins 147

H-bonding involving the side chain hydroxyl detected no difference. However, a higher fre- group and this results in hindered access of quency of atopy has been shown in patients who antibody to this structure. With this patient, diag- had a fatal reaction to penicillin so atopic indi- nostic employment of the penicilloyl specifi city viduals who are also allergic to penicillins may only would have produced an erroneous picture of be at increased risk of anaphylaxis to the drug. the patient’s true fi ne structural recognition Data clearly shows that more allergic reactions to sensitivity. penicillins occur following parenteral than after Of course, the question of the clinical oral administration and this is in keeping with the relevance of antibody responses to drugs detected well-known facts that anaphylaxis is a key risk of in vitro is pertinent to any discussion of the appli- peripheral IV therapy and that the parenteral cation and results of drug-specifi c IgE tests for route of administration increases the severity and the diagnosis of drug allergies. The detection of frequency of an anaphylactic reaction. The risk is penicillin- reactive IgE antibodies may prove rec- also higher for patients with histories of anaphy- ognition and even sensitization to a β-lactam laxis and urticaria compared to those with vague, structure(s) but not necessarily the existence in mild, or unknown histories of penicillin reac- the patient of allergy as a clinical disease. tions. However, allergic reactions to drugs on fi rst However, while the presence of a population of exposure are known with the frequency of this drug-reactive IgE antibodies does not guarantee occurrence varying between different groups of type I allergic sensitivity, such sensitivity does drugs—for example, reactions on fi rst exposure not seem to occur in the absence of IgE antibod- are commonly seen with neuromuscular blocking ies (but see Sects. 3.2.7 and 5.2.4.3.2 ). drugs, but there are no reports of reactions after fi rst contact with the induction agent thiopentone. Reactions provoked by the fi rst dose of a 5.1.4 Risk Factors for Immediate penicillin occur, but the question of prior expo- (Type I) Reactions to sure and its possible contribution to sensitization Penicillins is a diffi cult one to resolve given that penicillins have been found in milk, meats, other foodstuffs, 5.1.4.1 General Risk Factors human breast milk, and other environmental Young and middle-aged adults appear to be at sources. Patients with a history of prior reactions greatest risk of acute allergic reactions to penicil- to penicillins have a four- to sixfold increased lins, although the elderly may not cope as well risk of a reaction to penicillin compared to those with a reaction due to a generally poorer state of without a previous history to the drugs. In consid- health and children may generally have a lower ering penicillin exposure and risk, the persistence cumulative exposure to the antibiotic. It also of IgE antibodies to the drug is another poten- appears that IgE-based sensitivity may wane tially important factor. Penicillin- reactive anti- quicker in children with one study showing a bodies in human sera have been shown to have 33 % reduction in skin test positivity to penicillin half-lives from as little as 10 days to many years, 1 year after initial testing. While some studies suggesting that their disappearance is not simply have demonstrated a higher frequency of positive a consequence of IgE catabolism (see also skin tests in atopic individuals, others have Sect. 5.1.5.3.5 ).

Fig. 5.12 (continued) hydroxyaminobenzyl side chain close proximity between the hydroxy group on the side group and the peptide residues impossible. With the chain and the peptide residues. This close association per- amoxicilloyl determinant, however, opening the β-lactam mits H-bonding and, as a consequence, access of antibod- ring allows increased fl exibility and rotation about C-6 ies to the side chain of amoxicillin is hindered. From Zhao and C-7 and the resultant possibility of close association Z et al. β-Lactam drug allergens: fi ne structural recogni- between the side chain and the peptide carrier linked at tion patterns of cephalosporin-reactive IgE antibodies. C-7. Two possible confi gurations of the amoxicilloyl J Mol Recogn 2001 ; 14: 300. Reprinted with permission determinant are shown; the lower one demonstrates the from John Wiley and Sons

[email protected] 148 5 β-Lactam Antibiotics

5.1.4.2 Risk Factors Associated minor determinants, potassium benzylpenicillin, with Testing sodium benzylpenicilloate, and sodium benzyl- As might be expected, the risk of sensitization penilloate (and sometimes benzylpenicilloyl- and a systemic reaction is lower with the prick amine) were each used at a concentration of test than with intradermal testing. In general, the 10 −2 M. The maximum test concentration of the risk of a skin test-induced systemic reaction is benzylpenicilloyl-polylysine reagent was later rare, but it cannot be excluded especially in some adjusted to 6 × 10 −5 M in the USA and 5 × 10 −5 M highly susceptible subjects, for example, subjects in Europe. The maximum concentration for the with a previous history of anaphylaxis, uncon- minor determinants in Europe was changed to trolled asthmatics, pregnant women, and small 2 × 10−2 M; the minor determinant mix is not children. The rate of systemic reactions induced available in the USA where benzylpenicillin is by penicillin skin testing is said to be about 1 % used alone at a maximum concentration of (see also Sect. 5.1.5.3.2 ). This presumably refers 10,000 IU/ml. Note that these concentrations are to patients with a previous penicillin-induced the same for both prick and intradermal testing. reaction. Sullivan found none of 83 skin test- Originally sold as a research reagent marked “not negative patients given a β-lactam immediately for human use,” the major determinant was mar- after skin testing experienced an allergic reac- keted in the USA in 1973 after a large-scale tion. Although the risk appears to be small, skin cooperative skin testing prospective study spon- testing and challenge testing can each induce sored by the American Academy of Allergy. In resensitization to penicillins. This is so even Europe, penicilloyl-polylysine was fi rst produced though low concentrations of drugs are used in and distributed as a research reagent by the the tests. In fact, sensitization is believed to have Institute of Clinical Immunology, University of resulted from even lower concentrations of peni- Berne, Switzerland, before being registered in cillins in the environment. In one recent investi- France in 1974. Penicilloyl-polylysine (Pre- gation of over 300 cases, 2.5 % of skin Pen® ) was withdrawn from the market in the test-negative subjects became skin test positive USA from September 2000 to November 2001 after testing with benzylpenicillin, penicilloyl- and then again from September 2004 to polylysine, and minor determinant mix. September 18, 2009, when full regulatory approval was granted by the FDA. Production of this reagent and minor determinant mix ceased in 5.1.5 Skin Testing Today for Europe in 2005 but was replaced by a new com- Immediate Hypersensitivity mercial kit containing both the major determi- to Penicillin nant (5 × 10−5 M) and minor determinant mix (each component at a maximum concentration of 5.1.5.1 Historical Perspective 2 × 10 −2 M). In September 2011, Pre-Pen ® was Skin testing for allergic sensitivity to penicillins approved by Health Canada. In July 2011, an has not proved to be free of problems with the agreement with global distribution rights was practice being beset by diffi culties of regulatory reached for marketing the major determinant requirements, suitability of testing reagents, and together with a minor determinant mix currently interrupted supply. As outlined above, the favored under development in the USA. During the peri- skin testing reagents have their origins in the ods of lack of supply, some clinical and labora- early research that identifi ed penicillin metabo- tory investigators produced their own major and lites and breakdown products some of which minor skin test preparations. With the increasing were introduced and used for diagnostic testing. usage of some semisynthetic penicillins, in par- From the earliest introductions, the major deter- ticular amoxicillin and ampicillin, the increase in minant, benzylpenicilloyl-polylysine, fi rst devel- numbers of allergic responses to these drugs, and oped in 1961, was employed for skin testing at a the realization of the allergenic importance of maximum concentration of 10−6 M and later, the side chain structures, amoxicillin and ampicillin

[email protected] 5.1 Penicillins 149 are now often included in the standard battery of history of penicillin allergy have a positive skin skin tests. The maximum test concentration for test to either the major determinant or the minor these drugs is usually 20 mg/ml for prick and determinant mix. Overall though, responses to intradermal testing. Note, however, that patterns penicilloyl-polylysine alone or together with the of usage of amoxicillin and ampicillin, and con- response to the minor determinant mix were pos- sequently the numbers of allergic reactions, vary itive in more than 50 % of the patients. For those between different countries. Now, after many with a negative history of penicillin allergy, the years of uncertainty and on–off accessibility for incidence of positive skin tests is 2–7 %. For skin testing reagents, the immediate future of diagnos- test-positive patients given a therapeutic dose of tic skin testing for allergy to penicillin looks penicillin, the risk of an acute allergic reaction more secure than at any time in the past. ranges from 10 % in patients with a negative history to 50–70 % in patients with a positive his- 5.1.5.2 Experience So Far of Skin tory. Reactions occur rarely in patients with a Testing with Penicillin Test negative skin test (1–4 % in one study) and any Reagents reactions tend to be mild and self-limiting. The In the cooperative prospective skin testing study possibility of a life-threatening reaction is said to of 3,000 subjects (1,718 with symptoms of peni- be almost negligible and any β-lactam can be cillin allergy) sponsored by the American safely given. Severe allergic reactions to penicil- Academy of Allergy in 1977, 19 % of the cases lins, such as anaphylaxis, do not appear to have proved positive to benzylpenicilloyl-polylysine been reported in skin test-negative patients. With and/or benzylpenicillin. Fifty four percent were increasing prescribing of semisynthetic penicil- positive to the major determinant only, 22 % to lins over more recent years, many clinicians have benzylpenicillin, and 25 % to both reagents. It supplemented their panel of tests with these was in this study that the currently used concen- drugs, particularly amoxicillin. For example, trations of test antigens were established. amoxicillin minor determinants (amoxicillin, Addition of penicilloic acid to the panel of test amoxicilloic acid, and a derivative of diketopi- reagents in a study of 740 patients, 63 % of whom perazine formed from aminolysis of the parent had a positive skin test to at least one of the molecules and containing the hydroxyphenyl and reagents, revealed positive responses of 21 % to thiazolidine rings) have been used on patients the major determinant, 42 % to the mixture of with immediate hypersensitivity to amoxicillin. minor determinants, and 45.2 % to the major plus There is some data indicating that skin test sensi- the minor reagents. Some subjects were positive tivity to amoxicillin may not persist as long as the only to benzylpenicillin or penicilloic acid with skin test response to benzylpenicillin determi- 14.6 % of cases in the latter group. Addition of nants. In a 5-year follow-up investigation of ampicillin to the test panel provided no additional cases, 40 % of the benzylpenicillin-sensitive information since all ampicillin-positive patients group became skin test negative whereas all of also reacted to benzylpenicillin. These studies, the patients with side chain sensitivity to amoxi- together with many more investigations (includ- cillin became negative. These reagents did not ing some with large numbers of patients, e.g., increase the number of patients with positive 5,063 subjects, 776 of whom had a history of reactions to the drug. Even with increases in the penicillin allergy), revealed signiﬁ cant variations administrations and allergic responses to the in responses to the major and minor determi- semisynthetic penicillins, some skin test studies nants. Skin testing with only the major determi- have detected a signiﬁ cant number of patients nant is said to identify up to 97 % of allergic positive to only benzylpenicilloyl-polylysine patients and testing without inclusion of the and/or benzylpenicillin minor determinant mix. minor determinants misses from 3 to 10 % of A recent retrospective study of over 800 patients patients. From data assembled by Weiss and consulting for possible allergy to a β-lactam drug Adkinson, 7–63 % of patients with a positive revealed that the employment of these two test

[email protected] 150 5 β-Lactam Antibiotics reagents also detected an additional 27.6–32 % of episode of anaphylaxis, skin tests may be nega- positive reactions in patients allergic to other tive for up to 2 weeks or even longer. This can β-lactams. have important consequences if the culprit drug was not identifi ed. For patients who test negative 5.1.5.3 Some Important Aspects of after an anaphylactic episode, skin testing should Skin Testing for Penicillin- be repeated after 3–6 weeks. Allergic Sensitivity For a detailed description of the background, ratio- 5.1.5.3.3 Sensitivity and Specifi city nale and methodology of skin testing with drugs, of Skin Testing and for the interpretation and reading of results, the To discriminate true allergic from nonallergic reader is referred to Sect. 4.2 . Prick testing should responses, the drug provocation test is normally be done fi rst and should be followed by intradermal used but, with penicillins, the risk of challenging testing only if the prick test is negative. a patient with both a positive history and skin test is generally considered to be unacceptable. This, 5.1.5.3.1 Indications for Skin Testing of course, makes the determination of specifi city It seems prudent to skin test all patients with a and sensitivity of skin tests diffi cult. Although history of allergy to a penicillin if, at the time, skin tests to penicilloyl-polylysine have been con- penicillin remains the indicated drug of choice. sidered to be positive in up to 70 % of patients Skin testing should be carried out immediately with immediate type I responses to penicillin, before administration of the drug and repeated testing of 290 patients with a history of immediate before any subsequent courses. Skin testing with allergic reactivity to penicillin (71 % anaphylaxis, penicillins or any other β-lactam is absolutely 29 % urticaria) revealed skin test sensitivities of contraindicated in patients with a history of 22 % for the benzylpenicilloyl hapten, 21 % for Stevens–Johnson or Lyell’s syndrome (toxic minor determinant mix, 43 % for amoxicillin, and epidermal necrolysis), exfoliative dermatitis, or 33 % for ampicillin. Skin test positivity to at least other reactions where β-lactam drug administra- one determinant occurred in 70 % of the patients, tion is contraindicated. showing that 30 % of patients could be misdiagnosed without further diagnostic investigation. 5.1.5.3.2 Safety These results are not reassuring since even with Serious reactions and even death have been the employment of four different determinants, reported following skin testing with penicillin skin test sensitivity was a long way short of ideal. reagents, but if the test is done properly and A second unexpected and worrying fi nding was potential dangers (such as those that might be the number of patients with a negative skin test apparent in the patient’s history) are anticipated, but a positive drug provocation test. This does not skin testing is generally a safe procedure with a fi t with the currently accepted belief that the pos- systemic reaction rate of about 1 % or less. Most sibility of reacting to a penicillin is negligible in systemic reactions that do occur are mild. About subjects with negative skin tests to the major and 2–7 % of patients with no history of reactions to minor determinants. To establish the specifi city of a penicillin show a positive skin test and most skin testing for penicillin sensitivity, results from penicillin-induced anaphylactic deaths occur in tests on subjects with known tolerance to the drug patients with no apparent history of a reaction to must be obtained. When this is done, specifi city is the drug. Severe reactions have occurred to generally good and in the range 97–99 %. higher than recommended test concentrations or after intracutaneous testing without fi rst doing a 5.1.5.3.4 Reading Tests prick test. Skin testing should be done in the pres- Results are read 15–20 min after completing the ence of a physician capable of managing anaphy- skin test. A 3 mm wheal accompanied by ery- laxis and with ready access to the appropriate thema with a negative response to a saline control medications and equipment. Importantly, after an is generally taken as the threshold for a positive

[email protected] 5.1 Penicillins 151 prick test while a positive intradermal test result diagnostic tests for penicillin-induced type I is often considered to be an increase in wheal allergic reactions are essentially the same as the size (accompanied by erythema) of 3 mm or tests employed for the diagnosis of immediate more over the diameter of the bleb size formed hypersensitivities to other drugs. These tests are following injection (usually about 2 mm). presented in detail in Chap. 4 and this informa- A positive intradermal test result is therefore usu- tion should be referred to before proceeding with ally a wheal with a diameter of around 5 mm or this section. greater surrounded by erythema. Patients should be advised of the possibility of a late reaction. 5.1.6.1 Detection of Penicillin- A positive late reaction to intradermal testing Reactive IgE Antibodies may manifest as erythema, papulation, infi ltrate, While it is clear that the prime choice of tests for eczema, and swelling. Any infi ltrated erythema the diagnosis of penicillin-induced immediate with a diameter greater than 5 mm should be con- reactions is the skin test, co-employment of sidered a positive reaction. serum IgE tests for penicillin-reactive IgE antibodies is advisable since some cases of positive 5.1.5.3.5 Persistence of Skin Test IgE tests have been seen in patients with a history Reactivity to Penicillins of an immediate reaction to a penicillin but a Skin test reactivity to penicillins generally negative skin test to the drugs. decreases with time. Testing has shown that skin Soon after the development in 1967 of the tests carried out within 1–2 months of an acute radioallergosorbent test, known as the RAST, for allergic reaction to penicillin were positive the in vitro detection of allergen-reactive IgE 80–90 % of the time, but this was followed by a antibodies, Wide and Juhlin in Sweden applied time-dependent decline—in one study, positive the test to the sera of penicillin-allergic patients reactions to penicillins persisted for 7–12 months using solid phases prepared from benzylpenicil- in 93 % of subjects. In another early study, loyl and phenoxymethylpenicilloyl protein con- Sullivan and coworkers found a positive response jugates. IgE antibodies to the penicilloyl in 73 % of patients within 1 year, 57 % continued determinants were found in 9 of 11 patients and to show a positive reaction between 1 and 10 results from skin tests and RAST reactions agreed years, and there were still 22 % of positive reac- for positive and negative reactors. Subsequent tors after 10 years. The chance of a positive skin early applications of penicillin RASTs revealed test response therefore appears to decrease by rare positive reactions to penicillamine, cross- about 10 % per year, meaning that about half the reactivity between penicillin minor determinants patients who had an immediate reaction to peni- and the major determinant, and the fi nding that cillin will be skin test negative after 5 years. the penicillanyl determinant yielded no more Long-lasting IgE antibody formation to penicil- information than the penicilloyl determinant lins often occurs in patients who have had (compare Sect. 5.1.2.5 ). In perhaps the most penicillin- induced serum sickness reactions. informative of the early applications of the RAST to penicillin allergy, Dewdney’s group prepared and examined thiol-linked penicillamine, benzyl- 5.1.6 In Vitro Tests for Immediate penicillenic acid and the benzylpenicillanyl Hypersensitivity to Penicillins determinant. These reagents essentially confi rmed the importance of the penicilloyl determi- As with other drugs, but perhaps more so, a vari- nant, but, most importantly, the study also ety of humoral and cellular investigations have confi rmed that the heterogeneity of the immune been utilized over many years with the aim of response to penicillins extends to the specifi city aiding the diagnosis and elucidating underlying of the serum IgE antibodies. mechanisms of penicillin hypersensitivities. In more recent years, a number of laboratories The most commonly and widely used in vitro have developed and applied their own in-house

[email protected] 152 5 β-Lactam Antibiotics immunoassays to detect serum IgE antibodies to undertaken in the authors’ laboratory in 2001 the parent drug and some semisynthetic penicil- when, using both penicilloyl and penicillanyl lins, principally amoxicillin and ampicillin. derivatives of benzylpenicillin and amoxicillin, Perhaps the best known commercially available sensitivities for the detection of benzylpenicillin- test reagents for detecting penicillin-reactive IgE and amoxicillin-reactive IgE antibodies in the antibodies are the Phadia ImmunoCAP® (Thermo sera of 28 patients with diagnosed immediate Scientifi c) drug-solid phases for penicilloyl G hypersensitivity reactions to a β-lactam were and V, amoxicilloyl, and ampicilloyl determi- 57.1 % and 78.6 %, respectively, while the cor- nants which are widely distributed. These assays responding fi gures for the ImmunoCAP assays measure specifi c IgE antibodies in the range were 35.7 % and 28.6 %. Once again, however, 0.01–100 kUA/l with a cutoff value of 0.35 kUA/l specifi cities of 80.7–87.3 % for the laboratory for a positive result and levels higher than tests were less than the results of 86.3 and 98.2 % 0.1 kUA/l , indicating sensitization to the drug. obtained with the commercial assays. Clearly, One assessment of the performance of the ben- improvements in the IgE antibody in vitro assays zylpenicilloyl and amoxicilloyl ImmunoCAP are needed, especially in regard to sensitivities of assays using sera from patients with positive skin the tests for different penicillins. tests to amoxicillin and/or what was described as “other benzylpenicillin-derived agents” revealed 5.1.6.2 CAST-ELISA ® and Flow-CAST® sensitivity of 54 % with a specifi city of 95–100 %. Note that as a diagnostic test for β-lactam allergy, While the sensitivity of the amoxicilloyl serum IgE determinations are claimed to be less ImmunoCAP assay in tests on 29 sera from sensitive than the cellular allergy stimulation test patients skin test positive to amoxicillin but nega- (CAST® ) (Buhlmann Laboratories AG) which tive to benzylpenicilloyl-polylysine and minor measures the release of cysteinyl leukotrienes determinant mix was 41 % and 42 % of 26 skin from peripheral blood leukocytes following aller- test negative, provocation test-positive patients gen challenge (see Sect. 4.5.3 ). In a multicenter were positive in the immunoassay, showing that study of 181 patients with a history of immediate the provocations could have been avoided by hypersensitivity to a β-lactam, overall sensitivity doing the IgE test. In another IgE examination of with the CAST-ELISA® in skin test-positive sera from 58 patients who each experienced an patients was 41.7 % and 27.9 % in skin test-nega- immediate reaction to a β-lactam and had a positive patients. When these results were considered tive skin test to at least one of benzylpenicillin, together with Flow-CAST® (Sects. 4.5.3.1 and benzylpenicilloyl-polylysine, minor determinant 4.5.3.2 ) results, diagnostic sensitivity increased to mix, amoxicillin, ampicillin, and cephalosporins, 64.3 % with a specifi city for both tests combined the sensitivity and specifi city of the same reagents of 73–92 %. Sensitivity of specifi c IgE determina- were found to be only 37.9 and 86.7 %, respec- tions in the same population was 28.3 %, a fi gure tively. A similar study some years earlier on which seems extraordinarily low. Individual spec- patients with immediate reactions and positive ifi cities for specifi c IgE determinations, CAST- skin tests detected penicillin-reactive serum IgE ELISA® , and Flow-CAST ® were claimed to be antibodies in 37 % of the patients. As pointed out 86.5 %, 78.7 %, and 88.9 %, respectively. In the by the Blanca group, immunoassay sensitivities, multicenter study, a maximum sensitivity of but not necessarily specifi cities, for penicillins 85–90 % was reached in 112 of 124 patients with developed in individual laboratories can compare a history of allergy to amoxicillin when all four favorably with the commercial assay with one tests, skin tests, serum β-lactam-reactive IgE comparison showing specifi cities and sensitivi- assays, Flow-CAST® , and CAST-ELISA® were ties of 83.3–100 % and 12.5–24 %, respectively, applied in that order to patients with a negative for the commercial assay and 66.7–83.3 % and reaction to the previous test. On the downside, 42.9–75 %, respectively, for the laboratory test. however, the increase in sensitivity was matched These fi gures are similar to a comparison by a decrease in specifi city. Even then, the eight

[email protected] 5.1 Penicillins 153 negative patients responded positively to a con- Challenges should be performed only after trolled challenge. Although it was claimed that prior skin testing and preferably a drug-specifi c application of so many tests can cut down the IgE antibody test. If either of these tests returns a number of challenges and therefore reduce costs positive result that is in accordance with the and patient discomfort, one wonders about the patient’s history, the risk precludes provocation practicality of the routine extra investigations testing. According to the ENDA (European let alone the extra economic cost to so many Network for Drug Allergy) guidelines, in the fi rst patients and the national health care bill. A more instance, skin and IgE testing should be under- logical, potentially effective, and sensible taken with benzylpenicillin, and, if this is posi- approach may be to put greater effort into tive, the patient should be considered to be researching β-lactam allergenic determinants with allergic to the β-lactam group of drugs. If testing the aim of defi ning a more optimal set of determi- with the parent penicillin is negative, the patient nants that will increase the sensitivities of the rou- is then tested with the drug that provoked the tine skin and serum IgE antibody diagnostic tests. reaction if it is known. A positive reaction to a known drug confi rms selective allergy to the 5.1.6.3 The Basophil Activation Test drug. When the drug is not known, and in the in the Diagnosis of Penicillin case of a negative reaction to a known drug, a Immediate Hypersensitivity diagnosis cannot be made and further testing There have been at least fi ve studies of the perfor- should then proceed beginning with an aminope- mance of the basophil activation test in the diag- nicillin such as ampicillin or amoxicillin. nosis of immediate hypersensitivity to β-lactams. Figure 5.2a, b shows an example of a rash on a All of the studies demonstrated a sensitivity of patient’s neck, arms, and hands that developed about 50 % with specifi city in the range of ~90– after the last oral challenge dose of amoxicillin. 100 %, although sensitivities as high as 67 % and The patient had previously tested negative to the as low as 20 % were seen in an investigation in penicillin in the intradermal test. Challenges with which the basophil activation markers CD63 and drug and a placebo are performed in a single CD203c were compared (see Sect. 4.6.2 ) in the blind procedure by a physician able to manage diagnosis of amoxicillin allergy. Amoxicillin anaphylaxis preferably in an intensive care set- induced upregulation of CD203c in 60 %, or 12 ting in a hospital environment with all the neces- of 20 anaphylactic patients skin test positive to sary resuscitation facilities and medications amoxicillin, but upregulation of CD63 was sig- available to handle a possible emergency (see nifi cantly lower at 20 % (4 out of 20 patients). Sect. 4.4 ). Provocation testing of patients with Somewhat surprisingly, upregulation of CD203c exfoliative dermatitis, Stevens–Johnson syn- and CD63 by ampicillin was more than amoxicil- drome, or Lyell’s syndrome is contraindicated. lin, occurring in 67 % (8 of only 12) and 33 % Intervals between increasing doses of the drug (4 of 12), respectively, of the anaphylactic should be at least 30–60 min and progress to the patients. Also disconcerting was the report of ten next increase should not occur before each dose is false positives, confi rmed by negative provoca- clearly judged to be well tolerated. Any dose that tion tests, with both markers. causes systemic responses even if they are mild such as rhinitis, redness or pruritus should be repeated until tolerance is demonstrated. 5.1.7 Challenge (Provocation) Administration of an antihistamine is usually Testing for Penicillin enough to control the symptoms of these reac- Hypersensitivity tions. Any more severe reaction that looks like an allergic reaction such as swelling of the throat, This section should be read in conjunction wheezing, or a drop in blood pressure should with the discussion of challenge testing set out in be treated with appropriate measures including Sect. 4.4 . epinephrine, steroids, bronchodilators, etc.

[email protected] 154 5 β-Lactam Antibiotics

Table 5.3 ENDA protocols for penicillin parenteral and oral provocation tests Cumulative Route of Drug Dose dose administration Benzylpenicillin 103 IU/ml 103 IU/ml IM 10 4 IU/ml 1.1 × 104 IU/ml IM 10 5 IU/ml 1.1 × 105 IU/ml IM 5 × 10 5 IU/ml 6.1 × 105 IU/mla IM Phenoxymethylpenicillin 1 mgb 1 mg Oral and amoxicillin 5 mg b 6 mg Oral 50 mg c 56 mg Oral 100 mg d 156 mg Oral 250 mg e 406 mg Oral 400 mg f 806 mgg Oral Interval between doses 30–60 min 1 IU penicillin = 0.6 μg Cumulative dose needs to be adapted to children and patients with kidney or liver disease ENDA European Network for Drug Allergy, IM intramuscular a Cumulative dose should be no more than 106 IU/ml b Normally 1–5 mg but 0.1–5 mg for patients with history of a severe reaction c 50–65 mg d 100–150 mg e 250–300 mg f 400–800 mg g Cumulative dose should be no more than 1,000 mg

Such responses are interpreted as a positive aller- ability, bacteriostatic or bactericidal action. In gic reaction and the challenge is discontinued. such cases, the risk of infection may outweigh Table 5.3 sets out the recommended ENDA pro- the desensitization risks. Protocols using both the tocol for penicillin drug provocation testing with parenteral and oral routes have been developed the parent penicillin given parenterally and for penicillin desensitization. Desensitization can penicillin V and amoxicillin administered orally. be achieved safely by the former route, but oral Suggested dose ranges for each step and maxi- challenges have caused fewer severe reactions mum cumulative doses are shown. If the reaction and are generally considered to be safer. Again, to a penicillin is not an immediate type I hyper- patients with a history of exfoliative dermatitis or sensitivity, reactions following dosages may Stevens–Johnson or Lyell’s syndromes should occur with intervals of hours or days and this not be subjected to desensitization and the proce- must be considered before proceeding with the dure should be carried out in an intensive care next challenge step. setting with an IV line set up, β-adrenergic antagonists discontinued, and blood pressure, pulse, and respiratory rate recorded after each dose. 5.1.8 Penicillin Desensitization Note that Castells and coworkers have recommended that in addition to the exfoliative skin This section should be read in conjunction with reactions mentioned above, patients with other the presentations on desensitization in Sect. 3.5 . reactions including maculopapular rashes, ﬁ xed Although there are risks associated with drug eruptions, bullous erythema, drug reaction desensitization to a drug, a patient may, for with eosinophilia and systemic symptoms example, show drug resistance to a possible alter- (DRESS), transaminitis, acute interstitial nephri- native antibiotic and there may also be the possi- tis, serum sickness, hemolytic anemia, thrombo- bility of failure to control an infection by cytopenia, or neutropenia should not be subjected substituting a drug that provides poorer bioavail- to rapid IV desensitization. Before beginning a

[email protected] 5.1 Penicillins 155 desensitization procedure, the patient’s risk situa- Table 5.4 Rapid oral desensitization protocola for tion should be assessed and the history should be patients with a positive skin test to penicillin(s) consistent with a mast cell-IgE-mediated Stepb Dose (mgc ) Cumulative dose (mgc ) response. Penicillin skin testing with the major 1 0.03 0.03 and minor determinants has a high negative pre- 2 0.06 0.09 dictive value and these tests can be particularly 3 0.12 0.21 useful for patients with uncertain histories. 4 0.24 0.45 Patients with a negative skin test are usually not 5 0.50 0.95 candidates for desensitization while those with 6 1 1.95 positive tests are advised to avoid penicillins and 7 2 3.95 cephalosporins, especially the fi rst-generation 8 4 7.95 9 8 15.95 drugs. Desensitization should be considered for 10 16 31.95 the skin test-positive patients if administration of 11 32 63.95 these antibiotics is judged to be necessary. 12 64 127.95 Protocols developed in the USA by Sullivan 13 125 252.95 and his collaborators have been successfully used 14 250 502.95 and adapted for many years. In one demonstra- Patient should be observed for 2 h after last dose tion of the utility of the oral procedure using a For example, for benzylpenicillin or phenoxymethylpenicillin, 15 pregnant women, phenoxymethylpenicillin most infected with syphilis, were both desensi- b 15 min interval between steps cDoses obtained from freshly made solutions of (e.g.,) tized and cured of their infections. Reactions dur- concentrations 1 mg/ml (doses 1–7) and 100 mg/ml (doses ing the desensitization process and subsequent 8–14) therapy were confi ned to the skin and were not serious. The starting dose for desensitization is commonly about one ten-thousandth or less of a should be repeated until the patient tolerates the full therapeutic dose. Using penicillin in solid dose without adverse signs or symptoms. form and starting with a dose of 0.05 mg, Sullivan Reactions that are more serious such as hypoten- employed doubling doses at 15 min intervals in sion, asthma, or laryngeal edema need appropri- 14 steps up to a maximum dose of 400 mg and a ate treatment but, if the decision is made to cumulative dose of 800 mg before administering continue with the desensitization procedure, the the full therapeutic dose of the drug 30 min after patient should fi rst be stabilized before dosage is the last desensitizing dose. An example of a pro- continued with one-tenth the amount per dose. tocol for a rapid oral desensitization procedure is Skin test responses to penicillins diminish with shown in Table 5.4 . For desensitization via the IV desensitization and may become negative. The lit- route, doubling doses, starting with an initial erature contains many case reports and a few case dose of 0.01 mg, are administered at 15 min series on rapid desensitization to antibiotics in intervals in 17 steps until a maximum dose of cystic fi brosis patients where infections by 640 mg and a cumulative dose of 1,280 mg are Pseudomonas aeruginosa and antibiotic allergies reached. Again, the full therapeutic dose is are a common problem often requiring desensiti- administered 30 min after the fi nal dose. For a zation of the infected patients. These studies col- patient to remain in the desensitized state, peni- lectively provide important information on the cillin dosage will normally need to be main- feasibility, performance, and safety of β-lactam tained, often on a twice daily schedule. If antibiotics in a population of high-risk patients penicillin is discontinued for 48 h or more, it is with poor lung function. Rates of successful highly likely that desensitization will have to be desensitization ranged from 58 to 100 %. It should repeated. be emphasized that most patients require ongoing During the stepwise dosage procedure, any courses of desensitization with time. In desensiti- dose that provokes even a mild systemic reaction zations carried out in the Drug Desensitization

[email protected] 156 5 β-Lactam Antibiotics

Unit, Brigham and Women’s Hospital, Boston, 15 β-lactam drugs. Skin prick and patch tests on a patients completed 52 desensitizations, seven of large number of patients with a history of sus- which involved reactions. Six patients had limited pected cutaneous adverse drug reactions detected symptoms of immediate hypersensitivity and one 89 positive patch tests (10.8 %), mainly to patient experienced acute respiratory failure to β-lactams, trimethoprim, and clindamycin, in ceftazidime. Successful desensitizations were 829 patients and 10 positive prick tests (1.1 %) in obtained with benzylpenicillin, nafcillin, cefazo- 935 patients. Eight of 298 patients (2.7 %) were lin, and ceftriaxone. patch test positive to phenoxymethylpenicillin. Challenge tests on 17 patients who were skin test positive and 229 who were skin test negative pro- 5.1.9 Delayed-Type duced 14 and 22 positive results, respectively. Of Hypersensitivity Reactions the 22 (9.6 %) skin test-negative and challenge to Penicillins test-positive patients, 12 reacted with exanthema, seven with urticaria, and three with fi xed drug Delayed-type, non-IgE-mediated hypersensitiv- eruptions. In a study designed to assess the inci- ity, often manifesting as macular or maculopapu- dence of delayed allergy during penicillin ther- lar exanthemata (see Sect. 2.2.4.3 and Fig. 2.7 ), apy and to evaluate the diagnostic potential of may occur during treatment with penicillins, par- patch, intradermal (with delayed reading), and ticularly aminopenicillins. Incidences of hyper- challenge tests, ampicillin, amoxicillin, and a sensitivity to penicillin range up to 10 %, and for small number of other penicillins were employed maculopapular exanthemata occurring during in tests on 259 patients most of whom had expe- therapy with aminopenicillins, the incidence is rienced maculopapular rashes. Positive patch and about 9.5 %. At least some of the penicillin- intradermal tests with penicillins revealed 98 (94 induced exanthemata are due to T cells and can to aminopenicillins, four to piperacillin) patients be confi rmed by skin testing with aminopenicil- (37.8 %) with delayed reactions. Of the 98 skin lins. Other delayed reactions elicited by penicil- test-positive patients, 93 had experienced macu- lins include acute generalized exanthematous lopapular rashes. Of 125 patients with negative pustulosis (AGEP), delayed urticaria/angio- skin tests who underwent challenges, only three edema, exfoliative dermatitis, and the more reacted. The investigators concluded that patch severe bullous exanthems, Stevens–Johnson and intradermal test positivity can provide an and Lyell’s syndromes. Severe hypersensitivity indication of delayed hypersensitivity to penicil- responses to penicillin that are not primarily seen lins, and of the two tests, intradermal testing is on the skin include vasculitis, hepatitis, intersti- the more sensitive. tial nephritis, and pneumonitis while DRESS is a In a prospective investigation of allergic cross- combination of skin eruptions, fever, and visceral reactivity between aminopenicillins, phenoxy- involvement. More detailed descriptions of these methylpenicillin, and two cephalosporins with drug- induced reactions and what is currently different R1 side chains, 71 patients (57 with a understood about the underlying mechanisms are history of macular or maculopapular exanthema, contained in Sects. 2.2.4, 3.6.3 and 3.8 . two with erythema multiforme-like exanthema, nine with acute urticaria, and three with unclassi- 5.1.9.1 Diagnostic Tests fi ed symptoms) were evaluated with intradermal 5.1.9.1.1 Skin Tests and patch tests. Sixty eight of the 71 patients An extended presentation on skin testing is set (95.8 %) had at least one positive intradermal or out in Sect. 4.2 . patch test to ampicillin or amoxicillin, 48 reacted Intradermal tests with delayed reading, patch to the aminopenicillins only, four to the aminope- tests, and occasionally prick tests provide the nicillins and benzylpenicillin, and 16 to the three mainstay diagnostic procedures for the evalua- penicillins plus phenoxymethylpenicillin. From tion of delayed reactions to penicillins and other these results, intradermal and patch tests were

[email protected] 5.1 Penicillins 157 deemed to be reliable diagnostic tools with high thresholds are a more than 3 mm diameter wheal sensitivity for delayed-type hypersensitivity to for prick tests and an increase in initial bleb wheal aminopenicillins but, because tests were some- diameter of more than 5 mm for intradermal tests. times positive with only one of the methods, Reactions after intradermal testing are documented combined use of the two tests was recommended. by the diameter of the erythema, papulation, and It has been shown that positive skin tests can be infi ltrates together with descriptions of swelling, obtained with aminopenicillins years after expo- erythema, eczema, etc., and photodocumentation if sure to the drugs and this was confi rmed in this possible. For reading patch tests, see Sect. 4.2.4.3 . study with 20 of the 21 patients testing positive more than 1 year after their exanthematous reac- 5.1.9.1.2 Challenge (Provocation) Tests tions. Importantly, a positive test result was not Often restricted by ethical considerations, drug always obtained with ampicillin and amoxicillin challenge can be regarded as the best test for con- so testing with both drugs seems to be necessary. fi rming a drug-induced delayed hypersensitivity The usefulness of the penicillin major and response. Patients who are negative to all of the minor determinants in evaluating delayed reac- other tests for a β-lactam may undergo challenge tions to penicillins was assessed in intradermal testing with one-hundredth of the therapeutic and patch tests on 162 patients who experienced dose of the β-lactam as the initial dose, and, if delayed reactions to penicillins, mainly aminope- negative, 3 days to 1 week later, a one in ten dilu- nicillins. Positive intradermal and/or patch tests tion of the therapeutic dose should be given. If in 157 patients (96.9 %) to the responsible peni- the response is again negative, a full therapeutic cillin reagents indicated cell-mediated hypersen- dose is administered after the same interval cho- sitivity while only 9 (5.5 %) and 17 (10.5 %) sen before the second dose. All precautions set were positive to penicilloyl-polylysine and minor out in Chap. 4 and mentioned above must be determinant mix, respectively, demonstrating the observed and the test is contraindicated in limited usefulness of the two benzylpenicillin patients with DRESS, AGEP, bullous exanthe- determinant preparations. mas, Stevens–Johnson syndrome, and toxic epi- Individual penicillins, penicillin determinants dermal necrolysis. Tests on patients with and their concentrations (all diluted in sterile drug-induced vasculitis, anemia, and neutropenia physiological saline) used in skin tests: are also contraindicated. Benzylpenicillin at 100 and 10,000 IU/ml (0.06 and 6 mg/ml); ampicillin and amoxicillin, both at 5.1.9.1.3 Delayed Reactions to Penicillins concentrations of 1–2 and 20–25 mg/ml; other and the Lymphocyte penicillins, 1–20 mg/ml; cephalosporins 2 mg/ Transformation Test ml; commercially available benzylpenicilloyl- This test is discussed in Sect. 4.7.1 . polylysine solution (see Sect. 5.1.5.1 ) initially Although claimed to be a useful test in the diluted 1:10 and undiluted if negative; minor hands of some experienced with the technique, determinant mix diluted 1:100 and repeated undi- others have found it unreliable and diffi cult to luted (2 × 10−2 M) if initial test is negative. For standardize. Yet to be validated and still essen- patch testing, penicillins can be used at a concen- tially a research tool, the test is claimed to have a tration of 5 % w/w in petrolatum. Drugs in solid sensitivity of 74 % with a rather low specifi city of form such as tablets, capsule contents, pessaries, 85 %. One group found an overall sensitivity of etc., should be ground fi nely in a mortar and 62 % and a specifi city of 92.8 % in 51 patients formulated in petrolatum or in saline if soluble, with a well-documented history of β-lactam taking into account the ratio of active drug to allergy (31 immediate reactors and 19 non- nondrug components in the tablet/capsule etc. immediate). This was made up of sensitivities of Prick and intradermal tests with penicillins 64.5 % and 57.9 % for the immediate and non- should be read after 20 min and delayed reactions immediate groups, respectively, but the prolifera- in the intradermal test after 48 and 72 h. Positive tive responses, expressed as stimulation indices,

[email protected] 158 5 β-Lactam Antibiotics were higher in the cells from the delayed group. undergo processing to stimulate T cell clones The authors considered the test to be a useful in specifi c for this determinant. Extension of these vitro diagnostic tool to identify subjects allergic experiments to an examination of the specifi cities to penicillins, especially the delayed reactors of the T cells revealed two different recognition where, somewhat surprisingly, they found it patterns—one directed at the penicilloyl specifi c- superior to skin testing. Interestingly, T cell pro- ity plus the side chain structure and the other liferative responses were seen 10 or more years more broadly cross-reactive with recognition of after initial exposure to penicillin and without the aminopenicillins, ampicillin and amoxicillin, reexposure in the years between. as well as benzylpenicillin. Further investigations It has been suggested that the relatively poor of the structural features recognized by sensitivity of the lymphocyte transformation test benzylpenicillin- reactive T cell clones from dif- is due to background nonspecifi c proliferation of ferent patients identifi ed the benzyl side chain the cultured peripheral blood mononuclear cells and the thiazolidine ring as antigenic determi- and only those patients with the highest numbers nants. Precise positioning of covalently bound of antigen-specifi c T cells are detected with the benzylpenicillin via a lysine residue on designer test. This suggestion appears to have been sup- peptides containing a DRB1*0401-binding motif ported by results from a comparative study in was required to induce proliferation of penicillin- which cells from 22 patients with well- specifi c clones and to be necessary for optimal documented histories of T cell-mediated allergy T cell recognition but, even more interestingly, a to amoxicillin were examined with the lympho- peptide sequence derived from a natural cyte transformation test and an ELISPOT assay DRB1*1101-binding peptide with attached peni- (see Sect. 4.7.3.2 ) for the detection of amoxicillin- cillin also acquired antigenic properties. Study of specifi c T cells producing IFN-γ. The IFN-γ the specifi city of penicillin-specifi c T cell clones ELISPOT assay seems to distinguish between was recently taken up and extended in experi- immediate and delayed hypersensitivities and ments designed to characterize the antigen recog- may prove to be a sensitive test for improving the nition profi le of clones from cystic fi brosis diagnosis of delayed hypersensitivity to β-lactam patients hypersensitive to piperacillin. drugs. Piperacillin-responsive T cell clones (CD4+, CD4+D8+, CD8+ but mainly CD4+) secreted 5.1.9.2 Recognition of Penicillin high levels of the Th2 cytokines IL-4, IL-5, and Antigens by T Cells IL-13 as well as CCR4 or macrophage infl amma- One of the relatively recent applications of the tory protein-1β (MIP-1β), the chemokine with extraordinary advancement in knowledge of cel- specifi city for CCR5 receptors. Mass spectrome- lular immune processes over the last 25 years is try and other investigations revealed that piper- witnessed in the fi eld of drug allergy and, more acillin bound to albumin lysines in T cell cultures specifi cally, in studies aimed at understanding the at residues 190, 195, 199, 432, and 541 after 16 h role of T cells in delayed reactions to drugs. As incubation, but, after only 1 h, the drug was occurred with studies on drug-induced immedi- detected at only one site. These results fi t with the ate hypersensitivity, the drugs most commonly time needed to stimulate a T cell proliferative selected for the cellular investigations have been response and demonstrate the need for high the penicillins since their frequency of use, high piperacillin binding for T cell activation. Attempts incidence of reactions, and the accumulated to stimulate clones with other penicillins and knowledge of their antigenic structures make with cefoperazone which has a structurally them the logical choice. Early investigations on related side chain to piperacillin and a dihydro- T cell clones showed that benzylpenicillin-specifi c thiazine instead of a thiazolidine ring were unsuc- clones were HLA class I or class II restricted and cessful, demonstrating that the T cells specifi cally processing of the free drug was not required recognized the penicillin nucleus and piperacillin whereas benzylpenicilloyl–HSA conjugate must side chain structures. Piperacillin–albumin con-

[email protected] 5.2 Cephalosporins 159 jugate stimulated T cells via a MHC- and forms protein adducts with human serum albumin processing-dependent pathway but fl ucloxacillin, and mass spectroscopy and other techniques have which reacts with the same lysine residues on revealed up to nine modifi ed albumin lysine albumin as piperacillin, did not stimulate T cells, residues with Lys190 and Lys212 being particu- indicating structural specifi city of the T cell larly involved. Both the parent drug and its receptors expressed on the drug-specifi c clones. 5-hydroxymethyl metabolite reacted with the Flucloxacillin did, however, competitively reduce same lysine residues. Whether such drug–protein piperacillin binding and the piperacillin or peptide complexes play a central role in medi- conjugate- specifi c T cell response, suggesting ating fl ucloxacillin-induced liver injury remains that co-administration of such a competitor might to be seen. prevent allergic sensitization or even reduce the hypersensitivity response in patients with established sensitivity. 5.2 Cephalosporins

For the early cephalosporins, the names were 5.1.10 The Genetic Basis of spelt with a “ph” and this has continued in Penicillin-Induced English-speaking countries although the UK, Liver Injury along with European countries, appears to be adopting International Proprietary Names which Due particularly to studies from Australia and use the “cef” spelling. Cephalosporins are often Sweden, fl ucloxacillin has a well-known associa- referred to as fi rst- or subsequent-(second, third, tion with severe and debilitating cholestatic liver and so on) generation drugs, a classifi cation disease. The incidence of this association is esti- essentially based on both the sequence of devel- mated to be from about 1 in 10,000–100,000; a opment and antimicrobial action. The newer gen- Swedish study estimated the rate to be 1 case per eration drugs (and some newer fi rst-generation 10,000–30,000 prescriptions. The female sex, drugs) tend to be spelt with an “f.” First- patient age over 55 years, high daily dose, and generation drugs are predominately active against taking the drug for more than 14 days seem to be Gram-positive organisms while succeeding gen- associated with a higher risk of the penicillin- erations were endowed with increasing Gram- induced liver injury. A 1992 paper from Sweden negative activity. It is arguable whether or not the reported 77 liver reactions “probably or possibly” “generation” classifi cation has any clinical rele- induced by penicillinase-resistant penicillins, vance or use. including cloxacillin and dicloxacillin as well as Together with the penicillins, the cephalospo- fl ucloxacillin, which were reported spontane- rin β-lactam antibacterials are the most widely ously to the Swedish Adverse Drug Reactions used antibiotics for the treatment of common Advisory Committee. A genome-wide associa- infections. The origin of cephalosporin antibiot- tion study of 51 cases of fl ucloxacillin-induced ics dates back to 1948 with the demonstration of liver injury and 282 controls showed a strong antimicrobial action in a fungal extract from what association in the major histocompatibility com- was then called Cephalosporium acremonium . plex (MHC) region with a marker in complete The Cephalosporium genus is now known as disequilibrium with HLA-B*57:01. This associa- Acremonium. The cephalosporins used in medi- tion, confi rmed in follow-up genotype studies, cine today are semisynthetic derivatives of the offers new insights into understanding the condition natural antibiotic cephalosporin C. Soon after and may ultimately improve its diagnosis, but the cephalosporins were introduced into clinical use, mechanism underlying fl ucloxacillin cholestatic adverse reactions including apparent hypersensi- hepatitis remains incompletely understood. Some tivity reactions and obvious anaphylactic clones from patients allergic to fl ucloxacillin are responses began to appear, and although their CD8+ and positive for granzymes. Flucloxacillin similar structural features to the penicillins

[email protected] 160 5 β-Lactam Antibiotics

suggested that the two groups might cross-react cephalothin with the possibility of complicating strongly and display similar antigenic and aller- routine cross-match tests and immune hemolytic genic properties, clinical manifestations, and anemia have not been substantiated. diagnostic challenges, the reality has been somewhat different. Now, almost 50 years since the fi rst therapeutic use of cephalosporin C in 1963, 5.2.2 Clinical Aspects of Cross- there are still a number of important outstanding Reactivity of Cephalosporins questions. These include the nature of cephalo- and Penicillins sporin breakdown products and allergenic determinants; the relationship between penicillin- and Retrospective surveys of the published literature cephalosporin-allergic sensitivities including on work undertaken over 40 years ago and current their clinical cross-reactivity; the specifi city and fi ndings at the time on the relationship between predictive value of cephalosporin skin testing; allergy to cephalosporins and a history of penicil- the relationship between cephalosporin-reactive lin allergy revealed fi ndings that are still widely IgE antibodies and clinical allergy; and the speed quoted and often disputed. The incidence of of decline in skin test and serum IgE positivity in patients with a history of penicillin allergy who cephalosporin-allergic patients. had allergic reactions to cephalosporins was found to be 9.2 % (5.4–16.6 %) while patients who had a reaction but a negative history to penicillins 5.2.1 Incidence of Cephalosporin showed an incidence of 1.7 % (1–2.5 %), that is, Hypersensitivity and Clinical allergic reactions to cephalosporins appeared to be Aspects about 5.4 times more frequent in patients with a history of penicillin allergy. A survey of 15,987 From retrospective studies and information from patients treated with cephalothin, cephaloridine, pharmaceutical companies, the incidence of cephalexin, cefazolin, or cefamandole published immune-mediated adverse reactions to cephalo- in 1978 revealed that 8.1 % of patients with a his- sporins was calculated to be 1–3 %, although a tory of penicillin allergy had reactions compared fi gure of 1 % is commonly seen in the literature. with 1.9 % without such a history; this indicates a Dermatologic reactions, mainly rashes, urticaria, fourfold risk for patients allergic to penicillin. and pruritus, occur with a frequency of about However, as pointed out at the time and since then, 1–3 %. Maculopapular rashes that are not pruritic this large difference is probably not likely because are generally thought to be non-IgE-mediated and penicillin-allergic individuals show an increased therefore not a reason to avoid cephalosporins, but incidence of drug hypersensitivities, some reac- a rash with pruritus may be an indication of an tions included as allergic reactions to cephalospo- allergic reaction. Anaphylaxis to cephalosporins rins were probably not immune mediated and a is said to be rare with an incidence of up to 0.1 %, positive history of penicillin allergy may not but this may be a signifi cant underestimate since always be reliable. Over the years, there have been no reliable large surveys seem to have been under- many studies, large and small, of cephalosporin taken. Apart from anaphylaxis, other immediate administration to patients with a history of penicil- type I reactions induced by cephalosporins include lin allergy. These have delivered widely varying urticaria, laryngeal edema, bronchospasm, and results with incidences of positive reactions up to hypotension. Serum sickness-like reactions have 18 % and a risk up to about eight times that of been reported in children, fever and immunohe- patients with no penicillin allergy. Both these fi g- matological reactions are uncommon, and severe ures are likely to be overestimates, but it does skin reactions such as exfoliative dermatitis and seem reasonably certain that there is a signifi cantly Stevens–Johnson syndrome occur but again are higher risk of a reaction to a cephalosporin in uncommon. Early reports of a high incidence of a patients already allergic to a penicillin. A recent direct positive Coombs’ test in patients receiving retrospective cohort study using the United

[email protected] 5.2 Cephalosporins 161

Kingdom General Practice Research Database and penicillins and cross-reactivity for cephalo- looked at a total of over 3.3 million patients given sporins overall was highest with the fi rst- and penicillin and, in particular, at over half a million second-generation drugs. (~15 %) who were subsequently given a course of cephalosporin. Estimation of numbers of patients who experienced allergic-like events within 30 5.2.3 Structures and Classifi cation days revealed that the unadjusted risk ratio for of Cephalosporin Drugs those who had a prior reaction compared to those who did not have a reaction was 10. The absolute Although the natural antibiotic cephalosporin C risk for anaphylaxis after a cephalosporin was lacked suffi cient potency for therapeutic use, 0.001 %, leading to the conclusion that a markedly demonstration of the structure of its nucleus, increased risk existed for patients who had previ- 7-aminocephalosporanic acid, showed that the ously reacted to a penicillin, but cross- reactivity cephalosporins had a structural basis analogous was not an adequate explanation for the increased to the 6-aminopenicillanic acid core of the peni- risk. Recently it has been claimed that many cillins. Like the structurally related other major patients with adverse reactions quite unrelated to β-lactam antibiotic group, the penicillins, cepha- immune hypersensitivities were categorized as losporins also contain a β-lactam ring with a allergic in the early surveys. For this reason as well linked side chain (R1) but with a six-membered as those mentioned above, and the early use of dihydrothiazine ring instead of the fi ve- membered fi rst-generation cephalosporins, the risk of cross- thiazolidine ring found in the penicillins. In addi- reactivity is probably closer to 0.5 % than 10 %. tion, cephalosporins have a second side chain Second- and third-generation cephalosporins group (R2), which is attached at position 3 of the appear to carry less risk of provoking a reaction in dihydrothiazine ring. The general structure, penicillin-allergic subjects and, given the struc- structures of the two side chain groups, and the tural differences seen in the side chains, especially generation of some of the many cephalosporins for the third- and fourth-generation cephalosporins developed and marketed for therapeutic use are (see below), this is perhaps not surprising. Some shown in Fig. 5.13 . Classifi cation of the drugs recent studies have concluded that the use of third- into different generations has not always been or fourth-generation cephalosporins in penicillin- observed and consistent in all countries, e.g., allergic patients carries a negligible risk of cefaclor is regarded as a fi rst-generation drug in cross-allergenicity. Japan but a second-generation cephalosporin in A review of the use of cephalosporins in chil- the USA. The classifi cation of cephalosporins is dren with anaphylactic reactions to penicillins based on an intended broader spectrum of suc- concluded that there were no published case ceeding generations of drugs, but this has some- reports of anaphylaxis to these antibiotics in the times come at the cost of decreased Gram-positive assessed group and, in any case, anaphylaxis to activity. However, the fourth-generation cephalo- cephalosporins was extremely rare in children. sporins are said to be truly broad spectrum in A prospective study of over 1,000 children with antimicrobial activity. suspected immediate reactions to cephalosporins and/or penicillins showed that 58 % were skin or challenge test positive to a β-lactam with 94.4 % 5.2.4 Cephalosporin Antigens positive to penicillins and 35 % positive to cepha- and Allergenic Determinants losporins. Approximately one-third of penicillin- allergic children cross-reacted with a cephalosporin Despite the chemical similarity of the cephalo- and those allergic to a cephalosporin showed an sporins and penicillins, differences in chemical 84 % frequency of reactions to penicillins. Cross- reactivity and stability exist. This is most obvious reactivity between cephalosporins was lower than when conditions to prepare the penicilloyl deter- cross-reactivity observed between cephalosporins minant are applied to the cephalosporins.

[email protected] G1 H H R1- -R2 S R -CONH 1 1 7 6 2 1st Cephalexin CH CH3 8 5 3 NH N 4 2 R O 2 1st Cephaloglycin CH CH OCOCH 2 3 COOH NH2 1 G R - -R2 2nd Cefaclor CH Cl SO3H 1 H C NH 2 N C 2 N N 3rd Cefdinir H N N-OH CH=CH 2nd Cefonicid CH CH S 2 S 2 2 N N N C NH2 3rd Ceftizoxime H N N-OCH H 1st Cephradine CH CH3 2 S 3 N C NH2

OCH 3rd Cefetamet H N N-OCH CH3 1st Cefroxadine CH 3 2 S 3

NH2 N C CH 3rd Cefpodoxime 1st Cefadroxil HO CH 3 H N N-OCH CH OCH 2 S 3 2 3 NH 2 N C CH=CH-CH 2nd Cefprozil HO CH 3 3rd Cefotaxime H N N-OCH CH2OCOCH3 2 S 3 NH2 N CH N N C N 3 1st Cephatrizine HOCH CH S 2 CH S NH 3rd Cefodizime H N N-OCH 2 2 S 3 NH2 S CH2 COOH H3C CH CH OCOCH 1st Cephalothin 2 2 3 N S N C N 3rd Cefmenoxime CH2 S H N N-OCH3 N 2 CH OCONH 2 S N 2nd Cefoxitin CH2 2 2 S H C Na N C 3 N N CH +N 1st Cephaloridine CH2 2 H N N-OCH CH -S O 3rd Ceftriaxone 2 S 3 2 S N O N N N C + N CH S 4th CH N 1st Cefazoline CH2 2 Cefepime 2 N H N SCH3 2 N-OCH3 S H3C N N N N 1st Ceftezole N CH CH2 S N C 2 N+ N S 4th Cefpirome CH2 H N 2 N-OCH3 Cl S N N 1st OCHN CH S Cefazedone 2 2 N C CH N+ SCH4th Cefozopran 2 3 N N Cl H2N N-OCH3 S N

H3C 2nd CH N N C Cefamandole N CH CH2 S 5th Ceftobiprole OH N H N N N-OH N N 2 S H C O 3rd Cefoperazone CH 3 N N N C S H HN N CH S N S 2 N 5th Ceftaroline HN N-OCH CH O N S 2 3 N O N fosamil HOP OH N+ CH O 3 O N N C

CH CH Cefixime H N N-O-CH -COOH CH=CH 2 3 3rd 2 S 2 2 HO CH H3C 3rd Cefpiramide N C + N CH CH N HN N Ceftazidime 3 2 CH2 S 3rd H N N 2 S N-O-C-COOH O N O N CH3 N C NH H C 3rd Ceftibuten H N C-CH -COOH H 3 2 S 2 2 CH N 2nd Cefmethazole 3 N NCCH SCH CH2 S 2 2 N CH OCONH N 2nd Cefuroxime C 2 2 O H3C N-OCH3 H NOC S N 2 2 N 2nd Cefotetan C CH2 S N NSCH HOOC S N 1st Cephapirin 2 CH2OCOCH3 H3CCH3 N NCCH 1st Cefacetril 2 CH2OCOCH3 (CH2)2 H N N N 2 N 1 G, Generation. 2nd S CH CH2 S Cefotiam S 2 N 2 a N Cephamycin with an -methoxy-group (-OCH3) at the7-position.

Fig. 5.13 Structures of some of the more frequently used ﬁ rst-, second-, third-, fourth-, and ﬁ fth-generation (G) drugs out of the large number of cephalosporins developed as potential antibacterials

[email protected] 5.2 Cephalosporins 163

S R1 CONH CH CH CH2 CN C CH O C 2 R2 COOH ROUTE ROUTE 1 Cephalosporin 2

S S R1 CONH CH CH CH2 R1 CONH CH CH CH2 O CN C O CNH C C CH2 C CH2 R Lysine NH Lysine NH 2 COOH COOH

Cephalosporoyl derivatives

HS HS

R1 CONH C CH CH2 R1 CONH C CH CH2 O CN C O CNH C C CH2 C CH2 R Lysine NH Lysine NH 2 COOH COOH

R1 CONH C CHOH O C Lysine NH

Penaldate

R1 CONH C CHNH2 O C Lysine NH

Penamaldate

Fig. 5.14 Aminolysis of cephalosporins and decomposi- cephalexin, cefadroxil), immediate breakdown of the tion products. With a good leaving group at C-3 (as in dihydrothiazine ring does not occur and the intact cephalo- cefaclor and cephalothin), aminolysis in the presence of sporoyl structure forms, if only temporarily, as the reaction protein or polylysine leads to cleavage of the β-lactam proceeds via route 2. The unstable intermediates decom- ring via route 1. With a poor leaving group at C-3 (e.g., pose to form the penaldate and penamaldate structures

5.2.4.1 Aminolysis of Cephalosporins new chromophores) consistent with the forma- and Decomposition Products tion of unstable intermediates that decompose to In early studies on cephalosporin C in E. P. penaldate and ultimately penamaldate structures Abraham’s laboratory in Oxford, acid hydrolysis (Fig. 5.14 ). Analysis of the penamaldyl product yielded no penicillamine and alkali treatment led showed that it was the amino group from to fragmentation of the molecule. Aminolysis of the amino acid not from the nitrogen atom of the cephalosporins in the presence of polylysine or dihydrothiazine ring that was involved in the protein gave results (including the appearance of formation of the penaldate-like compound.

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The nature of the leaving group at position C-3 5.2.4.2.1 Studies Implicating the R1 Side on the dihydrothiazine ring has a signifi cant Chain in Clinical Hypersensitivity infl uence on the activity of the β-lactam ring and In the earliest demonstration of a useful in vitro the stability of the dihydrothiazine ring. immunoassay for the detection of cephalosporin- Nucleophilic- induced β-lactam ring cleavage of reactive IgE antibodies in the sera of patients cephalosporins with a good leaving group at C-3 allergic to a cephalosporin, cephalothin sodium is accompanied by elimination of the leaving was covalently coupled to bis -oxirane-activated group. The presence of a good leaving group as Sepharose® (Pharmacia) (see Sect. 4.3.1 ) and in cefaclor (Cl), cephalothin (acetoxy), and ceph- used to detect complementary IgE in the sera of aloridine (pyridinium) would be expected to pro- patients who experienced anaphylaxis after ceed via the formation of a cephalosproyl administration of cephalothin or cephalexin. derivative with a double bond at Δ4 and a methy- Quantitative hapten inhibition results (Table 5.5 ) lene group at C-3 (route 1, Fig. 5.14 ). With a poor for one of the cephalothin-sensitive patients leaving group at C-3, for example, the methyl revealed clear recognition of the side chain of group in cephalexin and cefadroxil, aminolysis cephalothin and cross-recognition of compounds does not appear to cause immediate breakdown with R1 groups showing a structural similarity to of the dihydrothiazine ring and the cephalospo- the cephalothin R1 group, in particular, benzyl- royl structure forms, if only temporarily before penicillin. This conclusion was reinforced by the eventually proceeding to the penaldate and per- signifi cant inhibition seen with the side chain haps penamaldate derivatives (route 2, Fig. 5.14 ). analogs 2-thiopheneacetic acid and phenylacetic In fact, even with a good leaving group such as acid. Bearing in mind the decomposition of the pyridinium in cephaloridine and ceftazidime, dihydrothiazine ring induced by the coupling departure of the leaving group may not be con- procedure at alkaline pH, the determinant pre- certed with β-lactam ring opening. The rate of sented in the assay may be the penaldate- (see decomposition increases as the concentration of below) and/or penamaldate-like structures the compound increases and the pH decreases. (Fig. 5.14 ), and therefore, any reactive IgE anti- Dewdney has pointed out that the still often-used bodies detected with the solid phase conjugate cephalothin may undergo breakdown by both would be complementary to remaining unde- routes, one via direct aminolysis of the parent graded structures, most likely the cephalothin compound (route 1) and the other via its major side chain structure, and this, in fact, was found. metabolite deacetylcephalothin (route 2). The usefulness of the activated Sepharose radioimmunoassay for the detection of IgE antibodies 5.2.4.2 Allergenic Signifi cance to cephalosporins was also demonstrated by of the Acyl (R1) Side Chain Romano and collaborators in assays on sera from of Cephalosporins 70 patients with immediate reactions to cephalo- The work outlined above, and the interpretation sporins and 40 control sera from healthy, non- of the chemical fi ndings, indicates that as far as atopic individuals. A positive result was recorded any resultant determinants are concerned, ami- for 52 (74.3 %) of the patients and the specifi city nolysis results in structures in which only the R1 of the assay was 100 %. Again, cross-reactions side chain, the attached amide, and remnants of were observed and these could be explained by the β-lactam ring remain from the original cepha- recognition of identical or similar side chain losporin molecule. The resultant penaldate- and groups. The authors acknowledged that cephalo- any penamaldate-like structures linked to carrier sporin–Sepharose conjugates provide a useful protein therefore represent hapten–carrier com- diagnostic tool and could be employed as a complexes that may interact with side chain (R1)- plementary test with skin testing to evaluate specifi c IgE antibodies in allergic responses to cephalosporin-induced immediate reactions. therapeutic dosage of cephalosporin drugs. Given the clear-cut demonstration of the utility

[email protected] 5.2 Cephalosporins 165

Table 5.5 Inhibition of IgE antibody binding to cephalothin-Sepharose by cephalosporins and penicillins Structure H H S R -CONH 1 7 N Inhibition (%) of binding of O R2 IgE antibodiesa with COOH 400 nmol of compound to b b Compound R1 R 2 cephalothin-Sepharose Cephalothin 57 CH2 -CH OCOCH S 23 Cephaloridine 58 CH +N CH2 2 S Cefoxitin c 33 CH2 -CH OCONH S 22 Cephalosporin C HOOC CH(CH ) 9 2 3 - CH23 OCOCH NH2 Cephaloglycin 28 CH -CH OCOCH 23 NH2 Cephalexin 0 CH -CH 3 NH2 Cephapirin 26 N SCH2 -CH OCOCH 23 Cefazolin N N N N 18 N CH 2 CH2S CH3 N S Cephradine 14 CH -CH 3 NH2 Benzylpenicillin S 44 CH2 CONH CH3

N CH3 O COOH Ampicillin S 12 CH CONH CH3

NH2 N CH3 O COOH Amoxicillin S 4 HO CH CONH CH3

NH2 N CH3 O COOH 2-Thiopheneacetic acid 31 CH2 COOH S Phenylacetic acid 20 CH2 COOH

From Harle DG, Baldo BA. Drugs as allergens: An immunoassay for detecting IgE antibodies to cephalosporins’. Int Arch Allergy Appl Immunol. 1990 ; 92: 439. Reprinted with permission from S. Karger AG, Basel a IgE in serum from patient with anaphylaxis to cephalothin b R1 and R2 refer to cephalosporins, not to penicillins and other compounds examined c Cefoxitin is further characterized by the presence of a methoxy residue (–OCH3 ) at position 7 of the β-lactam ring

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R 2

NH 6 R1 C S R C NH 1 1 CH CH CH (CH ) NH CH N COOH O 2 3 2 3 2 7 6 O 7 H 8 3 CN5 4 R2 C O O NH

COOH (CH2)3

CH3 Cephalosporin Cephalosporoyl

- R2

NH 6 R1 C ? CH O 7 C O NH

(CH2)3

CH3 Model determinant structure with intact side chain

Fig. 5.15 Determination, prior to intended synthesis, of Antigenic Determinants of Cephalosporins: In vitro model structure with intact side chain remaining after Studies of Chemical Structure−IgE Molecular Recognition aminolysis of cephalosporins. Adapted from Montañez Relationships. Chem Res Toxicol 2011 ; 24: 706. Adapted MI et al. Synthetic Approach to Gain Insight into with permission from © 2011 American Chemical Society and performance of the Sepharose assay, its ease highly likely, these prove to be penaldate- and/or of preparation, and proven capacity to detect side penamaldate-like, the procedure and test reagents chain groups on cephalosporins, the method can- should continue to be considered as useful and not be dismissed simply because the dihydrothi- convenient diagnostic tools. azine ring is lost during preparation of the In a study aimed at gaining insights and pre- drug-solid phases. The procedure has worked paring and ultimately utilizing antigenic determi- well to detect R1 determinants on cephalothin, nant structures of cephalosporins, Perez-Inestrosa cefaclor, cephalexin, ceftriaxone, cefotaxime, and collaborators set out to synthesize the pro- ceftazidime, cefonicid, cefuroxime, and cefa- posed structure remaining on the carrier protein mandole and is particularly effective when used after the chemical breakdown resulting from with cephalothin and cefaclor. Because of the aminolysis of the cephalosporin molecule. known disintegration of the dihydrothiazine ring, n -Butylamine was used as the nucleophile instead nucleophilic addition of the bis -oxirane-activated of the primary amino groups of lysine residues in solid phase at alkaline pH to the S or N atom of proteins. The proposed aminolysis pathway and the six-membered ring may not be possible, but it model structure to be utilized in allergenic deter- might proceed by reaction with the N of the ami- minant investigations are shown in Fig. 5.15 . nobenzyl and S of the thiophene groups of the R1 Note that the structure remaining after disintegra- side chains of cefaclor and cephalothin, respec- tion of the dihydrothiazine ring retains the entire tively, although this might be expected to inter- R1 side chain group of the original cephalosporin fere with IgE interaction with the side chain. so, in any diagnostic application, only side chain- Efforts should be made to characterize the struc- speciﬁ c IgE antibodies would be detected. From tures retained after coupling and if, as seems research in the late 1960s and early 1970s and

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-H, -H, -OH, -OH, [R = [R = H N CH2 R -SH, H N CH2 R -SH, 2 2 3 3 -(OCH ) ] -(OCH ) ] 2 3 2 2 3 2 1 1

O OH O NH (CH2)3

Derived a-amino acid CH3 with selected functional Core structure group +

R1 OH C Condensation O -H, Selected side -OH, chain structure [R = R1 NH CH2 R -SH, C 3 2 -(OCH3)2] O 1

O NH (CH2)3

CH3 Model determinant

Fig. 5.16 Synthesis of the core structure (determined into Antigenic Determinants of Cephalosporins: In vitro from products of the aminolysis pathway—see Fig. 5.15 ) Studies of Chemical Structure−IgE Molecular Recognition and model haptenic structures with different R1 side chain Relationships. Chem Res Toxicol 2011 ; 24: 706. Adapted groups and functional groups attached at C-3. From with permission from © 2011 American Chemical Society Montañez MI et al. Synthetic Approach to Gain Insight

NMR data, it can be inferred that the model respectively, are utilized. When these model determinant has a penaldate-like structure. Based determinants were used in hapten inhibition experi- on knowledge of the aminolysis pathway and the ments with sera from cephalosporin-allergic likely structure of this determinant, a synthesis patients and cephalosporin-solid phases, optimal was devised for a series of antigenic determinants inhibition was seen with model structures contain- with the same core structure (shown in bold, ing the R1 side chain of the cephalosporin that Fig. 5.16) but with differences in the R1 group induced the reaction. Molecules containing OH and the functional group at C-3 [C-6 in the and O(CH 3 )2 at C-3 were slightly better inhibitors, original, undegraded cephalosporin molecule suggesting to the investigators that these structures (Fig. 5.15 )]. Since the nature of the substituent at mimic the real antigenic determinant involved in C-3 is unknown, and this C atom in the intact the initial allergic reaction. By contrast, the presence parent molecule (attached to a S atom) was of an S atom at C-3 drastically decreased inhibi- capable of undergoing a number of different tion, often abolishing it and suggesting that, in reactions including reactions with O and S, vivo, cephalosporin antigenic determinants may hydroxyl, thiol, and acetal groups were investi- occur via breaking of the S-1–C-6 bond. A puzzling gated as functional groups at C-3. For the prepa- feature of this work and its strategy is the nature ration of a methyl group at C-3, an alanine of the cephalosporin conjugates used on the solid derivative can be used as the starting derived phases for the testing of the inhibitory activities α-amino acid, and for the OH and SH functional of the model determinants. The preparations of groups, the desired serine and cysteine derivatives, the cephalosporin–poly-L -lysine conjugates used

[email protected] 168 5 β-Lactam Antibiotics in the radioimmunoassay were not described, but conjugates. Early studies on rabbit antibodies to the references provided are for the preparation cephalothin and benzylpenicillin demonstrated and examination of benzylpenicilloyl and cross-reaction between the two drugs and this amoxicilloyl conjugates, suggesting that the cross-reactivity was confi rmed in humans by a equivalent conditions and procedures were number of anaphylactic reactions to cephalothin employed for the preparation of the cepha- in penicillin-allergic patients on fi rst exposure to losporin conjugates. If that was not the case and the cephalosporin. In addition, positive intrader- conditions leading to disintegration of the dihy- mal tests to cephalothin were seen in patients drothiazine ring were not employed, the nature of with known allergic reactions to penicillins, but the different cephalosporin test reagents used previously unexposed to cephalothin and passive remains unclear. After devising a strategy to transfer tests with sera from cephalosporin- avoid the lability of the cephalosporins and allergic and penicillin-allergic patients gave pos- undertaking chemical syntheses to produce and itive reactions to challenges with benzylpenicillin precisely defi ne the model determinants, the and cephalothin, respectively. Cross-reaction same thinking and approach are needed for the between the two β-lactams was also suggested preparation of the cephalosporin-solid phases by results of IgE antibody direct binding stud- if the studies’ conclusions are to be beyond ies in the authors’ laboratory where the frequen- criticism. cies of detection of benzylpenicillin- and There have been a number of reports of ana- cephalothin- reactive IgE antibodies in 1,797 phylaxis and other immediate reactions to cepha- routine tests on sera from patients with suspected losporins with a 2-aminothiazol-4-yl R1 group allergy to a penicillin(s) and/or a cephalosporin(s) seen, for example, in reactions to ceftriaxone, were compared. Sera from 123 patients (6.8 %) ceftazidime, cefepime, cefodizime, and cefi xime were positive to both benzylpenicillin and ceph- with cross-sensitivity between these drugs dem- alothin, 238 sera (13.2 %) proved positive to onstrated in skin tests. Some patients with imme- cephalothin and negative to benzylpenicillin, diate clinical and skin test reactions to cefuroxime, and only 15 sera (0.8 %) were positive to the which has a 2-furyl substituent in the R1 side penicillin but negative to the cephalosporin. chain, were also shown to be skin test positive to Inhibition experiments with rabbit antisera had cephalosporins with the 2-aminothiazolyl R1 implicated the side chains as the source of the group. The chemical basis for this cross- cross-recognition and this conclusion was sup- recognition appears to be a common alkoxyimino ported by results of quantitative hapten inhibi- substituent in the R1 groups. This can be con- tion experiments with patients’ sera containing cluded from positive skin test reactions to cefu- IgE antibodies which clearly revealed that cross- roxime, cefotaxime, cefepime, ceftriaxone, and recognition is due to the side chain (R1) groups oxacillin in a patient who experienced anaphylac- on each molecule with the methylene group as tic shock after receiving cefuroxime axetil. The the focus of the allergenic determinant and with fi rst four of these compounds have a methoxyimino the probable contribution of the bioisosteric ben- group while oxacillin has a bioisostere of the zene and thiophene rings. The exact confi nes of methoxyimino in the form of an isoxazolyl group the determinant at the β-lactam ring end of the with a methyl substituent (Fig. 5.17 ). side chain could not be determined, but inhibition fi ndings with some analogs suggested that it 5.2.4.2.2 In Vitro and In Vivo Cross- probably makes some contribution. Interestingly, Reactivity Between Cephalothin both direct binding and inhibition investigations and Benzylpenicillin showed that the cross-reactive IgE antibodies Antigenicity of the R1 side chain group has recognized benzylpenicillin more strongly than been amply demonstrated by examination of the cephalothin and this may be relevant to the specifi cities of antisera from laboratory ani- observation that many patients allergic to peni- mals immunized with cephalosporin–protein cillin have tolerated cephalothin.

[email protected] Cefuroxime CH3 O N O C HH H S C N O N ONH2 O COOH O Cefotaxime CH3 H2N N NO C HH Methoxyimino group S H S C N CH3 O N OCH3 N O O C COOH O Cefepime

CH3 H2N N NO HH C H S S C N O N N+ O CH3 COOH Ceftriaxone

CH3 H2N N NO C HH S H S C N O N S N O O COOH N H3C N O H

Methyl-substituted isoxazole group O CH Oxacillin N C 3 O CH C N C 3 C HH S C N CH3 H O N CH3 O H COOH

Fig. 5.17 Cephalosporins with a methoxyimino group in and the methyl-substituted isoxazole of the latter high- the R1 side chain responsible for skin test reactivity in a lighted in red. Three-dimensional models of these pro- patient who experienced anaphylaxis to cefuroxime axetil. posed cross-reactive groupings of atoms are also shown

Oxacillin, which contains a methyl-substituted isoxazolyl with the related sequence of atoms C=N–O–CH 3 (in the group, a bioisostere of the methoxyimino group, was also cephalosporins) and C=N–O–C–CH 3 (oxacillin) high- skin test-positive in the patient. Two-dimensional struc- lighted in orange . (Results from Hasdenteufel F et al. Ann tures of the four cross-reacting cephalosporins and oxacil- Pharmacother 2007; 41: 1069) lin are shown with the methoxyimino groups of the former

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5.2.4.3 Summary of the Current Excellent uptakes of IgE antibodies in patients’ Situation with Cephalosporin sera by the cephalosporin-HSA solid phases were Determinant Structures: Are seen in direct binding assays and specifi city was R2 Side Chains Recognized as demonstrated by clear inhibition of binding after Allergenic Determinants? preincubation of the sera with free cephalosporin For many years, knowledge of cephalosporin drugs but not with penicillins (benzylpenicillin, determinant structures and diagnostic application ampicillin, amoxicillin, and becampicillin), of such determinants has not matched the chemi- aztreonam (a monobactam), and clavulanic acid cal insights and clinical applications that began (a clavam). Imipenem (a carbapenem) and moxa- 50 years ago with the structurally related lactam (an oxacephem) showed weak inhibitory β-lactams, the penicillins. In recent years, a para- activity with a few sera. From the inhibition doxical situation has existed with the cephalo- results, it became clear that although the R1 sporins—IgE antibody tests for the specifi c group was the dominant specifi city, both side detection of cephalosporin allergenic determi- chains contributed to the IgE antibody recogni- nants have proved to be effective assays and use- tion profi les as shown by some results summa- ful diagnostically, but the chemical understanding rized in Table 5.6 and interpreted in Table 5.7 . In of the drug conjugates and solid phases employed particular, with serum 3,252, a comparison of the as test reagents has been lacking. Some recent inhibitory activities of cefaclor on the one hand chemical work discussed above has begun to and cephalexin and cephaloglycin on the other improve this situation, but doubts remain about demonstrated that the aminobenzyl group at R1 the apparent belief that, because of the lability of and Cl at R2 were the preferred structures for the dihydrothiazine ring, the R1 side chain group interaction with the cefaclor-reactive antibodies. is the only relevant allergenic structure of the These results suggested that the complementary cephalosporin molecule. IgE antibodies recognized the whole cefaclor molecule. Recognition of the ester group at R2, 5.2.4.3.1 Recognition of the R2 Side Chain demonstrated by strong inhibition with cephalo- Although the antigenic preeminence of the R1 glycin and cephalothin, was seen with serum side chain may ultimately prove to be correct, 3,323 and this, together with a clear requirement there is already evidence that with some for the aminobenzyl group at R1 and moderate cephalosporin- allergic patients, in addition to inhibition by loracarbef which has a Cl at R2, recognition of the R1 structure, the R2 side chain suggested the possible presence of a second, and/or the whole cephalosporin molecule are rec- cross-reactive population of IgE antibodies. The ognized by serum IgE antibodies and this may be strong requirement for the aminobenzyl group at by one or more populations of antibodies. This R1 for serum 4,679, together with clear-cut rec- can be seen by analysis of the quantitative inhibi- ognition of the ester group at R2 (demonstrated tion data selected from results of a study of fi ne by the strong inhibition with cephaloglycin), structural recognition patterns demonstrated by again suggested the possible recognition of a sec- cephalosporin-reactive IgE antibodies in the sera ond allergenic specifi city. With serum 4,764, rec- of cephalosporin-allergic patients (Table 5.6 ). In ognition of the aminobenzyl group at R1 was an attempt to avoid the uncertainties resulting accompanied by a requirement for a “small” from opening the β-lactam ring and the associ- group (Cl or CH3 ) at R2. ated dihydrothiazine ring fragmentation, cefa- Apart from fi ndings from the authors’ labora- clor, cephalexin, and cephalothin solid phases tory, some other observations made during the for use in radioimmunoassays to detect investigation of patients’ immediate reactions to cephalosporin-reactive IgE antibodies were cephalosporins suggest that specifi cities in addi- prepared by linking HSA to the carboxyl tion to, or other than, the R1 group are recognized group at position 4 with 1-ethyl-3-(3- by IgE antibodies. In inhibition studies under- dimethylaminopropyl)carbodiimide (EDAC). taken in Spain with the sera of three patients

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Table 5.6 Inhibition by cephalosporins of binding of IgE antibodies in the sera of cephalosporin-allergic patients to cefaclor–HSA a. Recognition patterns indicating drug IgE-binding determinants b Structure S R1-CONH N R O 2 Inhibition (%) of IgE antibody binding Cephalosporin COOH by 500 nmol of cephalosporin with serum inhibitor R1 R 2 3,252 3,323 4,679 4,764 Cefaclor - 82 95 77 92 CH Cl NH 2

Loracarbef CH - 58 33 35 91 Cl ( a carbacephem ) NH 2 Cephalexin 32 6 52 89 CH -CH NH 3 2 Cephradine 31 21 22 88 CH -CH NH 3 2 Cefadroxil 12 4 8 88 HO CH -CH NH 3 2 Cephaloglycin 10 69 88 9 CH -CH OCOCH NH 23 2 Cephalothin 5 57 11 3 CH2 -CH OCOCH S 23

Cephapirin N SCH 7 0 0 0 2 -CH OCOCH 23 Cefotaxime N C 11 14 3 2 -CH OCOCH H2N N-OCH 23 S 3 Cefoxitin ( a 0 14 5 11 CH2 -CH OCONH cephamycin ) S 22 a Prepared by coupling human serum albumin to the 4-carboxyl group of cefaclor b Refer to Table 5.7 for interpretation From Pham NH, Baldo BA. β-Lactam drug allergens: Fine structural recognition patterns of cephalosporin-reactive IgE antibodies. J Mol Recognition 1996 ;9:287. Reprinted with permission from John Wiley and Sons allergic to cefaclor, inhibition was almost exclu- allergic sensitivity to a cephalosporin occurs sively confi ned to cefaclor while no inhibition exclusively via recognition of the R1 side chain was seen with ampicillin even though this peni- are mentioned. In an Italian study, a patient who cillin has the same R1 group as cefaclor. This was had anaphylaxis provoked by cefodizime and explained by the suggestion that the allergenic who was skin test positive to the drug was found specifi city complementary to the IgE antibodies to be skin test negative to two other cephalospo- was the R1 group “plus the remaining cephalo- rins with the same R1 group (ceftriaxone and sporin structure.” Given the absence of inhibition cefotaxime) and to two other cephalosporins with by ampicillin, it is not clear why the R1 group different but structurally closely related R1 was still assumed to be central to recognition nor groups (ceftazidime and cefuroxime). In addi- was the “remaining cephalosporin structure” tion, challenges with ceftriaxone and cefuroxime explained further. From test results on a number were negative. The most likely explanation of different individual patients, two other clear seems to be allergic recognition of the entire examples diffi cult to reconcile with the view that cefodizime molecule (as previously demonstrated

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Table 5.7 Recognition by IgE of R1 and R2 side chains on cephalosporins and interpretation of recognition patterns (refer Table 5.6 )

Recognition at R1 and R2

S R1-CONH N O R 2 COOH Important features of inhibition

Serum R 1 R 2 ﬁ ndings 3,252 Entire molecule recognized. Little CH --Cl CH NH 3 tolerance for change at R1 2 (aminobenzyl) and R2 (–Cl) (c.f. inhibitions with cefaclor and cephalexin) a

3,323 Requirement for aminobenzyl at R1 CH ->-Cl CH OCOCH >- CH NH 233 and –Cl at R2 but with some clear 2 recognition of ester group—presence of a second cross-reactive (with cefaclor) IgE with some tolerance at

R 1 and ester at R2 ? (Note inhibitions with cephaloglycin and cephalothin)a 4,679 Strong requirement of aminobenzyl CH ->--CH OCOCH Cl CH 23 3at R1 but with tolerance at R2 . NH2 Inhibition with cephaloglycina may indicate

– Preference for ester at R2 – Or a second allergenic specifi city 4,764 , , Requirement for aminobenzyl or CH CH -=-Cl CH 3 closely related structure at R1 and a NH2 NH 2 “small” group at R (–Cl or –CH ) HO CH 2 3 NH2 From Pham NH, Baldo BA. β-Lactam drug allergens: Fine structural recognition patterns of cephalosporin-reactive IgE antibodies. J Mol Recognition 1996 ;9:287. Reprinted with permission from John Wiley and Sons a See Table 5.6 with cefaclor) or recognition of the R2 side chain sideration of the contribution of other possible of cefodizime which differs from the other allergenic structures. The fi rst is the known labil- cephalosporins tested. In fi ndings from the same ity of the six- membered ring with the associated group, a patient who experienced an anaphylactic belief that R2 groups are readily lost both in vitro reaction to the third-generation cephalosporin and probably in vivo and therefore the R2 side cefoperazone was skin test positive to this drug chain cannot contribute to the molecules allerge- and to the second- generation cefamandole. These nicity. The second factor, stemming directly from two drugs have a structurally unrelated R1 side the fi rst, is the nature of the structures prepared chain but an identical 1-methyltetrazol-5-yl-sul- for the detection of cephalosporin-reactive IgE fonyl R2 group (Fig. 5.13 ). antibodies. These structures tend to comprise Therefore, although it is clear that the R1 side essentially the R1 side chain only. Unvalidated chain is considered to be a dominant (perhaps the diagnostic reagents arising from research and dominant) allergenic specifi city found on cepha- discussed above are examples of the bias of tests losporins, two factors may have infl uenced and for detection of the R1 specifi city and it remains reinforced this belief to the exclusion of any con- to be seen if efforts to couple cephalosporins via

[email protected] 5.2 Cephalosporins 173 the 4-carboxyl group retain stable complexes suit- solid phases with the cephalosporin molecule able for identifying potential allergenic specifi ci- undegraded and intact. Of course, the presence of ties on the whole cephalosporin molecule and, in IgE antibodies to a particular structure does not particular, on the R2 side chain. always accurately refl ect allergic sensitivity to that structure but, conversely, the absence of IgE 5.2.4.3.2 Does the Spectrum of Anti- (confi rmed by in vitro and in vivo techniques that Cephalosporin IgE Antibodies are suffi ciently sensitive) is invariably an indica- Refl ect the Full Range of tion of the absence of type I allergy. By combin- Cephalosporin Antigens? ing information obtained from the identifi cation As discussed earlier (Sect. 5.1.3 ), the heterogene- of the heterogeneous spectrum of IgE antibodies ity of penicillin determinants extends to the spec- complementary to determinants identifi ed on the trum of serum IgE antibodies, so rather than complete and intact cephalosporin molecule with trying to identify likely allergenic metabolites or chemical analyses and, most importantly, skin breakdown products, preparing them in the labo- testing and perhaps challenge procedures, it may ratory, and then testing them, a more comprehen- be possible to more quickly and accurately defi ne sive and effi cient approach might be to identify the range of the most important cephalosporin immunochemically the full spectrum of determi- allergenic structures. One factor that might work nants recognized by a large population of against this proposed strategy is the possibility cephalosporin- allergic patients. For drugs such that the immune response is directed to a metabo- as the cephalosporins where questions concern- lite, breakdown product, or protein adduct not ing the stability and existence of some structures detected by the employment of free, unmodifi ed remain, assessment of the complete allergenic drug. This, however, is likely to be anticipated recognition profi le of the cephalosporin-allergic and unlikely to nullify the potential success of the population seems even more logical. At present it strategy but, in any case, no cephalosporin has is clear that the relatively mild procedures that yet been found to form a hapten–protein adduct lead to opening of the β-lactam ring generally without laboratory intervention. also induce disintegration of the dihydrothiazine ring, but it is not always clear how precisely concerted this process is, whether ring breakdown occurs with a variety of other procedures, and 5.2.5 Skin Testing with Cephalosporins what is the nature and range of structures patients are exposed to in vivo following ingestion or Skin testing for allergic sensitivity to cephalospo- injection of a cephalosporin. Since immediate, rins has not been straightforward for at least three type I allergic reactions are mediated by IgE anti- main reasons—the lack of standardized and bodies, the full spectrum of cephalosporin aller- characterized cephalosporin determinants of genic determinants could, in the fi rst instance, be known composition and structure, the employ- obtained by using both direct antibody binding ment of concentrations of free drug that elicit irri- and quantitative hapten inhibition procedures. tant reactions, and the widespread impression With a cephalosporin conjugate that retains the that skin testing with these drugs produces unre- structural identity of the free drug, clear inhibi- liable results. Concentrations of cephalosporins tions obtained with free, unmodifi ed drugs and up to as high as 250 mg/ml have been used intra- carefully selected structural analogs, both free of dermally and although concentrations up to the possibility of structural lability, would be reli- 50 mg/ml of some cephalosporins were nonirri- able indicators of the structures that are comple- tant in control subjects, a concentration of 2 mg/ mentary to the combining sites of the drug-reactive ml for both prick and intradermal testing seems antibodies. For now, the obstacle to progress with to be satisfactory in terms of sensitivity and this approach is the absence of suitable stable and specifi city. Using this concentration, Romano characterized cephalosporin conjugates and drug- performed prick and intradermal skin tests on

[email protected] 174 5 β-Lactam Antibiotics

76 patients with histories of immediate reactions and ampicillin, suggesting that testing with to cephalosporins. Individual cephalosporins penicillins does not reliably predict cephalospo- tested were cephalothin, cefamandole, ceftazi- rin allergy unless the side chains of the penicillin dime, ceftriaxone, cefuroxime, and cefotaxime. and the culprit cephalosporin are similar. In other Positive responses were obtained in 60 of the words, if the side chain of the penicillin is identi- patients (78.9 %) and the authors concluded that cal or similar, skin testing with a penicillin may skin testing at a cephalosporin concentration of be of value, but skin test and IgE antibody results 2 mg/ml is a useful tool for evaluating immediate have shown that about 90 % of patients with ben- reactions to this β-lactam. zylpenicillin or amoxicillin allergy tolerate ceph- Test solutions should be freshly prepared, alosporins even when the side chains (i.e., the R1 preferably using solutions of drugs for intrave- cephalosporin side chain) on the two classes of nous injection and with all manipulations carried drugs are the same or similar. Given that skin out under sterile conditions using aseptic precau- testing with cephalosporins at nonirritant concen- tions. If powdered drug is used in pure form or trations appears to be an effective diagnostic pro- from capsules or tablets, solutions in sterile phys- cedure, there seems little logic in employing iological saline should be prepared under sterile penicillins for the purpose. Some patients respond conditions and passed through a bacterial fi lter. with a positive skin test only to the causative In the prick test, read after 15–20 min, a wheal cephalosporin and this is probably to be expected greater than 3 mm in diameter accompanied by given the fi ndings that the whole cephalosporin erythema together with a negative response to a molecule or some individual structural features saline control is considered positive. In intrader- have been identifi ed as allergenic determinants mal testing, an increase in diameter larger than (see Sect. 5.2.4.3 ). In these patients, negative skin 3 mm is considered positive. Any increase is test results with other cephalosporins appear to obtained by comparing the marked diameter of be a reliable indicator of tolerance to those ceph- the wheal at 20 min with the marked diameter of alosporins, although more investigations of this the initial injection bleb. As usual for skin test- important observation are needed. ing, a histamine-positive control should be included. An important precaution to observe in testing for immediate reactions to cephalosporins 5.2.6 Delayed Reactions is the need to keep in mind the patient’s last expo- to Cephalosporins sure to the drug since the longer that exposure, the greater the chance the test will be negative. In Like penicillins, cephalosporins can cause fact, Romano has suggested that cephalosporin delayed hypersensitivity reactions manifesting IgE-mediated hypersensitivity may be a transient mainly as maculopapular or morbilliform rashes condition. Some patients have been shown to test and delayed urticaria/angioedema. Intradermal negative 1 year after an allergic reaction to tests with delayed readings and/or patch tests penicillins and the same may be true for cephalo- have been recommended for diagnosis and prick sporins so a negative skin test result may not be testing has also been used but less often and with reliable. This also raises the questions of whether less success. In one study of 290 patients with challenge tests or a repeat skin test should be cutaneous adverse reactions to drugs, cephalo- undertaken and, if so, is resensitization of the sporins elicited positive patch tests in 4.1 % of the patient a possibility? patients. A recent study is summarized to illus- Comment should be made on what sometimes trate the diagnostic application of skin, patch, and seems like a preoccupation with employing provocation tests in the diagnosis of delayed reac- penicillins in skin tests to predict allergy to ceph- tions to cephalosporins. Skin tests were under- alosporins. Investigations have shown that <20 % taken with cefaclor, cephalexin, and cefatrizine at of patients with immediate reactions to cephalo- concentrations of 2 and 20 mg/ml, the latter con- sporins react to benzylpenicillin, amoxicillin, centration having been found to be nonirritant in

[email protected] 5.3 Monobactams 175

30 controls. Late readings of intradermal tests were taken at 48 and 72 h. For patch tests, cepha- 5.3 Monobactams losporins were used at 5 % w/w in petrolatum, occlusion time was 48 h, and readings were made These monocyclic drugs contain only the ring 24 h later, 15 min after removing the strips. structure that gives the family of β-lactam antibi- Patients who gave negative results with skin and otics their name. The prototype drug of the group patch tests were challenged with the causative is the synthetic compound aztreonam which has drug. Oral doses used were: cefaclor 500 mg, the distinctive features of a sulfate group on the cephalexin 1 g, cefatrizine 500 mg, cefi xime nitrogen of the β-lactam ring and a thiazolyl 400 mg, cefuroxime axetil 500 mg, cefprozil group in the side chain (Fig. 5.18a ). In initial 500 mg, cefpodoxime 200 mg, and ceftibuten studies to assess whether aztreonam cross-reacted 400 mg, while 1 g of cefamandole, cefazolin, cef- with other β-lactam antibiotics, Adkinson and triaxone, ceftazidime, cefotaxime, cefepime, and colleagues used penicilloyl-, cephalosporoyl-, cefonicid were each administered intramuscu- and aztreonyl-protein conjugates in solid-phase larly. Initial challenge doses were one- hundredth radioimmunoassay inhibition experiments with of the therapeutic dose. One week later, negative rabbit antibodies to the benzylpenicilloyl, cepha- responders were given one-tenth of the normal losporoyl, and aztreonyl determinates. The dose and, if the response was again negative, a results clearly demonstrated that aztreonam full dose was administered 1 week later. Of 105 showed little, if any, cross-reaction, indicating patients, only fi ve (4.7 %) had delayed reactions that the β-lactam nucleus was not being recog- with intradermal tests positive in the fi ve patients nized and it was predicted that the side chain of and patch tests positive in three. The investigators the drug, rather than the core, would be the main concluded that intradermal tests are useful for immunogenic site. This prediction was confi rmed identifying the cephalosporin responsible for when ceftazidime, a cephalosporin with a side delayed reactions and that patch tests are not rec- chain identical to aztreonam, completely inhib- ommended for cephalosporin-induced maculo- ited the rabbit anti-aztreonyl antibodies. Further papular and urticarial rashes. evidence for the lack of cross-reactivity with

a S b H2N

N H H HO H H

CONH CH3 CH NH NH S HO ON N N O O SO3H COOH O

H N c N d

HO H H O H O CH CH2-OH S C N N H O O COOH H COOH

Fig. 5.18 Structures of the monobactam aztreonam (a ), oxazolidine ring instead of the thiazolodine ring found in the carbapenems, imipenem ( b) and meropenem ( c ), and penicillins the clavam, clavulanic acid (d ). Note that clavams have an

[email protected] 176 5 β-Lactam Antibiotics penicillin determinants was seen in the failure of the latter replaced by a carbon atom. The two aztreonam–protein conjugates and free drug to most used drugs from this group are the broad react with human anti-benzylpenicilloyl IgE anti- spectrum antibiotics imipenem (Fig. 5.18b ) and bodies and in negative skin tests to aztreonam meropenem (Fig. 5.18c ). Imipenem is formulated major and minor determinants (analogous to with cilastatin, a compound without antibacterial these determinants in penicillin) in patients who activity but a specifi c inhibitor of renal dihydro- were skin test positive to penicillins. peptidase 1, the enzyme responsible for the In general, aztreonam is well tolerated in metabolism of imipenem and all other naturally patients allergic to other β-lactam antibiotics, but occurring carbapenems. Meropenem is stable to usage has shown that aztreonam occasionally the enzyme and is administered without cilastatin. causes IgE antibody-mediated reactions includ- Imipenem, especially in minor determinant form, ing urticaria, angioedema, and anaphylaxis and cross-reacts with penicillins and should therefore this may occur on fi rst exposure. Other adverse be regarded with caution before administering it reactions include rashes, erythema multiforme, to patients with suspected penicillin allergy. From eosinophilia, and gastrointestinal effects. the initial investigations by Saxon and colleagues, Sensitization to aztreonam and its cross- reactivity the frequency of cross-reactivity between imipe- with other β-lactams were assessed in high-risk nem and penicillins as defi ned by skin testing was patients with cystic fi brosis where it was well tol- about 47 % and this dropped to 25 % for subjects erated, but anaphylactic reactions in two patients reporting a penicillin allergy, that is, those with on reexposure to the drug and a 5 % skin test positive and negative skin test results. Recent ret- cross-reactivity with other β-lactam antibiotics rospective investigations claim the fi gure is about suggested that caution should be exercised in 9–11 %. In the general population, the frequency administering aztreonam to cystic fi brosis of hypersensitivity to carbapenems has been esti- patients allergic to other β-lactam antibiotics mated as less than 3 %. For patients with delayed- because it is potentially allergenic on repeated type hypersensitivity to penicillins, cross-reactivity use. A retrospective study of hypersensitivity between imipenem and other β-lactams studied reactions to aztreonam and other β-lactams in by delayed reading intradermal and patch tests is cystic fi brosis patients revealed a wide incidence said to be 5.5 %. of reactions ranging from 50.9 % for piperacillin Results from more recent prospective studies to 4 and 6.5 % to imipenem and aztreonam, have continued to show that the formerly accepted respectively. There was no discernable cross- perception of risk of administering carbapenems reactivity and the reactions to aztreonam seemed to penicillin-allergic subjects was probably an to be restricted to a small group of reactors with a overestimation. Skin testing with imipenem high propensity for hypersensitivity to β-lactams. (0.5 mg/ml) of over 100 patients each with at A cross-reaction due to the identical side chains least one positive skin test to a penicillin reagent on aztreonam and ceftazidime was detected by revealed only one positive reactor. Intramuscular positive skin tests to both drugs in a patient aller- challenges with increasing concentrations of imi- gic to aztreonam but skin test and challenge test penem–cilastatin over a 3 h period showed no negative to a number of penicillins and other reactors in the skin test-negative group. The cephalosporins. By contrast, four cystic fi brosis 0.9 % incidence of positive reactions found is patients who were allergic to ceftazidime demon- signifi cantly lower than previous estimates and strated tolerance to aztreonam. lower than the 4.4 % rate of cross-reactivity claimed for penicillins and cephalosporins. The results bring into question the practice of avoid- 5.4 Carbapenems ing imipenem in penicillin-allergic patients and indicate that prophylactic skin tests can be useful In addition to the β-lactam ring, the carbapenem in patients who require treatment with imipe- nucleus contains an unsaturated fi ve-membered nem–cilastatin. If the negative predictive value of ring as for penicillins but with the sulfur atom of skin testing with imipenem is in doubt, patients

[email protected] 5.5 Clavams 177 allergic to penicillins and skin test negative to tially cross-reactive IgE antibodies to the imipenem can be subjected to graded challenge β-lactam ring either do not occur or are rare in with the drug. Similar studies on meropenem and patients allergic to penicillins. It therefore seems imipenem in children led to similar fi ndings and likely that IgE antibodies to imipenem and conclusions. Skin testing and challenges showed meropenem generally recognize the groups that reactivity to meropenem at 1 mg/ml and pen- attached at C-6 and to the sulfur atom at C-3 of icillins in patients with IgE-mediated allergy to the bicyclic nuclear structure. penicillins was no higher than 5.2 % and the data indicated that no more than 3.5 % of patients would show a positive challenge after a negative 5.5 Clavams skin test. On the basis of these fi ndings, it was suggested that the practice of avoiding merope- Like penicillins, clavulanic acid is a bicyclic nem therapy in penicillin-allergic patients should structure with a β-lactam ring but, unlike penicil- be reconsidered. Given the much lower frequency lins, it lacks an R1 side chain and has an oxazoli- of cross-reactivity found in this study, it may be dine ring instead of the thiazolidine ring signifi cant that only the parent carbapenem was (Fig. 5.18d ). The compound has negligible anti- employed for skin testing whereas in the earlier microbial activity but binds to the active site of investigation by Saxon, imipenem metabolites β-lactamase ultimately inactivating the enzyme. It were also used. A second proviso to be kept in is therefore sometimes formulated with penicil- mind is that after challenges, patients did not lins such as amoxicillin and ticarcillin to prevent receive a full therapeutic course of carbapenem. inactivation of the antibiotic and retain antimicro- The absence of allergic cross-reactions of car- bial action. Early investigations showed that the bapenems with penicillins has also been deduced compound was poorly immunogenic and, until following conclusions formed from extensive fairly recently, it was generally assumed that it clinical experience with several hundreds patients demonstrated low or no allergenicity. Over the last over a 12-year period. Little or no cross-reactivity few years, there seems to have been an increase in was seen even in patients with a history of ana- cases of immediate hypersensitivity to clavulanic phylaxis to penicillins and who were not skin acid, leading to speculation that this is a refl ection tested. of its increasing usage with amoxicillin. The pres- The question of tolerability of penicillins, ence of the antibiotic in the combination, and the monobactams, and carbapenems in patients with limited availability and stability of the enzyme IgE hypersensitivity to cephalosporins was inhibitor, has hampered ready diagnosis of hyper- recently assessed in 98 subjects by serum IgE sensitivity to clavulanic acid. To overcome its antibody assays, challenge tests, and skin testing lability, freshly preserved solutions should be pre- with penicillin reagents, aztreonam, imipenem– pared for testing and cellulose or silica-based cilastatin, and meropenem. Approximately 25 % excipients may be added. For skin prick testing, of cephalosporin-allergic subjects were positive clavulanic acid and excipient each at 10 mg/ml to penicillins, while 3.1, 2, and 1 % showed posi- have been used, and for intradermal tests, the con- tive results to aztreonam, imipenem, and merope- centration is reduced to 1 and 0.1 mg/ml. nem, respectively. A reaction to a cephalosporin In an evaluation of 276 patients who had a with a similar or identical side chain to penicillin reaction attributed to amoxicillin–clavulanic was a signifi cant predictor of cross-reactivity. For acid, 55 patients (19.9 %) reacted positively to skin testing, the following concentrations were different penicillin determinants. Of the 221 with used: ampicillin and amoxicillin 1 and 20 mg/ml; negative skin tests, 15 were positive to amoxi- cephalosporins 2 mg/ml; aztreonam 2 mg/ml; cillin and seven were judged to be allergic to imipenem–cilastatin 0.5 mg/ml; and meropenem clavulanic acid on the basis of tolerance to 1 mg/ml. benzylpenicillin and amoxicillin and an immedi- From the relatively few investigations carried ate reaction to clavulanic acid following chal- out on the carbapenems, it seems clear that poten- lenges. In the skin test-positive group, skin and

[email protected] 178 5 β-Lactam Antibiotics challenge tests showed negative reactions to ben- • Penicillins have long been known to be the zylpenicillin reagents and amoxicillin but posi- most common cause of drug-induced allergic tive skin tests to clavulanic acid in 16 patients. reactions including anaphylaxis. For skin testing, clavulanic acid was used at a • Penicillins can cause all four types of hyper- concentration of 20 mg/ml and amoxicillin–cla- sensitivity responses provoking type I vulanic acid at 20 mg/ml amoxicillin and 4 mg/ IgE- mediated reactions such as urticaria, ml clavulanic acid. This difference in concentra- angioedema, asthma, and anaphylaxis; type II tions between test solutions of clavulanic acid antibody-mediated hemolytic anemia and used alone or in combination with amoxicillin thrombocytopenia; type III immune complex- may explain why more positive skin test reac- mediated serum sickness-like reactions and tions to the clavam have not been detected. Even vasculitis; and type IV T cell-mediated so, the fi nding that 29 % of immediate allergic contact dermatitis, rashes, and other skin reactions to the amoxicillin–clavulanic combina- eruptions. tion are directed to clavulanic acid is surprisingly • Investigations of the formation of antigenic high. As well as skin tests, detection of CD63 and allergenic determinants of benzylpenicil- expression by basophils in the basophil activation lin have identifi ed a list of determinants that test using fl ow cytometry and sulfi doleukotriene includes the penicilloyl, penicillenate, peni- release by basophils in the cellular antigen stimu- cilloic acid, penilloic acid, penamaldate, lation test (CAST) have been used to demonstrate penicoyl, penicillanyl, and penicillamine positive allergic responses to amoxicillin–clavu- structures. lanic acid and clavulanic acid alone and negative • The side chain of penicillins plays a large part responses to amoxicillin alone and other β-lactam in the specifi city of immunological reactions antibiotics. to the drugs. Allergic recognition of side chain Hepatitis and jaundice caused by the combi- determinants highlights the importance of nation of amoxicillin and clavulanic acid were including different individual penicillins in fi rst identifi ed in 1988. The combination of the the battery of penicillin skin test reagents. two drugs is associated with a higher incidence of • IgE antibodies in the sera of patients allergic liver injury than the administration of amoxicillin to β-lactam antibiotics detect a spectrum of alone. The risk of this drug-induced liver injury, antigenic specifi cities and IgE in the sera of mostly cholestatic in nature, increases with age different allergic patients shows heteroge- and by about a factor of 3 after 2 or more con- neous recognition and cross-reactive secutive courses of drug. responses. Some antibodies recognize discrete There are at least three reports of delayed regions of the penicillin molecule such as the reactions to clavulanic acid manifesting as gener- side chain only, or the thiazolidine ring only, alized urticaria, generalized itchy erythema, and while others show combining sites comple- allergic contact dermatitis with pruritic erythem- mentary to compound structures made up of atous patches. Diagnoses were confi rmed by the side chain with the β-lactam ring, the com- patch testing with amoxicillin–clavulanic acid bination of the β-lactam and thiazolidine dried syrup at 10 and 1 mg/ml and clavulanic acid rings, or the whole molecule. 10 mg/ml and for one patient, by skin prick and • IgE may also distinguish fi ne structural fea- intradermal tests. tures on different penicillins, for example, IgE antibodies in the sera of allergic patients that distinguished amoxicilloyl and amoxicillanyl Summary determinants. • Patients with a history of prior reactions to • The β-lactam antibiotics comprise four main penicillins have a four- to sixfold increased classes of drugs that possess antibacterial action, risk of a reaction to a penicillin compared to viz., penams (penicillins), cephems (cephalo- those without a previous history of hypersen- sporins), monobactams, and carbapenems. sitivity to the drugs.

[email protected] Summary 179

• In September 2009, full regulatory approval • The starting dose for penicillin desensitization was granted for penicilloyl-polylysine (Pre- is commonly about one ten-thousandth or less Pen® ) by the FDA. In Europe, a new commer- of a full therapeutic dose. Starting with a dose cial kit containing both the major determinant of 0.05 mg, doubling doses are given at 15 min (5 × 10 −5 M) and minor determinant mix (each intervals in 14 steps up to a maximum dose of component at a maximum concentration of 400 mg and a cumulative dose of 800 mg 2 × 10 −2 M) is now available. In September before administering the full therapeutic dose 2011, Pre-Pen® was approved by Health of the drug 30 min after the last desensitizing Canada. In 2011, an agreement with global dose. distribution rights was reached for marketing • Delayed-type, non-IgE-mediated hypersensi- the major determinant together with a minor tivity, often manifesting as macular or macu- determinant mix currently under development lopapular exanthemata, may occur during in the USA. treatment with penicillins, particularly amino- • For those with a negative history of penicillin penicillins. Other delayed reactions include allergy, the incidence of positive skin tests is acute generalized exanthematous pustulosis, 2–7 %. For skin test-positive patients given a delayed urticaria/angioedema, exfoliative der- therapeutic dose of penicillin, the risk of an matitis, and the more severe bullous exan- acute allergic reaction ranges from 10 % in thems, Stevens–Johnson syndrome, and toxic patients with a negative history to 50–70 % in epidermal necrolysis. Severe systemic hyper- patients with a positive history. sensitivity responses include vasculitis, hepa- • The commercially available Phadia titis, interstitial nephritis, and pneumonitis. ImmunoCAP® drug-solid phases detect IgE Penicillin-induced drug reaction (rash) with antibodies to penicilloyl G and V, amoxicil- eosinophilia and systemic symptoms (DRESS) loyl, and ampicilloyl determinants. These also occurs. assays measure specifi c IgE antibodies in the • Intradermal tests with delayed reading, patch range 0.01–100 kUA/l with a cutoff value of tests, and occasionally prick tests provide the 0.35 kUA/l for a positive result. Levels higher mainstay diagnostic procedures for the evalu- than 0.1 kUA/l, indicate sensitization to the ation of delayed reactions to penicillins and drug. other β-lactam drugs. • When applied to the diagnosis of immediate • Investigations on benzylpenicillin-specifi c T hypersensitivity to β-lactams, the basophil cell clones showed that processing of the free activation test demonstrated a sensitivity of drug was not required whereas benzylpenicil- about 50 % and specifi city of ~90–100 %. loyl–HSA conjugate must undergo processing A high incidence of false positives has been to stimulate T cell clones specifi c for this observed. determinant. • Challenge (provocation) testing should be per- • The HLA-B*57:01 genotype is a major deter- formed only after prior skin testing and prefer- minant of drug-induced liver injury due to ably a drug-specifi c IgE antibody test. If either fl ucloxacillin. of these tests returns a positive result that is in • There is a signifi cantly higher risk of a reac- accordance with the patient’s history, the risk tion to a cephalosporin in patients already precludes provocation testing. In the fi rst allergic to penicillin. Second- and third- instance, skin and IgE testing should be under- generation cephalosporins appear to carry less taken with benzylpenicillin and, if this is posi- risk of provoking a reaction. tive, the patient should be considered to be • Aminolysis of cephalosporins in the presence allergic to the β-lactam group of drugs. If of polylysine or protein produces unstable testing with the parent penicillin is negative, intermediates that decompose to penaldate the patient is then tested with the drug that and ultimately penamaldate structures. This provoked the reaction if it is known. results in structures in which only the R1 side

[email protected] 180 5 β-Lactam Antibiotics

chain, the attached amide, and remnants of the • The 0.9 % incidence of positive reactions β-lactam ring remain from the original cepha- found for imipenem is signifi cantly lower than losporin molecule. Immunochemical studies previous estimates and lower than the 4.4 % revealed clear IgE antibody recognition of rate of cross-reactivity claimed for penicillins cephalosporin R1 side chains and cross- and cephalosporins. Skin testing and chal- recognition of β-lactam R1 groups showing lenges showed that reactivity to meropenem structural similarity. The allergenic impor- and penicillins in patients with IgE-mediated tance of cephalosporin R1 side chains was fur- allergy to penicillins was no higher than 5.2 % ther supported by IgE recognition of synthetic and the data indicated that no more than 3.5 % hapten structures. of patients would have a positive challenge • With some cephalosporin-allergic patients, in after a negative skin test. On the basis of these addition to recognition of the R1 structure, the fi ndings, it has been suggested that the prac- R2 side chain and/or the whole cephalosporin tice of avoiding imipenem and meropenem molecule are recognized by serum IgE anti- therapies in penicillin-allergic patients should bodies. This was demonstrated by employing be reconsidered. drug-solid phases prepared by linking HSA to • It seems likely that IgE antibodies to imipe- the carboxyl group at position 4 on the cepha- nem and meropenem generally recognize the losporin nucleus. groups attached at C-6 and to the sulfur atom • A cephalosporin concentration of 2 mg/ml for at C-3 of the bicyclic nuclear structure. both prick and intradermal testing seems to be • There appears to have been an increase in satisfactory in terms of sensitivity and speci- cases of immediate hypersensitivity to fi city. Less than 20 % of patients with immedi- clavulanic acid, leading to speculation that ate reactions to cephalosporins react to this is a refl ection of its increase in usage with benzylpenicillin, amoxicillin, and ampicillin, amoxicillin. suggesting that testing with penicillins does • There are a small number of reports of delayed not reliably predict cephalosporin allergy reactions to clavulanic acid manifesting as unless the side chains of the penicillin and the generalized urticaria, generalized itchy ery- culprit cephalosporin are similar. thema, and allergic contact dermatitis with • Cephalosporins can cause delayed hypersensi- pruritic erythematous patches. tivity reactions manifesting mainly as maculo- • The combination of amoxicillin and clavu- papular or morbilliform rashes and delayed lanic acid is associated with the risk of drug- urticaria/angioedema. Intradermal tests are induced liver injury, mostly cholestatic in useful for identifying the cephalosporin nature. responsible for delayed reactions. Patch tests are not recommended for cephalosporin- induced maculopapular and urticarial rashes. Further Reading • The monobactam aztreonam shows little, if any, cross-reaction with penicillins and cepha- Baldo BA, Pham NH, Weiner J. Detection and side-chain losporins, indicating that the β-lactam nucleus specifi city of IgE antibodies to fl ucloxacillin in aller- is not recognized. The side chain of the drug, gic subjects. J Mol Recognit. 1995;8:171–7. not the core, is the main immunogenic site. Daly AK, Donaldson PT, Bhatnagar P, et al. HLA-B*5701 genotype is a major determinant of drug-induced • Caution should be exercised in administering liver injury due to fl ucloxacillin. Nat Genet. 2009;41: aztreonam to cystic fi brosis patients allergic to 816–9. other β-lactam antibiotics because it is poten- Dewdney JM. Immunology of the antibiotics. In: Sela M, tially allergenic on repeated use. editor. The antigens, vol. 4. New York, NY: Academic; 1977. p. 73–228. • The two most used carbapenems are the broad De Weck AL. Penicillins and cephalosporins. In: De spectrum antibiotics imipenem and meropenem. Weck AL, Bundgaard H, editors. Allergic reactions to

[email protected] Further Reading 181

drugs, Handbook of experimental allergology. Berlin: Parker CW. Drug allergy. In: Parker CW, editor. Clinical Springer; 1983. p. 423–82. immunology, vol. 2. Philadelphia, PA: Saunders; 1980. Faraci WS, Pratt RF. Elimination of a good leaving group p. 1219–60. from the 3′-position of a cephalosporin need not be Pham NH, Baldo BA. β-Lactam drug allergens: fi ne struc- concerted with β-lactam ring opening: TEM-2 tural recognition patterns of cephalosporin-reactive β-lactamase-catalysed hydrolysis of pyridine-2-azo- IgE antibodies. J Mol Recognit. 1996;9:287–96. 4′-(N′, N′-dimethylaniline) cephalosporin (PADAC) Romano A, Gaeta F, Valluzzi RL, et al. IgE-mediated and of cephaloridine. J Am Chem Soc. 1984;106: hypersensitivity to cephalosporins: cross-reactivity 1489–90. and tolerability of penicillins, monobactams, and car- Hamilton-Miller JMT, Newton GGF, Abraham EP. bapenems. J Allergy Clin Immunol. 2010;126:994–9. Products of aminolysis and enzymic hydrolysis of the Romano A, Guéant-Rodriguez RM, Viola M, et al. cephalosporins. Biochem J. 1970;116:371–84. Diagnosing immediate reactions to cephalosporins. Harle DG, Baldo BA. Identifi cation of penicillin aller- Clin Exp Allergy. 2005;35:1234–42. genic determinants that bind IgE antibodies in the sera Salkind AR, Cuddy PG, Foxworth JW. The rational clini- of subjects with penicillin allergy. Mol Immunol. cal examination. Is this patient allergic to penicillin? 1990;27:1063–71. An evidence-based analysis of the likelihood of peni- Levine BB. Immunochemical mechanisms of drug allergy. cillin allergy. JAMA. 2001;285:2498–505. In: Miescher PA, Muller-Eberhard HJ, editors. Zhao Z, Baldo BA, Baumgart KW, et al. Fine structural Textbook of immunopathology, vol. 2. 2nd ed. New recognition specifi cities of IgE antibodies distinguish- York, NY: Grune and Stratton; 1976. p. 403–19. ing amoxicilloyl and amoxicillanyl determinants in Montanez MI, Mayorga C, Torres MJ, et al. Synthetic allergic subjects. J Mol Recognit. 2001;14:300–7. approach to gain insight into antigenic determinants of Zhao Z, Batley M, D’Ambrosio C, et al. In vitro reactivity cephalosporins: in vitro studies of chemical structure- of penicilloyl and penicillanyl albumin and polylysine IgE molecular recognition relationships. Chem Res conjugates with IgE-antibody. J Immunol Methods. Toxicol. 2011;24:706–17. 2000;242:43–51.

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Abstract Allergic reactions to commonly used antibiotics such as the tetracyclines, macrolides, and rifamycins are rare. Neomycin consistently ranks in the top 10 % of the most common causes of allergic contact dermatitis while the incidence of bacitracin allergy is in the range 8–9.5 %. The most commonly occurring adverse effects caused by vancomycin are termed, collectively, red man syndrome. Immediate type I allergy is the most well-defi ned sulfonamide-induced hypersensitivity reaction with the best defi ned allergenic drug structures. Antigen-presenting cells may produce sulfonamide hapten–protein antigen complexes and ultimately induce the T cell response to the drug. Cross-reactions with a wide variety of frequently used non- antibacterial drugs containing a common sulfonamide group are thought not to occur. Almost all reports of trimethoprim-induced hypersensitivities are of the immediate kind and three different trimethoprim allergenic determinants structures have been identifi ed. IgE antibodies apparently specifi c for quinolones and positive skin tests in apparently normal, healthy controls have been demonstrated. Immediate type I mechanisms are responsible for the most important chlorhexidine-induced allergic reactions. The whole molecule has been identifi ed as the allergenic determinant. In almost all cases of povidone–iodine hypersensitivity, polyvinylpyrrolidone not iodine has been implicated as the offending component.

For both the clinician and patient, an allergic provoke a spectrum of immediate and delayed reaction elicited by a β-lactam antibiotic is prob- allergic responses. Although collectively this ably considered the archetypal drug allergy but a group of drugs distinct from the β-lactams is range of other antibiotics, both naturally occur- large and chemically diverse, an attempt is made ring and semisynthetic, and a number of other in this chapter to consider them under the broad antimicrobials of varied origin and nature also heading of other antimicrobials.

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 183 Relationships, DOI 10.1007/978-1-4614-7261-2_6, © Springer Science+Business Media, LLC 2013

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There have been times when some non-β-lactam 6.1 Antibiotics antibiotics were described as being implicated in hypersensitivity reactions with moderate fre- The defi nition we will use here for an antibiotic quency. This was true for both streptomycin and (see also Chap. 5 ) is a chemically defi ned sub- gentamycin, but their decreasing usage has seen stance produced by, or derived from, certain bac- reactions to these drugs decline signifi cantly. The teria, fungi, or other organisms (not necessarily same decline related to usage has been seen with microorganisms), or produced semi-synthetically, kanamycin, lincomycin, and polymyxins which that can, in low concentrations, destroy or inhibit were implicated in reactions occasionally but the growth of other microorganisms. This defi ni- which are now rarely, if ever, a problem. By con- tion excludes chemotherapeutic antibacterials trast, bacitracin, once thought of as an offender of such as the sulfonamides that appear to be increas- low incidence, has now become recognized as the ingly referred to as antibiotics. Antibiotics seem largest cause of hypersensitivity reactions to have always been around and although our induced by drugs used topically. With this big immediate thoughts about them are positive and increase in cases there has also been an alarming we naturally think of them as one of the mainstays number of reports of anaphylaxis, something that of chemotherapy for infections of man and ani- could surely not have been anticipated. In rela- mals of all sorts, the nagging warnings of their tion to anaphylaxis, recently published large epi- overuse and consequent loss of effectiveness demiologic surveys from France of drugs causing against some major life-threatening bacteria are a anaphylaxis during anesthesia showed that sobering reminder that these agents can never be despite the number of reactions to neuromuscular taken for granted. In fact, antibiotics have been blocking drugs and latex (the agents responsible used for about 70 years during which time they for the large majority of cases) remaining rela- have revolutionized the treatment of infectious tively stable, the incidence of anaphylaxis to anti- diseases, but, in addition to loss of effectiveness biotics has increased signifi cantly during the that results from their selection of resistant organ- same time period. isms, like many drug, they have provoked a cata- log of adverse reactions from transient rashes to life-threatening anaphylaxis. 6.1.1 Macrolides In terms of usage, no other antibacterial agent rivals the prolonged and heavy prescribing of Erythromycin, from the actinomycete Saccharo- penicillins and cephalosporins so it might be polyspora (formerly Streptomyces ) erythraea, is expected that adverse reactions to non-β-lactam the fi rst member of this family of antibiotics to be antibacterials would be encountered less often marketed and successfully used clinically to treat and this appears to be the case. The word infections in humans. It has an antimicrobial “appears” has been used deliberately here; com- spectrum at least as wide as the penicillins, and paring, for example, the number of reports of interestingly, from our perspective, it is often allergic reactions to clindamycin and amoxicillin used as a replacement for patients allergic to that draws attention to the need for a standardized group of drugs. Besides erythromycin, other method of comparison and this ultimately members of the macrolide family of antibiotics depends on the number of reactions for a speci- that are clinically useful include azithromycin, fi ed number of administrations. Calculation and clarithromycin, dirithromycin, roxithromycin, comparisons of such frequencies of reactions are telithromycin (these six are approved by the not often done, but, even so, the sparse reports of FDA), oleandomycin, and spiramycin. Clarith- allergic reactions to commonly used antibiotics romycin, dirithromycin, and roxithromycin and such as the tetracyclines and macrolides and fre- the azalide azithromycin are more recent mem- quently used antimicrobials such as chlorhexi- bers of the group and can be regarded as newer dine do indeed suggest that the reactions are rare. generation macrolide antibiotics.

[email protected] 6.1 Antibiotics 185

a shows the structure of the 14-membered lactone O erythromycin. Changes to its structure, sometimes small, produce other useful members of H3C CH3 the family. The structure of azithromycin, now one of the most heavily used antibiotics, has a HO OH H3C OH CH3 nitrogen atom in the 15-membered macrolide H3CCH3 N H C ring making it an azalide (Fig. 6.1b ). 3 HO O C2H5 O O CH3 6.1.1.2 Macrolide Hypersensitivities

O O OCH3 Allergies to macrolide antibiotics are said to occur with an incidence of 0.4–3 %. Apart from CH3 CH3 OH the very occasional case report of anaphylaxis, O CH3 the most commonly seen symptoms of a hyper- Erythromycin sensitivity reaction to this group of drugs include urticaria, often generalized, angio- b edema, pruritus, asthma, tachycardia, and delayed cutaneous reactions presenting as H3C CH3 N maculopapular exanthema. Stevens–Johnson HO syndrome and toxic epidermal necrolysis have OH H3C HO CH3 CH N 3 been observed rarely. Drug-reactive serum IgE CH3 H C H C antibodies to erythromycin inhibited by the 3 3 HO O O O CH3 drug have been demonstrated in at least one C2H5 case and the Prausnitz–Kustner test was shown OOOCH3 to be positive in a case of anaphylaxis following ingestion of a single dose of the drug. The CH3 CH3 OH presence of macrolide-reactive IgE antibodies O CH3 in some patients has also been inferred from Azithromycin positive prick test results, in particular, in studies involving spiramycin and roxithromycin. Fig. 6.1 Structure of the macrolide antibiotic erythromycin (a ) which contains a 14-membered lactone ring. The Skin prick tests were also utilized to examine azalide, azithromycin ( b ), derived from erythromycin, has cross-reactivities of spiramycin and roxithro- an N -methyl group incorporated into the lactone ring, mycin each with clarithromycin and erythro- making it a 15-membered ring (compare highlighted mycin. Cross-reactivity was absent in the sections) former case but present in the latter. A number of cases of asthma to spiramycin, generally in an occupational setting in pharmaceutical 6.1.1.1 Structures plants, have been reported and confi rmed by Chemically, the macrolide antibiotics comprise a one or more of skin tests and/or challenge large cyclic lactone ring structure with one or more (nasal or inhalation) tests. An anaphylactoid molecules of the unusual sugars, L- cladinose and reaction to telithromycin, a semisynthetic D-desosamine, attached by glycosidic linkage. derivative of erythromycin and the fi rst ketolide The lactone ring structure may be from 12- to antibiotic to be used clinically, was notable 16-membered—clarithromycin, dirithromycin, because of the symptoms of shortness of breath, erythromycin, oleandomycin, roxithromycin, and wheezing, and angioedema that occurred after telithromycin each have a 14-membered lactone the fi rst dose even though the patient had taken ring while azithromycin is a 15-membered and erythromycin and azithromycin in the past spiramycin a 16-membered macrolide. Figure 6.1a without adverse effects.

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6.1.1.3 Diagnosis of Macrolide reexamination of the approach to the diagnosis of Hypersensitivities macrolide hypersensitivity are hard to disagree The foregoing brief summary indicates the gaps with and yet the surprisingly small numbers of in our knowledge of macrolide hypersensitivities, patients with true macrolide allergy distinguished gaps brought about not only by the paucity of by provocation tests and excluded by skin and patients to study but also by the lack of studies laboratory tests might be as much a refl ection of designed to investigate the underlying mecha- the starting cohort of patients as an indication of nisms, our ignorance of the allergenic structural the inappropriateness of applied tests. Starting determinants and the absence of good clinical with patients referred on by general practitioners follow-up. For the macrolides, skin tests, prick, and other clinicians, many of whom are not aller- intradermal, and patch, are said to yield positive gists, almost certainly means that the initial results in only 25–50 % of patients and this brings applied diagnostic criteria will be varied, perhaps into focus the question of which examinations highly so, and perhaps based on no more than the are of value in confi rming a diagnosis of macro- temporal relationship already referred to. It lide allergy. Skin testing in cases of suspected seems likely that the allergological assessments allergies to erythromycins is poorly standardized undertaken by the general practitioners will be as acknowledged in a recent study recommend- less thorough and appropriate than those under- ing 0.05 mg/ml as a suitable nonirritating con- taken by the allergy specialists. Under these cir- centration of clarithromycin for intradermal cumstances it seems hasty to conclude that skin testing. Erythromycin-reactive IgE antibodies and in vitro tests are not helpful in establishing a specifi cally inhibited by the antibiotic and cross- diagnosis. This would appear to be true for all reactive with the structurally related diace- allergens if the patients were poorly or inappro- tylmidecamycin were demonstrated with a solid priately assigned in the fi rst place. phase immunoassay prepared by reacting erythromycin lactobionate with bis-oxirane-activated Sepharose. 6.1.2 Tetracyclines Recent studies designed to test what was seen as the possible uncritical assignation of macro- 6.1.2.1 Adverse Reactions lide hypersensitivity concluded that the diagnosis Even though minocycline has been identifi ed was often misinterpreted and lacked the neces- with a number of serious adverse drug reactions sary confi rmatory data. Essentially, a diagnosis including hypersensitivity and there are a small was often said to be made without diagnostic number of reports of anaphylaxis to tetracyclines, evaluation and based on nothing more than a sug- this family of broad spectrum antibiotics, devel- gestive history of a temporal relationship between oped in the 1940s, is seen as being comparatively macrolide intake and symptoms. For the moment safe, especially when viewed against their long- at least, the conclusion seems to be that history term and frequent usage. Dermatologists fi rst alone has been too often used in the diagnosis of prescribed tetracyclines in the early 1950s, when macrolide hypersensitivity and the number of it became apparent that the drugs offered an patients so classifi ed is a healthy overestimate. In effective treatment for acne vulgaris. Minocycline, addition, skin and in vitro tests such as serum IgE now widely used for this purpose, has been antibody determination, the basophil activation implicated in a serum sickness-like reaction, test, lymphocyte transformation tests, and the drug-induced lupus, cases of single organ dys- tryptase assay were judged to be not very useful function, and the so-called drug hypersensitivity in identifying hypersensitive patients. The chal- syndrome reaction, a severe adverse drug reaction lenge test alone, regarded as “the gold standard,” that may occur following a number of different is advocated as necessary to defi nitely confi rm or medications including anti-epileptic drugs, sul- rule out allergy to macrolides. The design and fonamides, dapsone, azathioprine, allopurinol, logic of the thinking and the studies behind this and cyclosporin. A variety of clinical abnormalities

[email protected] 6.1 Antibiotics 187 may be seen in drug hypersensitivity syndrome, OHO OH O O in particular, fever, skin lesions, lymphadenopathy, OH 10 11 12 1 and internal organ involvement. Skin reactions 9 2 NH2 8 3 reported include exudative maculopapules, pur- 7 6 5 4 OH puratous macules, and erythema multiforme- like HH R R N plaques and other reported effects on organs R4 R3 2 1 H C CH include eosinophilia, agranulocytosis, atypical 3 3 lymphocytosis, autoimmune hyperthyroidism, -R1 -R2 -R3 -R4 hepatitis, nephritis, and myositis. In important Tetracycline -H -CH -OH -H studies from Canada, 19 cases of hypersensitivity 3 Oxytetracycline -OH -CH -OH -H reaction were assessed to be due to minocycline, 3 Doxycycline two to tetracycline, and one to doxycycline. Of -OH -CH3 -H -H 16 cases of serum sickness-like reaction, 11 were Demeclocycline -H -H -OH -Cl due to minocycline, 3 to tetracycline, and 2 to Minocycline -H -H -H -N(CH ) doxycycline. For single organ dysfunction, 40 3 2 were caused by minocycline, 37 by tetracycline, Fig. 6.2 Structures of tetracyclines used clinically. Note and 6 by doxycycline, and all 33 cases of that all four compounds contain a 4-dimethylamino group drug- induced lupus were found to be due to but minocycline has a second one, raising the possibility minocycline. Despite the infrequency of these of allergenic bivalency reactions, the large numbers of subjects, especially teenagers, taking minocycline is a reminder doxycycline. In addition, with minocycline, the for the ongoing need to remain aware of possible presence of a second dimethylamino group at adverse reactions with this drug. position 7 (position R 4 in Fig. 6.2 ) raises the possibility of allergenic bivalency, that is, the pres- 6.1.2.2 Structural Considerations ence of two potentially reactive determinants It has been suggested that these serious adverse that might not only bind to their complementary effects of minocycline might be due to a reactive IgE antibody combining sites but also cross-link metabolite(s), but this concept remains vaguely the antibodies on the surfaces of mast cells and defi ned and so far unsupported by experimental basophils. In other words, the free drug might fi ndings. When the structures of the clinically elicit the direct release of infl ammatory media- used members of the family are examined tors without the presumed necessary binding to a (Fig. 6.2 ), one feature in particular, the carrier to form an antigenic complex. The sub- 4-dimethylamino group present in all four com- ject of immunological recognition of allergeni- pounds shown, stands out as a likely immuno- cally bivalent drugs is discussed in greater detail logically reactive substituent. IgE antibody in Sect. 7.4.2.3 . recognition of tertiary and quaternary substituted ammonium groups is well known (Sects. 7.4.2 and 8.5) and reaction of tetracycline and doxycy- 6.1.3 Rifamycins cline with IgE in the sera of subjects with multiple allergic drug recognition profi les has been Rifamycins belong to a group of antibiotics some demonstrated. Drug interaction with IgE was of which are biosynthesized in fermentation cul- shown to be due to antibody combining site rec- tures by the bacterium Amycolatopsis rifamyci- ognition of tertiary and quaternary mono-, di-, nica while some are semisynthetic compounds and trialkyl groups, but only if the alkyl groups prepared by derivitization. The name of the bac- were “small,” namely, methyl or perhaps ethyl. terium that produces rifamycins has evolved from Using such haptens immobilized on a solid the original Streptomyces mediterranei to phase, IgE antibodies have also been detected in Nocardia mediterranei , then to Amycolatopsis sera from subjects with a suspected allergy to mediterranei, and in 2004 to A. rifamycinica .

[email protected] 188 6 Other Antimicrobial Drugs

CH CH 3 3 epidermal necrolysis. The FDA has warned that HO rifampicin should not be given intermittently and

H3C the daily dosage regimen should not be inter- O OH H O rupted since renal hypersensitivity reactions may CH3 H3C O OH OH CH occur when therapy is resumed. Rifamycin SV O 3 NH H3C and rifampicin have provoked a number of cases CH 3 of anaphylaxis, usually following local adminis- O R tration, and the presence of IgE antibodies in OH O some patients has been indicated by one or more O of a positive skin prick, Prausnitz–Kustner test, CH3 basophil activation, and/or a radioallergosorbent -R test. For rifampicin, an irritant-free intradermal Rifamycin SV -H test concentration of 0.002 mg/ml has been rec-

Rifampicin -CH=N-N N-CH ommended. In one procedure used for a rifamy- 3 cin radioallergosorbent test, rifamycin SV was Rifapentine -CH=N-N N conjugated to poly- L-lysine before employment of reductive amination to reduce the Schiff base. Fig. 6.3 Structures of rifamycin SV, a rifamycin B struc- The drug–carrier conjugate was then covalently ture, and starting point for rifampicin which has a coupled to activated cellulose and used in an 4-methyl-1-piperazinaminyl side chain. The semisynthetic rifapentine is also used clinically immunoassay to detect serum IgE antibodies in patients’ sera. Immunochemical identifi cation of IgE antibody-binding fi ne structural determi- Fermentation produces fi ve chemically different nants has not yet been attempted, but investiga- but structurally closely similar rifamycins desig- tions over 35 years ago in Italy identifi ed nated A, B, C, D and E. Rifamycin SV is a deriv- rifamycin SV-reactive IgE antibodies in the sera ative of the B structure and is the starting point of patients allergic to rifampicin. The antibodies for the formation of rifampicin that has an added recognized the chromophoric nucleus of the rifa- 4-methyl-1-piperazineaminyl side chain. The mycin molecules but not the 4-methylpiperazine chemical structures of rifamycin SV and rifampi- side chain (Fig. 6.3 ). The only other experimental cin are shown in Fig. 6.3. Three other semisyn- fi ndings relevant to the allergenic structures of thetic rifamycins, rifabutin, rifapentine, and rifamycins were the recent demonstration of rifaximin are also used clinically. The rifamycins cross-reacting IgE antibodies in the serum of a are broad spectrum antibiotics used to treat tuber- patient who experienced an anaphylactic reaction culosis, gonorrhea, leprosy, and respiratory and to rifamycin SV solution applied to a wound and biliary tract infections. They are active against an immediate response involving urticaria and Helicobacter pylori and used for anti-infective dyspnea following the ingestion of a tablet of prophylaxis against meningococcal infection. rifaximin. Rifamycin SV and rifampicin reacted Other important dosage forms are as eye drops directly with the serum IgE antibodies while for the treatment of infectious conjunctivitis and rifabutin, rifapentine, and rifaximin all demon- as local applications to treat infected surgical and strated signifi cant inhibition of antibody binding traumatic wounds. to a rifampicin–Sepharose solid phase. Adverse reactions to rifamycins suggested to be immune mediated include a “fl u”-like syndrome, acute renal failure, hemolytic anemia, and 6.1.4 Vancomycin and Teicoplanin thrombocytopenia. Other more typical manifestations of hypersensitivity include urticaria, con- 6.1.4.1 Structures tact dermatitis, erythema multiforme, vasculitis, Vancomycin, a tricyclic glycopeptide antibiotic and rarely Stevens–Johnson syndrome and toxic (Fig. 6.4a ) produced by Amycolatopsis (formerly

[email protected] a HO OH HO O H2N HO O O OCl H C 3 OO CH3

HO OH Cl O O O H H O N N NHCH3 N N N H H H HN O O CONH2 HOOC

OH HO OH Vancomycin

O b HN C R HO HO OCl HO O OH OO HO O HO O NHAc Cl O O O H H O N N N N N NH2 H O H H HN O HOOC O HO OH OH HO O OH O OH OH OH

Teicoplanin

R side chain

T Teicoplanin A2-1

T Teicoplanin A2-2

T Teicoplanin A2-3

T Teicoplanin A2-4

T Teicoplanin A2-5

Fig. 6.4 Vancomycin (a ) is a tricyclic glycopeptide anti- copeptide, is a mixture of several compounds sharing the biotic. The aglycone consists of a heptapeptide connected same glycopeptide core. Three sugar moieties are present, by six peptide bonds with several non-proteinogenic N -fattyacyl-β- D -glucosamine, N -acetyl-β- D -glucosamine, amino acids, ﬁ ve of which are aromatic. D -Glucose and and D -mannose. The common core structure shared by the the novel amino sugar vancosamine are linked to the agly- two antibiotics is highlighted in bold cone through glycosidic bonds. Teicoplanin ( b), a lipogly-

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Streptomyces and then Nocardia ) orientalis , is often thought of as the “drug of last resort” for the treatment of pathogens such as methicillin- resistant Staphylococcus aureus . The aglycone part of the vancomycin structure consists of a heptapeptide connected by six peptide bonds with several non-proteinogenic amino acids, ﬁ ve of which are aromatic. The sugars present, D- glucose and the novel amino sugar vancosamine, are linked to the aglycone through glycosidic bonds and are thought to contribute to ligand binding and a consequent enhancement of antimicrobial activity. Teicoplanin, a lipoglyco- peptide, is a mixture of several compounds sharing the same glycopeptide core. Three sugar moieties are present, N -fattyacyl- β- D-glucosamine, N -acetyl- β-D -glucosamine, and D -mannose (Fig. 6.4b ).

6.1.4.2 Red Man Syndrome The most commonly occurring and well-known adverse effects caused by vancomycin are referred to collectively as red man (or neck) syndrome. In terms of severity, reactions may range from mild Fig. 6.5 Red man syndrome on the upper part of the body pruritus, erythema, and fl ushing of the upper body following vancomycin administration (photograph courtesy of Dr. F. C. K. Thien and Department of Dermatology, (Fig. 6.5 ) to angioedema and rarely hypotension Alfred Hospital, Melbourne) and cardiovascular collapse. Patients often complain of diffuse itching with a burning feeling and experience headache, dizziness, chills, fever, and and Sect. 7.4.3.2 ). The incidence of reactions has general discomfort. Signs and symptoms usually been estimated to be from 3.7 to 47 % of patients occur 5–10 min after the commencement of infu- with the most severe reactions occurring in those sion of the drug or they may become apparent after less than 40 years old, particularly in children. Oral infusion is complete. The visual signs of the vanco- administration of vancomycin led to red man syn- mycin-induced reactions, the known pharmaco- drome in a neutropenic child with normal renal logical effects of histamine, demonstrations that function and it has been claimed that between 50 the antibiotic causes degranulation of rat peritoneal and 90 % of normal and infected adults might have mast cells, and the absence of drug-reactive IgE a reaction to the drug. Following administration of antibodies all indicate that red man syndrome is 1 g of vancomycin over 1 h to healthy volunteers, caused by the release of histamine as a result of 80–90 % showed signs and symptoms of red man degranulation of mast cells and basophils. A further syndrome. Patients being treated for infections indication that the reactions are anaphylactoid appear to have a lower and less severe reaction rate. rather than immune mediated, that is, anaphylactic, It became apparent that appearance of reac- is the demonstration that tryptase levels are gener- tions, and their severity, were subject to both the ally not increased during vancomycin-induced ana- dose of vancomycin and the rate of its infusion. phylactoid reactions although the drug induces A clinical study involving the measurement of tryptase release from mast cells in vitro. Note, plasma histamine levels every 10 min during the however, that this fi nding does not correlate with in fi rst infusion of each regimen revealed that the vivo studies. (For more on the relationship between largest increases in histamine levels occurred in tryptase and anaphylactoid reactions see Sect. 4.5.1 subjects given 1 g doses; those given half that

[email protected] 6.1 Antibiotics 191 amount showed only slight changes in histamine with teicoplanin, have been judged to be type I levels. A signifi cant relationship was also seen responses. A case of direct contact allergy involv- between histamine release and reaction severity. ing periorbital skin erosive rash, hyperemia of Findings such as these have led to the recommen- conjunctiva, and corneal stromal edema, and dation that vancomycin should not be adminis- manifesting as itch, soreness, burning, and photo- tered as a bolus and quantities such as 1 g should phobia, was induced by vancomycin eye drops. be infused over a 1-h period. Some other drugs Symptoms resolved upon withdrawal of the eye such as rifampicin, ciprofl oxacin, amphotericin drops and upon initiation of intravenous teico- B, and rarely teicoplanin (see also below) may planin which was tolerated, perhaps surprisingly cause red man syndrome and these drugs, and given its structural similarity to vancomycin. others such as neuromuscular blockers, opioid Skin tests with vancomycin were positive indi- analgesics, and contrast media, can accentuate cating a type I hypersensitivity but skin tests have reactions by provoking the release of histamine not yet been validated with vancomycin, a known (see Sects. 3.2.5.1 , 7.3 , 8.4 and 10.4.1.1 ). histamine releaser, so a number of questions Reactions may be prevented or their severity related to concentration and irritation remain. decreased by extending the infusion time and/or Other reports of adverse reactions to vanco- premedication with histamine antagonists such as mycin include renal disorders, drug fever, and the H1 receptor blocker diphenhydramine alone hematological disorders including eosinophilia, or combined with the H2 receptor antagonist immune thrombocytopenia, leukocytosis, neutro- cimetidine. Despite its chemical similarity with penia, and drug hypersensitivity syndrome (also vancomycin, teicoplanin is claimed not to cause called drug rash with eosinophilia and systemic red man syndrome and histamine release even symptoms [DRESS] syndrome; Sect. 2.2.4.1 ), when infused at rates signifi cantly faster than the the latter being resolved in one study by substi- rates employed for vancomycin and this has led tuting teicoplanin. The most common hypersen- to its recommendation as a substitute for patients sitivities to vancomycin are cutaneous reactions intolerant to the latter drug. In a comparison of which may be a mild skin rash, the more frequent the two drugs, vancomycin, 15 mg/kg adminis- maculopapular or urticarial skin eruptions, exfo- tered over 60 min, and teicoplanin, 15 mg/kg liative dermatitis, fi xed drug eruption, vasculitis, over 30 min, were compared in a double-blind, and the rare reactions such as toxic epidermal randomized, two-way crossover study to deter- necrolysis, Stevens–Johnson syndrome, and mine the occurrence and severity of red man syn- linear IgA bullous dermatosis which can be con- drome and histamine release. Vancomycin caused fused with the fi rst of these toxic bullous cutane- red man syndrome in 11 of 12 patients and pro- ous reactions. voked signifi cant release of histamine into the The chemically similar teicoplanin, not plasma. Teicoplanin did not cause either the syn- approved in the USA, is not inferior to vancomy- drome or signifi cant histamine release. cin with regard to effi ciency of treating gram- positive infections. It shows a lower rate of 6.1.4.3 Other Adverse Reactions to adverse reactions, particularly nephrotoxicity Vancomycin and Teicoplanin and, as already discussed, is used as a substitute Despite a number of reports that cover a range of for vancomycin in red man syndrome. When systemic and dermatologic reactions, severe hypersensitivity reactions do occur with teico- reactions including true type I responses are rare. planin they are generally of the delayed type, but An anaphylactoid reaction to infused vancomy- there are a few documented cases of apparent IgE cin reported in a patient with vancomycin- antibody-mediated reactions implicated, for induced red man syndrome was interpreted, example, by an immediate wheal and fl are skin somewhat obscurely, as a possible case of true reaction to the drug or by teicoplanin-induced vancomycin allergy. Other cases induced by van- histamine release from a patient’s basophils. comycin, but also showing cross-sensitization Despite the chemical and pharmacological

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similarities of the drugs, a reaction to teicoplanin poorly effective or ineffective and when no other with tolerance to vancomycin has been docu- available antimicrobial agent is effective against mented. The reaction occurred 2 days after the infective organism. Desensitization is usually administration of teicoplanin and presented as a used in cases of immunoglobulin E-mediated maculopapular exanthema on the trunk and arms reactions to antibiotics, and although it can effec- over a 7-day period. Skin prick, intradermal, and tively induce tolerance to the problematic drug, patch tests with the drug were negative, but an the mechanism by which this occurs with drugs intravenous challenge test with 400 mg of teico- such as vancomycin is still to be understood and planin elicited pruritic erythematous papular explained (see Sect. 3.5). Both rapid (carried out exanthemata on the elbows, forearms, and abdo- over several hours) and slow (over a period of men 1 h after administration. A later control chal- days) desensitization protocols have proved lenge with vancomycin up to the therapeutic dose effective, although the former is preferred since it demonstrated good tolerance to the drug. This enables therapy for acutely ill patients to con- case was unusual since the literature reveals that tinue within 24 h. Contraindications to be aware the two drugs cross-react almost invariably in of before the initiation of desensitization to van- conditions such as maculopapular exanthemata, comycin include a history of leukocytoclastic vasculitis, acute generalized exanthematous pus- vasculitis, extensive fi xed drug eruption, drug- tulosis, and drug hypersensitivity syndrome. induced hypersensitivity syndrome, and past exfoliative skin reaction such as Stevens–Johnson 6.1.4.4 Skin Tests with Vancomycin syndrome and toxic epidermal necrolysis. and Teicoplanin Desensitization should be undertaken immedi- Skin testing with vancomycin and teicoplanin ately before the desired therapy. Other important has not been well studied and the procedure considerations are the patient’s health issues, remains to be validated with both positive and particularly any preexisting cardiac and pulmo- negative predictive values unknown. Skin test nary conditions, the patient should not be taking results, and particularly details of drug concen- any drugs that might increase the chance of ana- trations used and methodologies employed, are phylaxis or interfere with its treatment (for exam- hard to fi nd in the vancomycin–teicoplanin litera- ple, beta-blockers and ACE-inhibitors) and ture on adverse reactions. In a case study of van- desensitization should be carried out with comycin anaphylaxis followed by successful informed consent in an appropriate setting such desensitization, intradermal skin tests with the as an intensive care unit with all necessary drug were positive at a concentration of 0.1 μg/ml. resuscitation equipment and drugs on hand. Control subjects showed positive responses at Concomitant administration of histamine releas- concentrations of 10 μg/ml or greater. A loss of ing drugs such as neuromuscular blockers, opioid skin test reactivity to vancomycin has been dem- analgesics, some plasma expanders, propofol, onstrated in one case study after successful contrast media, and antibiotics such as ciprofl ox- desensitization to the drug. acin should be avoided or the drugs discontinued or given in smaller doses. 6.1.4.5 Desensitization for Vancomycin Hypersensitivity 6.1.4.5.1 Protocols for Desensitization There are occasions with a drug like vancomycin In one early successful procedure that can be when desensitization is appropriate or even completed in approximately 4 h if each dose is required. Circumstances where desensitization tolerated and no repeats are needed, the patient is would be considered include anaphylaxis to van- premedicated with diphenhydramine 50 mg iv comycin and the diffi cult situation where a case and hydrocortisone 100 mg iv 15 min prior to of red man syndrome cannot be overcome by starting and 6 hourly thereafter. Starting with a slowing the infusion rate of vancomycin, where 250 ml solution of vancomycin 2 mg/ml, four premedication with histamine antagonists proves successive tenfold dilutions are made up to

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Table 6.1 Rapid desensitization protocol for vancomycin continue. This is what happened using the Dose number Dose (mg)a Dose number Dose (mg)a desensitization protocol set out in Table 6.1 1 0.005 11 5.00 where the patient experienced itching that 2 0.010 12 10.00 responded to intravenous diphenhydramine and 3 0.020 13 20.00 mild hypotension and oxygen desaturation that 4 0.040 14 40.00 required reversal with subcutaneous epinephrine 5 0.080 15 80.00 and intravenous diphenhydramine. In compari- 6 0.160 16 160.00 son to the Lerner and Dwyer procedure, this pro- 7 0.320 17 320.00 tocol requires the preparation of many more 8 0.640 18 640.00 dilutions of vancomycin. Not all patients tolerate 9 1.25 19 1,000.00 a rapid desensitization and the proportion that do 10 2.50 – – not is not known. In such patients, desensitization aEach dose, except the fi nal dose, was given in 50 ml of may still be achieved by employing so-called 5 % dextrose infused by infusion pump over 15 min. The last dose was 25 ml of a solution of 1 g vancomycin in slow protocols where infusions rates and buildup 250 ml of 5 % dextrose. This was infused over 15 min and of dose are far slower, often extending over many the remaining solution was then infused at a rate of days. In the event of intolerance to a rapid desen- 200 ml/h. Reproduced with permission from Villavicencio sitization procedure, desensitization to vancomy- AT et al. J Allergy Clin Immunol 1997;100:853 cin has ultimately been achieved in some cases by switching to a slow protocol. To maintain the desensitized state and avoid the possibility of produce solutions of vancomycin containing having to repeat the entire desensitization proce- concentrations of 0.2 mg/ml, 0.02 mg/ml, dure, care should be taken to maintain the admin- 0.002 mg/ml, and 0.0002 mg/ml. That is, fi ve dif- istration of vancomycin. ferent solutions are prepared ranging in concen- Long-term vancomycin therapy may induce tration from 2 to 0.0002 mg/ml. Beginning with neutropenia. A recent report of severe neutopenia the most dilute solution, the drug is infused at a following a prolonged course of vancomycin that rate of 0.5 ml/min and this is increased by 0.5 ml/ progressed to agranulocytosis after reexposure to min every 5 min as long as the dose is tolerated. the drug should be kept in mind and focus attention If an increased rate is not tolerated, the on the safety of rechallenging vancomycin concentration is stepped down to the previously patients with possible drug-induced neutropenia. highest tolerated rate. This step is repeated up to three times for any given concentration. Upon completion of infusion of the solution of highest 6.1.5 Antibiotics Used Topically concentration (2 mg/ml), the patient’s required with Emphasis on Neomycin dose of vancomycin is administered over 2 h with and Bacitracin diphenhydramine 50 mg given orally 1 h before each dose. This procedure devised over 25 years 6.1.5.1 Neomycin ago (Lerner and Dwyer 1984 ) may be compared Neomycin, produced by the bacterium with a more recent rapid protocol summarized in Streptomyces fradiae , is an aminoglycoside anti- Table 6.1 and successfully applied in a patient biotic that shows good activity against gram- who experienced acute cardiac and pulmonary negative bacteria and some gram-positives. arrest after infusion of vancomycin. It should be Neomycin is made up of neomycin B and recognized that mild and transient hypersensitiv- neomycin C. Hydrolysis of neomysin B yields ity reactions such as rash, pruritus, fl ushing, and neamine made up of neosamine B and the amino- erythema occur in about 30 % of patients during cyclitol, 2-deoxystreptamine, and the disaccha- desensitization procedures, but if these symp- ride neobiosamine B composed of D -ribose and toms are tolerated by the patient and do not neosamine B. Hydrolysis of neomycin C produces cause too much discomfort, desensitization can neamine and neobiosamine C, a disaccharide

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eczematous contact dermatitis, neomycin is also NH 2 known to cause generalized reactions such as O Neamine exfoliative dermatitis and erythroderma. Patch HO H N HO 2 tests on patients with chronic otorrhoea have NH2 neosamine O NH 2 shown positive reactions in up to nearly 60 % of HO O OH O 2-deoxystreptamine subjects with neomycin and framycetin the major ribose offenders. In a prospective study of delayed Neobiosamine hypersensitivity reactions to topical aminoglyco- O OH R NH HO 1 2 sides in patients undergoing middle ear surgery, HO 119 patients with chronic otitis media and 30 with O R 2 neosamine otosclerosis were skin tested with a panel of ami- -R1 -R2 Neomycin B - noglycoside antibiotics. Overall, 14 % of patients CH2-NH2 -H were positive to at least one aminoglycoside with Neomycin C -H -CH -NH 2 2 13.4 % positive to gentamycin, 12.8 % to neomy- Fig. 6.6 Structure of the aminoglycoside antibiotic neo- cin, and 4.5 % to gramicidin. Sixteen percent of mycin, made up of neomycin B and neomycin C. Each patients with chronic otitis media were allergic to contains 2-deoxystreptamine while neomycin B is made one of the aminoglycosides commonly found in up of neosamine B and neobiosamine B and neomycin C antibiotic eardrops. Findings such as these have is composed of neosamine C and neobiosamine C. Neosamines B and C are stereoisomers relative to the led to the suggestions that patch testing is almost amino group obligatory in patients with long-standing otitis that does not respond to local therapy and, because of their high risk of sensitization, topical composed of D -ribose and neosamine C. neomycin (particularly as eye drops—see Neosamine B and neosamine C are stereoisomers Fig. 6.7 ) and framycetin should not be used rou- relative to the amino group (Fig. 6.6 ). With the tinely if at all. Attention has also been drawn to presence of a number of free amino and hydroxyl the need to keep patch tests in place for up to groups, the molecule readily lends itself to chem- 7 days since the aminoglycosides need this longer ical manipulation for the preparation of antigens time interval to reveal positive responders. for diagnostic use. 6.1.5.2 Bacitracin Delayed Hypersensitivity Reactions to Neomycin The high levels of allergic sensitivity provoked Allergic contact dermatitis, a type IV delayed by neomycin and gentamycin naturally led to hypersensitivity reaction, is an increasing prob- efforts to ﬁ nd a less allergenic, but equally effec- lem arising from the application of agents during tive, substitute. Initially, the antibiotic bacitracin the care of postoperative wounds, both closed and seemed to satisfy these requirements. Bacitracin open. Reports of neomycin as a causative agent is not an aminoglycoside but a mixture of related date back to at least the 1960s. In fact, neomycin cyclic polypeptides produced from the Tracy consistently ranks in the top 10 % of the most strain of Bacillus subtilis. Its high rate of cure, common allergenic causes of allergic contact der- apparent low incidence of allergic reactions (at matitis with patch test studies revealing sensitivi- least relative to penicillins), and its nephrotoxic- ties of 10–12 % in general patch test populations, ity more or less guaranteed that the antibiotic in the postsurgical population, and in patients would be restricted to topical use. Bacitracin’s suspected of the condition. Allergic sensitization effectiveness against gram-positive bacteria, its to neomycin in patients with chronic venous applicability to infections of the skin, eyes, and insufﬁ ciency has been reported to be as high as ears, and its lower frequency of sensitization rela- 34 % and it has been claimed that the drug ranks tive to neomycin led to its enthusiastic adoption near the top with nickel as the most tested drug as a topical antibacterial, but the antibiotic is not over the last 30 years. In addition to delayed without allergenic properties.

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are not restricted to allergic contact dermatitis but may include localized eczema-like reactions and acute vesicular and chronic dermatitis. As with neomycin, patch tests with bacitracin should be read after a relatively long delay, usually 2–4 days after application. The drug is applied at a concentration of 20 % weight: weight in petrolatum on unbroken skin and, because bacitracin now has a well- established reputation for causing anaphylaxis, it is recommended that patients should be observed for 1 h after patches have been applied.

6.1.5.2.2 Immediate Reactions As well as causing delayed-type IV reactions, bacitracin induces type I immediate hypersensitivities including contact urticaria and anaphylaxis. In fact, with what appears to be a constantly increasing frequency of allergic reactions to bacitracin, it has become, in a relatively short time, the topical antibiotic most recognized for eliciting anaphylaxis. Severe immediate reactions have eventuated following the application of ointments, creams, eye drops, lotions, powders, and irrigations containing the drug and the number of such Fig. 6.7 Allergic contact dermatitis (a ) and severe con- cases now recorded in the literature is approach- junctivitis ( b ) caused by neomycin eye drops (photograph ing 50. The presence of bacitracin- reactive IgE courtesy of P. J. Frosch). Reproduced with permission antibodies has been inferred in a number of the from Brandão FM, in Contact Dermatitis, Springer- cases by positive patch tests and, in at least one Verlag, Berlin, 2011. case, presence of the antibodies was demonstrated in a fl uorescent enzyme immunoassay with 6.1.5.2.1 Delayed Reactions patient’s serum by employing biotinylated baci- Bacitracin’s effectiveness increases in direct pro- tracin coupled to a streptavidin ImmunoCAP® portion to its concentration, and as this and its (Thermo Scientifi c) solid phase. usage rapidly increased, it soon became apparent that the drug was a signifi cant inducer of allergic 6.1.5.2.3 Cross-reactions Between contact dermatitis. Measured incidences of bacitra- Bacitracin and Aminoglycoside cin sensitivity, as low as 0.3 % in 1973, increased to Antibiotics a lower range estimate of 1.5 % in the last 20 years, Immediate allergic reactions including anaphy- but more recent estimates are consistently in the laxis also occur to other aminoglycoside antibiot- range 8–9.5 %. When used for conditions where ics with cases recorded for neomycin, gentamycin, wounds are open or on diseased skin such as tobramycin, framycetin, streptomycin, and dihy- chronic leg ulcers, the sensitization level of 24 %, drostreptomycin. In some of these cases where although signifi cantly less than the 34 % seen with tests were undertaken, patch and/or skin tests neomycin, was still unacceptably high. Apart from proved positive to the culprit aminoglycoside, but patients sensitized by cutaneous application of bac- cross-reactivity with bacitracin has not been itracin, others who contact the antibiotic in the reported or, it seems, looked for. course of their occupation, particularly nurses, are Unexpectedly, some patients demonstrate likely to become sensitized. Cutaneous reactions contact allergy to both bacitracin and neomycin

[email protected] 196 6 Other Antimicrobial Drugs even though the two antibiotics have markedly ear disease reached a number of important different structures. Such co-sensitivities even conclusions that are still relevant today. In the extend to the cyclic peptide antibiotic polymyxin. study group, 35 % of patients were found to have Since structural similarities do not appear to contact dermatitis to drugs used for treatment. account for this phenomenon, the explanation for Patch tests revealed neomycin and framycetin as the coincident reactions to all three drugs seems the most frequent sensitizers with an incidence to lie in the similar exposure patterns both in of 15 %, followed by gentamycin (10 %), poly- terms of the nature of the topical exposure sites myxin (5 %), and, surprising for today, bacitra- and the frequency of exposure. Reactions to baci- cin (2 %). The number of positive reactions to tracin and the aminoglycoside antibiotics seem to neomycin was probably higher than the observed occur particularly when the skin barrier is not incidence since tests were read after 4 rather intact and after prolonged use so applications in than 7 days, and because 90 % of subjects sensi- the form of ointments, creams, irrigations, and tized to neomycin are also allergic to framycetin other dosage forms to open wounds, ulcers, exco- and 40 % are allergic to gentamycin, reactions to riations, and skin grafts should be subject to the latter two aminoglycosides were attributed to caution. Known allergy to either bacitracin or primary neomycin sensitivity. A positive skin neomycin should also preclude the use of the test to gentamycin and a negative test to neomycin other antibiotics. There is a growing belief that were, however, observed and this has been sup- the application of topical antibiotics to closed ported by more recent fi ndings. Although the wounds should be strongly discouraged, that pet- low incidence of reactions to gentamycin is often rolatum is a suitable cost-effective protective put down to its low allergenic potency, this may substitute and for open wounds, and that neomy- again simply refl ect its infrequent use. A doubt cin should be avoided and bacitracin used instead commented on was the diffi culty of determining although its risks should be anticipated and whether the positive reactions to polymyxin and explained to patients. bacitracin were due to separate sensitizations or cross- reactions with neomycin and the conclu- 6.1.5.3 Other Aminoglycoside sions from the study are still of interest today. Antibiotics Firstly, since 17 % of patients were allergic to Emphasis here on neomycin and bacitracin in neomycin or framycetin, their withdrawal from topical preparations refl ects their usage over routine use was recommended; secondly, many years and this in turn has contributed to the because of the high incidence of contact sensi- frequency of occurrence of their now well- tivity to the topical antibacterials and the known adverse effects. Adverse reactions includ- uncertainty that cultured organisms are both ing hypersensitive responses to other pathogenic and relevant, these topical antibiotics aminoglycosides, namely, kanamycin, tobramy- should be avoided; lastly, patients with persistent cin, dihydrostreptomycin, streptomycin, framy- infl ammatory disease should be investigated for cetin, and gentamycin are also known, and again, drug-induced contact dermatitis and the fi ve little usage has probably infl uenced their reported topical antibiotics mentioned here should be low incidence of reactions since kanamycin and included in testing. Thirty years on from this tobramycin are infrequently administered and investigation these conclusions remain relevant. the streptomycins are now largely withdrawn from human administration. Even though the 6.1.5.4 Streptomycin and a Note on aminoglycosides exhibit cytotoxicity, they are Cross-sensitivity Between often formulated with the cyclic peptide antibac- Aminoglycoside Antibiotics terials bacitracin and polymyxin for use in ear The aminoglycoside antibiotics can be divided and eye preparations. An early study of medica- into two groups each made up of amino sugars ment (including aminoglcosides)-induced contact linked glycosidically to an aminocyclitol which dermatitis in patients with chronic infl ammatory is the base streptidine in the case of streptomycin

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NH HO OH NH O NH2 OH O HN NH2 H3C CHO Streptidine NH Streptose OH O

HOH2C O Streptobiosamine HO

NHCH3 HO N-methyl-L-glucosamine

Streptomycin

Fig. 6.8 Structure of streptomycin which is structur- sidically linked to the aminocyclitol streptidine. ally and antigenically similar to neomycin (see Streptobiosamine consists of the nitrogen-containing Fig. 6.6 ). The aminoglycoside antibiotic streptomycin sugar N -methyl-L -glucosamine and the cyclic alcohol consists of the disaccharide streptobiosamine glyco- streptose and 2-deoxystreptamine for the other aminogly- during in vitro fertilization and immunotherapy cosides considered here (see Sect. 6.1.5 and procedures. Fig. 6.6 ). In streptomycin, streptidine is linked to Reports of cross-reactions between neomycin a nitrogen-containing disaccharide, streptobiosa- and some other aminoglycosides, particularly mine composed of N -methyl- L -glucosamine and gentamycin and framycetin, are well known, the fi ve-membered cyclic base alcohol streptose but, consistent with the absence of common (Fig. 6.8 ). Chemically then, streptomycin and, antigenic structures between these molecules for example, neomycin share no structural simi- and streptomycin (compare Figs. 6.6 and 6.8 ), larities and this is also true antigenically. Unlike allergenic cross-sensitization has not been the other aminoglycoside antibiotics considered observed between the two different aminoglyco- here, streptomycin was not primarily used topi- side groups. cally. Hypersensitivity reactions to streptomycin Antibiotics discussed in the following sec- include maculopapular, morbilliform, erythema- tions are little, or only occasionally used and/or tous and urticarial rashes, pruritus, exfoliative poorly allergenic although clindamycin, because dermatitis, eosinophilia, stomatitis, angioedema, of its broad spectrum and suitability as a satisfac- and anaphylactic shock. Because of its greatly tory alternative for patients allergic to penicillins reduced usage and unlike neomycin and genta- and cephalosporins, is administered more fre- mycin, allergic contact dermatitis is currently not quently than the others. a problem with streptomycin, but reports on anaphylaxis to the antibacterial go back over 50 years with a relatively high incidence of reactions 6.1.6 Ribostamycin occurring in the 1960s. However, with the decline in streptomycin therapy in humans, occasional Ribostamycin is a broad spectrum aminoglyco- indications of the antibiotic’s allergenicity are side antibiotic isolated from Streptomyces still evident in the form of veterinary therapeutic ribosifi dicus often used by intramuscular injec- agents, nontherapeutic contact from tiny quanti- tion, particularly in some Asian countries where ties of the drug in foods and culture media, and it is administered to treat pelvic infl ammatory

[email protected] 198 6 Other Antimicrobial Drugs disease. After a third injection of the drug, a reported cases, usually after topical application. female patient developed shortness of breath, Chloramphenicol is still prescribed as eye drops fl ushing, and generalized pruritus, all of which and eye ointment for the treatment of bacterial spontaneously resolved. After a fourth injection conjunctivitis because of its broad spectrum, low of ribostamycin, epinephrine, hydrocortisine, corneal toxicity, its property of providing thera- diphenhydramine, and salbutamol were required peutic levels in aqueous humor, and its avail- to reverse the resultant severe hypotension, ability in preservative-free dosage forms. It is angioedema, dyspnea, dizziness, and generalized from its topical use that sensitization and hyper- urticaria. Prick and intradermal tests revealed sensitivity reactions occasionally occur including positive results with ribostamycin 1 mg/ml but urticaria, angioedema, contact dermatitis, and, as negative responses to two other aminoglycosides mentioned, anaphylaxis. In the face of its declin- tobramycin and micronomicin, suggesting immu- ing usage, especially in the developed nations, noglobulin E-mediated hypersensitivity. A case chloramphenicol has retained its worth as a treat- of erythroderma following intramuscular ribosta- ment for meningitis and this value is most appar- mycin revealed an interesting cross-reaction with ent in meningitis patients with penicillin and neomycin. Patch tests were positive to both cephalosporin allergy. In what fi rst appeared to be ribostamycin and neomycin, suggesting that the an unusual but intriguing fi nding, cross-reactivity three-ring identity the two aminoglycosides between dinitrochlorobenzene and chloramphen- share was responsible for the cross-reaction. icol was reported in 40 % of patients studied for Ribostamycin shares the neamine structure com- contact sensitivities. Since the molecule of chlor- posed of neosamine and 2-deoxystreptamine and amphenicol contains both a nitrobenzene group the ribose ring with neomycin but lacks the and two terminally linked chlorine atoms, this neosamine ring which is part of the neobiosa- claim may have been given some initial credence. mine structure in neomycin (see Fig. 6.6 ). These Follow-up investigations found no positive patch fi ndings suggest the possibility of a potential tests to chloramphenicol ointment 1 % in 100 hazard if a systemic aminoglycoside is given to a patients with dinitrochlorobenzene sensitivity patient sensitized by a topical aminoglycoside and none of 15 patients with eczema and a such as neomycin. delayed-type reaction to chloramphenicol cross- reacted with dinitrochlorobenzene in acetone or petrolatum. An additional 27 patients primarily 6.1.7 Chloramphenicol sensitized to dinitrochlorobenzene also showed no reaction to chloramphenicol or its salts. Originally from the bacterium Streptomyces ven- Misinterpretation of a primary irritant reaction as ezuelae , chloramphenicol shows a broad spec- an allergic contact reaction was suggested as the trum of antimicrobial activity and, despite being explanation for the originally reported industrial widely and frequently used for a number of years chemical–antibiotic cross-reaction. following its introduction in 1949, the antibiotic has provoked a remarkably small number of hypersensitive reactions. This is refl ected in the 6.1.8 Clindamycin infrequency of references to the drug in both broad surveys of drug-induced adverse reactions Clindamycin is formed by substituting a chlorine and in the tiny number of individual case reports. atom for the 7-hydroxy substituent of the natu- In recent years, administrations of the drug have rally occurring lincosamide antibiotic lincomy- declined due to the fear of resistant organisms and cin (Fig. 6.9 ). Both compounds contain the some safety concerns. The most serious adverse unusual sulfur-containing sugar moiety methyl- reactions are aplastic anemia, bone marrow sup- thiolincosamide. Clindamycin is used to treat pression, and rarely, anaphylaxis for which there anaerobes, protozoa, and some methicillin resis- appears to have been up to about a dozen tant S. aureus infections and is applied topically

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HO CH3 determinant structures been identifi ed. Skin tests CH3 O OH H H are not standardized and the optimum N OH N concentration(s) of drug to use have yet to be H determined and agreed upon. For skin prick test- O OH ing, a concentration of 150 mg/ml was used to H C detect positive reactions, and for patch tests, a 3 SCH Lincomycin 3 concentration of 10 % in petrolatum. Intradermal testing has been employed in the concentration Cl CH 3 range 0.15–1,500 μg/ml, but its use has been CH3 O OH H H N OH limited. Diagnosis of clindamycin hypersensi- N tivity often relies on temporal associations with H O drug-induced signs and symptoms. Some studies OH have concluded that skin tests are not adequate H C 3 SCH Clindamycin 3 to identify suspected allergic reactions and provocation testing is a suggested superior alterna- Fig. 6.9 Structures of the naturally occurring antibiotic tive. In one study, a negative predictive value of lincomycin, which contains the sulfur-containing sugar only 68 % was calculated for prick and intrader- methylthiolincosamide, and the closely related clindamy- mal tests. Oral challenge has also been suggested cin in which the 7-hydroxy substituent is replaced by a chlorine atom to overcome false positive and negative skin responses to the drug. Positive skin prick and intradermal tests to clindamycin were obtained for acne. Although it is a valuable anti-infection in a patient who developed erythroderma a week agent especially as a substitute for the β-lactam after receiving the antibiotic intravenously. No antibiotics, clindamycin demonstrates some immediate skin responses to the tests were toxicity, particularly pseudomembranous colitis, observed, but 12 h delayed responses were seen and occasionally elicits some hypersensitivity following skin prick tests with clindamycin at a responses. Delayed-type cutaneous reactions to concentration of 150 mg/ml and intradermal tests clindamycin including pruritus, exanthematous with the drug in the range 0.015–1.5 mg/ml. The rash, generalized maculopapular exanthema, same tests were negative in fi ve control patients. erythroderma, generalized exanthematous pustu- Patch testing with clindamycin 10 % in petrola- losis, and Stevens–Johnson syndrome have been tum yielded positive responses in patients with reported and contact dermatitis may occur after delayed cutaneous adverse reactions in the form topical application. However, to retain some of generalized maculopapular exanthema follow- perspective, attention should be drawn to a 1999 ing administration of the antibiotic. No positives hospital study in Chicago of clindamycin-induced were detected in control subjects, but as with skin adverse drug reactions for the period 1995–1997 prick and intradermal tests, patch testing with which concluded that clindamycin hypersensi- clindamycin needs standardization and validation tivity appears to be rare with an incidence of studies. Patch tests using a suspension of ground adverse reactions lower than reported 25 years clindamycin tablet in saline (150 mg/ml) and earlier. Anaphylaxis to the drug is said to occur in prick tests with a parenteral solution of the drug less than 0.1 % of treated patients, and with a (150 mg/ml) were performed on 33 patients with literature search revealing only two cases, the a history of skin reactions in temporal association fi rst apparently in 1977, this reported incidence with treatment with clindamycin. To exclude IgE may also be an overestimate. antibody-mediated hypersensitivity, prick tests Diagnostic tests for clindamycin-induced were read after 20 min. Patch tests were removed hypersensitivities have not been widely employed after 1 day and late reactions were assessed after and validated. No clindamycin-specifi c IgE anti- 2, 3, and 4 days. In four of the patients, matching bodies have been found nor have any allergenic positive delayed reactions were seen with both

[email protected] 200 6 Other Antimicrobial Drugs skin tests and patch testing identifi ed an addi- bacterium of the Streptomyces genus. The tional positive reactor. Challenge tests elicited structure, a methyloxirane derivative of phos- rashes in a further six patients—three patients phonic acid, is quite unlike any of the other anti- with exanthema, two with symmetrical drug- biotics. Fosfomycin shows promise for the related intertriginous and fl exural exanthema, treatment of multidrug-resistant Enterobacteria- and one with a non-pigmented fi xed drug erup- ceae infections including extended spectrum tion. The investigators concluded that combined β-lactamase-producing organisms and is admin- skin tests plus challenge testing are necessary to istered, usually as a single large dose, for infec- rule out allergic clindamycin hypersensitivity. tions of the urinary tract. At least three cases of anaphylactic shock to the antibiotic, two recently to fosfomycin tromethamine, have been reported, 6.1.9 Pristinamycin but there appears to be no other reports of adverse reactions presumably refl ecting the paucity of Pristinamycin is a member of the streptogramin clinical research, infrequent usage of the drug, class of antibiotics effective against vancomycin- and/or a low incidence of reactions. resistant S. aureus and vancomycin-resistant enterococcus. It is biosynthesized by the bacterium Streptomyces pristinaespiralis and consists 6.2 Antimicrobials Other than of two components, pristinamycin IA (a macro- Antibiotics lide) and the depsipeptide pristinamycin IIA (also known as streptogramin A) in the ratio of 30:70. 6.2.1 Sulfonamides Streptogramins, also called synergistins, are made up of two groups, streptogramins A and B. As already stated, sulfonamides used as chemo- There are at least four documented cases of therapeutic agents to treat infections will be anaphylaxis to the drug, but hypersensitivity referred to here as antimicrobials (or antibacteri- reactions are otherwise rare, usually cutaneous, als) not antibiotics. Other drugs with a sulfon- and of the delayed type. In one study of 29 amide structure but no antibacterial action will patients with cutaneous reactions to pristinamy- be termed, for example, a sulfonamide diuretic cin, maculopapular rash occurred in 18, erythro- (like furosemide and hydrochlorothiazide) or derma in 9, angioedema in 1, and Stevens–Johnson a sulfonamide rheumatologic agent (such as syndrome in 1 patient. The patients’ skin test sulfasalazine). responses were assessed to pristinamycin, and related streptogramins, quinupristin/dalfopristin 6.2.1.1 Structure–Activity and virginiamycin, were also tested on some Considerations patients to assess cross-reactions between the A sulfonamide contains a sulfonyl group con- drugs. Patch and intradermal but not prick tests nected to an amine and has the general formula proved to be of diagnostic value and detection of R1 SO2 NR2 R3 . In everyday clinical medicine and some positive reactions to quinupristin/dalfopris- amongst the public, a sulfonamide used to treat tin and virginiamycin as well as to pristinamycin infections is often called a “sulfa drug”. led to the conclusion that cross-reactivity existed Chemically, for the antibacterial drugs, the between the drugs and therefore all strepto- generic name sulfonamide refers to derivatives gramins should be avoided in patients with of para -aminobenzenesulfonamide or sulfanil- adverse cutaneous reactions to pristinamycin. amide, a structural analog of para -aminobenzoic acid (Fig. 6.10 ). Structural prerequisites for antibacterial action are, to greater or lesser extent, 6.1.10 Fosfomycin refl ected in the structural features of sulfanilamide. The arylamine group at the N4 position of Originally called phosphonomycin, fosfomycin the sulfonamide structure must be para to the is a broad spectrum antibiotic produced by a position of direct attachment of the sulfur to the

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O R = NH2 R2 1 R1 SN R2=H R3 or O R3=H heteroaromatic ring General formula for sulfonamide drugs

4 1 H2N SO2N H2 H2N COOH Sulfanilamide Para-aminobenzoic acid (para-aminobenzenesulfonamide)

R1 R2 R3 (N1 substituent) N

H2N SO2N H N Sulfadiazine

CH3

H2N SO2N H O N Sulfamethoxazole

Fig. 6.10 General structure of a sulfonamide antibacte- ment of the sulfur to the benzene ring. The amine must rial drug, general formula R1 SO2 NR2 R3, containing a be unsubstituted or in a form that can be converted back sulfonyl group connected to an amine. Sulfonamide to the free amine. The nature of the attached group at N 1 antibacterials are derivatives of para -aminobenzenesul- (R3 substituent) strongly infl uences the antibacterial fonamide or sulfanilamide, a structural analog of activity of the molecule. For example, attachment of the para -aminobenzoic acid. For antibacterial action, the methyl substituted isoxazolyl group at N 1 as in sulfa- arylamine group at the N 4 position of the sulfonamide methoxazole produces a sulfonamide with marked anti- structure must be para to the position of direct attach- bacterial action benzene ring and, as in para -aminobenzoic acid, microorganisms that must synthesize their own the amine must be unsubstituted or at least in a folic acid are sensitive to sulfonamide antibacte- form that can be converted in the tissues back to rials. Substitution of aromatic heterocyclic the free amine (Fig. 6.10 ). In general, the addi- groups at N1 , for example, a diazine nucleus in tion of other substituents to the benzene ring or sulfadiazine (Fig. 6.10), often produces sulfon- replacement of the ring with another ring leads to amides of higher potency (for example, com- a loss of antibacterial activity. The –SO 2 NH2 pared to sulfanilamide) even though such group attached directly to the ring is essential heterocyclic derivatives are chemically less like and not only is substitution at the N 1 nitrogen the metabolite para -aminobenzoic acid than sul- essential, but the nature of the attached group fanilamide. Ionization and dissociation measure- strongly infl uences the antibacterial activity of ments seem to provide an explanation for this. It the molecule. The sulfonamide antibacterials, by seems that the addition of such weakly basic virtue of their similarity in chemical structure to substituents to the –SO 2 NH2 group changes the para -aminobenzoic acid, competitively inhibit electrical properties of the modifi ed group, that the enzyme-assisted incorporation of this is, the –SO2 NHR group, in such a way that its essential metabolite into dihydropteroic acid, properties become more like the carboxylic acid the immediate precursor of folic acid. Hence, group of para - aminobenzoic acid.

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6.2.1.2 Hypersensitivity Reactions drug-induced immediate allergic reaction. A The overall incidence of adverse reactions to detailed analysis of the structure–activity fi nd- sulfonamide antibacterials is about 3–5 %. ings in the form of comparative inhibition results Reactions are many and varied and may involve obtained with selected analogs is provided as a almost every organ system of the body including guide for how quantitative hapten inhibition stud- the blood, bone marrow, liver, kidney, skin, and ies can be used to defi ne the allergenic fi ne struc- peripheral nerves. Individual adverse responses tural features of a drug, that is, the structural include nausea, vomiting and anorexia, hemolytic features most complementary to the induced IgE anemia, aplastic anemia, agranulocytosis, throm- antibody combining sites. Put more simply, this bocytopenia, eosinophilia, renal damage due to means the structural features responsible for the crystalluria (with older sulfonamides), hepatitis, “goodness of fi t” between a drug and its comple- goiter, hyperthyroidism, rarely peripheral neuri- mentary antibodies. Interpretations of the results tis, and hypersensitivity reactions. The latter are are reproduced from the author’s studies with said to occur in about 3 % of courses of the drug permission. and in approximately 50–60 % of sulfonamide- Skin prick testing was carried out using treated patients with human immunodefi ciency Septrin Parenteral Infusion® (Wellcome) contain- virus (HIV) infection. Many of the reactions ing 16 mg/ml of trimethoprim and 80 mg/ml of involve the skin and mucous membranes. A vari- sulfamethoxazole. For intradermal testing, trim- ety of more severe reactions may also occur ethoprim and sulfamethoxazole were dissolved including potentially lethal toxidermias and a in the minimum quantity of 0.1 M sodium delayed hypersensitivity-type syndrome charac- hydroxide and benzyl alcohol, respectively, terized by fever, skin rash, and multi-organ toxic- before diluting with physiological saline to a ity. All of the above reactions are distinct from the dilution of 1 mg/ml. Physiological saline solu- type I true allergic reactions which are immediate tions containing the same amounts of sodium in onset, usually non- febrile, mediated by drug- hydroxide or benzyl alcohol were used as con- reactive IgE antibodies, and may be accompanied trols. Tests were performed by injecting 0.02 ml by urticaria and symptoms of anaphylaxis includ- of test or control solution and generally testing ing wheezing, shortness of breath, hypotension, was commenced with the test solutions diluted angioedema, bronchospasm, and ultimately car- 1:100. Sera from patients with a history of an diovascular collapse. The involvement of sulfon- immediate response and/or a positive skin test to amide antibacterials in these hypersensitivities of sulfamethoxazole were examined in vitro in the immediate type will be dealt with fi rst. radioimmunoassays using a sulfamethoxazole– Sepharose solid phase covalent complex and an 6.2.1.2.1 Type I, IgE Antibody-Mediated 125 I-labeled anti-human IgE antibody as second Reactions to Sulfonamide antibody to detect binding of patients’ serum IgE Antimicrobials antibodies to the immobilized drug. Sera from These are the best worked out and defi ned normal, healthy subjects, cord blood, and patients sulfonamide- induced hypersensitivity reactions highly allergic to mites and pollens were used as with the best defi ned allergenic drug structures. controls. As an additional control, no signifi cant Patients with symptoms characteristic of an binding was observed when the patients’ sera immediate type I reaction such as generalized were tested with ethanolamine–Sepharose and itch, urticaria, angioedema, wheezing, and other Sepharose alone as control solid phases. To check anaphylactic-like symptoms following ingestion on the absolute specifi city of antibody binding, of standard oral doses of co-trimoxazole (trime- quantitative hapten inhibition experiments were thoprim 80 mg and sulfamethoxazole 400 mg per carried out using a range of sulfonamide analogs. tablet) were investigated and the investigative In the typical example shown (Table 6.2 ), easily procedures undertaken and the results obtained the most potent inhibitors were sulfamethoxazole are provided as a general protocol for studying a and sulfamerazine, the former requiring half the

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Table 6.2 Inhibition by antibacterial sulfonamides of the binding to a sulfamethoxazole-solid phase a of IgE antibodies in the serum of a patientb allergic to sulfamethoxazole Amount (nmol/tube) of sulfonamide for Inhibition (%) of binding 50 % inhibition of to sulfamethoxazole– binding of IgE Sepharose by 1 μmol/ Compound Structure antibodies tube of compound

Sulfamethoxazole CH3 265 66 H NSONH 2 2 O N

Sulfamerazine CH3 540 62 N H2NSO2NH N

Sulfamethizole S CH3 >1,000 39 H NSONH 2 2 N N Sulfachloropyridazine >1,000 46 H2NSO2NH Cl NN

Sulfamethazine CH3 840 52 N H2NSO2NH N CH3

Sulﬁ soxazole H3C >1,000 40 CH3 H NSONH 2 2 N O

Sulfadimethoxine OCH3 900 51

H2NSO2NH N N OCH3 Sulfamethoxypyridazine >1,000 31 H2NSO2NH OCH3 NN

Sulfamoxole O CH3 >1,000 33 H2NSO2NH N CH3 Sulfathiazole S >1,000 42 H2NSO2NH N Sulfapyridine >1,000 37 H2NSO2NH N Sulfanilic acid >1,000 2 H2NSO3H

Sulfanilamide >1,000 14 H NSONH 2 2 2 a Sulfamethoxazole–Sepharose covalent complex used in radioimmunoassay with a 125 I-labeled second antibody b Patient experienced severe itch and rash to Septrin® (two tablets) 1 year after experiencing an anaphylactic-like reaction to Septrin ® tablets (trimethoprim 80 mg and sulfamethoxazole 400 mg) Adapted from Harle DG et al. Mol Immunol 1988 ; 25:1347 with permission

[email protected] 204 6 Other Antimicrobial Drugs amount of drug as the latter to achieve 50 % ring which may sterically hinder the binding of inhibition, followed by sulfamethazine. Ten other the IgE antibodies particularly to the methyl analogs were far less well recognized with, in group linked to position fi ve. The relative bulk general, 600–900 nmol per tube of drug needed and close proximity of the methyl groups are for just 30–40 % inhibition. Sulfanilic acid and readily seen in the model shown in Fig. 6.11 . sulfanilamide, compounds without a heterocyclic Each of the above considerations, together ring attached at the N1 position, were virtually with the fi nding that sulfanilamide was virtually inactive producing only 2 and 14 % inhibition, without inhibitory effect, led to the overall con- respectively, at 1,000 nmol per tube. Close com- clusion that the 5-methyl-3-isoxazolyl group on parisons of the structures of the sulfonamides the sulfamethoxazole molecule is the allergenic examined and the corresponding inhibitory val- determinant with the methyl substituent on the ues revealed clearly that compounds with one ring being a dominant fi ne structural feature of methyl substituent on a fi ve- or six-membered the determinant. aromatic heterocyclic ring containing at least one nitrogen atom immediately adjacent to the point 6.2.1.2.2 Delayed Onset Sulfonamide of attachment to the N1 sulfonamido nitrogen Hypersensitivity Reactions were the structures most complementary to the These reactions, characterized by fever, a morbil- IgE antibody combining sites (Table 6.2 ). The liform or maculopapular non-urticarial skin rash, structures of sulfamethoxazole and sulfamera- occasionally multi-organ toxicity including the zine show a 5-methyl-3-isoxazolyl N1 substituent liver and blood, and often eosinophilia, generally in the former and a 4-methyl-2-pyrimidinyl group occur 1–2 weeks after the initiation of therapy. in the latter compound. Sulfamethazine, or N ′- Rare patients may progress to life-threatening (4,6-dimethyl-2-pyrimidinyl)sulfanilamide, the reactions such as Stevens–Johnson syndrome and dimethyl derivative of sulfamerazine was the toxic epidermal necrolysis. The number of these third best inhibitor. Each of the three best inhibi- lethal reactions to all drugs totals about ten cases tors has a methyl group on the carbon β to the per million persons per year with the incidence of sulfonamido substituent and either a fi ve- or a sulfonamide-induced reactions being one of the six-membered heterocyclic ring (Fig. 6.11 ). The highest. All drug-induced hypersensitivity reac- methyl group and the position of the nitrogen tions must occur via direct toxic effects on tissues atom are common to all three structures and these or via immune processes, so, bearing in mind the features are clearly important for recognition by multitude of effects seen in these non-type I, the complementary IgE antibodies. The fact that delayed hypersensitivities, it seems that the sulfamethazine, the dimethyl derivative of sulfa- involvement of any one or more of types II, III, or merazine, proved such a relatively poor inhibitor IV as well as direct cytotoxic effects of the drug suggested that only one methyl group is neces- are likely or possible in provoking reactions. sary for recognition. This is further supported by Other possibilities are direct cytotoxic effects or results obtained with the close analog of sulfa- the induction of an immune response by the drug methoxazole, sulfamoxole which contains a fi ve- or its metabolite(s) bound to an endogenous pro- membered 4,5-dimethyl-2-oxazolyl ring with the tein, probably of cellular origin, or the parent oxygen atom adjacent to the point of attachment unmetabolized sulfonamide might interact of the ring to the sulfonamido nitrogen atom directly with antibodies and/or T cells. (Table 6.2 ). Unlike sulfamethoxazole, sulfamox- Current thinking and most research efforts are ole contains two methyl groups at the four and based on the belief that the pathogenesis of the fi ve positions of the heterocyclic ring and the reactions involves bioactivation of the sulfon- lower inhibitory potency of sulfamoxole can amide, forming reactive metabolites that act as probably be explained by the presence of the the initial reactive agents in a sequence of inter- additional methyl group at position four of the actions that ultimately lead to the patients’

[email protected] Sulfamethoxazole

N N O O H2N SO2NH H2N SO2NH

CH3 CH3

Sulfamoxole

O CH3 O CH3

H2N SO2NH H2N SO2NH N CH N 3 CH3

Sulfamerazine

N N

H2N SO2NH H2N SO2NH N N

CH3 CH3

Sulfamethazine

CH3 CH3 N N

H2N SO2NH H2N SO2NH N N

CH3 CH3

Fig. 6.11 Two-dimensional and three-dimensional CPK structural features identiﬁ ed for cross-reactivity are an N1 space-ﬁ lling molecular models of sulfamethoxazole and aromatic heterocyclic ring containing at least one nitrogen three allergenically cross-reactive sulfonamides, sulfa- and with a methyl substituent ( arrowed ) β to the point of moxole, sulfamerazine, and sulfamethazine. Important attachment of the ring to the nitrogen (see also Table 6.2 )

[email protected] 206 6 Other Antimicrobial Drugs adverse responses. Metabolism of sulfonamide antibodies and/or T cells had rarely been demon- antimicrobial drugs proceeds via a number of strated in patients with sulfonamide hypersensi- processes. In humans, the largest proportion of an tivity. It now seems accepted that T cells are administered sulfonamide, for example, sulfa- involved although the nature of the antigen(s) methoxazole, is metabolized in the liver by reacting with the specifi c T cell receptor remains N -acetyltransferase to the N4 acetylated form that incompletely defi ned. The precise mechanisms is nontoxic and eliminated in the urine while underlying the hypersensitivity/adverse reactions about 15 % of the drug is glucuronidated at the in the patients are also yet to be worked out in N 1 nitrogen and also renally excreted. detail, although again T cells are thought to be Glucuronidation can occur at the N4 nitrogen but, involved since hapten–protein antigens stimulate unlike the N1 metabolite, the product is unstable. CD4(+) regulatory and CD8(+) effector T cells A small fraction of sulfamethoxazole, about from hypersensitive patients. In drug hypersensi- 10 %, is metabolized to a reactive hydroxylamine tivity research where bioactivation of the drug is intermediate by several enzymes, particularly involved, the relationship of drug metabolism to CYP2C9 (cytochrome P450), myeloperoxidase the immune response has usually been obscure. (MPO), and perhaps cyclooxygenase (COX), to Recent results suggest that antigen-presenting the unstable hydroxylamine intermediate that cells alone are suffi cient to generate metabolites auto-oxidizes to the highly reactive nitroso of sulfamethoxazole, produce the hapten–protein derivative nitroso-sulfamethoxazole (Fig. 6.12 ). antigen complexes, and ultimately induce the Although cyclooxygenase was thought to be T cell response to the drug. Dendritic cells are involved in the metabolic conversion to the thought to activate sulfamethoxazole intracellu- hydroxylamine, that is now in doubt. The hydrox- larly to nitroso-sulfamethoxazole which is not ylamine derivative is thought by some to be only chemically more reactive but also more involved in a number of the adverse reactions immunogenic/allergenic. Stimulation of human including hepatitis, nephritis, thrombocytopenia, T cells with sulfamethoxazole, and with its lupus erythematosus, and the sulfonamide hyper- nitroso and hydroxylamine metabolites, gener- sensitivity syndrome. Nitroso-sulfamethoxazole ated T cell clones that showed three recognition can itself be acetylated and eliminated or it patterns: 14 % were sulfamethoxazole-specifi c, can react with glutathione and be reduced back 44 % were sulfamethoxazole metabolite-specifi c, to the hydroxylamine derivative. Nitroso- and 43 % were stimulated with sulfame- sulfamethoxazole, but not the parent drug, can thoxazole and its metabolites. Although the react covalently with cysteine residues of cellular sulfamethoxazole- responsive clones were spe- surface proteins including skin keratinocytes, cir- cifi c for the stimulating sulfonamide, a large pro- culating peripheral blood mononuclear cells and portion of nitroso-sulfamethoxazole-specifi c splenocytes, and serum proteins, predominantly clones also responded to nitroso metabolites of immunoglobulins and albumin, to form hapten– sulfadiazine and sulfapyridine but not nitroso- protein complexes. In the latter case, reaction benzene, that is, T cell responses can occur via probably proceeds via Cys34 which is highly cross-reactivity with the haptenic immunogen. reactive with electrophiles. The ultimate drug The complexity of the fi ndings and some inter- determinant involved in the sulfonamide-induced pretations in the large body of research on delayed hypersensitivity reaction is believed to sulfonamide hypersensitivity involving the be the sulfonamide hapten made antigenic by enzyme-induced generation of drug metabolites; linkage to carrier proteins but, from this point on, unstable intermediates; protein-reactive species; the question becomes, what specifi c immunolog- the formation of cell surface and intracellular ical processes are involved in the pathogenesis of protein adducts; co-stimulatory signaling; and the reaction? A little more than a decade ago the development of an antigen-specifi c T cell some investigators pointed out that drug-reactive response leave some doubts and questions. For

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O NH NH2 HN CH 2 3

N-Acetyltransferase UDP-Glucuronosyl OOS OOS OOS HOOC O NH (NAT1) NH transferase HO N HO OH N N N O O O CH CH CH3 3 3 N4-Acetylsulfamethoxazole Sulfamethoxazole Sulfamethoxazole-N1-glucuronide (Detoxification product) [45-70%] (Detoxification product) [5-15%] CYP2C9/MPO/COX [~10%] CYP2C9 Hydroxylation OH NH2 HN

? Adverse OOS OOS reactions NH NH

N N O O CH 3 CH2OH Sulfamethoxazolehydroxylamine 5-Hydroxysulfamethoxazole [5-10%] [O] Reduction Glutathione O Cysteine N

Cell damage OOS NH

N O CH 3 Nitrososulfamethoxazole

Protein-SH Antigen formation

S OH Protein N

OOS NH

N O

CH3 Unstable mercapto intermediate

O S Protein NH

Antigen Hyper- recognition Antigen T sensitivity/ OOS cell and presentation Adverse NH processing reactions N O CH 3 Sulfamethoxazole–protein adduct

Fig. 6.12 Bioactivation of sulfamethoxazole to its reactive metabolite and subsequent steps leading to the metabolite- speciﬁ c immune response and hypersensitivity/tissue damage

[email protected] 208 6 Other Antimicrobial Drugs example, the disparity of some results in labora- 6.2.1.3 Reactions to Sulfamethoxazole tory animals and humans produces doubts about in Patients Infected with Human the relevance of the laboratory findings nor is Immunodefi ciency Virus it clear why apparently all sulfamethoxazole- With the exception of sulfamethoxazole (gener- allergic patients show T cell responses to both ally in combination with trimethoprim), sulfon- the nitroso derivative of sulfamethoxazole and amides are no longer heavily and widely used. the parent drug. Attempted obscure explanations However, the drug, again with trimethoprim, has involving the amorphous concept of “avidity found specialized application due to its effective- spreading” to the continually present parent drug ness against Pneumocystis jirovecii in immuno- do nothing to aid understanding. One is still left compromised HIV-infected patients with with the question of individual susceptibilities. Pneumocystis pneumonia (PCP) and other poten- Sulfamethoxazole induces hypersensitivity reac- tially life-threatening opportunistic infections. tions in 1–3 % of those exposed to it (and in 50 % For effective prophylaxis of PCP, the drug com- of HIV patients). Every patient given the drug is bination is given as a single double-strength daily exposed to it and its nitroso metabolite so why dose of 160 mg of trimethoprim and 800 mg of don’t all patients develop a hypersensitivity reac- sulfamethoxazole. A serious drawback to the use tion to the antigen formed by the metabolite of these drugs in HIV-infected patients is a high reacting with protein? Possible explanations rate of adverse reactions that can range up to advanced to account for different susceptibilities 50–60 % and require discontinuation of therapy. of individuals include patients with a slow acety- The withdrawal of the drugs is lower during dos- lator phenotype that reduces their capacity to age for prophylaxis than during treatment, but detoxify reactive metabolites; glutathione poly- patients receiving secondary prophylaxis may morphisms; the need for what has vaguely been experience reactions despite successful previous described as “co-stimulation” for T cell receptor acute treatment. For patients with a history of activation; and the possible need for complemen- reactions, rechallenge provoked adverse reac- tary bidirectional drug binding domains within the tions in 13–47 % of subjects, rates that are similar major histocompatibility complex (MHC) and to the incidences of reactions in patients with pri- the T cell receptor. The latter possible explana- mary adverse reactions. Regardless of previous tions add an even more bewildering layer of com- reaction history, adverse reactions are similar and plexity on an already densely complex narrative. include rash, fever, fl u-like symptoms, and gas- An apparent high risk of sulfonamide hyper- trointestinal disturbances. In two early trials, sensitivity in patients with hematological malig- reactions to high doses of sulfamethoxazole–tri- nancies was the stimulus for a recent study methoprim resolved in more than 80 % of patients designed to look for defi ciencies in sulfonamide despite the therapy continuing. Because of the detoxifi cation pathways. Patients were examined effi cacy of sulfamethoxazole–trimethoprim for for levels of glutathione, ascorbate, cytochrome PCP prophylaxis, the reintroduction of the drug b5 , and cytochrome b 5 reductase, but no defi cien- combination in patients who had a prior adverse cies of the blood antioxidants and cytochromes reaction is sometimes desired. With this in mind, were found. In a secondary study, the incidence a randomized, controlled trial was undertaken to of drug hypersensitivity following intermittent compare dose escalation with direct rechallenge trimethoprim–sulfamethoxazole prophylaxis was regimens. With the primary end point being the compared to the incidence reported for high dose patient’s tolerance of single-strength sulfa- regimens. After 3–4 weeks of the administration methoxazole–trimethoprim for 6 months, 75 % of trimethoprim–sulfamethoxazole 960 mg three of the dose-escalation group and 57 % of the to four times weekly to 22 patients, no patient direct rechallenge group were able to receive the developed sulfamethoxazole-specifi c T cells and single-strength dosage for the selected time, only one patient developed a rash. demonstrating that it is possible to successfully

[email protected] 6.2 Antimicrobials Other than Antibiotics 209 reintroduction the drugs to a high proportion of AIDS patients, glutathione levels are progressively HIV-infected patients who previously experi- depleted and this parallels the higher incidence enced treatment-limiting adverse reactions. of sulfamethoxazole-induced hypersensitivity in The rate of drug-induced adverse reactions in these patients. A polymorphism in glutamate HIV-infected patients is over fi ve times higher cysteine ligase catalytic subunit (GCLC), also than the rate for HIV-negative subjects, and known as gamma-glutamylcysteine synthetase, although the reason for this is not known, some the rate-limiting enzyme of glutathione synthesis, observations related to reduction of the reactive was recently found to be associated with sulfameth- nitroso-sulfamethoxazole metabolite may be oxazole-induced hypersensitivity in HIV/AIDS pertinent. Defi ciencies of ascorbate and thiols patients. A single nucleotide polymorphism (for example glutathione), two agents that effect (SNP), reference SNP number rs761142 T > G, in reduction of the metabolite in vivo, have been GCLC was signifi cantly associated with reduced reported in HIV-infected subjects and it has been mRNA expression in liver and B lymphocytes and speculated that this may result in increased with the drug hypersensitivity. This fi nding was metabolite-mediated lymphocyte toxicity and a interpreted as support for the role of reactive signifi cantly increased risk of hypersensitivity metabolites in the pathogenesis of sulfamethoxa- reactions. zole-induced hypersensitivity and led to the specu- An unusual acute anaphylactic-like syndrome lation that GCLC may be associated with reactions that resembles septic shock has been observed in caused by some other drugs. some HIV-infected patients following the administration of co-trimoxazole. The severe systemic 6.2.1.4 The Question of Cross- reactivity reaction is characterized by fever, hypotension, Between Antimicrobial and pulmonary infi ltrates, but absence of bron- and Non-antimicrobial chospasm and laryngeal edema are points of dif- Sulfonamides ference with classic anaphylaxis. Because of the Chemically, a sulfonamide is a compound that similarities between this hypotensive syndrome contains a sulfonyl group connected to an amine and septic shock, it has been suggested that and which has the general formula shown in tumor necrosis factor (TNF), a mediator of septic Fig. 6.13 (see also Fig. 6.10 ). Besides the sulfon- shock, is released during episodes of the syn- amide antimicrobials, many drugs of diverse phar- drome. However, TNF and IgE antibodies to co- macological action used in medicine today are trimoxazole were not detected and there was no sulfonamides. Some that are widely used include depression of complement in a patient who the diuretics furosemide and hydrochlorothiazide; responded with a second episode of shock after sulfonylureas such as glyburide and tolbutamide; being rechallenged with the drug combination. uricosurics, e.g., probenecid; sulfasalazine used as A number of trials have confi rmed the effi cacy a rheumatologic agent and for infl ammatory bowel of desensitization with trimethoprim–sulfamethox- disease; the selective serotonin-1 receptor agonist azole in HIV-infected patients. Successful desensi- sumatriptan; and the cyclooxygenase-2 inhibitor tizations, both rapid and slow, were achieved with celecoxib (Fig. 6.13 ). These drugs are distin- success rates often in the 70–100 % range depend- guished from the antimicrobials by the absence of ing on the number of patients involved and the an unsubstituted arylamine group (or a group CD4+ and CD8+ counts. Desensitization seemed capable of being converted to the free amine) more often successful with lower CD4+ percent- attached directly to the benzene ring at the N4 ages and CD4+:CD8+ ratios. position of the antimicrobial compounds (com- Polymorphisms in genes involved in sulfa- pare Fig. 6.10 ). Another important difference is methoxazole biosynthesis, metabolite detoxifi ca- the presence in the sulfonamide antimicrobials of tion, and the regulation of glutathione levels are of an aromatic fi ve- or six- membered heterocyclic interest in attempts to understand the risk of ring (except for the “parent” antimicrobial sulfa- hypersensitivity induced by the drug. In HIV/ nilamide), generally containing at least one nitro-

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O R2 R1 SN R O 3 Sulfonamide general formula

O Cl Cl

H H HN SO 2 N HN SO2 N H H O S OH HN O O

Furosemide (Frusemide) Hydrochlorothiazide

O O

HN H H H H Cl N SO2 N SO2 N N

O O

Glyburide (Glibenclamide) Tolbutamide

O O HO H SO2 N N SO N HO 2 HO N N

Probenecid Sulfasalazine

H F3C N N H SO2 N N SO2 N CH3 H N H

Sumatriptan Celecoxib

Fig. 6.13 Examples of some non-antimicrobial sulfon- tuted arylamine attached at N 4 (see Fig. 6.10 ) and (except amides containing a sulfonyl group connected to an amine for sulfasalazine) an aromatic ﬁ ve- or six-membered het- but which are not recognized by sulfamethoxazole-reac- erocyclic ring at N 1 tive IgE antibodies. Note the absence of both an unsubsti-

[email protected] 6.2 Antimicrobials Other than Antibiotics 211 gen atom, attached to the N 1 nitrogen of the matory drug containing the sulfonamide group. sulfonamide group (Fig. 6.10 , Table 6.2 ). The same reaction on the patient’s lips occurred 4 Loose usage of the term “sulfa allergy,” the months later after taking rofecoxib and a chal- generally poor understanding of the mechanism lenge 1 month later with a low dose of the drug of sulfonamide-related adverse reactions, and again elicited the same response. Most interest- some reports of patients reacting to non- ing of all was the demonstration of an identical antimicrobial sulfonamides have sometimes led reaction at the same location 2 weeks later when to a situation where all sulfonamides, regardless the patient was challenged orally with sulfa- of chemical structure, are considered contraindi- methoxazole–trimethoprim. Findings such as cated in patients with a history of so-called “sulfa these suggested that cross-reaction between dif- allergy.” The presence of the sulfonamide group ferent antimicrobial and non-antimicrobial sul- in such a wide variety of frequently used drugs fonamides may occasionally occur, or are at least raises the question of possible allergenic cross- possible, but based on our current knowledge of reactivity between any, or perhaps even some, the chemistry, metabolism, immune responses, drugs because of a common sulfonamide group. and clinical fi ndings, cross- reactions are cur- In immediate hypersensitivity reactions to anti- rently thought not to occur. microbial sulfonamides, IgE antibody-mediated In the light of this background, it is interesting cross-reactivity may occur between drugs in this and worthwhile to consider a retrospective cohort group and cross- reactivity has been demonstrated study that examined the risk of an allergic reaction in some delayed reactions, so is the presence of a within 30 days after receipt of a non-antimicrobial common sulfonamide structure in different non- sulfonamide in 969 patients who had a prior aller- antimicrobial drugs also recognized in some gic reaction to a sulfonamide antimicrobial and in immediate and delayed reactions? This does not 12,257 patients who experienced no allergic reac- seem likely since compounds such as furose- tion after receiving a non-antimicrobial sulfon- mide, tolbutamide, celecoxib, and so on amide. Analysis showed that patients with a prior (Fig. 6.13 ) do not contain the N1 aromatic hetero- allergy to an antimicrobial sulfonamide were more cyclic ring substituent necessary for recognition likely than the nonallergic patients to react with a by IgE antibodies and apparently necessary for non-antimicrobial sulfonamide with the percent- adverse reactions such as hypersensitivity syn- ages of reactors being 9.9 and 1.6 %, respectively. drome and lethal toxidermias to occur. However, These fi ndings indicated that allergy to an antimi- a few case reports indicated that cross-reactions crobial sulfonamide is a risk factor for a subsequent seem to occur among various sulfonamide- reaction to a non-antimicrobial sulfonamide, but containing drugs, but there has been no universal the risk factor is even greater for patients with a acceptance of this conclusion. In one case, skin prior hypersensitivity to a sulfonamide antimicro- tests on a patient following an anaphylactic reac- bial who received penicillin (14 %) compared with tion to furosemide showed positive reactions to patients with no prior hypersensitivity to a sulfon- the culprit drug and sulfamethoxazole 0.03 mg/ amide antimicrobial who received penicillin (2 %). ml. In an attempt to rule out allergic recognition Interestingly, in trying to get these results into some of the para-amino group linked to the benzene sort of overall perspective of drug allergy, the risk ring, an epicutaneous test with p -phenylenedi- of an allergic reaction to a non-antimicrobial sul- amine and parenteral challenge with acetamino- fonamide in patients previously allergic to a sulfon- phen (paracetamol) and procaine were carried amide antimicrobial proved to be lower than the out. Both challenges were tolerated. Another risk of a reaction to a penicillin in sulfonamide example that seems to indicate cross- reactivity antimicrobial-allergic patients. Lastly, the risk of between a non-antimicrobial sulfonamide and an allergic reaction to a non-antimicrobial sulfon- sulfamethoxazole is contained in a report describ- amide was lower in patients with a history of sul- ing a fi xed drug eruption on the lips of a patient fonamide antimicrobial hypersensitivity than in after taking rofecoxib, a nonsteroidal anti-infl am- those with a history of penicillin hypersensitivity.

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Summing up then, it seems that hypersensitivity to navir commonly causing skin rashes and rarely a sulfonamide antimicrobial is a risk for a subse- Stevens–Johnson syndrome, and warnings about quent reaction to a non-antimicrobial sulfonamide, taking these protease inhibitors if there is a but prior penicillin allergy is at least as great, or an known hypersensitivity to a sulfonamide, there even greater, risk. The most likely explanation for so far appear to be no reports of clear cases of the conclusions of this study is that the patients allergic cross- reactivity with other sulfonamide showed a general predisposition to allergic reac- drugs. Dapsone, or diamino-diphenyl-sulfone, is tions rather than simply allergenic cross- reactivity not, by deﬁ nition, a sulfonamide, but it contains with drugs containing the sulfonamide group. The a sulfone group and the equivalent of the N 4 message for prescribers who are anxious to know arylamine moiety of sulfonamide antibacterials. whether or not there is broad cross- reactivity Dapsone is well known to cause the so-called amongst sulfonamide drugs is simply that there is dapsone hypersensitivity syndrome with fever, an increased risk to drugs in general, rather than rash, lymphadenopathy, and some organ involve- just sulfonamides, in patients with a history of ment (this syndrome may also be referred to as allergy of any type to sulfonamides or penicillins. DRESS—drug reaction [or rash] with eosino- The general message for drug allergy in the clinic philia and systemic symptoms; see Sects. 2.2.4.5 is not new and has been demonstrated before with and 3.6.3.5 ). Probably as a consequence of the a variety of different drugs: the history of an introduction of multidrug therapy for leprosy adverse drug reaction increases the risk of a subse- worldwide, reports of dapsone hypersensitivity quent adverse drug reaction. syndrome have increased dramatically in recent years. Other non-antimicrobial arylamines with The Possibility of Cross-reactions with Non- the equivalent N4 structure but without a sulfone antimicrobial Sulfonylarylamines and Arylamines group such as benzocaine and procainamide Some early research investigators concluded (in should also be looked at closely (Fig. 6.14 ). the absence of convincing and clear supporting Procainamide is known to occasionally induce data) that the N 4 arylamine moiety of sulfon- drug fever and some other allergic reactions amides like sulfamethoxazole is a major deter- including anaphylaxis and reports of anaphy- minant recognized by IgE antibodies. This laxis to benzocaine are readily found in the lit- therefore raises the questions of the safety in erature. There appears to be few, if any, reports patients allergic to antibacterial sulfonamides of of hypersensitivity responses to the β-blocker non- antibacterial sulfonamides containing an acebutolol. Hence, with all of these other sulfon- unsubstituted arylamine group at the N4 position amides and arylamines without a sulfone group, as well as drugs with such an unsubstituted no available data indicate that adverse reac- arylamine group but lacking a sulfonyl group. tions to the drugs are linked to their structure or The sulfonylarylamine antiretrovirals amprena- that they cross-react allergenically with sulfon- vir and fosamprenavir, used in the management amide antimicrobials. Nevertheless, it is certain of HIV-infected patients, contain a 4-amino ben- that reactions to the antiretrovirals, dapsone, and zenesulfonamido group, and although they lack both local anesthetics have not been investigated the required aromatic heterocyclic ring structure with the aim of identifying the precise structures attached at the N 1 nitrogen of sulfonamide anti- provoking hypersensitivity reactions nor have microbials, it seems possible, on structural investigations of possible allergenic cross-reac- grounds, that cross-reactions might occur tivity with sulfonamide antimicrobials been pur- with sulfonamide-reactive IgE antibodies and sued at the structural level. All of this means that in delayed hypersensitivity responses. while the risk of reactions to sulfonylarylamines Fosamprenavir is a pro-drug metabolized to like fosamprenavir, to dapsone, and to arylamines form amprenavir (Fig. 6.14 ), a protease inhibitor like benzocaine appears small, prescribing these used to treat HIV infection. Although there are drugs for patients allergic to sulfonamide antimi- some references in the drug literature of ampre- crobials should not be avoided. Nevertheless, it

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CH 3 HO OH P H C O O O 3 H O N NO O CH S 3 O O O H2N H2N Benzocaine

Fosamprenavir

CH3 CH3 O

N CH3 H3C OH N H H O N NO S H2N O O O Procainamide H2N

Amprenavir OH H OO O N CH3 S O

CH3 CH3 H3C N H H2N NH2 O Dapsone Acebutolol

Fig. 6.14 The sulfonylarylamine antiretrovirals amprenavir attached at the N1 nitrogen necessary for allergenic cross- and fosamprenavir both contain a 4- aminobenzenesulfona reactivity with sulfamethoxazole-reactive IgE antibodies mido group. Both drugs and dapsone, diamino-diphenyl- (see Fig. 6.10). Arylamines such as benzocaine, procain- sulfone, used in the management of HIV-infected patients amide, and acebutolol which lack a sulfonyl group also fail lack the required aromatic heterocyclic ring structure to react with anti-sulfamethoxazole IgE antibodies

should be remembered that drug allergy is personal experiences of laboratory testing and replete with examples of interesting reactions investigations over many years, one might expect and speciﬁ cities detected in the rare individual trimethoprim drug allergy to be far more inten- and the clinician should be aware of this and sively studied and worked out than it is. This con- remain watchful. clusion is supported by patient responses, written and verbal, to questions about apparent trimethoprim “allergy,” by numerous reports of adverse 6.2.2 Trimethoprim reactions to co-trimoxazole over a long period and the early demonstrations of true, IgE- 6.2.2.1 Trimethoprim antibody- mediated reactions to the drug. There and Hypersensitivities seems to be a major contributing factor to this The subject of trimethoprim-induced hypersensi- state of affairs. Sulfonamide antimicrobials, tivities is not straightforward in that the literature particularly sulfamethoxazole, have been the leaves one with the impression that the real situa- subject of interest and study by many clinical tion may not be in full view. From the authors’ and laboratory- based research groups since

[email protected] 214 6 Other Antimicrobial Drugs sulfonamides were introduced into medicine and and sulfamethoxazole. It was therefore surprising especially since co-trimoxazole, the combination to see a published study as late as 1998 with the of sulfamethoxazole and trimethoprim, was stated aim of determining whether the trime- introduced (see sulfonamide antimicrobials, this thoprim component of co-trimoxazole can cause chapter). Adverse reactions to the sulfonamide an anaphylactic reaction and the conclusion that quickly became apparent and interest in these anaphylaxis to the drug combination is not reactions was spurred by the drug combination’s always caused by sulfamethoxazole was equally high incidence of adverse reactions in immuno- surprising since it had already been clearly compromised HIV-infected patients where it had established. This was again, a further example of become recognized as a highly valuable treat- a mind-set that prejudged the inherent allergenic- ment in combating P. jirovecii infections. ity of sulfamethoxazole to the exclusion of a drug Although there is little doubt that many already well known to occasionally provoke co-trimoxazole-induced adverse reactions are severe type I allergic responses. provoked by the sulfamethoxazole component, perusal of many studies shows that it is often 6.2.2.2 Type I IgE Antibody-Mediated assumed to be the culprit drug without thorough Reactions to Trimethoprim investigation of the possible contribution of the Studies have been few but the diagnostic proce- trimethoprim component. It is not unusual to read dures applied so far for trimethoprim and sulfa- papers on hypersensitivity reactions to co- methoxazole, and the immunochemical defi nition trimoxazole where all clinical or laboratory of the drug allergenic determinants, have pro- investigations are directed at sulfamethoxazole vided a fi rm basis for the clinician to confi dently and trimethoprim is not even mentioned let alone diagnose and distinguish immediate allergic investigated. reactions to these two drugs. Whether trimethoprim is relatively free of Skin testing details for trimethoprim are hard delayed hypersensitivity effects or whether its to fi nd and no validation studies for skin testing apparently less troubling adverse effects are due with this drug appear to have been done, but for to investigative neglect, it does appear that the prick tests on patients with suspected allergy to drug has not greatly worried allergists and der- co-trimoxazole, Septrin Parenteral Infusion® matologists. Apart from rare reports of hypersen- (Wellcome) containing 16 mg/ml of trimethoprim sitivity pneumonitis, some adverse skin reactions (0.055 M) and 80 mg/ml of sulfamethoxazole is including erythematous papular skin eruption, useful in an initial examination. For intradermal and toxidermias including toxic epidermal testing, trimethoprim is dissolved in the mini- necrolysis, almost all reports of trimethoprim- mum quantity of 0.1 M sodium hydroxide before induced hypersensitivities are of the immediate diluting with physiological saline to dilutions of kind. In 936 reports of drugs frequently associ- 0.01, 0.1, and 1 mg/ml (0.034 M) and injecting ated with anaphylaxis (that is, when a causal rela- 0.02 ml quantities. Physiological saline contain- tionship was judged to be certain or probable) in ing the same amount of sodium hydroxide is used The Netherlands between 1974 and 1994, sulfa- as a control solution. There are other reports of methoxazole with trimethoprim was implicated successfully employing trimethoprim at a con- in 12 and trimethoprim alone in 11 cases. In addi- centration of 20 mg/ml, that is, 0.069 M, for skin tion, a number of individual case reports of ana- prick testing. phylaxis to trimethoprim, some with and some IgE antibodies to trimethoprim were fi rst without accompanying investigations for evi- demonstrated in the mid-1980s. Immunochemical dence of IgE antibodies, have been published. A investigations employing trimethoprim cova- 1996 study of eight patients who experienced lently coupled via a spacer arm to Sepharose in anaphylaxis to co-trimoxazole (discussed below) quantitative hapten inhibition experiments with revealed seven patients with IgE antibodies to tri- carefully selected analogs provided insights into methoprim and one with IgE to both trimethoprim the precise structures of the drug recognized by

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with an extensive range of analogs of the trimethoxybenzyl and diaminopyrimidine rings of trimethoprim identifi ed two structures complementary to the IgE antibody combining sites, again the 3,4- dimethoxybenzyl group but also the combined trimethoxybenzyl and diaminopyrimidine rings of trimethoprim. In an attempt to more precisely defi ne the fi ne structural specifi city differences of trimethoprim allergenic determinants, further quantitative hapten inhibition studies Fig. 6.15 Specifi c inhibition by trimethoprim and were undertaken with sera from eight patients, all some structurally related compounds of the reaction of females, who presented after immediate allergic IgE antibodies from a trimethoprim-allergic patient with the drug. The IgE antibodies showed specifi city reactions to co-trimoxazole. Immunoassays with for the 3,4- dimethoxybenzyl group of trimethoprim. trimethoprim and sulfamethoxazole-solid phases Key to symbols: (open circle ) trimethoprim; (fi lled revealed that all the patients had IgE antibodies to circle ) 6- hydroxytrimethoprim; (open square ) trimethoprim while one also showed a weak IgE 6- chlorotrimethoprim; ( fi lled square) diaveridine; ( fi lled triangle ) 3,4-dimethoxyphenylethylamine; (vertical open antibody response to the sulfonamide. Three dis- diamond) 3-(3′,4′,5′-trimethoxyphenyl)-propionic acid; tinct patterns of inhibition were seen (Table 6.3 ). ( fi lled diamond ) 3,4-dimethoxybenzoic acid; (inverted In group 1, comprising sera I1, I2, A1, A2, and I3, triangle) 4-methoxyphenylethylamine; ( horizontal open diaveridine, which differs structurally from trim- diamond) 3,4,5-trimethoxycinnamic acid. See also Table 6.3 and Fig. 6.16 (reproduced with permission from ethoprim by the absence of a single methoxy group, Smal MA et al. Allergy 1988; 43: 184) was essentially equally as potent an inhibitor as the “parent” compound. Other compounds representing the trimethoxybenzyl end of the trimethoprim molecule such as 3,4,5-trimethoxycinnamic acid complementary IgE antibodies in the sera of and 3,4-dimethoxyphenylethylamine were also trimethoprim- allergic patients. Initial examina- signifi cant inhibitors while structures with only tions with sera from two allergic patients showed one methoxy group attached to the ring, e.g., the clear presence of trimethoprim-binding IgE 4-methoxyphenylacetic acid, and structures rep- antibodies and inhibition patterns of antibody resenting the other end of the trimethoprim mol- binding indicated specifi city of binding and ecule, e.g., 2,4,6-triaminopyrimidine and the probable existence of more than one aller- 2-amino-4-hydroxypyrimidine, were essentially genic determinant on the trimethoprim molecule. inactive. With serum I4, once again trimethoprim In the example shown (Fig. 6.15), the most potent and diaveridine were equally well recognized, inhibitors (that is, the structures showing the best but other compounds lacking one end or the other fi t for the IgE combining sites) were diaveridine of the trimethoprim structure were either very and 3,4-dimethoxyphenylethylamine and these and weak or non-inhibitors. Group 3 showed the sim- the other inhibition results indicated that the tri- plest profi le with only trimethoprim showing methoprim determinant recognized was the 3,4- inhibitory activity; signifi cantly, diaveridine was dimethoxybenzyl group which is almost identi- without activity. Sulfamethoxazole showed no cal to one-half of the trimethoprim molecule. signifi cant inhibition with six of the sera, but the Reinforcing this conclusion was the almost com- small amount of inhibition seen with sera I4 and plete absence of inhibition seen with structures I5 might indicate weak cross-reactivity with the representing the other half of the trimethoprim pyrimidine ring of trimethoprim (Table 6.3 ). molecule, viz., 2-amino-4-chloro-6-methylpyrim- Taken together, these data demonstrate the exis- idine, 2-amino-4-hydroxy-6-methylpyrimidine, tence of at least three different trimethoprim and 4-amino-5-aminomethyl-2-methylpyrimi- allergenic determinant structures complementary dine. More detailed follow-up investigations to the combining sites of trimethoprim-reactive

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Table 6.3 Inhibition of trimethoprim–IgE antibody interactions in the sera of trimethoprim-allergic patients by trimethoprim and some structurally-related compounds % Inhibition of IgE antibody binding to TMP–Sepharose with 100 nmol of compound in sera Group 1 Group 2 Group 3 Compound and structure I1 I2 A1 A2 I3 I4 I5 A3 Trimethoprim 94 96 92 84 91 94 72 49

CH3O H2N N CH3O CH2 NH2 N CH3O Diaveridine 76 90 97 85 92 86 3 0

CH3O H2N N CH3O CH2 NH2 N 3,4,5-Trimethoxycinnamic acid 87 60 84 40 50 14 10 0

CH3O

CH3O CH=CH COOH

CH3O 3,4-Dimethoxyphenylethylamine 63 76 93 60 80 24 11 0 CH3O

CH OCHCH NH 3 2 2 2 3,4-Dimethoxybenzoic acid 35 41 68 0 26 9 20 0

CH3O

CH OCOOH 3 4-Methoxyphenylacetic acid 3 0 5 0 0 17 18 0

CH OCHCOOH 3 2 2,4,6-Triaminopyrimidine 2 10 0 3 10 10 25 0

H2N N NH2 N H2N 2-Amino-4-hydroxypyrimidine 9 0 0 2 0 0 13 0 HO N NH2 N Sulfamethoxazole 6 3 10 13 13 24 25 0

CH3 H NSONH 2 2 O N (continued)

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Table 6.3 (continued) % Inhibition of IgE antibody binding to TMP–Sepharose with 100 nmol of compound in sera Group 1 Group 2 Group 3 Compound and structure I1 I2 A1 A2 I3 I4 I5 A3

Allergenic determinants identiﬁ ed NH2 NH2 NH2 NN NN NN

H2N H2N H2N

CH2 CH2 CH2

H3CO OCH3 H3CO OCH3 H3CO OCH3 OCH OCH 3 3 OCH3 3,4-Dimethoxybenzyl group 2,4-Diamino-5-(3′,4′- Entire dimethoxybenzyl) trimethoprim pyrimidine group molecule Figures highlighted in bold indicate inhibition percentages considered to be clear, unequivocal, and signiﬁ cant in interpreting antibody combining site structure–activity relationships. See also Fig. 6.16 ) Reproduced from Pham NH et al. Clin Exp Allergy 1996;26:1155 with permission

IgE antibodies in the sera of patients allergic to the drug (Fig. 6.16 ). One of the determinants, the Space-filling model 3,4-dimethoxybenzyl structure represents one- half of the trimethoprim molecule while the other two comprise the entire, or almost the entire, molecule. Of the latter two determinants, 2,4-diamino-5-(3′,4′-dimethoxybenzyl)pyrimidine differs from the third determinant, the entire trimethoprim structure, by a single methoxy group demonstrating the importance a single, a small structural feature can have in antibody– drug recognition. Drug allergenic determinants, like antigenic determinants on a wide variety of peptide and non-peptide structures, show structural heterogeneity and it is likely that the IgE antibody response to trimethoprim in individual b patients is also heterogeneous. While sera I5 and A3 appeared to contain antibodies of only one specifi city, serum I4 may also have contained these antibodies in addition to the identifi ed population and sera in group 1 probably contained different sized populations of antibodies of all three specifi cities. c 6.2.2.3 T Cell Studies Fig. 6.16 CPK space-fi lling models of trimethoprim with These studies are in their early days. An attempt the three IgE antibody-binding determinants shown in to look at trimethoprim hypersensitivity by green , yellow, and blue . ( a ) The 3,4-dimethoxybenzyl investigating drug metabolism, processing in determinant shown in green; ( b ) 2,4-diamino-5-(3′,4′- dimethoxybenzyl)pyrimidine determinant shown in yel- antigen presentation, and cross-reactivity pat- low ; ( c) the whole trimethoprim molecule shown in blue terns has recently been reported. It seems that comprises the allergenic determinant for some anti- antigen presentation can be dependent on or trimethoprim IgE antibodies. See also Table 6.3

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Fig. 6.17 Structures of a O O some quinolones, a broad 5 4 COOH family of synthetic 6 3 F chemotherapeutic 7 antibacterials based on the 2 N1 N N 4-quinolone ( a ) and 8 H C H 1,8- naphthyridine ( b ) HN 2 5 structures. Norfl oxacin and nalidixic acid, respectively, 4-Quinolone Norfloxacin are examples of a drug from each of these groups. b O Cinoxacin is an example of 5 4 a quinolone antibacterial COOH 6 3 based on cinnoline, an aromatic heterocyclic with 7 2 H C N N 3 N N two attached six-mem- 8 1 bered rings containing C2H5 adjacent nitrogens at 1,8-Naphthyridine Nalidixic acid positions one and two ( d ). The basic fl uoroquinolone pharmacophore is shown in ( c ). The addition of a c R fl uorine atom at position 2 O 5 4 six of the two-ring nucleus F 6 COOH produced a 100-fold 3 increase in the antibacterial 2 minimum inhibitory 7 R3 X N concentration 8 1 R X = C or N 1 Fluoroquinolone pharmacophore

d O 5 4 O COOH 6 3

7 N N 2 N O N 8 1

C2H5 Cinnoline Cinoxacin

independent of processing. Cross-reactivity 6.2.3 Quinolones studies showed that trimethoprim clones were stimulated by the close analog diaveridine and 6.2.3.1 Structures by pyrimethamine but not by other closely Quinolones is the name given to a broad family related structures. Presumably attempts will be of synthetic chemotherapeutic antibacterials made to work out a possible immune chemically based on the 4-quinolone and mechanism(s) for trimethoprim hypersensitiv- 1,8-naphthyridine structures. The structures of ity based on the involvement of metabolites and these two quinolone nuclei and an example of a hapten–protein complexes as was done for drug from each of these groups are shown in sulfamethoxazole. Fig. 6.17a, b. Cinoxacin is an example of a

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1.O 2. O COOH F COOH N

N N N N N

HN C2H5 HN Pipemidic acid Ciprofloxacin

3. O F COOH

N N

N O CH H C 3 3 H Levofloxacin

4. O O F COOH F COOH

H N H N N N HN F OCH3 H H N 2 H Moxifloxacin Trovafloxacin F Fig. 6.18 Examples of some quinolone/naphthyridine antibacterials illustrating the structural developments over the four generations (1–4) of the drugs quinolone antibacterial based on cinnoline, an lococci and streptococci; and produce agents aromatic heterocyclic with two attached six- with adequate pharmacodynamic profi les. These membered rings containing adjacent nitrogens changes were effected by chemical incorpora- at positions one and two (Fig. 6.17d ). The addition of judiciously selected groups such as the tion of a fl uorine atom at position six of the two- cyclopropyl substituent used to replace the N -1 ring nucleus produced a 100-fold increase in the ethyl of norfl oxacin to gain the increased bio- antibacterial minimum inhibitory concentration availability seen with ciprofl oxacin. Another of the resultant fl uoroquinolones. The basic fl u- example is the addition of a piperazine ring to oroquinolone pharmacophore is shown in the C-7 position (Fig. 6.18 ) to increase activity Fig. 6.17c . Nalidixic acid, generally regarded as against gram-negative organisms. The range of the fi rst of the quinolone antibacterials, was quinolone and naphthyridines now available in used for urinary tract infections. It was quickly oral and parenteral forms provide far greater followed by other so- called fi rst generation and bioavailability and work against a much broader then by second-, third-, and fourth-generation range of organisms, including many diffi cult drugs as efforts were made to increase low tis- anaerobes, than the earlier generation drugs. sue concentrations; increase short half-lives and Examples of some quinolone/naphthyridine decrease the need for frequent dosage; broaden antibacterials illustrating the structural develop- the spectrum against Pseudomonas aeruginosa ments over the four generations of drugs are and gram-positive organisms such as the staphy- shown in Fig. 6.18 .

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6.2.3.2 Quinolones and 6.2.3.3 Immediate Hypersensitivity Hypersensitivities to Quinolones Chemists, pharmacologists, and toxicologists Immediate hypersensitivity reactions that may are always looking for likely structural corre- involve skin rashes, pruritus, respiratory distress, lates between chemical groupings and adverse and shock are the most frequently reported reactions to a drug. Some examples of this with immune-mediated reactions with a frequency of the quinolones include speculations concerning from 0.4 to 2 %. Descriptions of anaphylactic trovafl oxacin and hepatic eosinophilia, eosino- reactions to a number of different quinolone philia of the lungs and other immunological drugs, especially ciprofl oxacin but also including effects caused by tosufl oxacin, and the sus- nalidixic acid, pipemidic acid, pefl oxacin, ofl ox- pected involvement of temafl oxacin in a hemo- acin, norfl oxacin, levofl oxacin, and moxifl oxa- lytic uremic-like syndrome. Metabolites that cin, are not hard to fi nd. As with many other share structural features not yet identifi ed on drugs, reactions can occur on fi rst exposure so some quinolones especially the naphthyridones, sensitization by previously taking a quinolone are suspected of being the agents responsible for antibacterial does not seem to be required. some immunologically mediated reactions to Ciprofl oxacin has been the most commonly used these drugs. Many reports state that reactions to quinolone and the drug responsible for most of quinolone antibacterials are rare, but the litera- the anaphylactic/anaphylactoid reactions. A lit- ture does not always refl ect this. An incidence of erature search for the period 1960–2009 identi- adverse reactions of 2–10 % has been described fi ed 64 cases of anaphylactoid reactions as “fairly safe.” Included in the list of reactions considered to be probably related to ciprofl oxa- are gastrointestinal complaints, central nervous cin. Although the manufacturer of ciprofl oxacin system symptoms, anaphylactoid reactions, and lists pulmonary edema as an adverse event asso- skin conditions such as maculopapular and ciated with the drug, no reports of this reaction urticarial skin rashes, dermatitis, and vasculitis. were identifi ed in the survey. A frequency of 1 in 50,000 treatments has been reported for quinolone-induced immune- 6.2.3.4 Skin Tests mediated hypersensitivities while a recent study For the diagnosis of immediate allergic reactions of T cell-mediated reactions to quinolones to quinolones, skin tests including prick, intra- stated a combined incidence of 2–3 % for imme- dermal, and patch tests have been employed. diate and delayed hypersensitivities. One recent Skin tests with these drugs have been described large survey from Thailand covering a 4-year by a number of investigators as not useful for period looked at 166,736 patients treated with diagnosis, providing little information, and not fl uoroquinolones and found prevalences of suitable for determining specifi c tolerances to adverse and cutaneous adverse reactions of 0.13 individual drugs. Authors from a few other stud- and 0.09 %, respectively. Prevalences of ies, however, have concluded that quinolone skin between 0.04 and 0.37 % were seen for cutane- tests are a useful tool for the study of hypersensi- ous reactions to individual fl uoroquinolones tivity to quinolones. A highly suggestive history although the full range of quinolone drugs to of quinolone allergy was found to be associated which the population had been exposed over the with positive skin tests and skin tests were there- period and prior to it was not provided. Skin fore claimed to be helpful in predicting the results reactions have been reported with incidences of of challenge tests. The following quinolone drug up to 2.5 % with maculopapular exanthema and concentrations have been used for prick testing fi xed drug eruptions being the most common in subjects with suspected quinolone allergy: manifestations. Rare cases of Stevens–Johnson Ciprofl oxacin 0.02 mg/ml; levofl oxacin 5 mg/ml; syndrome and toxic epidermal necrolysis have pipemidic acid, ofl oxacin, norfl oxacin, levofl oxa- occurred. cin, moxifl oxacin 400 mg tablet ground up and

[email protected] 6.2 Antimicrobials Other than Antibiotics 221 suspended in physiological saline; trovafl oxacin deciding whether or not a challenge test should 200 mg tablet in saline. For intradermal tests: be employed. In one retrospective analysis, a Ciprofl oxacin 0.005, 0.02, and 0.05 mg/ml; levo- negative skin test preceded a negative challenge fl oxacin 0.005 and 0.05 mg/ml; moxifl oxacin test in 94 % of challenged patients and only 5 % 0.005 and 0.05 mg/ml. All the drugs were ini- of patients had a negative skin test and a positive tially used in prick tests and then injected challenge test. On the other hand, only 50 % of (0.05 ml) intradermally if the prick test result patients with a positive skin test had a positive proved negative. A recent study recommended a challenge test. In a typical challenge test, increas- nonirritant intradermal test concentration of ing doses of the drug are ingested or injected 0.0067 mg/ml for ciprofl oxacin. Skin testing with every 30 min until the therapeutic dose is reached quinolones has revealed a curious fi nding that so or until symptoms of a reaction occur. The patient far seems to have been neglected or ignored. At is kept in the clinic under observation for 45 min least fi ve separate studies have found positive after the last dose. A test is considered positive if skin test results to quinolone antibacterials in any signs or symptoms of the patient’s previous healthy control subjects. This observation is reaction occur within 24 h of the last challenge highly interesting and will be returned to below dose. The result is scored as negative if no sign of in the discussion on IgE antibodies to quinolones. drug hypersensitivity occurs after the usual thera- Patch testing does not seem to have been widely peutic dose is administered. Doses of some qui- used. In 101 patients with a history of hypersen- nolone drugs administered in a typical study are sitivity (immediate or delayed) to quinolones in ciprofl oxacin and levofl oxacin 50, 125, 250, and temporal relation to administration of fl uoroqui- 500 mg (one tablet); moxifl oxacin 40, 100, 200, nolones, 71 were excluded by tolerated oral chal- and 400 mg (one tablet). Blood pressure and lenge tests and patch testing was consistently heart rate are monitored after each dose, resusci- negative. Six patients developed anaphylaxis, and tation equipment and drugs are available, and interestingly, three of these were skin prick test challenges should not be undertaken without the positive and three were skin prick test negative to patient’s written informed consent. fl uoroquinolones. There appears to be no delayed reactions to In summary, skin testing to diagnose immedi- quinolone antibacterial drugs following chal- ate allergies to quinolones is currently not widely lenge tests or skin tests. and confi dently accepted and practiced although some investigators advocate its use, and even in 6.2.3.6 IgE Antibodies to Quinolones cases where correlations with challenge tests are Early attempts to demonstrate specifi c IgE anti- not perfect, results with some patients show good bodies to quinolones appear to have been correlation. unsuccessful due to diffi culties associated with binding the drugs to a suitable solid phase carrier. 6.2.3.5 Challenge Tests The fi rst successful demonstration of IgE anti- See Sect. 4.4 for a detailed discussion of drug bodies apparently specifi c for quinolone antibac- challenge tests. terials employed bis-oxirane coupling of drug to A number of investigators advocate the need the solid phase Sepharose and was carried out in to carry out challenge tests to confi rm allergic 1995–1996 in the authors’ laboratory in Sydney sensitivity or tolerability to quinolones. Some employing sera from Italian patients supplied by negative skin test results with positive oral chal- Professor R. Zerboni of Florence. Unlike Europe, lenge results have indicated the advisability of quinolones are not extensively prescribed in performing oral challenge tests before selecting Australia and cases of hypersensitivity are conse- quinolone as safe to administer or as a safe quently rare. Experiments were initially directed alternative drug. However, quinolone skin tests at determining the optimum position(s) for have still been claimed to be useful for the study coupling on the quinolone or naphthyridine of type I allergic responses since they help in nucleus and the best chemical procedure to

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Preparation of quinolone-solid phase conjugates:

R2 COOH

R3 N

Bis-Oxirane-activated R1 1. Carbodiimide/HSA Sepharose 2. Adsorb to NC discs Quinolone

O O

R2 COOH R2 CONH Protein Solid phase Solid phase N R N R3 3 R R1 1

N R3 = N : reactive non-reactive IgE binding N R3 N : non-reactive

Direct IgE-binding activity of different analogues of pipemidic acid:

Increasing similarity to pipemidic acid

O O N COOH N CH3 N NH N N N N N N N N NN HN HN HN HN HN HN C2H5

Piperazine Phenyl- Pyridinyl- Pyrimidinyl- Acetopyrimidinyl- Pipemidic acid piperazine piperazine piperazine piperazine

Little or no reactivity observed Increasing reactivity

Fig. 6.19 Preparation of quinolone-solid phase cova- ring, compounds representing increasing portions of the lent conjugates. Reactions were selected to attach an pipemidic acid molecule were selected, coupled to a insoluble carrier to each side of the two-ring nucleus as solid support, and examined for IgE antibody binding part of a strategy directed at determining the optimum with patients’ sera. Note that the names of these latter position(s) for coupling on the quinolone or naphthyri- compounds have been simpliﬁ ed for ease of inclusion in dine nucleus and ultimately achieving maximum binding the ﬁ gure (compare text) (adapted and reproduced with to drug-reactive IgE antibodies. To further evaluate the permission from Baldo BA Curr Opin Allergy Clin binding and importance of the substituted 4-quinolone Immunol 2001; 1: 327) achieve maximum binding to drug-reactive IgE bodiimide activation of the 3-carboxyl group antibodies. Reactions were selected to attach an and coupling to human serum albumin before insoluble carrier to each side of the two-ring adsorption of the drug–albumin complex to nitro- nucleus. With pipemidic acid for example, car- cellulose discs were effected by reactions on only

[email protected] 6.2 Antimicrobials Other than Antibiotics 223 one side of the molecule. For linkage of carrier nonallergic, healthy control subjects was to the other side of the molecule, base-catalyzed unexpected and diffi cult to interpret. In addition, nucleophilic addition of the nitrogen of the “normal” sera that reacted with the drug conju- piperazine ring attached at position seven of gate were inhibited by free pipemidic acid and the quinolone nucleus, to the epoxide of bis- some other quinolones, suggesting that the reac- oxirane-activated Sepharose, was utilized. Both tions seen were quinolone specifi c. These fi nd- strategies for linkage are represented in Fig. 6.19 . ings seemed to confi rm that quinolone-reactive Experiments using the drug-solid phases pre- IgE antibodies occur not only in sera from pared by coupling functional groups on each side quinolone-allergic patients but also in some of the molecule showed unequivocally that con- apparently “normal” controls, i.e., sera from indi- jugates prepared by coupling through the 3-linked viduals with no known or apparent allergic sensi- carboxyl group were not reactive with IgE anti- tivity to quinolone antibacterial drugs. The origin bodies in allergic patients’ sera while conjugates of these antibodies remains obscure but it should prepared by linkage to the opposite side of the be remembered that immediate allergic reactions molecule were clearly reactive. This indicated to quinolones, and to a number of other drugs that the allergenic determinant structure comple- (with the best example being neuromuscular mentary to the IgE antibody combining sites blocking drugs), often occur on fi rst exposure in comprised part of, or perhaps, the entire 3-car- subjects with no previously suspected allergy to boxy-4-quinoline ring. Total absence of antibody the drug. It would be interesting to determine the binding after coupling to the 3-carboxyl group frequency of occurrence of such subjects and sera suggests that it is probably an important feature in the normal and drug-allergic populations and of the allergenic site. To further evaluate the speculate on the likely possible source of sensiti- binding and importance of the substituted zation. This curious fi nding also adds interest to 4-quinolone ring, compounds representing the results showing positive skin tests in appar- increasing portions of the pipemidic acid mole- ently normal, healthy controls, results obtained in cule (Fig. 6.18) were selected, coupled to bis- a number of careful investigations. Sera from oxirane-activated Sepharose and examined for these subjects too should be tested for IgE anti- IgE antibody binding with patients’ sera. Little or body binding to quinolones. “Nonspecifi c” hista- no reactivity was found with piperazine and the mine release by direct action of quinolones on next three simplest derivatives of piperazine, mast cells has been offered by some as an expla- 1-phenylpiperazine, 1-(2-pyridyl)piperazine, nation for the “false” positive skin test results. and 1-(2-pyrimidinyl)piperazine (Fig. 6.19 ), This may indeed be the explanation but in one although one serum from a patient allergic to skin test study with quinolones where numbers quinolones reacted with 1-(2-pyridyl)piperazine. were given, 5 of 37 subjects who were classed as However, with the structure of the selected com- not hypersensitive to quinolones on the basis of pounds that showed the closest similarity to the negative challenge tests had positive skin tests. If parent drug, 1-(5-aceto-(2-pyrimidinyl))pipera- quinolone-induced histamine release were zine, clear IgE antibody-binding was obtained involved one might expect all, or perhaps even although antibody binding was weaker than the most, of the subjects to show skin reactions. binding seen with pipemidic acid. Although all of Some radioimmunoassay results obtained these fi ndings pointed to the development of a with bis-oxirane-linked quinolones using the successful assay for the detection of quinolone- strategy and chemical procedures described reactive IgE antibodies and inhibition studies above, were published from Europe in 2004 and with free quinolone drugs demonstrated specifi c- another drug–Sepharose radioimmunoassay for ity to quinolones and cross-reactivities between detection of IgE antibodies to quinolones has different quinolones, reactivity of the Sepharose recently appeared. In the former study, 30 of 55 conjugate with occasional sera from a normal, patients (54.5 %) who showed an immediate

[email protected] 224 6 Other Antimicrobial Drugs reaction to a quinolone drug yielded positive 6.2.3.7 Cross-reactions of Quinolones results when tested in the immunoassay 1–48 There is no doubt that immunologic cross- months after the reaction. The interval between reactivity between quinolone antibacterials exist; blood sampling and the reaction to the drug the questions are how extensive is it and what is may be important since radioactive uptakes were the clinical relevance? Views range from the need signifi cantly higher in subjects sampled within to avoid any quinolone if a patient is allergic to 8 months of the adverse reaction. In the more one, to the belief that some patients show good recent Sepharose-bis-oxirane-based radioimmu- tolerance to quinolones selected by drug chal- noassay tests for ciprofl oxacin, levofl oxacin, and lenge tests. For example, levofl oxacin was found moxifl oxacin were undertaken and shown to be to be a tolerated alternative for four out of fi ve positive for quinolone-reactive IgE antibodies in patients allergic to ciprofl oxacin and for two 12 of 38 patients with severe immediate allergic patients who reacted to norfl oxacin. A high reactions to quinolones. With the same subjects, degree of cross-reactivity between the fl uoroqui- 27 patients (71 %) were positive in the basophil nolones has been reported and cross-reactivity activation test when ciprofl oxacin, levofl oxacin, between fi rst- and second-generation quinolones and moxifl oxacin were analyzed together. The seems to be generally accepted. Overall though, high sensitivity of the basophil activation test in there seems to be no reliable and confi dent way this study indicates that the test may be a valuable of predicting cross-reactivity short of employing adjunct for drug provocation in the diagnosis of series of laborious and potentially dangerous immediate allergic reactions to quinolones, but challenges and this means that cross-reactions results with the test in some other hands have not need to be looked at patient by patient. been as encouraging. In three separate investiga- At the level of serum IgE antibodies, 9 of the tions, the test was positive in none of 12 patients 55 patients discussed above reacted to more than who had immediate reactions after oral adminis- one quinolone and 24 of 30 patients (80 %) tration of a quinolone, in none of four patients showed IgE antibody reactivity with more than with symptoms of anaphylaxis after an oral chal- one quinolone. The comment has been made that lenge with fl uoroquinolones, and in 17 of 34 cross-reactivity between quinolones “seems to patients who experienced an immediate hyper- be related to the molecular ring common to all sensitivity reaction within 1 h of quinolone quinolones,” and although this seems self-evi- administration. In the fi rst of these investigations, dent since cross- reactivity must ultimately have the authors concluded that the basophil activa- some common chemical basis, the chemical pic- tion test along with skin testing was not helpful ture is not so simple and straightforward. As the in establishing a diagnosis or in predicting cross- structures set out in Fig. 6.17 show, three differ- reactivity to quinolones. A very recent BAT ent ring systems are found in the so-called qui- study on 34 patients with immediate hypersensi- nolone antibacterial drugs and, regardless of the tivity to quinolones found 17 positive and 17 particular nucleus, different added substituents, negative to the suspected quinolone. Fifteen of particularly at positions one, six, seven, and the negative group tolerated the reintroduction of eight, lead to differences in chemical, physical, the drug (two were skin test positive) causing the and pharmacological properties between many investigators to state that a negative BAT is valu- individual drugs. These differences will include able information to consider when deciding the extent to which individual drugs are recog- whether or not to perform challenge tests on nized by IgE antibodies induced by a particular patients with a history of an immediate reaction quinolone. As outlined above and summarized in to quinolones. More studies are needed with the Fig. 6.19, the position of attachment of the solid basophil activation test before its diagnostic phase carrier necessary for development of an application to quinolone immediate hypersen- immunoassay to detect IgE demonstrated the sitivity is accepted and validated or further importance of the 3-carboxy-4-quinoline “face” of questioned. quinoline and naphthyridine derivative structures

[email protected] 6.2 Antimicrobials Other than Antibiotics 225 in reacting with antibodies. This immediately mononuclear cells proliferated in vitro in explains the poor recognition of some quinolone- response to ciprofl oxacin, norfl oxacin, or moxi- reactive IgE antibodies with cinoxacin that has fl oxacin in six patients with delayed hypersensi- an extra nitrogen at position two on that face. No tivity to the drugs. Patch tests were positive after doubt other IgE antibodies with different fi ne 24 and 48 h in three of the patients. Ciprofl oxacin- structural recognition spectra occur so one can and moxifl oxacin-specifi c T cell clones were predict that depending on this antibody heteroge- generated and used with different quinolone neity, on the point of attachment to the quinolone drugs to examine cross-reactions. Three patterns molecule of drug-solid phases, and on the selec- of cross-reactivity were observed: clones that tion of analogs chosen for side-by-side inhibi- reacted only with the eliciting drug and clones tion studies, fi ndings will differ, sometimes quite with limited or broad cross-reactivity. T cell markedly, depending on the quinolones exam- clones were said to be recognized directly with- ined for cross-reactivity. Once again, the most out processing within the immune system. informative and effi cient way to clarify the apparently complex question of allergenic cross- reactivity of a family of drugs is to carry out 6.2.4 Chlorhexidine carefully planned in vitro quantitative hapten inhibition studies using suitable IgE antibody Chlorhexidine is a synthetic cationic bis- immunoassays and a judiciously selected panel biguanide that can be viewed as two biguanide of quinolone drugs and other structural analogs groups each linked to a terminal 4-chlorophenyl some of which will not be therapeutic agents. group with the resultant chlorguanide structures For clinical relevance comparisons, the results connected by a hexamethylene bridge (Fig. 6.20 ). of such examinations should be looked at along- Since its introduction in 1954 as an antiseptic and side results of complementary skin test and disinfectant, chlorhexidine, usually as the gluco- perhaps basophil activation tests. It seems that nate or acetate salt, has found widespread appli- the picture of IgE antibody-mediated allergic cation in a myriad of products used domestically, recognition of the quinolone drugs is still sub- industrially, and in the medical environment. stantially incomplete. Some everyday products that include chlorhexidine are toothpastes, gargles, antiseptic creams, 6.2.3.8 Delayed Reactions liquid disinfectants, mouthwashes, contact lens to Quinolones fl uid, contraceptives, lubricants, and cosmetics. Non-immediate reactions to quinolones occur Medically, as an antimicrobial agent, it is in but they are not encountered as often as immedi- liquids, gels and creams, ointments, plasters, ate reactions and in-depth studies are so far few. dressings, and suppositories and is used on skin, Some of the more often-seen delayed reactions mucous membranes, wounds, and burns and to are fi xed drug eruptions and maculopapular disinfect surfaces and instruments. These lists are exanthemas where a T cell mechanism has nowhere near complete indicating how common been demonstrated. Specifi c T cell clones were and extensive is human contact with chlorhexi- identifi ed from patients with ciprofl oxacin- dine and as a result how easy is the opportunity induced maculopapular exanthems and about for contact with the compound. half of the clones proved cross-reactive with Anaphylaxis to chlorhexidine can occur, and related drugs. Reexposure studies in patients although rare, it is these reactions that have with exanthems revealed that cross-reactivity is aroused allergists’ attention to the agent. Many in fact lower than this. Cellular tests such as drugs provoke an occasional anaphylactic or lymphocyte transformation tests were judged to anaphylactoid response but with chlorhexidine be not very useful. However, the lymphocyte there are some features of the cases that were transformation test was said to have confi rmed unanticipated, surprising, and, initially at least, the involvement of T cells when peripheral blood obscure. The fi rst reported cases of anaphylaxis

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Model showing IgE-binding determinant Structure Space-filling model chlorhexidine and analogues

Chlorhexidine Cl H H H NH NH N N N N N N NH NH H H H Cl

4-chloro- bigua- hexame- bigua- 4-chlorophenyl nide thylene nide phenyl

Chlorguanide

H H H N N NCH3 NH NH CH Cl 3

4-chloro- bigua- phenyl nide

Alexidine

CH3CH2 H H H NH NH N N N (CH2)3CH3 CH3(CH2)3 N N N H H H NH NH CH2CH3

2-ethyl- bigua- hexame- bigua- 2-ethyl- hexyl nide thylene nide hexyl

Fig. 6.20 Chlorhexidine is a synthetic cationic bis-bigua- of chlorhexidine and the subterminal guanide structures nide that can be viewed as two biguanide groups each were non- inhibitory. Clear antibody recognition of alexi- linked to a terminal 4-chlorophenyl group ( yellow) with the dine demonstrated that the antibody combining sites resultant chlorguanide structures connected by a hexameth- extended beyond the chlorguanide structure and included ylene bridge (blue ). Space-ﬁ lling models showing the rela- the hexamethylene central sequence. Taken together, these tive allergenic (IgE antibody-binding) activities of different results showed clear antibody combining site recognition of regions of the chlorhexidine model. Only chlorhexidine, the identical arms of the chlorhexidine molecule, but the chlorguanide, and alexidine showed inhibitory activity of structure most complementary to the sites was the entire anti-chlorhexidine IgE with the parent compound being chlorhexidine molecule. See also Table 6.4 (adapted and most active (yellow and blue ). All of the compounds reproduced with permission from Pham NH et al. Clin Exp selected to represent the terminal chlorinated aromatic ring Allergy 2000 ; 30: 1001)

were in 1985 in Australia and Japan and the Japanese Ministry of Welfare between 1967 and ﬁ rst description of chlorhexidine-induced ana- 1984, nine cases were of anaphylactic shock and phylaxis via the urethral route appeared in 1992. all 50 cases were associated with mucosal appli- The latter paper appears to have been the fore- cation of the drug. The agency’s response was a runner of a number of reports of anaphylaxis recommendation that chlorhexidine should not elicited during urologic procedures when be used on mucous membranes. In one of the chlorhexidine was exposed to urethral and blad- original reports of anaphylaxis, cleansing the der mucosa. A review of the literature in 2002 skin with 0.5 % chlorhexidine acetate prior to a revealed that 21 of 66 cases of anaphylaxis to skin graft induced the reaction. This example, the chlorhexidine up to that time were precipitated fact that chlorhexidine is not a drug administered by urethral exposure to the drug. The same review by anesthetists, and its presence in what has been revealed that 27 patients had an immediate reac- described as “covert” forms all make it easy to tion after chlorhexidine was applied to mucous see why this widely distributed and used antimi- membranes and 16 patients experienced anaphy- crobial agent was, and presumably still is, often laxis after introduction of a chlorhexidine-coated overlooked as a provoking source of anaphylaxis. central venous catheter. Among 50 cases of The presence of chlorhexidine as a coating for adverse reactions to chlorhexidine reported to the catheters or in urethral lubricants, for example,

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Instillagel® which contains 0.25 % chlorhexidine, to the drug showed no positive reactions in 104 2 % lidocaine, and mixed hydroxybenzoates, subjects. This is at marked variance with a demonstrates why its involvement in inducing Japanese study of healthcare workers that allergic reactions might in some cases remain reported a 7 % incidence of contact dermatitis unrecognized. A survey in 2005 concluded that to chlorhexidine. chlorhexidine accounted for 27 % of overlooked perioperative hypersensitivity reactions. 6.2.4.2 Immediate Hypersensitivity Reactions to Chlorhexidine 6.2.4.1 Delayed Hypersensitivity Incidences of anaphylaxis to chlorhexidine are Reactions to Chlorhexidine not necessarily readily found or widespread and, Although reactions with an immediate type I in fact, there are no reports of any cases from mechanism are the most important chlorhexidine- many countries. This may refl ect the local induced allergic reactions medically and receive absence of chlorhexidine preparations, failure most attention, delayed hypersensitivity reactions to recognize and/or investigate reactions, the to the drug do occur and immediate and delayed different concentrations used in antiseptics in reactions can occur in the same patient. Exposure different countries, or absence of sensitivity to to chlorhexidine, usually prolonged, can lead to the agent. The Danish Anaesthesia Allergy contact sensitization and allergic contact dermati- Centre, established in 1998 to investigate patients tis and stomatitis. In a study designed to examine referred from all over Denmark, found 4 men of sources of exposure and sensitization to chlorhex- 21 patients with positive skin tests to various idine and to obtain information on the prevalence substances tested positive to chlorhexidine. of sensitization and contact allergy, over 7,500 Three cases of anaphylaxis to chlorhexidine general dermatology patients with suspected were reported in Finland up to 1999, 11 cases contact allergy in Finland were patch tested with were seen in Australia in the period 1985–1994 chlorhexidine digluconate 0.5 %. A positive test and 9 in Japan between 1967 and 1984. was seen in 0.47 % of patients with fi ve patients showing dermatitis or stomatitis caused by topical 6.2.4.2.1 Skin Tests for Diagnosis medicaments containing chlorhexidine and the In the case of the immediate reaction to 0.5 % antiseptic was judged to have contributed to chlorhexidine acetate aqueous solution men- current dermatitis in 11 patients. Only 16 sensi- tioned above, the same solution produced no tized patients could recall a history of previous reaction when tested on the patient’s skin but a exposure while four appeared to have had no 1:100 dilution injected intradermally produced, exposure. The conclusion was that creams, disin- within 1 min, a wheal that lasted more than fectants, and so-called oral hygiene products 30 min. In another diagnostic investigation of are the main sources of contact sensitization to chlorhexidine-induced anaphylaxis, application chlorhexidine and its presence in cosmetics may of chlorhexidine solution to the skin in dilutions delay the improvement in eczema in some of 1:10,000 to 1:1,000 proved negative but a 1 % patients. Chlorhexidine skin swabs, irrigation solution was strongly positive. In the same study, solutions, and skin “prep” solutions used prior skin prick tests using dilutions of 1:100 to to surgery have been associated with contact 1:10,000 were negative but stimulation with dermatitis which was found by patch testing to chlorhexidine was high in the sulfi doleukotriene occur with incidences of 2.5 % in a Danish skin stimulation test (CAST® , Buhlmann Laboratories, clinic and 5.4 % in atopic patients. In another Switzerland). In Korea, positive prick tests to study of dermatology patients, the incidence of chlorhexidine solution 5 % were obtained when type IV hypersensitivity to chlorhexidine was tested undiluted and at 1:10 and 1:100 and intra- determined to be between 1 and 2.5 %. In health- dermal tests with 1:1, 1:10, and 1:100 dilutions care workers in Denmark, skin tests (including also gave positive results. For prick testing, inves- patch testing) to detect type IV hypersensitivity tigators often utilize available stock solutions

[email protected] 228 6 Other Antimicrobial Drugs

(usually 1–5 %) either neat or a little diluted, years later when chlorhexidine linked to a bis- e.g., up to about 1:10. The Danish Anaesthesia oxirane-activated solid phase was used in quanti- Allergy Centre employs a standardized investiga- tative hapten inhibition studies with a serum from tion program in which the more widely used a patient who experienced anaphylactic episodes chlorhexidine digluconate is used at a concentra- on three separate occasions after exposure to ure- tion of 0.5 % in physiological saline for prick thral gel containing chlorhexidine and ligno- testing and 0.0002 % for intradermal testing. caine. Chlorguanide, which represents almost Prick tests are performed on the forearm with half the chlorhexidine molecule, alexidine, ami- saline and histamine controls. Development of a noguanidine, and arginine, which are similar to wheal greater than half the diameter of the the interior structure and some substituted chlo- wheal of the positive histamine control together rophenyl compounds that mimic the terminal with a negative saline control is considered posi- group at each end of the chlorhexidine molecule tive. Intradermal tests are performed on the back were selected for study of the chlorhexidine- or forearm with a saline negative control. A posi- reactive IgE antibody combining sites in the tive test is the development of a wheal (with fl are) patient’s serum. Only chlorhexidine, chlorgua- equal to or greater than twice the diameter of the nide, and alexidine proved inhibitory with, bleb formed from the volume of the injected expectedly, the parent compound being most solution. In practice, a positive test is a bleb with active (Table 6.4 ). A comparison of the 50 % a diameter of 8 mm or greater. These concentra- inhibitory concentrations of the three compounds tions used for skin testing are based on more than showed that chlorhexidine was from 100 to 200 800 negative controls for prick tests with 0.5 % times as potent as the two analogs. All of the chlorhexidine digluconate and more than 300 compounds selected to represent the terminal negative reactors to 0.0002 % chlorhexidine chlorinated aromatic ring of chlorhexidine and digluconate in intradermal tests. the subterminal guanide structures represented Skin prick tests on a patient who experienced by aminoguanidine and arginine were non- an anaphylactic reaction to Instillagel® produced inhibitory. Clear antibody recognition of alexi- results that draw attention to a potential pitfall dine demonstrated that the antibody combining when testing for sensitivity to this preparation. sites extended beyond the chlorguanide structure The demonstration of a positive skin test to and included the hexamethylene central sequence. chlorhexidine after the fi nding of a negative skin Taken together, these results showed clear anti- prick test to Instillagel® was attributed to the body combining site recognition of the identical presence of lidocaine in the gel and its effect in arms of the chlorhexidine molecule but the ablating the neurogenic wheal response required structure most complementary to the sites, that for a positive prick test. is, the structure showing the “best fi t,” was the entire chlorhexidine molecule (Table 6.4 ). 6.2.4.2.2 IgE Antibody Tests for Diagnosis Figure 6.20 shows the dominant features of the and Drug Allergen Studies fi ne structural recognition of chlorhexidine by IgE antibodies were fi rst implicated in an ana- the complementary IgE antibodies studied. phylactic reaction to chlorhexidine in Japan in Many of the drug allergenic determinants 1986. Specifi c skin-sensitizing IgE antibodies to identifi ed so far comprise parts of rather than chlorhexidine were demonstrated in the patients’ the entire molecule and complementary IgE sera by passive transfer and by an immunoassay antibodies show combining site heterogeneity using paper discs conjugated to chlorhexidine sometimes identifying two or more different linked to human serum albumin. Dose-dependent structures on a drug molecule (see Chap. 5 , inhibition of IgE antibody binding to the Sect. 5.1.2 , Chap. 6 , Sect. 6.2.2 and Chap. 7 , chlorhexidine-solid phase by both chlorguanide Sects. 7.4.2.2 and 7.5.1.2 for examples). It will be and the parent drug demonstrated specifi city of interesting to see if chlorhexidine demonstrates the reaction. Attempts to identify chlorhexidine such recognition heterogeneity. In the 1986 allergenic determinants were initiated nearly 20 Japanese study, the authors concluded that

[email protected] 6.2 Antimicrobials Other than Antibiotics 229

Table 6.4 Inhibition of the interaction of chlorhexidine–Sepharose complex with IgE antibodies by chlorhexidine and some structurally related compounds % Inhibition of IgE antibody binding to chlorhexidine − Sepharose in serum at Compound Structure 10 nmol 100 nmol Chlorhexidine H H 81.3 85.9 N N NH(CH2)3

NH NH Cl 2 Chlorguanide H H 34.5 64.0 N N NHCH(CH3)2

NH NH Cl Alexidine H H 40.5 53.2 CH (CH ) 3 2 3 N NNH(CH2)3 CHCH2 CH3CH2 NH NH 2 4-Chloro-acetanilide H 0 0 NCH3

O Cl 4-Guanidino-benzoic acid H 0 0 NNH2

NH HOOC

4-Chloroaniline NH2 0 0

4-Chlorophenyl- hydrazine NH NH2 0 0

Cl 4-Chlorophenol OH 0 0

Aminoguanidine H2NNHNH2 0 0

Arginine H2NNH(CH2)3CH(NH2)COOH 0 0

NH Compounds with % inhibition values in bold showed signiﬁ cant inhibition Reproduced from Pham NH et al. Clin Exp Allergy 2000 ;30:1001 with permission chlorhexidine and chlorguanide share the same protein binding might allow bridging of combin- antigenic determinant. In looking at the structure ing sites of adjacent mast cell-bound IgE mole- of chlorhexidine from the viewpoint of binding to cules leading to mediator release. This prediction IgE molecules, the identical structural features at was made and conﬁ rmed for neuromuscular each end of the molecule raise the possibility blocking agents where there are at least two that this inherent divalency without the need for allergenic determinants at distances apart of

[email protected] 230 6 Other Antimicrobial Drugs

1–1.45 nm (see Sect. 7.4.2.3 ). The terminal problem in some rare and hard to identify chlorophenyl groups of chlorhexidine are sepa- hypersensitivities. rated by 16 atoms of carbon and nitrogen and Hypersensitivity responses to povidone– the IgE-reactive chlorguanide groups are bridged iodine include a few clear-cut cases of anaphy- by a six carbon chain so the requirements of dis- laxis, generalized urticaria–angioedema, and tance and fl exibility needed for cross-linking at contact dermatitis, and, in almost all cases, PVP the mast cell surface may be satisfi ed. The chlor- not iodine has been implicated as the offending guanide structure has been shown to interact with component. Patch testing has been used to IgE antibody combining sites; it remains to be identify delayed reactions and for immediate seen if IgE antibodies with specificity restricted responses skin prick testing is the diagnostic to the terminal 4-chlorophenyl group occur. method of choice since it has been shown to per- A specifi c test to detect serum IgE antibodies form well at concentrations of povidone–iodine to chlorhexidine is now available in the form of 1 mg/ml (0.1 %) and PVP 35 mg/ml. Skin tests an ImmunoCAP ® (Thermo Scientifi c) test. The have generally been positive to povidone–iodine drug- solid phase is prepared by coupling 1-[ N5 - and PVP but not iodine, and in some other cases ( p - chlorophenyl)biguanido]-6-aminohexane where PVP was present as an excipient with other which represents half of the symmetrical bivalent drugs or agents, again PVP induced a positive chlorhexidine molecule to cyanogen bromide- response. It is therefore diffi cult to escape the activated cellulose sponge. conclusion that in the rare cases of hypersensitivity to preparations such as Betadine™, and perhaps in some other preparations where PVP has 6.2.5 Povidone–Iodine been included as a solubilizing, dispersing, sus- pending, or binding agent, as much attention Povidone–iodine is a chemical complex of ele- should be directed at PVP as the preparation’s mental iodone, 9–12 % (w/w), with polyvinyl- active agent(s). There is a report of hypersensi- pyrrolidone (PVP) producing an iodophor tivities to povidone–iodine where agents other preparation that slowly releases free iodine in than PVP seem to have been responsible. In a solution. PVP, also called polyvidone or povidone, patch test study of suspected contact dermatitis is a water-soluble polymer of N -vinylpyrrolidone, induced by the antimicrobial agent, all ten chains of which can range in molecular weight patients reacted positively to povidone–iodine from 10,000 to 700,000 Da. Sold under the name but no positive reactions were seen with PVP and Betadine™, povidone–iodine is a very effective one patient was positive to iodine. The authors antimicrobial agent, more stable than tincture of did state, however, that it was diffi cult to distin- iodine, and with the added advantages of lower guish between allergic responses and irritation. irritancy and toxicity. Bacteria do not develop Detection of IgE antibodies to PVP has been resistance to the preparation, it is active against reported in a subject who experienced an imme- Chlamydia, fungi, and viruses, including HIV diate reaction that included urticaria and angio- and Herpes , and its sensitization rate of 0.7 % is edema following topical application of low. Povidone–iodine is therefore widely used in povidone–iodine, but the weak reaction, inter- medicine for skin cleansing pre- and postopera- preted as a positive result after simple adsorption tively and for the prevention and treatment of of PVP to a microtiter plate, is not entirely infections of wounds, burns, ulcers, etc. The convincing. properties of PVP make it a useful agent for In summary, with povidone–iodine, the appar- formulating many pharmaceutical and other ently inert substance PVP added to improve the products, and because statement of its inclusion formulation, rather than the active component, is in formulations is not always mandatory, its likely to be the cause of cases of hypersensitivity presence is frequently overlooked, a potential occasionally seen with its use.

[email protected] Summary 231

range estimate of 1.5 % in the last 20 years but Summary more recent estimates are consistently in the range 8–9.5 %. • Allergies to macrolide antibiotics are said to • Some patients demonstrate contact allergy to occur with an incidence of 0.4–3 %. Apart both bacitracin and neomycin even though the from the very occasional case report of ana- two antibiotics have markedly different struc- phylaxis, the most commonly seen symptoms tures. Such co-sensitivities even extend to the include urticaria, often generalized, angio- cyclic peptide antibiotic polymyxin. edema, pruritus, asthma, tachycardia, and • Because of its greatly reduced usage, allergic delayed cutaneous reactions presenting as contact dermatitis is currently not a problem maculopapular exanthema. with streptomycin. Reports on anaphylaxis • Tetracyclines are viewed as being compara- to the antibacterial go back over 50 years tively safe drugs. Minocycline, widely used with a relatively high incidence of reactions for acne vulgaris, has been implicated in a occurring in the 1960s. serum sickness-like reaction, drug-induced • Delayed-type cutaneous reactions to clinda- lupus, cases of single organ dysfunction, and mycin include pruritus, exanthematous rash, drug hypersensitivity syndrome reaction. generalized maculopapular exanthema, eryth- • Adverse reactions to rifamycins suggested to roderma, generalized exanthematous pustulo- be immune mediated include a ‘flu’-like syn- sis, and Stevens–Johnson syndrome. drome, acute renal failure, hemolytic anemia, • Many of the reactions to sulfonamides involve and thrombocytopenia. Other more typical the skin and mucous membranes. The more manifestations of hypersensitivity include severe reactions that occur include potentially urticaria, contact dermatitis, erythema multi- lethal toxidermias and a delayed hypersensi- forme, and vasculitis. tivity-type syndrome characterized by fever, • The most commonly occurring adverse effects skin rash, and multi-organ toxicity. caused by vancomycin are referred to collec- • Immediate type I reactions are the most well- tively as red man syndrome. Reactions may defi ned sulfonamide-induced hypersensitivity range from mild pruritus, erythema, and fl ush- reactions with the best defi ned allergenic drug ing of the upper body to angioedema and structures. Sulfonamides with one methyl sub- rarely hypotension and cardiovascular col- stituent on a fi ve- or six-membered aromatic lapse. Reactions may be prevented or their heterocyclic ring on the carbon β to the sulfon- severity decreased by extending the infusion amido substituent are the structures most

time and/or premedication with histamine H1 complementary to anti-sulfamethoxazole IgE

and H 2 receptor antagonists. antibody combining sites. • Despite its chemical similarity with vancomy- • About 10 % of sulfamethoxazole is metabolized cin, teicoplanin does not appear to cause red to a reactive hydroxylamine intermediate by man syndrome. CYP2C9 and myeloperoxidase. The unstable • Both rapid (carried out over several hours) and hydroxylamine intermediate auto- oxidizes to slow (over a period of days) desensitization the highly reactive, toxic, immunogenic, and protocols for vancomycin sensitivity have allergenic nitroso-sulfamethoxazole. proved effective. • Nitroso-sulfamethoxazole, but not the parent • Neomycin consistently ranks in the top 10 % drug, can react covalently with cysteine resi- of the most common causes of allergic contact dues of cellular surface proteins including skin dermatitis. Neomycin is also known to cause keratinocytes, circulating peripheral blood generalized reactions such as exfoliative mononuclear cells and splenocytes, and serum dermatitis and erythroderma. proteins, predominantly immunoglobulins and • Measured incidences of bacitracin sensitivity, albumin, to form hapten–protein complexes. as low as 0.3 % in 1973, increased to a lower The hydroxylamine derivative may also be

[email protected] 232 6 Other Antimicrobial Drugs

involved in a number of adverse reactions Sepharose solid phase complex covalently including hepatitis, nephritis, thrombocytope- linked with bis-oxirane. Quinolone-reactive nia, lupus erythematosis, and the sulfonamide IgE antibodies are found in some “normal” hypersensitivity syndrome. sera. This fi nding adds interest to the results • Antigen-presenting cells alone may be suffi - showing positive skin tests in apparently normal, cient to generate metabolites of sulfamethoxa- healthy controls. zole, produce the hapten–protein antigen • Although reactions with an immediate type I complexes, and ultimately induce the T cell mechanism are the most important chlor- response to the drug. hexidine-induced allergic reactions and • A serious drawback to the use of sulfamethox- receive most attention, delayed hypersensitivity azole–trimethoprim in HIV-infected patients reactions to the drug do occur and immediate is a high rate of adverse reactions that can and delayed reactions can occur in the same range in frequency up to 50–60 % and require patient. discontinuation of therapy. • A specifi c assay to detect IgE antibodies to the • The presence of the sulfonamide group in a drug has been developed. The structure shown wide variety of frequently used non- to be most complementary to the antibody antibacterial drugs raises the question of combining sites was the entire chlorhexidine possible allergenic cross-reactivity between molecule. drugs with a common sulfonamide group. • Hypersensitivity responses to povidone– Based on the chemistry, metabolism, immune iodine include a few clear-cut cases of ana- responses, and clinical fi ndings, cross-reac- phylaxis, generalized urticaria–angioedema, tions are thought not to occur. This conclusion and contact dermatitis. Polyvinylpyrrolidone also seems to apply to the sulfonylarylamine not iodine has been implicated as the offending antiretrovirals amprenavir and fosamprenavir component. both of which contain a 4-aminobenzenesul- fomamido group but nevertheless, clinicians should remain watchful. • Almost all reports of trimethoprim-induced Further Reading hypersensitivities are of the immediate kind. At least three different trimethoprim allergenic Castrejon JL, Berry N, El-Ghaiesh S, et al. Stimulation of human T cells with sulfonamides and sulfonamide determinant structures complementary to the metabolites. J Allergy Clin Immunol. 2010;125: combining sites of trimethoprim- reactive IgE 411–8. antibodies have been identifi ed. One of the Cribb AE, Miller M, Leeder JS, et al. Reactions of the determinants, the 3,4-dimethoxybenzyl struc- nitroso and hydroxylamine metabolites of sulfamethoxazole with reduced glutathione. Implications ture represents one-half of the trimethoprim for idiosyncratic toxicity. Drug Metab Dispos. 1991; molecule; the other two comprise structures 19:900–6. that make up the entire, and almost the entire, Cribb AE, Lee BL, Trepanier LA, et al. Adverse reactions molecule. to sulfonamide and sulfonamide-trimethoprim antimicrobials: clinical syndromes and pathogenesis. • Immediate hypersensitivity reactions that may Adverse Drug React Toxicol Rev. 1996;15:9–50. involve skin rashes, pruritus, respiratory dis- Harle DG, Baldo BA, Wells JV. Drugs as allergens: detec- tress, and shock are the most frequently tion and combining site specifi cities of IgE antibodies reported immune-mediated reactions to qui- to sulfamethoxazole. Mol Immunol. 1988;25:1347–54. Lerner A, Dwyer JM. Desensitization to vancomycin. Ann nolones with a frequency of from 0.4 to 2 %. Intern Med. 1984;100:157. Skin testing to diagnose immediate allergies Lim KS, Kam PCA. Chlorhexidine – pharmacology and to quinolones is currently not widely and con- clinical applications. Anaesth Intensive Care. fi dently accepted and practiced. 2008;36:502–12. Parkes AW, Harper N, Herwadkar A, Pumphrey R. • IgE antibodies apparently specifi c for quino- Anaphylaxis to chlorhexidine component of Instillagel®: lones have been detected using a drug– a case series. Br J Anaesth. 2009;102:65–8.

[email protected] Further Reading 233

Pham NH, Weiner JM, Reisner GS, Baldo BA. Anaphylaxis Strom BL, Schinnar R, Apter AJ, et al. Absence of cross- to chlorhexidine. Case report. Implication of immuno- reactivity between sulfonamide antibiotics and sulfon- globulin E antibodies and identiﬁ cation of an allergenic amide nonantibiotics. N Engl J Med. 2003;349: determinant. Clin Exp Allergy. 2000;30:1001–7. 1628–35. Sahai J, Healy DP, Shelton MJ, et al. Comparison of van- Svensson CK, Cowen EW, Gaspari AA. Cutaneous drug comycin- and teicoplanin-induced histamine release reactions. Pharmacol Rev. 2000;53:357–79. and “red man syndrome”. Antimicrob Agents Wazny LD, Daghigh B. Desensitization protocols for van- Chemother. 1990;34:765–9. comycin hypersensitivity. Ann Pharmacother. 2001; Sheth VM, Weitzul S. Postoperative topical antimicrobial 35:1458–64. use. Dermatitis. 2008;19:181–9.

[email protected] Drugs and Other Agents Used in Anesthesia and Surgery 7

Abstract Neuromuscular blocking drugs (NMBDs) are the most common cause of anaphylaxis during anesthesia representing ~60 % of reactions and an incidence of 1 in 1,000–20,000. Reactions are mediated by IgE antibodies with speciﬁ city for tertiary and quaternary ammonium ions, but adjoining structures may also be recognized. Recognition of the substituted ammonium groups accounts for the extensive cross-reactivity between the NMBDs. Diagnosis of reactions is effected by skin testing with free drugs, IgE antibody assays (especially using a morphine-solid phase), and the tryptase assay. Reversal of rocuronium-induced NM block with the cyclodextrin sugammadex has highlighted the question of changed allergenicity of such chemically sequestered drugs in host–guest complexes. Anaphylactic reactions to the hypnotics thiopentone and propofol are rare. Both are diagnosed by skin testing and the former also by a speciﬁ c IgE test. True IgE-mediated reactions to local anesthetics are extremely rare; many reactions appear to be vasovagal responses, but delayed reactions are well known. Anaphylactic and other adverse reactions occasionally occur to colloids such as hydroxyethyl starch, gelatin, and dextrans, to the polypeptides protamine and aprotinin, and to heparins and patent blue V. Pre-injection of small MW dextran 1 reduces the incidence of dextran- induced anaphylaxis from 25 to 3 per 100,000.

7.1 Drug-Induced Reactions already sick patients. The administration of a During Anesthesia variety of drugs that alter patients’ normal physiological functions in a short time together with a Although anesthesia today is safer than it has range of symptoms, mild to severe, can make the ever been, it still involves signiﬁ cant risk due to a task of pinpointing the cause of an adverse reac- number of factors including exposure of patients tion a difﬁ cult one. Achieving this will depend on to a mixture of different drugs often given intra- careful consideration of the drug(s) involved, the venously and over a short time period, systems signs and symptoms of the patient’s reaction, the failure, and the presence of abnormalities in some temporal relationship of drug administration to

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 235 Relationships, DOI 10.1007/978-1-4614-7261-2_7, © Springer Science+Business Media, LLC 2013

[email protected] 236 7 Drugs and Other Agents Used in Anesthesia and Surgery symptoms, and the diagnostic tests employed. As basis for the reaction exists. This is a prime well as application of appropriate investigative objective once successful treatment of the reac- tests, the effi ciency and effectiveness in classify- tion has been effected. Data on the proportion of ing a reaction may depend on some factors that anaphylactic and anaphylactoid reactions from are not readily apparent. These include the edu- different surveys sometimes yield surprisingly cation of clinicians in managing adverse reac- different results, results that often originate in tions during and after the perioperative period, differences between a centralized recording and whether appropriate guidelines have been issued investigative methodology and a far less orga- and followed and whether a suitable reporting nized approach by a few individuals relying on process, spontaneous and/or statutory, is in place. spontaneous reporting of cases. In an example of Predictably, practices, guidelines, and regula- the latter approach, the drugs responsible for tions vary widely with some localities far in adverse reactions in 350 consecutive patients advance of others. In France for example, more during anesthesia were identifi ed and the under- attention is paid to the safety of anesthesia as a lying mechanisms of the reactions were studied result of rare serious drug-induced adverse events by intradermal testing, serum tryptase determina- and the reporting, registering, and subsequent tions, and drug-reactive serum IgE antibody tests. investigation of such events is more organized The reactions in 139 patients (39.7 %) were than elsewhere. This well-organized strategy has shown to be anaphylactic in nature, that is, enabled the regular and ongoing publication of evidence for an immunologic cause was found data on substances responsible for anaphylaxis in and the fi gures of 39.7 and 60.3 % for those expe- France from 1984 to 2004 with more recent riencing a drug-induced anaphylactic and anaphy- fi ndings no doubt following soon. When infor- lactoid reaction, respectively, are almost the exact mation is needed or sought on epidemiological, reverse of the fi gures obtained in three French clinical, and a number of associated features of surveys where the corresponding results are reactions under anesthesia, the value of the 1999–2000 66 and 34 %, 2001–2002 69 and French approach is apparent. However, whether 31 %, and 1997–2004 72.2 and 27.8 %. This dis- the organization and arrangements are as devel- parity may be a result of a reporting and investi- oped and sophisticated as in France or carried out gative system that, in the French surveys, sorts by only a few, or perhaps even a single individ- and selects patients more likely to have experi- ual, the interest and application of those in the enced a true type I immediate reaction. While the profession of anesthesiology dedicated to keep- French survey fi gures refl ect organization and ing up with the ever-present problem of adverse procedures that are preeminent for the study of reactions are essential. anaphylaxis to drugs during anesthesia, they may Symptoms of a pseudoallergic reaction, for not reveal the everyday situation of the wide example, one resulting from direct histamine range of adverse patient responses, many mild to release, may give the same clinical picture as a moderately severe, to the variety of drugs admin- true anaphylactic reaction, thus making it diffi - istered. For drug reactions during anesthesia one cult to distinguish the two. The same clinical pic- of the fi rst and most pertinent questions is what ture requires the same immediate management proportions of everyday adverse reactions are but, if for no other reason than patient safety dur- anaphylactic and non-anaphylactic (anaphylac- ing subsequent drug treatments and anesthesia, it toid)? Whether the proportion is 40:60 or 70:30 is apparent that the underlying mechanism of any will depend on a number of factors, in particular, adverse reaction must be identifi ed. This means criteria and guidelines for reporting, selection, establishing whether a drug-induced reaction and testing and this should be remembered when during anesthesia is anaphylactic, that is, immune considering drug allergy data collection and mediated, or anaphylactoid, where no immune interpretation.

[email protected] 7.2 Incidences of Drug- and Other Agent-Induced Anaphylaxis During Anesthesia 237

Table 7.1 Agents responsible for type I immediate 7.2 Incidences of Drug- and Other allergic reactions during anesthesia Agent-Induced Anaphylaxis Reaction (%) Reaction (%) During Anesthesia Agent Francea Australiab Neuromuscular blocking drugs 58.1 61.9 Succinylcholine 33.4 32.8 With the above proviso in mind, the most compre- Rocuronium 29.3 16.8 hensive collection of data on drugs involved in Atracurium 19.3 9.1 immunoglobulin E-mediated reactions during Vecuronium 10.2 5.6 anesthesia is contained in the ongoing French Pancuronium 3.6 1.9 Mivacurium 2.5 0.5 studies beginning with results of the 1984–1989 Cisatracurium 1.7 0.5 survey and in the long-standing Australian series Alcuroniumc 24.8 gathered and maintained for over 30 years by d-Tubocurarine 2.9 Gallamine 2.1 Malcolm Fisher at the Royal North Shore Hospital More than one drugd 2.1 of Sydney. In France, from 1984 until the most Hypnotics/Induction agents 2.3 10.4 recently published survey that covers the years Propofol 55.8 6.3 1997–2004, 60–70 % of immediate hypersensi- Midazolam 32.6 52.4 Thiopentone 9.3 30.2 tivity reactions that occur have been classifi ed as Ketamine 2.3 9.5 true type I IgE antibody-mediated reactions. Alfathesin 1.6 Mortality resulting from anaphylactic reactions Propanidid Methohexitone during anesthesia is in the range of 3–9 % depend- Latex 19.7 0.8 ing on the country. Neuromuscular blocking Antibiotics 12.9 8.6 drugs (NMBDs) have consistently been the most Penicillins 49.0 15.4 common cause of anaphylaxis far exceeding the Cephalosporins 37.0 73.1 Vancomycin 5.8 next most implicated agents latex, antibiotics, Others 14.0 5.8 colloids, hypnotics, and opioids in that order Colloids 3.4 4.6 (Table 7.1 ). Over the 20-year period of seven con- Gelatin 89.9 85.7 secutive surveys the incidence of reactions to Hetastarch 9.5 Albumin 1.6 NMBDs has ranged from a high of 81 % in the Dextran 70 14.3 1984–1989 survey to a low of 58.2 % in the 1999– Opioids 1.7 2.6 2000 survey, with the fi gure for 1997–2004 of Morphine 35.5 50.0 58.1 % being almost identical. IgE antibody- Fentanyl 22.6 25.0 Sufentanil 22.6 mediated reactions to latex (see Sect. 7.8.3) have Nalbuphine 12.9 shown upward movements since the fi rst survey Remifentanil 6.5 despite increasing awareness of the risk of latex Meperidine (Pethidine) 18.7 Omnopon 6.3 sensitization in children with spina bifi da and Other agents e,f 2.7 e 3.8 f healthcare workers and the adoption of measures No causal drug detected 7.4 to minimize risk such as the upgraded require- aSurvey in France 1997–2004; 1,816 patients. Data from ments for surgical gloves and making surgery a Mertes PM et al. J Allergy Clin Immunol. 2011;128:366 latex-safe environment. The incidence of type I b Ongoing Australian survey; 606 patients. Data from reactions to antibiotics also markedly increased Fisher MM et al. Acta Anaesthesiol Scand. 2011;55:99 c from only 2 % of the total in 1984–1989 to 15.1 % Discontinued d Eight reactions with two different neuromuscular blocking in the 1999–2000 survey. Allergic reactions to drugs administered antibiotics appear to be increasing with time with eMade up largely of patent blue, propacetamol, local penicillins and cephalosporins (see Chap. 5 ) anesthetics, aprotinin, and protamine f accounting for most of the increase. Comparing Made up largely of induction agent plus neuromuscular blocker (four patients), protamine, local anesthetics, patent the Australian survey involving 606 patients with blue, chlorhexidine, contrast media, and ondansetron

[email protected] 238 7 Drugs and Other Agents Used in Anesthesia and Surgery the French series, three features in particular are Table 7.2 Clinical features of anaphylactic and anaphy- noteworthy. NMBDs again predominate with an lactoid reactions during anesthesia incidence close to the French fi gure, reactions to Anaphylactic Anaphylactoid hypnotics, principally due to the induction agents Symptoms reactions (%)a reactions (%)b thiopentone and alfathesin, are markedly higher Cardiovascular 74.7 33.9 in the Australian fi gures and the difference in the Arterial hypotension 17.3 18.4 c incidence of latex anaphylaxis is remarkable Cardiovascular collapse 50.8 11.1 Bradycardia 1.3 0.7 (Table 7.1 ). Within the NMBDs, the surveys agree Cardiac arrest 5.9 0 that succinylcholine accounts for a third of reac- Bronchospasm 39.8 19.2 tions, but otherwise there are some striking differ- Cutaneous 71.9d 93.7e ences. Rocuronium, considered by some in Angioedema 12.3 7.7 Europe to be a risk for anaphylaxis (see Reactions in France 1999–2000 Sect. 7.4.4.3 ) and introduced in 1994, in other Data from Mertes PM et al. Anesthesiology 2003; 99: 536 words, many years after succinylcholine, still a 518 anaphylactic patients accounted for nearly 30 % of reactions in France. b 271 anaphylactoid patients c The Australian incidence is almost half that, but Sole feature in 6.2 % of cases d Sole feature in 9.7 % of cases alcuronium (see Sect. 7.4.4.3 ), now off the market e Sole feature in 50.2 % of cases for nearly two decades, represents almost one- quarter of the 375 Australian reactions to NMBDs. Both surveys for reactions to antibiotics should not be relied upon in making a diagnosis. reveal the β-lactam antibiotics to be the domi- However, symptoms in anaphylactic patients do nant culprit drugs with incidences of 86 and seem to be more severe—on a graded scale of 88.5 % in the French and Australian fi gures, clinical manifestations most anaphylactic reac- respectively. However, while penicillins were tions during anesthesia fall into grades 2 and 3 implicated a little more often than cephalospo- (see Sect. 2.2.1.1.4 ) while anaphylactoid reac- rins in the French experience (49–37 % of reactions are mainly grade 1. Most anaphylactic reactions to antibiotics), the number of reactions to tions during anesthesia occur within minutes of cephalosporins in Australian patients was nearly induction and, mainly, occur following intrave- fi ve times as great as the penicillins fi gure nous administration of the drug or agent. For both (73.1–15.4 %). anaphylactic and anaphylactoid episodes, adverse Adverse reactions to the penicillin and cephalo- cardiovascular reactions are a common and seri- sporin antibiotics, two of the clinically most impor- ous symptom which may progress to cardio- tant families of antimicrobial drugs but also two of vascular collapse if not treated. Multi-organ the most allergenic, are examined in detail in involvement is usually the case, but for anaphy- Chap. 5 and will not be discussed further here. The lactic patients in particular, cardiovascular col- opioid group of drugs are responsible for around lapse may be the only symptom with frequencies 2 % of reactions during the perioperative period. of occurrence of up to more than 80 % and the These histamine-releasing, clinically important sole feature in up to 60 % of cases. As with car- analgesics are dealt with separately in Chap. 8 . diovascular symptoms, bronchospasm is more often seen in anaphylaxis, but the incidence of cutaneous symptoms is far greater in anaphylac- 7.3 Clinical Features toid reactions (Table 7.2 ). In one survey, bron- of Anaphylactic and chospasm was seen in 19 % of patients who Anaphylactoid Reactions experienced anaphylaxis during anesthesia and it During Anesthesia was the sole clinical feature in 4.5 % of patients. Findings from a multivariant analysis placed in a Symptoms of anaphylaxis and an anaphylactoid clinical context suggest that a patient with response are often suffi ciently similar to make bronchospasm and hypotension in the periopera- distinguishing them diffi cult and symptoms alone tive situation is 27 times more likely to prove skin

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 239 test positive compared to those who do not not only revolutionized the practice of anesthesia develop hypotension, and bronchospasm in but also started the modern era of surgery and anaphylactic patients is more likely to be severe made possible the explosive development of than in those patients where the bronchospasm cardiothoracic, neurological and organ transplant was provoked by a nonimmune mechanism. The surgery.” role of direct histamine release in the etiology of bronchospasm during anesthesia is unclear since although the autacoid exerts a powerful effect on 7.4.1 Some Epidemiological the bronchi when inhaled in nebulized form, its Background effects are far less obvious when it enters the general circulation. It therefore seems likely that Most case series, surveys, and diagnostic and the histamine- releasing capacities of anesthetic mechanistic studies on NMBDs, the most fre- drugs as a factor in the induction of broncho- quent instigators of drug-induced anaphylaxis spasm are not as important as sometimes assumed during anesthesia, have been undertaken in and expressed. For an expanded discussion of Australia, France, New Zealand, the United direct histamine release by drugs, see Chap. 8 . Kingdom, and, more recently, Scandinavia and Spain. Few investigations have been pursued in the USA where reactions are presumably rare or 7.4 A naphylaxis to Neuromuscular go unrecognized or unreported. Incidences of Blocking Drugs reactions vary between countries and estimates have ranged from 1 in 1,000–2,000 to about 1 in In 1942 Griffi th and Johnson in Montreal intro- 20,000. In the United Kingdom about 500 reac- duced their landmark paper (Anesthesiology tions a year are thought to occur while the inci- 1942; 3: 418) on the fi rst use of drug-induced dences in France, Norway, and Australia are said muscle relaxation in anesthesia with the state- to be ~1 in 5,500, 1 in 5,200, and 1 in 10,000, ment—“Every anesthetist has wished at times respectively. With regard to prior sensitization of that he might be able to produce rapid and com- reactors, estimates of previous exposure to an plete muscular relaxation in resistant patients NMBD range from about 15 to 50 % and atopy under general anesthesia.” Using what was does not seem to be a significant risk factor. described as a “purifi ed curare” preparation or A history of previous anesthesia is also not a risk “Intocostrin” (Extract of Unauthenticated Curare, factor, but a history of an adverse reaction during Squibb), Griffi th and Johnson administered the a previous anesthesia is. Many studies over the preparation intravenously at a dose of 10–20 mg last 30 years have concluded that female reactors per 20 lbs body weight to 25 patients and obtained predominate with ratios compared to males of up temporary but complete muscular relaxation with to four to one. The median annual incidences of no apparent harmful effects. They concluded: allergic reactions to NMBDs estimated in a “…curare may prove to be a drug which will 1997–2004 French survey were 105.5 and 250.9 occasionally be of great value, and will give us a per million procedures for men and women, means of providing the surgeon rapidly with respectively. The overall annual incidence per excellent muscular relaxation at critical times million for both sexes was 184 and for children, during certain operations.” Within a year of the males and females showed an equal incidence of initial report, the “curare” preparation, essen- 61 cases. The number of children sensitized to tially an early purifi cation of d -tubocurarine, had NMBDs was found to increase with adolescence. been used in 131 patients during general anesthe- The distribution of anaphylactic reactions accord- sia and in 1952 Foldes et al. (N Engl J Med 1952; ing to age for adults showed peaks for males in 247: 596) summed up a decade’s use of muscle the 10–20 and 40–50 years age ranges, and for relaxants in anesthesia in the following females, the highest incidences occurred between words—“…[the] fi rst use of muscle relaxants in 30 and 60 years with the peak in the 40–50 anesthesiology by Griffi th and Johnson in 1942 year range.

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Fig. 7.1 Demonstration of allergenic cross-reactivity nium ( fi lled square ); (5) gallamine (open triangle); (6) pan- between NMBD. Inhibition by six different NMBDs of the curonium (fi lled triangle). From Baldo BA, Fisher MM. binding of IgE antibodies in the serum from an alcuronium Anaphylaxis to muscle relaxant drugs: Cross-reactivity and allergic patient to an alcuronium-solid phase conjugate. molecular basis of binding of IgE antibodies detected by Key: (1) alcuronium (open circle ); (2) d -tubocurarine (fi lled radioimmunoassay. Mol Immunol.1983;20:1393. Reprinted circle ); (3) succinylcholine (open square ); (4) decametho- with kind permission from Elsevier Limited

7.4.2 Mechanisms Underlying Table 7.3 Serological cross-reactivity between neuro- Anaphylaxis to muscular blocking drugs Neuromuscular Blocking Neuromuscular Amount (nmol/tube) for 60 % Drugs blocking drug inhibition of binding a Alcuronium 1.5 For information on histamine and its metabolism, Succinylcholine 7.0 Decamethonium 9.0 see Sect. 3.2.5.1 . Gallamine 16.5 d-Tubocurarine 17.0 7.4.2.1 Immunoglobulin E Pancuronium 14.0 Recognition of Allergenic Serum from a patient who experienced anaphylaxis to Determinants alcuronium The ﬁ rst reports of anaphylactic-like reactions Data from Baldo BA, Fisher MM. Mol Immunol 1983;20:1393 to NMBDs appear to have been in the late 1960s, a Inhibition of binding to an alcuronium-solid phase and by the early 1980s skin testing with the free drugs was being routinely used to diagnose reactions, identify sensitivity to the drugs, and muscle relaxant properties examined in inhibi- look for cross-reactivity between the different tion experiments showed that the cross-reactive NMBDs. Utilizing sera taken from patients who antibody- binding structures on the NMBDs experienced an anaphylactic reaction to an were quaternary and tertiary ammonium ions, NMBD, Baldo and Fisher in 1983 developed a the former groups being the structure involved radioimmunoassay to demonstrate the presence in conferring neuromuscular blocking activity of serum IgE antibodies to a solid phase cova- on the muscle relaxant drugs. Apart from the lent complex of alcuronium. Quantitative bind- presence of ammonium ions, some of the crossing and hapten inhibition studies demonstrated reacting drugs showed little or no structural that antibodies to one NMBD generally also similarity. Drugs with diverse pharmacological recognized and reacted with other NMBDs activities were inhibitory including an antihista- (Fig. 7.1 , Table 7.3 ). Also, results with panels of mine, a neuroleptic, a ganglionic blocking carefully selected drugs and chemicals with no agent, an opioid analgesic, an acetylcholine

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Fig. 7.2 Cross-reactivity of NMBD-reactive IgE antibod- ( open square ); pentolineum tartrate (fi lled square ); tri- ies with some widely used drugs of different pharmaco- methaphan camphorsulfonate (open triangle ); morphine logical action but which all contain quaternary and/or HCl ( fi lled triangle). From Baldo BA, Fisher MM. tertiary ammonium groups. Inhibition of alcuronium-reac- Anaphylaxis to muscle relaxant drugs: Cross-reactivity tive IgE antibody binding to an alcuronium-solid phase by and molecular basis of binding of IgE antibodies detected alcuronium (plus sign ); promethazine HCl (open circle ); by radioimmunoassay. Mol Immunol.1983;20:1393. chlorpromazine HCl (fi lled circle ); neostigmine bromide Reprinted with kind permission from Elsevier Limited

Table 7.4 Recognition of some frequently used drugs of trimethylammonium salts in inhibition studies diverse pharmacological activities by neuromuscular were generally clearly recognized by the blocking drugs-reactive IgE antibodies NMBD-reactive IgE antibodies showing differ- Amount (nmol/tube) ent inhibitory potencies with different sera. For of drug needed for 60 % example, in the results shown (Fig. 7.3 , Drug inhibition of binding a Table 7.5), the tetraalkylammonium salt octyl- Alcuronium 2.0 Promethazine HCl 90 trimethylammonium bromide was almost Chlorpromazine HCl 90 equally potent in inhibiting the IgE in the Neostigmine bromide <17 patient’s serum as the NMBD implicated in the Pentolineum tartrate 15 patient’s anaphylactic reaction and, on a molar Trimethaphan 30 basis, was over three times as potent as the ethyl camphorsulphonate and dodecyl derivatives and nearly ﬁ ve times as Morphine HCl 19 potent as the hexadecyl derivative. These results IgE antibodies in serum from a patient who experienced suggested that as the alkyl chain increased in anaphylaxis to alcuronium. Serum is the same as used in length, the optimum “ﬁ t” for the complemen- inhibitions with neuromuscular blocking drugs—see tary antibody combining sites was reached with results Table 7.3 Data from Baldo BA, Fisher MM. Mol Immunol a chain length of around eight carbons (bearing 1983;20:1393 in mind that chain lengths of 3–7 and 11 carbons a Inhibition of binding to an alcuronium-solid phase were not tested), and with the addition of more carbons, inhibitory activity declined. Also, as receptor antagonist, and an acetylcholinesterase one might expect, the nature of the alkyl group inhibitor (Fig. 7.2 , Table 7.4 ). Quaternary within the ammonium ion is important for anti- ammonium compounds including some tetraa- body recognition. Of the three tetraalkylammo- lkylammonium bromides and a series of alkyl- nium salts tested, the tetramethylammonium

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Fig. 7.3 Cross-reactivity of NMBD-reactive IgE antibod- bromide (open square ); ethyltrimethylammonium ies with quaternary ammonium compounds. Inhibition by bromide (open triangle); octyltrimethylammonium bro- tetraalkylammonium and alkyltrimethylammonium salts mide (fi lled triangle ); dodecyltrimethylammonium bro- of the binding of alcuronium-reactive IgE antibodies to an mide ( fi lled diamond); hexadecyltrimethylammonium alcuronium-solid phase conjugate. The serum was from a bromide ( open diamond). From Baldo BA, Fisher MM. patient who experienced anaphylaxis to alcuronium and Anaphylaxis to muscle relaxant drugs: Cross-reactivity is the same serum used in experiments summarized in and molecular basis of binding of IgE antibodies Figs 7.1 and 7.2 . Key: alcuronium (plus sign ); tetrameth- detected by radioimmunoassay. Mol Immunol.1983; ylammonium bromide (open circle ); tetrapropylammo- 20:1393. Reprinted with kind permission from Elsevier nium bromide (fi lled square); tetrapentylammonium Limited

Table 7.5 Recognition of quaternary alkyl ammonium salts by neuromuscular blocking drug-reactive IgE antibodies Amount (nmol/tube) needed Quaternary ammonium compound for 60 % inhibition of binding a Alcuronium (neuromuscular blocking drug) <2.0 Ethyltrimethylammonium bromide 8.1 Octyltrimethylammonium bromide 2.5 Dodecyltrimethylammonium bromide 8.6 Hexadecyltrimethylammonium bromide 12.2 Tetramethylammonium bromide 8.6 Tetrapropylammonium bromide 230 Tetrapentylammonium bromide 500 IgE antibodies in serum from a patient who experienced anaphylaxis to alcuronium. Serum is the same as used in inhibitions with neuromuscular blocking drugs—see results Table 7.3 Data from Baldo BA, Fisher MM. Mol Immunol 1983;20:1393 aInhibition of binding to an alcuronium-solid phase. Results for alcuronium shown in Tables 7.3 , 7.4 and 7.5 were obtained in three separate studies salt was ~27 and 58 times as active as the tetra- 7.4.2.2 Fine Structural Speciﬁ cities propyl and tetrapentyl derivatives, respectively of IgE Antibodies that React (Fig. 7.3 , Table 7.5 ). Table 7.6 summarizes with Neuromuscular examples of compounds without neuromuscular Blocking Drugs blocking properties that are recognized and Extensive IgE antibody combining site speciﬁ c- interact with NMBD-reactive IgE antibodies in ity studies employing sera from patients with sera from NMBD-allergic patients. NMBD-induced anaphylaxis together with all

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Table 7.6 Examples of some drugs and chemicals recognized by IgE antibodies that react with neuromuscular blocking drugs Structure of compound or cation group Compound with ammonium group highlighteda b Trialkylamines NR 3 + c d + e Tetraalkylammonium salts N R 4 , RN (R') 3 + Choline (CH 3 ) 3 N CH2 CH2 OH + Acetylcholine (CH 3 ) 3 N CH2 CH2 OCOCH3 Promethazine

SNCH2 CH N(CH3)2

CH3

Neostigmine + N (CH3)3

OOCN(CH3)2 Morphine HO

O N CH3 HO

Pentolineum + + N (CH2)5 N CH3 H3C Procaine H2NCOO(CH2)2N(C2H5)2

From Baldo BA, Pham NH. Structure–activity studies on drug-induced anaphylactic reactions. Chem Res Toxicol 1994; 7: 703. Reproduced with permission from American Chemical Society aDeterminants show heterogeneity. They may be solely the ammonium group or extend to attached or nearby atoms or groupings. The exact confi nes of IgE-binding determinants are not always clear and depend on the particular serum (IgE) studied b R = methyl or ethyl c R = methyl, ethyl, propyl, etc. d R = methyl, ethyl, …, hexadecyl, etc. e R ′ = methyl, ethyl, propyl, etc. available NMBDs and selected analogs led to the Category 2. Antibodies with recognition profi les conclusion that the combining site specifi cities of confi ned to the ammonium groups but which NMBD-reactive IgE antibodies fall into fi ve main cross-react with, and are inhibited almost equally groups (Fig. 7.4 ): well by, each of the NMBDs with the same or similar groups linked to the nitrogens. Examples Category 1. Antibodies that react with the include succinylcholine and decamethonium; ammonium group(s) of only one NMBD and are d -tubocurarine and atracurium; and pancuronium inhibited by that NMBD only. It is assumed that and vecuronium. However, since differences the specifi city of such antibodies is precisely occur in the structures attached to the nitrogens in complementary to some fi ne structural detail(s) some NMBDs, antibodies to one NMBD, for present uniquely on the group(s) of the NMBD example, succinylcholine, may not readily cross- (see for example discussion on alcuronium and react with, and be inhibited by, some other rocuronium, Sect. 7.4.4.3 ). NMBDs, for example, rocuronium.

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H&L chain antigen-binding fragment of IgE

Determinants on different NMBDs

Category 1 Category 2

Category 3 Category 4a

Category 5 Category 4b

Fig. 7.4 Diagrammatic representation of different possi- antibodies. Some antibodies may be promiscuous in their ble fi ne structural recognition patterns of IgE antibodies recognition profi le, reacting with all or most NMBDs via that react with neuromuscular blocking drugs. Category 1. a small combining site complementary to a small determi- Antibodies that react with the ammonium group(s) of only nant structure present on a number of NMBDs (4a) or via one NMBD. Category 2. Antibodies with recognition pro- a large combining site able to accomodate similar (but not fi les confi ned to the ammonium groups but which cross- necessarily identical) structures on different NMBDs (4b). react with each of the NMBDs with the same or similar Category 5. Antibodies that recognize a structure on an groups linked to the nitrogens. Category 3. Antibodies that NMBD other than the ammonium ions that is not found on recognize ammonium groups together with adjoining and/ other NMBDs or adjacent structures. Category 4. Highly cross-reactive

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 245

Category 3. Antibodies that recognize ammonium capacity. In anaphylactic responses such as those groups together with adjoining and/or adjacent induced by NMBDs, mediators of the allergic structures. Here inhibitory activities of different reaction are released by the cross-linking of NMBDs may vary widely depending on the FcεRI-bound IgE molecules on mast cells and ammonium group and the uniqueness or other- basophils. This occurs by allergen molecules wise of the neighboring structure(s) recognized. reacting with the combining sites of the divalent antibody molecules, but to do this an allergenic Category 4 . Highly cross-reactive antibodies. molecule must be at least divalent. Even without Some antibodies may be promiscuous in their being protein bound, allergenic divalency (or recognition profi le, reacting with all or most greater) is a requirement that all NMBDs fulfi ll NMBDs. This may occur in two ways: (a) via by virtue of the substituted ammonium ions pres- recognition of a small fi ne structural feature com- ent in the molecules. The optimum molecular mon to all NMBDs or (b) via antibodies with length of NMBDs, depolarizing or not, is 2.0– larger combining sites lacking recognition of 2.1 nm with the ten-carbon 2.0 nm length of common fi ne structural detail but showing broad, decamethonium showing potent activity. The less precise recognition of larger similar general ammonium ions in NMBDs are, of course, structural features found an all or most NMBDs. responsible for the neuromuscular blocking property of the drugs, and at a distance apart of about Category 5 . Antibodies that recognize a structure 1–1.6 nm, bridging of adjacent antibody mole- on an NMBD other than the ammonium ions that cules can be effected. Experimental verifi cation is not found on other NMBDs. of this predicted free drug-induced release from So far, we have found only one serum contain- human leukocytes has been provided. In in vitro ing IgE antibodies that reacted with, and were histamine release studies comparing the NMBDs inhibited by, only one NMBD, but because the succinylcholine and pancuronium with a series of precise structure recognized remained unidenti- diammonium salts of increasing chain length, fi ed, it was not possible to say whether the anti- lengths greater than 4 Å (0.4 nm) were required bodies belonged to category 1 or category 5. for release to occur and a length of 0.6 nm pro- Figure 7.4 sets out simplifi ed diagrammatic duced optimum release. In addition, and as might representations of these fi ve different antibody be expected, NMBDs with rigid structures such recognition profi les. It does not attempt to depict as pancuronium were less active in promoting the fi ner structural differences referred to. Given the release of histamine than fl exible straight chain known heterogeneity of antibody combining molecules like succinylcholine with its widely sites, some sera may contain mixtures of speci- spaced terminal determinant groups. For a further fi cities from one or more of the fi ve different discussion of allergen bridging of cell-bound IgE categories. molecules for mediator release, see Sect. 3.1.2 .

7.4.2.3 Neuromuscular Blocking 7.4.2.4 Inhibition of Histamine-N - Drug-Induced Release of Methyltransferase by NMBDs Mediators of Anaphylaxis All NMBDs inhibit histamine-N- methyl- Simply binding speciﬁ cally to IgE antibody transferase (HMT), the primary catabolic enzyme combining sites is not enough by itself to describe for histamine in humans (see Fig. 3.6 ). Inhibition a molecule as an “allergen.” For an immediate, is competitive with respect to the methyl donor type I reaction, ‘allergenic activity’ is the prop- and noncompetitive with respect to histamine. erty of speciﬁ cally provoking an allergic response Six different NMBDs, alcuronium, pancuronium, via the release of the biologically active mole- d-tubocurarine, gallamine, succinylcholine, and cules that cause the signs and symptoms of a decamethonium, inhibited enzyme activity in the hypersensitivity reaction and a true allergen has concentration range 10−7 –10 −3 M with alcuronium −6 this property as well as IgE antibody-binding being the most potent inhibitor (ID50 = 2 × 10 M)

[email protected] 246 7 Drugs and Other Agents Used in Anesthesia and Surgery and with the activity of the other NMBDs follow- that ~5 % of blood donors and 10 % of allergic ing in the above-listed order. Alcuronium proved subjects had serum IgE antibodies that reacted to be of similar potency to the dimaprit analog with a morphine solid phase (for the signifi cance SKF91488, one of the most potent HMT inhibi- of this, see Sects. 7.4.3.4.2 and 7.4.5.2 ). These tors known. In an investigation of structure– fi ndings need to be kept in mind when skin tests activity relationships of potent inhibitors of and serum IgE antibody tests are being used to HMT, alcuronium was compared to the antima- investigate suspected NMBD hypersensitivities. larials amodiaquine, quinacrine, and chloroquine and to the antiseptic agent chlorhexidine. Each of 7.4.3.1 Persistence of IgE Antibodies the compounds showed marked similarities to the to Neuromuscular Blocking conformations of the arrangement of the three Drugs nitrogen atoms of histamine. Following the In six patients investigated by skin testing and release of vecuronium, it was shown to be a better detection of serum IgE antibodies from 4 to nearly inhibitor (K i = 1) than pancuronium (K i = 2). 30 years after NMBD-induced anaphylaxis during Calculations showed that administration of anesthesia, the positive skin test to an NMBD(s) vecuronium in doses of 0.1–0.15 mg/kg could obtained soon after the reaction was found to per- inhibit HMT for up to 14 min by 25–50 %. This sist while NMBD-reactive antibodies were may explain the occasional severe bronchospasm detected in the fi ve patients tested. In a study of seen in some patients given vecuronium. changes over a 4–13 year period of intradermal test results, the positive tests for an NMBD(s) incrimi- nated at the time of the reaction remained positive 7.4.3 Diagnosis of Anaphylaxis in 15 of 18 patients. An investigation of changes in to Neuromuscular Blocking NMBD-reactive IgE antibodies in seven NMBD- Drugs allergic patients showed six had persisting positive tests with the antibody levels increasing in one A challenge (provocation) test with an NMBD for patient and decreasing in fi ve. Results with one the diagnosis of allergic sensitization to NMBDs patient were interesting and somewhat unexpected. is of course not acceptable, so this means that After initially reacting to decamethonium and test- other diagnostic methods must be relied upon and, ing skin test positive to this drug, negative to suc- for preference, the employment of more than one cinylcholine, and positive to succinylcholine in the method is desirable. Because most, if not all, IgE immunoassay, all of the tests proved positive 5 patients who experience an anaphylactic reaction years later, but after a further 2 years the intrader- to an NMBD are not allergically sensitized by mal test for succinylcholine was negative while the prior exposure to one of these drugs, one can serum IgE antibody test for the drug remained assume that the NMBD-reactive IgE antibodies positive. In many direct binding IgE antibody and were already present in the patients’ sera prior to inhibition studies employing the full range of their anesthesia. This means that sensitization available NMBDs, reactivities of decamethonium probably occurs through exposure to a stimulating and succinylcholine invariably seemed to go allergen source or perhaps via a “natural” antibody together. This suggested that the patient’s sensitiv- (see below, Sect. 7.4.5.3 ) and one might therefore ity to decamethonium/succinylcholine detected by predict that the presence of such antibodies in the the antibody immunoassay was persisting, and general population might not necessarily be rare. although skin testing did not pick it up, the wisest In fact, this appears to be the case. A study to advice was to avoid the responsible drugs for the determine the prevalence of NMBD reactivity in a rest of the patient’s life. Skin tests, leukocyte hista- sample of the general population in France showed mine release experiments, and IgE antibody immu- that 9.3 % of 258 subjects had either a positive noassays were utilized to investigate persistence of skin test to one or more NMBDs or the presence of allergy to NMBDs in 21 patients who reacted to quaternary ammonium ion-reactive serum IgE. succinylcholine. Skin tests showed sensitization to Another relevant study in Scandinavia revealed the drug persisted in 18 patients 1–4 years later,

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 247 and histamine release and IgE determinations also a diagnosis and identify the underlying mecha- remained positive in most of the patients. Seasonal nism of a suspected drug-induced reaction during variations to allergens such as pollens and other anesthesia. investigations involving a variety of different allergens have shown that IgE antibody production 7.4.3.3 Skin Tests increases with antigen exposure, but some obser- For a detailed discussion of the employment of vations such as the transfer of allergy in humans by skin testing in the diagnosis of drug allergy, the bone marrow transplants show that cells retain the reader is referred to Sect. 4.2 . capacity to secrete specifi c IgE antibodies in the There is general agreement on the value of absence of continuing exposure. This is supported skin testing with NMBDs for the diagnosis of by the demonstration of immunoglobulin E secret- allergic sensitivity to the drugs but, as mentioned, ing cells in both the marrow and lymph nodes the so-called gold standard of challenge with the draining the site of antigenic challenge in labora- drug delivered in incremental doses cannot be tory animals. done so it is not possible to validate the test. Despite this, the positive and negative predictive 7.4.3.2 Serum Tryptase value of the skin test appears to be good even Determination without validation. In one recent diagnostic inves- The tryptase test is discussed in detail in Sect. tigation of rocuronium allergy the authors con- 4.5.1 where details of the assays for total and sidered diagnosis was established by a positive mature tryptase are given. skin test for the drug without further tests. Skin Upon the induction of what appears to be an testing then is the preeminent diagnostic test for anaphylactic-like response during anesthesia and NMBD- allergic sensitivity, but therein lays the once the reaction has been brought under control nagging doubt that some critics have for the test. and the patient stabilized, the primary question False- positive results with prick tests, and more must be was the reaction immune mediated, that likely with intradermal tests, occur and some is, anaphylactic, or nonimmune, that is, anaphy- believe that in the diagnosis of anaphylaxis to lactoid in nature? Although elevation of the con- NMBDs false positives occur more often than centration of mature and total (>11.4 μg/l) they should and sometimes this is not recognized. tryptase in serum is not always a certain indica- In particular, doubts have been expressed about tion of a true anaphylactic reaction since its pres- the concentrations of rocuronium and vecuronium ence, like histamine, can sometimes be released recommended for prick testing and intradermal from mast cells via nonimmunologic mecha- testing. As well as false positives, negative skin nisms, an increased tryptase level suggests that tests have occasionally led to an anaphylactic the reaction is more likely to be IgE antibody reaction to a second NMBD but the end result mediated and the greater the increase the higher is that whenever a false positive skin test or a the probability that the reaction is anaphylactic failure to detect an allergic sensitivity occurs, the rather than anaphylactoid. It should also be culprit drug and the underlying mechanism pointed out that the release of tryptase by nonim- remain unidentifi ed. mune mechanisms is not unequivocally established since serum tryptase levels are not, for 7.4.3.3.1 Methodology example, increased during vancomycin-induced Skin testing of patients who experienced a sus- anaphylactoid reactions even though the drug pected immediate allergic reaction to an NMBD induces tryptase release from mast cells in vitro should be undertaken no earlier than 4–6 weeks (see also, Sect. 6.1.4.2). In summary, the in vivo after the reaction. Since most of the IgE antibod- serum tryptase test result, whether it is positive ies may have been consumed during the allergic or negative, provides invaluable information to event, testing within the 4–6 week period be considered along with other important test increases the risk of a false-positive fi nding, so results, in particular skin test and drug-specifi c only positive results should be taken notice of IgE antibody determinations, in efforts to establish and a negative test should be repeated after the

[email protected] 248 7 Drugs and Other Agents Used in Anesthesia and Surgery recommended delay. For prick testing, the com- Table 7.7 Concentrationsa of neuromuscular blocking mercially available preparations of the NMBDs drugs used for skin testing are used diluted if necessary with sterile physio- Skin prick testb Intradermal testc logical saline. If a bacteriostatic is required phenol concentration concentration Drug (mg/ml) (μg/ml) 0.5 % w/v may be added. A positive histamine Succinylcholine 10 100 control (10 mg/ml in saline) and a negative Rocuronium d 10 50 (vehicle) control are always included. Information Vecuroniumd 4 400 on the shelf life of diluted NMBD preparations is Pancuronium 2 200 lacking, an indication that solutions for skin Atracurium 1 10 testing should be freshly prepared at the time Cisatracurium 2 20 of testing and rejected upon completion but, Mivacurium 0.2 2 except for atracurium, cisatracurium, mivacurium Data from Mertes PM et al. J Invest Allergol Clin Immunol and rocuronium, each of which should be freshly 2011;21:442 diluted, storage of test solutions for up to 3 a Concentrations normally nonreactive in subjects not months at 4 °C has been suggested by some. The allergic to a neuromuscular blocking drug b A positive test is a wheal after 20 min with a diameter NMBD solutions are used diagnostically on the 3 mm greater than the negative control or a diameter at forearm or back in skin prick and intradermal least half the diameter of the positive control testing; the former is a little more specifi c but less c 0.02–0.05 ml injected to give a 4 mm diameter bleb. sensitive, that is, it has a small tendency to give A positive test is the appearance of an erythematous wheal (often pruritic) after 20 min with a diameter at least twice false negative results while the latter test is more that of the initial bleb sensitive but less specifi c, that is, it produces Positive control for prick test: Histamine 10 mg/ml or more false positives. Up to 97 % concordance and codeine phosphate 9 % w/v. Negative control for prick a similar diagnostic value have been found in and intradermal tests: Same volume of solvent used for drugs studies comparing the two methods. A commonly d A high proportion of positive reactions in normal con- used prick test procedure employs side by side the trols has led to suggestions that these prick test concen- commercial NMBD injectable solutions neat and trations are too high (see text) diluted one in ten, the latter to reduce false- positive reactions. Atracurium, mivacurium, and is used to initiate intradermal testing and thereafter rocuronium are more likely to provoke histamine- ten times stronger concentrations (up to the induced nonspecifi c wheals leading some to maximum concentration) of solution are used at recommend starting their testing at a one in ten 20 min intervals until a positive reaction is seen. dilution. A wheal diameter of at least 2 mm is The maximum concentration should not be generally taken as a positive test, but because exceeded. Some other practitioners carry out NMBDs often produce smaller wheals than com- intradermal testing at the highest concentration of mon inhalant and other allergens, a positive wheal NMBD that does not cause a reaction in normal to neat test solution but a negative to the one in subjects. For intradermal testing 0.02–0.05 ml of ten dilution may be considered diagnostic. In a solution is injected into the dermis to give a bleb slightly different approach, the skin prick test is of ~4 mm diameter. The SFAR-SFA criterion for used to guide the choice of the fi rst concentration a positive reaction is the appearance after 20 min selected for intradermal injection. In this protocol of an erythematous wheal with a diameter at least issued by the French Society for Anaesthesia and twice that of the original bleb. Table 7.7 lists the Intensive Care (SFAR) and French Society of concentrations of NMBDs for skin prick and Allergology (SFA), a positive prick test is defi ned intradermal testing together with criteria for a as a wheal with a diameter 3 mm greater than the positive test in guidelines issued by the SFAR negative control or a diameter at least half that of and SFA. The reliability of prick testing using the the positive control 20 min after completion of recommended concentrations for rocuronium the test. When the prick test is negative, a 1 in (10 mg/ml undiluted) and vecuronium (4 mg/ml 1,000 dilution of the commercial NMBD solution undiluted) has been questioned following the

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 249 demonstration of positive skin prick test heavily slanted to NMBD reactors. The results responses to these concentrations in approxi- therefore, should not be assumed to be immedi- mately half of 30 healthy, non-atopic, anesthesia- ately applicable to other agents used in anesthesia naïve male and female volunteers. This result or, indeed, to other drugs in general. appears to add weight to the earlier report of positive cutaneous reactions without mast cell 7.4.3.3.3 Cross-Sensitivity Between degranulation in almost all of 30 normal volun- Neuromuscular Blocking Drugs teers following intradermal injection of a 1:100 Skin testing with NMBDs is undertaken not only stock solution (100 μg/ml) of rocuronium. This to establish whether or not a reaction during anes- report led to a change in the recommended intra- thesia was provoked by the drug administered and, dermal test concentration for rocuronium from at the same time whether the reaction was immune 100 to 50 μg/ml. mediated or due to direct mediator release, but also to investigate cross-sensitivity between differ- 7.4.3.3.2 Comparison of Prick and ent NMBDs with the view of identifying not only Intradermal Skin Testing in the the causative drug, but also any other NMBDs that Diagnosis of Anesthetic Allergy do not cross-react in the skin and which may In a prospective, non-randomized study involv- therefore be possible alternative drugs for safe ing 212 consecutive patients over a 4-year period, future use. Skin testing for this purpose must prick testing with undiluted drugs was compared therefore include all other commercially available to intradermal testing with diluted drug solutions NMBDs. Cross-reactivities of NMBDs detected in of standard, accepted concentrations. A positive skin tests, unlike cross-reactions demonstrated by prick test was taken as a wheal of diameter serum IgE antibodies, are regarded as the most >4 mm while a positive intradermal test was a relevant indication of a patient’s in vivo recogni- persistent wheal greater than 8 mm. Results of tion and likely response to individual NMBDs. the comparison are summarized in Table 7.8 . Despite the sometimes apparently clinically non- Overall, there was 93 % agreement between the applicable nature of information on NMBD cross- paired tests and differences between the tests reactivities derived from antibody investigations, were not signifi cantly different. Severe reactions better fi ne-structural defi nitions of individual accounted for 135 of the patients and there were NMBD allergenic determinants combined with 29 patients with minor reactions. Of the 135 skin test results are likely to provide a better basis severe reactors, the majority (93 patients, 69 %) for interpreting cross-reactivity data and thereby reacted to NMBDs. In this group of severe reac- increase the predictability of potentially unsafe tors, intradermal tests distinguished more posi- reactions to NMBDs and the identifi cation of tive reactors, but this fi nding would have been NMBDs safe for administration (see Sect. 7.4.4 reversed if a prick test wheal size of <4 mm had for an extended discussion). been selected as the positive cutoff. It should be As for skin testing in general, the concentra- pointed out that some practitioners of skin testing tions used to identify cross-reacting drugs consider that any wheal should be considered (Table 7.7 ) are important but with NMBDs the positive for prick testing. Employment of both added complication of possible cross-reactions tests produced more clear diagnostic outcomes caused by the histamine-releasing properties of than either test alone and it is likely that the safety drugs such as atracurium and mivacurium must of subsequent anesthesia would be improved by be taken into account. Some guidelines recom- performing both tests. When doubt exists over mend that the skin prick test can be used to detect the result of either test, both should be carried cross-reactions between NMBDs if the causative out. This study provides an interesting and valu- drug produces a positive prick test reaction but if able comparison of the diagnostic performances the drug is only positive in the intradermal test, of the two different skin test methods but it should then all NMBDs must be tested intradermally. be remembered that the patient numbers were Cross-reactions are more likely between NMBDs

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Table 7.8 Comparison of results of prick and intradermal tests in 212 consecutive patients in an anesthetic allergy clinic No. of Drug responsible detected by Agreement between tests patients No. of Group tested comparisons Prick Intradermal Both Either Neither Agree (%) Disagree (%) Minor reactions 29 117 8 5 4 9 20 113 (97 %) 4 (3 %) Severe reactions 135 965 108 112 104 116 19 890 (92 %) 75 (8 %) Preoperative sample 13 82 1 1 0 2 11 75 (90 %) 7 (9 %) Not anaphylactic 35 228 0 0 0 0 35 216 (95 %) 12 (5 %) Total 212 1,392 117 118 108 127 85 1,294 (93 %) 98 (7 %) Drugs tested: neuromuscular blocking drugs, induction agents, opioids, colloids, antibiotics, protamine, neostigmine From Fisher MM, Bowey CJ. Intradermal compared with prick testing in the diagnosis of anaesthetic allergy. Br J Aneasth. 1997;79:59. Reprinted with permission from Oxford University Press

of similar structure such as the benzylisoquino- alcuronium and d -tubocurarine, direct chemical liniums but again, a detailed understanding of the coupling of the drug to the insoluble carrier was IgE antibody-binding determinant structures at achieved but with NMBDs such as succinylcho- the fi ne structural level is needed to put the sub- line, decamethonium and gallamine, suitable ject of clinical cross-sensitivity between NMBDs functional groups for coupling are not present on a better scientifi c base. making the formation of drug-solid phases or drug–protein covalent complexes a diffi cult task 7.4.3.4 Detection of IgE Antibodies without fairly lengthy and complex chemical Reactive with Neuromuscular manipulations. This problem was overcome by Blocking Drugs utilizing structural analogs with the same termi- 7.4.3.4.1 Detection with Neuromuscular nal allergenic groups as the NMBDs and which Blocking Drugs and Analogs could be easily chemically coupled to a solid Following the development and application of phase carrier. Succinylcholine can be thought of the initial immunoassays to detect IgE antibod- as two molecules of choline linked by a carbon ies to alcuronium, immunoassays designed to chain derived from succinic acid and likewise detect IgE to other NMBDs were soon devel- decamethonium has a choline-like structure at oped and used alongside skin tests to diagnose each end of the molecule linked by a six-carbon allergic reactions to the range of NMBDs then chain (Fig. 7.5 ). The strategy pursued was therein use. Concurrently, assays for the detection of fore to covalently link choline via its available d -tubocurarine, succinylcholine and gallamine hydroxyl group to the solid phase by bis - oxirane were developed and used to examine, at the coupling to produce a complex mimicking the molecular level, the extent of cross-reactivity terminal groups of succinylcholine and decame- and quantitative relationships for the recogni- thonium. It was reasoned that such a complex tion of different drugs by IgE antibodies in indi- should be antigenically similar, if not identical, vidual sera. For each different drug assay the to succinylcholine- and decamethonium- solid methodology was essentially the same as that phase complexes. The same strategy was used to used for alcuronium. The drug, or an appropri- prepare a complex to detect IgE antibodies to ate selected analog, was covalently coupled to gallamine. Gallamine’s three attached quater- the solid phase via a spacer arm to form a chem- nary ammonium groups can be viewed as three ical complex without fi rst linking the drug/ana- molecules of triethylcholine attached as anten- log to a carrier protein. Patients’ sera were nae to an aromatic ring. The ethyl analog of added to small quantities of the drug complex choline was therefore synthesized and coupled and any specifically bound IgE antibodies directly to the solid support to produce a com- were detected by the addition of a radiolabeled plex suitable for the detection of gallamine- anti-human IgE antiserum. For NMBDs like reactive IgE antibodies. Figure 7.6 shows typical

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Choline Succinylcholine Decamethonium

O CH3 CH3 H3C H3C + H3C + CH2 OH CH2 O CH2 CH2 N CH3 CH2 CH2 CH2 CH2 CH2 N CH3 + CH + + H3C N 2 H3C N CH2 CH2 O CH2 H3C N CH2 CH2 CH2 CH2 CH2 CH3 CH3 H3C H3C O H3C

Fig. 7.5 Example of a strategy to overcome the difﬁ culty succinic acid and likewise decamethonium has a choline- of preparing solid phase complexes of drugs lacking a like structure at each end of the molecule linked by a six- suitable functional group for chemical coupling. An carbon chain. Choline was linked via its available example of utilizing structural analogs with the same hydroxyl group to the solid phase by bis -oxirane coupling terminal allergenic groups as the NMBDs to prepare drug to produce a complex mimicking the terminal groups of conjugates for use in assays to detect NMBD-reactive succinylcholine and decamethonium. Such a complex is IgE antibodies. Succinylcholine can be thought of as two antigenically similar, if not identical, to succinylcholine- molecules of choline (shown in blue in the 2D structures and decamethonium-solid phase complexes and CPK models) linked by a carbon chain derived from

Fig. 7.6 Employment of choline-Sepharose-solid phase ( fi lled square); gallamine (open triangle ); pancuronium conjugate to detect IgE antibodies to the NMBD succinyl- ( fi lled triangle ); triethylcholine (open diamond ); choline choline and other NMBD-reactive IgE in sera of patients ( fi lled diamond ). From Harle DG et al. Assays for, and allergic to NMBDs. Inhibition of IgE antibody binding to cross-reactivities of, IgE antibodies to the muscle relax- the solid phase by NMBD, choline, and triethylcholine. ants gallamine, decamethonium and succinylcholine Key: alcuronium ( open circle ); d-tubocurarine ( fi lled cir- (suxamethonium). J Immunol Methods 1985;78:293. cle); succinylcholine (open square ); decamethonium Reprinted with kind permission from Elsevier Limited

[email protected] 252 7 Drugs and Other Agents Used in Anesthesia and Surgery concentration-dependent inhibition profi les N -hydroxysuccininide ester and fi nal attachment obtained when the choline-solid phase is used to the solid phase after linkage to a carrier mole- with serum from a succinylcholine-allergic cule. Recent employment of the ImmunoCap® - patient, NMBDs and choline. Typically, cross- based test has produced encouraging results. An reactivity between NMBDs is apparent and ImmunoCap ® (Phadia) test for the detection of IgE although succinylcholine is a good inhibitor, antibodies to succinylcholine is also available. decamethonium, d -tubocurarine and gallamine The specifi city of assays for the detection of show superior inhibitory potencies and pan- NMBD-reactive IgE antibodies in patients’ sera curonium and alcuronium are relatively poor generally exceeds 90 % but the specifi city may inhibitors. These results indicate that detection vary from around 50 % reaching a maximum of of IgE antibody combining sites specifi c for about 90 %. In one study of assay performance of ammonium ion determinants with attached radioimmunoassays for the detection of IgE anti- “small” methyl or ethyl alkyl groups. These bodies to NMBDs a value of 100 % was obtained groups occur on decamethonium, d -tubocura- for the positive predictive value of a positive rine, gallamine, and succinylcholine, but the result. Skin tests remain the current reference test ammonium ion determinants on pancuronium, for NMBD-allergic sensitivity but their specifi c- and especially alcuronium, are markedly differ- ity is occasionally in question and their sensitiv- ent in structure and therefore poorly cross-reac- ity sometimes less than expected. In addition, tive with the former NMBDs. The clear, but skin tests are said to be unreliable in the fi rst 4–6 inferior to NMBD, inhibition demonstrated by weeks after the reaction to an NMBD and certain choline is probably related to its single quater- medications may preclude their application in nary ammonium group compared to two groups some patients. In these cases and in situations on decamethonium, succinylcholine, and where skin tests cannot be employed, for exam- d -tubocurarine and three on gallamine. The ple in attempts to assess a patient’s allergic status excellent inhibition shown by gallamine sup- before a completed operation or after a fatal reac- ports this interpretation. For some years, and tion, IgE antibody tests on serum from blood since the introduction of this choline-Sepharose samples taken at the time offer the only chance of support for detecting NMBD-reactive antibod- obtaining the relevant diagnostic data. ies, most tests to detect IgE to succinylcholine have been undertaken with what have been 7.4.3.4.2 The Use of Morphine for the described and reported, especially in the French Detection of IgE Antibodies to literature, as “new” solid phases consisting of Neuromuscular Blocking Drugs choline coupled to Sepharose via an ether link- In the original study where IgE antibodies to age or coupled p -aminophenylphosphorylcholine NMBDs were fi rst detected, morphine was found rather than the originally reported epoxy-cou- to potently inhibit IgE antibody binding to the pled solid phase. Apart from offering no con- NMBD-solid phase via antibody recognition of its ceptual advancement and, in our hands, no tertiary methylamino group (Fig. 7.2 ). Subsequent signifi cant advantages in performance, these investigations revealed that morphine reacts read- tests can hardly be described as “new.” ily with sera from patients allergic to NMBDs and Brief early attempts to prepare drug-solid in comparative inhibition experiments its potency phases for the detection of IgE antibodies to the often exceeds that of all NMBDs including the competitive, non-depolarizing, bulky and structur- drug that induced the anaphylactic reaction. This ally rigid aminosteroid NMBDs pancuronium, indicates that morphine might more closely com- vecuronium and rocuronium, met with only partial plement the combining sites of the NMBD- success but with the increasing popularity and use reactive IgE antibodies than the NMBDs of rocuronium in recent years, a Phadia rocuronium themselves. Along with morphine, d -tubocurarine ImmunoCap® has been prepared by reacting also reacts readily and strongly with many sera the drug with succinic anhydride to prepare the from NMBD-allergic patients and the two drugs hemisuccinate followed by activation to the are often the most strongly recognized compounds

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 253

a b H C O 3 OCH3 HO CH + 3 N OH

H O H H O NCH3 N

H CO OH H C HO 3 3

c d

Fig. 7.7 Two-dimensional and CPK space-fi lling models reactive IgE in sera from NMBD-allergic patients of morphine (a , c , e ) and d -tubocurarine (b , d , f ). The (see Harle et al. Mol Immunol 1990: 27:1039). From Baldo circled groups show the striking similarity of the shapes BA et al. On the origin and specifi city of antibodies to neu- and orientation of the tertiary N -methylamino groups on romuscular blocking (muscle relaxant) drugs: an immuno- each compound. This similarity is refl ected in the equal chemical perspective. Clin Exp Allergy 2009;39:325. reactivity of morphine and d -tubocurarine with NMBD- Reprinted with permission from John Wiley and Sons

of all the drugs and other chemicals so far exam- molecule. This very close structural similarity in ined. The almost equal capacities of morphine and the antibody- binding regions of the two drugs d-tubocurarine for interaction with NMBD- accounts for their almost equal performance in reactive IgE antibodies from patients following direct antibody- binding and inhibition tests, but, in anaphylaxis to an NMBD prompted further side- addition, both morphine and d -tubocurarine by-side comparisons of the drugs including the exhibit pronounced histamine-releasing properties construction of Corey, Pauling, Koltun (CPK) and the near- identical structural grouping of atoms models. The models (Fig. 7.7 ) revealed an almost identiﬁ ed on both drugs may be complementary exact likeness in the conformations of atoms of the to a mast cell receptor site involved in direct substituted ammonium groups on one face of each histamine release by the drugs.

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Table 7.9 Incidences of positive IgE antibody test for neuromuscular blocking drugs in the sera of a homogeneous group of patents a who experienced an anaphylactic reaction during anesthesia. Comparison of the morphine immunoassay with immunoassays for individual drugs Number detected positive for IgE antibodies with Neuromuscular blocking Patients skin test positive to Number positive Morphine testb drug (or analog) testc One or more neuromuscular 118 100 (85 %) 56/108 (52 %) blocking drugs Succinylcholine 69 67 (97 %) 47 (68 %) Rocuronium 15 15 3/13 Vecuronium 10 9 2 Pancuronium 2 0 0 Atracurium 12 3 2/6 Alcuronium 8 4 2 Gallamine 1 1 0 Mivacurium 1 1 0 a Patients (118) deﬁ ned by a positive skin test to one or more neuromuscular blocking drugs and an elevated mast cell tryptase test result b Data from Fisher MM, Baldo BA. Anaesth Intensive Care. 2000;28:167 c Tests employing individual neuromuscular blocking drug or analog solid phases

Because of morphine’s striking capacity to immunoassays used at that time to aid the diagno- detect and cross-react with NMBD-reactive anti- sis of allergic reactions to an NMBD and recently bodies in patients’ sera, it was used in solid phase this conclusion has been endorsed by the commer- form over a number of years side by side with cial release of the so-called QAM ImmunoCAP® other immunoassays to detect NMBD-reactive (Thermo Scientifi c), a form of immobilized mor- IgE antibodies. This culminated in a study of 347 phine designed for use in a fl uorescent enzyme patients with suspected anaphylaxis to an NMBD immunoassay. Assessment of this test on 168 where the morphine-solid phase proved a better patients in two hospitals revealed a sensitivity of test for the detection of the antibodies than the 84.2 %, almost exactly the same as the sensitivity other test materials prepared from individual fi gure obtained in the original study and the fi nd- NMBDs or analogs. In an homogeneous group of ing that positive reactions to IgE antibodies were 118 patients distinguished by an elevated tryptase signifi cantly higher in skin test-negative reactors level and a positive skin test to an NMBD, the (24.6 %) than in controls (9.3 %) suggested that morphine immunoassay detected NMBD-reactive some allergic reactors to NMBDs can be identi- IgE antibodies in 100 (84.7 %) of the subjects fi ed by tryptase and serum IgE antibody measure- with a specifi city of 98 % and a positive predictive ments in some skin test negative patients. It was value of 96 %. Effi ciency of the assay, that is, the concluded that the simplicity of the commercial percentage of all results that are true results, was morphine- solid phase assay and its suitability for calculated to be 94 %. The morphine test’s fi gures routine laboratory use made it a valuable addition for the detection of IgE to individual NMBDs to skin testing in diagnosing NMBD-allergic were 67 out of 69 (97 %) for succinylcholine, 9 sensitivity. out of 10 for vecuronium and 15 out of 15 for rocuronium but only 3 out of 12 for atracurium 7.4.3.5 Basophil Activation Test (Table 7.9 ). In the assay for succinylcholine- When used as a diagnostic aid for allergy to reactive antibodies (using a choline-solid phase), NMBDs, the basophil activation test (BAT) has 47 of the 69 sera (68 %) were positive. These been found to be specifi c but disappointingly results led to the conclusion that the morphine test lacking in sensitivity. Results for specifi city have was an improvement on the battery of individual generally been above 90 %, and although a fi gure

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 255 of 92 % was obtained for sensitivity in one study 7.4.3.6 Leukocyte Histamine Release on rocuronium-induced anaphylaxis, fi ndings Tests have been as low as 36 % and <60 % in fi ve sepa- Although not widely used, in vitro leukocyte rate studies. Various suggestions to explain the histamine release tests have been applied with often poor and variable sensitivity results have some encouraging results to the diagnosis of been advanced and include arbitrarily chosen allergy to NMBDs. Histamine release was dem- decision thresholds often based on experience onstrated in 8 of 25 (32 %) and 26 of 40 (65 %; with protein allergens rather than drugs, the fail- maximum release 43.8 ± 23.3 %, spontaneous ure to include well-characterized control sub- release 1.7 ± 1.1 %) patients with anaphylaxis to jects, the possible interference of other an NMBD. The specifi city was not determined in medications and the concentrations of NMBDs the former study but in the latter investigation no used. Another contributing factor might be the histamine release was demonstrated from the time elapsed between patient reactions and the leukocytes of 44 control subjects. The applica- performance of the test although NMBD-reactive tion of drug-induced histamine release in drug IgE antibodies are known to be long lasting in allergy diagnosis is discussed further in Sect. 4.5.2 . some patients (see Sect. 7.4.3.1 ). In one study the sensitivity increased from a low of 36.1–47.6 % 7.4.3.7 Best Current Combination for reactions occurring in ten patients 4–8 years of Tests for the Diagnosis before testing and up to 85.7 % for reactions in six of Immediate Hypersensitivity patients within the last 3 years. However, the to Neuromuscular Blocking question does not, as yet, seem to have been sys- Drugs tematically investigated and longitudinal studies For the diagnostic investigation of an adverse reac- to determine optimal time intervals are needed. tion to an NMBD, a reaction that all too often Application of the BAT to aid the management of manifests as a life-threatening response with anaphylaxis to rocuronium led to the conclusion anaphylactic- like symptoms, tests with the capac- that the method is a reliable diagnostic aid and ity to identify the culprit drug and identify any superior to serum IgE antibody inhibition assays cross-sensitivity with other NMBDs are required. in complementing skin tests for the identifi cation To confi rm the release of infl ammatory mediators of clinically relevant cross-reactions between from mast cells, the serum tryptase assay, which NMBDs. Although the BAT can be said to refl ect measures the concentration of the released enzyme, the in vivo pathways leading to allergen-induced is currently the best available relevant test. Note mediator release and the resultant allergic mani- that assays for total tryptase released from mast festations, there seems to be no compelling argu- cells and mature tryptase which is a direct measure ment at present for it to replace or add to skin of mast cell activation now exist and care should testing and tests for serum IgE antibodies in drug- be taken to distinguish the two when requesting induced allergy diagnosis. It should also be diagnostic tests, interpreting tryptase concentra- remembered that the BAT and the skin test are not tions in results provided and when reviewing the directly comparable. Even though both tests pro- literature (see Sect. 4.5.1.3 ). Although mast cell ceed via FcεRI receptors on the surfaces of baso- tryptase is said to be elevated in some anaphylac- phils and mast cells, the cells involved are different, toid or nonimmune-mediated reactions (eg. red they are at different stages of differentiation and man syndrome following vancomycin), evidence mast cells at different anatomical sites show sig- for this is confl icting and elevated levels of the nifi cant heterogeneity. There are, however, occa- enzyme in patients’ sera show strong correlation sions when an allergic reaction to an NMBD can with the involvement of NMBD-reactive IgE anti- be identifi ed by clearly positive tryptase and IgE bodies in reactions to these drugs during anesthe- antibody tests despite a negative skin test and in sia. In fact, guidelines issued by some professional this situation the BAT may be useful to confi rm the bodies regard the employment of the tryptase test diagnosis and identify a safe alternative NMBD. as “mandatory” in the diagnostic protocol for

[email protected] 256 7 Drugs and Other Agents Used in Anesthesia and Surgery drug-induced suspected allergic reactions. Skin tions of the drug with antibodies in the sera of testing with NMBDs is the reference test and cen- NMBD-allergic patients are due to antigenic cross- tral to diagnosis such that if only one test were to reactivity of the shared tertiary ammonium groups. be used, it would be selected. However, skin test- In summary, in addition to a meticulously ing with NMBDs is not problem- or criticism-free gathered and recorded history of an NMBD- with its sensitivity for some drugs sometimes lack- induced reaction and expert clinical assessment ing and its specifi city occasionally controversial. of the signs and symptoms of the reaction, and In addition to positive responses obtained in cases regardless of the particular NMBD implicated, involving an IgE-dependent mechanism, positive the “best combination” of investigative proce- skin tests may occur as a result of nonspecifi c his- dures currently available comprises: tamine release. False-positive results obtained 1. The tryptase test carried out on at least one, with the known histamine releaser atracurium but ideally two or more, serum samples drawn illustrate this point. Importantly, skin tests are also at optimum time intervals, namely 30 min up unreliable in the fi rst 4–6 weeks after a reaction to to 4–6 h after the onset of symptoms. an anesthetic agent and in patients receiving cer- 2. Skin tests with unconjugated NMBDs presented tain medications. Direct binding and inhibition either percutaneously or intradermally or both. tests for the detection of IgE antibodies that react The skin test is central to identifying the culprit with NMBDs are valuable adjuncts to skin tests drug as well as any cross-reacting NMBD. and the tryptase assay. The value of the IgE test is 3. The morphine-solid phase assay for detecting most obvious in cases where clinical data indicates IgE antibodies that react with any of the an allergic reaction but skin tests are negative or NMBDs. This assay may be supplemented equivocal and when, unlike skin testing, it can be with other immunoassays prepared with a spe- used in the 4–6 week period immediately follow- cifi c NMBD (e.g., rocuronium) or an analog ing a patient’s reaction. Other important applica- such as choline (for succinylcholine). In the tions are tests on preoperative serum samples taken right hands, judicious application of quantita- from patients who subsequently experienced a tive inhibition studies of antibody binding can reaction during anesthesia and sera taken just add valuable information by identifying the before and after death. Because of its apparent fi ne structural recognition features of the most capacity to react with IgE antibodies to all of the reactive NMBDs and the NMBDs most likely NMBDs, its sensitivity, simplicity and suitability to cross-react clinically. for routine laboratory use, the morphine immuno- Application of this combination of tests offers assay (Sect. 7.4.3.4.2 ) is the best single choice for the best chance of successfully investigating a the diagnostic detection of NMBD-reactive IgE hypersensitivity reaction to an NMBD during anes- antibodies in patients’ sera. If available, other thesia, confi rming or eliminating the occurrence of “specifi c” immunoassays for individual NMBDs a true IgE antibody-mediated reaction and thus may also be used. Note, however, if IgE antibody gaining insights into the underlying mechanism. immunoassays are to yield their full potential as a diagnostic aid for NMBD- induced hypersensitivity responses, much greater attention will need to 7.4.4 Cross-Reactions Between be paid to gaining immunochemical insights for Neuromuscular Blocking the interpretation of antibody recognition of indi- Drugs vidual NMBDs as well as the many factors relevant to the design and interpretation of direct 7.4.4.1 Reactivity of Patients’ Sera binding and inhibition studies (see Sect. 7.4.4 ). It with Different Neuromuscular should also be remembered that in most cases, the Blocking Drugs detection of morphine-reactive IgE antibodies is Both skin and IgE antibody tests can be used to not an indication of clinical hypersensitivity to the detect allergenic cross-reactivity between drug. Type I IgE antibody-mediated hypersensitiv- NMBDs but the frequency of cross-reactions is ity reactions to morphine occur rarely and the reac- generally higher when determined by interaction

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 257 with antibody than skin testing. The clinical rel- at each end of an extremely fl exible ten-carbon evance of serum antibody test results has been chain while succinylcholine with the same termi- questioned and this refl ects the different underly- nal structures is less fl exible due to the presence ing processes of the two tests with the in vitro of two ester groups in the chain. One might there- method detecting interaction of the drugs with fore expect that in free solution decamethonium the antibody combining sites and the cutaneous would be more accessible to antibody binding test detecting the cross-linking of cell-bound IgE and this would be refl ected in its superior inhibi- by NMBDs and the subsequent release of infl am- tory performance. The N -methyl groups of deca- matory mediators. Even so, it is true that much of methonium might also help explain its excellent the cross-reactivity data from the in vitro inhibi- inhibitory results. When investigated at the fi ne tion experiments remains poorly understood and structural level, the combining sites of many interpreted. As emphasized in this chapter, under- NMBD-reactive IgE antibodies show specifi city standing and interpreting immunological recog- for the N-methyl rather than N -alkyl groups nition and cross-reactivity of NMBDs depends with more carbons, and although the higher on defi ning the precise determinant structure on alkyl substituents are often recognized, the rec- an individual suspected NMBD as well as taking ognition is clearly weaker. The likely recogni- into account a number of other important factors tion preference for N -methyl determinants is such as the structure of the antibody-reactive also revealed in Table 7.10 by the unexpectedly solid phase used in the inhibition study and the better than expected inhibitory fi ndings with fl exibilities, conformations, and binding affi ni- d-tubocurarine (which has a quaternary ammo- ties of the all the NMBDs examined. If the struc- nium ion with two methyl groups and a tertiary tures on NMBDs recognized by their methylamino group) and by recent results with complementary antibody combining sites were atracurium. The quaternary allylpyrrolidinium confi ned to the tertiary and quaternary ammo- groups on alcuronium and rocuronium, the nium groups one would expect that, regardless of morpholino group of rocuronium and the the particular NMBD involved (and assuming N-piperidinium groups on vecuronium and pan- equal accessibility for the antibodies), all NMBDs curonium (Figs. 7.8 and 7.9 ) would also be less would show approximately equal reactivity. This well recognized by antibody combining sites is clearly not the case as even a quick perusal of complementary to determinants carrying the the extensive quantitative inhibition fi ndings set N-methyl group. out in Table 7.10 shows, but three main, and The second main conclusion drawn from the interrelated, conclusions can be drawn from the data set out in Table 7.10 is that there is, not 150 inhibition results obtained when six different unexpectedly, a relationship between the NMBD NMBDs were used with sera from 13 different that induced the anaphylactic reaction, the drug- patients. Firstly, regardless of the drug that solid phase used for testing and the most active caused the patient’s reaction and the drug on the inhibitors. The most secure results and results solid phase test material, the clear recognition most likely to be relevant to productive immuno- and superior inhibitory potency of decametho- chemical and clinical interpretation are those nium is apparent. The reasons for this may be obtained in inhibition studies where the NMBD related to the relative fl exibility of the decame- that produced the anaphylaxis matches the thonium molecule and the chemical composition NMBD attached to the solid phase. The problem, of its ammonium groups. The substituted ammo- however, in pursuing this analysis and coming to nium ions on the fl exible, straight chain depolar- any fi rm conclusions is a factor that cannot be izing NMBDs like decamethonium and ignored, viz., the sensitizing agent(s) for most, if succinylcholine (Fig. 7.8 ) are more accessible to not all, patients who reacted to an NMBD is not antibody binding than the ammonium ions on the known. This is the third, and obvious, conclusion bulky, rigid molecules such as alcuronium and drawn from the results shown here and which the aminosteroid NMBDs (Figs. 7.8 and 7.9 ). must be kept in mind when NMBD–IgE antibody Decamethonium has trimethylammonium groups interactions are being considered. The conundrum

[email protected] 258 7 Drugs and Other Agents Used in Anesthesia and Surgery Gallamine ar blocking drugs ar blocking drugs -Tubocurarine -Tubocurarine Pancuronium Succinylcholine Decamethonium d 39 17 16 29 130 195 11 240 105 100 75 19 180 220 10.1 12.5 42.1 110 52 67 14 46 61 85 320 90 470 525 1000 1000 Amount (nmol/tube) of NMBD to produce 50 % inhibition of uptake of anti-human IgE Amount (nmol/tube) of NMBD to produce 50 % inhibition uptake Alcuronium

c 1.1 1.8 0.4 1.3 0.9 1.2 d city of antibodies to neuromuscular blocking (muscle relaxant) drugs: an immunochemical perspective. Clin Exp Allergy Allergy Clin Exp city of antibodies to neuromuscular blocking (muscle relaxant) drugs: an immunochemical perspective. -Tubocurarine -Tubocurarine -Tubocurarine -Tubocurarine -Tubocurarine -Tubocurarine -Tubocurarine Triethylcholine Triethylcholine 15 Triethylcholine Triethylcholine 89 25 Triethylcholine 50 d Choline 35 0.6 Triethylcholine d 74 68 9.4 58 5.9 15 8 12 24 0.7 4.9 6.2 18 6.7 4.3 7.7 21 0.4 12 9.8 3 14 3.2 13 4.5 1.9 5.6 5.9 5.6 8.4 d Alcuronium d Choline Triethylcholine 39 d Alcuronium 28 Vecuronium 23 34 2 28 5 130 6 5 4 100 15 19 13 160 73 2.9 1.1 105 59 6 11 42 on solid phase Drug or analog -Tubocurarine -Tubocurarine -Tubocurarine -Tubocurarine Decamethonium d d d Alcuronium Drug reacted to Inhibition of IgE antibody binding to different neuromuscular blocking drug or analog solid phases by individual neuromuscul neuromuscular blocking drug or analog solid phases by individual Inhibition of IgE antibody binding to different

a :325). Reprinted with permission from John Wiley and Sons. Wiley :325). Reprinted with permission from John 39 Neuromuscular blocking drug (NMBD) or analog—choline for succinylcholine and decamethonium; triethylcholine for gallamine Neuromuscular blocking drug (NMBD) or analog—choline for succinylcholine produced 50 % inhibition at 0.2 nmol/tube Vecuronium Serum from patients who reacted with anaphylaxis after receiving the indicated drug Serum from patients who reacted with anaphylaxis after receiving coupled to Sepharose Drug or analog covalently 4 5 6 7 Alcuronium 8 Alcuronium Alcuronium 9 Choline Succinylcholine Choline 10 Succinylcholine Choline Choline Choline Succinylcholine 42 30 Succinylcholine Choline 15 41 Choline 86 12 9 3.3 4.3 10 37 1.3 9.7 9.4 2.9 10 1.8 14 17 4.1 9.8 2.9 6.1 4.7 7.3 4.7 11 4 1.6 6 2 1.3 0.5 0.8 8.7 8.7 0.9 7.8 6.9 1.6 9.6 6.8 11 12 Succinylcholine 13 et al. On the origin and speciﬁ From Baldo BA 2009; Gallamine Gallamine Triethylcholine Triethylcholine 74 750 12 40 14 54 8.1 55 7.3 32 4.2 37 1 2 3 Table Table 7.10 Patient’s a b c d

serum

[email protected] Depolarizing NMBDs–flexible molecules Decamethonium

Succinylcholine

Competitive NMBDs–bulky, rigid molecules

Alcuronium

d-Tubocurarine

Fig. 7.8 Comparison of two-dimensional structures and containing an sp2 carbon linked to an sp2 oxygen. Two rela- space-fi lling models of straight chain, fl exible (decametho- tively large allylammonium groups are not easily accessible nium and succinylcholine) and bulky, rigid (alcuronium and in the bulky structure of alcuronium while the quaternary d-tubocurarine) NMBDs. Relative accessibilities of substi- ammonium group, and especially the tertiary methyamino tuted ammonium groups on depolarizing and competitive group on d -tubocurarine, are overtly exposed. From Baldo NMBDs. The freehand fl exibility of decamethonium is due BA et al. On the origin and specifi city of antibodies to neu- to two trimethylammonium groups at the ends of a ten-sp3 - romuscular blocking (muscle relaxant) drugs: an immuno- carbon chain while fl exibility of succinylcholine is some- chemical perspective. Clin Exp Allergy 2009;39:325. what more restricted by the presence of two ester groups Reprinted with permission from John Wiley and Sons

[email protected] 260 7 Drugs and Other Agents Used in Anesthesia and Surgery

R1 R2 R3

O + Vecuronium N CH3COO- N O CCH CH 3 CH 3 H 3 CH 16 R 3 CH3 3 + + R1 Pancuronium N CH3COO- N 2 H CH3 3 O R2 H Rocuronium N HO- N+

CH2CHCH2

R1 Vecuronium

Pancuronium

Rocuronium

Fig. 7.9 Two-dimensional and space-ﬁ lling models of origin and speciﬁ city of antibodies to neuromuscular the bulky and rigid competitive aminosteroid NMBDs blocking (muscle relaxant) drugs: an immunochemical vecuronium, pancuronium, and rocuronium. Note the perspective. Clin Exp Allergy 2009;39:325. Reprinted structural differences of the three compounds at R 1 and at with permission from John Wiley and Sons R2 and R 3 for rocuronium. From Baldo BA et al. On the

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 261 of the sensitizing agent(s) is examined in more detail below.

7.4.4.2 Cross-Reactions of Neuromuscular Blocking Drugs at the Clinical Level The frequently used aminosteroid NMBD rocuronium provides a good example of our current understanding of cross-sensitivity of NMBDs from the clinical perspective. Soon after rocuronium’s release, its cross-reactivity with fi ve other NMBDs was assessed by skin testing, reactivity with serum IgE antibodies and leukocyte histamine release. Of the 31 patients included in the study, 30 showed some cross-reactivity with at least one NMBD and ten did not cross-react with rocuronium leading to the suggestion that the NMBD might be safe for those patients. In addi- Fig. 7.10 Concentration-dependent inhibition of binding tion, of fi ve patients positive to all the NMBDs of IgE antibodies specifi c for tertiary and quaternary methyl (mainly) and ethyl amino groups by neuromuscu- examined, one patient showed no cross-reactivity lar blocking drugs. Side-by-side quantitative comparisons at all with rocuronium. A recent assessment of of inhibitory potencies of rocuronium and alcuronium cross-reactivity between rocuronium and with six other neuromuscular blocking drugs. Refer to vecuronium, succinylcholine and benzylisoquin- Table 7.11 for the precise quantitative relationships. The dashed line indicates 50 % inhibition. From Pham NH oline employing skin tests, serum IgE antibody et al. Studies on the mechanism of multiple drug allergies. assays and the BAT found that 41 patients (69 %) Structural basis of drug recognition. J Immunoassay with established rocuronium allergy showed evi- Immunochem. 2001;22:47. Reprinted with permission dence of cross-reactivity with vecuronium. For from Taylor & Francis succinylcholine, 76 % of the rocuronium-allergic group had serum IgE antibodies that reacted with the drug but only one-third of these patients frequency. Increasing concern about the drug’s showing serum IgE-reactivity had a positive skin safety culminated in the Norwegian Medicines test and/or BAT to succinylcholine. Cross- Agency publishing an alert and recommending reactivity with the benzylisoquinolines cisatracu- restriction of usage to urgent intubations but oth- rium and atracurium was either entirely absent or, ers have claimed that false-positive skin tests at most, equivocal in four patients. overestimating the incidence of anaphylactic cases and/or the increased usage and market share 7.4.4.3 A Topical Practical Example of are the reasons for rocuronium’s alleged risk. the Value of Insights Obtained An alternative possible explanation based on a from Cross-Reaction Studies: structural perspective has recently been advanced. Rocuronium and the Risk In an immunochemical investigation, the relative of Anaphylaxis inhibitory potencies of rocuronium and seven other Rocuronium also provides a good example of NMBDs were compared side by side in quantita- allergenic cross-reactivity detected by IgE anti- tive hapten inhibition experiments using a serum bodies from the immunological perspective. With containing IgE antibodies with a precisely defi ned rocuronium’s increasing popularity and wide specifi city for tertiary and quaternary “small,” that usage has come a coincident increase in reports of is, methyl and ethyl (ethyl less strongly recognized), anaphylaxis to the drug leading some clinicians to mono, di and tri (di and tri less strongly recog- call for closer monitoring of the reports and their nized), alkyl amino groups (Fig. 7.10 ). In keeping

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Table 7.11 Demonstration of cross-reactivity between neuromuscular blocking drugs. Results of immunoassay inhibition studiesa with a patient’s serum containing IgE antibodies reactive with “small” ammonium groupsb Neuromuscular Alkylamino or alkylammonium Amount (nmol/tube) of drug for 50 % blocking drug group Structure(s) of group(s) inhibition of binding of IgE antibodies Atracurium 2 Quat methylammonium + 0.25 N CH 3 d -Tubocurarine 1 Quat dimethylammonium + 0.25 N (CH3)2

1 Tert methylammonium NH+ CH 3 Vecuronium 1 Quat methylammonium + 1.4 N CH3

N H 1 Tert amino + 1.5 Pancuronium 2 Quat methylammonium N CH 3 Succinylcholine 2 Quat trimethylammonium + 2 N (CH3)3 Gallamine 3 Quat triethylammonium 4 N+(C H ) 2 5 3 Rocuronium 1 Quat allylammonium + 10 N -CH2-CH=CH2

1 Tert amino N−H

Alcuronium 2 Quat allylammonium 30 N+-CH -CH=CH 2 2 Data from Pham NH et al. J Immunoassay Immunochem 2001; 22:47 Tert tertiary, Quat quaternary a For experimental details see Pham NH et al. J Immunoassay Immunochem 2001;22:47 b Tertiary and quaternary mono-, di- and tri-alkylamino groups but only if alkyl groups are “small”, viz., methyl or perhaps ethyl with the recognition speciﬁ city of the antibody ylammonium ions of greater complexity (the combining sites and its complementary dimethyl- methylpiperidinium structure in vecuronium and amino antigen-solid phase, NMBDs containing pancuronium and trimethylammonium ions of methylammonium groups were the most strongly succinylcholine), recognition by antibody declines recognized structures. Thus, atracurium and (Table 7.11). The markedly poorer inhibitory d-tubocurarine were the most active inhibitors fol- potencies exhibited by rocuronium and alcuronium lowed by vecuronium, pancuronium, succinylcho- can be explained by the presence in both drugs of line, and gallamine in that order. Table 7.11 relates the N -allyl (also called propenyl) pyrrolidinium the structures of the ammonium ions on the quaternary ammonium ion (Fig. 7.11a, b) and this NMBDs to the inhibitory potencies of all eight conclusion is reinforced by the signiﬁ cantly NMBDs examined. The most poorly recognized weaker inhibition obtained with alcuronium which structures were rocuronium and alcuronium, has two of these groups compared to rocuronium requiring 40 and 120 times as much drug respec- which has one. A comparison of the structures set tively as alcuronium and d-tubocurarine for out in Fig. 7.11 provides a visual perspective to 50 % inhibition. These results show clearly that this structure–activity interpretation revealing the best recognized structures contain at least one the marked differences in size, shape and orien- quaternary methyl or dimethylammonium ion of tation between the methylammonium group on simple structure (atracurium and d -tubocurarine) most NMBDs, for example the methylammonium and as these groups are replaced by a tertiary ion of vecuronium (Fig. 7.11c, e) and the allylam- amino group (vecuronium) and quaternary meth- monium ion of rocuronium and alcuronium

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 263 a bc H2 C + CH2 N C O OCOCH H O CH CH 3 CH C 3 3 H N 3 H O + + CH N HO CH CH OH CH N N 3 N 3 N H2C H CH2 H CH C H 3 + HO H C N H C C O H 3 H H2C C O H2 Rocuronium Alcuronium Vecuronium de

quat. allyl- quat. allyl- quat. methyl- quat. methylammonium ion pyrrolidinium ion ammonium ion piperidinium ion

Fig. 7.11 Two-dimensional structures of ( a ) rocuronium, ( d) and the quaternary methylammonium and methylpi- ( b ) alcuronium, and (c ) vecuronium with the quaternary peridinium ions are shown in ( e ). In (d ) and (e ), hydrogen allylpyrrolidinium ions of rocuronium and alcuronium and carbon atoms of the pyrrolidine and piperidinium shown in red (a and b) and the quaternary methylpiperi- rings are colored light blue and gray, respectively, to dis- dinium ion of vecuronium shown in blue (c ). Three- tinguish these groups from the allylammonium and meth- dimensional CPK molecular models of the quaternary ylammonium ions allylammonium and allylpyrrolidinium ions are shown in

(Fig. 7.11d). In the late 1970s and early 1980s of the NMBDs involved and the structure–activity alcuronium was, like rocuronium now, also said to relationships demonstrated in quantitative IgE be more likely to provoke allergic reactions than antibody binding and recognition (from inhibi- other NMBDs. It may be, for reasons not under- tion studies) experiments. stood, that the allylpyrrolidinium group makes these two NMBDs somewhat more anaphylacto- genic and therefore a greater risk than other 7.4.5 The NMBD–IgE Conundrum: NMBDs. By combining these structural insights The Origin of IgE Antibodies with experiments designed to compare IgE anti- to Neuromuscular Blocking body-mediated release of mediators (for example, Drugs in basophil activation and histamine release studies) it might be possible to implicate or exonerate 7.4.5.1 The Speculations the allylpyrrolidinium specifi city as an added risk and Evidence so Far for NMBD-induced anaphylaxis. In both clinical and research areas of NMBD The above fi ndings demonstrate that as well as hypersensitivity, much of the diffi culty in inter- viewing and interpreting cross-reactions of preting and explaining results of IgE recognition NMBDs from the purely clinical perspective pro- studies stems from our lack of information on the vided by a patient’s symptomatology, skin tests precise specifi city(ies) of the preexisting antibod- and the tryptase assay, it remains important to ies found in the sera of patients exhibiting aller- consider both the underlying structural features gic sensitivity to the drugs. These IgE antibodies

[email protected] 264 7 Drugs and Other Agents Used in Anesthesia and Surgery are present in the sera of apparently normal, and found positive reactions to the NMBD in healthy subjects before they are exposed to an 0.4 % and 3.7 % and to morphine in 5 % and NMBD and, as discussed above, appear to occur 10 % of Norwegian blood donors and allergic in a signifi cant proportion of the general popula- subjects, respectively. No serum from Sweden tion, at least in France and Scandinavia. To be in was positive to either drug. When the morphine a position to better understand some of the seem- analog pholcodine, a cough suppressant sold ingly anomalous results of direct binding, inhibi- without restriction in Norway but not in Sweden, tion and cross-reactivity experiments, knowledge was tested, sera from 6 % of Norwegian blood of the antigenic source of the NMBD-reactive donors, but no Swedish sera, reacted positively. antibodies is essential. The answer to a second In addition, tests for IgE antibodies to pholcodine intriguing and important question—why are the and morphine on sera from 65 anaphylactic antibodies of the IgE class?—might bring with it Norwegians showed a similar incidence of posi- clues for the evolution and role of immunoglobu- tive reactions. This suggested that pholcodine lin E in humans and perhaps contribute to a better might be the source of sensitization to NMBDs in understanding of the presence of preexisting Norway and this appeared to be supported by a allergic sensitivities to some drugs and other large boost in IgE antibodies to pholcodine, mor- allergens and even a role for IgE in the pathogen- phine, and succinylcholine following dosage with esis of some infectious diseases. In seeking a cough syrup containing pholcodine. answers to these two questions, the most obvious A follow-up study involving the administration of existing clue seems to be the substituted ammo- pholcodine to patients with IgE-mediated ana- nium (tertiary and quaternary) groups present in phylaxis to NMBDs again revealed increases in all the NMBDs and complementary, at least in IgE antibody levels, but a boost to total IgE levels part, to the NMBD-reactive IgE antibodies. An was also seen. Maximum serum concentrations of early suggestion in 1983 of the origin of the anti- immunoglobulin E were much higher than the bodies, viz. stimulation by environmental agents sum of the identifi ed antibodies, suggesting at in the form of cosmetics, hair products, house- least some non-pholcodine-specifi c stimulation hold chemicals and toilet items etc remains of IgE. This seems like an important point. appealing and appears to explain a number of Experiments 20 years ago with sera containing aspects of sensitization but little or no substantive high levels of total IgE from subjects with no his- evidence has been forthcoming and, more tory of allergy to NMBDs but with eczema, asper- recently, other speculative explanations involv- gillosis, food allergies, and/or asthma revealed ing mammalian rhesus (Rh) ammonium trans- clear reactions of IgE with NMBDs and morphine port proteins and phosphorylcholine antigens in direct binding and inhibition experiments and (see Sect. 7.4.5.3 ) have been advanced. Again, recent tests on 32 subjects with no hypersensitiv- these two suggestions remain unsupported by ity to NMBDs but total IgE concentrations in evidence but both are amenable to experimental excess of 1,500 kU/l showed positive reactions to investigation. A new theory with some evidence pholcodine and morphine in approximately one- has, however, recently been advanced from third of the subjects. In the latter study, IgE anti- Scandinavia and this will be outlined in the fol- bodies to morphine and pholcodine were found in lowing section. 88 and 86 %, respectively, of patients allergic to rocuronium but positive reactions to these drugs 7.4.5.2 Pholcodine and Anaphylaxis were detected in only 0.5–1 % of sera from 95 to Neuromuscular Blocking healthy controls, 95 atopic controls, and nine sub- Drugs in Scandinavia jects exposed to morphine. Starting with the knowledge that anaphylactic In morphine, codeine, and pholcodine the reactions to NMBDs are six times more common determinant structure recognized by NMBD- in Norway than in Sweden, researchers looked for reactive IgE antibodies, that is, the tertiary IgE antibodies to succinylcholine and morphine N-methyl group, is freely accessible so there

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 265 seems to be no reason why all three drugs would “benefi ts outweigh its risks” and “all marketing not react equally well with antibodies. The three authorizations for medicines containing pholco- compounds differ in structure only at position 3 dine should be maintained throughout the where morphine has a hydroxyl group, codeine a European Union.” methoxy, and pholcodine a morpholinylethyl group (Fig. 7.12 ) so if pholcodine, but not its two 7.4.5.3 Phosphorylcholine and analogs, induces allergic sensitization and has an Neuromuscular Blocking immunological boostering effect on IgE antibody Drug-Reactive IgE Antibodies levels, the conclusion seems inescapable, viz., if Phosphorylcholine occurs widely in natural these effects are real, only the morpholinoethyl products, connected via phosphodiester linkages group can account for the differences. A possible to N-acetylglucosamine in proteoglycans and mechanism for such sensitization remains to be glycolipids. It also occurs in macromolecules elaborated. In 2009, results of a multinational, that have immunomodulatory and anti-infl amma- multicenter study on national pholcodine con- tory properties such as promotion of Th2 sumption and the prevalence of IgE sensitization responses and inhibition of pro-infl ammatory were published with the conclusions that the cytokine production by macrophages. “Natural” result “lends additional support to the PHO antibodies, that is, antibodies formed without (pholcodine) hypothesis” and “that other, yet exposure to foreign antigens via infection or pas- unknown, substances may lead to IgE- sive or active immunization, also occur widely sensitization towards NMBAs.” A signifi cant and such antibodies to phosphorylcholine positive association between pholcodine con- develop under sterile conditions in mice and are sumption and prevalences of IgE sensitization to found as nonpathogenic autoantibodies produced pholcodine and morphine but not to succinylcho- by CD5+/B-1 B cells in humans. Some monoclo- line or choline hapten (used as mimic for the suc- nal antibodies in both the mouse and humans and cinylcholine IgE-binding determinant) was the acute phase protein, C-reactive protein, have found, but absence of succinylcholine sensitivity combining sites complementary to phosphoryl- in some countries with signifi cant levels of anti- choline, the immunodominant structure in pholcodine IgE antibody raises doubts about the “C-substances” a teichoic acid of Streptococcus hypothesis. pneumoniae, and a component of other In February 2011 the European Medicines C-substance-like antigens widely distributed in Agency’s Committee for Medicinal Products for some other bacteria, fungi, protozoa, plants, Human Use (CHMP) began a review of the safety arthropods, and helminthes. C-substances, iso- and effectiveness of pholcodine following lated by affi nity chromatography on C-reactive concerns that the drug might put people at risk protein and mouse anti-phosphorylcholine IgA from anaphylaxis during anesthesia. At the time antibodies from house dust mites, intestinal of the review, medicines containing pholcodine worms, and fungi, especially dermatophytes, had been withdrawn from the market in Norway were shown to have allergenic properties by skin in 2007 (in Sweden this occurred in the 1980s), prick test, IgE antibody binding. and histamine further publications from Scandinavia had stated release studies. C-substances precipitate with that pholcodine use may increase the likelihood C-reactive protein and anti-phosphorylcholine of anaphylaxis to NMBDs and the French medi- monoclonal antibodies, both reactions are inhib- cines regulatory agency had asked the CHMP to ited by phosphorylcholine, and there is evidence assess the risk-benefi t balance of pholcodine and that anti-phosphorylcholine antibodies have a decide whether or not the marketing authoriza- protective effect against S . pneumoniae infections for the drug should be changed. In tions and artherosclerosis. Antigens containing November 2011 the European Medicines Agency phosphorylcholine are common in helminthes as completed its review concluding that the existing are anti-phosphorylcholine-specifi c IgE antibod- evidence against pholcodine is “weak,” the drug’s ies in infected hosts, so, together with natural IgE

[email protected] 266 7 Drugs and Other Agents Used in Anesthesia and Surgery

Morphine Codeine

Pholcodine

Fig. 7.12 Two-dimensional structure and space-ﬁ lling 17 on the opposite side of each of the three drugs. From models of morphine, codeine, and pholcodine showing the Baldo BA et al. On the origin and speciﬁ city of antibodies to different substituents (R) at position 3. Note that any sub- neuromuscular blocking (muscle relaxant) drugs: an immu- stituent at position 3 on all three compounds cannot restrict nochemical perspective. Clin Exp Allergy 2009;39:325. access of antibodies to the N -methyl substituent at position Reprinted with permission from John Wiley and Sons

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 267

Fig. 7.13 Structural a identity (shown in bold ) of CH OH ( a ) the polar head region of 3 the phosphorylcholine CH3 N CH2 CH2 O POH molecule and (b ) both O termini of the NMBD, CH3 succinylcholine b CH3 O CH3 CH3 NCH2 CH2 O CH2 C C CH2 O CH2CH2 NCH3 CH3 O CH3 antibodies to this commonly encountered hapten, ylcholine, can bind to preexisting IgE antibodies phosphorylcholine-reactive immunoglobulins E in patients’ sera and precipitate an anaphylactic antibodies are likely to be not uncommon in reaction. Figure 7.13 shows the structural iden- human populations. In the normal situation, the tity of both termini of the NMBD succinylcholine IgE antibody-binding so-called cross-reactive and the polar head region of the phosphorylcho- carbohydrate determinants derived from some line molecule. Cross-reaction between the substi- plant-derived foods, pollens, and Hymenoptera tuted ammonium groups of succinylcholine and venoms, and the phosphorylcholine determinant, the ammonium groups of other NMBDs means act like “dormant” allergens demonstrating no that anti-phosphorylcholine IgE antibodies will clinical relevance as evidenced by absence of also react, to equal or lesser extent, with these clinical symptoms and negative skin tests in NMBDs. Investigations undertaken with this patients. However, in both laboratory animals speculation in mind should ultimately reveal its and humans, intravenous challenge with some correctness or otherwise. antigens, for example, parasite glycoconjugates in rats and a galactose disaccharide in humans, provokes an anaphylactic response mediated by 7.4.6 Sugammadex and preexisting IgE antibodies in the patients’ sera. In Anaphylaxis to Rocuronium these cases, the host has gone from a clinically benign or dormant situation where pre- 7.4.6.1 Sugammadex and Its Binding sensitization to an allergen exists, to an unsus- to Rocuronium pected and severe immediate type I anaphylactic The naturally occurring γ-cyclodextrin is a response. It seems likely that a similar response circular oligosaccharide made up of eight 4 might occur if an antigen containing phosphoryl- D-glucopyranoside units in rigid C1 conforma- choline was presented intravenously to subjects tion and linked α(1-4) around a central cavity with natural or immune anti-phosphorylcholine (Fig. 7.14a). The oligosaccharide has a truncated IgE antibodies. However, with little likelihood of cone or toroidal shape with a total of eight pri- C-substances being administered intravenously mary hydroxyl groups on carbon 6 at the narrow and with this potential allergenic determinant or so-called primary end and 16 secondary usually inaccessible as a component of phospha- hydroxyls on carbons two and three of each glu- tidylcholine and sphingomyelin, the conversion cose unit at the wider or secondary rim of the of a dormant to an active allergic response medi- molecule (Fig. 7.14b). Cyclodextrins are used in ated by a phosphorylcholine–IgE antibody inter- the pharmaceutical, food, and cosmetic industries action seems unlikely. The intravenous to encapsulate molecules within their hydropho- administration of an NMBD during anesthesia bic cavities, thus forming stable guest-inclusion creates, however, the circumstances whereby complexes to improve solubility, stability, and structures that closely resemble, or are identical delivery of drugs, prevent drug interactions, and to, the quaternary ammonium group of phosphor- reduce irritation, unpleasant tastes, and smells.

[email protected] a 6 OH 5 O O 1 OH 4 O 1 2 4 O 3 OH OH HO O OH 4 HO 1 O OH HO O 1 HO OH 4 O HO

OH O 4 HO OH 1 O HO OH O 1 OH 4 HO O OH HO HO O 4 1 O HO O 1 O 4 HO b

-COONa -COONa 2)2 ) 2 Thio(2- 2 S-(CH )2-COONa S-(CH 2 -COONa S-(CH carboxyethyl) ) 2 sodium group 2 S-(CH ) -COONa 2 2 6 S-(CH 6 6 Primary rim 6 ) -COONa -COONa 6 2 2 ) 2 S-(CH 2 ) -COONa 2 2 S-(CH S-(CH

6 6 ~11-11.5Å 6 Hydrophilic exterior 7.8Å

Secondary 2 3 2 3 rim 2 3 2 3 OH OH OH OH OH OH 2 OH OH 3

2 OH 3 OH Secondary 2 3 hydroxys 3 2 OH OH OH OH OH OH

7.5-8.3Å

17.5Å

Fig. 7.14 ( a ) Two-dimensional chemical structure of length by 3.2–3.7 Å (7.8 Å to 11–11.5 Å) by addition of γ-cyclodextrin, made up of eight glucopyranose units con- these groups. See also Fig. 7.16. From Baldo BA et al. The nected in a circle by α-1,4 bonds. ( b) Diagammatic repre- cyclodextrin sugammadex and anaphylaxis to rocuronium: sentation of the toroidal shape of sugammadex showing the Is rocuronium still potentially allergenic in the inclusion attachment of eight thio(2-carboxyethyl)sodium groups at complex form? Mini Rev Med Chem 2012;12:701. the primary face and indicating the extension of the cavity Reprinted with permission from Bentham Science Publishers

[email protected] 7.4 Anaphylaxis to Neuromuscular Blocking Drugs 269

COONa For a stable inclusion complex to form, the guest S molecule must preferably be fully accommodated CH2CH2 6 by the cyclodextrin and this sometimes involves 4 O extension of the cyclodextrin carrier by chemical 5 2 means. This was the strategy employed to encap- O 3 1 sulate and accommodate rocuronium. In an H O attempt to encompass the entire six rings of H rocuronium, chemical modiﬁ cations were carried 8 out to elongate the cavity by per-6 substitution of each of the primary hydroxyls with propionic acid side chains each linked by a thiol-ether group Fig. 7.15 Chemical structure of one of the eight α-1-4- linked glucopyranose units of sugammadex, 6-perdeoxy- (Fig. 7.15). This achieved a lengthening of the 6-per(2-carboxyethyl)thio-γ-cyclodextrin sodium salt, cyclodextrin carrier from ~7.8 Å to about 11.5 Å, showing the thio(2-carboxyethyl)sodium group linked at the approximate distance between the C3 carbon position 6 of the glucose molecule. From Baldo BA et al. of ring A and C16 of ring D on the rocuronium Drug-speciﬁ c cyclodextrins with emphasis on sugammadex, the neuromuscular blocker rocuronium and periop- molecule (Fig. 7.16 ), with the four rings A, B, C, erative anaphylaxis: implications for drug allergy. Clin and D within the cavity and the pyrrolidinium Exp Allergy 2011;41:1663. Reprinted with permission quaternary ammonium and the morpholine from John Wiley and Sons groups visible at the primary and secondary faces, respectively (Fig. 7.17 ). A method to encapsulate the NMBD had been sought to aid solubility and

OCOCH3 Fig. 7.16 Two-dimensional CH3 chemical structure and H space-ﬁ lling CPK O 17 molecular model of CD16 N+ rocuronium showing the CH3 N 15 ~11 Å distance between the C3 atom on ring A of the 2 CH CHCH AB H 2 2 gonane nucleus and the C16 3 atom on ring D. The propenyl (allyl) pyrrolidin- HO ium quaternary ammonium H group is shown in magenta ~11Å on the two-dimensional structure and the CPK model. See also Figs. 7.14b and 7.17 for the signiﬁ cance of the ~11 Å distance. From Baldo BA et al. The cyclodextrin sugammadex and anaphylaxis to rocuronium: Is rocuronium still potentially allergenic in the inclusion complex form? Mini Rev Med Chem 2012;12:701. Reprinted with permission from Bentham Science Publishers ~11Å

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Fig. 7.17 Space-ﬁ lling CPK and ball-and-stick three- of sugammadex (see Fig. 7.15 ) is ringed in black . dimensional molecular models ( a and b, normal CPK col- Substituents attached to ring A of the steroid backbone of oring; c and d , mono-colors to distinguish molecules) rocuronium (see Fig. 7.16) are partially visible at the sec- illustrating the encapsulation of rocuronium by sugamma- ondary rim ( left-hand side here) of the cavity, rings B, C, dex to form a sugammadex–rocuronium inclusion com- and D of rocuronium are within the cavity, and the quater- plex. (a ) Left-hand structure, sugammadex; right-hand nary ammonium group attached to ring D is surrounded structure, rocuronium. ( b ) Sugammadex–rocuronium by carboxyl groups at the opposite, primary rim end inclusion complex. ( c ) Left-hand structure (green ), ( right-hand side here), of the sugammadex carrier. From sugammadex; right-hand structure (blue ), rocuronium. (d ) Baldo BA et al. Drug-speciﬁ c cyclodextrins with empha- Sugammadex–rocuronium inclusion complex. The posi- sis on sugammadex, the neuromuscular blocker tively charged propenyl pyrrolidinium quaternary ammo- rocuronium and perioperative anaphylaxis: implications nium group on rocuronium (see Fig. 7.16 ) is ringed in red for drug allergy. Clin Exp Allergy 2011;41:1663. and one of the eight thio(2-carboxyethyl) sodium groups Reprinted with permission from John Wiley and Sons

[email protected] 271 decrease pain when injected, but the effectiveness 7.4.6.3 Relative Affi nities of the of the binding led to the application of sugam- Interaction of Rocuronium madex for the reversal of rocuronium-induced with Sugammadex and IgE blockade by sequestering the drug and removing Antibodies 7 −1 it from the neuromuscular junction. With an association constant K a of 1.8 × 10 M sugammadex forms a stable complex with its 7.4.6.2 Rocuronium, Sugammadex, guest molecule rocuronium. There is no informa- and Anaphylaxis tion available on the affi nities and dissociation Coincident with rocuronium’s popularity to constants of IgE antibody–rocuronium com- induce neuromuscular blockade has been an plexes, but the average association constants for increase in reports of anaphylaxis to the drug and multideterminant, multivalent allergens such as the suggestion that sugammadex, as well as rap- pollen proteins for example with their comple- idly reversing neuromuscular block, might also mentary IgE antibodies are often high and in the offer a novel treatment to reverse anaphylaxis range 1010 –10 11 M−1 . By virtue of their tertiary caused by rocuronium. However, because the and quaternary ammonium groups, NMBDs are allergenic ammonium group of the encapsulated probably at most bideterminant and bivalent and rocuronium molecule is visible at the primary one might therefore expect lower average affi ni- face of the sugammadex carrier, doubts were ties (in terms of the average association con- expressed that sequestered rocuronium mole- stants) and avidities (the overall stability or cules would be protected from interacting with strength of the complex) for rocuronium–serum IgE antibodies. Case reports quickly appeared IgE antibody interactions. In fact, the affi nities apparently supporting the suggestion that sugam- and avidities of rocuronium–IgE complexes may madex might be a new and useful treatment to be even lower than fi rst expected because of the manage rocuronium-induced anaphylaxis and at specifi city of the drug–antibody interaction. The the time of writing there are at least seven reports absolute specifi cities of NMBD-reactive IgE from six different countries of the mitigation of antibodies in patients’ sera are not known since anaphylaxis provoked by the drug. Even so, the the source of the sensitizing agent(s) is also ammonium ion at position 16 on the steroid unknown, but, whatever the original antigenic nucleus of encapsulated rocuronium is loosely stimulus, it is probably not an NMBD. This in surrounded by (2-carboxyethyl)thio groups of turn leads to antigen–antibody combining site sugammadex, and with the tertiary ammonium complexes of poorer complementarity or “fi t” group partly visible at the opposite end of the than seen in reactions between antibody combin- inclusion complex, both potentially reactive ing sites and the antigen that stimulated the pro- ammonium groups might still be accessible for duction of the antibody in the fi rst place. If binding with complementary IgE molecules sugammadex is to successfully mitigate an ongo- (Figs. 7.16 and 7.17 ). These are questions that ing anaphylactic response induced by rocuronium should be amenable to experimental in vitro in a patient, its association constant for reaction investigation in the laboratory. In the meantime, with the NMBD has to be higher than the average since challenge studies with encapsulated association constant of the patient’s IgE antibod- rocuronium or induction of hypersensitivity with ies for rocuronium. Affi nities for antibodies the NMBD followed by administration of sugam- reacting with the same hapten can differ by up to madex are not ethically permissible, further a factor of 10 5 so the effectiveness of sugamma- information on the effectiveness or otherwise of dex in reversing rocuronium-induced anaphy- sugammadex in mitigating rocuronium-induced laxis in different patients may vary. In general anaphylaxis will have to await the accumulation though, one may predict that higher affi nities for of more case reports. the IgE–rocuronium complexes than for the

[email protected] 272 7 Drugs and Other Agents Used in Anesthesia and Surgery sugammadex complex would result in the failure 7.4.7 Antigenic Similarity Between of sugammadex administration to mitigate a α-Bungarotoxin and reaction while mitigation of anaphylaxis would Neuromuscular Blocking follow if the sugammadex–rocuronium affi nity Drugs were higher. Elapid snake neurotoxins bind specifi cally and 7.4.6.4 Allergy to Sugammadex with high affi nity to acetylcholine receptors on Sugammadex itself may occasionally cause ana- muscle cells and the electroplax of electric fi shes. phylaxis and the withholding of approval by the α-Bungarotoxin (α-BT) is a 74 amino acid (MW United States Food and Drug Administration in 7,983 Da) basic poypeptide α-neurotoxin from 2008, even though the drug is approved for use in the Taiwanese banded krait that binds irrevers- Europe, was due to concerns about allergic reac- ibly and competitively to the nicotinic acetylcho- tions to the modifi ed cyclodextrin. In a recent line receptor at the motor end plate producing a report of such a reaction, intense erythema of the non-depolarizing block of neuromuscular thorax, severe lip and palpebral edema, a fall in transmission. Competitive NMBDs like blood pressure, tachycardia, and bilateral wheeze d - tubocurarine also act at the postjunctional were reported in a young adult 1 min after receiv- membrane blocking the transmitter action of ace- ing a low dose of sugammadex (3.2 mg/kg). The tylcholine. Investigations of the affi nities of patient proved skin test positive to sugammadex. NMBDs for the α-BT binding sites at the Three other cases of sugammadex-induced hyper- α-subunit of the acetylcholine receptor showed sensitivity were recently reported from Japan that the drugs had some of the highest affi nities when the drug was administered at doses ranging for the binding sites, in fact, even higher than from 1.9 to 2.2 mg/kg. In one case, facial ery- acetylcholine and nicotine. These fi ndings sug- thema and blepharedema developed 3 min after gested that the NMBDs and α-BT probably share sugammadex and the patient was skin test positive some properties of, for example, shape, confor- to sugammadex. The second patient developed mation, and charge. The possibility that a similar- hypotension and generalized erythema 3 min after ity between the widely chemically different the cyclodextrin and again the skin test proved ligands might be detected immunologically was positive. The third patient, not skin tested, therefore investigated in competitive binding responded with wheeze and intense erythema studies with sera from 16 different patients who 4 min after sugammadex. The three reactions experienced anaphylaxis to NMBDs. For two occurred out of a total of 1,864 instances in which patients, both of whom reacted to succinylcho- sugammadex was administered during general line, α-BT clearly inhibited the binding of IgE anesthesia. Sugammadex-specifi c IgE antibodies antibodies to a choline-solid phase, but little or were not directly identifi ed in these cases which no inhibition was seen when triethylcholine-, might have been due to direct drug- induced medi- alcuronium-, d -tubocurarine-, or vecuronim- ator release. It has been pointed out that the use of solid phases were used. For one patient, host molecules such as sugammadex, other cyclo- d- tubocurarine was the most potent inhibitor dextrins, dendrimers, vesicles, cell ghosts, hydro- (IC50 0.72 nmol), alcuronium was the weakest gels, and so on to “carry” other molecules and (IC50 58 nmol) and α-BT was intermediate in improve drug delivery will sometimes produce inhibitory potency (IC50 16 nmol) but more active changed immunological recognition including the than choline and triethylcholine (IC50 23 and potentiation and reduction of allergenic proper- 20 nmol, respectively). Fifty percent inhibitory ties. There will be a need to take this into account concentrations for succinylcholine, decametho- in preclinical drug safety assessments and the nium, gallamine, and pancuronium were 9.8, 1.9, possibility of altered allergenic behavior of even 8.4, and 7.7 nmol, respectively. For the second some well-known drugs should not be overlooked serum, a similar inhibitory pattern was seen with by allergists and dermatologists. α-BT being more potent than alcuronium while

[email protected] 7.5 Anaphylaxis to Hypnotic Drugs Used in Anesthesia 273 decamethonium and d -tubocurarine were the best anesthesia by propofol, but it is still the classic inhibitors. Competition between polypeptides drug used in rapid sequence inductions and it is and small ligands at protein binding sites are sometimes used in electroconvulsive therapy. known with nucleases and proteases. Changeux postulated that the sizes of α-BT (diameter 27 Å) 7.5.1.1 Incidence of Reactions, and NMBDs such as d -tubocurarine (distance Clinical Features, and between nitrogens 14 Å) did not rule out the pos- Epidemiology of Anaphylactic sibility of competitive binding at the cholinergic Reactions to Thiopentone receptor. Further investigation of the possible Hypersensitivity reactions to thiopentone are gen- antigenic similarity between the peptide toxin erally regarded as rare, with incidences of ana- and NMBDs seems warranted. Identifi cation of phylaxis of about 1 in 22,000 and 1 in 29,000 sera showing clear IgE antibody recognition of suggested in two prospective studies and only 1 in α-BT might prove useful in attempts to better 400 in a third. After its introduction in 1934, 41 defi ne and ultimately understand the different cases of anaphylaxis were attributed to the drug in and often puzzling antibody NMBD recognition 1980, only 13 published cases were identifi ed in patterns from different patients allergic to the 1974 while 2 years later it was concluded that 30 same NMBD. cases had occurred since the drug’s introduction. In 1985 in France 258 cases of anaphylaxis were identifi ed in a literature search. The true incidence 7.5 Anaphylaxis to Hypnotic of allergic reactions to thiopentone is therefore Drugs Used in Anesthesia not clear, but it seems likely that after the many millions of injections of the drug given annually, Table 7.1 shows that amongst the anesthetic many cases of anaphylaxis may have gone unrec- agents used in the years covered by the main sur- ognized or been misdiagnosed. veys of anaphylactic responses, the Cremophor- Unlike NMBDs, previous exposure to thio- based drugs alfathesin and propanidid contributed pentone is a predisposing factor in producing signifi cantly, at least in the Australian series, to anaphylaxis. Anaphylaxis on fi rst exposure is the list of induction agents causing reactions with unusual; at least six exposures are usually an incidence as high as 1 in 875 cases. The drugs required and up to 37 uneventful exposures have that replaced them, etomidate and propofol, been reported prior to the fi rst reaction. Patients rarely produce severe hypersensitivity reactions who react to thiopentone are older than reactors and that is also true for ketamine and midazolam. to NMBDs (43 vs. 36 years), women predomi- Therefore, because the Cremophor-based induc- nate with a female to male ratio of three to one, tion agents are of little or no relevance today and and the incidences of a history of allergy (42 %), allergic or allergic-like reactions to etomidate, atopy (36 %), and asthma (22 %) are similar to ketamine, and midazolam are so few, only the NMBD reactors. Also, unlike NMBDs, hyper- commonly and widely used propofol will be con- sensitivity reactions ranging from rashes to urti- sidered here along with the long-standing induc- caria and severe exfoliative dermatitis have been tion agent thiopentone. seen with thiopentone.

7.5.1.2 Diagnosis of Immediate 7.5.1 Thiopentone Allergic Reactions 7.5.1.2.1 Challenge Tests Used in the induction phase of anesthesia, thio- Challenge tests with thiopentone have been used pentone is an ultrashort acting barbiturate. The to conﬁ rm the diagnosis of anaphylaxis to the drug acts within 30–45 s producing its peak con- drug. Results obtained in challenge tests on two centration in the brain about 1 min after injection. patients give some indication of the responses Thiopentone has now been largely replaced in that might be expected. After building up the

[email protected] 274 7 Drugs and Other Agents Used in Anesthesia and Surgery challenge dose of thiopentone to 64 mg in a ent allergenic determinants on opposite sides of female patient 4 weeks after an anaphylactic-like the thiopentone molecule, the ethyl and second- reaction to the drug, the patient experienced ary pentyl groups at position 5 of the pyrimidine “head spinning,” turned bright red with erythema ring nucleus and the region of the ring encom- spreading down the body, the conjunctivae passing the attached sulfur atom (Fig. 7.18 ). became injected and the pulse rate rose abruptly Pentobarbitone, which differs from thiopentone to 135. Administration of incremental increases only in the hetero atom (O for the former, S for of thiopentone up to a dose of 220 mg to an adult the latter), was a key inhibitor in identifying the male who experienced a suspected anaphylactic alkyl group determinants while good inhibition reaction to thiopentone and was skin test negative with 2-mercaptopyrimidine, and to a lesser extent to the drug produced severe itching and erythema thiouracil, served the same purpose in identifying within 17 min. A systemic reaction following a the thio region as a second IgE antibody-binding skin test with thiopentone has also been reported. structure. Although the immunoassay is a valu- Therefore, although challenge tests can and have able supplement to skin testing for the detection been used to confi rm diagnosis, their use should of thiopentone-allergic sensitivity, the method probably be reserved for patients in remote areas can sometimes detect “false-positive” reactions where other tests are unavailable or for cases and the interpretation of results is therefore not where other tests are negative. This conclusion is always completely straightforward. Sera from reinforced by the fact that other useful and reli- NMBD-allergic subjects containing high levels able tests are available to diagnose sensitivity to of IgE antibodies to substituted ammonium thiopentone. groups react with the thiopentone-solid phase, but this reaction is not inhibited by preincubation 7.5.1.2.2 Skin Tests of the sera with NMBDs. Lability of thiopentone Diagnosis of suspected anaphylaxis to thiopen- at the high pH used to prepare the drug-solid tone generally rests on skin testing, a procedure phase appears to be the explanation why the that has proved its applicability. In a prospective NMBD-reactive IgE antibodies bind in the assay. skin test study on 83 patients, no signifi cant dif- The molecular basis of this reaction was eluci- ferences were seen between prick testing and the dated by further inhibition investigations. intradermal test. Prick testing is performed with 2-Mercaptopyrimidine but not thiopentone or undiluted ready-prepared drug solution, that is, thiobarbituric acid inhibited binding of the 25 mg/ml, while for intradermal testing, testing NMBD-positive sera indicating that the ring starts at a 1 in 10,000 dilution and proceeds up to nitrogens of the pyrimidine nucleus are the com- a maximum of 1 in 10, that is, a concentration of plementary binding structures for the NMBD- 2.5 mg/ml maximum. reactive IgE. These groups are presumably accessible to antibody binding on the drug-solid 7.5.1.2.3 Serum IgE Antibody Tests phase but sterically hindered on free thiopentone In the fi rst clear-cut demonstration of the involve- by the alkyl and, perhaps, keto groups. Table 7.12 ment of IgE antibodies in suspected anaphylactic summarizes the IgE antibody binding structures reactions to thiopentone, the drug was covalently on the thiopentone molecule identifi ed with sera coupled to bis -oxirane-activated Sepharose and from thiopentone-allergic patients and sera from used in a solid phase radioimmunoassays with patients allergic to NMBDs. Figure 7.19 shows sera from patients who experienced anaphylaxis diagrammatically how the pyrimidine ring nitro- to the drug. Specifi city of the binding of IgE antigens become accessible for antibody binding bodies in patients’ sera to the drug-solid phase once the bulky alkyl and keto groups are removed. was demonstrated by inhibition of binding with free thiopentone; 5 μmol of drug generally pro- 7.5.1.2.4 Leukocyte Histamine Release duced up to 65 % inhibition of binding. Extensive Injection of anesthetic doses of thiopentone into quantitative inhibition studies with a number of healthy, nonallergic humans leads to release of selected barbiturate analogs identifi ed two differ- histamine into the plasma, but the amount is

[email protected] ab

O NH CH3CH2 S CH CH CH CH 3 2 2 NH

CH3 O

Fig. 7.18 Two-dimensional chemical structure (a ) and pentyl groups at position 5 of the pyrimidine ring (shown CPK space-ﬁ lling model (b ) of thiopentone showing two in blue ; a and c ) and the region of the ring encompassing different allergenic determinants identiﬁ ed on opposite the attached sulfur atom (shown in magenta ; a and d ) sides of the thiopentone molecule, the ethyl and secondary

Table 7.12 IgE antibody-binding structures identifi ed so far on thiopentone Dominant structure in IgE-binding determinanta (in bold font) Serological fi ndings c Clinical relevance O Free drug (thiopentone) inhibits Presence of IgE of this specifi city binding of patient’s serum to indicates allergy to thiopentone CH3CH2 NH b d S “thiopentone”-solid phase CH3CH2CH2CH NH O CH3 O As above As above b CH3CH2 NH S CH3CH2CH2CH NH O CH3 N 2-Mercaptopyrimidinee unlike Reactive IgE in sera from some SH b thiopentone inhibits binding of NMBD-allergic subjects. Subjects IgE to “thiopentone”-solid phased not allergic to thiopentone N From Baldo BA & Pham NH. Structure–activity studies on drug-induced anaphylactic reactions. Chem Res Toxicol 1994;7: 703. Reproduced with permission from American Chemical Society a Exact confi nes of determinant not always clearly defi ned with all sera b Dominant feature of determinant shown. Other structures, in particular the pyrimidine ring, probably has an auxiliary function c Findings when used with sera from subjects allergic to thiopentone or an NMBD dThiopentone is used to prepare drug-solid phase, but coupling conditions probably lead to some decomposition of the thiopentone. Exact structure of attached species is therefore uncertain e 2-Mercaptopyrimidine also inhibits binding of IgE antibodies reactive with the thio region of thiopentone

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patient who experienced a life-threatening reaction to the drug. Thiopentone at a concentration as low as 7 × 10−6 M released histamine from leukocytes taken from a patient who reacted to the drug. Serum from the patient failed to passively sensitize cells from five different control subjects. This was interpreted as evidence for an anaphylactoid rather than an IgE-mediated anaphylactic reaction.

7.5.2 Propofol

Propofol, 2,6-diisopropylphenol, is administered intravenously and used in general anesthesia as a short-acting induction and maintenance agent. The drug is formulated as an oil–water emulsion containing 1 % propofol, soybean oil 10 %, glycerol, and egg phospholipid as emulgent.

7.5.2.1 Propofol and Anaphylaxis Although propofol is generally said to be a remarkably safe drug, 14 patients were reported to have life-threatening reactions within a few Fig. 7.19 Space-ﬁ lling models showing how the pyrimi- minutes of the administration of propofol, numer- dine ring nitrogens become exposed and accessible for ous other anaphylactic-like reactions have been antibody binding once the bulky alkyl and keto groups are reported, and other occasionally observed hyper- removed from thiopentone. Models for (a ) thiopentone and ( b) 2-mercaptopyrimidine. Removal from thiopentone of sensitivity reactions include bronchospasm, the bulky secondary pentyl group ( light blue area) and the angioedema, urticaria, and erythematous rash. ethyl (green ) and two keto groups (red ), the latter two par- The overall incidence of anaphylaxis induced by tially obscured in (a ), allows accessibility to the ring nitro- propofol in France is about 1 % and 0.65 % in the gens (dark blue ). The nitrogens are clearly visible in (a ), but the view from the opposite side of the molecule shows Australian survey (Table 7.1 ). Another survey that accessibility to the nitrogens is impeded by the keto estimated that 1.2 % of cases of perioperative groups. In (a ), the nitrogens show the attached hydrogens anaphylactic shock were attributable to propofol. which are absent in ( b ). The large sulfur atom (yellow ) is Risk factors for a reaction are said to be a history freely accessible from both sides of the molecule of previous drug allergy and the use with atracurium, the latter because of possibly enhanced his- relatively small and too little to affect the circu- tamine release. latory system. Thiopentone-induced histamine The picture of propofol’s involvement in release from leukocytes has been studied in evoking anaphylaxis is confusing with some vitro by a number of groups using leukocytes aspects of the suggested underlying mechanism from subjects who reacted to the drug and to open to doubt. In the study of the 14 patients with barbiturate analogs for cross-reactivity studies. life-threatening reactions, positive intradermal In one study, pentobarbitone also produced his- skin tests were obtained in eight patients at con- tamine release, but methohexitone did not. In centrations of 10 μg/ml propofol or less, and keeping with the in vivo ﬁ ndings, thiopentone because a concentration of 100 μg/ml had been induced only a moderate increase in histamine shown to be negative in 100 patients who showed from peripheral blood leukocytes taken from a no adverse reaction to the drug, the eight positive

[email protected] 7.5 Anaphylaxis to Hypnotic Drugs Used in Anesthesia 277 tests were regarded as an indication of the pres- CH3 OH CH3 ence of specifi c IgE antibodies. In an attempt to detect IgE antibodies to propfol, Phenyl- H3C CH3 Sepharose® was used as a solid support to bind the drug to the support by hydrophobic interaction. After incubation of the propofol-Sepharose-solid Fig. 7.20 Chemical structure of propofol. The aromatic phase with patients’ sera, uptakes of IgE anti- ring nucleus and two isopropyl groups give the molecule body were indeed observed, but background its hydrophobic properties binding levels and binding of NMBD-reactive IgE in sera from some subjects were also alarm- ingly high, suggesting that some nonspecifi c groups on opposite sides of the six-membered binding might be involved. Signifi cantly, no ionic ring structure (Fig. 7.20 ). It has been speculated detergent was included in the washing proce- that the isopropyl groups confer allergenic diva- dures during the assay and no steps (other than lency on the molecule, and although this has been the addition of 1 M sodium chloride) specifi cally repeated many times in the literature, there is as directed at minimizing nonspecifi c binding yet no convincing evidence that it is so. appear to have been undertaken. A direct and convincing way to establish specifi city of a drug– 7.5.2.4 Propofol and Allergies to Egg antibody reaction is by inhibition experiments, and Soybean Oil but this approach was precluded by propofol’s In 1994 an allergic reaction to propofol was insolubility in aqueous media. In interpreting the reported in an egg-allergic patient. This led to the results of the study, it was concluded that one suggestion that egg allergy might be a possible positive test was suffi cient to indicate an IgE- risk to consider prior to propofol administration. dependent mechanism and, on this basis, 13 of Following the hospitalization for treatment of the 14 patients had therefore experienced a true respiratory symptoms of a 14-month-old boy anaphylactic type I allergic reaction. Some years with a history of airways disease and allergies to after this study, the same investigators announced egg, peanut, and molds, propofol was adminis- that the solid phase drug-Phenyl-Sepharose IgE tered for sedation. The condition progressed to immunoassay for propofol “has to be reconsid- anaphylaxis with the patient becoming hypoten- ered as non-specifi c binding of hydrophobic sive and tachycardic. Because the propofol for- drugs.” It was also said that propofol and NMBDs mulation contains both soybean oil and egg may potentiate mediator release by a “non-eluci- phospholipid, it was concluded that, if possible, dated mechanism,” but what that mechanism propofol should be avoided in patients with aller- might be remains obscure. gies to egg and/or soybean oil. In a recent retrospective review of cases for the period 1999–2010 7.5.2.2 Skin Tests of children with egg and/or soy allergy, 28 egg- Prick testing with propofol is said to be unreliable allergic subjects with a total of 43 propofol and intradermal testing is recommended. Prick administrations were identifi ed. No soy-allergic testing is performed with the undiluted propofol children were found. One non-anaphylactic formulation (10 mg/ml). For intradermal testing, immediate reaction 15 min after propofol admin- the maximum concentration is a 1 in 10 dilution, istration was identifi ed in a 7-year-old boy with a that is, propofol 1 mg/ml. Testing should start at history of egg anaphylaxis and multiple other 1 in 1,000 and proceed up to the 1 in 10 dilution. IgE-mediated food allergies. A skin prick test to propofol (performed within 12 months of the 7.5.2.3 Chemical Structure reaction) was positive with a wheal of 3 mm. It and Allergenicity was concluded, “Propofol is likely to be safe in The hydrophobicity of propofol is due to the aro- the majority of egg-allergic children who do not matic phenyl nucleus and the two isopropyl have a history of egg anaphylaxis.” The discovery

[email protected] 278 7 Drugs and Other Agents Used in Anesthesia and Surgery of the single patient with egg allergy who reacted 7.6.1.1 Chemistry to propofol more or less guarantees that egg- HES, a nonionic preparation, is synthesized allergic sensitivity will continue to be regarded as from amylopectin and is made up of a backbone a risk factor that cannot be ignored when propo- of d -glucose units linked α-(1-4) and with fol is administered. Rather than singling out egg, branches of d -glucose units attached by α-(1-6) some have suggested that patients with multiple glycosidic bonds. Hydroxyethyl groups are food allergies including soy, and a history of attached at glucose carbons 2, 3, and 6 (when eczema and asthma, might increase the risk of free). The attached hydroxyl groups retard allergy to propofol. hydrolysis of the polysaccharide by amylase, In Asia allergies to soy are well known and an thereby delaying its elimination from the circula- association of soybean allergy with propofol- tion. The degree of branching of a HES polymer induced anaphylaxis has been suggested. In a refers to the ratio of α-(1-6) branches to recent convincing case, anaphylaxis with severe d -glu cose units, and the degree of substitution, oropharyngeal edema and bronchospasm generally expressed as a number between 0 and occurred in a 74-year-old woman with a history 1, gives the fraction of d -glucose units bearing a of soy allergy a few minutes after receiving hydroxyl group. For example, a degree of propofol. Skin prick tests revealed positive reac- branching of ratio 1:20 signifi es one α-(1-6) tions to propofol and 20 % Intralipid® which, like branch for every 20 d -glucose units; a degree of the propofol formulation, contains soybean oil. substitution of 0.7 indicates seven hydroxyethyl It was concluded that the anaphylactic reaction groups for every 10 d -glucose units. HES can be was caused by the soybean oil present in the monodisperse, that is, molecules of one molecu- administered propofol. lar weight only, or polydisperse where a range of molecular weights make up the colloid. For polydisperse colloids, molecular weight can be 7.6 Anaphylaxis to Colloids expressed as the weight average molecular weight Mw or the number average molecular The risks associated with blood transfusions and weight Mn. These give different number values, the shortage of available blood have led to for example, pentastarch range 10–1,000 kDa, increased administration of substitute solutions Mw 280, is equivalent to Mn 120. for volume expansion. Colloids are commonly used and three of the main colloids commonly 7.6.1.2 Risk and Adverse Reactions administered, gelatin, hydroxyethyl starch, and HES is well tolerated with an incidence of dextran will be discussed. Each one may cause adverse events less than gelatin and dextran. adverse reactions ranging from grade I with skin One report estimated the incidence of allergic manifestations to grade IV involving cardiac and/ reactions to be 0.0004 % while fi gures for the or respiratory arrest. risk of life-threatening reactions are in the region of 0.006–0.085 % (1 in 1,172). For comparison, the risk of anaphylactoid reactions to 7.6.1 Hydroxyethyl Starch albumin is 0.011 %. Clinical manifestations of reactions include anaphylactic and anaphylac- Hydroxyethyl starch (HES), also known as hetas- toid shock, erythema, urticaria, and pruritus, the tarch, is most often administered for intravascular latter having an incidence of 10–40 %. In a pro- volume expansion during the perioperative period spective, randomized, controlled study, HES and for resuscitation from trauma and shock. 6 % (200/0.5) and Ringer lactate solution were Because of its property of increasing blood fl uid- compared for the induction of anaphylactoid ity, hydroxyethyl starch is sometimes infused to reactions and pruritus. Results showed no dif- treat patients with disturbed microcirculation. ferences although there was an incidence of

[email protected] 7.6 Anaphylaxis to Colloids 279 more than 10 % for pruritus in both groups. 7.6.2 Gelatin Pruritus, which may persist for up to 1–2 years, is generally refractory to usual treatments and Gelatin is a protein obtained by hydrolysis of is increasingly being recognized as a common, animal (usually cow and pig) collagen from skin, frequently severe, and protracted adverse effect bone, and connective tissue. Allergic reactions to of HES administration. It has been claimed that gelatin have been reported after eating fl avored all currently available HES solutions of diverse fruit gums and condiments, following injection molecular weights and substitutions are subject of vaccines containing gelatin as a heat stabilizer to the risk of pruritus, thought to result from the and after infusion of plasma expanders contain- deposition of HES in tissues particularly macro- ing the protein. The overall frequency of ana- phages. Because of the relative infrequency of phylactoid reactions (grades I–IV) to colloid reactions to HES and the usual diffi culties of intravascular infusions containing gelatin was identifying an allergic reaction and the caus- estimated to be 0.115 % and for severe reactions ative agent during the perioperative period, it (grades III and IV) 0.038 %. A more recent esti- has been pointed out that HES may be over- mate for severe reactions gave a fi gure of 0.345 % looked as a cause. There are some reports in the for gelatin. Risk factors for allergy to gelatin literature that bear that out. include allergy following ingestion of the protein and drug allergy in general. Male gender has also 7.6.1.3 Reaction Mechanisms been mentioned. and Diagnosis As a blood volume expander, gelatin has The possibilities of antibody formation by been or is marketed as Haemaccel® and patients given HES and involvement of antibod- Gelofusine® . Both are derived from bovine ies in adverse reactions to the agent have been spongiform encephalitis-free bovine herds in investigated in a small number of studies. In one the USA, Haemaccel is cross-linked with urea investigation of just over 1,000 patients 14 days and has a molecular weight of about 35,000 Da, after HES administration, antibodies of the IgM and Gelofusine is succinate-linked with a mean class were found in only one patient, and despite molecular weight of about 30,000 Da. Each has repeated HES infusions, no clinical reaction been associated with reports of allergic reac- eventuated. The investigators concluded that tions and at least one study has demonstrated antibodies to HES are extremely rare and they do allergic cross-reactivity between the two prep- not necessarily provoke an anaphylactic response. arations. Clinical manifestations of reactions In a similar study of 1,056 patients, the investiga- to gelatin include anaphylaxis, sneezing, bron- tors concluded that preformed antibodies to HES chospasm, and urticaria. The mechanism of do not exist in humans or are extremely rare. A allergic reactions to gelatin colloids has been claim to have detected anti-HES IgE antibodies said to be IgE antibody and non-IgE antibody was made following retrospective testing of sera mediated and diagnosis has been based on skin from an anaphylactic episode. Details of the testing which, for intradermal testing at least, patient’s exposure and specifi city of the antibody carries its own risk of anaphylaxis. Recently, are lacking. Pentastarch-specifi c antibodies, pre- the BAT has been successfully applied to con- sumably IgE, that bound to and activated baso- fi rm anaphylaxis and, together with positive phils in the BAT were found in the serum of a skin tests, the involvement of gelatin-reactive woman investigated for a severe anaphylactic/ IgE antibodies. A Phadia ImmunoCAP ® test for anaphylactoid episode. the detection of IgE antibodies to gelatin is For skin testing, solutions are used undiluted also available. Direct histamine release by gela- for the prick test and at a dilution of from 1 in 100 tin has been proposed to explain some reactions to undiluted for intradermal testing. to the protein.

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For skin test diagnosis, gelatin solutions are A well-known side effect since the mid–late used undiluted (for Gelofusine, Haemaccel, and 1940s is dextran-induced anaphylactic reactions gelatin solutions ~35 mg/ml) in the skin prick test (DIAR) that can range in severity from mild ery- and at dilutions of from 1 in 1,000 to 1 in 10 for thema (grade I) to death (grade V). DIARs are intradermal tests. caused by the formation of immune complexes formed by reaction of preexisting circulating antibodies to dextran, primarily of the IgG class, 7.6.3 Dextrans with the injected dextran. In grade III and IV reactions at least, the immune complexes formed Dextrans are polysaccharides of varying chain from the cross-linking of dextran by the dextran- lengths synthesized from sucrose by lactic acid specifi c IgG antibodies bind to receptors on the bacteria Leuconostoc mesenteroides and cell surfaces of mast cells and basophils leading Streptococcus mutans . Dextran chains are com- to the release of mediators that cause circulatory posed of d -glucose units linked α-(1-6) with collapse and bronchoconstriction. In the fi eld of branches linked α-(1-3). Two intravenous solu- quantitative immunochemistry dating back to the tions containing the high molecular weight dex- foundations of the modern science of immunol- trans 40 and 70 are used for plasma volume ogy, dextran has its own special place as a model expansion in the treatment of hypovolemic shock, antigen in the study of antibody–carbohydrate for postoperative thromboembolic prophylaxis, as reactions and for its central role in experiments a component of the pump prime for cardiopulmo- that defi ned the size of the antibody combining nary bypass, and to promote blood fl ow in the site. By the early 1950s, a project group led by microcirculation. In a large-scale multicenter Elvin Kabat in New York and carried out with study involving nearly 201,000 infusions of intra- the backing of the Offi ce of the Surgeon General, vascular colloid volume substitutes, the overall US Army, had shown that signifi cant correla- frequency of anaphylactoid reactions (grades I– tions existed between: (1) skin test sensitivity IV) for dextrans was 0.032 %, while for severe and allergic reactions to some dextrans; (2) the reactions (grades III and IV) the fi gure was presence of antibodies to some dextrans and sys- 0.008 %. In a more recent prospective screen of temic allergic reactions; and (3) the structural nearly 20,000 patients, the incidence of anaphy- complexity of some dextrans and the incidence lactoid reactions to dextrans was 0.273 %. Reports of positive skin tests in the general population. It to the FDA from 1969 to 2004 of adverse events was further demonstrated that partial hydrolysis involving dextrans numbered 366, one quarter of of native dextrans produced a reduction in their which was anaphylactic/anaphylactoid in nature. capacity to produce a positive wheal and fl are Anti-dextran IgG antibodies cross the placenta so reaction and that a low proportion of non-1-6 dextran administration should normally be avoided linkages and elimination of higher molecular in pregnant women. Cases of neurological impair- weight fractions produced a low incidence of ment and deaths in neonates following anti-dex- systemic allergic reactions. By the 1970s, accu- tran administration have been reported. Identifi ed mulated knowledge of the humoral recognition risk factors are few with atopy linked to milder of dextran polysaccharides and oligosaccharides reactions and high levels of anti-dextran antibod- of varying sizes laid the groundwork for an ies associated with severe reactions. Clinical signs animal model of DIAR that showed that the and symptoms range from anaphylactic shock, hapten dextran 1 (molecular weight 1,000 Da), fever, and death to nausea and minor fl ushing. administered immediately before dextran 40, Cutaneous symptoms include urticaria, pruritus, greatly reduced the incidence of severe hypoten- angioedema, and macular rash. Respiratory mani- sion. The mechanism of this effect is due to the festations are wheezing, chest tightness, coughing, monovalent hapten nature of dextran 1 that inter- dyspnea, and pulmonary edema. acts with anti-dextran antibody combining sites

[email protected] 7.7 Local Anesthetics 281 without forming an antigen–antibody lattice often seems that the general public and the medi- complex and therefore no detrimental immune cal profession perceive the situation differently. complexes. This can be viewed as competitive The public’s perception is that local anesthetics hapten inhibition carried out in vivo. are a frequent cause of reactions while allergists Subsequently, large multicenter clinical trials at least know that although these agents are well found that pre-injection of 20 ml dextran 1 down any list of drugs causing true allergic reac- reduced the incidence of severe DIAR from 25 tions, immediate or delayed, referrals for local cases per 100,000 units dextran 40/70 to three anesthetic allergy are often as high as for known cases per 100,000 units. A review of the 10-year allergenic drugs such as the penicillins. period 1983–1992 revealed that the prophylactic use of dextran 1as hapten was associated with a 35-fold reduction in the incidence of severe 7.7.1 Chemistry DIAR and a 90-fold reduction of lethal DIAR. It has been claimed that the introduction of All local anesthetics possess similar structural fea- dextran 1 for hapten inhibition prior to infusion tures of a hydrophobic aromatic ring linked to a of dextrans 40 and 70 has made these plasma hydrophilic amine group and the linkage group volume expanders the safest of all the volume may be an ester, amide, ether, or ketone. In clinical expanders in clinical use. practice today, the esters and amides predominate Adverse reactions to dextran 1 were investi- with the latter group by far the most widely used. gated in trials involving over 70,000 patients. Well-known esters are cocaine, benzocaine, pro- The following reactions and incidences were caine, and tetracaine and amides include lidocaine recorded: cutaneous reactions 0.016 %; moderate (lignocaine), prilocaine, etidocaine, mepivacaine, hypotension 0.014 %; severe hypotension bupivacaine, ropivacaine, and articaine (Fig. 7.21 ). 0.001 %; bradycardia and moderate hypotension As with the sulfonamide drugs, all of the local 0.013 %; bradycardia and severe hypotension anesthetic esters are derivatives of p -aminoben- 0.001 %; bradycardia alone 0.004 %; and mild zoic acid (see Sect. 6.2.1.1 ), and this compound is symptoms (nausea, pallor, shivering) 0.011 %. released upon hydolysis. Prilocaine, etidocaine, For skin test diagnostic investigation, dextran mepivacaine, bupivacaine, and ropivacaine exist (6–10 mg/ml) is used undiluted in prick tests and as stereoisomers (R-(+) and S-(−) isomers) and at a maximum concentration of a 1 in 100 dilu- most are marketed as racemic mixtures, although tion of the prick test concentration for intrader- ropivacaine is used as the S-isomer and mal testing. S-bupivacaine is less toxic than R-bupivacaine.

7.7 Local Anesthetics 7.7.2 Adverse Reactions to Local Anesthetics Local anesthetics are the key components that enable the application of local and regional anes- Following the ﬁ rst description of “allergy” to a thetic techniques for the treatment of acute pain local anesthetic over 90 years ago, there was ini- and the drugs are also used in the management of tially a steady stream of reports of reactions to chronic pain where they may have a prolonged the drugs consisting mainly of erythema or effect. It can be said that local anesthetics revolu- edema. With the introduction of the amide local tionized the practice of anesthesia, surgery, den- anesthetics, the number of hypersensitivity reac- tistry, and ophthalmology and with their daily tions tapered off signiﬁ cantly, indicating that routine use in countless minor procedures and ester compounds were less well tolerated. Even their applications in obstetrics they are known to today, however, reports of adverse reactions to affect almost everyone at some time in their life. local anesthetics occasionally appear, but the With regard to “allergy” to local anesthetics, it nature of the reactions cannot always be described

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ESTERS: O O

N OH O

H2N N p-Aminobenzoic acid Aromatic Ester Amine ring group group

ON O

N O O

OO H2N Cocaine Procaine

AMIDES:

O O

N N N N H H Lignocaine O (Lidocaine) S N N H O Aromatic Amide Amine ring group group N N H

O H Mepivacaine S N N H

O COCH3 Articaine

Fig. 7.21 Chemical structures of two main categories of local anesthetics, esters and amides, with some structures of important members of each category as hypersensitivity responses. In fact, many of Apart from the presence of epinephrine, local the reactions, if not the majority, may be vasova- anesthetics may evoke sympathetic effects that gal reactions, anxiety attacks, and some may be include palpitations, light-headedness, syncope, reactions to epinephrine which is frequently or tachycardia. Local anesthetics are heavily used added to local anesthetics as a vasoconstricting in dental procedures and many reports of adverse agent for dental and some minor procedures. reactions following their administration emanate

[email protected] 7.7 Local Anesthetics 283 from that source. A prospective study of just over toms occurred within 1 h of administration and 5,000 patients who received local anesthetics included respiratory, cardiovascular, and neuro- during dental treatment revealed only 25 adverse logical signs. It was concluded that both clinical reactions (~0.5 %) with none of the reactions history and skin testing are necessary to confi rm allergic. Twenty-two of the 25 reactions were an immediate reaction and, in fact, the latter test mild, vasovagal in nature, or what was described is considered mandatory. Five delayed-type reac- as quickly reversible psychogenic reactions. tions with mainly cutaneous signs of erythema, Other additives to formulations of local anesthet- pruritus, urticaria, and eczema appearing 6 h to 1 ics such as parahydroxybenzoates (parabens) and month after local anesthetic administration were the antioxidant sodium metabisulfi te have come also identifi ed in the French study. Type IV under suspicion as causative agents for adverse hypersensitivity reactions particularly to ester- reactions. The former compounds (which are no type local anesthetics are well known and the longer added to many formulations) are known to involvement of amides in delayed reactions is cause contact sensitivities but, together with also recognized. Of the six cross-reactivity cases sodium metabisulfi te, evidence of involvement in mentioned above, four were type I reactions, and immediate reactions is lacking. The incidence of although cross- reactivity between ester local what has been called “systemic toxicity” to local anesthetics is well known, this was not the case anesthetics is said to have decreased from 0.3 to for the amides. Lack of cross-reactivity between 0.01 % over the past 30 years. articaine and the other amides is probably due to the presence of a thiophene and not a phenyl ring 7.7.2.1 Immediate Reactions in the former drug (Fig. 7.21 ). True type I immediate reactions to local anesthet- The rarity of immediate reactions to local ics, especially the amides, are rare, but they have anesthetics is further emphasized in results been documented even for the amides. A number obtained in two large studies of a total of 354 of reactions, well short of 1 % of reported allergic patients with a history of reacting to the drugs. No reactions to local anesthetics, are thought to be evidence for an IgE-mediated reaction was found immune mediated and, of these, amide com- in one of the studies involving 157 patients while pounds make up a small proportion. In perhaps three patients, two with an immediate reaction the most comprehensive review of data related to and one with a delayed response, were seen in the local anesthetic allergy undertaken so far, reports review of the other 197 patients. No IgE antibod- in the French Pharmacovigilance and the Groupe ies were detected in the two immediate reactors. d’Etudes des Reactions Anaphylactiques Peranesthesiques (GERAP) databases for the 7.7.2.2 Delayed Reactions 12-year period 1995–2006 were analyzed for Lidocaine-specifi c T cell lines and clones gener- clinical features, skin test results, delayed- and ated from patients with contact dermatitis to the immediate-type allergic reactions, and cross- drug proved to be mainly MHS class II restricted reactions. Of 16 relevant cases identifi ed, an and CD4+, but some were MHC class I restricted immediate reaction occurred in 11 patients, lido- CD8+ clones. The T cell lines cross-reacted with caine was the drug most involved (11/16), and mepivacaine but not with procaine, oxyprocaine, cross-reactivity between the amides lidocaine bupivacaine, and tetracaine. Most of the CD4+ and mepivacaine was found in six cases. clones produced a Th2-like pattern of cytokines Reactions occurred mostly in young females with a high IL-5 component. Cellular recognition (F:M ratio 14:2) and diagnoses were confi rmed studies with the rather chemically nonreactive with prick tests, intradermal tests, and challenges. lidocaine showed that it was recognized directly The fi nding that lidocaine was the drug most by αβ+ T cells in an HLA-DT restricted manner often involved in immediate reactions is consis- with neither drug metabolites nor breakdown tent with the fact that it is the most often used products involved or protein processing required. local anesthetic in medical practice. First symp- Examination of the cross-reactivity between

[email protected] 284 7 Drugs and Other Agents Used in Anesthesia and Surgery lidocaine and mepivacaine at the cloned cell level of only 43 % and 14 %, respectively, for the showed cross-recognition with small structural sensitivity of intradermal and prick tests. In addi- changes affecting T cell stimulation. For exam- tion to its “gold standard” status as a diagnostic ple, one clone recognized lidocaine and mepiva- test, provocation testing for the investigation of caine but no hydroxy metabolite, others adverse reactions to local anesthetics has been responded to 3-hydroxy metabolites, and only described as safe and well tolerated. some amines and a broadly reactive clone toler- IgE antibodies to local anesthetics appear to ated hydroxy substituents and reacted to linear be so rare that some have even doubted that they and cyclic amines. Findings with the broadly occur, but occasional case reports indicate an reactive clone suggested that the structure of the underlying mechanism of type I hypersensitivity amine side chain of local anesthetics is essential and some skin test results suggest that these anti- for recognition by the T cell receptor. bodies do exist. Evidence from a dot-blot procedure for an IgE-mediated reaction to lignocaine has been presented, and more recently, IgE 7.7.3 Diagnosis of Reactions antibodies to mepivacaine were reliably detected to Local Anesthetics by a Phadia immunoassay shown by inhibition studies to be specifi c. For skin testing, solutions of local anesthetics are used undiluted in the prick test and either at or up to a dilution of 1:10 in the intradermal test. A few 7.8 Polypeptides positive intradermal tests have been reported with dilutions of 1:100 down to 1:10,000. 7.8.1 Protamine Examples of some undiluted starting concentrations are lidocaine 10 mg/ml, bupicacaine 2.5 mg/ Protamines are arginine-rich nuclear proteins with ml, mepivacaine 10 mg/ml, and ropivacaine molecular weights in the range 4,500–5,000 Da. 2 mg/ml. Although it has been stated that skin Generally purifi ed from sperm heads in salmon testing in the diagnosis of hypersensitivity to milt and used as the sulfate, protamines are used to local anesthetics is mandatory, it should be said reverse heparin’s anticoagulant effect via binding that there is no universal agreement on the value of its basic guanidine groups in arginine to acidic and place of skin testing in the assessment of heparin molecules and to retard the absorption of patients’ adverse reactions to these drugs. A insulin (as neutral protamine Hagedorn, NPH) and recent review of the literature over the period increase its duration of action. A study nearly 40 1978–2009, focusing on sensitivity and specifi c- years ago on the cardiorespiratory effects of prot- ity of skin testing and provocation challenge, amine after cardiopulmonary bypass surgery in found only three immediate reactors in 1,094 humans showed that the protein at a concentration patients when the local anesthetic was repre- of 6 mg/kg produced a marked fall in cardiac out- sented. None of these three reactors were origi- put and a brief fall in systemic arterial pressure. As nally detected by skin prick or intradermal tests. a result of its use during cardiac catheterization Over all the studies, false-positive skin tests var- and cardiopulmonary bypass in very large num- ied from 0 to 27 %. From such results, some have bers of patients, its interaction with heparin, and concluded that skin tests are a poor predictor of the occasional reactions it provokes, protamine’s positive challenge since most adverse reactions adverse effects have received a good deal of atten- to local anesthetics are not allergic in nature and tion. From numerous clinical and experimental skin tests are sometimes positive in patients who investigations it is known that protamine induces tolerate challenge with the suspected drug. For increased pulmonary artery pressures and falls in patients confi rmed by positive challenge, the blood pressure, heart rate, cardiac output, myocar- specifi city and sensitivity of skin tests have also dial oxygen consumption, and systemic vascular been questioned with claims (from small numbers) resistance.

[email protected] 7.8 Polypeptides 285

7.8.1.1 Adverse Reactions 7.8.1.2 Immune-Mediated to Protamine Hypersensitivities When administered too quickly, protamine may to Protamine cause transient fl ushing, a feeling of warmth, bra- In what may be the fi rst evidence for an immune- dycardia, and hypotension and this has resulted in mediated hypersensitivity reaction to protamine, the practice of injecting the drug slowly, usually data were presented in 1978 for an anaphylactic over a 5–10 min period. Adverse reactions to prot- reaction mediated by complement-dependent amine can manifest as fl ushing, rash, urticaria, skin-sensitizing IgG antibodies to the protein. angioedema, wheezing, hypotension, broncho- Soon after, descriptions of clinical manifesta- spasm, cardiovascular collapse, and sometimes tions, temporal details, positive intradermal tests death. Until the late 1970s, reactions to protamine to protamine, known previous exposure to the were generally considered to be non-immunologic agent, and immediate responses to IV adrenaline in nature and probably due to direct effects includ- led to a diagnosis of type I anaphylactic responses ing mast cell degranulation. Experiments with in three patients following IV administration of human basophils and lung mast cells have not protamine sulfate. Published incidences of the always clearly demonstrated histamine release by risk of immediate adverse reactions to protamine protamine or protamine–heparin complexes, but sulfate during cardiopulmonary bypass surgery Marone’s group in Naples showed that protamine vary from 0.06 to 10.7 %. For reactions regarded released the preformed mediators histamine and as “clinically signifi cant” rather than “immedi- tryptase from human basophils but did not stimu- ate,” reported incidences range from 0.1 to 24 % late de novo synthesis of eicosanoids. Protamine and the incidence of systemic hypotension fol- also caused the release of histamine and tryptase lowing protamine administration is said to be from human heart mast cells and, to a lesser extent, 1.76–2.88 %. A signifi cantly higher incidence of from synovial mast cells but not lung mast cells. anaphylaxis to protamine in insulin-dependent As for basophils, protamine did not induce de diabetics than in patients not receiving insulin novo synthesis of LTC4 and PDG2 from lung and suggested sensitization by protamine in NPH- skin mast cells. Fulminating non-cardiogenic pul- insulin preparations. In one early review of 1,150 monary edema, also referred to as adult respiratory patients given protamine, anaphylactoid reac- distress syndrome or ARDS, has been described as tions occurred in 3 % of diabetic patients but in a rare event occurring in 0.2 % of cardiopulmo- only 0.2 % of nondiabetics. Other assessments of nary bypass patients with mortality rates approach- the incidence of anaphylaxis after protamine ing 30 %. Protamine-induced direct release of reversal of heparin in patients on protamine- histamine and/or complement activation is the sus- insulin therapy range from 0.6 to 27 %. To deter- pected mechanism. What has been termed prot- mine whether anti-protamine IgG and/or IgE amine-induced pulmonary artery vasoconstriction antibodies mediated the reactions in protamine- with cardiovascular collapse after cardiac surgery insulin-dependent diabetics, diabetics and non- and thought to involve the potent vasoconstrictor diabetics who reacted to IV protamine and thromboxane A2 is reported to have an incidence diabetics who tolerated protamine during surgery of 1.5 %. This may be mediated by complement were studied. Anti-protamine IgE and IgG anti- activation resulting from the interaction of prot- bodies were judged to be signifi cant risks for amine with heparin or anti-protamine IgG and acute protamine reactions in diabetics who had C5-mediated thromboxane generation. In vitro received protamine-insulin. IgE antibodies were experimental results demonstrating inhibition of not found in any patient without previous expo- carboxypeptidase N (kininase I), the inactivator of sure to protamine-insulin. In nondiabetic patients, anaphylatoxin and kinin mediators released in the presence of anti-protamine IgG antibodies shock reactions, have been advanced as another was shown to be a signifi cant risk, but about 30 % possible mechanism that may contribute to the of patients who reacted had neither anti- spectrum of adverse reactions to protamine. protamine IgG or IgE antibodies. In a follow-up

[email protected] 286 7 Drugs and Other Agents Used in Anesthesia and Surgery study of a single patient who had a life- threatening Intradermal tests on 32 patients with protamine reaction to IV protamine, serum IgE and IgG 1 μg/ml were positive in four (13 %) despite the antibody levels showed a twofold and 70-fold absence of a clinical reaction in all of the patients. rise, respectively, 1 month after the reaction. This 87 % specifi city for protamine was almost Intradermal skin tests with protamine sulfate did the same as the 91 % specifi city obtained with not discriminate between the test subject and intradermal saline. At a concentration of 10 μg/ nine normal subjects who had no previous expo- ml, intradermal protamine injections were posi- sure to protamine and no protamine-reactive anti- tive in ten (31 %) of the patients. Tests for bodies. In vitro challenge of basophils with protamine-reactive IgE and IgG serum antibodies protamine in histamine release experiments were positive in 46–54 % and 100 % of patients, proved inconclusive. While it was clear that respectively. These high incidences of false- subcutaneous injection of protamine in insulin positive reactions demonstrated the unsuitability preparations induced protamine-specifi c anti- of skin and antibody tests for screening patients body responses, it remained important to know before administration of protamine. whether single-dose IV protamine administration could provoke an antibody response. IgE and IgG antibodies were found in 18 and 16 %, respec- 7.8.2 Aprotinin tively, of previously sero-negative patients 4–6 weeks after a single IV dose of protamine. Aprotinin, a protease inhibitor isolated from Appearance of antibodies was associated with bovine lung, is a single chain polypeptide of 58 insulin-dependent diabetes and male gender. amino acid residues, molecular weight 6,512 Da. Other possible risk factors for protamine sensitiv- Marketed as Trasylol (Bayer) and Antagosan ity include allergy to fi sh (protamine is obtained (Aventis Pharma), aprotinin inhibits fi brinolysis, from fi sh testes), men who have undergone reduces thrombin generation, and maintains vasectomy, and infertile men. Evidence for each platelet function, properties that explain its pro- of these is not compelling. Fish muscle and skin, phylactic intravenous use in cardiac surgery, not testes, are usually consumed, and in the latter organ transplantations, and hip surgery where two cases, antibodies to sperm and hence prot- reductions in bleeding, blood loss, and transfu- amine have been suggested to be increased risks, sion needs are important. It is also thought that but clear demonstrations of associations have not aprotinin may help to reduce infl ammatory reac- been forthcoming so far. tions after cardiopulmonary bypass surgery by interacting with enzymes activated during the 7.8.1.3 Diagnostic Tests surgery. In addition, the polypeptide’s antifi bri- Diagnostic tests for hypersensitivity to protamine nolytic action has led to its topical application in are problematic. Protamine skin tests have been ready-to-use tissue sealants or so-called fi brin shown to have poor specifi city with irritant glues to maintain hemostasis. Given its protein responses in normal controls resulting from intra- nature and bovine origin, aprotinin as a foreign dermal injections of concentrations of 100– protein can induce an immune response in 1,000 μg/ml. The recommended concentration humans with the formation of specifi c antibodies, for intradermal testing is 1 μg/ml, although this both IgG and IgE, and occasionally anaphylaxis. concentration, and a concentration of 10 μg/ml, Previous exposure is considered to be the major has been found to give false-positive results. The risk factor for a severe adverse reaction to the uncertainty associated with the specifi cities of polypeptide, with most reactions generally tests for protamine were highlighted in a pro- occurring within some months of initial or previ- spective study of patients undergoing elective ous administration. Other risk factors are said to cardiac surgery and subjected to skin and serum be the consumption of beef, milk in its different antibody tests for protamine reactivity. forms, and egg albumin.

[email protected] 7.8 Polypeptides 287

7.8.2.1 Adverse Reactions adult serum samples from cardiosurgical patients to Aprotinin for aprotinin-reactive IgG antibodies revealed According to a 2007 analysis of aprotinin- that of 22 positive sera (4 %), only three were induced anaphylaxis in over 12,000 patients from patients with documented aprotinin preex- exposed to the drug in cardiac surgery, the inci- posure. Only one of the 22 positive sera was also dences of hypersensitivity reactions were 4.1 %, positive for IgE antibodies to aprotinin. This 1.9 %, and 0.4 % in less than 6 months, 6–12 patient experienced anaphylaxis and also had months, and more than 12 months reexposure recent IV preexposure to the drug. The investiga- intervals, respectively. In addition to anaphylaxis, tors concluded that the clinical signifi cance of clinical manifestations of hypersensitivity IgG antibodies to aprotinin is questionable and include bronchospasm and cutaneous symptoms the presence of the antibodies is not a reliable of pruritus, urticaria, and exanthema. Of 124 prediction of previous exposure. What makes the cases of aprotinin-induced anaphylaxis with 11 interpretation of the signifi cance of serum anti- deaths identifi ed in the period 1963–2003, the bodies to aprotinin even more diffi cult is the pres- risk of anaphylaxis in previously exposed patients ence of IgG and IgE antibodies in 55 % of patients was ~2.8 %. The reexposure interval was <3 with an allergic reaction and 32 % of non- months in 72 % (38) of the 53 affected patients. reactors. In addition, half the patients still have An adverse reaction incidence of 2.8 % (seven serum IgG to aprotinin 4 years after receiving the patients) was also found in an analysis of 248 drug. In a study of anaphylaxis after reexposure reexposures to aprotinin over the period 1988– to aprotinin during cardiac surgery, the incidence 1995. Five of the seven patients reacted within 3 of anaphylaxis was found to be 2.5 % (3 of 121 months of reexposure and two reacted to a load- patients) but detected IgG and IgE anti-aprotinin ing dose following a test dose. The same outcome antibodies were not always clinically relevant. was seen in two patients who reacted within All three anaphylactic patients had high levels of 5 min of the loading dose given after a test dose aprotinin-reactive IgG and two had high levels of even though the patients had been pretreated with IgE antibodies. According to the authors, a com- corticosteroids and antihistamines. The safety of parison of the propensity of a patient to react the use of aprotinin, and its reuse, in pediatric adversely, and the length of the aprotinin expo- cardiothoracic surgery was assessed in a retro- sure—reexposure interval, indicated that within a spective review of 681 fi rst exposures, 150 sec- reexposure interval of 6 months, aprotinin should ond exposures, and 34 third or higher exposures be used with caution only in exceptional cases to the agent. The incidences of reactions were such as patients with a high risk of bleeding. found to be low—specifi cally 1 %, 1.3 %, and Aprotinin in fi brin sealants has also been reported 2.9 %, respectively, with reactions no more likely to trigger anaphylaxis. For example in one case, to occur in any of the three categories. Skin test- topical reexposures to fi brin sealants containing ing showed a negative predictive value of 99 % aprotinin did not provoke adverse reactions, but and a positive predictive value of 20 %. Aprotinin- anaphylaxis did result following IV administra- reactive IgE antibodies were not detected in tion of aprotinin during coronary artery bypass seven of eight reactive patients tested, leading to graft despite a negative test dose 5 min before. the conclusion that the test “would not be clini- Postoperative serological tests revealed elevated cally useful.” In view of the small number tested aprotinin-specifi c IgG and IgE antibodies. and the absence of any details of the test method- In 2007, aprotinin was temporarily withdrawn ology, this conclusion should not yet be taken as worldwide until results from the Canadian BART fi nal. However, on the basis of the results of this (Blood conservation using antifi brinolytics: a ran- study, the overall conclusion that the use and domized trial) study conducted in high-risk car- reuse of aprotinin in children in cardiothoracic diac surgery patients was completed and evaluated. surgery is essentially safe and that any reactions Sales of the drug were suspended in May 2008, that do occur can generally be managed success- but this suspension was lifted in Europe by the fully seems sound. Screening of 520 preoperative European Medicines Agency in February 2012.

[email protected] 288 7 Drugs and Other Agents Used in Anesthesia and Surgery

For skin testing, aprotinin can be used at a in part, a refl ection of the draping of patients concentration of 10,000 IU/ml in prick tests and during surgery. In addition, to the well-known up to a maximum of 100 IU/ml intradermally. at-risk groups of healthcare workers and spina bifi da patients, other risk factors include atopy, allergies to fruits, previous exposures such as 7.8.3 Latex repeated insertions of latex catheters and indwell- ing catheters, and patients who have undergone Natural rubber latex, from the Hevea brasiliensis multiple surgical procedures. Pretreatment with tree, is used widely in a vast array of rubber antihistamines and corticosteroids is not always goods including, until recently, many medical effective in preventing anaphylactic reactions to products. Beginning in the 1980s and extending latex so an emphasis should be placed on preven- into the 1990s, the number of reports of allergy to tion. This, according to Phil Lieberman, involves latex, particularly anaphylaxis, increased spec- the introduction of measures and changes in tacularly. Two main reasons for this appear to operating, treatment, and recovery rooms that have been increasing concern about transmissible includes the employment of non-latex or latex- infections like AIDS, leading to a dramatic free gloves (most important), catheters, bandages, increase in the use of rubber gloves and the so- tapes, tubing, bite blocks, breathing system, and called bias of ascertainment , that is, the greater electrocardiogram and pulse oximetry leads. The ease of recognizing a condition once it has been pharmacy should institute latex-free protocols identifi ed and defi ned diagnostically. The inci- and the hospital should set up a latex-free cart dence of allergic sensitivity to latex in the general with non-latex gloves, stethoscope, masks, pres- population is estimated to be about 2.1–3.7 % but sure cuffs, neoprine bags, uncuffed polyvinyl can be much higher in risk groups such as den- chloride endotracheal tubes, Webril™ tourni- tists (up to 15 %) and spina bifi da patients (up to quets, and so on. 64 %). The steady increases in the reports of ana- For the diagnosis of latex allergy, a detailed phylaxis to latex appear to have peaked presum- history, including domestic and occupational his- ably because of widespread increased awareness tory, is an essential and prime requirement. Skin of the problem, more stringent requirements for tests with commercially available standardized glove manufacture, the decrease in numbers of latex extracts can be used and immunoassays for latex surgical products, the setting up of “latex- latex-reactive IgE antibodies are available, for free” medical environments, warning labels, and example, the Phadia’s ImmunoCAP® assay regulatory requirements. Incidences of anaphy- (Thermo Scientifi c). Extracts of latex gloves for laxis to latex during anesthesia and surgery, esti- prick and patch tests are easily prepared by mated to represent about 20 % of all cases, may extracting minced material in physiological now vary greatly as shown by the widely con- saline followed by protein determinations and trasting incidences revealed, for example, by stanardization for IgE antibody binding with a recent surveys in France where the fi gure was 25 pooled standard serum sample from allergic sub- times the Australian incidence (see Sect. 7.2 and jects. Although delayed-type reactions are occa- Table 7.1 ). A comparison of clinical manifesta- sionally seen, most reactions of concern are IgE tions of latex anaphylaxis in 1,158 cases not antibody mediated. H . brasiliensis extracts con- related to anesthesia and surgery with 583 cases tain at least 13 IgE-binding components that react occurring during surgery revealed no instances of with sera from latex-allergic subjects. About half cardiovascular collapse in the former group but of these can be considered major allergens or an incidence of 50 % in the latter group (compare important for cross-reactivities with, for exam- Table 7.2). Respiratory symptoms were more ple, some fruits. An interesting relationship exists evenly distributed, but cutaneous reactions between some allergies to natural rubber latex occurred with much higher frequency (98 %) in and some fruits. The latex components responsi- the non-anesthesia/surgery group. This may be, ble for this cross-recognition include hevein, a

[email protected] 7.9 Heparin 289 wound-induced protein from the rubber tree, and induced thrombocytopenia, rare cases of anaphy- a class 1 chitinase (Hev b 11). The major latex laxis, a few delayed reactions, and some adverse allergens identifi ed and purifi ed so far are Hev b skin reactions making up most of the reports. 1 (rubber elongation factor), Hev b 3 (small rub- Heparin can bind to surface-bound platelet factor ber particle protein), and Hev b 4 (a glucosidase), 4 (PF4) on the platelet surface. IgG antibody for- important for reactions in spina bifi da patients; mation is common after heparin administration Hev b 2 (β-1,3-glucanase) and Hev b 6 (prohev- and some antibodies react with the heparin–PF4 ein/hevein), important in latex-fruit cross-reac- complex on the platelets activating the platelets tions; and major allergens Hev b 5 (an acidic via their Fc pieces and causing the release of mic- protein) and Hev b 13 (a lipolytic esterase). roparticles that promotes thrombin formation. A minimum of 12–14 saccharide units of heparin are required to form the heparin–PF4 complex 7.9 Heparin and elicit antibody formation to the complex. Although this reaction is often classifi ed as a type Heparin is a sulfated glycosaminoglycan poly- II cytotoxic hypersensitivity response, the forma- mer of repeating disaccharides most commonly tion of immune complexes on the platelet surface composed of 2- O -sulfo-α-L -iduronic acid and also suggests a type III mechanism (see also Sect. 2-deoxy-2-sulfamido-α- d -glucopyranosyl-6- 3.6 ). Up to about 50 % of heparin-treated patients sulfate. Disaccharides containing 3-O -sulfated may form antibodies reactive with the heparin– d -glucosamine or d -glucosamine with its free PF4 complex. A few cases of heparin-induced amino group are more rarely found. Preparations anaphylaxis have been seen in hemodialysis can range from unfractionated to low molecular patients. Other recorded immediate reactions weight (LMW) heparins which include certopa- include rhinitis, urticaria, pruritus, and broncho- rin, dalteparine, enoxaparine, nadroparine, tinza- spasm. Given the quite marked antigenicity of parine, and reviparine. The polymers used heparin, the rarity of immediate reactions is medicinally and supplied commercially gener- somewhat surprising. Delayed hypersensitivity ally have molecular weights in the range reactions to LMW heparins are occasionally seen 12–15 kDa. Heparin is stored in the granules of with generalized maculopapular rashes, skin human mast cells and basophils. It is used as an necrosis, baboon syndrome, DRESS, and Lyell’s anticoagulant, preventing but not breaking down syndrome recorded. Cross-reactions between clots by binding to antithrombin III and activat- unfractionated and LMW heparins occur and ing the enzyme which then inactivates thrombin even pentosan polysulfate and danaparoid, both and other proteases involved in clotting, particu- chemically distinct from heparin, may show larly factor Xa. It is often administered to patients some cross-reactivity. In one study of delayed during cardiac surgery including pulmonary heparin allergy, 81 and 45 % of patients showed bypass surgery and for acute coronary syndrome, cross-reactivity to danaparoid and pentosan poly- atrial fi brillation, deep vein thrombosis, and pul- sulfate, respectively. In a skin test study of cross- monary embolism. Heparin is given parenterally, reactivity between unfractionated and LMW usually by infusion, since it is not well absorbed heparins in patients with suspected delayed from the gut and has a half-life of about 1 h hypersensitivity to heparin, 11 of 15 patients (4–5 h for LMW heparins and 1–2 h for unfrac- (73 %) showed cross-reactivity between heparins tionated heparin). and/or danaparoid, six patients (40 %) reacted to LMW heparins only, and nine patients (60 %) reacted to both the unfractionated and LMW 7.9.1 Adverse Reactions to Heparin preparations. Danaparoid was tolerated in six of eight patients and hirudin in all three patients The overall incidence of adverse reactions to tested. Other reported adverse cutaneous reac- heparin has been estimated at 0.2 % with heparin- tions to LMW heparins include skin necrosis due

[email protected] 290 7 Drugs and Other Agents Used in Anesthesia and Surgery to vasculitis (described as a type III Arthus reac- with dilutions of from 1–1,000 to 1–10. For patch tion) and erythematous, well-circumscribed testing, undiluted or 10 % aqueous solutions are lesions without necrosis, usually secondary to a applied for 24 or 48 h with readings at 24 or 48, delayed-type IV reaction. 72, and 96 h and again at 7 days. Hirudin is a naturally occurring 65 amino acid polypeptide anticoagulant from the salivary 7.9.2 Diagnostic Methods glands of the leech used in medicine, Hirudo medicinalis. It is the most potent inhibitor Diagnostic skin testing for immediate reactions is of thrombin and has thrombolytic properties, undertaken with undiluted commercial heparin preventing and dissolving clots and thrombi. preparations as prick test solutions while a starting Desirudin and lepirudin are recombinant forms. dilution of 1–1,000 stepping up to 1–10 is Hirudin is a mixture of isoforms of the protein employed for intradermal tests. With cross- whereas the two recombinant forms are homoge- reactivity and possible alternative drugs in mind, neous preparations. Desirudin differs from hiru- skin tests should include unfractionated and LMW din only by the absence of a sulfate group on heparins, danaparoid, hirudin, enoxaparin, and Tyr-63; lepirudin differs by absence of the same perhaps synthetic pentasaccharides such as sulfate but also by substitution of a leucine for fondaparinux. In one patient with type I hypersen- isoleucine at the N-terminal of hirudin. Being sitivity and a positive skin test to dalteparine, other completely different in chemical structure from LMW heparins were also skin test positive, but heparins, there is no cross-reaction between unfractionated heparins, fondaparinux, and the hirudins and heparins. Figures of 0.015 % and recombinant hirudin lepirudin were skin test nega- 0.16 % have been reported for anaphylaxis on tive and tolerated by the patient. For delayed reac- fi rst and subsequent exposures, respectively. tions, prick testing is undertaken with undiluted Urticaria and angioedema are other reported commercial preparations and for intradermal tests type I reactions. Injection site reactions may a 1–10 dilution can be used. Readings should be occur and eczematous plaque and granuloma- carried out after 30 min, 24, 48, 72, and 96 h or tous delayed hypersensitivity reactions have even longer if a very late response is thought to be been described. Clinical trials showed allergic a possibility. Undiluted and 10 % aqueous solu- reactions to desuridin in 1.6 % of treated patients. tions may be applied for 24 or 48 h as patch tests Since both recombinant hirudins are prepared in and read at 24 or 48, 72, and 96 h and then again genetically modifi ed yeast cells, allergic reac- at 1 week. Patch tests are said to yield a high tions may occur in subjects allergic to yeast. rate of negative results so the very late readings Antibodies have been reported in patients treated (72–96 h and beyond) may prove important. with hirudins so there is potential for immunological cross-reactivity between the three preparations. Fatal anaphylactic/anaphylactoid 7.9.3 Danaparoid and Hirudins reactions have occurred with hirudin therapy. The product information for desirudin (Revasc) Danaparoid is chemically distinct from heparin states that hirudin-specifi c IgE evaluations may and generally shows little or no cross-reactivity not be indicative of sensitivity to desirudin since in heparin-intolerant patients. It is a mixture of the test was not always positive in the presence heparan sulfate, dermatan sulfate, and chondroi- of symptoms. Undiluted commercial prepara- tin sulfate and works by inhibiting factor Xa. tions are employed for prick testing and a dilu- Danaparoid should be included in tests on tion of 1–100 for intradermal tests. Pure and patients who cannot tolerate LMW heparins. 10 % aqueous solutions are used for patch testing Reported reactions to the drug include rash, pru- with application and reading times as for danap- ritus, and reactions (some delayed) at the injec- aroid. Despite the statement on Revasc (above), tion site. Prick tests are undertaken with undiluted the peptide nature of hirudins makes tests for IgE commercial preparations and intradermal tests and IgG antibodies feasible.

[email protected] 7.10 Patent Blue V, Isosulfan Blue, and Methylene Blue 291

7.9.4 Fondaparinux ogy used for patent blue V and isosulfan blue . The chemical structures of the two triarylmethane Fondaparinux, a synthetic pentasaacharide with dyes are shown in Fig. 7.22 where it can be seen structural identity to a sequence of fi ve sugar that patent blue V, usually obtained as a calcium- units of heparin, is a new class of antithrombotic chelated dimer or sodium salt, differs from iso- agents that selectively inhibits coagulation factor sulfan blue in the hydroxyl group at position 5 Xa. Unlike heparin, fondaparinux does not inhibit and the sulfonate groups at positions 1 and 4 thrombin and it has proved more effective than while isosulfan blue has the sulfonate groups at enoxaparin in preventing venous thromboembo- positions 2 and 5. The literature information on lism after major surgery. Tolerance to the two dyes is often contradictory, especially in fondaparinux in some heparin-sensitized patients relation to synonyms and sometimes even CAS has been demonstrated and antibody-induced numbers, but the information summarized in thrombocytopenia caused by fondaparinux has so Table 7.13 appears to be correct. far not been reported. Results indicate that In a recent study of adverse reactions to pat- fondaparinux has a low allergenic potential with ent blue V in 7,917 patients with breast carci- an incidence of allergic skin reactions of only noma, patients were given patent blue V and 0.4 %. In one recent prospective study involving technetium- 99 m (99m T) colloid as part of senti- 231 patients, no cross allergies were observed in nel lymph node biopsy. The study was part of a patients with delayed-type hypersensitivity reac- UK-wide sentinel lymph node biopsy NEW tions to heparin. The incidence of allergic cutane- START surgical training program and the ous reactions to the drug was one-twentieth of the Axillary Lymphatic mapping Against Nodal heparin fi gure, leading the investigators to con- Axillary Clearance (ALMANAC) multicenter clude that in selected patients, fondaparinux trial under the auspices of the Medical Research might substantially improve patient care, thera- Council of the UK. Adverse reactions were seen peutic safety, and cost-effectiveness of anticoag- in 72 of the 7,917 (0.91 %) patients and no ulant therapy. Delayed-type reactions to patients died. Four patients had nonallergic reac- pentosanpolysulfate have been reported without tions (0.05 %), 23 (0.29 %) experienced minor exposure to the drug if patients are sensitized to (grade I) allergic skin reactions, 16 (0.2 %) had structurally related heparins. In these cases, and grade II reactions, and fi ve (0.06 %) had severe when hirudin must be avoided, fondaparinux grade III reactions. In 24 (0.3 %) patients the appears to be a valuable alternative. adverse reaction was not further described or graded. By comparison, collective results from American studies in which isosulfan was used 7.10 Patent Blue V, Isosulfan Blue, for sentinel lymph node biopsies in breast and and Methylene Blue melanoma patients showed allergic reactions in 119 (1.42 %) of 8,372 patients. With regard to Water-soluble blue dyes with and without iso- the severe allergic reactions (grade III), the fi g- tope, in particular, patent blue V, isosulfan blue, ures were 0.44 % and 0.06 % for isosulfan blue and methylene blue, are being increasingly used and patent blue V, respectively. Pharmacovigilance to identify sentinel lymph nodes in melanoma fi les provided by the manufacturer on the patients and in cases of breast, bladder, cervical, involvement of patent blue V in 158 adverse endometrial, and other cancers. Reports of events collected worldwide and from the litera- adverse reactions to blue dyes date to at least the ture over the 5 year period 2002–2007 revealed 1960s and it is now clear that type I allergic reac- two adverse events for every 10,000 patients. tions including anaphylaxis occur occasionally Note that biphasic reactions to both dyes have with all three of the above-mentioned drugs. been observed with a second episode occurring Unfortunately, confusion surrounds the terminol- 3–8 h after the initial reaction.

[email protected] 292 7 Drugs and Other Agents Used in Anesthesia and Surgery

H3CH2C CH2CH3 H CH C CH CH N+ 3 2 N+ 2 3

.1/2Ca2+ .Na+ - - O3S O3S C C

- CH2CH3 CH2CH3 O3S N N - OH CH2CH3 SO3 CH2CH3

Patent blue V Isosulfan blue

- CH3 Cl CH3 N S+ N H3C CH3 .3H O N 2 Methylene blue

Fig. 7.22 Chemical structures of dyes used for sentinel blue. The latter dye is not always approved for sentinel lymph node biopsies—the triarylmethane dyes, patent lymph node localization blue V and isosulfan blue, and the thiazine dye, methylene

Table 7.13 Comparison of blue dyes that have been used For diagnosing immediate hypersensitivity to for sentinel lymph node localization patent blue V and isosulfan blue, intradermal Dyea Other names CAS nō b /CI nōc testing is generally satisfactory using a 1:100 Patent blue Vd Disulfi ne blue 3536-49- 0/42051 dilution of the stock solution (1 %). Flow cyto- Acid blue 3 metric methods using CD63-positive basophils Patent blue violet Food blue 5 have also been successfully used to demonstrate Trade name—Bleu true type I reactions to patent blue V, but the Patenté V: Guerbete ImmunoCAP® assay for isosulfan blue and some Isosulfan blue Trade name— 68238-36- 8/– g other immunoassay methods for detecting IgE f Lymphazurin™ antibodies to patent blue V have proved negative Methylene blueh Methylthioninium 7220-73- 3/52015 chloride in the few studies carried out. Immunological Basic blue 9 cross-reactivity between the two dyes has been Swiss blue demonstrated in both skin and CD63 expression Aniline violet studies and, as reactions frequently occur on Solvent blue 8 what is apparently fi rst exposure, sensitization by a See Erratum on nomenclature. J Nucl Med 2003;44:649 exposure to triarylmethane dyes in various prod- b CAS nō, Chemical Abstract Service unique numeral identifi er number ucts encountered in everyday life is assumed. c CI nō, Color Index number Methylene blue , or methylthioninium chlo- dAlso used as food colorant with number E131. Banned as ride, is a thiazine dye (Fig. 7.22 , Table 7.13 ) a food dye in Australia and USA because of possibility of unrelated in structure to patent blue V and isosul- allergies e 25 mg/ml fan blue. Although not always approved for the f 10 mg/ml purpose, methylene blue has been used for senti- g No CI number yet assigned nel lymph node localization. Because it can cause h 10 mg/ml injection necrosis on subcutaneous injection, it is generally

[email protected] Summary 293 given IV. In a prospective study of 30 patients in reactive IgE antibodies. Solid phase com- the USA, the dye was used instead of isosulfan plexes of NMBDs and some analogs (such as blue to localize lymph nodes. It proved success- choline for succinylcholine) have also been ful in 90 % of the cases giving results similar to used to detect IgE. isosulfan blue. The investigators pointed out the • When used as a diagnostic aid for allergy to substantial cheaper cost of methylene blue. NMBDs, the BAT has been found to be specifi c but disappointingly lacking in sensitivity. • Reversal of rocuronium-induced NM block Summary with the cyclodextrin sugammadex has highlighted the question of changed allergenicity • NMBDs are responsible for ~60 % of anaphy- of such chemically sequestered drugs in host– lactic episodes in the perioperative period. guest complexes. • Succinylcholine accounts for about one-third • With at least seven current reports in six dif- of the reactions. Reactions to rocuronium are ferent countries of the mitigation of signifi cantly higher in Europe than in rocuronium-induced anaphylaxis by sugam- Australia. madex, this specifi cally modifi ed cyclodextrin • Cardiovascular reactions are common and might prove to be a new and useful treatment serious symptoms during anaphylaxis to to manage rocuronium-induced anaphylaxis. NMBDs. Reactions may progress to cardio- • Many patients who experience anaphylaxis to vascular collapse in up to 80 % of cases and an NMBD do so on fi rst exposure, raising the may be the only symptom in 60 % of cases. question of the sensitizing source. Pholcodine, Bronchospasm is seen more often in anaphy- which cross-reacts with NMBDs, has been lactic than in anaphylactoid reactions to suggested to be this source and this appears to NMBDs. be supported by a large boost in IgE antibod- • Incidences of anaphylaxis to NMBDs are 1 in ies to pholcodine, morphine, and succinylcho- 5,500 in France, 1 in 5,200 in Norway, and 1 line following dosage with a cough syrup in 10,000 in Australia. containing pholcodine. • Reactions to NMBDs are mediated by IgE • “Natural” IgE antibodies to phosphorylcho- antibodies with specifi city for tertiary and line have been suggested as an alternative quaternary ammonium ions, but adjoining explanation for preexisting NMBD allergic structures may also be recognized. Recognition sensitivity. of the substituted ammonium groups accounts • Anaphylactic reactions occasionally occur to for the extensive cross-reactivity between the the hypnotics thiopentone and propofol. Both NMBDs. are diagnosed by skin testing and the former • Substituted ammonium groups occur widely also by a specifi c IgE test. in many drugs and chemicals which means • The colloid hydroxyethyl starch is well toler- that NMBD-reactive IgE antibodies cross- ated with an incidence of adverse events less react with many other drugs. Reaction with than gelatin and dextran. The incidence of morphine is particularly pronounced. allergic reactions is 0.0004 % and the risk of • Considering fi ne structural recognition, the life-threatening reactions 0.006–0.085 %. combining site specifi cities of NMBD-reactive • Gelatin, marketed as Haemaccel® and IgE antibodies fall into fi ve main groups. Gelofusine® , can cause both IgE- and non- • Diagnosis of reactions is effected by skin test- IgE- mediated reactions. Clinical manifesta- ing with free drugs, IgE antibody assays, and tions include anaphylaxis, urticaria, the tryptase assay. Because of morphine’s bronchospasm, and sneezing. striking capacity to detect and cross-react with • Pre-injection of small MW dextran 1 reduces NMBD-reactive antibodies in patients’ sera, it the incidence of dextran-induced anaphylaxis is used in solid phase form to detect NMBD- from 25 to 3 per 100,000.

[email protected] 294 7 Drugs and Other Agents Used in Anesthesia and Surgery

• True IgE-mediated reactions to local anesthet- Baldo BA, Fisher MM. Anaphylaxis to muscle relaxant ics are extremely rare; many reactions appear drugs: Cross-reactivity and molecular basis of binding of IgE antibodies detected by radioimmunoassay. Mol to be vasovagal responses but delayed reac- Immunol. 1983b;20:1393–400. tions are well known. Baldo BA, Fisher MM. Diagnosis of IgE-dependent ana- • Anaphylactic reactions to protamine and apro- phylaxis to neuromuscular blocking drugs, thiopen- tinin occur infrequently and anaphylaxis to tone and opioids. Ann Fr Anesth Reanim. 1993;12:173–81. latex has decreased markedly in recent years. Baldo BA, Fisher MM, Pham NH. On the origin and spec- • The overall incidence of adverse reactions to ifi city of antibodies to neuromuscular blocking (mus- heparin has been estimated at 0.2 % with cle relaxant) drugs: an immunochemical perspective. heparin-induced thrombocytopenia, rare cases Clin Exp Allergy. 2009;39:325–44. Baldo BA, McDonnell NJ, Pham NH. The cyclodextrin of anaphylaxis, a few delayed reactions, and sugammadex and anaphylaxis to rocuronium: is some adverse skin reactions making up most rocuronium still potentially allergenic in the inclusion of the reports. complex form? Mini Rev Med Chem. 2012;12: • Danaparoid and hirudins are chemically dis- 701–12. Ewan PW, Dugue P, Mirakian R, et al. BSACI guidelines tinct from heparin and generally show little or for the investigation of suspected anaphylaxis during no cross-reactivity in heparin-intolerant general anaesthesia. Clin Exp Allergy. 2009;40: patients. Reported reactions to danaparoid 15–31. include rash, pruritus, and reactions (some Fisher MM, Baldo BA. Anaphylaxis during anaesthesia: current aspects of diagnosis and prevention. Eur J delayed) at the injection site. Figures of 0.015 Anaesthesiol. 1994;11:263–84. and 0.16 % have been reported for anaphy- Fisher MM, Baldo BA. Immunoassays in the diagnosis of laxis to hirudins on fi rst and subsequent expo- anaphylaxis to neuromuscular blocking drugs: the sures, respectively. Urticaria and angioedema value of morphine for the detection of IgE antibodies in allergic subjects. Anaesth Intensive Care. 2000;28: are other reported type I reactions. 167–70. • Fondaparinux selectively inhibits coagulation Florvaag E, Johansson SGO. The pholcodine story. factor Xa and has a low allergenic potential. Immunol Allergy Clin North Am. 2009;29:419–27. • In a multicenter trial on patent blue V, adverse Fuzier R, Lapeyre-Mestre M, Mertes PM, et al. Immediate- and delayed-type allergic reactions to amide local reactions occurred in 0.91 % of patients and anesthetics: clinical features and skin testing. no patients died. Nonallergic reactions Pharmacoepidemiol Drug Saf. 2009;18:595–601. occurred in 0.05 % of patients, 0.29 % experi- Harle DG, Baldo BA, Fisher MM. The molecular basis of enced minor (grade I) allergic skin reactions, IgE antibody binding to thiopentone. Binding of IgE from thiopentone-allergic and non-allergic subjects. 0.2 % had grade II reactions, and 0.06 % had Mol Immunol. 1990;27:853–8. severe grade III reactions. Harper NJN, Dixon T, Dugue P, et al. Guidelines. • Studies in which isosulfan was used for senti- Suspected anaphylactic reactions associated with nel lymph node biopsies in breast and mela- anaesthesia. Anaesthesia. 2009;64:199–211. Kroigaard M, Garvey LH, Gillberg L, et al. Scandinavian noma patients showed allergic reactions in clinical practice guidelines on the diagnosis, manage- 1.42 % of patients. With regard to the severe ment and follow-up of anaphylaxis during aneasthesia. allergic reactions (grade III), the fi gures were Acta Anaesthesiol Scand. 2007;51:655–7. 0.44 % and 0.06 % for isosulfan blue and pat- Levy JH. Anaphylaxis during cardiac surgery: implications for clinicians. Anesth Analg. 2008;106: ent blue V, respectively. 392–403. Mertes PM, Alla F, Trechot P, et al. Anaphylaxis during anesthesia in France: an 8-year national survey. Further Reading J Allergy Clin Immunol. 2011a;128:366–73. Mertes PM, Malinovsky JM, Jouffroy L, et al. Reducing the risk of anaphylaxis during anesthesia: 2011 Baldo BA, Fisher MM. Substituted ammonium ions as updated guidelines for clinical practice. J Investig allergenic determinants in drug allergy. Nature. Allergol Clin Immunol. 2011b;21:442–53. 1983a;306:262–4.

[email protected] Opioid Analgesic Drugs 8

Abstract Opioid analgesics are one of the most commonly administered groups of drugs in hospitals. These drugs show common structural features, bind specifi cally to opioid receptors and possess morphine-like pharmacologic action. Tramadol differs from other opioid analgesics in its monoaminergic activity as well as its affi nity for the μ opioid receptor. Many opioids are potent histamine releasers producing hemodynamic changes and anaphylactoid reactions, but there seems to be no direct relationship between the histamine plasma concentrations and these changes. True IgE antibody- mediated immediate allergic reactions to opioids are uncommon, although some anaphylactoid reactions are interpreted as allergic, emphasizing the need to investigate whether or not reactions have an immune basis. The histamine-releasing properties of opioid drugs sometimes hamper skin testing, and general unavailability of specifi c IgE antibody tests contributes to the failure to investigate reactions. Reactions to tramadol, whether anaphylactoid or IgE antibody-mediated, are rare, and the drug is generally considered to be safe with a low potential for adverse reactions. Clinical implications for the diagnosis of opioid drug-induced anaphylactoid and anaphylactic reactions are discussed.

Opioid analgesic drugs (OADs), particularly pain relief are required. OADs might, therefore, fentanyl and its analogs alfentanil, remifentanil, be considered along with other agents used in and sufentanil, are extensively used for anesthe- general anesthesia (Chap. 7 ), but their general sia and analgesia. Administration of intravenous usefulness as analgesics apart from their applica- hypnotics and OADs as a high dose opioid, low tion in the operating room together with their his- dose hypnotic (usually a benzodiazepine such as tamine releasing properties and the large number midazolam) has found wide application in car- of members of the opioid family of drugs, both diac anesthesia and to produce procedural seda- naturally occurring and synthetic, make these tion for endoscopy, catheterizations, and a variety drugs worthy of closer and more individual of other surgical procedures where sedation and examination.

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 295 Relationships, DOI 10.1007/978-1-4614-7261-2_8, © Springer Science+Business Media, LLC 2013

[email protected] 296 8 Opioid Analgesic Drugs

alkaloids and related synthetic drugs led to the 8.1 Terminology identifi cation of endogenous peptide ligands, in particular, enkephalins, β-endorphin, and dynor- The naturally occurring analgesic drug morphine phins for the receptors. These peptides are called along with a number of structurally related alka- opioid peptides since they have effects resem- loids including codeine, noscapine (narcotine), bling opiate drugs and, correspondingly, the thebaine, and papaverine, and together with about complementary receptors are called “opioid.” 25 other minor alkaloids of unknown or little-or- The term “opioid drug” then includes naturally no useful pharmacological action, are obtained occurring and semi- and fully synthetic drugs from the latex from immature seed pods of the that produce their effects by binding specifi cally opium poppy plant Papaver somniferum . The to any of several different opioid receptors and medically important morphine and codeine, and which are competitively antagonized by nalox- the semisynthetic derivatives such as heroin (dia- one. Not all opioid drugs show a receptor recog- morphine), hydromorphone, oxymorphone, nition pattern identical to morphine and hence hydrocodone, and oxycodone, prepared from opioid drugs may, or may not, have similar phar- some of the natural alkaloids (mainly morphine, macological actions as the prototypic opioid. codeine, and thebaine), were originally desig- Opioid receptors do not form a homogeneous nated opiates , a generic term used for both natural population and occur in two major branches. The and synthetic drugs with morphine-like actions. receptors named mu (μ), kappa (κ), and delta (δ), The term narcotic , derived from the Greek where naloxone acts as an antagonist, forms the “narko,” (from narkoun, to numb; narkē, numb- main branch while the other branch comprises ness) was originally applied to any substance that the nociceptin or ORL 1 receptor which shows no relieved pain, dulled the senses, or induced sleep. recognition of naloxone. In 1999 an International The defi nition covered substances that produced Union of Basic and clinical Pharmacology at least some morphine-like actions including (IUPHAR) opioid receptor subcommittee of the unwanted side effects such as drowsiness, eupho- Receptor Nomenclature and Drug Classifi cation ria, nausea, constipation, and dependence and, Committee (NC-IUPHAR) recommended that: therefore, also encompassed some substances not “The well-established Greek terminology for necessarily derived from the opium poppy such as opioid receptor types using the descriptors μ cocaine and coca leaves (in the USA at least). The (mu), δ (delta) or κ (kappa), is recommended, but term now carries with it negative connotations the receptor type should be additionally defi ned since in a legal context, almost universally in the as MOP, DOP, KOP, or NOP when fi rst men- media, and to the layman, it indicates a prohibited tioned in publication.” drug such as morphine, heroin, or the potent Opioid drugs and peptides that bind to the opi- oxy- and hydro-derivatives of morphine and oid receptors produce the primary general and codeine. Therefore, although the terms opiate and specifi c clinical effects summarized in Table 8.1 . narcotic are still often used, they are no longer The opioids relevant to this review are those that useful in a pharmacological context. are used in medicine today meaning, therefore, Nalorphine (N -allylnormorphine), synthe- the frequently administered analgesics, particu- sized in 1942, proved to have not only some anal- larly morphine, codeine, and synthetic analogs gesic action but also strong antagonistic effects such as fentanyl, meperidine, and methadone. on many of the actions of morphine. It was becoming clear that opioid agonists, antagonists (such as naloxone), and mixed agonist–antago- 8.2 Structure–Activity nists must act on multiple receptors and by the Relationships early 1970s, binding studies with radiolabeled drugs revealed stereospecifi c opioid binding sites Morphine, the prototypic opioid, is a phenan- in the central nervous system. Confi rmation of threne alkaloid with a benzylisoquinoline back- the existence of specifi c receptors for opium bone. It is composed of fi ve fused rings A–E,

[email protected] 8.2 Structure–Activity Relationships 297

Table 8.1 Clinical effects resulting from some opioid ligands binding specifi cally to opioid receptors Examples of some drugs and endogenous peptides showing Receptors preferential binding Primary general effect Clinical effects μ Morphine Central depression Analgesia; euphoria; respira- Heroin tory depression; bradycardia; Oxymorphone hypothermia; constipation; Hydrocodone physical dependence Fentanyl, etc.a Endomorphins κ Nalbuphineb Sedation Sedation; analgesia; dysphoria; Butorphanol dissociative, deliriant and Pentazocine hallucinogenic effects Dynorphins δ Leu-enkephalin Analgesia Regulation of analgesia and Met-enkephalin behavior; antidepressant; endocrine function Some data from Freye E. Opioids in Medicine. Dordrecht: Springer; 2008 a Also alfentanil, remifentanil, sufentanil b Drugs that show preferential binding to κ receptors show lower abuse potential three of which are in approximately the same to three to fi ve, especially with unsaturated plane (rings A, B, and E) and with the piperidine bonds, produces compounds such as naloxone ring (D) at right angles (Table 8.2 ). The naturally and naltrexone with high affi nity for μ receptors occurring levorotatory isomer is the active form; but with antagonistic not agonistic properties. the dextrorotatory isomer is devoid of opioid Substitution of a methoxy at position 3, or acetyl activities. For morphine and other closely related groups at positions 3 and 6 instead of hydroxyls, rigid structures, three structural features in par- produces the weak μ receptor agonists codeine ticular are important for analgesic activity, that is, and heroin, respectively. Changes in the C ring for binding to opioid receptors. They are: a ter- can also lead to compounds of increased analge- tiary amine that is part of a piperidine ring; an sic potency as demonstrated with hydromor- aromatic ring (A) axially connected to the piperi- phone where a 6-keto group replaces the hydroxyl dine ring at the carbon para to the nitrogen (C12– and a 7–8 dihydro linkage replaces the double C13 morphine structure Table 8.2 ) thereby bond found in morphine. Addition of a hydroxyl maintaining a fi xed geometry between the aro- group at position 14 generally enhances μ recep- matic ring and the elevated piperidine ring; and tor agonist activity. Oxymorphone and oxyco- polar groups containing an oxygen at C3 of the done are examples, both being equal to, or more phenyl ring. Minor changes in the structure of potent than, morphine. Nalbuphine, also with a morphine usually cause changes, not only in the 14-hydroxyl group but with an N -cyclobutylmethyl physicochemical and pharmacokinetic properties instead of an N -methyl substituent, is agonistic at but also in the selectivity profi le for opioid recep- the κ receptor and antagonistic at the μ receptor. tors. Ring A and the protonated tertiary nitrogen A quick perusal of the two-dimensional struc- of ring D at physiological pH are two dominant tures of a range of important opioid drugs reveals structural features of μ opioid receptor agonists. that while many of the clinically relevant Changes in the groups attached to nitrogen and at compounds show clear structural similarities to the 3 and 6 positions can produce marked changes morphine (Table 8.2 ), such similarities are not in both pharmacological action and potency. For immediately obvious with some other widely example, increasing the length of the carbon used drugs such as the synthesized, and structur- chain attached to the nitrogen from one (methyl) ally fl exible, opioids meperidine (pethidine),

[email protected] 298 8 Opioid Analgesic Drugs

Table 8.2 Structure of morphine and some chemically related naturally occurring or semisynthetic clinically important opioid drugs

17 N CH3 16 D 10 9

1 11 15 14 8 B

2 A C 7 12 13

E 3 4 5 6

OH O OH Morphine

Substituent Substituent Substituent at Substituent at Bond(s) at Drug at position 3 at position 6 position 14 position 17 positions 7–8

Morphine –OH –OH –H –CH3 Double

Codeine –OCH3 –OH –H –CH3 Double

Heroin –OCOCH3 –OCOCH3 –H –CH3 Double

Hydromorphone –OH ═ O –H –CH3 Single

Oxymorphone –OH ═ O –OH –CH3 Single

Hydrocodone –OCH3 ═O –H –CH3 Single

Oxycodone –OCH3 ═O –OH –CH3 Single a b b Buprenorphine –OH –OCH3 Single –CH2

Naloxone –OH ═ O –OH –CH2 CH═CH3 Single From Baldo BA, Pham NH. Histamine-releasing and allergenic properties of opioid analgesic drugs: Resolving the two. Anaesth Intensive Care. 2012;40:216. Reproduced with permission from Australian Society of Anaesthetists a Has a 1-hydroxy-1,2,2-trimethylpropyl substituent at C-7 b Endo-ethano bridge between C-6 and C-14

methadone, and fentanyl that also act at the μ ring via the central carbon C13 in morphine and opioid receptors like the rigid morphine-like struc- C4 in meperidine and terminating in an N -methyl tures. With meperidine, for example, conforma- group. Viewed in isolation, this phenylpropyl- tional studies have shown that the aromatic ring amine group is seen in morphine in the sequence can be axial or equatorial to the piperidine ring, of positions 13, 15, 16, and 17 attached to the aro- there is free rotation between them, and the meper- matic ring A at C12. The corresponding sequences idine structure can be fairly well superimposed on in meperidine are positions 4, 3, 2, and 1 attached the equivalent structural features of morphine to the aromatic ring. These phenylpropylamine (Table 8.2; Fig. 8.1). Both morphine and meperi- groups are highlighted on the structure of mor- dine have a phenylpiperidine substituent, a phine in Table 8.2 and the meperidine structure sequence of three carbons linked to an aromatic (Fig. 8.1 ). Fentanyl has a 4- anilidopiperidine

[email protected] 8.2 Structure–Activity Relationships 299

Phenylpropylamine structure: Anilidopropylamine structure: (Phenyl links to the propylamine 17 N CH3 sequence via a nitrogen) 16 3 1 H2 2 N C C R 15 2 4 H2 N 13 R1

C CH3 O C O H2 HO OH R1 R2 3 1 2 N H CH3 4 N N

2 CH2OCH3 N N C C CH2CH3 O O 3 O

COOCH3 COOCH3

CH2OCH3 N S CH 3 a (5 ,6 )-7,8-didehydro-4,5-epoxy-17-methylmorphinan-3,6-diol b Ethyl 1-methyl-4-phenylpiperidine-4-carboxylate 4 c (RS)-6-(dimethylamino)-4,4-diphenylheptane-3-one d N-[1-(2-phenylethyl)-4-piperidinyl]-N-phenylpropanamide C e N-{1-[2-(4-ethyl-5-oxo-4,5-dihydro-1H-1,2,3,4-tetrazol-1-yl)ethyl]- O C 4-(methoxymethyl)piperidin-4-yl}-N-phenylpropanamide f Methyl 1-(3-methoxy-3-oxopropyl)-4-(N-phenylpropanamido)- piperidine-4-carboxylate g N-[4-(methoxymethyl)-1-(2-thiophen-2-ylethyl)-4-piperidyl]-N- phenylpropanamide

Fig. 8.1 Comparison of the structures of morphine and but their conformational similarities to morphine and meperi- some important synthetic opioid analgesics. While clear dine are readily apparent. Although methadone lacks a piper- similarities in structure are not always obvious, both mor- idine ring, it retains a phenylpropylamine substituent phine and meperidine have a phenylpiperidine structure and ( highlighted), and it is thought that the enol tautomer of a phenylpropylamine grouping, highlighted here as a l - methadone may form a morphine-like conformer by intra- sequence of three carbons linked to the aromatic ring via the molecular hydrogen bonding with the nitrogen (see Figs. 8.2 central C13 carbon in morphine and C4 in meperidine and and 8.3 ). From Baldo BA, Pham NH. Histamine-releasing terminating in an N -methyl group. Fentanyl, alfentanil, remi- and allergenic properties of opioid analgesic drugs: Resolving fentanil, and sufentanil contain a 4-anilidophenylpiperidine the two. Anaesth Intensive Care. 2012 ;40:216. Reproduced and a 4-anilidophenylpropylamine ( highlighted) structure, with permission from Australian Society of Anaesthetists rather than a 4- phenylpiperidine sequence but, phine and meperidine, methadone retains a phen- again, its conformational similarity to meperidine ylpropylamine structure (highlighted, Fig. 8.1 ) and hence morphine is readily seen although link- and has 11 degrees of rotational freedom. The age of the aromatic ring to the propylamine molecule has a heptane backbone and can be sequence is via a nitrogen (Fig. 8.1 ). Methadone is viewed as a central carbon (C4) linked to two phe- similar to morphine in receptor binding and phar- nyl groups, a ketone and a propyl group with an macological action; however, the similarity is not, attached acyclic tertiary amine. A number of mor- at ﬁ rst sight, readily reﬂ ected in structural confor- phine-like conformers of this structure have been mations of the two compounds. Although a advanced usually based on intramolecular interac- piperidine ring is no longer present, like both mor- tions between the carbonyl and amine groups. In

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H3C Methadone Enol tautomer CH N 3 of l-methadone H3C CH3 H H3C O N CH 3 CH CH C 3 H 2

C CH 3 O C H 2 1a 1c 2a 2c

1b 1d 2b 2d

Fig. 8.2 Two-dimensional structures of l -methadone and hydrogen bond between the positive charge of the hydroxyl the enol tautomer of l- methadone together with three- hydrogen and the unshared electrons of nitrogen to give a dimensional space-ﬁ lling ball-and-stick (1a, c and 2a, c) and seven-membered ring, which may be seen as the counterpart CPK models (1b, d and 2b, d) for l -methadone and its enol of the morphine piperidine ring. In 1c, d and 2c, d the mor- tautomer, respectively. The models of l -methadone show phine pharmacophore of a tertiary alkylamine at least three one of many possible conformations while the models of the atoms away from, and including, an aromatic ring is shown enol form show the molecule locked by the intramolecular in blue while the rest of the molecule is shown in light gray one proposed conformer a pseudopiperidine ring clear difference in the position and orientation of is formed by hydrogen bonding between one of the morphine pharmacophore between the two the methyl groups on the nitrogen and the car- conformations is clearly visible. Figure 8.3 , 2a–d bonyl oxygen. In another suggested conformer, reveals the close similarity between the conforma- the enol tautomer of l -methadone is thought to tion of the phenylpropylamine sequence in the form an intramolecular hydrogen bond between enol tautomer of l - methadone and the same the nitrogen proton and the oxygen thus producing sequence in selected conformations of morphine a seven-membered ring that is seen as the counter- (Fig. 8.3 , 1a–d) and meperidine (Fig. 8.3 , 3a–d) part of the piperidine ring of morphine. Models of and the anilidopropylamine sequence of fentanyl l -methadone (Fig. 8.2 , 1a–d) and its enol tautomer (Fig. 8.3 , 4a–d). It should be remembered though (Fig. 8.2, 2a–d) reveal clear differences in shape that no conclusive evidence for the existence, one and orientation of structures within the methadone way or the other, of the postulated conformers has molecule. Figure 8.2 , 1a–d shows one of many been forthcoming and the actual conformation(s) possible conformations due to free rotation about of methadone at the opioid receptor sites is yet to the sp 3-hybridized central C-4 atom of methadone be ﬁ rmly established. with its tetrahedral symmetry. In the enol form The synthetic, centrally acting analgesic trama- (Fig. 8.2 , 2a–d), however, the molecule is locked dol with its phenylpropylamine sequence (high- by the intramolecular hydrogen bond, and the lighted in Fig. 8.4a, b) shows some structural

[email protected] 8.3 Classiﬁ cation of Opioid Drugs 301

Fig. 8.3 Space-ﬁ lling ball-and-stick (1a-4a, 1c-4c) and seven-membered ring, predicted to form by intramolecular CPK (1b-4b, 1d-4d) three-dimensional molecular models H-bonding of the enol tautomer form with the nitrogen, is of the structures of morphine (1a-1d), methadone (2a-2d), shown most clearly in 2a and 2c. This “virtual” ring is seen meperidine (3a-3d), and fentanyl (4a-4d). (For two-dimen- as the counterpart of the piperidine ring in morphine, sional structures of morphine, methadone, meperidine, and meperidine, and fentanyl. Conformations were selected to fentanyl refer to Fig. 8.1 ). Conventional colors used for show the clear similarities in shape, size, and orientation of different atoms are shown in structures 1a–4a and 1b–4b. In the pharmacophore in all four drugs. From Baldo BA, structures 1c–4c and 1d–4d the morphine pharmacophore Pham NH. Histamine-releasing and allergenic properties of of a tertiary alkylamine at least three atoms away from, and opioid analgesic drugs: Resolving the two. Anaesth including, an aromatic ring is shown in blue while the rest Intensive Care. 2012 ;40:216. Reproduced with permission of the molecule is shown in light gray . For methadone, the from Australian Society of Anaesthetists

resemblances to codeine and morphine and is (5-hydroxytryptamine) and norepinephrine (nor- classed as an opioid despite showing some signifi - adrenaline) uptakes. (+)-Tramadol is approxi- cant differences to the other opioid analgesics. mately four times as potent as (−)-tramadol in Tramadol exists as four stereoisomers, but the mar- inhibiting serotonin reuptake while (−)-tramadol is keted compound is a racemic mixture of the about ten times stronger than the (+)-enantiomer in (1R,2R) or (+)-enantiomer and the (1S,2S) or inhibiting norepinephrine reuptake. These actions (−)-enantiomer. The drug has a low affi nity for are complementary and synergistic and together the μ opioid receptor, but it can be considered to with the receptor agonist activity lead to inhibitory be a prodrug with the active metabolite effects on pain transmission in the spinal cord. O-desmethyltramadol (Fig. 8.4c, d ), formed by demethylation of the methoxyphenyl group, being a more potent μ receptor agonist than the parent. 8.3 Classifi cation of Opioid Drugs Only partial inhibition of tramadol’s analgesic action by the opioid antagonist naloxone suggested Opioid drugs can be classifi ed in a number of the involvement of a second mechanism of action, ways but, from the medical point of view, the and this was explained by the demonstration of most obvious, simple, and useful classifi cation is monoaminergic activity with effects on serotonin on the basis of their clinical importance. To this

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acO O OH CH 3 H3C

H H H HO H3C OH HO CH3 CH3 1 N 1 1 N 2 N 2 2 CH 3 CH3 CH3

(1R,2R)- (1S,2S)-

CH H C b 3 3 d CH3 N N N 1 1 1 CH 2 CH3 H3C 2 2 3

H 3 H H HO 3 OH HO 3 4 4 4

OCH3 H3CO OH

Tramadol O-Desmethyltramadol

Fig. 8.4 Tramadol, 2-(dimethylaminomethyl)-1-(3-meth- retains a phenylpropylamine structure (see Fig. 8.1 ), that is, oxyphenyl)cyclohexanol (a , b), can exist in four different a sequence of three carbons linked to the aromatic ring and stereoisomeric forms, (1R,2R), (1S,2S), (1R,2S), and terminating in a methyl ammonium group (marked in bold (1S,2R), but the drug used (as the hydrochloride) is a race- in a and drawn in b to show similarities with structures mic mixture of the fi rst two of these isomers that are shown shown in Fig. 8.1 ). O -Desmethyltramadol (c , d ), the phar- here. Like methadone, tramadol lacks a piperidine ring but macologically active metabolite of tramadol, has a hydroxyl like morphine, codeine, meperidine, and methadone it instead of the methoxy group at position 3 on the phenyl ring initial classifi cation one can add the origin of a classed as a strong agonist and methadone a weak drug, namely, whether it is naturally occurring, agonist. Naloxone is an antagonist while pentaz- semisynthetic, that is, synthesized from a natural ocine is a mixed agonist–antagonist and nalor- product, or prepared by total synthesis. The phine is classed as an antagonist with a little chemical structure of the compound provides a agonist activity. Most of the opioids used clini- further useful classifi cation as does the traditional cally, like morphine, are relatively selective for μ way of comparing the drug on the basis of anal- receptors but drugs may interact with additional gesic potency. Finally, individual drug selectivi- receptors if given at higher dosages. This can ties for receptors, that is, agonist or antagonist lead to changes in a drug’s pharmacological pro- actions at the different opioid receptors, add per- fi le. Even at normal clinical doses, agonist– haps the most important information that leads to antagonist opioids in particular may recognize an understanding of action and hence clinical more than one receptor, acting as an agonist at applications of the various opioid drugs. one and an antagonist at the other. Table 8.3 sum- Opioid drugs show a wide range of agonist marizes side-by-side the above-listed different and antagonist effects. For example, morphine is classifi cation categories for 13 different clini- an agonist but in relative terms, phenazocine is cally relevant opioids.

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Table 8.3 Classifi cation of some clinically important opioid drugs Main receptor(s) Function at Drug Origin Chemical classa Analgesic potency recognizedb receptorb Morphine Naturally Phenanthrene Intermediate–strong μ Agonist occurring Codeine Naturally Phenanthrene Weak–intermediatec μ, δ Weak agonist occurring Heroind Semisynthetic Phenanthrene Strong μ Agonist Hydromorphone Semisynthetic Phenanthrene Strong μ Agonist Oxymorphone Semisynthetic Phenanthrene Strong μ Agonist Hydrocodone Semisynthetic Phenanthrene Intermediate–strong μ Agonist Oxycodone Semisynthetic Phenanthrene Intermediate μ Agonist e Buprenorphine Semisynthetic Phenanthrene Intermediate μ , δ, ORL1 Partial agonist Naloxone Semisynthetic Phenanthrene –f μ, δ, κ g Antagonist Meperidine Synthetic Phenylpiperidine Weak–intermediate μ Weak agonist (pethidine) Methadone Synthetic Phenylpropylamineh Intermediate μ Agonist Fentanyli Synthetic Anilidopiperidine Strong μ Agonist Tramadol j Synthetic Phenylpropylamine Weak–intermediate μ Weak agonist a Some fi t into more than one chemical class b May have weak agonist/antagonist action at other opioid receptors c Acts after metabolized to 6-glucuronide derivative d Metabolized to morphine e Agonist at μ and ORL1 , and antagonist at κ f Antagonist action of opioid analgesics g Highest affi nity at μ receptor, low affi nity at κ h Also classifi ed as a phenylheptylamine i Similar properties for alfentanil, remifentanil, and sufentanil j Also has monoaminergic activity and metabolized to O -desmethyltramadol which is more active at μ receptor than parent drug

8.4 Opioids and Histamine Release 8.4.1 Histamine Receptors

Many opioid drugs are potent releasers of hista- Refer to Sect. 3.2.5.1 for an extended discussion mine from animal tissues producing, for exam- of histamine and its receptors. The range of dif- ple, rash, fl ushing, urticaria, pruritus, hypotension, ferent biological effects displayed by histamine and mucous. This property continues to cause suggested that some of them must proceed via diffi culties and confusion in diagnosis and treat- different receptors and application of selected ment of various conditions ranging from a simple specifi c agonists and antagonists ultimately itch to life-threatening anaphylaxis since many of revealed three G protein-coupled receptors desig- the symptoms elicited by histamine are similar to nated H1 , H2 , and H3 . Each receptor mediates dif- those of antibody-mediated type I allergic hyper- ferent effects, for example, smooth muscle sensitivity. With the view of distinguishing ana- contraction and some allergic effects including phylactoid from true anaphylactic reactions, the vasodilation and bronchoconstriction proceed histamine releasing properties of those opioid under H1 receptor control; H 2 receptors regulate analgesic drugs (OADs) so far studied for this gastric acid secretion and smooth muscle relax- effect will, therefore, be covered in some detail ation and inhibit some immunological cellular and the fi ndings compared with features of opioid processes; and H 3 receptors, fi rst cloned in 1999, drug-induced immediate IgE-antibody mediated control histamine turnover, act as receptors in allergic reactions. histaminergic neurons, and inhibit the release of

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a number of neurotransmitters including acetyl- nation of H 4 and H1 receptor antagonism might choline, dopamine, GABA, norepinephrine, and offer a valuable new approach for the treatment serotonin. Even with this variety of effects, not of pruritus-related conditions such as atopic all of histamine’s actions could be accounted for dermatitis. by the three known receptors and, in 2000, a As work on the H4 receptor progresses, evi- fourth receptor H4 was cloned and characterized. dence is accumulating for an immunomodulating In humans, mast cells, and basophils, role for histamine in the immune response, and enterochromafﬁ n- like cells of the gut and there is much optimism that further insights into

histaminergic nerves in the brain constitute the the role of H 4 receptors in immune and infl amma- three main sources of histamine. H1 and H 2 recep- tory disorders will yield novel therapeutic agents tors are widely expressed in the body, H3 recep- for the alleviation of a range of allergic condi- tors are found in the peripheral and central tions. Other demonstrations of the expression of nervous systems, and H 4 receptors in bone mar- functional histamine receptors on human T and row and white blood cells as well as in the small dendritic cells also point to an immunomodula- intestine, colon, and in organs such as the lung, tory effect of histamine. spleen, liver, and tonsils. Histamine is, of course, a key mediator in allergic processes and, of all the body tissues that show some connection to this 8.4.2 Early Studies on Opioid Drug- prime mediator, the mast cell is probably of Induced Release of Histamine greatest interest to allergists and dermatologists. Perhaps surprisingly, expression of histamine In 1907 Windaus and Vogt completed the fi rst receptors on mast cells had been diffi cult to show, chemical synthesis of histamine and soon after the effect of histamine on mast cells remained Sir Henry Dale and coworkers began investiga- unclear, and, intriguingly, effects on cells by tions that showed that histamine was a powerful some antihistamines did not appear to be medi- vasodepressant, it stimulated smooth muscle ated by H1 receptors. The relationship between from the gut and respiratory tract and caused histamine and the mast cell became a little clearer shock when injected into laboratory animals with the fi nding that the autacoid induces chemo- mimicking the systemic effects of anaphylaxis. taxis of mouse mast cells without affecting These early results were followed by demonstra- degranulation of the cells, and this affect can be tion of the involvement of histamine in vascular blocked by a dual H3 /H4 antagonist but not by H1 reactions of the skin and the observation that or H 2 antagonists. The chemotactic response, as morphine caused the so-called triple response in well as calcium mobilization, is mediated by H 4 human skin, that is, the event sequence of an ini- but not H 3 receptors with the former, but not the tial red spot followed by a red irregular fl are and latter, being expressed on the mast cells. In fact, a fl uid-fi lled wheal. Over 30 years later antihista- histamine does not seem to have any effect on mines were shown to reduce morphine-induced degranulation of mast cells either alone or in skin wheals, and histamine itself was detected in combination with anti-IgE complexes. H2 and H4 effl uents of isolated perfused cat gastrocnemius receptors have been shown to be expressed on muscle after arterial injection of opium alkaloids. human skin and leukemic mast cells, and evi- Released histamine was also detected in cat skin, dence is accumulating that the H 4 receptor, to a and raised levels were found in plasma after much greater extent than the H1 receptor, is intravenous injection of morphine. implicated in some pruritic responses. The signifi cance of the release of histamine in Experiments employing models of allergic con- humans stems from its actions of increasing both tact dermatitis have shown that although H 4 heart rate and the force of myocardial contrac- receptor antagonism fails to reduce the allergic tion, thus increasing cardiac output, and from its infl ammatory response, allergen-induced itch is dilatator effect on small blood vessels that leads strongly inhibited, and this suggests that a combi- to fl ushing, decreased vascular resistance, and

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Table 8.4 Monitoring cutaneous mast cell degranulation intradermal challenge with histamine and antigen. in vivo in humans. Histamine levels in venous blood Two interesting fi ndings were that morphine following intradermal challenge with histamine, morphine, and antigen liberated histamine concentrations suffi cient to produce adverse symptoms and the time taken for Time (min) Range of Time (min) Challenge to reach peak histamine to return to histamine levels to return to baseline following agent histamine level a levels (ng/ml) baselineb antigen injection was over twice the 30-min Histamine 2–10 1.4–85.2 30 period seen with histamine and morphine. (2 mg) A review of the many human studies on the Morphine 1–8 2.3–12.7 30 histamine releasing properties of OADs (the vast sulfate majority employing morphine) reveals not only Antigen 5–15 1.1–24.4 >60 signifi cant progress in elucidating hemodynamic Results summarized from McBride P et al. J Allergy Clin and cutaneous changes but also a picture that is Immunol. 1989;83:374 a For all three challenges, histamine levels peaked sometimes confusing and contradictory and with 5–10 min before maximum development of wheal and some important questions still in need of fl are reactions convincing answers. Table 8.5 is an attempt to b Histamine baseline levels, 0.6 ng/ml summarize results obtained from the most signifi cant in vivo and in vitro studies designed to examine morphine-induced hemodynamic and hypotension. Despite the many early relevant cutaneous changes in patients over the last 35 demonstrations of the physiological actions of years. Different fi ndings of plasma histamine histamine in laboratory animals, and of morphine- concentrations, peripheral and central hemody- induced release of the autacoid in human skin, an namic vascular effects, and cutaneous changes understanding of the clinical relevance of hista- can often be explained by the dose of opioid mine liberation by opioid drugs was slow in com- used, its mode and rate of administration, appli- ing. Studies in the late 1950s on urticaria cation of different methodologies for measuring pigmentosa linked opioids with the release of the released histamine, the site of opioid injec- histamine and fl ushing of the skin and then in the tion, the distribution of histamine receptors, the early 1970s, studies in healthy humans demon- effects of other medications, and the variation in strated signifi cant decreases in peripheral and patients’ physiological responses to histamine. total systemic vascular resistance following the For a given dose of morphine, variations between injection of doses of up to 2 mg/kg of morphine. individuals in plasma concentrations of histamine and hemodynamic and skin changes can be signifi cant and extend over a wide range. 8.4.3 Summary of Morphine-Induced Therefore, in reviewing the relevance of plasma Hemodynamic and Cutaneous histamine levels to hypotension, one needs to Changes in Humans take into account that morphine itself has direct effects on the vascular, individual responses to Infusion studies designed to examine the relation- histamine can vary widely, the relative magni- ship between human plasma histamine levels and tudes of histamine release and cardiovascular symptoms revealed that from resting levels of effects are unpredictable, and apparent correla- 0.62 ng/ml, 1.61 ng/ml of histamine produced a tions of morphine- induced histamine levels with 30 % increase in heart rate, 2.39 ng/ml elicited decreases in systemic vascular resistance do not fl ush and headache, and 2.45 ng/ml was followed imply causation. Because of morphine’s capacity by a 30 % increase in pulse pressure. Venous to directly induce vascular responses, the use of blood plasma histamine levels were monitored antihistamines to antagonize the adverse hemo- after intradermal injection of morphine to induce dynamic effects of histamine-releasing opioids cutaneous mast cell degranulation and the fi nd- is not likely to be completely effective. As well ings (Table 8.4 ) were compared with the results of as morphine’s multiple direct effects on the

[email protected] Table 8.5 Summary of studies on morphine-induced histamine release. Hemodynamic and skin test changes in patients and in vitro fi ndings Route of administration (or in vitro study) and dose (or concentration) of Histamine Hemodynamic and other morphine used releasea effectsb Reference Intravenous 0.5 or 1 mg/kg rapid IV NTc ↓ PVR d , ↑ capacitance Hsu et al. Anesthesiology. 1979;50:98 1 mg/kg IV at 5–10 mg/ ↑ ↓ SVR, ↓ DBP, ↑ CIe Philbin et al. Anesthesiology. min 1981;55:292 0.3 mg/kg IV at 10 mg/ ↑ Hypotension, tachycardia, Fahmy. Anesthesiology. min erythema 1981;55:329 ↓ SVR, ↑ CO, ↑ catecholamines 0.3 mg/kg IV at ↑ ↓ SVR, ↓ MAP, ↓ SBP, ↑ CO, ↑ Fahmy et al. Clin Pharmacol 5–10 mg/min HR, ↑ SV, ↑ plasma Ther. 1983;33:615 epinephrine 1 mg/kg IV at 100 μg/kg/ ↑f ↓ MAP, ↓ SVRf Rosow et al. Anesthesiology. min over 10 min 1982;56:93 0.6 mg/kg IV ↑g Hypotension, tachycardiag Flacke et al. Anesth Analg. over ≤ 10 min 1987;66:723 1 mg/kg IV over 10 min → – Warner et al. J Cardiothorac Vasc Anesth. 1991;5:481 0.16 mg/kg IV ↑ h − Withington et al. Anesthesia. 1983;48:26 0.15 mg/kg IV ↑ i “Symptoms”i of histamine Doenicke et al. Clin release but no hemodynamic Pharmacol Ther. 1995;58:81 changes 0.07–0.14 mg/kg IV over → j ↑ MAPk , ↑ HR, → Mildh et al. Anesth Analg. 2 min catecholamine 2000;91:51 Intradermal 5 × 10−6 M to 1.5 × 10−3 M ↑l wheal and fl are Levy et al. Anesthesiology. intradermal skin tests 1989;70:756 In vitro studies 1.5 × 10 −5 M to 4.5 × 10−3 M ↑m – Hermens et al. Anesthesiology. in vitro with leukocytes and 1985;62:124 skin mast cells 10 −5 M to 3 × 10−4 M in ↑n – Stellato et al. Anesthesiology. vitro with human basophils, 1992;77:932 skin, lung, and heart mast Marone et al. Ann Fr Anesth cells Reanim. 1993;12:116 Adapted from Baldo BA, Pham NH. Histamine-releasing and allergenic properties of opioid analgesic drugs: Resolving the two. Anaesth Intensive Care. 2012 ;40:216. With permission from Australian Society of Anaesthetists a Histamine or effect elevated ↑; decreased ↓; unchanged → b CI cardiac index, CO cardiac output, DBP diastolic blood pressure, HR heart rate, MAP mean arterial pressure, PVR peripheral vascular resistance, SBP systolic blood pressure, SO stroke volume, SVR systemic vascular resistance c Not measured d 46 % increase at 2 min. Returned to control values at 9 min. When promethazine preceded morphine decrease in PVR was 25 % e Minimal effects on responses by H1 and H2 blockers alone but signifi cant attenuation by H1 + H2 f Biggest decreases in SVR occurred in patients with highest levels of plasma histamine g In 1 of 10 patients h In 9 of 38 patients i In 13 of 15 patients within 5 min of injection j Local signs (redness, itching) at injection site k Transient increase l Relative to histamine control m Histamine released from skin mast cells only—detected at 1.5 × 10−4 M and maximum at 5 × 10−4 M morphine n From skin mast cells only

[email protected] 8.4 Opioids and Histamine Release 307 vasculature, other hemodynamically active and mast cells and no release with associated infl ammation-producing mediators may be cardiovascular effects. The consensus is that released along with histamine following mor- fentanyl, sufentanil, alfentanil, and remifentanil phine-induced mast cell degranulation and this do not cause histamine release when normal too contributes to the variable response to it and doses of the drugs are given intravenously. some other OADs. In addition to histamine, other Apart from morphine and fentanyl, meperi- preformed biologically active mediators and pro- dine is the only other opioid analgesic that has teins are liberated from mast cell granules. These been fairly well studied for histamine liberating include the proteases chymase and tryptase which properties. This synthetic opioid is a potent cause tissue damage, serine esterases, cathepsin releaser of histamine. In the skin it induces itch G, carboxypeptidase, eosinophil chemotactic and wheal and fl are reactions and erythema, factor, neutrophil chemotactic factor, some hypotension, tachycardia, and epinephrine release interleukins, the anticoagulant heparin, platelet- following intravenous injection. Of the other clin- activating factor (PAF), and tumor necrosis ically important opioid analgesic drugs only oxy- factor, although most of the latter is newly syn- codone, oxymorphone, buprenorphine, alfentanil, thesized by activated mast cells. remifentanil, sufentanil, methadone, and codeine In addition to studying hemodynamic have been studied to any extent. The histamine- changes caused by liberated histamine, plasma releasing capacity of codeine is well known with epinephrine and norepinephrine levels have been the drug sometimes employed as a positive con- investigated following injection of morphine. In a trol alongside allergen extracts in skin tests. study of a patient who experienced an anaphylac- Interestingly, some opioid drugs reveal some toid response following the intravenous injection functional differences between human mast cells of morphine 0.3 mg/kg, plasma catecholamines and basophils and between mast cells from dif- were increased and this was accompanied by ferent anatomical sites. This is illustrated by mor- decreases in systemic vascular resistance and phine’s selective release of histamine and tryptase arterial blood pressure. In another study, intrave- from human skin mast cells but not from lung, nous morphine increased cardiac output, hista- intestinal, and heart mast cells or from blood mine, and epinephrine plasma concentrations and basophils, while buprenorphine releases hista- decreased arterial blood pressure and systemic mine and tryptase from human lung but not skin vasculature resistance in adult subjects with no mast cells. Despite the belief by some that high history of drug allergy or clinical evidence of car- doses of injected morphine can induce an ana- diovascular, pulmonary, or metabolic disease. phylactoid reaction via the release of histamine Although most studies of opioid analgesics from skin mast cells, the above observational dif- have been done with morphine, it is clear that not ferences may indicate why morphine rarely pro- all opioid drugs show the same capacity to release duces systemic responses resembling anaphylaxis histamine. In both in vivo studies in humans and even though it strongly induces histamine release in in vitro experiments with human leukocytes; in the skin and why opioid drug-induced direct basophils; and skin, lung, and heart mast cells, histamine release does not, or rarely, produces fentanyl caused no change in plasma histamine bronchospasm. In fact, if bronchospasm does levels and nor did it, or oxymorphone, release the occur in an anaphylactoid reaction, the reaction is mediator from mast cells in vitro. The fi nding more likely to be immune-mediated (Sect. 7.3 ). that naloxone did not release histamine from skin Findings so far with mast cells derived from heart mast cells and did not inhibit morphine-induced tissue indicate that the human heart may be both histamine release suggested that the release of the site and target of anaphylactic reactions. histamine by morphine is a direct degranulating Human heart mast cells are activated in vitro by a effect that does not proceed via opioid receptor variety of drugs including some anesthetic agents recognition. Overall, studies on fentanyl have and radiocontrast media to produce non-IgE- revealed little or no release of histamine from mediated “anaphylactic” reactions; anti-IgE,

[email protected] 308 8 Opioid Analgesic Drugs anti-FcεI, and C5 induce the release of histamine non- immediate hypersensitivity reactions (Sect. and tryptase, and it has been suggested that ana- 3.2.5.1), and since many opioid drugs are potent phylactic reactions could be particularly severe in releasers of histamine, confusion and diffi culties patients with certain cardiovascular diseases. arise in distinguishing and managing what pres- Because of its effects on the heart and vascu- ents as the same clinical picture, namely, anaphy- lature, in particular its venodilation properties lactoid and anaphylactic reactions. When one and its capacity to produce small reductions in considers the variable cutaneous histamine heart rate, morphine is now often indicated in the releasing properties of opioid analgesic from initial treatment of acute coronary syndromes strong for morphine, codeine, and meperidine to (ACS). Although no randomized, controlled tri- weak or absent for fentanyl, diffi culties involved als have been undertaken, bodies such as the in employing the drugs in skin tests at the diag- American Heart Association in a Scientifi c nostic level are readily apparent. This has led to Statement of guidelines and under the heading the recommendation that placebo-controlled “Management strategies: Basic therapy for acute challenge is necessary for the accurate diagnosis coronary syndrome (ACS)” has recommended of allergic sensitivity to opioid drugs, but the the administration of morphine when symptoms drugs may sometimes be used for this purpose if are not immediately relieved by nitroglycerine diluted beyond their histamine releasing concen- and a β-blocker or when acute pulmonary con- trations. Before considering the question of how gestion or agitation is present. This relatively opioid drug-induced anaphylactoid and anaphy- recent therapeutic application of morphine and lactic reactions can be resolved, their allergenic- its vasodilation effect in human veins also ity and frequency of allergic reactions will be reminds us that the drug’s direct effect on the considered. peripheral arterial vasculature is not well under- One might assume that given the widespread stood. Recently, intra-arterial infusion of mor- prescribing and frequent administration as anal- phine was shown to elicit dose-dependent gesics of morphine, codeine, their hydro- and vasodilation mediated by histamine and nitric oxy-derivatives, meperidine, fentanyl, and trama- oxide. Thus, morphine can be considered to be a dol, and the use of heroin (often by intravenous vasodilator of both arteries and veins reinforcing injection) and methadone by addicts, IgE- the belief that the opioid might be useful for the antibody-mediated allergy to these drugs would treatment of some cardiovascular diseases. not be uncommon. This is not the case. The rea- Unlike morphine, intravenous tramadol pro- sons why are examined below. duces no change in plasma histamine concentration and no systemic anaphylactoid reactions or fl ushing of the skin. Only slight and transient 8.5.1 Naturally Occurring and elevations in blood pressure and heart rate with- Semisynthetic Opioid Drugs out abnormal ECG fi ndings have been noticed so the drug can be considered to be hemodynami- There is only a handful of reports of adverse reac- cally stable. Also, unlike nonsteroidal anti- tions of the immediate kind following the admin- infl ammatory drugs, tramadol does not inhibit istration, usually by intravenous injection, of prostaglandin synthesis. morphine and other naturally occurring opioid drugs, but, because of the failure to apply basic diagnostic tests, a confi dent diagnosis distin- 8.5 Allergenicity of Opioid guishing an anaphylactoid from an immune- Analgesic Drugs mediated reaction is often not possible. In the most comprehensive and informative immuno- Many of the symptoms elicited by histamine are logical study so far, IgE antibodies that reacted the same or similar to those of antibody-mediated equally well with morphine and codeine were type I allergic responses and some other detected in the serum of a suspected anaphylactic

[email protected] 8.5 Allergenicity of Opioid Analgesic Drugs 309 patient following intramuscular injection of papaveretum–hyoscine as preanesthetic medication. In the late 1980s when the reaction occurred, papaveretum, a mixture of opium alkaloids, was standardized to contain 47.5–52.5 % morphine, 2.5–5.0 % codeine, 16.0–22.0 % narcotine, and 2.5–7.0 % papaverine. Intradermal tests revealed positive wheal and fl are responses to papaveretum 200 ng/ml and morphine 100 ng/ml (0.3 × 10−6 M) but negative results with hyoscine, meperidine, and fentanyl. Skin testing of normal subjects was not undertaken. Liberation of histamine by morphine from human skin mast cells in vitro and in human skin is fi rst detected at 1.5 × 10 −4 M and 5 × 10 −6 M, respectively, so the skin test result with morphine probably refl ected Fig. 8.5 Specifi c inhibition by morphine and some structurally related opioid drugs of IgE antibodies in the antibody-mediated histamine release by mast serum of a patient allergic to papaveretum. Drugs were cells. Direct binding immunoassays employing used to inhibit antibody binding to a morphine–solid morphine, codeine, and hyoscine solid phases to phase covalent complex. Symbols: ( open circle ) mor- detect specifi c IgE antibodies revealed strong phine, ( fi lled circle ) codeine, (open square ) nalorphine, ( fi lled square) naloxone, ( open triangle) naltrexone, ( fi lled positive reactions to both opium alkaloids and a triangle) buprenorphine, (inverted open triangle ) meperi- weak to equivocal reaction with hyoscine. dine, ( inverted fi lled triangle) methadone, ( open diamond ) Quantitative hapten inhibition studies with some fentanyl, ( fi lled diamond) hyoscine. From Harle DG et al. analogs of morphine (Figs. 8.5 and 8.6 ) showed Anaphylaxis following Administration of Papaveretum. Case Report: Implication of IgE Antibodies that React that the two best inhibitors, morphine and with Morphine and Codeine, and Identifi cation of an codeine, which differ only at position 3 where Allergenic Determinant. Anesthesiology. 1989;71:489. morphine has a hydroxyl and codeine a methoxy Reproduced with permission from Lippincott Williams & group, were of equal potency. This suggested that Wilkins the IgE antibodies did not recognize this region of the morphine and codeine molecules. Some has an N -methyl and methadone a dimethylamino recognition of nalorphine was also obvious but group. Analysis of the inhibition results sug- the decrease in inhibitory potency indicated that gested, therefore, that the important structural the composition of the group attached to the features of the morphine (and codeine) allergenic piperidine nitrogen was also important for IgE (that is, IgE antibody binding) determinant are antibody recognition. Evidence of the impor- the cyclohexenyl ring with a hydroxyl at C6 and, tance of the cyclohexenyl ring for antibody bind- most importantly, a methyl substituent attached ing was the weak inhibition shown by naloxone to the nitrogen (Fig. 8.7 ). These fi ne structural that shares an N -propyl group with nalorphine features of the morphine allergenic determinant but differs in having a keto group at C6, a single are similar to the complementary structures bond at C7–C8, and a hydroxyl group at C14. recognized by antibodies to morphine raised in Further evidence of the importance of the sub- laboratory animals. stituent attached to the nitrogen was the lack of At normal doses, codeine , even more so than recognition of naltrexone which is structurally morphine and despite its capacity to release his- the same as naloxone except for the presence of tamine in the skin (but perhaps also refl ecting the an N -cyclopropylmethyl instead of an N -allyl differences in parenteral administration), has substituent. Surprisingly, meperidine and metha- rarely been implicated in anaphylactoid or ana- done showed signifi cant inhibition of IgE bind- phylactic reactions. However, an unusual case of ing but like morphine and codeine, meperidine fever with urticaria, generalized pruritus, and

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Inhibition (%) of IgE antibody Inhibition (%) of IgE antibody Compound Structure binding to morphine-Sepharose Compound Structure binding to morphine-Sepharose with 200 nmol of compound with 200 nmol of compound

HO HO * Buprenorphine O 31 O Morphine 72 NCH2 NCH3 H3CO HO

H3CO CH2CH2 N Fentanyl 3 Codeine O 72 NCH3 N CCH2CH3 HO O

CH3 HO N

O CH OH Nalorphine O 46 Hyoscine 2 9 NCH2CH=CH2 OCHC O HO

HO CH3 N Naloxone O OH 15 Meperidine 31 NCH2CH=CH2 COOCH2CH3 O

O O OH Naltrexone 3 Methadone CH3CH2C CCH2CHN(CH3)2 31 NCH2 CH3 O

* Buprenorphine contains an endoetheno bridge between C6 and C14 and a 1-hydroxy-1,2,2-trimethylpropyl substitution on C7.

Fig. 8.6 Comparative inhibitory potencies of morphine Implication of IgE Antibodies that React with Morphine and some other opioids in quantitative hapten inhibition and Codeine, and Identiﬁ cation of an Allergenic studies with serum containing IgE antibodies to papaver- Determinant. Anesthesiology. 1989;71:489. Reproduced etum (see Fig. 8.5 ). From Harle DG et al. Anaphylaxis with permission from Lippincott Williams & Wilkins following Administration of Papaveretum. Case Report:

erythema occurring 6 h after ingestion of a single and, more recently, this recommendation has tablet of 30 mg of codeine has been reported. been extended to adults following the reporting Specifi city of the reaction was confi rmed by oral of life-threatening hypotension in three patients challenge 1 month later with 5 mg of codeine given codeine phosphate intravenously. phosphate when the patient responded 3 h later Despite heroin’s notoriety and its widespread with hot and cold sensations, oral pruritus, gener- misuse intravenously, allergic reactions to the alized urticaria, palpebral and labial edema, and drug appear to be far less common than one might petechia. At higher doses maculopapular and expect. A diagnosis of anaphylaxis, confi rmed by urticarial rashes, generalized dermatitis, and a positive tryptase test at the time and positive eczema are seen. Codeine has also been reported prick testing several weeks later, was made on a to cause pseudo scarlet fever and adverse reac- patient following intrathecal injection of diamor- tions ranging from tachycardia and cutaneous phine. In recent years there have been a number vasodilation to severe hypotension and apnoea in of reports of heroin inhalation inducing or exac- children given codeine phosphate intravenously. erbating life-threatening asthma. In one case AMA Drug Evaluations , the American Medical diagnosed by bronchial provocation testing, Association’s evaluation of the most commonly inhalation of heroin powder provoked allergy prescribed drugs, concluded that codeine phos- involving asthma and urticaria. No primary cause phate should not be used intravenously in children can be identifi ed in a signifi cant number of deaths

[email protected] 8.5 Allergenicity of Opioid Analgesic Drugs 311

a 17N CH3

16 10 D 9

1 11 15 14 8 B

2 A C 7 12 13

E 3 4 56

OH O OH

Fig. 8.7 Two-dimensional (a ) and space-fi lling three- hexenyl ring (ring C) are both shown in green ; the double dimensional ball-and-stick (b , left hand side) and CPK bond at positions 7–8 of ring C is in magenta ; the (b , right hand side ) molecular models of morphine show- hydroxyl at C6 is red ; and the tertiary methylamine at ing in color the identifi ed allergenic determinant recog- position 17 is blue . From Baldo BA, Pham NH. nized by the IgE antibodies in the papaveretum-allergic Histamine-releasing and allergenic properties of opioid patient (refer Figs. 8.5 and 8.6 ). The individual fi ne-struc- analgesic drugs: Resolving the two. Anaesth Intensive tural features of the determinant are shown in different Care. 2012 ;40:216. Reproduced with permission from colors: the piperidine ring (ring D) attached to the cyclo- Australian Society of Anaesthetists in heroin drug addicts and speculation that tions (refer Sects. 6.1.4.2 and 7.4.3.2 ), without anaphylaxis might be involved in at least some tests for morphine- or heroin-specifi c IgE anti- cases remains. Morphine is known to bind to the bodies, there is still no direct evidence for the gamma-globulin fraction of serum from addicts involvement of immunological processes, that is, so it is speculated that its close analog diamor- type I allergy, in the deaths. phine, complexed with serum proteins, might act as a sensitizing allergen in some users. Mast cell tryptase determinations on postmortem blood 8.5.2 Synthetic Opioid Drugs samples from addicts and in immunohistological investigations demonstrated elevated tryptase Meperidine, methadone, and fentanyl (and its concentrations, and it was concluded that many close analogs alfentanil, remifentanil, and sufent- of the fatalities might be due to drug-induced anil) are widely used but again allergic reactions direct liberation of mediators from mast cells. are rare. In what may be the fi rst reported case of However, given that tryptase release has not been true type I allergy to meperidine, an intramuscu- totally disproved in some anaphylactoid reac- lar injection of 100 mg of the drug to a woman in

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Table 8.6 Demonstration of cross-reactive recognition of morphine, meperidine, and fentanyl by human IgE antibodiesa Serum 1 Serum 2 Inhibition (%) of Inhibition (%) of Drug used for Amount (nmol) for IgE-bindingb with Amount (nmol) IgE-bindingb with inhibition 50 % inhibition 200 nmol of drug for 50 % inhibition 200 nmol of drug Morphine 24 72 <1 98 Meperidine 830 31 22 74 Fentanyl >2,000 3 43 62 Nalorphine 240 46 58 57 Naltrexone >2,000 3 ~800 17 Data adapted from Baldo BA et al. Chemistry of drug allergenicity. Curr Opin Allergy Clin Immunol. 2001;1:327. With permission from Wolters Kluwer Health a IgE antibodies detected in two different sera from patients with suspected allergy to opioid analgesic drugs b IgE antibodies detected by binding to a morphine–solid phase complex labor induced a dramatic fall in blood pressure, cases, immunoglobulin E antibodies occur that cyanosis, and urticarial rash. A further 50 mg of detect cross-reactive allergenic determinants on meperidine after recovery again led to hypoten- the naturally occurring phenanthrene morphine sion. Skin tests or specifi c IgE antibody assays and some of the opioid synthetic drugs. have confi rmed other immediate allergic reac- Fentanyl, perhaps again refl ecting usage, is tions to meperidine, but in what appears to be the the subject of most of the few reports of anaphy- only claim of methadone allergy, no supporting lactoid/anaphylaxis reactions to synthetic opioid diagnostic evidence was advanced to back up the analgesic drugs, but in a number of cases again belief that the observed localized urticaria was no appropriate diagnostic data were advanced to the result of a true type I allergy. Some IgE anti- support the diagnosis of fentanyl-induced ana- bodies to morphine can also cross-react with phylaxis. A curious feature noticed in two differ- meperidine and methadone (see previous section ent anaphylactic reactions to fentanyl was an above and Figs. 8.5 and 8.6 ), while others react apparent delay between the administration of the with meperidine and fentanyl. The latter fi nding drug and the appearance of symptoms. A 15-min was demonstrated in quantitative inhibition stud- gap occurred in one case while in the other, pro- ies with two sera from subjects with “suspected found hypotension was observed only after an narcotic allergy” (Table 8.6 ) and suggests that hour. In the latter case, although hypotension IgE antibodies that react with the synthetic opioid may have occurred immediately and not observed analgesics may occur more often than previously because it was successfully contained by the ini- thought if suitable IgE direct binding and quanti- tial treatment, another possible explanation is the tative inhibition immunoassays are more widely small amount of fentanyl-induced release as a employed to investigate suspected opioid allergic result of the low concentration of the potent drug subjects. A serum from a subject examined for (compared to say morphine) acting at the mast multiple drug allergies provided further evidence cell membrane. that methadone cross-reacts immunologically Tramadol is generally considered to be a with morphine, and IgE antibodies can detect “safe” drug with a low risk (put at less than 0.1 % this. Antibodies in the subject’s serum strongly in one report) of adverse reactions. Eleven cases recognized morphine, codeine, and methadone, of angioedema, possibly related to tramadol, reacted less strongly with nalorphine, and not at were reported in Sweden and in 5 of 28 patients all with fentanyl and the antagonists naloxone (18 %) with chronic urticaria exacerbated by non- and naltrexone (Fig. 8.8). The inhibitory poten- steroidal anti-infl ammatory drugs, tramadol- cies for morphine, codeine, and methadone were induced urticaria with one of the patients of the same order again indicating that, in rare developing laryngeal edema. Hypersensitivity

[email protected] 8.6 Resolving the Histamine- Releasing and Allergenic Effects in Diagnosing Reactions to Opioid Drugs 313

Fig. 8.8 Immunological cross-reactivity between metha- mechanism of multiple drug allergies. Structural basis done, morphine, and codeine demonstrated by serum of drug recognition. J Immunoassay Immunochem. IgE antibodies. Serum was from a patient showing multi- 2001;22:47. Reproduced with permission from Taylor & ple drug allergies. From Pham NH et al. Studies on the Francis pneumonitis and a maculopapulous toxic skin guishing a pseudoallergic reaction resulting reaction with secondary erythroderma have also from the direct release of histamine from a true been ascribed to the drug. Assessment of the pos- anaphylactic reaction lies in the application of sible involvement of tramadol in Stevens–Johnson diagnostic investigations that reveal underlying and Lyell’s syndromes showed a high univariate immunological processes. Table 8.7 that sum- relative risk for recent use, but comedication with marizes a number of cases where reactions to highly suspected drugs led the authors of the opioid analgesics were described, compares the study to “doubt causal association for tramadol.” diagnostic conclusions of the authors of each In general, skin reactions caused by tramadol are study with a retrospective diagnostic assessment said to be uncommon and usually benign. based on our necessary information requirements and tests for classifying a reaction as anaphylactic, that is, a true type I IgE antibody- 8.6 Resolving the Histamine- mediated allergic response. These requirements Releasing and Allergenic (see also Chap. 4 ) are: Effects in Diagnosing • A carefully gathered and recorded history and Reactions to Opioid Drugs lists of the drugs administered and the drug- induced signs and symptoms. In reviewing the published ﬁ ndings on adverse • The use of appropriate dilutions of suspected reactions induced by all the opioid analgesics drugs in skin tests (skin prick and/or intrader- implicated so far, it is clear that the key to distin- mal tests) and/or placebo-controlled challenge

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dent skin prick test dently rmly estab- rmly SPT ed to confi rmatory tests lacking Retrospective Retrospective assessment of diagnosis and comments Anaphylactoid reaction. No mediated of immune- evidence reaction tests to check for irritant reaction not done confi could be Anaphylactic but cross-reaction with MNBDs Absence of appropriate test results precludes confi diagnosis Diagnosis not fi lished. Could be anaphylactoid. skin test for No positive of LTT Relevance fentanyl. and meperidine SPT? Conc. of meperidine SPT? Inconclusive. No immune basis Inconclusive. identifi diagnose anaphylaxis neuromuscular blocking drug, NMBD Direct effect of morphine. Direct effect 1 reaction unlikely Type Anaphylactic reaction Anaphylactic reaction. Skin Anaphylactic reaction Anaphylactic reaction to Anaphylactic reaction fentanyl Anaphylaxis to fentanyl Anaphylaxis to fentanyl and propofol meperidine are are are ness to “Hypersensitivity” BP, face puffi face BP, 2 cm wheal Anaphylactoid reaction. 1 cm wheal Morphine and codeine positive 5 mm wheal and fl 6 mm wheal and fl to fentanyl and to fentanyl suxamethonium Positive for fentanyl for fentanyl Positive and proposol for meperidine Positive ↓ lymphocyte transformation test, lymphocyte LTT M) M) −2 −6 intradermal test, IDT IgE Abs with morphine– solid phase Diagnostic tests employed Test results Authors’ diagnosis IDT with fentanyl IDT with fentanyl 5 ng/ml 0.5 ng/ml IDT with morphine (0.35 × 10 Tryptase Tryptase test Positive Anaphylaxis Scratch and IDT LTT wheal and fl Positive Possibly anaphylaxis SPT but None – Anaphylactoid reaction Anaphylactoid or anaphylactic. IDT with morphine (0.3 × 10 Challenge with 50 mg meperidine heart rate, HR culties, ushing BP, BP, ↓ HR, pulse, shortness pulse, cyanosis, pulse, cyanosis, ↑ ↑ ↓ BP, BP, sinus tachycardia, BP, BP, BP, BP, BP, blood pressure, Symptoms erythema, urticarial rash, cyanosis Hypotension, tachycardia, fl ↓ tachycardia ↓ wheezing wheezing, frothy secretions ↓ of breath, wheezing Hypotension, sweating, nausea, palpitations ↓ urticarial rash BP g IV g Delayed g μ decreased, μ ↓ Summary of some case studies of anaphylactoid/anaphylactic reactions to opioid analgesic drugs and diagnostic conclusions Summary of some case studies anaphylactoid/anaphylactic reactions to opioid analgesic drugs and diagnostic conclusions Elevated, Elevated, Table 8.7 Drug Morphine 0.3 mg/kg IV at 10 mg/min Morphine 10 mg bolus IV Fentanyl 50 Morphine 3 mg bolus IV Fentanyl Fentanyl 100 (no details) Fentanyl Breathing diffi Papaveretum–hyoseine Papaveretum–hyoseine IM Meperidine 100 mg IM ↑

[email protected] 8.7 Why Are Opioid Analgesic Drugs So Poorly Allergenic? 315

tests. Skin tests with OADs are carried out assessment of 975 so-called “immediate with the knowledge that they are, in general, anaphylactoid reactions” to parenterally adminis- histamine releasers; some are more potent in tered anesthetic drugs where the mechanism was doing this than others and therefore the dilu- confi rmed in only half of the patients. tions of drug used are critical. The concentrations (mg/ml and molarity) of the fi ve most commonly used OADs that are normally non- 8.7 Why Are Opioid Analgesic reactive in the skin of normal, healthy subjects Drugs So Poorly Allergenic? are for skin prick and intradermal testing, respectively: OADs are some of the most commonly adminis- Morphine 1 mg/ml, 3.5 × 10 −3 M; 10 μg/ml, tered drugs in hospitals and together with their 3.5 × 10 −5 to 10 −7 M frequent prescribing (e.g., morphine, codeine, Fentanyl 0.05 mg/ml, 1.5 × 10 −4 M; 5 μg/ml, and tramadol) in the community worldwide, and 1.5 × 10 −5 M their rampant misuse by addicts, the question of Alfentanil 0.5 mg/ml, 1.2 × 10 −3 M; 50 μg/ml, why IgE-antibody-mediated allergic reactions to 1.2 × 10 −4 M them are so rare is intriguing. Numerous surveys Remifentanil 0.05 mg/ml, 1.3 × 10 −4 M; 5 μg/ml, have shown that OADs account for only about 1.3 × 10 −5 M 1 % of anaphylactic/anaphylactoid reactions dur- Sufentanil 0.005 mg/ml, 1.3 × 10−5 M; 0.5 mg/ml, ing the perioperative period ranking them behind 1.3 × 10−6 M colloids, hypnotics, antibiotics, latex, and A histamine control should be included and NMBDs, the most frequently involved group of drugs should also be tested for histamine releas- agents. It has long been recognized that “small” ing/nonspecifi c irritant effects in the skin of nor- organic molecules of molecular weight less than mal subjects. about 1,000 Da need to be presented to the • Determination of serum tryptase levels sam- antibody-forming cellular machinery as haptens pled at appropriate time intervals preferably bound to macromolecular carriers (usually pro- within 6–8 h of the reaction. More than one teins) for antibody formation to result, although sample should be taken if possible. this may not always hold true (see Sect. 3.1 ). • Application of specifi c IgE antibody immuno- Morphine is known to bind to the γ-globulin frac- assays for individual drugs together with tion of human serum, morphine-reactive antibod- appropriate inhibition studies to confi rm species have been found in normal, unimmunized ifi city and identify possible cross-reactive subjects, and morphine coupled to carrier prodrugs. teins have readily induced IgG and IgM antibody The selected examples summarized in responses in laboratory animals. All this suggests Table 8.7 are only a few of many case studies that that morphine, and probably other opioid drugs, show confusion with, or a misunderstanding of, are not lacking immunogenicity. Although there the terms “anaphylactoid” and “anaphylactic.” is only a single report of the identifi cation of, and The studies reveal the often inadequate or even specifi city study on, an opioid drug-reactive IgE complete absence of, appropriate diagnostic antibody in a patient allergic to an OAD(s) (see investigations necessary for an accurate diagno- Sect. 8.5.1 and Figs. 8.5 , 8.6 , and 8.7 ), this may sis. In the absence of evidence defi ning the under- be as much a refl ection of the lack of immuno- lining mechanism of a reaction, it is likely that chemically based investigations in the drug some, if not many, reactions caused by direct allergy fi eld as an indication of the rarity of such release of histamine and those that were medi- antibodies. For allergen-induced release of the ated by IgE antibodies have been misdiagnosed. mediators of allergy from mast cells and baso- This conclusion is supported by a search of the phils, cross-linking by multivalent allergens of English and French language literatures over the FcεRI receptor-bound complementary IgE anti- 20-year period 1964–1984 and a retrospective bodies on the cell surfaces is required. Drugs

[email protected] 316 8 Opioid Analgesic Drugs might do this if they are at least bivalent in terms collapse is more common in true anaphylactic of their allergenic determinant sites, for example responses and cutaneous symptoms more fre- the NMBDs, or if they are in a multivalent form quently seen in anaphylactoid reactions. bound to a macromolecular carrier (see Sects. Historically, in prescribing an alternative 3.1.2 and 7.4.2.3). It seems likely, if not clear, OADs in cases of known existing hypersensitiv- that OADs are not allergenically bivalent and ity to an OAD, the phenanthrene drugs on the one cross-linking of cell-bound IgE antibodies by any hand and the synthetics meperidine, methadone, OAD has yet to be shown to occur. and fentanyl on the other have been grouped Opioids are said to modulate both innate and together, and it is usually recommended that a acquired immunity, and among a vast array of member of the opposite group should replace a claimed effects on immune cells, the drugs have known non-tolerated opioid. However, possible been shown to suppress antibody formation by allergenic cross-reactivities among all of the human B lymphocytes in vitro. Developments in OADs should be kept in mind. Although cross- this area of research may be relevant to the rare reactivity between the structurally similar natu- and infrequent occurrence of allergic antibodies rally occurring alkaloids morphine and codeine to OADs in humans and are awaited with interest. and their semisynthetic oxy- and hydro-deriva- A more conventional explanation than suppres- tive analogs is to be expected, cross-reactions sion of antibody formation for this apparent between these morphine analogs and the synthet- restricted immune response may be that OADs ics meperidine, methadone, and fentanyl might are seen as “self” components and are not seen as be much more common than currently believed. foreign antigens. Opioid peptides and the natural Some compelling evidence for this has been pre- and synthetic OADs show the same biological sented (see above under Sect. 8.5.2 ). Before an activities in vivo and act on the same receptors, alternative opioid drug is selected, it should be so it seems possible that from the immunological used to skin test the patient. perspective, the shape and conformation in space If appropriate OAD-reactive IgE antibody of the endogenous and exogenous opioids might tests are available, they should be used as an be seen as essentially the same. adjunct to skin testing but an understanding of the immunochemical basis of cross-reactivity between opioids and NMBDs shows that a cau- 8.8 Some Important Clinical tionary approach to interpretation is necessary. Implications Related to the For example, a morphine solid phase is routinely Use of Opioid Analgesic Drugs employed to detect NMBD-reactive IgE antibodies in the sera of subjects allergic to NMBDs (see Because the signs and symptoms of an OAD- Sect. 7.4.3.4.2). The structural basis of the cross- induced pseudoallergic reaction and a true, recognition is the tertiary methyl ammonium antibody-mediated allergic reaction to an OAD group of morphine and the substituted ammo- can be so similar, distinguishing them is not nium groups on NMBDs. always easy unless appropriate tests to identify an When an OAD-induced reaction is known or underlying immunological mechanism are suspected to be due to histamine release and the applied. Routine application of skin tests with continued administration of an OAD is judged to controls and/or placebo-controlled challenge tests be necessary, pretreatment with H1 and H2 (and, in with the suspected drug(s), serum tryptase deter- future, perhaps an H4 ) histamine antagonists is an minations, and speciﬁ c immunoassays to detect option. Some opioid analgesics show differences drug-speciﬁ c IgE antibodies will take much of the in the amount of histamine they release and in the uncertainty out of resolving the different responses. anatomical site where release occurs. Substitution If bronchospasm occurs in a reaction to an of OADs can be made in both cases: for example, OAD, the reaction is more likely to involve OAD- fentanyl for morphine in the former case and fen- reactive IgE antibodies. Likewise, cardiovascular tanyl or morphine for buprenorphine if histamine

[email protected] Further Reading 317 release occurs in the lungs and fentanyl for • Cardiovascular collapse and bronchospasm morphine following cutaneous release. If an alter- are more frequent in anaphylactic reactions to native to an OAD is sought, a nonsteroidal OADs anti-infl ammatory drug or regional analgesic • Cutaneous symptoms occur more often in techniques might be substituted. anaphylactoid reactions • OAD-induced histamine release rarely if ever produces bronchospasm Summary • Adverse skin reactions to OADs sometimes occur but as is the case for type I OAD allergic Histamine releasing (anaphylactoid) and allergic reactions, other hypersensitivity reactions are (including anaphylactic) reactions to opioid anal- rare gesic drugs • Tramadol is generally considered to be a ‘safe’ • Anaphylactoid and anaphylactic reactions to drug with a low risk (< 0.1%) of adverse OADs: same clinical picture; same immediate reactions management • Anaphylactoid and anaphylactic reactions are distinguished by the underlying mechanism: Further Reading Anaphylactic reactions are immune (IgE antibody)-mediated; anaphylactoid reactions Baldo BA, Pham NH. Histamine-releasing and allergenic properties of opioid analgesic drugs: resolving the are not two. Anaesth Intensive Care. 2012;40:216–35. • Reactions can be distinguished by appropri- Casy AF, Parfi tt RT. Opioid analgesics. Chemistry and ately diluted skin tests with suspected drug(s) receptors. New York, NY: Plenum Press; 1986. and specifi c IgE antibody tests Eguchi M. Recent advances in selective opioid receptor agonists and antagonists. Med Res Rev. 2004;24: • Tryptase assays are useful but may not distin- 182–212. guish anaphylactic and anaphylactoid reactions Freye E. Opioids in medicine. Dordrecht: Springer; 2008. • Potency of the direct histamine releasing Grond S, Sablotzki A. Clinical pharmacology of trama- effect of different OADs varies widely dol. Clin Pharmacokinet. 2004;43:879–923. Portoghese PS. From models to molecules: opioid recep- • Direct release of histamine may provoke an ana- tor dimers, bivalent ligands, and selective opioid phylactoid reaction especially if the OAD is receptor probes. J Med Chem. 2001;44:2259–69. given IV as a bolus, but correlation between Trescot AM, Datta S, Lee M, Hansen H. Opioid pharma- plasma histamine levels and hypotension is poor cology. Pain Physician. 2008;11:S133–53. Wu Z, Hruby VJ. Backbone alignment modeling of the • OADs are poorly allergenic and anaphylactic structure-activity relationships of opioid ligands. J and other less severe type I reactions are also Chem Inf Model. 2011;51:1151–64. rare Yaksh TL, Wallace MS. Opioids, analgesia and pain man- • Clinical manifestations are generally more agement. In: Brunton LL, Chabner BA, Knollmann BC, editors. Goodman and Gilman’s the pharmaco- severe and more often life-threatening in ana- logical basis of therapeutics. 12th ed. New York, NY: phylactic patients McGraw Hill; 2011. p. 481–526.

[email protected] Nonsteroidal Anti-Inﬂ ammatory Drugs 9

Abstract Nonsteroidal anti-inﬂ ammatory drugs (NSAIDs) are responsible for 20–25 % of ADRs. Cyclooxygenase isoenzymes COX-1 and COX-2 cata-

lyze the formation of PGG2 from arachidonic acid. Ultimate products of the

metabolic pathway are PGD2 , PGE2 , PGF2 α, PGI2 , and TXA2 . NSAIDs can be classiﬁ ed on the basis of their COX inhibitory and selective properties. Most are mainly COX-1 inhibitory, e.g., aspirin and ibuprofen, while some are COX-2 inhibitory, e.g., celecoxib. NSAIDs with the highest GI toxicity have the highest COX-1 selectivity. COX-2 is expressed in inﬂ ammation and selective COX-2 inhibitors show fewer GI effects but can produce cardiovascular effects. The mechanism of NSAID-induced respiratory reactions appears to be due to the redirection of arachidonic acid metabolism from the COX to the lipoxygenase synthetic pathway with associated production of cysteinyl leukotrienes. Aspirin-induced asthma, which makes up 3–5 % of adult asthmatics, has symptoms of chronic asthma, rhinosinusitis, and nasal polyps. NSAID-induced cutaneous reactions occur in a number of different clinical patterns. Challenge testing is the only way to diagnose sensitivity to an NSAID. Desensitization can be induced by repeated oral administration. Delayed reactions are seen and may take the form of contact dermatitis, FDE, DRESS, AGEP, SJS, TEN, or nephritis.

Anti-infl ammatory drugs have a special, if not of willow bark and leaves as an analgesic. During pivotal, place in the history of Western medicine the time of Galen and thereafter, willow became and drug therapy and in the birth and evolution of widely used throughout the Roman world, an the pharmaceutical industry. From at least the important component of the materia medica of time of early Chinese and Egyptian civilizations Western medicine and a standard entry in the over 5,000 years ago, humans sought plants and early pharmacopoeias. After more than two mil- other natural materials for the relief of the four lennia of the use of barks of various Salix spp. for signs of infl ammation—redness, heat, swelling, pain, fever, and infl ammation in both Europe and and pain. We know that as early as 400 BC, the the new world, salicin was isolated in 1828 and Greek physician Hippocrates advocated the use by the mid to late nineteenth century the

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 319 Relationships, DOI 10.1007/978-1-4614-7261-2_9, © Springer Science+Business Media, LLC 2013

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properties and value of salicylate medicine were conditions involving pain including headache, becoming increasingly appreciated. The chemi- migraine, pain due to infl ammation and tissue cal and early pharmacological work on salicin, injury, dysmenorrhoea, muscle strains, surgical, salicylic acid, and sodium salicylate led to the dental and postoperative pain, and so on. The production of salicylic acid by 1874 and the drugs are also commonly taken for colds and introduction of acetylsalicylic acid, named aspi- infl uenza to relieve fever, temperature, and pain. rin, by Bayer in 1899. In the twentieth century, Important indications for prescribed NSAIDs are other organic acids and later nonacidic anti- rheumatoid arthritis and osteoarthritis where, in infl ammatory compounds followed and, although the former case, anti-infl ammatory action is the mechanisms of action of aspirin and other refl ected in reductions of pain, swelling of joints anti-infl ammatories in the pre-prostaglandin age and morning stiffness, increased mobility, and remained obscure, usage of what became known enhanced functional capacity (such as grip as the nonsteroidal anti-infl ammatory drugs strength). No single NSAID has stood out as a (NSAIDs) grew until this group was among the superior drug for rheumatoid arthritis, rather, most commonly used drugs worldwide. individual patients may respond better or worse Severe bronchospasm following the ingestion to certain members of the group. For osteoarthri- of aspirin was described soon after its introduc- tis, improvements may occur in pain at rest, tion. Now, with NSAIDs said to be used by more stiffness, pain from movement, and night pain, than 30 million people every day, 111 million and movement overall may become easier. Again, prescriptions written annually in the USA and the no individual NSAID seems superior, but indo- drugs making up approximately 60 % of the USA methacin should be avoided because of its over-the-counter analgesic market, NSAIDs are greater toxicity and its possible potential for said to be responsible for 21–25 % of reported accelerating progression of the disease. NSAIDs adverse drug events. This chapter will present a are also widely administered for infl ammatory summary of important NSAIDs, their mechanism arthropathies such as ankylosing spondylitis and of action, the clinical manifestations of NSAID Reiter’s syndrome, for acute gout, metastatic sensitivity reactions, diagnostic tools for, and bone pain, renal colic, and postoperative pain, management of, the reactions, and the pathomech- sometimes as a substitute for poorly tolerated anisms thought to be involved. The therapeutic opioid analgesics. and chemical properties of these drugs and their mechanism of action are presented before the clinical picture and pathogenesis of intolerances 9.2 Patient Usage to NSAIDs are discussed later in this chapter. Over-the-counter usage of NSAIDs is common in most countries, although one survey reported that 9.1 Therapeutic Applications about half of NSAID users were unaware of the of NSAIDs drugs’ potential side effects; over-dosing is more common with over-the-counter than with pre- NSAIDs have analgesic and anti-infl ammatory scription users and approximately 40 % of pre- properties and are, therefore, widely used for the scription users take over-the-counter NSAIDs at treatment of both minor and more severe pain and the same time. A survey in the USA found that for the management of swelling and joint infl am- approximately 30 % of adults take over-the- mation. Some NSAIDs also possess antipyretic counter analgesics on a regular basis for chronic activity and are, therefore, useful for the treat- pain. Use for this purpose increases with age—an ment of fever. Specifi cally, NSAIDs available estimated 10–40 % of people over 65 years take over the counter such as aspirin and ibuprofen are prescribed or over-the-counter NSAIDs on a heavily used for self-medication of a host of daily basis. Ibuprofen and diclofenac are the most

[email protected] 9.3 Classifi cation of NSAIDs 321 commonly consumed NSAIDs throughout the turn of the nineteenth century by its acetylated world accounting for 40 % of global sales. derivative, acetylsalicylic acid or aspirin Following, in order, are naproxen, meloxicam, (Fig. 9.1a ). Esterifi cation of the phenolic celecoxib, ketoprofen, and etorocoxib. For pre- hydroxyl group and substitution of a hydropho- scription drugs, ibuprofen and naproxen head the bic group at C-5 enhances the pharmacological list in the USA while diclofenac is the leader in action of the anti-infl ammatory salicylates. the UK. Ibuprofen leads over-the-counter NSAID sales in the USA. An extraordinary increase in NSAID use 9.3.2 Propionic Acid Derivatives occurred with the introduction of the COX-2- selective drugs but, following the withdrawal of These drugs are sometimes referred to as “pro- rofecoxib and valdecoxib in 2004–2005, there fens,” the term originating from the name of the was a marked reduction in the prescribing of all group’s original member, ibuprofen (Fig. 9.1b ). NSAIDs. Between 2003 and 2005, the decrease Like the salicylates, they are organic acids in usage of COX-2-selective agents for arthritis (pKa = 3–5) and form soluble salts. Other members in the USA went from 55.1 to 29.2 % while in of the group include naproxen, fenoprofen, keto- Germany, COX-2 prescription numbers dropped profen, dexketoprofen, carprofen, fl urbiprofen, by 37.1 million between 2004 and 2005, and dexibuprofen, loxoprofen, and oxaprozin. Although overall NSAID usage decreased by 8.4 %. the drugs, except for naproxen, are usually marketed as racemates, in vivo they invert from the inactive R-enantiomer to the active S-enantiomer 9.3 Classifi cation of NSAIDs and this is the isomer found in plasma.

It is informative to classify NSAIDs on the bases of both their pharmacological and chemical prop- 9.3.3 Aryl and Heteroaryl Acetic erties but, before considering pharmacological Acids mechanisms of these drugs, a classifi cation based on chemical structures is presented. Many This group can be further subdivided into indene/ NSAIDs are organic acids with pKas between indoles, pyrroles, and oxazoles. Indomethacin about three and fi ve. Many are carboxylic acids and sundilac are the most well known of the and therefore form salts that ionize at physiologi- indene/indole group. Indomethacin (Fig. 9.1c ) is cal pH. The acidic group also accounts for the administered for rheumatoid arthritis, osteoar- ionic binding with plasma proteins generally thritis, and ankylosing spondylitis and to sup- seen with NSAIDs, and it serves as a major site press uterine contraction but has a number of for the metabolism and clearance (e.g., as gluc- undesirable actions including gastrointestinal uronides) of the drugs. By virtue of their aryl ulceration and hemorrhage, CNS effects, aplastic groups and hydrophobic substituents, NSAIDs anemia, bone marrow suppression, and thrombo- show varying degrees of lipophilicity. cytopenia. Sulindac (Clinoril®), which lacks the indole nitrogen of indomethacin, is a prodrug that is 50 times as active in its metabolized sulfi de 9.3.1 Salicylates form. A third well known drug in this group is etodolac (Lodine®). Tolmetin (Tolectin®), with a The salicylate NSAIDs, for example, aspirin, short half-life of less than 5 h is perhaps the best difl unisal, and salsalate, are derived from the rel- known pyrrole acetic acid NSAID while oxapro- atively strong organic acid, salicylic acid, used zin (Daypro®) is an example from the oxazole fi rst as the sodium salt and then replaced at the class of acetic acids.

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O O a b CH CH 6 3 3 1 C C 5 OH OH OH OH C CH C 2 3 4 O O OH O CH3 H C 3 C R 3 O General Salicylic acid Acetylsalicylic acid Ibuprofen structure Salicylates Propionic acid derivatives c OH OH d O O H3CO C R C C C O O OH OH N O NH2 NH General Anthranilic acid structure R (R=aryl or Cl heteroaryl) Indomethacin General anthranilate structure Aryl & hetero arylacetic acids O C e OH OH O NH C N N CH H 3 N S CH3 O O CH3

Piroxicam Mefenamic acid Oxicams (Enolic acid derivatives) Anthranilates (Fenamic acid derivatives)

O O O f g h OO O H C H S N CH3 N CH3 H3C S H3C N O NH2 O N N N

R NO2 CF3 Phenylbutazone General structure Nimesulide Celecoxib of anilides

Phenylpyrazolones Anilides COX-2-selective inhibitors

Fig. 9.1 Classiﬁ cation of nonsteroidal anti-inﬂ ammatory in bold in category a and the central core structure of a drugs based on chemical structure with important exam- diaryl-5-membered heterocyclic ring with two adjacent ples from each category. The structure common to sali- phenyl substituents is highlighted in category h cylic acid and acetylsalicylic acid (aspirin) is highlighted

[email protected] 9.3 Classiﬁ cation of NSAIDs 323

9.3.4 Anthranilates (Fenamic Acid agranulocytosis, bone marrow suppression, and Derivatives) bleeding. Oxyphenbutazone, a metabolite of phenylbutazone was also used as a NSAID. Other Also known as fenamates, these compounds, phenylpyrazolones in use include phenazone, pro- which include mefenamic acid (Ponstel®; Ponstan®) pyphenazone, aminophenazone, and metamizole. (Fig. 9.1d ) and meclofenamic acid (Meclomen), are N -aryl derivatives of anthranilic acid. Note that anthranilic acid is a bioisostere of salicylic acid 9.3.7 Anilides and diclofenac is a derivative of 2-arylacetic acid. The presence of small alkyl groups or halogen These drugs are simple acetamides of aniline atoms on the N -aryl ring increases activity, for (Fig. 9.1g) with the heavily used acetaminophen example, meclofenamate with Cl at positions 2, 5, (paracetamol) and phenacetin (now rarely, if ever, and 6 on the ring is 25 times as potent as mefe- used) being the best known examples. They are namic acid. Other anthranilates administered are neutral compounds without an acid group and fl ufenamic acid and tolfenamic acid. The anthrani- with little if any inhibitory activity for cyclooxy- late NSAIDs are well absorbed from the gastroin- genase. Their mechanism of action is different to testinal tract but produce a number of adverse other NSAIDs and their classifi cation here along reactions including ulceration, diarrhea, nausea, with other groups of NSAIDs is not a perfect fi t. vomiting, headache, and hematopoietic toxicity. Nevertheless, acetaminophen is now an important and widely used drug and is often substituted for NSAIDs that are not tolerated (see below). 9.3.5 Oxicams (Enolic Acid Discovered as part of a program to identify anti- Derivatives) infl ammatory drugs based on sulfonamides and developed as a clinically effective drug in the The usual carboxyl group seen in many NSAIDs 1960s before the COX-isoforms were discovered, is absent from the oxicams which, for compounds nimesulide (4-nitro-2-phenoxymethane sulfo- such as piroxicam and meloxicam, derive nanilide) (Fig. 9.1g ) is a COX-2-selective NSAID their acidity from the 4-hydroxyl group. Both of with analgesic and antipyretic properties used for these drugs are characterized by a a number of conditions including the treatment of 4- hydroxybenzothiazine group as shown in the acute pain and osteoarthritis. This drug, now in structure of piroxicam (Fig. 9.1e). Lack of a car- the European Pharmacopoeia, appears to have boxyl substituent means that the drugs cannot be more than one mechanism of action in addition to readily glucuronidated and excreted, and this its inhibitory activity of the COX-2 isoenzyme. In results in a longer half-life and once a day dosing. a 2006 assessment of the clinical and safety pro- Other administered drugs in this group are tenoxi- fi les of numesulide, it was concluded that the cam, droxicam, lornoxicam, and isoxicam. drug is an effective and safe therapeutic choice for the treatment of various painful infl ammatory conditions, and it has a rapid onset of action and 9.3.6 Phenylpyrazolones an overall positive to risk profi le.

Viewing aspirin as the progenitor drug, the ﬁ rst member of what we now call the NSAIDs is phen- 9.3.8 COX-2 Selective Inhibitors ylbutazone (Fig. 9.1f), a compound with a 1-aryl- 3,5-pyrazolidinedione structure. Introduced by The enzymes that catalyze the synthesis of Geigy in Basel in 1946, phenylbutazone was prostaglandins from arachidonic acid are extensively used for over 30 years for arthritis known as cyclooxygenases, and the term COX and other pain until replaced by newer NSAIDs refers to cyclooxygenase activity. By convention, and because of its adverse effects that included the enzyme responsible for the production of

[email protected] 324 9 Nonsteroidal Anti-Inﬂ ammatory Drugs

prostaglandins and thromboxane A2, important infl ammation, pyrexia, asthma, osteoporosis, for the regulation of stomach and kidney blood cardiovascular disease, uterine function, and can- fl ow and gastric acid secretion as well as homeo- cer. NSAIDs, a chemically diverse range of drugs static maintenance, is termed COX-1. COX-2, with common analgesic, anti-infl ammatory, and induced by various infl ammatory stimuli, is also antipyretic properties exert their pharmacological responsible for prostaglandin production and action by interfering in the arachidonic biosyn- plays an important part in pain and infl ammatory thetic pathways that produce the prostanoids. processes. COX-2 inhibitors (often termed coxibs from cox -inhib itors) are diaryl-substituted pyr- azoles/furanones/oxazoles. Celecoxib (Fig. 9.1h ), 9.4.1 Biosynthesis of the for example, has a central pyrazole ring con- Prostanoids nected to two phenyl rings, one with a methyl and the other a sulfonamide group. Rofecoxib has a Arachidonic acid, an unsaturated 20-carbon fatty central furanone ring and valdecoxib a central acid located in the cell membrane as a phospho- oxazole ring. The diaryl tricyclic structure with a lipid is released by either secretory or cytoplas-

fi ve-membered heterocyclic ring and a phenyl mic phospholipases (sPLA2 or cPLA2 ). ring with a sulfonamide or methylsulfone sub- Cyclooxygenase (COX) or prostaglandin H2 syn- stituent are critical in determining specifi c COX-2 thase (PGHS) catalyses the fi rst two steps in the inhibition. The central core of a diaryl-5- biosynthesis. There are two PGHS isoenzymes, membered heterocyclic ring with two attached PGHS-1 (COX-1; prostaglandin H synthase 1) phenyl rings found in the COX-2 selective inhibi- and PGHS-2 (COX-2; prostaglandin H synthase tors is highlighted in Fig. 9.1h . Coxibs have high 2) which exhibit two different but complemen- pKa values (8–9) compared to conventional tary enzymatic activities—a cyclooxygenase that NSAIDs, are well absorbed providing peak catalyzes the formation of the hydroperoxy endo- plasma levels within 3 h, and have a relatively peroxide prostaglandin G2 (PGG2 ) from arachi- long duration of action due to slow clearance. donic acid and two molecules of oxygen (O2 ) and COX-2 inhibitors are used to treat pain and a peroxidase that facilitates its subsequent reduc- infl ammation in acute and chronic conditions like tion to the hydroxyendoperoxide PGH2 . PGH2 is rheumatoid arthritis, osteoarthritis, surgical, den- then converted by specifi c synthases and isomer- tal and postoperative pain, acute injuries, and ases to fi ve biologically active primary pros- dysmenorrhea. Besides celecoxib, other COX-2 tanoids, PGD2 , PGE 2 , PGF 2α, prostacyclin PGI 2 , inhibitors include rofecoxib, valdecoxib, and thromboxane A2, TXA 2 (Fig. 9.2 ). Whereas parecoxib, etoricoxib, fi rocoxib, and lumiracoxib. TXA2 is a vasoconstrictor and stimulates platelet

Due to the induction of a so-called cardio-renal activation and aggregation, PGI2 is a vasodilator syndrome involving an apparent high risk of that inhibits platelet activation. This suggests that myocardial infarction, exacerbation of hyperten- in relation to the vascular system, the two highly sion and elevation of blood pressure, rofecoxib, biologically active molecules participate in a bal- and valdecoxib were withdrawn from the market ancing process of homeostasis that normally pro- a relatively short time after their introduction. tects against vascular damage.

9.4 Mechanism of Action 9.4.2 The Cyclooxygenase Isoforms of NSAIDs COX-1 and COX-2

Prostanoids, the prostaglandins, and thrombox- In 1971, Vane and Piper found that NSAIDs ane are end products of fatty acid metabolism inhibit the biosynthesis of prostaglandins from with important physiological functions and arachidonic acid by preventing the substrate from pathological effects in areas relevant to pain, binding to the COX enzyme active site. As it was

[email protected] 9.4 Mechanism of Action of NSAIDs 325

Fig. 9.2 Biosynthetic pathways of the synthesis of prostanoids from arachidonic acid by isoenzymes COX-1 and COX-2 catalysis

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Lipoxygenase Pathway 12-HPETE 12-HETE FLAP ARACHIDONIC 12-Lipoxygenase 5-HPETE LTA4 ACID 5-Lipoxygenase 15-Lipoxygenase 15-HPETE 15-HETE 5-HETE Physiologic Inflammatory LTB , LTC , COX-1 COX-2 4 4 stimulus PGG2 stimulus LTD4, LTE4, E.g.Aspirin E.g.Aspirin Indomethacin Indomethacin Ibuprofen PGH2 Ibuprofen Meloxicam Ketoprofen Numesulide Piroxicam Celecoxib Rofecoxib

TXA2 PGI2

PGF PGD2 2a

PGE2

Fig. 9.3 Overall scheme summarizing the relationship biosynthetic pathways together with intervention of COX between arachidonic acid and the cyclooxygenase, inhibitors and their selectivity properties 5-lipoxygenase, and 15- (and 12-) lipoxygenase (eoxin) known then, the COX enzyme was purifi ed in ation, but these drugs can produce cardiovascular 1976 before being cloned in 1988. Soon after in effects. This is thought to be due to suppression the early 1990s a second isoform of COX, now of the cardio-protective PGI2 . In addition, and termed COX-2, was discovered and, as Vane has unlike the consequences of inhibiting of COX-1, pointed out, in a clear sign of the therapeutic PGE2 and PGI 2 produced via the COX-2 pathway potential of this work, several highly effective add to the infl ammation initiated by mediators anti-infl ammatory agents and new therapeutic such as histamine and bradykinin leading to areas became the subject of investigation within 5 increased vascular permeability and edema. years of the discovery. COX-1 is constitutively expressed at high levels in the endothelium, platelets, monocytes, renal tubules, and seminal 9.4.3 Classifi cation of NSAIDs by vesicles indicating that the prostanoids it helps to COX Isoenzyme Selectivity produce are involved in numerous physiological functions in the gastrointestinal tract, kidney, and NSAIDs can be classifi ed on the basis of their platelets. For example, inhibition of COX-1 COX inhibitory and selectivity properties. In reduces the production of protective PGE2 and general, most NSAIDs inhibit both COX-1 and

PGI2 in the stomach, which may lead to gastric COX-2 to some extent, but most are mainly ulceration. COX-2 expression is induced by cyto- COX-1-selective (e.g., aspirin, sulindac, indo- kines and other inﬂ ammatory mediators such as methacin, ketoprofen, piroxicam); some are interleukin (IL)-1, tumor necrosis factor (TNF), somewhat selective for COX-1 (e.g., ibuprofen, and lipopolysaccharides (LPS) in tissues and diclofenac, naproxen); others are slightly selec- cells as diverse as macrophages, monocytes, tive for COX-2 (etodolac, meloxicam); while endothelial cells, vascular endothelial cells and NSAIDs like lumiracoxib, rofecoxib, and cele- rheumatoid synovial endothelial cells. Since coxib are clear selective inhibitors of COX-2 COX-2 is expressed only in inﬂ ammation, selec- (Fig. 9.3 ). Clearly, knowing the relative inhibi- tive COX-2 inhibitors might be expected to show tory potencies of different NSAIDs for COX-1 fewer side effects, for example, less gastric ulcer- and COX-2 would be useful in attempting to

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Table 9.1 Classifi cation of NSAIDs according to their inhibitory potencies of COX-1 and COX-2a Group Inhibitory potencies of NSAIDs for COX-1 and COX-2b Examples of NSAID 1 Full inhibition of COX-1 and COX-2 with poor selectivity Aspirin, ibuprofen, diclofenac, naproxen, indomethacin, sulindac, piroxicamc 2 Inhibition of COX-1 and COX-2 but preferential selectivity Etodolac, meloxicam, nimesulide, celecoxib for COX-2 3 Weak activity for COX-1; strong inhibition of COX-2 Rofecoxib, [diisopropylfl uorophosphate] 4 Weak inhibitors of COX-1 and COX-2 Many salicylates, nabumetone, sulfasalazine Results taken from Warner TD et al. Proc Natl Acad Sci. 1999;96:7563 a Human whole blood assays measuring formation of TXB2 and PGE2 by radioimmunoassay b Calculated as IC50 and IC80 values (refer Table 9.2 ) c Group 1 contains most of the currently used NSAIDs establish COX-1 and COX-2 selectivities and be fi ve times more toxic in this respect than any distinguishing benefi cial from harmful effects of other NSAID. Group 2 contains preferential the different drugs. This comparison was under- COX-2 inhibitors with, for example, etodolac and taken by the Vane group using human whole meloxicam producing 80 % inhibition of COX-2 blood assays for COX-1 (with added NSAID and and only 25 % inhibition of COX-1 at the same calcium ionophore A23187) and COX-2 (treated concentration. Compounds in this group have an with aspirin to inactivate COX-1 and then LPS improved gastrointestinal toxicity profi le but, plus NSAID or exposed to IL-1β plus NSAID, since they do inhibit COX-1 to some extent, then A23187 and diclofenac). Concentrations of increases in dosage could increase gastrointesti-

TXB2 (as a measure of TXA2 formation) and nal toxicity. NSAIDs in group 3 have very little

PGE 2 were determined by radioimmunoassay as effect on COX-1 and this is refl ected in their low measures of COX activity. The NSAIDs tested gastrointestinal toxicity, but attention has been could be divided into four groups on the basis of drawn to the possibility that their use in the pres- their inhibitory potencies of COX-1 and COX-2 ence of existing gastrointestinal damage may (Table 9.1 ). Determination of the concentrations slow the healing process due to inhibited produc- of individual NSAIDs needed to inhibit the tion of protective COX-2 products. Compounds COX-1 and COX-2 isoforms and comparison of in group 4 showed weak activity in inhibiting the activities on a molar basis revealed the COX-1 prostanoid production—sodium salicylate, for and COX-2 selectivities of each drug and enabled example, inhibited prostanoid formation at far their ranking in order of potency (Table 9.2 ). higher concentrations than those achieved in vivo. A comparison of these fi ndings with epidemiological studies of NSAID-induced gastrointestinal toxicity revealed that NSAIDs associated 9.4.4 The So-Called COX-3 Isoform with the highest gastrointestinal toxicity have the greatest COX-1 selectivity. Surveys have shown After discovery of a COX-1 variant in some this order of toxicity (from the least to the most regions of the brain, it was suggested that acet- damaging NSAID) to be: ibuprofen, diclofenac, aminophen, a weak inhibitor of COX-1 and difl unisil, fenoprofen, aspirin, sulindac, naproxen, COX-2, acts by inhibiting this variant, termed indomethacin, piroxicam, ketoprofen, tolmetin. COX-3. Discovered in 2002, encoded by the NSAIDs associated with the greatest COX-1 COX-1 gene and with an intron not retained in selectivity in group 1 in the table (Table 9.2 ), viz., COX-1, COX-3 was shown to be inhibited by ketorolac, fl urbiprofen, ketoprofen, and indo- acetaminophen, phenacetin, metamizole (dipy- methacin, are among the NSAIDs with the great- rone), antipyrine, and some NSAIDs in rodent est gastrointestinal toxicity. Ketorolac, the most experiments. It turned out, however, that results COX-1-selective drug in the study, is known to with canine COX-3 were not applicable to mouse

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Table 9.2 COX selectivities of some important NSAIDs determined from their inhibitory potencies of COX-1 and COX-2 Whole blood COX selectivityc Rank orderd of a b assay IC80 (μM) index IC 80 ratio selectivity for NSAID COX-1 COX-2 COX-2:COX-1 COX-2 COX-1 GROUP 1e Aspirin 8.0 30.0 3.8 16 5 Diclofenac 1.0 0.23 0.23 7 14 Fenopropen 230 24.0 1.0 12 9 Flubiprofen 1.0 51.0 51.0 19 2 Ibuprofen 58.0 150.0 2.6 14 7 Indomethacin 0.46 2.0 4.3 17 4 Ketoprofen 1.0 6.0 6.0 18 3 Ketorolac 0.0034 1.0 294.0 20 1 Naproxen 110.0 330.0 3.0 15 6 Piroxicam 15.0 7.0 0.47 9 12 Sulindac sulﬁ de 38.0 11.0 0.29 8 13 GROUP 2e Celecoxib 28.0 3.0 0.11 5 16 Etodolac 69.0 3.0 0.043 3 18 Meloxicam 22.0 2.0 0.091 4 17 Nimesulide 41.0 7.0 0.17 6 15 GROUP 3e NS398 65.0 1.0 0.015 1 20 Rofecoxib >100 5.0 <0.05 2 19 GROUP 4e Ampyrone 270 670 2.5 13 8 Diﬂ unisal 530 400 0.75 10 11 Sodium salycilate 4,956 45,000 0.92 11 10 Data from Warner TD et al. Proc Natl Acad Sci. 1999;96:7563 a William Harvey human whole blood assay. Radioimmunoassays of TXB2 and PGE2 used as a measure of COX-1 and COX-2 activities (Warner TD et al. Proc Natl Acad Sci. 1999;96:7563) b IC80 used since relative potency of NSAID varies with concentration and NSAIDs are used therapeutically at doses that produce more than 50 % reduction of prostanoid formation c Selectivities of NSAIDs toward COX-1 and COX-2 were determined by IC80 ratios. Higher ratios indicate increasing inhibitory (IC80 ) amounts for COX-2 compared to COX-1 d Lower numbers are associated with increased selectivity toward the COX isoform e For details of the four groups see Table 9.1

and human which have COX-1- and COX-2 analgesics does not seem worthwhile but perhaps proteins of completely different amino acid there remains more to learn about COX isoforms sequence to the canine COX-3 protein. COX-3 is and their applicability to medical science. a splice variant of COX-1. In dogs, the protein resembles the other two COX isoforms but in mice and humans it does not. Some results sug- 9.5 Sensitivities to NSAIDs gest that acetaminophen inhibits COX activity via an antioxidant action seen with phenolic com- It was the progenitor drug aspirin that was the ﬁ rst pounds. At present, therefore, the utilization of NSAID implicated in an allergic-like reaction COX-3 as a potential drug target in the search for when, in 1902, Hirschberg reported acute

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angioedema provoked by the newly marketed appears to be a higher risk of anaphylaxis with compound. Twenty years later, Widal, Abrani, and aryl and heteroaryl acetic acid and propionic acid Lermoyez described the association of aspirin derivatives; pyrazolones may induce more imme- with episodes of asthma and nasal polyps, but it diate reactions and COX-2 inhibitors have also was another 44 years before this “aspirin triad,” as been implicated in sensitivity reactions. Delayed- it became known, was recognized and defi ned as a type responses to NSAIDs with cutaneous mani- distinct clinical syndrome when Samter and Beers festations are well recognized and although the published their clinical studies on aspirin intoler- severe necrotic epidermal reactions such as ance together with a consideration of its pathogen- Stevens–Johnson syndrome occur, they are rare. esis. A number of different sensitivities to NSAIDs In the general population, aspirin sensitivity is involving different mechanisms and a wide range thought to affect about 0.5–1.9 % of subjects. of symptoms from true type I anaphylaxis to The incidence of aspirin sensitivity in asthmatic delayed-type cutaneous reactions are now recog- adults is said to be 3–5 % when history alone is nized with adverse respiratory and cutaneous considered, but aspirin challenges boost these responses of unknown pathogenesis in between. fi gures up to two or threefold. It has been claimed that asthma induced by aspirin is widely underdi- agnosed, and a number of surveys comparing 9.5.1 Defi nition and Epidemiology patients with physician-diagnosed asthma with subjects without asthma appear to back this up, In relation to aspirin, Szczeklik and Stevenson showing incidences of aspirin-induced asthma have proposed the name “ASA [aspirin]-exacer- four to ten times higher in the former group. In bated respiratory disease” as the best designation one European survey, aspirin provocation tests of what they describe as “the aggressive and con- revealed positive responses in 15 % of subjects tinuous infl ammatory disease of the airways, previously unaware of their sensitivity to the combined with exacerbation of asthma and rhini- drug. For patients with bronchial asthma and tis attacks” after ingestion of aspirin and most nasal polyps, aspirin sensitivity has been shown NSAIDs. Other names commonly used are to be present in about one in four subjects and in aspirin-induced asthma, aspirin sensitivity, the chronic urticaria, an incidence of 27–35 % has aspirin triad, aspirin-intolerant asthma, Widal’s been reported. Considering other NSAIDs as syndrome, and Samter’s syndrome. Several well as aspirin, the prevalence of sensitivity to names have been used to cover adverse reactions this group of drugs is estimated to be 0.3–0.9 % to the many different drugs collectively called of the general population, making NSAIDs the NSAIDs, viz., NSAID hypersensitivity, pseudo- second-most common cause of drug-induced allergy, idiosyncrasy, intolerance, and sensitivity. intolerance after antibiotics. NSAIDs have also The two latter terms will be employed here. been reported to be fi rst or second on the list of Depending on the symptoms and what is known drugs that provoke anaphylaxis. Patients with of the underlying mechanisms, several types of chronic idiopathic urticaria show an increased sensitivity responses to NSAIDs have been frequency of intolerance to aspirin and female defi ned (see below) but the mechanistic and asso- gender, atopy, young adulthood, and intermittent ciated clinical pictures are still vague enough not use of NSAIDs are additional risk factors for to be able to distinguish some cases where mixed NSAID intolerance. reactions from separate categories are seen in Sensitivity to COX inhibitors is rare in chil- some patients. dren with most reactions occurring to ibuprofen, Sensitivity reactions occur with all NSAIDs aspirin, and acetaminophen in that order. Cross- regardless of structure, chemical and physical reactions between NSAIDs are frequent, but only properties, anti-infl ammatory potency, COX about 10 % of NSAID-sensitive children react to selectivity, and differences in mechanism that acetaminophen and all of these patients react to may exist (e.g., with acetaminophen). There other NSAIDs.

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9.5.2 Clinical Classifi cation asthmatic attack that is often accompanied by of Sensitivities to NSAIDs rhinorrhea, infl ammation of the conjunctiva, fl ushing, abdominal pain, and perhaps some urti- The natural history of aspirin-induced asthma, or caria. Most patients suffer from chronic rhinosi- aspirin triad, begins with rhinorrhea and nasal nusitis and severe, persistent, steroid-dependent congestion. This proceeds to rhinitis which asthma with nasal polyps. Reactions to aspirin or becomes perennial and associated with chronic a NSAID can precipitate an asthma attack that is sinusitis and nasal polyps. Asthma develops 1–5 severe and life-threatening. years after the onset of rhinitis. The disease usually appears at an age of 30–34 years and its 9.5.2.1.2 Mechanisms course is usually more severe and progressive in Although the mechanism(s) underlying these women who outnumber men. NSAID-induced respiratory reactions is not immu- For many investigators, the sheer number and nological, the precise cellular and molecular pro- range of clinical manifestations observed in cesses have not yet been fully defi ned and proven. reviewing cases of NSAID-induced sensitivities For some years now, the most favored explanation have made classifi cation of the reactions diffi cult. is a pharmacologic mechanism originally proposed This is understandable since symptoms may by Szczeklik in 1975. Asthmatic attacks were include nasal congestion, rhinorrhea, sinusitis, attributed to the inhibition of COX (now known to dyspnea, wheezing, urticaria, angioedema, and be COX-1 not COX-2) by aspirin and aspirin-like skin rashes as well as patients with a combination drugs in the airways of sensitive patients leading to of respiratory and skin symptoms. In addition to a shift of arachidonic acid metabolism from the the number of different clinical signs and symp- cyclooxygenase to the lipoxygenase pathway with toms, a consideration of the time of onset of reac- an associated increased production of cysteinyl tions and the apparent underlying mechanisms leukotrienes (Sect. 9.4.1 ; Figs. 9.2 and 9.3 ; see also should be undertaken before any classifi cation of Sect. 3.2.5.2 and Fig. 3.8 ). The consequent sensitivities to NSAIDs is attempted. These con- decreased amount of PGE2 , which normally helps siderations have been taken into account in the to dampen the production of LTC4 , LTD4 , and LTE4 following classifi cations. and modulate mediator release from mast cells and other infl ammatory cells, is also thought to be a 9.5.2.1 NSAID-Induced Respiratory contributing factor. Observations advanced to sup- Reactions port this theory include: 9.5.2.1.1 Clinical Features 1. Respiratory reactions in aspirin-induced The classic “aspirin triad” or NSAID sensitivity asthma are triggered by COX-1 and not consists of asthma, rhinosinusitis, and nasal pol- COX-2 inhibitors. yps. Symptoms may develop stepwise over a 2. Airways symptoms tend to correlate with the number of years. In some patients only two of the potency of the inducing COX-1 NSAID. main symptoms may be present. It is one of two 3. Leukotriene receptor antagonists and inhibi- cross-reactive types of NSAID sensitivity. This tors of leukotriene synthesis prevent, or par- state of intolerance, which occurs more often in tially prevent, symptoms following aspirin women than men, is observed in patients with challenge. aspirin-induced asthma, but the recently sug- 4. Abnormalities in the lipoxygenase pathway gested name, aspirin-exacerbated respiratory dis- may exist in patients hypersensitive to COX-1 ease, is probably a more appropriate designation inhibitors since they show higher levels of for the condition that is essentially not a true drug leukotrienes even before exposure to aspirin hypersensitivity but an underlying chronic infl am- and other NSAIDs. Patients with aspirin- matory respiratory disease occasionally exacer- induced asthma may show increased baseline bated by aspirin or some other NSAID. Ingestion levels of urinary LTE4 that increase after chal- of aspirin or some NSAIDs by these patients pro- lenge with aspirin and correlate with the vokes, within 3 h (usually 30 min to 2 h), an acute severity of the induced reaction.

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5. Challenge with aspirin increases leukotriene tive” to NSAIDs revealed that 50 % of the patients levels in nasal and bronchial secretions. were misdiagnosed. This result indicates that 6. There is some evidence that the leukotrienes NSAID sensitivity is probably overestimated are major mediators of clinical symptoms (presumably because of misinterpretation of clin- seen in asthma exacerbated by aspirin/ ical histories) and, at the same time, emphasizes NSAIDs, and this evidence has been bolstered the importance of challenge testing for patients by the important ﬁ nding of over-expression of suspected of sensitivity to these drugs.

LTC4 synthase in bronchial biopsies from Without a clear history, single or double- patients with aspirin-intolerant asthma. blinded challenge tests are obligatory. The tests 7. In aspirin-sensitive rhinosinusitis, the number are best done in a hospital environment by trained of cells expressing the leukotriene receptor specialists (see Sect. 4.4 ). American and European

CysLT1 R (Sect. 3.2.5.2.2 ) is signiﬁ cantly guidelines on aspirin challenge tests have been higher in aspirin-sensitive than in aspirin- published (see Ann Allergy Asthma Immunol tolerant patients, and desensitization is associ- 2007;98:172 and Allergy 2007;62:1111). Oral, ated with a decrease in the numbers of cells inhalation, and nasal aspirin challenges are used

expressing CysLT1 R. and, although the sensitivities of the three tests are 8. Japanese researchers recently demonstrated similar (89, 77–90, 80–86.7 %), oral challenge is

signiﬁ cantly higher baseline levels of LTE4 and generally considered to be the preferred conﬁ rma-

the abundant urinary metabolite of PGD2, tory test for NSAIDs. Oral or inhalation challenge tetranor-PGDM (1,15-dioxo-9α-hydroxy- should not be performed in patients with unstable 2,3,4,5-tetranorprostan-1,20-dioc acid), in asthma or FEV1 % lower than 1.5 L or 70 % of the patients with aspirin-induced asthma. This may predicted value. The threshold dose of aspirin for be an indication of increased mast cell activa- oral challenge, that is, the smallest dose evoking a tion in the pathophysiology of the disease. signifi cant fall in FEV1, varies with the individual Genetic polymorphisms associated with NSAID patient and may range from <10 mg to 600 mg. intolerance and most related to enzymes in the ara- For oral challenge with aspirin, placebo and drug chidonic acid metabolic pathways have been are given at one and a half to 2 h intervals, and sought but results so far have not always been con- FEV1 is measured at baseline and then every 30 sistent or led to major insights (see Sect. 9.5.5 ). min. On day one, placebo is given in four doses; on day two, four increasing doses of aspirin are 9.5.2.1.3 Diagnosis given until a maximum single dose of 500 mg is Sánchez-Borges believes that the information on reached or a clinical reaction results. Other proto- symptoms and exposure to NSAIDs is of the cols recommend a maximum dose of aspirin of utmost importance in determining the temporal 325 mg. A positive reaction is the appearance of relationship between the clinical picture and the clinical symptoms or a decrease in FEV1 equal to probability of a drug etiology—the diagnosis of a or greater than 20 % of the baseline fi gure. For NSAID-induced respiratory reaction is reliable in inhalation challenge, lysine–aspirin is adminis- a patient who shows repeated episodes of asthma tered in increasing doses using a dosimeter-con- after taking cross-reactive NSAIDs. Confi rmation trolled nebulizer. Doses are given every 30 min is obtained by oral challenge testing. and FEV1 measurements are taken every 10 min after each drug administration. A positive response 9.5.2.1.3.1 Challenge (Provocation) Tests is again the appearance of symptoms or a 20 % Although reactions to NSAIDs are said to be one fall in FEV1, the latter obtained from construction of the most common drug hypersensitivities and of a dose response curve. For patients with severe challenge testing is currently the only sure way to asthma, nasal challenge with lysine-aspirin can be diagnose or exclude true hypersensitivity, a care- considered. Nasal symptoms are evaluated with ful evaluation involving challenges of 260 the aid of peak nasal inspiratory fl ow, acoustic rhi- patients previously diagnosed as “hypersensi- nometry, or active anterior rhinomanometry.

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9.5.2.1.3.2 Basophil Activation Test for aspirin showed sensitivity of 42 % and a speci- Because of the occasional presence of mixed fi city of 100 %. Sensitivities with other NSAIDs symptoms, blurred demarcation lines between were not impressive ranging from 15 % for met- classifi ed NSAID sensitivities and uncertainty amizol to 55 % for naproxen. Adding the CAST about the putative mechanism(s) underling the results to the sensitivity fi nding for all four NSAIDs reactions, there is at present an absence of obvious tested, increased the sensitivity to 73 % with a in vitro complementary diagnostic tests for the dif- specifi city of only 71 % making the conclusion that ferent NSAID-induced responses. The basophil the basophil test might “help avoid some cumber- activation test (BAT) has been applied to the diag- some and dangerous provocation challenges” nosis of reactions and its use advocated, but there is debatable. In at least two other investigations of no common agreement about the test’s general use- cysteinyl-leukotriene release in aspirin-induced fulness or even applicability. Only a minority of asthmatics, higher amounts of the liberated media- hypersensitive reactions to NSAIDs are IgE anti- tors in the aspirin-sensitive patients were recorded, body-mediated with most of the reactions being but the low sensitivity and predictive values were respiratory and cutaneous responses apparently seen as limiting the clinical usefulness of the test in resulting from drug-induced inhibition of COX-1, the diagnosis of aspirin sensitivity. depletion of PGE2 , and a resultant increase in production of cysteinyl leukotrienes and other mast 9.5.2.1.3.4 Generation of 15-HETE from cell mediators. When BAT has been employed Peripheral Blood Leukocytes with suspected cases of NSAID sensitivity, the par- Proceeding from the observation that aspirin ticipating patients have not always formed a clini- specifi cally triggers the release of cally homogeneous group sometimes with mixed 15- hydroxyeicosatetraenoic acid (15-HETE) respiratory, cutaneous, immediate, and delayed (Fig. 9.3 ; Sect. 3.2.5.2 and Fig. 3.8) from epithe- reactions among the selected cases. Despite this, a lial cells of nasal polyps and peripheral blood number of studies from Spain claim that basophil leukocytes from patients with aspirin-induced activation induced by aspirin and other NSAIDs is asthma/rhinosinusitis, 15-HETE generation was useful for the in vitro diagnosis of what has some- utilized as a test for the identifi cation of aspirin- times been termed the “NSAID hypersensitivity sensitive patients. Stimulation in vitro of periph- syndrome” (see below, Sect. 9.5.4 ). eral blood leukocytes with 200 μM of aspirin resulted in a mean increase of over 400 % in 9.5.2.1.3.3 Test for Release of Cysteinyl 15-HETE generation but only small to insignifi - Leukotrioenes cant responses were seen in aspirin-tolerant asth- The Cellular Allergy Stimulation Test (CAST® ), matic and control subjects. Sensitivity of the test designed to measure the release of cysteinyl leukot- was 83 % and specifi city 82 %; positive and neg- rienes from leukocytes following allergen chal- ative predictive values were 0.79 and 0.86, lenge (see Sect. 4.5.3.1 ), has been applied as an respectively. The NSAID COX-1 inhibitor ELISA or combined with fl ow cytometry in the naproxen also triggered 15-HETE release but Flow CAST® for the in vitro diagnosis of aspirin- COX-2-selective NSAIDs did not. and other NSAID-induced sensitivity. In a Polish investigation, application of the CAST to aspirin- 9.5.2.1.4 Patient Management intolerant and tolerant asthmatics after stimulation Patients who experience exacerbation of their with lysine–aspirin revealed a weak stimulatory asthma after taking aspirin or other NSAID effect on leukotriene release in both groups of COX-1 inhibitors should avoid these drugs as patients. The verdict of the investigators was that well as drugs that increase leukotriene levels and the test had no value for diagnosis of aspirin- topical or systemic corticosteroids. NSAIDs that induced asthma. In their continuing application of have only weak inhibitory activity for the ultimate the CAST assay, the de Weck/Sanz group in synthesis of prostaglandins such as acetamino- Pamplona studied 60 “aspirin- and/or NSAID- phen and selective COX-2 inhibitors seem to be hypersensitive” patients. A fl ow cytometric BAT tolerated by most patients with aspirin- induced

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Table 9.3 Oral desensitization protocol for patients with challenges, cutaneous reactions make up a major aspirin-induced asthma (aspirin-exacerbated respiratory component of symptoms seen in patients with the disease) following clinical patterns: Time Placebo or aspirin dose (mg) on (Hours) Day 1 Day 2 Day 3 9.5.2.2.1 Cross-Reacting NSAID-Induced 0 Placebo 30 150 Urticaria and Angioedema 3 Placebo 60 325 For an extended discussion of urticaria and 6 Placebo 120 650 angioedema see Sect. 3.2.8 . Following successful desensitization, daily aspirin tablet In patients with asthma or chronic urticaria, treatment is maintained Adapted from Szczeklik A, Stevenson DD. J Allergy Clin the incidence of NSAID sensitivity is said to be Immunol. 1999;104:5 23–28 %. Some patients with chronic idiopathic urticaria develop wheals and angioedema or an increase in urticaria after receiving aspirin. Such asthma. Caution should be exercised, however, reactions generally develop 1–4 h after adminis- with high doses of acetaminophen and preferen- tration of the drug although the time may be as tial COX-2 inhibitors like nimesulide and meloxi- short as 15 min and as long as 24 h. Reactions cam since loss of tolerance and sensitivity usually resolve within a few hours but may con- reactions may result. Oral tolerance tests with tinue for up to 10 days. In contrast to aspirin- these drugs are therefore recommended before induced asthma, elucidation of the mechanisms the drugs are employed for regular use. involved in aspirin-induced urticaria/angioedema has been a slow process. As well as aspirin, 9.5.2.1.5 Desensitization NSAIDs, like ibuprofen, naproxen, and diclofe- Tolerance to aspirin (or other NSAIDs) can be nac that inhibit COX-1, may cause an increase in induced by repeated oral administrations of drug, wheals and swelling in up to one-third of patients but to maintain tolerance, or desensitization, the with chronic urticaria. In an evaluation of cross- patient needs to ingest the drug on a regular (usu- sensitivity between aspirin and selective COX-2 ally daily) basis. The “Scripps Clinic Protocol,” inhibitors in chronic idiopathic urticaria patients, devised by Szczeklik and Stevenson for oral Szczeklik and collaborators found that rofecoxib desensitization of patients with aspirin-induced (37.5 mg) and celecoxib (300 mg) did not elicit asthma is based on the administration of small skin eruptions in any aspirin-sensitive patients incremental doses of aspirin over a 3 day period (see also below under Management) indicating until 400–650 mg of the drug is tolerated that there is no cross-reaction between these two (Table 9.3 ). After successful desensitization, COX-2 inhibitors and aspirin, and aspirin- daily aspirin treatment is maintained. Szczeklik induced urticaria-angioedema is a COX-1- and Stevenson recommend that aspirin desensiti- dependent process. Caution should be exercised, zation followed by a daily maintenance dose be however, when considering the use of COX-2 considered for aspirin-induced asthma patients inhibitors in patients with NSAID-induced urti- whose disease is controlled only with unaccept- caria and angioedema. Some recent results with ably high doses of corticosteroids, patients who etoricoxib suggest that COX-2 inhibitors may not require repeated nasal polypectomies or sinus be safe in such patients who are intolerant to a surgery, and those who need aspirin or other number of NSAIDs including acetaminophen.

NSAIDs for treatment of, for example, their cor- Urinary LTE4 measurements indicate that onary artery disease, thromoembolism, etc. aspirin-sensitive chronic idiopathic urticaria is associated with overproduction of cysteinyl leu- 9.5.2.2 NSAID-Induced Cutaneous kotrienes. NSAID sensitivity in urticaria is Reactions reﬂ ected in the overproduction of cysteinyl leu-

Although urticaria and angioedema may be seen kotrienes including basal production of LTE 4 . as minor symptoms in about 5 % of patients with Inhibition of COX-1 and alterations in eicosanoid aspirin-induced asthma and during some aspirin production, including an increase in PGD2 levels,

[email protected] 334 9 Nonsteroidal Anti-Infl ammatory Drugs following aspirin challenge in patients with urti- Table 9.4 Oral desensitization protocols for patients caria and angioedema, suggests a similar mecha- with aspirin/NSAID-induced cutaneous reactions nism to NSAID-induced asthma with release of Placebo or mediators from infl ammatory cells in the skin. Protocol Time aspirin dose (mg) This was confi rmed by the detection of signifi - Wong et ala 0 min 0.1 15 min 0.3 cant increases in urinary LTE4 and the stable 30 min 10 metabolite of PGD 2 , 9α, 11β-PGF 2 in patients’ plasma. Patients with chronic urticaria and 45 min 30 60 min 40 NSAID sensitivity show a higher incidence of 85 min 81 asthma and nasal polyps compared to those with 110 min 162 chronic urticaria alone. 135 min 325 For diagnosis, skin testing with aspirin and Schaefer-Goreb 0 h Placebo other NSAIDs is not indicated and in the absence 1 h 150 of proven, validated, and standardized in vitro 2 h 325 tests, oral challenge is currently the only way to 3 h Placebo diagnose or exclude NSAID sensitivity. Even so, 4 h 325 the sensitivity of challenge testing is not 100 % 5 h Placebo and false negative results occur. Oral provocation 6 h END should not be performed during periods of active Patients may be maintained on aspirin 81 mg daily if urticaria; a single-blind placebo-controlled pro- necessary after careful assessment tocol should be employed at least 1–2 weeks after Note: These protocols to be used only if there is zero-to- minimum risk of a respiratory or anaphylactoid reaction skin reactions, and testing should be undertaken a Data from Wong JT et al. J Allergy Clin Immunol. over 2 days. After four separate doses of the pla- 2000;105:997 cebo on day one, four increasing doses of 71, b Data from Schaefer OP, Gore JM. Cardiology. 1999;91:8 117, 312, and 500 mg (total 1,000 mg) of aspirin are administered at intervals of one and a half to two hours. Testing is interrupted if symptoms of sensitivity to NSAIDs appear. Most challenged 9.5.2.2.1.2 Desensitization patients react to an aspirin dose of 325–650 mg. Although desensitization of patients with aspirin- For challenge testing with other NSAIDs, see induced urticaria-angioedema has been said to be Chap. 4 , Table 4.3 . diffi cult to achieve and maintain, successful oral protocols, both rapid and long, have been pub- 9.5.2.2.1.1 Management lished. To shorten the previously published In the management of patients with cutaneous sen- desensitization procedures involving dosage sitivity to NSAIDs, avoidance measures are similar intervals of 2–24 h, a starting dose of 0.1 mg was to those employed for aspirin-/other NSAID- selected, and this was increased 3–3.3-fold in induced asthma, viz., avoid selective COX-1 inhib- early steps and twofold in later steps at 10–30 min itors but acetaminophen is generally well tolerated. intervals allowing the whole procedure to be The selective COX-2 inhibitors etoricoxib, cele- completed within a few hours (Table 9.4 ). In a coxib, and rofecoxib appear to be well tolerated; study involving 11 patients with histories of aspi- however, in one assessment of the tolerability of rin- or NSAID-induced urticaria or angioedema, some COX-2 inhibitors, reactions to the latter two nine patients tolerated the procedure without drugs were seen in 33 % and 3 % percent of adverse effects and continued taking aspirin for patients, respectively, indicating that controlled from 1 to 24 months without developing urticaria oral provocation testing should still be performed. or angioedema. The two intolerant patients had The importance of provocation testing is also high- urticaria due to other agents as well as NSAIDs. lighted by the intolerance of etoricoxib in cross- In some other published protocols, the maximum reactive patients intolerant to acetaminophen. dose of aspirin administered is 325 mg (Table 9.4 )

[email protected] 9.5 Sensitivities to NSAIDs 335

9.5.2.2.2 Multiple NSAID-Induced aspirin) involves two placebo doses given 2 h Urticaria and Angioedema apart on day one, two doses of drug, 100 mg and Note that in a recent “update,” Sánchez-Borges 200 mg, given 2 h apart on day two, one dose of et al. (Pharmaceuticals. 2010;3:10) include ana- 325 mg on day three, and 650 mg on day four. phylaxis along with urticaria and angioedema as Skin scores are recorded every 2 h. one of the manifestations of this intolerance. Successful desensitization of otherwise This variant of NSAID-induced intolerance is healthy individuals with multiple NSAID- more prevalent in atopics and occurs in otherwise induced urticaria and angioedema but no under- normal, healthy cross-reactive subjects with lying skin disorder does not appear to have been acute urticaria and/or angioedema but with no reported. other underlying skin or respiratory disease. Facial angioedema, occurring within minutes or 9.5.2.2.3 Single NSAID-Induced Urticaria, up to 24 h after ingestion of an NSAID, is the Angioedema, and/or Anaphylaxis most frequent clinical manifestation. The These reactions, said to constitute about 30 % of mechanism(s) of these reactions has not been all cases of NSAID sensitivity, are induced by a elucidated but the most likely mechanism may be single or chemically closely related NSAID and related to COX-1 inhibition as is the case with may occur more often in subjects who are atopic, cross-reactive NSAID sensitivities. IgE antibod- female, and have a history of food or drug allergy. ies do not seem to be involved because of the Reactions occur to pyrazolones in particular but variety of different NSAID structures involved. also to diclofenac and less often to acetaminophen, For diagnosis, skin tests and in vitro tests are not aspirin, ibuprofen, indomethacin, ketorolac, considered relevant, so diagnosis tends to be on sulindac, fenoprofen, tolmetin, meclofenamate, the basis of reactions to more than one NSAID in naproxen, piroxicam, and celecoxib. Clinical patients who are otherwise healthy. Up to 80 % of manifestations include urticaria, angioedema, patients with sensitivity to a number of NSAIDs laryngeal edema, generalized pruritus, rhinitis, are said to tolerate acetaminophen and nimesu- bronchospasm, and anaphylaxis. Urticaria/angio- lide but tolerance of COX-2 inhibitors varies, and edema generally occurs within minutes of oral or these drugs may induce reactions in some NSAID IV exposure, and anaphylactic shock and death cross-reactive patients. The capacity to induce have been reported. Sensitivity to a single NSAID reactions may depend on the relative COX-1 with tolerance of other chemically unrelated inhibitory activities of the COX-2 drugs, for NSAIDs indicates that a COX-1-related mecha- example, nimesulide, meloxicam, and celecoxib nism for these reactions is unlikely but the timing each possess some COX-1 inhibitory activity and characteristics of the symptoms points to an while selective COX-2 inhibitors rofecoxob and IgE antibody-mediated mechanism. Evidence for valdecoxib appear to be well tolerated by the this is strongest with the pyrazolone drugs where majority of NSAID-sensitive patients with cuta- positive skin tests and drug-reactive IgE antibod- neous reactions. However, after studying a case ies have been detected. of rofecoxib-induced urticaria and angioedema in an aspirin-sensitive patient, Ring and collabora- 9.5.2.2.3.1 Sensitivity to Pyrazolones tors suggested that COX-2 inhibition may play a There are two forms of pyrazolone sensitivity, role in the pathogenesis of urticaria and they con- both apparently non-dose-related—reactions cluded that use of a selective COX-2 inhibitor in resembling aspirin-induced asthma, probably aspirin-sensitive patients is no guarantee of involving inhibition of prostanoid synthesis safety. Controlled challenges are recommended and overproduction of cysteinyl leukotrienes before administration of any of the so-called spe- and immune type I immediate reactions mani- ciﬁ c COX-2 inhibitors to NSAID- sensitive festing as urticaria, angioedema, and anaphy- patients. A typical oral provocation protocol (for laxis. Reports of anaphylactic shock reactions

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ab H3C H3C

N CH3 N CH3 N N

CH3 CH O O CH3 Phenazone Propyphenazone (Antipyrine) (4-Isopropylpyrine)

cd H3C H3C CH N 3 N CH3 N N O CH3 N N S O– Na+ O O O CH3 CH3 Aminophenazone Metamizole (Aminopyrine, Amidopyrine) (Dipyrone)

Fig. 9.4 Some important pyrazolone nonsteroidal anti-infl ammatory drugs showing the 1-phenyl-2,3-dimethyl-3- pyrazoline-5-one common core structure in bold following administration of aminophenazone ophenazone, or metamizol showed that the IgE (aminopyrine; amidopyrine) and metamizol antibody assay was specifi c for propyphenazone, (dipyrone) date back over 40 years but there is demonstrating lack of cross-reactivity between some evidence that such reactions to pyrazolones the different pyrazolone drugs, a point to be (including the above two drugs, phenazone and borne in mind for skin testing anaphylactic propyphenazone) (Fig. 9.4 ), were observed much patients with the culprit drug. A solid phase earlier. IgE antibodies to propyphenazone, well immunoassay for demonstrating the presence of known for occasional severe adverse reactions, IgE antibodies to pyrazolone drugs has also been were detected in a study of patients with symp- developed and applied for the detection of the toms suggestive of immediate allergic reaction to 1-phenyl-2,3-dimethyl-3-pyrazoline-5-one com- the drug. After linking a spacer arm to mon core structure found in aminophenazone, N -demethylpropyphenazone and conjugating to propyphenazone, and metamizol as well as the human serum albumin with carbodiimide (EDC)/ “parent” drug phenazone (Fig. 9.4 ). A strong S- N-hydroxysuccinamide, the drug conjugate association between pyrazolone drug hypersensi- was employed to detect specifi c IgE antibodies in tivity and HLA-DQ and DR antigens has been an ELISA system alongside skin tests with pro- reported. pyphenazone 0.25 %. Positive wheal and fl are Of other NSAIDs, those more often impli- skin test reactions were seen in 44 of the 53 cated in single NSAID-induced anaphylactic patients (83 %) and propyphenazone-specifi c IgE reactions include diclofenac, acetaminophen, antibodies were detected in 58 % of the patients, ibuprofen, and naproxen. Skin testing of diclofe- but seven of nine skin test-negative patients also nac hypersensitive patients has persistently given had propyphenazone-reactive IgE antibodies. negative results leading investigators to conclude Inhibition of IgE binding to the drug conjugate that a drug antigenic determinant(s) results from by propyphenazone but not by phenazone, amin- protein binding of a metabolite(s) generated

[email protected] 9.5 Sensitivities to NSAIDs 337 in vivo. The nature of the putative metabolite(s) Application of BAT using CD63 expression as is uncertain but 5-hydroxydiclofenac and auto- marker in two studies of patients with oxidation products 5-hydroxydiclofenac quino- hypersensitivity to the pyrazolone, metamizol, an neimine and 4 -hydroxydiclofenac quinoneimine analgesic known to cause IgE- mediated immedi- have been advanced as possible antigenic deter- ate reactions, showed sensitivities of 55 and 42 % minants following experiments in mice. However, and specifi cities of 86 and 100 %, respectively. experiments with human serum albumin conju- In the latter study, CAST® had a sensitivity of gates of diclofenac and its 3-, 4-, and 5-OH phase 52 % with a specifi city of 90 % and when consid- I metabolites did not detect any patients with spe- ered along with the BAT and skin test fi ndings, the cifi c IgE antibodies to the drug or its metabolites three tests together identifi ed 77 % of the drug- in sera from 59 patients with a history of acute allergic patients. Both investigations found a time- hypersensitivity to diclofenac. In addition, BAT dependent decrease in BAT- positive reactions and CAST assays with representative patient with approximately 60 % of patients becoming samples detected no upregulation of CD63 with negative after 6 months. BAT and no signifi cant production of cysteinyl For management of the patient, strict avoid- leukotrienes with CAST. The overall conclusion ance of the culprit drug and cross-reactive drugs was that there was no evidence for an IgE- (unless shown to be tolerated) is necessary, and mediated mechanism based on hapten–protein alternative NSAIDs should only be administered conjugates in diclofenac hypersensitive patients, if fi rst shown to be tolerated by challenge testing. and the involvement of the most relevant metabolites of the drug could be excluded. There 9.5.2.3 Delayed Reactions to NSAIDs appears to be about 40–50 cases of anaphylactic These are responses that generally develop more reactions to acetaminophen. Oral challenge is the than 24 h after exposure to a NSAID and which most successful diagnostic method applied, there appear to be immune-based type IV reactions are only a few reports of positive skin tests to mediated by T cells. Table 9.5 lists cutaneous the drug, and at least one report of the detection and/or systemic reactions that may be provoked of specifi c IgE antibodies. Females predomi- by NSAIDs together with the drugs most com- nate in reported reactions. In a few cases, the monly implicated. Diagnosis is based on the clin- need to dose beyond a threshold (e.g., 100 mg in ical picture of symptoms and their recurrence, one patient) to induce an anaphylactic reaction appearance, and location of skin lesions; involve- has been observed. Collective data on the risks ment of other organs; and, of course, temporal of different NSAIDs is hard to obtain but, relationships. Patch tests and the lymphocyte between 1,985 and 2,000, The Netherlands transformation test may be employed, although Pharmacovigilance Foundation received reports the sensitivity and specifi city of the former is of 76 cases of NSAID-induced anaphylaxis with likely to be unknown and/or in question and the naproxen, ibuprofen, and diclofenac dispropor- validation, reliability, and cost of the latter tionately represented. The results strengthen pre- method are considerations (see Chap. 4). Again, vious fi ndings of a higher risk of anaphylaxis drug provocation testing is the gold standard associated with these three drugs compared to although it is, of course, contraindicated in other NSAIDs. severe, generalized reactions (Chap. 4). In the For reliable skin test diagnosis, testing should management of delayed-type reactions, early be undertaken as soon as possible after the reac- withdrawal of the offending NSAID is essential tion as a loss of sensitivity with time is apparent. and systemic treatment with corticosteroids and To establish the safety of alternative drugs, oral antihistamines may be warranted. Combined challenges are necessary, but oral challenge t esting immediate- and delayed-type hypersensitivity to with the culprit drug and structurally similar drugs NSAIDs may occur indicating that patch as well should be assessed against the risks involved. as prick and intradermal testing should be kept in

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Table 9.5 Delayed reactions to NSAIDs. Clinical manifestations and drugs implicated Clinical manifestations Drugs commonly implicated Comments Maculopapular eruptions Ibuprofen, fl urbiprofen, diclofenac, Probably specifi c T cell hypersensitivities pyrazolones, celecoxib Fixed drug eruptions (FDE) Aspirin, ibuprofen, naproxen, NSAIDs are among the most common indomethacin, diclofenac, difl unisal, causes of FDE mefenamic acid, piroxicam, Cross reactions may occur phenylbutazone, pyrazolones, acetaminophen, nimesulide Contact and photo-contact Diclofenac, indomethacin, ibuprofen, Topical ketoprofen frequently implicated dermatitis fl urbiprofen, ketoprofen, fl ufenamic Chemically related cross-reactivity observed acid, etofenamate, bufexamac, Patients sensitized topically may develop tiaprofenic acid cutaneous reactions to same drug given orally or parenterally Acute generalized Ibuprofen, nimesulide, etoricoxib, Reactions rare exanthematous valdecoxib, celecoxib, aspirin, pustulosis (AGEP) acetaminophen Drug reaction (rash) with Oxicams, celecoxib, ibuprofen Reactions rare eosinophilia and systemic symptoms (DRESS) Severe bullous cutaneous Oxicams, phenylbutazone, Reactions rare reactions (SJS, TEN) oxyphenylbutazone, some Symptoms may occur up to 8 weeks after COX-2 inhibitors, difl unisal drug administration Pneumonitis Naproxen, sulindac, fenbufen, Symptoms of cough, fever, dyspnea, ibuprofen, tolfenamic acid, malaise. Typically responds to drug diclofenac, phenylbutazone, withdrawal but corticosteroids may be oxyphenylbutazone needed. Exact etiology not known Nephritis Indomethacin, sulindac, ibuprofen, Symptoms—rash, fever, eosinophilia or tolmetin, piroxicam, rofecoxib, eosinophiluria, acute renal failure valdecoxib, celecoxib Progression of disease may depend on balance between relative strengths of immune reaction and anti-infl ammatory effects of drug Aseptic meningitis Ibuprofen, sulindac, naproxen, tolmetin, Rare. No obvious cross-reactivity of ketoprofen, diclofenac, indomethacin, NSAIDs. Mechanism may be type III or IV piroxicam, rofecoxib, celecoxib hypersensitivity to drug SJS Stevens–Johnson syndrome, TEN toxic epidermal necrolysis

mind in assessing some patients. Patients with three further sub-patterns each involving urticaria/ severe bullous cutaneous reactions such as angioedema), and delayed, Sánchez- Borges has Stevens–Johnson syndrome and toxic epidermal categorized so-called non-immediate allergic and necrolysis are treated as for burns victims in pseudoallergic adverse reactions to NSAIDs into intensive care. four main patterns—respiratory, cutaneous (again with three sub-patterns manifesting urticaria/ 9.5.2.4 Mixed and Systemic Patterns angioedema), mixed, and systemic. of NSAID-Induced Sensitivity Illustrating how difﬁ cult it can sometimes be to 9.5.2.4.1 Mixed Pattern classify a patient who has reacted adversely to an The mixed pattern, said to be seen in about 30 % NSAID into one of the three main clinical patterns of NSAID-sensitive patients during controlled presented here, viz., respiratory, cutaneous (with challenge, is a blend of respiratory and cutaneous

[email protected] 9.5 Sensitivities to NSAIDs 339 symptoms that include cough, breathlessness, 9.5.3 IgE Antibodies to Aspirin hoarseness, rhinorrhea, wheezing conjunctival itch, urticaria, and angioedema. Absence of clear In 1981, a new medical graduate beginning a signs and symptoms of chronic severe asthma, Ph.D in the authors’ laboratory, was disturbed by sinusitis, nasal polyps, chronic urticaria, and ana- the plight of an unusually high number of aspirin- phylaxis is another indication that these patients sensitive patients with chronic urticaria attending need to be categorized into a separate group. the weekly allergy clinic. Thinking that, in at least some of the patients, there was a high prob- 9.5.2.4.2 Systemic Pattern ability of an underlying IgE-mediated mecha- In this category, reactions may include fl ushing, nism for the distressing reactions, a strategy was nasal and ocular symptoms, bronchospasm, urti- devised to examine the patients’ sera for the pres- caria, abdominal pain, and, occasionally, vasomo- ence or absence of such antibodies. Nineteen tor collapse. The reactions may be anaphylactoid patients, mainly non-atopic with chronic urticaria or anaphylactic, the latter usually observed in were tested for specifi c anti-salicyloyl serum IgE patients who reacted to a single NSAID while tol- antibodies using a freshly prepared salicyloyl- erating other chemically unrelated NSAIDs. The polylysine solid phase conjugate. With polyly- involvement of drug-reactive IgE antibodies is sine succinate as control, skin prick tests and in shown by positive prick or intradermal tests and, vitro histamine release were also investigated if available, immunoassays for drug-specifi c IgE using the aspirin- polylysine antigen. No positive and/or BAT. Some surveys have concluded that responses were obtained in any of the tests lead- NSAIDs constitute the second largest group of ing to the conclusion that the clinical symptoms drugs responsible for anaphylactic reactions. In and signs in patients with chronic urticaria asso- addition to the pyrazolones, there are case reports ciated with aspirin sensitivity are not mediated by of anaphylactic reactions to just about every specifi c IgE antibodies. In view of what has been known NSAID such as aspirin (see below), ibu- learned of the classifi cation and mechanisms profen, diclofenac, indomethacin, naproxen, involved in NSAID sensitivities, it seems likely fenoprofen, sulindac, zomepirac, piroxicam, that the 19 patients studied over 30 years ago meclofenamate, tolmetin, ketorolac, glafenine, would today be placed in the cutaneous pattern acetaminophen, and a number of coxibs including group in the classifi cation of NSAID-sensitive celecoxib, valdecoxib, and etoricoxib. patients. The salicyloyl determinant and the After reviewing the classifi cation of the vari- O -methylsalicyloyl derivative, each linked to a ous intolerant states provoked by NSAIDs, two solid phase by a spacer arm, were later employed specifi c aspects pertaining to mechanisms of by Zhu in Beijing to look for aspirin-specifi c IgE action and diagnosis of hypersensitive responses antibodies in the sera of 28 patients with positive to these drugs, namely, IgE antibody responses to histories of aspirin sensitivity, most confi rmed by aspirin and application of BAT to diagnosis, are oral challenge. Ten patients had asthma, 16 urti- considered. The subject of serum IgE antibodies caria/angioedema, one generalized fl ushing with to aspirin is considered separately since they may pruritus, and one rhinoconjunctivitis. The sera of be found not only in patients classifi ed into dif- 27 (96.4 %) and 20 (71.4 %) patients were shown ferent sensitivity categories but also in patients to have salicyloyl- and O -methylsalicyloyl- where more than one mechanism is implicated. specifi c IgE antibodies. The former antibodies As a diagnostic method for NSAID intolerances, were strongly inhibited by salicylic acid and good BAT has been applied to patients with a heteroge- inhibition was also seen with 2-aminophenol, indi- neous mix of symptoms, some of which are asso- cating antibody recognition of the hydroxy group ciated with IgE-independent respiratory and ortho to the attached spacer group. cutaneous reactions. Many of these patients seem O -Methylsalicylic acid also proved to be a good inappropriate candidates for the test. inhibitor of its complementary antibodies as was

[email protected] 340 9 Nonsteroidal Anti-Infl ammatory Drugs indomethacin which was weaker but still surpris- 27 % had airway symptoms, 67 % had skin symp- ingly active. At the clinical level, indomethacin in toms of urticaria/angioedema, and 6 % presented both low and therapeutic doses has been shown to with both. Given the mixed symptom patterns elicit positive provocation reactions in aspirin- and what is currently understood of the mecha- sensitive asthmatics and cross-desensitization nisms underlying the variously classifi ed sensi- with the two drugs has been reported. This cross- tivities to NSAIDs, the high proportion of positive recognition seen both in vitro and in vivo seems BAT responses found in this study is surprising. to have its basis in the structural similarity Again, it was concluded that BAT would be par- between O -methylsalicylic and the ticularly appropriate for patients with a clinical O -methylphenol structure that is part of the history of NSAID intolerance but in whom chal- indole nucleus of indomethacin. lenge tests are not advisable. In what seems to be Starting with a population of more than 100 a more understandable application of BAT, viz., patients suspected of aspirin intolerance and employment with cells from patients with imme- suffering from asthma with or without nasal pol- diate reactions (anaphylaxis, urticaria, angio- yposis, Sainte-Laudy in Paris employed the clini- edema, asthma, conjunctivitis) to one or more cal histories, skin tests, IgE determinations, NSAIDs, results of a study on 43 patients with and BAT to fi nd nine cases involving an IgE the NSAIDs aspirin, ibuprofen, metamizol, antibody-dependent mechanism. For the IgE diclofenac, acetaminophen, and ketorolac assay, aspirin–lysine was mixed with epoxy (bis- showed a sensitivity of 43 %, specifi city of oxirane)-activated Sepharose (Sect. 4.3.1 ) to 100 %, and positive and negative predictive val- form the aspirin solid phase. ues of 100 % and 54 %, respectively. Two other BAT-based diagnostic investigations, however, one examining different NSAIDs and the other 9.5.4 NSAID Sensitivities and BAT looking only at diclofenac, concluded that the value of the test in NSAID intolerances is yet to In a fl ow cytometric determination of basophil be fi rmly established. Interestingly, although activation induced by aspirin and other NSAIDs, diclofenac induced basophil degranulation, leukocytes from 60 NSAID sensitive patients (38 upregulation of the CD63 marker antigen did not with cutaneous, 20 with airway, and 2 with cuta- occur in NSAID-sensitive patients. neous and airway symptoms) were stimulated with aspirin, acetaminophen, metamizol, diclofenac, and naproxen. Sensitivities and specifi ci- 9.5.5 Genetic Mechanisms of ties, respectively, obtained with the different Aspirin-Induced Sensitivities drugs were: aspirin 43 and 100 %; acetaminophen 12 and 100 %; metamizol 15 and 100 %; A study in a polish population of possible genetic diclofenac 43 and 93 %; naproxen 55 and 74 %. markers for aspirin-induced asthma revealed an These results led the investigators to conclude association between HLA-DPB1*0301 and this that the test might help to avoid some challenge phenotype. This result was later confi rmed in a tests with NSAIDs. In a later extension of this in Korean population by H-S Park and coworkers vitro diagnostic investigation of what was termed where it was found that patients with DPB1*0301 the NSAID “hypersensitivity syndrome,” a multi- were more often females with a lower FEV1 and center study performed within the framework of a higher incidence of rhinosinusitis and nasal the European Network for Drug Allergy (ENDA) polyps than those lacking this marker. Enhanced found 57 % of 140 patients were BAT-positive to expression of LTC4 synthase due to overactive multiple NSAIDs, 27 % were positive with only transcription of an allelic variant (LTC 4 S- one or two concentrations of a single NSAID, 444A > C) associated with aspirin-induced and 16 % were negative. Of the patients described asthma has also been reported from Poland. as having a history of hypersensitivity to NSAIDs, Patients with the C allele had a greater risk of

[email protected] Summary 341 developing aspirin-induced asthma, and those take over-the-counter NSAIDs on a regular homozygous for LTC4 S-444A had higher basis for pain. increases in urinary LTE4 after aspirin challenge. • Ibuprofen and diclofenac are the most com- Note, however, that this association has not been monly consumed NSAIDs. Naproxen, melox- observed in some other populations including icab, celecoxib, ketoprofen, and etorocoxib American, Japanese, and Korean. A suggested are also high on the list. possible involvement of the 5-lipoxygenase gene • Many NSAIDs are organic acids that bind to (ALOX5) in aspirin-induced asthma has been plasma proteins. Different chemical groups supported by the detection of a signifi cant asso- are recognized: salicylates, propionic acid ciation between ALOX5 promoter polymorphism derivatives, aryl and heteroaryl acetic acids, and the severity of airway hyperresponsiveness in anthranilates (fenamic acid derivatives), oxi- a Korean population. Genetic associations cams (enolic acids), phenylpyrazolones, between the leukotriene receptor genes CYSLTR1 analides, COX-2-selective inhibitors. and CYSLTR2 and aspirin-induced asthma were • Prostaglandin endoperoxide H synthases also established in Park’s laboratory where three (PGHS-1, PGHS-2; cyclooxygenases COX-1, single nucleotide polymorphisms (SNPs) in the COX-2) catalyze the formation of prostaglan- promotor region of CYSLTR1 were signifi cantly din G2 (PGG2 ) from arachidonic acid. PGG2 is associated with the asthma phenotype, especially then reduced to PGH2 which is converted by in males. A functional SNP of the PGE2 subtype specifi c synthases to PGD2 , PGE 2 , PGF 2α ,

2 gene, PTGER2, and two SNPs of TBXA2R prostacyclin PGI2 (a vasodilator), and throm- were also found to be associated with aspirin- boxane A2, TXA2 (a vasoconstrictor). induced asthma. • Inhibition of COX-1 reduces production of

A strong positive association has been shown protective PGE2 and PGI 2 in the stomach, between aspirin-induced urticaria/angioedema which may lead to gastric ulceration. and the HLA DRB1*1302 and HLA DQB1*0609 • COX-2, expressed only in infl ammation, is alleles, and it has been suggested that HLA- induced by the cytokines IL-1 and TNF and DRB1*1302-DQB1*0609-DPB1*0201 may be a by LPS. Selective COX-2 inhibitors show genetic marker for determining this phenotype. fewer gastrointestinal side effects but can pro- Promoter polymorphisms of ALOX5 were duce cardiovascular effects, probably by sup- signifi cantly different between aspirin-induced pressing cardio-protective PGI2 . urticaria/angioedema and aspirin-induced asthma • NSAIDs can be classifi ed on the basis of their and an FcεRIα gene promoter polymorphism COX inhibitory and selective properties. Most was signifi cantly associated with the former NSAIDs inhibit both COX-1 and COX-2 to phenotype. some extent. Most are mainly COX-1 inhibi- It is hoped that further genetic studies will tory, e.g., aspirin, ibuprofen, diclofenac, provide more insights into the molecular genetic naproxen, while some are COX-2 inhibitory, mechanisms of aspirin-induced sensitivities and e.g., celecoxib, rofecoxib, and lumiracoxib. fi nd reliable genetic markers for predicting drug • Vane et al. established the relative inhibitory responses. This, in turn, might lead to improved potencies of NSAID COX-1 and COX-2 diagnostic approaches and therapy. inhibitors. NSAIDs with the highest gastrointestinal toxicity have the highest COX-1 selectivity. Summary • Discovered in 2002, the so-called COX-3 isoenzyme, inhibited by acetaminophen, phen- • NSAIDs are one of the most widely and com- acetin, and metamizole, resembles the other monly used drugs worldwide and are respon- two isoforms in dogs but not in humans or sible for about 20–25 % of adverse drug mice. COX-3 is, in fact, a splice variant of reactions. About 30 % of adults in the USA COX-1.

[email protected] 342 9 Nonsteroidal Anti-Inﬂ ammatory Drugs

• In sensitive subjects, aspirin (and other NSAIDs) • IgE antibodies to aspirin occur and specifi c is associated with the so-called “aspirin triad” or assays for their measurement have been aspirin-induced asthma (AIA; also called aspi- developed. rin-exacerbated respiratory disease). Classic • An association of AIA with HLA-DPB1*0301 AIA consists of chronic asthma and rhinosinus- has been demonstrated in Polish and Korean itis together with nasal polyps. The incidence of populations. Other positive associations AIA in adult asthmatics is about 3–5 %. between aspirin-induced urticaria/angioedema • Aspirin together with other NSAIDs form the and HLA alleles have been demonstrated or second biggest group causing drug-induced suggested. intolerance after antibiotics. • The mechanism of NSAID-induced respiratory reactions appears to be due to the redirec- Further Reading tion of arachidonic acid metabolism from the COX to the lipoxygenase synthetic pathway Berges-Gimeno MP, Simon RA, Stevenson DD. Long- with associated production of cysteinyl leu- term treatment with aspirin desensitization in asth- kotrienes. PGE normally helps to dampen the matic patients with aspirin-exacerbated respiratory 2 disease. J Allergy Clin Immunol. 2003;111:180–6. production of the leukotrienes. Harris RE, editor. Infl ammation in the pathogenesis of • Challenge testing is the only sure way to diag- chronic diseases. The COX-2 controversy. Subcellular nose or exclude true sensitivity to an NSAID. Biochemistry, vol. 42. New York: Springer; 2007. Oral challenge with drug and placebo is usually Higashi N, Taniguchi M, Mita H, et al. Aspirin-intolerant asthma (AIA). Assessment using the urinary biomark- undertaken over a 2 day period. A maximum ers, leukotriene E4 (LTE4) and prostaglandin D2 dose of 325 mg or 500 mg aspirin is often used. (PGD2) metabolites. Allergol Int. 2012;61:393–403. • Other diagnostic methods sometimes Kim S-H, Ye Y-M, Lee S-K, et al. Genetic mechanism of employed include BAT (often of doubtful aspirin-induced urticaria/angioedema. Curr Opin Allergy Clin Immunol. 2006;6:266–70. value) and measurement of released cysteinyl Kowalski ML, Ptasinska A, Jedrzejczak M, et al. Aspirin- leukotrienes and 15-HETE. triggered 15-HETE generation in peripheral blood leu- • Desensitization can be induced by repeated kocytes is a specifi c and sensitive aspirin-sensitive oral administration of drug, e.g., the “Scripp’s patients identifi cation test (ASPITest). Allergy. 2005; 60:1139–45. Protocol.” To maintain tolerance, patients Kowalski ML, Makowska JS, Blanca M, et al. need a daily dose of drug. Hypersensitivity to nonsteroidal anti-infl ammatory • NSAID-induced cutaneous reactions occur in drugs (NSAIDs) – classifi cation, diagnosis and man- a number of different clinical patterns—cross- agement: review of the EAACI/ENDA and GA2LEN/ HANNA. Allergy. 2011;66:818–29. reacting NSAID-induced urticaria and angio- Palikhe NS, Kim S-H, Jin HJ et al. (2012) Genetic mecha- edema; multiple NSAID-induced urticaria and nisms in aspirin-exacerbated respiratory disease. angioedema; single NSAID-induced urticaria J Allergy (Cairo). Article ID 794890, 6 pages and angioedema or anaphylaxis. doi. 10.1155/2012/794890 . Sánchez-Borges M, Capriles-Hulett A, Caballero-Fonseca • Reactions to pyrazolone drugs can resem- F. The multiple faces of nonsteroiod antiinfl ammatory ble AIA or be type I IgE-mediated drug hypersensitivity. J Investig Allergol Clin hypersensitivity. Immunol. 2004;14:329–34. • Mixed cutaneous and systemic patterns of Schubert B, Grosse Perdekamp MT, Pfeuffer P, et al. Non- steroidal anti-infl ammatory drug hypersensitivity: NSAID sensitivity occur and are said to make fable or reality? Eur J Dermatol. 2005;15:164–7. up about 30 % of NSAID-sensitive patients. Szczeklik A, Stevenson DD. Aspirin-induced asthma: Patients show a blend of respiratory and cuta- advances in pathogenesis, dagnosis, and management. neous symptoms. J Allergy Clin Immunol. 2003;111:913–21. Vane JR, Bakhle YS, Botting RM. Cyclooxygenases 1 and • Delayed, that is type IV, T cell-mediated reac- 2. Annu Rev Pharmacol Toxicol. 1998;38:97–120. tions are seen. Some NSAID-induced delayed Warner TD, Giuliano F, Vojnovic I, et al. Nonsteroid drug reactions occur as contact dermatitis, fi xed drug selectivities for cyclo-oxygenase-1 rather than cyclo- eruption, DRESS, acute generalized exanthem- oxygenase-2 are associated with human gastointesti- nal toxicity: a full in vitro analysis. Proc Natl Acad Sci atous pustulosis, SJS, TEN, and nephritis. USA. 1999;96:7563–8.

[email protected] Contrast Media 1 0

Abstract Reactions to iodinated contrast media range from mild inconvenience to life-threatening emergency. Histamine release can account for many of the symptoms and nonionic agents are tolerated better than ionics. Reactions can be immediate (IR) or delayed (DR). Incidences of the former are 3–4 % (ionics), 0.2–0.7 % (non-ionics), severe reactions 0.1–0.4 % (ionics), and 0.02–0.04 % (non-ionics). Up to 80 % of reactions can be avoided by using nonionic agents. For DRs, there is no difference between the incidences of reactions to each of the agents. Risk factors for IRs are a previous reaction to a contrast medium, bronchial asthma, cardiac disease, and highly allergic subjects; for DRs, a previous reaction, use of β-blockers, treatment with IL-2, history of drug allergy, and contact allergy. Diagnosis of IRs is based largely on skin tests; IgE antibodies have not been convincingly demonstrated. Breakthrough reactions have occurred following

corticosteroid and H1 antagonist premedication. Gadolinium-based agents, especially the linear chelates, have been associated with nephrogenic systemic ﬁ brosis. They show an adverse reaction incidence of about 0.48 % and 0.01 % for anaphylaxis. Overall, given the large number of contrast media administered, they are one of the safest drugs.

Along with the development and introduction of The agents are now so widely and frequently powerful investigative techniques and advanced used that they are said to be the most commonly diagnostic equipment, contrast media have used drugs in the history of modern medicine. helped place clinical diagnostic radiology irre- Contrast media are not dyes. For contrast media- placeably at the center of diagnostic medicine aided visualization of the body’s internal struc- today. By increasing the contrast of anatomical tures, X-ray imaging techniques are most often structures that are otherwise not easily seen used. Information was originally recorded on and discriminated, contrast media allow the visu- X-ray ﬁ lm, but that has now been largely super- alization of details of internal tissues such as ceded by digitized images using computer-based blood vessels, intestine and the various organs. methods of recording, storage, and display.

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 343 Relationships, DOI 10.1007/978-1-4614-7261-2_10, © Springer Science+Business Media, LLC 2013

[email protected] 344 10 Contrast Media

Instead of passing a single X-ray beam through the body, a computerized tomography scanner 10.1 Iodinated Contrast Media takes X-ray images at many different angles. A computer is used to work out the relative density Contrast media are substances that affect the of the emerging X-rays and ultimately a 3D attenuation of X-rays, thus changing the contrast image can be constructed. Sometimes structures seen in X-ray images. Introduction of gases, as in cannot be visualized by X-rays alone, even with the examination of the gastrointestinal tract, is an the aid of contrast media, for example, the cord of example of the application of negative contrast nerve roots. Here, specifi c contrast media can be media where there is a reduction in the attenua- employed and visualized directly using magnetic tion of X-rays, but most contrast media are posi- resonance imaging (MRI). tive or radio-opaque, that is, they increase the In an X-ray examination of a patient some of attenuation of X-rays. Since X-ray absorption the X-rays are scattered in all directions and increases with the number of electrons (that is the some are absorbed by the different tissues: this is atomic number), the presence of atoms of high known as attenuation of the rays. The amount of atomic number will absorb more X-rays than X-rays absorbed depends on the thickness and atoms of low atomic number such as hydrogen density of the material (for example, gas in the ( Z = 1), carbon (6), nitrogen (7), and oxygen (8). lungs verse lung tissue) in the path of the rays In one of the most successful examples of and, importantly, its chemical composition. Since improvement in clinical diagnostics, the sciences the absorption of X-rays increases with the num- of pharmacology and synthetic medicinal chem- ber of electrons, the chemical composition of a istry cooperated to increase the water solubility tissue can be thought of as the average or effec- and attenuation of contrast media while at the tive atomic numbers of all the atoms involved. same time reducing toxicity. This was achieved This is so since the atomic number ( Z) of an atom by the introduction of iodinated compounds in an is the number of protons in the nucleus of an evolving program of increasing effectiveness. atom and, in an electrically neutral atom, Z is The use of iodine in a water-soluble form for con- equal to the number of electrons in the atom. In trast imaging began in the 1920s with sodium addition to the contribution of density, being able iodide and then monoiodinated pyridine to distinguish, for example, soft tissue from bone derivatives, but toxicity and poor contrast results on a radiograph, is a consequence of the low led on to the second-generation di-iodinated pyri- average atomic number of the soft tissue con- dines. The fi rst big breakthrough came in the trasted with the signifi cantly higher average 1950s with the lower toxicity but still very hyper- atomic number of the calcium-containing bone. osmolar sodium and meglumine (an amino sugar Effective atomic numbers and densities derived from sorbitol) salts of triiodinated ben- (expressed as g/cm3 ) are 7.42 and 1 for water, zoic acid. Since, for better tolerance, the osmo- 7.46 and 1 for muscle, 5.92 and 0.91 for fat, and lalities of the injected contrast medium and body 20 and 1.55 for calcium. If two tissues have simi- fl uids should be as close as possible, another sig- lar densities, thickness, and average atomic num- nifi cant advance was the introduction in the 1970s bers, visualizing and distinguishing the tissues of nonionic iodinated contrast media. By convert- are more diffi cult and may not be possible. ing the carboxyl group of triiodobenzoic acid to Although this situation occurs commonly in the amide, dissociation in solution could no lon- diagnostic radiology, visualization and contrast ger occur and the iodine:ions/particle ratio was can be artifi cially altered by increasing the aver- changed from 1.5:1 (three iodine atoms:two ions) age atomic number of a structure. This is often for high-osmolality contrast media for, example, achieved by administering a liquid of high aver- diatrizoate, to 3:1, a contrast medium of lower age atomic number, for example, to the blood to osmolality, for example, iohexol (Tables 10.1 and visualize blood vessels. 10.2 ). In addition to the low-osmolar nonionic

[email protected] 10.1 Iodinated Contrast Media 345

Table 10.1 Examples of structures from each of the four different categories of iodinated contrast mediaa Category Drug name and structure Ionic monomer Diatrizoateb Cation Acetrizoate O O c Na/Meglumine Diatrizoate Iodamide I I Ioglicate O O Iothalmate H3C N N CH3 Metrizoate H H I Ionic dimer O Ioxaglate Cation Ioxaglate O O c N Na/Meglumine I I I I H O H H N N N HO N O H OO I I Nonionic monomer OH Iohexol H Iobitridol O N OH Iohexol Iomeprol I I Iopamidol OH H Iopentol N OH Iopromide HO N Ioversol O OH I O Nonionic dimer Iodixanol Iodixanol OH H H OH Iotroland HO N O O N OH

I I I I OH H H OH HO N N OH N N OH O I I O O O a All for angiography or urography unless otherwise stated b Also called amidotrizoate c Meglumine: OH OH OH N H OH OH d A myelographic contrast medium compounds, low osmolality was also achieved by for example, the nonionic monomer iohexol, but preparing ionic dimers such as ioxaglate both are still twice as osmolar as human blood. (Tables 10.1 and 10.2 ) where only one of the car- Contrast media with approximately the same boxyls of the two linked triiodinated aromatic osmolality of blood were ﬁ nally produced in the rings was converted to an amide, thus producing 1980s with the introduction of nonionic dimers two ions in solution with a total of six iodine containing six iodine atoms for each non-dissoci- atoms and a ratio of 3:1. The osmolality of this ating molecule, for example, iodixanol ionic dimer is a little less than the osmolality of, (Tables 10.1 and 10.2 ).

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Table 10.2 Evolution of the development of contrast media with fewer side effects and low toxicity. Comparison of some important properties inﬂ uencing the effectiveness and toxicity of drugs in the different categories of iodinated contrast media Iodine Number of timesc content Ratio iodine atoms Osmolality more osmolar Viscosity a b b,d Category Drug example (mg/ml) to number of ions (mOsmol/kg H2 O) than blood (cPs at 37 °C) Ionic monomer Diatrizoatee,f 306 3:2 or 1.5:1 1,530 5 5.0 Ionic dimer Ioxaglatef 320 6:2 or 3:1 580 2 7.5 Nonionic Iohexol 300 3:1 640 2 6.3 monomer Nonionic dimer Iodixanol 320 6:1 290 Iso-osmolarb 11.4 a See structures Table 10.1 b Different contrast media in the same group may show different osmolalities, e.g., iotrolan osmolality = 320 mOsmol/kg H2 O and is more osmolar than blood (290 mOsmol/kg H2 O) c Approximately d At 300 mg iodine/ml e Also known as amidotrizoate f Used as the sodium and/or meglumine salts

The iodine atom has an atomic radius of better. With normal renal function, the iodinated 140 pm (1.4 Å) (empirically measured) which is agents are cleared almost completely by the kid- signifi cantly larger than the atomic radii from, for neys with a half-life of only 1–2 h. Although most example, carbon (70 pm), hydrogen (25 pm), often used intravenously, contrast media can be oxygen (60 pm), nitrogen (65 pm), sulfur given into arteries, the abdomen, and intrathecally, (100 pm), and chlorine (100 pm). To obtain a and numerous surveys in hospitals throughout the visual image of how the presence of iodine atoms Western world attest to their heavy and widespread dominates the volume in space of iodinated con- usage. As diagnostic technologies increase in trast media molecules, the two-dimensional struc- effectiveness and sophistication, new agents and tures of the four contrast agents, containing either procedures become accepted as standard practice, three or six iodine atoms and shown in Table 10.1 , and costs become more affordable, contrast media are represented three-dimensionally as CPK mod- usage will inevitably increase. A trend that will els in Fig. 10.1 . The presence of the bulky iodine almost certainly continue is the preference for the atoms signifi cantly infl uences the physical and better-tolerated nonionic compounds and this will chemical properties of the contrast media and be driven by demand from both doctor and patient. produces structures that, as potential immunogens For example, even in the late 1990s, surveys (antigens and allergens), are relatively unique. revealed that this preference was already clear in many areas of the USA. In one report, 43 % of hospitals surveyed in the southeast of the country 10.2 Usage and Safety of Contrast used nonionic contrast media 100 % of the time Media while 71 % used them more than 75 % of the time. To reduce the usage of the nonionic agents, hospi- Iodinated contrast media are one of the most often tals often introduced selective protocols. administered and safest pharmaceutical products As with every administered pharmaceutical used today not only in radiology but in all areas of agent, adverse reactions to iodinated contrast media medicine. In 2005 it was estimated that worldwide do occur, but the incidence is low. Reactions can be administrations of contrast media exceeded 75 mil- unrelated to the dose or concentration of the solu- lion per year and it is safe to assume that this fi gure tion administered (see below) or they can be dose grows annually. Unlike most drugs, they are not dependent. An important contributor to the latter designed to have a specifi c therapeutic action; in category is the osmolality of the administered agent fact, the more pharmacologically inert they are the which is responsible for the feelings of discomfort,

[email protected] 10.3 Adverse Reactions 347

- + O O Na Diatrizoate Sodium I I O O (Ionic monomer) H C N N CH3 3 H H I

O - + O Na O N Ioxaglate Sodium I I I I H O H H (Ionic dimer) N N N HO N O H I OOI

H OH O N OH Iohexol I I (Non-ionic monomer) H OH N OH HO N OH O O I

OH H H OH HO N O O N OH Iodixanol I I I I OH H H OH (Non-ionic dimer) HO N N OH N N O OH O I O O I

Fig. 10.1 2D structures and CPK models of some commonly used ionic and nonionic iodinated contrast media. Note the bulk of the iodine atoms relative to the rest of the molecules heat, and pain and which may also provoke distur- bance of the electrolyte balance in small children, 10.3 Adverse Reactions renal problems, and damage to the blood–brain barrier. As already indicated, the tolerance of con- 10.3.1 Classifi cation and Symptoms trast media increases as the osmolality approaches the osmolality of serum. Also related to the con- 10.3.1.1 Acute (Immediate) Reactions centration used are the viscosity, the hydrophilic- Iodinated contrast media used today have been ity/lipophilicity balance, protein-binding capacity, carefully developed with the aim of maximizing and histamine- releasing properties of the contrast their effectiveness for tissue visualization while at medium. Viscosity increases with molecular the same time minimizing toxic effects. With the weight so the viscosities of the dimer solutions are relatively safe agents used today, of more concern higher than the solutions of monomers. The practi- than dose-related intolerance and toxicity out- cal implications of a more viscous solution are the lined above are adverse reactions covering a range greater force required for injection, especially of severities that are mostly independent of dose through thin catheters, and slow fl ow infl uencing or concentration. Severity is, in fact, a convenient the visualization of tissues. Some additional adjust- and useful way of categorizing these reactions ments made to reduce toxicity include adjusting the since this approach is clinically relevant and pro- pH to neutral, adding calcium ions to reduce car- vides a guide for subsequent treatment. As with diac toxicity, and altering the number of hydroxyl the so-called toxic reactions, the adverse reac- groups to decrease neural toxicity. tions are small in number relative to the millions

[email protected] 348 10 Contrast Media

Table 10.3 Adverse reactions to contrast media. Some of the main symptoms seen in acute and late reactions Acute (immediate) reactionsa Late (delayed) reactionsb Mild Moderate Severe Reactions to: Reactions to: iodinated gadolinium e contrastc,d media contrast media Nausea, mild vomiting Severe vomiting Hypotensive shock Nausea, vomiting Nephrogenic Urticaria Marked urticaria Respiratory arrest Headache systemic ﬁ brosisf Itching Bronchospasm Cardiac arrest Musculoskeletal pain Facial/laryngeal edema Convulsions Fever Vasovagal attack Skin reactiong Data adapted from ESUR Guidelines of Contrast Media. European Society of Urogenital Radiology, Version 7.0. View at http://www.esur.org/ESUR-Guidelines.6.0.html a Reactions occur within 1 h of injection of contrast media b Reactions occur 1 h to 1 week after injection of contrast media c Many symptoms described not related to contrast media d Risk factors: previous contrast media reaction; interleukin-2 treatment. Prophylaxis generally not recommended Patients who had a previous serious late reaction can be given steroids as premedication (see Table 10.4 ) e Reactions to gadolinium contrast media of lower risk than with an iodinated contrast media f Nephrogenic systemic ﬁ brosis usually presents after 1 week but may occur earlier (see Sect. 10.7.2 ) g Usually mild to moderate and self-limiting. Management is similar to other drug-induced skin reactions

of doses administered each year. A number of marked urticaria; and severe reactions (life- professional bodies including, for example, The threatening)—hypotensive shock, cardiac and American College of Radiology and the European respiratory arrest, severe laryngeal edema, and Society of Urogenital Radiology, have issued convulsions. Table 10.3 sets out the classifi cation classifi cations and guidelines based on a primary of acute non-renal adverse reactions to contrast division of acute (or immediate) and late reac- media and the associated symptoms listed by the tions with the former division subdivided further European Society of Urogenital Radiology in into mild, moderate, or severe reactions. Acute their ESUR Guidelines on Contrast Media. reactions are those that occur within about 1 h of the administration of contrast media. The cutoff 10.3.1.2 Late Reactions time of 1 h for a reaction to be classifi ed as acute Late reactions become apparent more than 1 h and is, of course, somewhat arbitrary and it is a fi gure up to about 1 week after contrast media exposure. disputed by some who argue that fi ndings with Excluding contrast media-induced nephropathy, some patients show that the cutoff point should be the symptoms most commonly seen include nau- extended to 2 or even 3 h. A similar problem of sea, vomiting, headache, and cutaneous reactions deciding the time when the designations “imme- (Table 10.3 ) which tend to be self-limiting and diate” ends and “delayed” begins is seen in other include maculopapular rash in over 50 % of drug allergies. The question is probably best affected patients, xanthema, urticaria, and usually resolved by the patient’s symptomatology. pruritus. In rare cases, cutaneous reactions may Signs and symptoms commonly listed for the progress to a cutaneous vasculitis or even a three different acute reaction categories include Stevens–Johnson-like syndrome. Late reactions mild reactions (generally self-limiting without may often be missed since patients generally leave evidence of progression)—nausea, vomiting, the department sooner than an hour after adminis- cough, headache, itching, pallor, fl ushing, and tration of the contrast preparation, and because the chills; moderate reactions (require treatment but delayed reactions are so often self- limiting, the not immediately life-threatening)—tachycardia/ radiologist may remain unaware of them. Recent bradycardia, bronchospasm, laryngeal edema, reports, however, of delayed skin reactions and a

[email protected] 10.3 Adverse Reactions 349 few cases of serious delayed reactions involving the low- and high-osmolality agents. The mortality hypotension, shock, and angioedema following rate has been estimated to be in the range 1 in intravascular injection of nonionic iodinated 100,000 to 1 in 170,000. dimers highlight the potential dangers of late reac- In summary then, it can be said that as well as tions (see below) occurring in the absence of provoking a higher incidence of adverse reactions, direct medical awareness and supervision. high-osmolar ionic contrast media cause reac- Gadolinium-based contrast media are referred to tions that are more severe than the low-osmolar in Table 10.3 for the sake of completeness. These nonionic contrast media-induced reactions and agents are discussed later in this chapter. the nonionic preparations are less distressing for the patient overall. Although severe reactions with high-osmolar ionic contrast media are still 10.3.2 Incidence of Reactions rare, they are more frequent than severe reactions to low-osmolar nonionic media. Up to 80 % of 10.3.2.1 Acute Reactions the reactions to the ionic agents can be avoided The largest study so far of the incidences of by substituting a nonionic medium. adverse reactions to different contrast media was reported by the Japanese Committee on the 10.3.2.2 Late Reactions Safety of Contrast Media in 1990. In this pro- Obtaining reliable and relevant information on spective study of 337,647 cases, 169,284 cases the frequency of late reactions to contrast media (50.1 %) received ionic contrast media and is not easy for a number of the usual reasons 168,363 (49.9 %) received nonionic contrast related to data collection but particularly because media. Adverse drug reactions occurred in of the relatively larger time interval between the 12.66 % of the ionic contrast media group and injection of the agent and the appearance of 3.13 % of the nonionic group. For severe adverse symptoms. And, of course, the bigger the time reactions the corresponding fi gures were 0.22 % interval, the more diffi cult it is to be sure that the and 0.04 %, respectively, with one death occurred symptoms were caused by the contrast medium. in each group. The authors of the study concluded Most studies show that there is no signifi cant dif- that “non-ionic contrast media signifi cantly ference in the incidences of late reactions between reduce the frequency of severe and potentially ionic and nonionic media or between the differ- life-threatening adverse drug reactions to con- ent nonionic preparations. Although fi gures as trast media at all levels of risk and that use of low as 0.52 % and as high as 23 % have been these media represents the most effective means reported, the incidence of reactions in the fi rst of increasing the safety of contrast media exami- 24 h appears to be about 4 % settling to about nations.” While the incidence of reactions to 1–3 % over a 7-day period. The nonionic com- high-osmolar ionic contrast media in the Japanese pound iopamidol showed an incidence of 5.5 % survey is higher than most estimates, it is clear of late skin rashes in a survey of 1,381 patients. that reactions to these agents occur within the A curious seasonal variation in the occurrence range of about 2–8 % with a fi gure of around 3 or of late adverse skin reactions has been reported 4 % perhaps being most likely. The estimated from Finland. In a study of a possible relation- reaction frequency for the low-osmolar nonionic ship between sun exposure and late reactions in agents ranges up to a maximum of about 3 %, but 4,875 adults who had received an iodinated confi gures of 0.2–0.7 % have been deduced in a trast medium, a 3-month (April to June) peak in number of studies. Figures for the incidences of the incidence of reactions was seen. This period severe reactions are much more settled being included 35 % of all events and most of the reac- 0.1–0.4 % for ionic and 0.02–0.04 % for nonionic tions occurred on sun-exposed areas of the body, agents. For reactions judged to be severe, the cor- leading the authors to conclude that a possible responding fi gures are 0.04 % and 0.004 %. Fatal explanation for the observations was the photo- reactions occur rarely and do not differ between sensitizing effect of the contrast media.

[email protected] 350 10 Contrast Media

Table 10.4 Risk factors for acute reactions to iodinated Table 10.5 Drugs and instruments that should be in the contrast media and procedures and strategies to reduce examination room following injection of contrast media the risks Oxygen Risk factors Epinephrine (adrenaline) 1–1,000

Patient related Patient with history of Histamine H1 -receptor antagonist in injection form − Previous reaction to iodinated Atropine contrast media Beta2-agonist in metered dose inhaler form − Asthma − Allergy requiring medical treatment IV fl uids—physiological saline or Ringer’s solution Contrast High-osmolality ionic contrast media Anti-convulsive drugs (eg., diazepam) media related Sphygmomanometer To reduce the risk One-way mouth breather apparatus For all patients Use a nonionic contrast medium Information from ESUR Guidelines on Contrast Media. Keep patient in Radiology Dept. European Society of Urogenital Radiology, Version 7.0. for 30 min after injection of contrast View at http://www.esur.org/ESUR-Guidelines.6.0.html media Have drugs and equipment for resuscitation readily available (see Table 10.5 ) the points, for example, use of nonionic media, For patients Consider alternative test, i.e., substituting a different contrast medium, and at increased not requiring a contrast medium keeping patients under surveillance longer, have risk of reaction Use different iodinated contrast media been considered above. Physicians using contrast for previous reactors media should be trained to recognize, test for, and Consider premedicationa treat anaphylaxis. Drugs and instruments that Data adapted from ESUR Guidelines on Contrast Media. should be close at hand for acute reaction emer- European Society of Urogenital Radiology, Version 7.0. View at http://www.esur.org/ESUR-Guidelines.6.0.html gencies following administration of contrast a Suitable regime: prednisolone 30 mg (or methyl- media are listed in Table 10.5 . prednisolone 32 mg) orally given 12–2 h before contrast media (see discussion in Sect. 10.6 ) 10.3.3.2 Late Reactions Risk factors for a late skin reaction following administration of an iodinated contrast medium 10.3.3 Risk Factors include current and up to 2 years past treatment with interleukin-2 (IL-2), a history of drug allergy 10.3.3.1 Acute Reactions or contact hypersensitivity, and a history of reac- Risk factors for acute reactions to contrast media tion to a previous contrast medium. Late reac- are summarized in Table 10.4 . The most signifi - tions are more common in patients who reacted cant risk is for patients who have experienced a previously, especially if the same contrast previous immediate reaction to an iodinated con- medium is administered (see below). The latter trast medium. Reexposure to the same or struc- fact is interesting since it suggests that the mech- turally similar ionic preparation is said to carry anism of the late reaction with its demonstration with it a 21–60 % risk of a repeat reaction. This of memory may be immunologically and, in par- risk is one-tenth as great if a nonionic contrast ticular, T cell mediated. medium is substituted for the repeat injection. Comparable fi gures for nonionic media used for the initial and the repeat administrations do not 10.3.4 Biphasic Reactions seem to be available. Other important risks are bronchial asthma, the use of β-blockers, cardiac About 20 % of adverse reactions to iodinated con- disease, and subjects who are highly allergic. trast media are biphasic in nature and although Procedures and efforts to reduce the risks of an severe biphasic reactions are rare they are of con- acute reaction are set out in Table 10.4. Some of cern since they can be life-threatening. The second

[email protected] 10.4 Mechanisms of Adverse Reactions to Iodinated Contrast Media 351

Table 10.6 Fatal biphasic reaction following anaphylaxis to a nonionic contrast medium

Data from Choudhury M et al. Indian J Anaesth. 2011;55:631

or late phase usually occurs after an asymptomatic period of from about 1 h up to 3 days or more (see 10.4 Mechanisms of Adverse Sect. 10.5.1 ) and it can be less, equal, or more Reactions to Iodinated severe than the immediate reaction. A recently Contrast Media reported case summarized in Table 10.6 dramatically illustrates why, after an anaphylactic reaction The range and diversity of adverse effects pro- to a contrast medium, a physician should be wary voked by contrast media remain poorly under- of a second-phase response and patients should be stood and hence difﬁ cult to categorize. For the made aware of the risk on discharge. Although allergist and clinical immunologist used to think- there appears to be no clinical features that can ing of immediate reactions as type I allergic indicate the possibility of a biphasic acute reac- responses mediated by IgE antibodies and tion, it seems that patients who experience the delayed reactions as type IV hypersensitivity delayed response require higher doses of epineph- reactions mediated by antigen-speciﬁ c effector T rine to control their initial reaction. cells, adverse reactions to contrast media, divided

[email protected] 352 10 Contrast Media as they often are into acute and late reactions, do reactions than the low-osmolar compounds not fi t neatly into the conventional mechanism- seems to fi t with research fi ndings showing that based classifi cation (refer to chapter 2 ). the high-osmolar compounds release more histamine than the low- and iso-osmolar contrast media, although some studies have concluded 10.4.1 Anaphylactoid that hypertonicity is not an absolute requirement and Anaphylactic Reactions for contrast media-induced histamine release from human basophils in vitro. Another signifi - The serious severe reactions induced by contrast cant conclusion from in vitro experiments was media show close similarity to other drug- the fi nding that bloods from previous reactors induced anaphylactoid and anaphylactic reac- release a larger percentage of histamine than tions discussed at length in earlier chapters, but, bloods from nonreactors. Just as the opioid drugs as with drugs such as neuromuscular blocking show differences in the amount of histamine they agents, the opioids and other histamine-releasing release and in the sites where release occurs agents (Chaps. 7 and 8), distinguishing the true, (Sect. 8.4 ), contrast media show similar anatomi- immunologically based anaphylactic reactions cal selectivity releasing histamine and tryptase from pseudoallergic or anaphylactoid reactions from human lung and heart mast cells but not caused by release of infl ammatory mediators is from skin mast cells. Depending on the particular generally diffi cult. contrast medium injected, this selectivity may infl uence the symptoms and severity of any sub- 10.4.1.1 Histamine Release sequent reaction to the drug. Among the ionic Preformed histamine when newly and rapidly agents, meglumine salts are more potent releasers released by degranulation of mast cells and baso- of histamine than the corresponding sodium salts. phils accounts for most of the primary Of course, with any case of drug-induced his- manifestations seen in an anaphylactic reaction tamine release, the key question to consider is the (Sect. 3.2.5.1 ), but the usefulness of assessing mechanism underlying the release. Release might histamine release in vitro has not led to general proceed in an immediate reaction by direct action application of the strategy for the diagnosis of of the drug on the mast cells and/or basophils or drug allergies (see Sect. 4.5.2 ). Contrast media of it might be immunologically mediated as in ana- high-, low-, and iso-osmolality are well-known phylaxis mediated by drug-reactive IgE antibod- releasers of histamine, a property demonstrated ies. However, no cell membrane receptors for any in many studies over a period of more than 40 iodinated contrast medium have been identifi ed years, so it is not surprising that the experimental so far and proving an immunological basis even fi ndings have led to the suggestion that released for the most serious acute reactions has been histamine may be the mechanism of severe diffi cult. For the large majority of patients with anaphylactic-like reactions to these agents. High- contrast medium-induced symptoms appearing and low-osmolality contrast media have been within 1 h, IgE antibodies complementary to the shown to produce a rise in plasma histamine that culprit drug cannot be demonstrated (see below peaks and falls back to baseline over a period of for a further discussion of IgE antibodies and about 10 min, but there seems to be no direct contrast media). relationship between the magnitude of the rise Activation of complement by contrast media and the severity of the reaction. Even so, a rela- to produce the anaphylatoxins C3a and C5a has tionship to moderately severe symptoms such as also been proposed as the mechanism for contrast vomiting and rash was suggested and this raised media-induced histamine release. These pro- the question of the probability of histamine’s infl ammatory complement fragments act via spe- contribution to the more severe reactions to con- cifi c receptors on endothelial and mast cells and trast media. The fact that high-osmolar contrast can induce a shock-like reaction similar to that media are responsible for more severe acute seen in type I allergic responses. As yet, however,

[email protected] 10.4 Mechanisms of Adverse Reactions to Iodinated Contrast Media 353 with apparently only a single study showing no reactions (see Sect. 10.5.2 below). The subject has differences in anaphylatoxin levels between been bedeviled by the inconsistencies of results patients and controls, there appears to be no com- obtained from investigations of fatal reactions and pelling evidence either way to accept or refute from a broad group of mild to severe reactors suf- this proposal. fering an acute (immediate) adverse reaction. There are many obvious problems to confront There are also the questions of who is to be inves- in any study designed to examine the role of his- tigated and when investigations should be pursued. tamine (and other infl ammatory mediators) in Despite the fact that many drug reactions occur on adverse reactions to contrast media. Plasma his- fi rst exposure to the drug (for example with neuro- tamine levels peak within 1–8 min following muscular blockers, quinolones, and a wide range of direct treatment with most histamine-liberating different drugs in some individuals), a belief per- drugs (see Sect. 8.4.3 ) and within 5–15 min after sists that without prior sensitization to the drug no antigen challenge, returning to baseline about immunological response, and in particular an IgE 30 min and 60 min later, respectively. There are antibody response, can occur. Much the same atti- obvious diffi culties in being able to select and tude can sometimes be found toward breakthrough study the right patients at the right time and reactions to contrast media after premedication within the required short time frame. Obtaining with antihistamine and cortisone. The problem results, even from small numbers of patients with these poorly informed approaches is that experiencing a severe immediate reaction, some patients with genuine acute reactions that involves many diffi culties plus an element of luck may be antibody mediated, and even potentially on the investigator’s part. The rarity of severe anaphylactic, remain unstudied and undetected. reactions emphasizes the paucity of suitable sub- The question of whether or not iodinated con- jects available for contrast media-induced hista- trast media elicit antibody formation, and in par- mine release studies and probably results in the ticular IgE antibodies, has been considered from examination of too many patients undergoing the viewpoint of whether the drugs can stimulate minor reactions. Ideally, one would like to be antibody formation in the fi rst place. Using what able to perform specifi c skin tests, specifi c IgE was described as “highly favorable conditions for antibody assays (both with the appropriate con- the production of antibodies” that included trols which include skin testing normal subjects Nippostrongylus brasiliensis -infected Hooded with contrast media and IgE-contrast media inhi- Lister rats that are said to be better at producing bition studies), tryptase sampling at suitable antibodies than other strains and the proven adju- times, and quantitation of released histamine. vant Bordetella pertussis, meglumine ioglyca- mate and sodium/meglumine diatrizoate 10.4.1.2 The Question of the conjugated to carrier proteins were employed as Involvement of Contrast immunogens and any subsequent homocytotropic Media-Reactive IgE antibody formation was monitored by the passive Antibodies cutaneous anaphylaxis (PCA) technique. No evi- In true type I immediate allergic responses to dence for the formation of reaginic antibodies drugs, just as with immediate reactions to com- was found leading to the conclusion that “it there- mon inhalant, food, and venom allergens, IgE fore seems unlikely that the majority of adverse antibodies mediate the reactions and one would reactions to radiographic contrast media are aller- therefore anticipate the presence of contrast gic in nature”. Apart from the obvious doubts media-reactive IgE antibodies in the sera of sub- about extrapolating results in rodents to humans, jects showing immediate reactions, especially a number of different drugs in their original severe ones, following injection of contrast media. unbound state may allergically sensitize and elicit Intriguingly, however, positive tests for serum antibody responses by direct interaction with antibodies have been extremely rare and when immune cells and, as with the neuromuscular found they have been in patients with severe acute blocking drugs for example, the sensitizing agent

[email protected] 354 10 Contrast Media may not be the drug in question. Perhaps another prediction was advanced for the neuromuscular study that should be considered is the injection of blocking drugs (Sect. 7.4.2.3 ). a large number of rats or mice with contrast media in an attempt to mimic the situation with 10.4.1.3 Activation of the Kinin human patients and then look for reaginic anti- System and Bradykinin body responses. Considering the human situa- See Sect. 3.2.8.5.2 and Fig. 3.13 ) for a summary tion, one might expect that for this experiment to of the kallikrein–kinin system. be informative, a large number of animals would Bradykinin, a potent vasoactive nonapeptide, have to be examined. A clue to what might be a is formed by interaction of factor XII (Hageman possible immunological function of contrast factor), high molecular weight kininogens and media in vivo is the fi nding that iopamidol had a prekallikrein on negatively charged surfaces (for marked adjuvant effect on the production of anti- example, silicates), on macromolecular surfaces hapten IgE and IgG1 antibodies in mice and such as collagen of connective tissue, heparin and enhancement of antibody production was associ- mucopolysaccharides, and on the surfaces of cells ated with IL-4 release. Antibodies to the contrast together with some specialized proteins including medium were not detected. complement component C1q . Bradykinin is also In the absence of easy-to-perform and reliable produced by a mechanism bypassing factor XII assays for the detection of contrast media- reactive that involves protease activation of prekallikrein. IgE antibodies in patients’ sera, some other less Cell activation during infl ammation and heparin direct but arguably more biologically and clini- release is thought to activate the plasma cascade cally relevant test procedures have occasionally leading to bradykinin release and, via its infl am- been employed. With both the basophil activation matory and hypotensive effects and capacity to and Prausnitz–Kustner (P–K) tests, the interac- induce tissue hyperresponsiveness, its detrimental tion between allergen (contrast medium) and IgE role in asthma, anaphylaxis, and other allergic antibodies takes place at the basophil (or mast conditions. Bradykinin is degraded by carboxy- cell) surface, thus mimicking the in vivo situa- peptidase N and angiotensin-converting enzyme tion. However, for different reasons, neither test and it has been claimed that the latter enzyme is has become, nor is likely to soon become, a rou- inhibited in asthmatics with active bronchospasm tinely and widely applied procedure for the diag- and by ionic contrast media at concentrations nosis of adverse reactions to contrast media or attainable in the circulation. Such an action reduc- any other drug (see below). ing or preventing the hydrolysis of bradykinin and An interesting fi nding with the low-osmolar therefore limiting its effects might help to explain dimer ioxaglate may have some relevance to the the increased susceptibility of asthmatics to con- question of whether or not contrast media- reactive trast media. More fi ndings advanced to support IgE antibodies are part of the mechanism underly- the role of the kinin system in elucidating the ing some reactions to contrast media. Some (but mechanism(s) of contrast media-induced adverse not all) study comparisons have reported more effects are the reported increases in the plasma of reactions to the dimers than to higher osmolar negatively charged heparin-like contact activators ionic media, a similar incidence of severe reactions and so-called cryptic soluble negatively charged by the two, and a lower incidence of monomer- surfaces in subjects who react to contrast media induced fatal reactions. This has prompted the and in asthmatics. An indirect action of bradyki- speculation that the dimers may be antigenically nin also contributes to its pro-infl ammatory divalent, thus allowing them to bridge adjacent effects. By activating phospholipase A2 , the pep- antibody combining sites of mast cell-bound IgE tide stimulates the release of arachidonic acid molecules (see Sect. 3.1.2). The prediction is that from phospholipids leading to the production of if this is so, dimeric iodinated contrast media might prostaglandins and leukotrienes via the cyclooxy- be more likely to induce mediator release and ana- genase (Sect. 9.4.1 ) and lipoxygenase (Sect. phylaxis than univalent monomers. A similar 3.2.5.2.1 ) pathways. Currently it is diffi cult to

[email protected] 10.4 Mechanisms of Adverse Reactions to Iodinated Contrast Media 355 judge the importance of these proposed mecha- non-immediate reactions to iodinated contrast nisms to explain contrast media reactivity since media but a number of different cellular and other supporting and follow-up evidence has not cytokine- driven processes may be involved, mak- been forthcoming. ing this a diffi cult and complex problem to study. Positive patch and delayed intradermal tests, the presence of T cells at skin test sites, positive 10.4.2 Delayed Reactions responses to provocation testing, immunohistological fi ndings, and contrast media-induced prolifer- Delayed-type hypersensitivity reactions to iodin- ation of T cells from patients with delayed reac ated contrast media are said to be rare, but there tions to the agents all give weight to the belief that are some claims that 1–3 % of patients injected these late reactions are mediated by T cells. Results intravenously with the agents experience such from a recent investigation of possible pathways reactions. Symptoms include persistent pain at for recognition of iodinated contrast media by T the injection site, nausea, vomiting, fl u-like cells suggested that two mechanisms of T cell symptoms, angioedema, dyspnea, fi xed drug stimulation were operative. Contrast media-spe- eruption, and maculopapular exanthema. Only a cifi c T cell clones (TCC) were generated from con- small number of cases showing documented evi- trast media-allergic patients and a specifi c T cell dence supporting a diagnosis of delayed hyper- receptor (TCR) was transfected into a mouse T cell sensitivity reactions with positive delayed skin hybridoma. Proliferation and IL-2 and Ca2+ assays tests have been reported. Maculopapular exan- were performed using HLA-DR- matched or mis- thema is the most commonly seen reaction, matched antigen-presenting cells (APC). An accounting for over 50 % of patients with a increase in intracellular Ca2+ within seconds of the delayed reaction to a contrast medium. In one addition of drug, cell proliferation, and IL-2 secre- well-documented case, a 61-year-old patient with tion in the presence of glutaraldehyde-fi xed APCs no history of allergy or prior exposure to contrast suggested that stimulation occurred by direct bind- media developed generalized maculopapular ing to the major histocompatibility complex exanthema 7 days after injection of the nonionic (MHC)–TCR complex. With other TCCs, abroga- agent iopamidol. Three months after the reaction, tion of presentation by glutaraldehyde-fi xed APCs, iopamidol was again administered. Despite pre- failure to wash away drug from APCs preincubated medication with prednisone and cetirizine com- with contrast media, and an optimal pulsing time mencing 3 days before injection, the patient of 10–20 h suggested processing by APCs. reacted 1 day later with generalized, confl uent The precise mediators of contrast media- macular exanthema accompanied by severe itch- induced allergic reaction are also largely ing and enanthema of the oral mucosa. Patch test- unknown, but cytokines are known to participate ing showed a positive allergic reaction to in both immediate and late reactions. iopamidol. In follow-up patch tests, iohexol and Investigations so far of possible cytokine involve- ioversol as well as iopamidol gave positive reac- ment following contrast media injection revealed tions, but the ionic agent sodium amidotrizoate an early increase (after 1 h) in IL-2 followed by a proved negative. Although the ionic compound delayed increase of IL-4 and IL-6, indicating a shares a 2,4,6-triiodobenzene core nucleus with T h 1 to Th 2 shift in late adverse reactions. The the three nonionic drugs, it is structurally differ- highest histamine levels were seen in late reac- ent in the groups attached at positions 1 and 3 of tors 24 h after injection of the contrast medium. the ring where the latter compounds each have TNF-α did not show any signifi cant change. T acetamido groups. This structural difference cell studies, still in their early days and so far on accounts for cross-reactivity of the nonionic small numbers of patients, have generated iodin- agents and the absence of it for the ionic drug. ated contrast media-specifi c T cell clones and Some investigators believe that cell-mediated demonstrated cross-reactivity with some other hypersensitivities are responsible for most of the contrast media by some of the CD4(+) clones.

[email protected] 356 10 Contrast Media

Iomeprol-specifi c peripheral T cells for example patients and prick tests in only 4. An early and were shown to occur with a frequency of 0.6 %. obvious fi nding was the relationship between the skin test result and the time between reaction and testing. Within 2–6 months of the adverse reac- 10.5 Tests for the Diagnosis tion, the percentage of positive reactors increased and Study of Adverse from the overall fi gure of 26 to 50 % (14/28). For Reactions to Contrast Media patients tested at other times, that is, earlier than 2 months and later than 6 months, the fi gure was As always, a meticulously recorded and studied only 18 % (17/92). history is desirable, but additional clinical and For non-immediate reactors, a combination of laboratory tests can contribute greatly to an accu- intradermal and patch tests were needed to iden- rate diagnosis and help to identify the tify the maximum number of reactants. Delayed mechanism(s) of the reaction. Skin tests, detec- skin tests were positive in 37 of 98 patients tion, and quantitation of released mediators and (38 %) with only three positive in the skin prick serum IgE antibodies may all contribute to estab- test. Of 31 patients positive in the intradermal lishing a more precise diagnosis of an adverse test, 9 delayed reactions were detected on day 1, response to an iodinated contrast medium. 16 on day 2, and 6 on day 3. Patch tests required times of up to 3 days to detect all of the 22 positive patients out of 79 tested (28 %). Some 10.5.1 Skin Tests patients were positive in the intradermal but not in the patch test while with some others it was the For a general discussion of skin testing, see reverse. As with the immediate reactors, most Sect. 4.2 . reactors (29 of 62; 47 %) were detected when the Although there is no long-established and tests were performed within 6 months of the widespread diagnostic practice of skin testing reactions to contrast media; only 8 of 36 (22 %) patients with contrast media, results from recent were positive at later times. Cross-reactions studies indicate that skin testing with iodinated between some contrast media were detected contrast media is a useful tool in efforts to especially between those of similar structure, for improve the diagnosis of allergy to these agents. example, iophexol, iomeprol, iopentol, ioversol, Some small skin test studies and tests on indi- and the nonionic dimer iodixanol. From the viduals have been carried out intermittently over wider perspective of the mystery of how some the years, but a recent European multicenter pro- patients become allergically sensitized to some spective study carried out under the auspices of drugs (see for example, Sect. 3.1 ), it is interest- the European Network of Drug Allergy and the ing to note that one-third of the patients with a European Academy of Allergy and Clinical positive delayed skin test in this study reacted to Immunology Interest Group on Drug contrast media on their fi rst exposure to the Hypersensitivity set out, for the fi rst time, to agent. In what may turn out to be a useful obser- determine in a large study the specifi city and sen- vation, patients with a delayed positive skin test sitivity of skin tests in patients who experienced showed a higher number with maculopapular reactions to iodinated contrast media. Skin prick exanthema and a lower number with urticaria- tests followed by intradermal tests and patch tests like exanthema than the patients who were skin were performed on 220 patients with a reported test negative. previous hypersensitivity reaction, either imme- The authors of the European multicenter study diate or delayed, to contrast media using the dilu- pointed out some limitations in their report. tions, procedures, and controls summarized in These included lack of provocation testing, the Table 10.7 . Overall, positive skin tests were seen need for more control subjects exposed to a in 32 of 122 patients (26 %) with immediate reac- contrast medium but without clinical signs of a tions. Intradermal tests were positive in 30 hypersensitivity reaction, the culprit drug was not

[email protected] 10.5 Tests for the Diagnosis and Study of Adverse Reactions to Contrast Media 357

Table 10.7 Skin testing details and procedures for testing patients with suspected hypersensitivity reactions to iodinated contrast media Concentration of test solutiona Test and how applied Site of test Reading times Positive reactionb,c Skin prick testd Undiluted contrast media Volar forearm After 20 min Immediate reaction: solution. and on days 2 and 3 Wheal ≥ 3 mm after 20 min For standard procedure, Delayed reaction: see Sect. 4.2.2 Erythematous induration at site on day 2 or 3 Intradermal teste Contrast media solution Volar forearm After 20 min and on Immediate reaction: Initial diluted 1–10. or back days 1, 2 and 3 wheal increased by at least Inject 0.03–0.05 ml to give 3 mm diam. and surrounded 4–5 mm diam. bleb . by erythema after 20 min Delayed reaction: Erythematous induration at site at delayed reading Patch testf Undiluted contrast media Back Up to 3 days. Read For reading and scoring Filter paper soaked in 15 min after removal patch test reactions, see contrast media and placed and 24 h later Sect. 4.2.4.3 , Fig. 4.5 and in 12 mm aluminum Finn Table 4.1 chamber Fixed on back with adhesive tape for up to 3 days Data from Brockow K et al. Allergy. 2009 ;64:234 a All solutions were 300–320 mg iodine/ml b Controls: positive—histamine solution 0.01 %; negative—saline 0.9 % c Skin test sensitivity—the percentage of skin test-positive patients showing a typical hypersensitivity reaction after administration of contrast media; skin test speciﬁ city—percentage of negative controls with a positive skin test to contrast media d See Sect. 4.2.2 e See Sect. 4.2.3 f See Sect. 4.2.4

always identifi ed, a possible lack of test sensitiv- 10.5.2 IgE Antibody Tests ity, and the fact that the negative predictive value of the skin tests had not been determined. In rela- 10.5.2.1 Prausnitz–Kustner Test tion to the last point, a negative predictive value This test relies on the capacity of homocytotropic of 96.6 % has recently been claimed for tests with antibodies or reagins to fi x to human skin and be iodinated contrast media (see Sect. 10.5.4 ), but, detected by subsequent injection of the suspected to be confi dent of this fi gure, studies with larger allergenic drug. Although a positive reaction is numbers of patients need to be done. Despite not immediately indicative of the involvement of these limitations and some others, most of which IgE antibodies, their presence is often inferred if are not necessarily easy to overcome, the main skin sensitization is prevented by prior heating of and most important conclusions from the fi rst 4 the serum to 56 °C. Prior to the realization of the years of this prospective study are that about half possibility of the transfer of blood-borne viruses, of the hypersensitivity reactions to iodinated con- and the introduction of easier-to-carry-out tests trast media have an immunological basis and skin that specifi cally identify IgE, the P–K test was testing, especially intradermal and patch testing, used in studies aimed at determining whether or is a useful diagnostic tool that may aid the selec- not the antibody was implicated in some adverse tion of a safe contrast agent for those patients reactions to iodinated contrast media. In one case who have experienced a previous reaction. study, a positive P–K test was taken as evidence

[email protected] 358 10 Contrast Media that an immediate reaction to ioglycamic acid (see Sect. 4.6 ) is an in vitro activation of a was mediated by IgE antibodies. patient’s basophils and as such the test mimics the interaction between the allergen and circulating 10.5.2.2 Serum Tests basophils in the patient’s body. The test is not, For details of the detection of drug-reactive IgE however, a primary diagnostic tool and is essen- antibodies in sera, see Sect. 4.3 . As already out- tially complementary to skin tests and quantita- lined, IgE antibodies to iodinated contrast media tion of allergen-reactive IgE antibodies. Despite have not been convincingly and consistently references to its application to the investigation of demonstrated and the current consensus is that contrast media adverse reactions, there currently true type I IgE antibody-mediated reactions to appears to be very few published studies where it the drugs are rare, but they do occur, generally in has been applied in this way. In a 2008 study the the most severe immediate cases. This conclu- contrast media iomeprol and iopromide were sion, or impression, may be correct, based as it is diluted and used over a broad range up to 1 μg/ml on the antibody detection methodologies applied and a minimum of 500 basophils per sample were to date, but consistent failures to detect the anti- activated (CD63+, IgE++) and assessed by fl ow bodies may be due to the inadequacies or inap- cytometry. Because drugs give lower activation propriateness of those methodologies. Even with percentages than inhalant and venom allergens, the small number of positive sera detected in activation was considered positive if data analysis some studies, reactions are often weak, quantita- showed more than 5 % activated basophils. In tive inhibition results to demonstrate specifi city three patients the test revealed 15 % maximum are not shown, and details of the materials and activation of basophils at 1 μg/ml. Two patients methods used are not provided. The average showed positive results only with iomeprol while association constant of IgE for contrast media the third was positive to both contrast media. In was low in what appears to be the only study the control samples, activation remained negative where the affi nity of the reaction was looked at. at all contrast media concentrations. A recent The fi rst more convincing demonstration of the study in Thailand on 26 patients with diagnosed detection of contrast media-reactive IgE antibod- immediate reactions to contrast media and 43 ies involved activation of the hemisuccinate of healthy volunteers found signifi cantly higher per- ioxaglic acid to form the N -hydroxysuccinimide centage activations at drug dilutions of 1:10 and ester before linking to amino groups on human 1:100 with the patients’ than the volunteers’ cells. serum albumin as carrier protein (Fig. 10.2 ) and Once again the conclusion was reached that the using the drug–carrier complex in immunoassays basophil activation test has potential as a diagnos- with patients’ sera. Although binding uptakes tic tool, especially as a confi rmatory test. were weak, specifi city of the reaction with the contrast medium was demonstrated by dose- dependent inhibition in the range 25–80 % and 10.5.3 Tests to Detect the Release IgE antibody was detected in 16 of 34 patients of Mediators (47 %) with a history of adverse reactions to ioxaglate and in 14 of 68 patients whose sera were From at least the early 1980s, studies relevant to collected at the time of the adverse reaction to the the possibility of monitoring contrast media- contrast agent. The frequency of 47 % seems induced histamine release either directly or indi- consistent with a previous claim of 42 % for the rectly via measurement of urinary methylhistamine presence of IgE antibodies in patients with an have been pursued. Although concentrations of adverse reaction to a contrast medium. histamine and its metabolite have been shown to increase in some patients who had an adverse reac- 10.5.2.3 Basophil Activation Test tion to a contrast medium, diagnostic tests for Unlike in vitro methodologies that detect binding these mediators have not often been used and it of serum IgE antibodies, usually to an allergen in was not until an assay for tryptase (see Sect. 4.5.1 ) solid phase form, the basophil activation test became widely available that routine measurement

[email protected] 10.5 Tests for the Diagnosis and Study of Adverse Reactions to Contrast Media 359

CH3 COOH N COCH3 III I

HO -CH2CH2NHCO NHCOCH2NHCO CONHCH3 I I Ioxaglic acid (IOX)

DMF / triethylamine Succinic anhydride

CH3

COOH N COCH3 III I

HOOC - CH 2 CH 2 COO -CH2CH2NHCO NHCOCH2NHCO CONHCH3 I I Succinyl IOX

Dicyclohexylcarbodiimide N-hydroxysuccinimide + HSA

CH3

COOH N COCH3 III I

HSA - NHCO - CH2 CH2 COO -CH2CH2NHCO NHCOCH2NHCO CONHCH3 I I HSA-IOX Conjugate

Fig. 10.2 Preparation of a drug–carrier complex of ioxaglic acid with human serum albumin for use in a solid phase assay for the detection of drug-reactive IgE antibodies. Structures in bold show reactive groups and point of attachment of albumin of a mast cell mediator became almost standard produced by the human mast cell, is preferred in practice. In one of the most informative studies, diagnostic investigations of severe immediate but released histamine and tryptase levels correlated not delayed allergic reactions. signiﬁ cantly with the severity of symptoms to contrast media and it was suggested that less clear results from many previous investigations may 10.5.4 Challenge Tests have been due to the recruitment of patients with minor or only moderate reactions. The relative Although considered to be the “gold standard” in half-lives of histamine (about 2 min) and tryptase the diagnosis of drug hypersensitivity, challenge (about 90 min) give another indication of why tests (Sect. 4.4) are time-consuming, are poten- the latter mediator, the most abundant protein tially dangerous, and tend to be reserved for use

[email protected] 360 10 Contrast Media

Table 10.8 Challenge protocol for iodinated contrast media probably be said that the practice has not received Written informed consent obtained from each patient wide support. There are many reports of break- Time interval should be at least 6 weeks since anaphylactic through reactions—some detailing that hypersen- reaction sitivity responses were not prevented in a number Patient observed during whole period of challenge of patients; some reporting a signiﬁ cantly higher Emergency drugs and equipment available recurrence rate in those who had a previous mild Challenge doses increased stepwise at 30 min intervals reaction but prevention of a reaction in those who up to a normal dose. Doses used: 0.05, 0.5, 1, 5, 7.5, 10, 25 ml. Total = 49.05 ml had a severe previous reaction; and a frequent ﬁ nding that breakthrough reactions were of simi- From Trcka J et al. Am J Roentgenol. 2008 ;190:666 lar severity to the patients’ initial reactions. The theory behind the inclusion of histamine

H 1 antagonists in the premedication is obvious but when the information gathered from other tests the mode of action of corticosteroids is not com- yields inconclusive results or contradictions. pletely understood so some believe its inclusion Challenge tests with contrast media have so far cannot be explained and justiﬁ ed. Corticosteroids also been employed rarely but appear to be valu- ultimately inhibit kallikrein, a peptide that lowers able to identify contrast media that are tolerated blood pressure and liberates bradykinin. by challenging intravenously skin test-positive Corticosteroids also act in the arachidonic acid patients with skin test-negative agents. In cascade to inhibit the production of prostaglan- addressing the paucity of data on the negative dins and leukotrienes, so there does seem to be predictive value for skin tests with iodinated con- some rationale for their use. There are some indi- trast media, 29 skin test-negative patients need- cations that premedication prevents the recurrence ing a new contrast medium were rechallenged of many minor reactions. Some, or even many, of without premedication. Mild reactions resulted in these reactions may not be immune mediated, pro- 1 of 24 patients with a history of an immediate ceeding instead via a nonspeciﬁ c and low-level reaction and one of four with a history of a non- histamine release. In the case of severe immediate immediate reaction, giving a negative predictive reactions, IgE antibody-mediated explosive hista- value of 96 %. Two patients with a positive skin mine release from mast cells may overwhelm the test to an iodinated contrast medium tolerated an potential effectiveness of premedication. alternative drug without experiencing a reaction. Despite calls to discontinue the prophylactic A protocol for challenge with contrast media is use of corticosteroids and antihistamines for con- set out in Table 10.8 . trast medium-induced anaphylactoid reactions in the USA, the recommendation for their use is still unaltered. This situation is in contrast to the atti- 10.6 Premedication for the tude in some other countries, for example, Prevention of Anaphylactoid/ France, but one wonders what the attitude of the Anaphylactic Reactions to critics of premedication would be if they faced Iodinated Contrast Media injection of an iodinated contrast medium for angiography after experiencing a life-threatening From the results of studies in the USA dating reaction knowing they were allergic to both ionic back to the mid-1970s, it was recommended that and nonionic media of all osmolalities. Given our lower osmolality contrast media should be given knowledge of the pharmacological effects of his- to patients who had previously experienced what tamine H 1 antagonists and the steroids, could was called an “immediate generalized reaction.” there be anything to lose in opting for premedica- In addition, the prophylactic use of prednisone tion in such a situation? A suitable premedication and diphenhydramine was recommended to regime is set out in the legend of Table 10.4 . reduce the chance of a reaction in high-risk Of course, for high-risk patients when admin- patients. Such premedication is often given, but istration of an iodinated contrast medium is opinion of its effectiveness is divided and it could regarded as essential, a nonionic agent would be

[email protected] 10.7 Gadolinium-Based Contrast Agents 361 the ﬁ rst choice, but if that choice is also poten- set out in Fig. 10.4 . Stability of gadolinium che- tially dangerous and cannot be avoided, the lates is a major concern and great emphasis has patient should be informed of the risks, patient been placed on this because of the possibility of approval should be obtained, and resuscitation transmetallation, that is, exchange or release of arrangements should be fully in place. free Gd3+ . With the Gd3+ caged within the chelate Gadolinium-based contrast media may also be complex, the macrocyclic compounds are gener- considered, but with these agents some extra fac- ally more stable than their acyclic counterparts. tors need to be considered.

10.7.2 Nephrogenic Systemic 10.7 Gadolinium-Based Contrast Fibrosis Agents Identifi ed in 1997 and reported in 2000, nephro- Gadolinium, a rare earth metal, forms trivalent genic systemic fi brosis (NSF) was fi rst associated ions with paramagnetic properties that make solu- with gadolinium contrast agents in 2006. Chronic tions of chelated gadolinium complexes with large kidney disease, hepato-renal syndrome with renal organic molecules useful as intravenously admin- insuffi ciency, and acute kidney injury were istered contrast agents detected by magnetic reso- described as the clinical settings in early case nance imaging (MRI). Gadolinium- based contrast reports and it soon became apparent that the dis- agents are approved by the FDA (and many other ease affects multiple organs including the lungs, licensing authorities) for use with MRI as a con- heart, liver, and muscles. In patients with reduced trast agent, but, although they can be used for renal function, the prevalence of NSF after expo- magnetic resonance angiography (MRA), there is sure to gadodiamide is reported to be 3–7 %. Of no approval for this use. The usual dose for many 589 patients who developed NSF associated with MRI applications is 0.1 mmol/kg up to a maxi- gadolinium contrast agents between 1997 and mum approved dose of 0.3 mmol/kg for intrave- 2007, all were to linear chelates—68 % with nous use. Above that fi gure the agents may induce gadodiamide, 26 % with gadopentetate, and 5 % potentially fatal nephrogenic systemic fi brosis with gadoversetamide. Subsequent to 2006, of (NSF), a scleroderma- and eosinophilic fasciitis- 1,603 cases reported to the FDA, 93 % were from like disease of the joints, skin, eyes, and organs 60 hospitals in the USA and 4 % from 2 hospitals particularly in patients with kidney failure. in Denmark. Gadodiamide was revealed to be a key factor in the relatively high incidence of NSF found in Denmark. The macrocyclic agents are 10.7.1 Molecular Structures regarded as low-risk compounds. By 2009, there of Gadolinium-Based had been no cases of NSF after exposure to a Contrast Agents macrocyclic, but three cases were reported in Denmark in 2011. These agents may be acyclic (linear) or macrocyclic and ionic or nonionic. Examples from each category are acyclic, ionic—gadopentetate 10.7.3 Other Adverse Reactions (Gd-DTPA) dimeglumine; acyclic, nonionic— gadodiamide (Gd-DTPA-BMA); macrocyclic, Risk factors for acute reactions to gadolinium- ionic—gadoterate (Gd-DOTA) meglumine; and based contrast media and procedures and strate- macrocyclic, nonionic—gadoteridol (Gd-HP- gies to reduce those risks are summarized in DO3A). The structures of the acyclic agent gado- Table 10.9 . Note that the risk of an adverse reac- pentetic acid (gadopentetate dimeglumine is the tion to a gadolinium-based contrast medium is salt) are shown in Fig. 10.3 and structures of three eight times higher in patients who have experi- macrocyclics, gadoterate meglumine, gadoteridol, enced a previous reaction to a gadolinium agent. and the nonionic gadobutrol (Gd-BT- DO3A) are Gadolinium chelates have been used parenterally

[email protected] 362 10 Contrast Media

a b O

O N - O O N - - O O - N - O O O - O O O O NNGd3+ Gd3+ - - O O N - O OH OO2 - O O

OH2

Fig. 10.3 2D structure (a , b), ball-and-stick (c ), and CPK dimeglumine, a complex of Gd3+ with diethylenetriamine- ( d ) models of gadopentetic acid, the ﬁ rst gadolinium- pentacetate (DTPA 5− ) (Gd-DTPA) with the nine-coordi- based magnetic resonance imaging contrast agent intro- nate Gd ion surrounded by three nitrogens and ﬁ ve duced in 1987. This linear, ionic agent is usually oxygens and the ninth site occupied by a water molecule administered as the meglumine salt gadopentetate ( a , b ) for over 30 years and are well tolerated in the vast site. “Allergic-type” reactions have an even lower majority of patients. Adverse reactions to gado- frequency of 0.004–0.7 % with symptoms of linium contrast media appear to be less frequent rash, urticaria, and rarely bronchospasm. than reactions to the iodinated media. The Anaphylactic/anaphylactoid reactions are American College of Radiology (ACR) extremely rare (0.001–0.01 %). In one study, an Committee on Drugs and Contrast Media has adverse reaction rate of 0.48 % and an incidence reported that the frequency of all adverse events of 0.01 % for severe anaphylactoid reactions after injection of 0.1–0.2 mmol/kg of gadolinium were reported for gadolinium chelates. Of the 45 chelate is in the range 0.07–2.4 % with the vast patients with 46 adverse reactions, 96 % were majority of these reactions being mild, for exam- mild reactions, 2 % were moderate, and 2 % ple, nausea, vomiting, headache, paresthesia, severe. Three (6.7 %) of the 45 patients had prior dizziness, itching, and coldness at the injection reactions to iodinated contrast media. In an

[email protected] 10.7 Gadolinium-Based Contrast Agents 363

Macrocyclic- structure MRI Structure Trade name contrast agent

O-

Gadoterate N N O Artirem, - 3+ - or sodium O Gd O Dotarem (Gd-DOTA) O N N (ionic) +(1) Meglumine + -O or Na

O-

N N Gadoteridol - 3+ (Gd-HP-DO3A) O Gd OH ProHance (non-ionic) O N N

-O

O-

N N O Gadobutrol Gadovist, 3+ - (Gd-BT-DO3A) HO Gd O Gadavist, Gadograft, (non-ionic) HO N N Protovist

-O HO

(1) Meglumine: OH OH OH N H OH OH

Fig. 10.4 Structures of three macrocyclic gadolinium-based magnetic resonance imaging contrast agents

[email protected] 364 10 Contrast Media

Table 10.9 Risk factors for acute reactions to iodinated contrast media which is consistent with gadolinium- based contrast media a and procedures and a previous conclusion that the risk of adverse strategies to reduce the risksb,c reactions to gadopentetate is 3.7 times higher in Risk factors patients with a history of reaction to iodinated Patient related Patient with history of: contrast media. Gadopentetate dimeglumine was − Previous acute reaction to gadolinium contrast media the agent most often implicated being the admin- − Asthma istered drug in 29 of the 36 patients (0.138 %) − Allergy requiring medical treatment including both of the life-threatening reactions Contrast Note: Risk not related to osmolality. (0.01 %). These 2 % for gadopentetate differ sig- media related Low dose makes osmolar load small nifi cantly from an earlier investigation of the To reduce the risk safety of the same agent where the overall inci- For all patients Keep patients in Radiology Dept. for 30 min after injection of contrast media dence of adverse events was higher (1–2 %) Have drugs and equipment for while the incidence for anaphylactoid reactions resuscitation readily availabled was only 0.0003 %. For comparison, incidences For patients at Consider alternative test not requiring a of life-threatening reactions to iodinated contrast increased risk gadolinium agent media are said to be 0.031 % for low-osmolarity of reaction Use a different gadolinium agent for previous reactors agents and 0.157 % for high-osmolality agents. Consider premedicatione A recent survey of the incidences of immediate a Non organ specifi c reactions to gadolinium contrast media revealed bData adapted from ESUR Guidelines on Contrast Media. rates of 0.2, 0.5, 1.2, and 3.3 per 1,000 injections European Society of Urogenital Radiology, Version 7.0. of gadodiamide, gadopentetate dimeglumine, View at http://www.esur.org/ESUR-Guidelines.6.0.html gadobenate dimeglumine, and gadoteridol, cRisk of an acute reaction to gadolinium contrast media is signifi cantly lower than the risk with an iodinated contrast respectively. For the period 2004–2009, the FDA media received reports of 40 NSF- unrelated deaths d See Table 10.5 resulting from 51 million administrations of gad- e See legend of Table 10.4 for a suitable premedication olinium contrast agents with incidences of 0.15, regime 0.19, 0.7, 0.97, and 2.7 per million for gadodiamide, gadoversetamide, gadoteridol, gadopen- assessment of the clinical safety and diagnostic tetate dimeglumine, and gadobenate dimeglumine, value of the gadolinium chelate gadoterate respectively. It has been pointed out that these meglumine in 24,308 patients injected with the fi gures represent a similar risk of death from trav- agent, adverse reactions were seen in 0.4 % of the eling 86 miles by car! With gadoterate meglu- examinations and were mostly rated as minor, mine, positive skin tests, sometimes with a that is, feelings of warmth or taste alterations. positive leukocyte histamine release test, indi- Only one serious adverse reaction was seen. A cated that the reactions were almost certainly IgE review of 21,000 patients administered gadolin- antibody mediated. Positive skin tests and a posi- ium contrast media in a Michigan hospital tive tryptase fi nding were also found in a case of revealed 36 adverse reactions (0.17 %) classifi ed anaphylaxis to gadobenate (Gd-BOPTA) into four groups: mild, nonallergic reactions dimeglumine. Skin testing with other gadolinium (nausea, vomiting) 15 patients; mild reactions chelates generally revealed mono-sensitization resembling allergy (hives, erythema, skin irrita- with none of the other agents showing cross-reaction) 12 patients; moderate reactions resembling tions. In some of the investigations, skin test-neg- allergy (respiratory symptoms) 7 patients; and ative fi ndings with high concentrations of life-threatening reactions resembling allergy meglumine ruled it out as the provoking agent and (chest tightness, respiratory distress, periorbital direct and explosive mast cell degranulation edema) 2 patients (0.01 %). Reactions resem- by gadolinium chelates in the reported cases bling allergy therefore occurred in 21 patients. seems unlikely since in vitro experiments have Four of the patients had previous reactions to shown that the concentrations needed for direct

[email protected] Summary 365 histamine release are about 100–400 times the • Contrast media are better tolerated when the normal serum concentrations found in patients. osmolalities of the injected media and body The skin test fi ndings so far with the gadolinium fl uids are as close as possible. Nonionic media chelates suggest that this simple and easy-to- are better tolerated than the ionics. carry-out test might be useful for identifying • Adverse reactions are divided into acute or alternative MRI contrast agents, but more exten- immediate and late or delayed. The former sive testing with many more patients is needed to occur within an hour and the latter from about validate the procedures before the predictive 1 h up to a week but usually 1–3 days. value of skin testing can be established. As with • Acute reactions are conveniently divided into the iodinated contrast media, breakthrough reac- mild (with symptoms including nausea, vom- tions to gadolinium contrast agents, sometimes iting, and headache), moderate (tachycardia/ described as “allergic-like,” have occurred after brachycardia, marked urticaria, severe vomit- corticosteroid and antihistamine premedication. ing), and severe (hypotensive shock, cardiac In some cases where the history was reliable and respiratory arrest, laryngeal edema). Late enough and where questioning about exposure to reactions include nausea, vomiting, and espe- products containing gadolinium (for example, in cially (and usually self-limiting) maculopapu- CDs, electronic components, nuclear materials, lar rash, exanthema, urticaria, and pruritus. alloys, phosphors, optical glass, ceramics, and • Severe biphasic reactions to iodinated contrast manufacturing plants where the element is used) media, that is, a life-threatening late reaction was undertaken, it became apparent that reac- after an initial acute immediate reaction, are tions to gadolinium chelates occurred on fi rst rare but can occur. After discharge following exposure. As discussed before in this volume, an acute immediate reaction to a contrast this is not unusual in drug allergy and while a medium, patients should be made aware of the number of different speculative explanations risk of a second-phase response. have been offered for the mechanism of sensitiza- • The incidence of acute (immediate) reactions tion for some other drugs such as neuromuscular to the ionic media is about 3–4 % with up to blockers, the advancement of any sort of plausi- about 12 % reported. For the low-osmolar ble speculation accounting for allergic sensitiza- nonionic agents, the fi gure is 0.2–0.7 % (up to tion to complexes containing a rare earth metal about 3 %). For severe immediate reactions seems even more diffi cult. (mainly anaphylactic), incidences are 0.1– 0.4 % for ionic and 0.02–0.04 % for nonionic media. For very severe reactions the percent- Summary ages drop to 0.04–0.004 %. • Fatal reactions (1 in 100,000 to 1 in 170,000) Iodinated Contrast Media are extremely rare and show no differences • Considering the very large number of admin- between low- and high-osmolar agents. istrations worldwide, iodinated contrast media • Up to 80 % of reactions to an ionic agent can be are one of the safest of all drugs. avoided by substituting a nonionic medium. • Reactions to iodinated contrast media can be • For delayed reactions, there is no difference dose-dependent (toxic reaction) or unrelated between the incidences of reactions to ionic to the dose (for example, an immunological and nonionic media or between the different reaction). nonionic agents. Incidences of delayed reac- • Reactions range from a mild inconvenience tions occurring in the fi rst 24 h and over a such as heat sensation and nausea to a life- 7-day period are approximately 4 % and threatening emergency. 1–3 %, respectively. • In the great majority of cases reactions are • Risk factors: for immediate reactors—a previ- mild and direct histamine release can account ous immediate reaction to an iodinated confor the symptoms. trast medium; bronchial asthma; use of

[email protected] 366 10 Contrast Media

β-blockers; cardiac disease; highly allergic • A signifi cant number of immediate and subject. For late reactors–a previous reaction; delayed reactors with positive skin tests to treatment with IL-2; a history of drug allergy; contrast media reacted on fi rst exposure to the contact allergy. agents. • Evidence that immediate reactions to a con- Gadolinium-Based Contrast Agents trast medium is IgE antibody mediated is • Gadolinium, a rare earth metal, forms ions based largely on skin test results with or with- with paramagnetic properties making the toxic out tryptase measurements and very occasion- metal in chelate form a useful contrast agent ally serum IgE and basophil activation tests. that can be detected by magnetic resonance • Delayed reactions mainly manifest as exan- imaging. thematous skin eruptions. They are mediated • Gadolinium-based contrast agents were associ- by antigen-specifi c effector T cells with as yet ated with nephrogenic systemic fi brosis in 2006. poorly defi ned cytokine involvement. The linear chelates, and especially gadodiamide, • Contrast media-induced hypersensitivity has are most commonly implicated. The macrocy- traditionally been regarded as nonallergic in clic agents are regarded as relatively safe. nature with skin testing not relevant. Skin tests • Gadolinium chelates show an adverse reaction with iodinated contrast media are, however, incidence of about 0.48 % and an incidence of positive in a subgroup of reactors. about 0.01 % for anaphylactoid reactions. • In a large multicenter study, 50 % of immedi- This is lower than the corresponding fi gures ate reactors tested within 2–6 months of the for iodinated contrast media. reaction showed a positive skin test. The prick • There are a number of reports of anaphylaxis test was only rarely positive. The intradermal to gadolinium-based contrast media with the test was clearly more informative. diagnosis supported in some cases by positive • For delayed reactors, a combination of intra- skin tests to the agents. dermal and patch testing identifi ed the maxi- • Some adverse reactions occurred on fi rst mum number of positive reactions (47 %). exposure to the contrast agents. The highest number of positives was detected • Gadolinium contrast media are well tolerated when tests were performed within 6 months of by the vast majority of patients and are the reaction. regarded as remarkably safe drugs. • Cross-reactions between iodinated contrast media were detected in skin tests on delayed reactors making intradermal and patch tests Further Reading useful tools for selecting a safe contrast medium. ACR manual on contrast media, version 7. ACR committee on drugs and contrast media. American College of • IgE antibodies to iodinated contrast media have Radiology; 2010 not been consistently and convincingly demon- Brockow K, Romano A, Aberer W, et al. Skin testing in strated. This raises doubts about patient selec- patients with hypersensitivity reactions to iodinated tion (that is, the degree of severity of patient contrast media – a European multicenter study. Allergy. 2009;64:234–41. reactions) and the adequacy and appropriate- Caravan P, Ellison JJ, McMurry TJ, et al. Gadolinium (III) ness of the present IgE test methodologies. chelates as MRI contrast agents: structure, dynamics, • Challenge tests with contrast media, rarely and applications. Chem Rev. 1999;99:2293–352. employed and refused by most patients, are ESUR guidelines on contrast media. European society of urogenital radiology, version 7.0 view. http://www. valuable for the identifi cation of tolerated skin esur.org/Contrast-media.51.0.html test-negative agents. Morcos SK. Acute serious and fatal reactions to contrast • Opinion on the effectiveness of premedication media: our current understanding. Br J Radiol. with corticosteroids and a histamine H - 2005;78:686–93. 1 Murphy KJ, Brunberg JA, Cohan RH. Adverse reactions antagonist is divided since many breakthrough to gadolinium contrast media: a review of 36 cases. reactions have been reported. AJR Am J Roentgenol. 1996;167:847–9.

[email protected] Further Reading 367

Prince MR, Zhang H, Zou Z, et al. Incidence of immediate Trcka J, Schmidt C, Seitz CS, et al. Anaphylaxis to iodin- gadolinium contrast media reactions. AJR Am J ated contrast material: nonallergic hypersensitivity or Roentgenol. 2011;196:W138–43. IgE-mediated allergy? AJR Am J Roentgenol. 2008; Thomsen HS. ESUR guideline: gadolinium-based con- 190:666–70. trast media and nephrogenic systemic ﬁ brosis. Eur Webb JAW, Stacul F, Thomsen HS, et al. Late adverse Radiol. 2007;17:2692–6. reactions to intravascular iodinated contrast media. Thomsen HS. Nephrogenic systemic ﬁ brosis: history and Eur Radiol. 2003;13:181–4. epidemiology. Radiol Clin North Am. 2009;47:827–31.

[email protected] Biologics 1 1

Abstract Currently, ~28–30 mAbs are approved or under consideration for approval as specifi c therapies in the USA or European Union, although about 350 new mAbs for therapeutic application in humans are in the commercial pipeline. So far, the number of target antigens for the mAbs is surprisingly small with more than one of the approved antibodies specifi c for TNF, HER2, CD20, EGFR, or VEGF. Other specifi cities include EpCAM, glycoprotein IIb/IIIa, CD30, CD52, C5, α-4 integrin, IgE, IL-6R, BLys, IL-1β, and RANK-L. Initial infusion reactions to some mAbs may provoke tumor lysis syndrome, cytokine release syndrome, and systemic infl ammatory response syndrome. Systemic and cutaneous reactions also occur to mAbs. Rituximab, for example, may cause serum sickness, vasculitis, cutaneous reactions, interstitial pneumonitis, and ARDS as well as post-infusion reactions. Some patients receiving cetuximab experienced severe immediate hypersensitivity reactions. The antibodies involved are IgE specifi c for α-D -galactose-(1–3)- β-D - galactose and reactive with this disaccharide present on the Fab portion of the chimeric antibody. The nature of, and main adverse reactions to, etanercept, the synthetic IFNs pegylated IFNα-2a and pegylated IFNα-2b, IL-2, denileukin diftitox, anakinra, afl ibercept, anti-thymocyte globulin, epoetins, and recombinant human insulin are discussed.

Until the late 1990s, the backbone of treatments diseases, systemic lupus erythematosus, asthma, for malignancies was the administration of cyto- and for the prevention of organ transplant rejec- toxic chemotherapeutic agents. However, from tion. This new class of therapeutic drug, often about 1998 and with ever expanding usage over referred to simply as “biologics,” encompasses an the last 14 years, passive immunotherapy with tar- expanding diversity of agents, many of which are geted agents, generally monoclonal antibodies, is genetically engineered copies or modiﬁ cations of being used to manage various human cancers, natural products of the body’s immune system. some autoimmune diseases such as rheumatoid Biologics may be composed of proteins, nucleic arthritis and Crohn’s disease, some cardiovascular acids, sugars, or combinations of these; they may

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 369 Relationships, DOI 10.1007/978-1-4614-7261-2_11, © Springer Science+Business Media, LLC 2013

[email protected] 370 11 Biologics be of human, animal, or microbial origin; and, in 11.1.1 Nomenclature the vast majority of cases, they are created by biological processes, or biotechnology, rather than The nomenclature of monoclonal antibodies is chemical synthesis. Besides recombinant thera- the one used by both the U.S. Adopted Names peutic proteins such as some antibodies, cyto- (USAN) and World Health Organization’s kines, and receptors, included in any list of International Proprietary Names (INN) for phar- biologics would be gene therapies, somatic cells, maceuticals. All of the monoclonal antibody adult and embryonic stem cells, vaccines, tissues, names end with the stem -mab and use preceding and blood products and components. substems depending on the structure and function of the antibodies. Different modifi ed suffi xes or stems are added to distinguish the origins of 11.1 Monoclonal Antibodies mabs. Those of murine origin are designated by for Therapy the stem -omab; chimeric antibodies in which the variable region is spliced into a human constant Monoclonal antibodies (mAbs) for the treatment region are given the -ximab stem; humanized of diseases belong to a new class of therapeutic antibodies with the murine hypervariable regions agents called biologics. From their announce- spliced into a human antibody have the -zumab ment in the scientifi c literature in 1975 and for stem and antibodies with a complete human many years thereafter until more recent times, sequence are given the -mumab or -umab suffi x. these antibodies have been produced using mouse hybridoma cells prepared by fusing spleen cells from an immunized mouse with mouse myeloma 11.1.2 Monoclonal Antibodies cells. The hybridoma cells so produced retain the Approved for Therapy capacity to make specifi c antibody while the myeloma cells impart the capacity of the cells to Only 11 years after the publication of Köhler and grow indefi nitely in culture, continuously secret- Milstein’s paper on the development of hybrid- ing antibody. For therapeutic use in humans, the oma technology, the fi rst therapeutic monoclonal mouse antigens rapidly induce an immune antibody OKT3 (muromonab, Orthoclone) response which not only inhibits the mouse anti- received regulatory approval. Over the last 15 body’s action but can also provoke allergic reac- years, specially designed and produced mAbs tions including anaphylaxis. This has led have become one of the most important and suc- increasingly to the development of methods to cessful therapies for patients with hematological humanize mAbs. One approach involves the pro- malignancies and solid tumors. These targeted duction of chimeric antibodies by splicing the agents are also fi nding increasing application for variable regions encoding the antigen- recognition the treatment of other diseases such as chronic determinants from a mouse antibody into the asthma, psoriasis, systemic lupus erythematosus, constant and Fc regions of human IgG. While this and macular degeneration; the prevention of the is one step forward in attempts to eliminate or rejection of transplanted organs and graft-versus- attenuate an immune reaction, mouse antigens in host reactions; inhibition of platelet aggregation the variable regions can sometimes still stimulate in some cardiovascular diseases; treatment of an immune response. Another approach involves autoimmune diseases such as Crohn’s disease and the creation of a phage display library by the rheumatoid arthritis; for paroxymal nocturnal fusion of gene segments encoding human antigen- hemoglobinuria and cryopin-associated periodic binding variable regions to genes encoding bacte- syndrome; and for inhibiting respiratory syncytial riophage protein coat. Newer methods for the virus. Most are produced in Chinese hamster production of human mAbs utilize transformed ovary cells, Sp2/0 cells, or NSO cells. The sub- human B cells, plasmablasts secreting antibody, class of immunoglobulin used in therapeutic or by generating human B cell hybridomas. mAbs is an important consideration, especially in

[email protected] 11.1 Monoclonal Antibodies for Therapy 371 treating tumors. A glance at a list of mAbs shows like reactions, cytokine release syndrome, and that IgG1 is frequently selected, IgG2 and IgG4 tumor lysis syndrome (refer Sect. 13.5 ). The are occasionally employed, and IgG3 is rarely if range of clinical manifestations include those ever used. This relates to the different biological seen in local skin reactions at the injection site properties of the antibody subclasses—IgG1 is through to cutaneous and systemic hypersensi- the subclass of choice for antibody-dependent tivities and sometimes pyrexia, an infl uenza-like cell-mediated cytotoxicity which makes it emi- syndrome, and the potentially fatal systemic nently suitable for treating cancer cells while infl ammatory response syndrome. Acute anaphy- IgG4, which does not aid cytotoxicity, is the laxis and anaphylactoid reactions are well known choice when cell killing is not wanted. IgG3 is to occur with mAbs. Anaphylaxis to some mAbs seldom used since it has a signifi cantly decreased such as cetuximab and omalizumab has been fre- half-life. quently described and is of special interest (see At the time of writing, the ~28–30 mAbs below), but there are reports of reactions to others approved or under consideration for approval as including muromonab, basiliximab and OKT3. specifi c therapies in the USA or European Union Incidences of immediate hypersensitivity have are listed in Table 11.1. The great majority of been reported for infl iximab (2–3 %), rituximab these mAbs are for cancer immunotherapy. (5–10 %), trastuzumab (0.6–5 %), omalizumab Humanized and human monoclonal antibodies (0.1–0.2 %), and from <1 to 3 % for cetuximab with each comprise about one-third of the approved much higher rates reported in some regions of the total. Two of the antibodies are administered with USA (see Sect. 11.1.3.2 ). Serum sickness has radiolabels, ibrutumomab tiuxetan with been described for a number of mAbs including yttrium-90 or indium-111 and tositumomab with the chimeric antibodies infl iximab and rituximab 131 I; catamaxomab has dual specifi city, for and the humanized mAbs alemtuzumab and EpCAM (epithelial cell adhesion molecule) and natalizumab. For both the initial reactions and CD3; and only one, brentuximab vedotin, is an overall, cutaneous reactions are the most fre- antibody–drug conjugate. However, it is true to quently seen adverse responses to mAbs. say that the number of target antigens is surpris- Reactions following initial infusions of anti- ingly small with more than one of the approved body are common, but these can usually be han- antibodies specifi c for TNF, HER2, CD20, dled by a cautious rate of infusion, appropriate EGFR, or VEGF. It is said that about 350 new hydration and diuresis, and, if necessary, premed- mAbs for therapeutic application in humans are ication. Twenty six percent of initial reactions are in the commercial pipeline and it is likely that reported to be mild, 48 % moderate, and 26 % many of these will be antibody–drug conjugates, severe. The initial infusion reaction to some bispecifi c, and/or specifi cally engineered frag- mAbs, for example, rituximab (see below), may ments or domains. There may be at least some provoke tumor lysis syndrome, cytokine release truth in those who predict that the future of ther- syndrome, and systemic infl ammatory response apy belongs to the emerging biologics. syndrome. Tumor lysis syndrome, noted particularly with rituximab, can occur following cancer treatment and sometimes without treatment. It is 11.1.3 Immune Reactions believed to be the result of breakdown products to Monoclonal Antibodies of cancer cells leading to increased levels of some metabolites and refl ected in conditions Reactions, both immune and innate and to human such as hypercalcemia, hyperkalemia, hyper- as well as foreign proteins, may occur to mAbs. phosphatemia, acute uric acid nephropathy, and Acute reactions caused by a number of different acute renal failure. The syndrome can occur in mechanisms have been reported. These reactions the early stages of mAb therapy and is potentially include true, type I anaphylaxis, delayed reac- life-threatening. Cytokine release syndrome, also tions, anaphylactoid responses, serum-sickness- called cytokine storm, is commonly seen after

[email protected] 372 11 Biologics

® Rituxan

® ®

® ® ® ® ® ® ® ® Trade Trade name Malignant ascites Removab Non-Hodgkin lymphoma Zevalin

c /ADCC cking MS and Crohn’s Tysabri ammation RA Actemra b ALCL and Hodgkin lymphoma Adcetris I Non-Hodgkin lymphoma Bexxar d 131 Autoimmune disease (RA, Crohn’s) Humira Autoimmune disease (RA, Crohn’s) Remicade and RA Crohn’s Cimzia α α α

c and CD3 Reduces T cell activation T cell activation Reduces transplant rejection organ Prevent Simulect and CDC Blocks lymphocyte trafﬁ i cell division off Turns Colorectal and head neck cancers Erbitux Angiogenesis inhibitor -A Colorectal cancer Angiogenesis inhibitor Macular degeneration Avastin Lucentis of HER2 expression over Prevents Breast cancer Herceptin Mechanism of action Approved indication Inhibits virus entering cell RSV Synagis f f e a j k Inhibits immuno- stimulant BLys Systemic lupus erythematosus Benlysta l receptor (CD25) of C5 Inhibits cleavage nocturnal hemoglobinuria Paroxymal Soliris h -4 integrin -4 integrin chain IL-2 TNF Inhibits TNF- EpCAM/CD3 Binds both EpCAM α α VEGF

E. coli Hybrid hybridoma E. coli NSO C5 κ CHO CHO CD52 VEGF Eliminates lymphocytes CHO Chronic lymphocytic HER2 leukemia Campath-1H

NSO CHO Fab IgE Removes IgE Chronic asthma Xolair NSO CHO RSVF IL-6R Blocks IL-6-induced inﬂ κ κ κ κ κ κ κ κ κ CHO CD30 Antimitotic MMAE Sp2/0 TNF Inhibits TNF- Sp2/0 EGFR d

CHO CD20 B cell killing Non-Hodgkin lymphoma MabThera Sp2/0 Fab Fab Sp2/0 Glycoprotein IIb/IIIa inhibitor Platelet aggregation disease Cardiovascular Reopro Hybridoma CD20 Kills B cells with

κ κ κ κ κ κ g CHO TNF Inhibits TNF- CHO CD20 Binds malignant B cells NSO BLys λ c κ κ λ Fab, pegylated pegylated Fab, IgG2a bispecﬁ Type of mAb Type Cell line Target Chimeric IgG1 d Humanized IgG1 g I Murine IgG2a 131 Therapeutic monoclonal antibodies (mAbs) marketed or under review in the USA or European Union (As at June 2012) in the USA or European Union (As at June 2012) or under review Therapeutic monoclonal antibodies (mAbs) marketed iximab IgG1 Chimeric Cetuximab Chimeric IgG1 Rituximab -zumabs Alemtuzumab Bevacizumab Chimeric IgG1 Humanized IgG1 Humanized IgG1 Trastuzumab -(m)umabs Adalimumab Humanized IgG1 Human IgG1 -omabs Catumaxomab Rat IgG2b/Mouse Table Table 11.1 Generic name Tositumomab- Tositumomab- Inﬂ Natalizumab Humanized IgG4 Brentuximab vedotin Eculizumab Humanized IgG2/4 Ibritumomab tiuxetan Murine IgG1 Basiliximab Chimeric IgG1 Certolizumab pegol Omalizumab Ranibizumab Humanized IgG1 Humanized IgG1 Belimumab Human IgG1 -ximabs Abciximab Chimeric IgG1 Palivizumab Tocilizumab Humanized IgG1 Humanized IgG1

[email protected] 11.1 Monoclonal Antibodies for Therapy 373

® ®

® ® Xgeva ®

® ® 8-azaguanine, and are HAT sensitive. sensitive. 8-azaguanine, and are HAT Ilaris m Bone loss Prolia Metastatic melanoma Yervoy CAPS β

o Autoimmune disease Simponi α ammation by IL-12, IL-23 Psoriasis Stelara ammation by IL-1 with its ligands by RANK-L Antitoxin activity. In review In review Antitoxin activity. I nhibits dimerization of HER2 I with other HER receptors

Blocks interaction of CTLA-4 n cell division off Turns Colorectal cancer Vectibix

e k Blocks infl Blocks β B. anthracis Ag protective complement-dependent cytotoxicity complement-dependent cytotoxicity CDC CHO HER2 κ Sp2/0 TNF Inhibits TNF- CHO EGFR CHO CTLA-4 Sp2/0 IL-1 NSO CD20 B cell killing Chronic lymphocytic leukemia Arzerra CHO RANK-L NSO Inhibits activation of osteoclasts Sp2/0 IL-12, IL-23 Inhibits infl κ κ κ κ κ κ κ κ Chinese hamster ovary cells; Sp2/0, BALB/c mouse spleen cells fused with P3 myeloma. Cells do not secrete Ig, are resistant to cells; Sp2/0, BALB/c Chinese hamster ovary -chain synthesizing. 8-Azaguanine-resistant and HAT-sensitive mouse myeloma cell line -chain synthesizing. 8-Azaguanine-resistant and HAT-sensitive L CHO cytotoxic T-lymphocyte antigen 4; CD152 T-lymphocyte cytotoxic antibody-dependent cell-mediated cytotoxicity, antibody-dependent cell-mediated cytotoxicity, cryopyrin-associated periodic syndrome; Muckle–Wells syndrome periodic syndrome; Muckle–Wells cryopyrin-associated epidermal growth factor receptor factor epidermal growth city of mAb vascular endothelial growth factor (a subfamily of growth factors; includes VEGF-A) includes factors; of growth (a subfamily factor endothelial growth vascular human epidermal growth factor receptor. Also known as Neu, ErB-2, CD340, or p185 Also known receptor. factor human epidermal growth B lymphocyte stimulator; B-cell activating factor, BAFF BAFF factor, stimulator; B-cell activating B lymphocyte Human respiratory syncytial virus; F (viral protein coat antigen) Human respiratory syncytial rheumatoid arthritis. -4 integrin (CD49d) -4 integrin CAPS With Yttrium-90 or Indium-111 With agent monomethyl auristatin E (MMAE) Conjugated to the cytotoxic Attached to PEG (polyethylene glycol) Complement component 5 HER2 CTLA-4 Ligands for CTLA-4—CD80/CD86 name, Perjeta Trade June 2012 for metastatic breast cancer. by FDA Approved Specifi ADCC EGFR VEGF α RSV BLys NSO, non-Ig secreting, non-

RA Golimumab Human IgG1 Panitumumab Panitumumab Human IgG2 Ipilimumab Human IgG1 Canakinumab Human IgG1 a b c d e f g h i j k l m n o p

Ofatumumab Ofatumumab Human IgG1

Denosumab Human IgG2 Raxibacumab Human IgG1 Ustekinumab Human IgG1 Pending Pertuzumab Humanized IgG1

[email protected] 374 11 Biologics infusions of anti-immune cell mAbs (again, such sensitive to allergen stimulation. Secondly, anti- as rituximab). It is thought to be a consequence of gen trapping by IgE and subsequent presentation antibody binding to, and activation of, the cells by dendritic cells are markedly reduced or pre- producing a systemic infl ammatory response vented resulting in no further activation of aller- together with high fever. The reaction, which is gen-specifi c Th2 cells. similar in some respects to infection, can induce Despite the clear effi ciency of omalizumab in life-threatening pulmonary edema and possibly reducing levels of IgE antibodies, the question of death. Systemic infl ammatory response syn- its safety remains paramount. In one assessment drome affects the whole body and resembles the of the mAb’s safety and tolerability, data from response seen to sepsis. It may lead to respiratory completed clinical studies involving more than distress syndrome, renal failure, gastrointestinal 7,500 patients with asthma, rhinitis, and other bleeding, and dysfunction of the central nervous conditions were reviewed with a focus on hyper- system. sensitivity reactions, other immune effects, Although there are many reports of adverse thrombocytopenia, malignant neoplasia, and par- reactions, especially infusion reactions, to many asitic infections. Findings revealed that omali- of the mAbs in current use, most information is zumab had good safety and tolerability records available on fi ve widely and frequently used that were maintained for up to 4 years in one antibodies and they will therefore be considered study. The incidence of anaphylaxis to the agent in some detail. Reactions seen to these four was 0.14%, twice the fi gure seen in control mAbs, ranging from mild skin rashes to full- patients, but based on an estimated exposure of blown anaphylaxis and including infusion reac- 57,300 patients, the frequency of anaphylaxis tions, are similar to those seen with the other was estimated to be at least 0.2 %. Increased risks antibodies. of malignant neoplasia and thrombocytopenia were not detected. A review undertaken by the 11.1.3.1 Omalizumab U.S. Food and Drug Administration Adverse Omalizumab, a humanized IgG1κ mAb with Event Reporting System of cases of anaphylaxis specifi city for human IgE antibodies (Table 11.1 ), to omalizumab for the period June 2003 and is approved for the treatment of severe allergic December 2006 revealed 124 cases, many of asthma in patients 12 years or older. It binds to whom experienced delayed (>2 h) onset of the free, circulating IgE antibodies and membrane- reaction or protracted progression of the symp- bound IgE molecules on some cells such as B toms. A similar review for the period June 2003 lymphocytes expressing the antibody, but it does to December 2005 carried out by a joint task not bind to IgE already bound to mast cells, baso- force of the major U.S. allergy societies found 41 phils, and dendritic cells. This selectivity of bind- cases of anaphylaxis in 35 of 39,510 patients ing results from omalizumab binding to a given omalizumab, a rate of 0.104 %. These fi g- determinant in the Cε3 region of the free anti- ures show that in 2006, another 83 episodes of body, the same region involved in binding to the anaphylaxis were recorded and this fi nding, FcεRI receptor on the mast cell. Interference with together with the higher incidence of anaphylaxis binding due to steric hindrance also occurs when in the post-marketing period than in the pre- IgE is bound to the receptor, in this case prevent- marketing clinical trials, led the Omalizumab ing the binding of the mAb to the patient’s IgE Joint Task Force to issue guidelines for the molecules. Omalizumab has proven extremely administration of the agent. The Task Force effi cient in depleting free circulating IgE to recommended: (1) Prior to administering almost negligible levels with two interesting con- omalizumab, patients should be assessed for vital sequences. As IgE levels are reduced, the com- signs, asthma control, and lung function; (2) plementary receptors on mast cells, basophils, Informed consent should be obtained; (3) Patients and dendritic cells fall correspondingly and this should be advised how to recognize anaphylaxis, has the consequence of rendering the cells less how to use an epinephrine auto-injector and to

[email protected] 11.1 Monoclonal Antibodies for Therapy 375 ensure that the injector is always available before administration of cetuximab, but how they during the administration of omalizumab; (4) became sensitized to the disaccharide in the fi rst Omalizumab should only be administered in a place remains uncertain. A possible explanation facility that has both the staff and equipment to was suggested by the occurrence in patients given treat anaphylaxis; and (5) Patients should be cetuximab of delayed-onset anaphylaxis, angio- observed for 2 h after each of the fi rst three injec- edema, and/or urticaria 3–6 h after consuming tions and for 30 min after subsequent injections red meat. Following some investigations and of the mAb. questioning of patients in a large area of the U.S. South East, it has been speculated that IgE anti- 11.1.3.2 Cetuximab bodies to the α-linked D -galactose disaccharide Cetuximab, a chimeric mouse-human IgG1κ present in the meat may be linked to prior tick mAb to the epidermal growth factor receptor bites. Previous results by Van Nunen and col- (EGFR) used to treat colorectal cancer and squa- leagues in Sydney who reported an association mous cell cancer of the head and neck, has been between reactions to tick bites and allergy to red shown to be associated with anaphylaxis in a meats appear to support this suggestion. The most curious way. Natural antibodies, mostly of mechanism of this association is, as yet, unknown the IgG class, specifi c for an α-1,3-linked and is itself subject to speculation. An anti-cetux- D-galactose disaccharide, a structure found in imab IgE ELISA was recently reported with the many animals but not humans, are found in all claim that it could be a valuable test to identify individuals as a result, it is thought, of inactiva- potential cases of anaphylaxis following cetux- tion of the gene for the enzyme α-1,3- imab infusion. Cetuximab-reactive IgE antibod- galactosyltransferase. Presumably, this resulted ies were detected in 24 of 92 (26.1 %) of in the loss of immune tolerance to the α-D - pretreatment patients and 33 of 117 (28.2 %) galactose determinant and the production of anti- healthy blood donors. Hypersensitivity reactions bodies to it. Although the biological role of this occurred in 14 of the 92 patients (15.2 %) and 8 antibody remains unclear, it may provide some of these were grade 3–4 reactions. Seven of the protection against gastrointestinal bacteria and eight patients (87.5 %) with severe hypersensitiv- contribute to the removal of senescent red cells ity reactions had anti-cetuximab IgE antibodies via recognition of cryptic α- D - galactosyl residues while 14 of 78 (17.9 %) with no signs of hyper- exposed in the course of cell aging. Humans of sensitivity showed the presence of antibodies. blood group B have a terminal α-1,3-linked- d - With the greatly expanding administration of galactose on their blood group substances in biologic agents, it is already clear that infusion secretions and on red cells, but the presence of a reactions occur frequently, and because many penultimate α-1,2-linked L -fucose prevents bind- patients being treated with mAbs often have few ing to the antibody. By the early 1990s it was or even no other therapeutic alternatives, success- known that the anti-D -galactosyl antibodies in ful desensitization to mAbs is likely to be increas- humans were potentially capable of interacting ingly sought. Claim of a successful desensitization with therapeutic recombinant proteins expressing protocol to cetuximab has been published. The the complementary α-linked determinant and this patient was premedicated with prednisolone 12 h became a reality just a few years ago when and 1 h before desensitization and with diphen- patients receiving cetuximab experienced severe hydramine 30 min before desensitization. immediate hypersensitivity reactions. The anti- Beginning with an infusion dose of 0.001 mg of bodies involved were found to be IgE, specifi c for cetuximab, doses were doubled every 15 min α-D - galactose-(1–3)-β-d -galactose and reactive with each dose tolerated until a total of 64 mg of with this disaccharide present on the Fab portion cetuximab was reached. The appearance of a pru- of the chimeric antibody at asparagine 88 of the ritic cutaneous reaction was managed with heavy chains. Most of the patients who reacted diphenhydramine, a 30 min waiting period, and already had the IgE antibodies in their serum dose and infusion rate reductions. The fi nal dose

[email protected] 376 11 Biologics

leukocytoclastic vasculitis (Fig. 11.1 ), serum sickness, and anaphylaxis. In a large center study of 165 consecutive patients who received 479 inf- liximab infusions, the overall incidence of infusion reactions to the mAb was 6.1 % with 9.7 % of the patients affected. Mild, moderate, and severe reactions occurred in 3.1, 1.2, and 1 % of infl iximab infusions, respectively. Serum tryptase levels suggested that acute reactions in 11 of 14 patients were not type I hypersensitivity reactions. Delayed reactions were rare, resulting from only 0.6 % of infusions. An examination of incidences of systemic and delayed reactions to infl iximab in children as well as adults with Crohn’s disease revealed that 14 % of 86 patients experienced severe systemic reactions from a total of 304 infusions. A signifi cant difference between the results for adults and children was noted—severe systemic reactions were seen in 11 of 52 adults (21.2 %) and in only one of 34 (2.9 %) children. Fig. 11.1 Palpable purpura in a patient with hypersensi- These reactions were characterized by hypoten- tivity vasculitis (leukocytoclastic vasculitis), a small vessel vasculitis usually involving post-capillary venules in sion, mucosal irritability, and laryngospasm the dermis. This cutaneous manifestation of vasculitis requiring epinephrine, antihistamines, and/or cor- occurs occasionally following treatment with mAbs and ticosteroids. Delayed reactions, which manifested some other biologic agents (Photograph of Dr. John as arthralgia, fever, and myalgia requiring cortico- Stone) (Reproduced with permission from Weyand CM, Goronzy J, in Klippel JH, Stone JH, Crofford LeJ, White steroids were seen in eight adults (9.3 %); no PH, editors. Primer on the Rheumatic Diseases, 13th ed. delayed reactions occurred in children. An exami- New York: Springer, 2008) nation of the extent of, and reasons for, discontinuation of infl iximab treatment in 84 patients of cetuximab (325 mg) was tolerated giving a with established rheumatoid arthritis revealed that cumulative dose of 844 mg. When challenged 28 (33 %) discontinued the therapy. The main rea- with cetuximab 1 week later, the patient tolerated son for discontinuation was an adverse reaction in the dose without diffi culty. This was expected 16 of the 84 patients (19 %). In this group, 9 of the since the half-life of cetuximab is approximately 16 patients (10.7 %) experienced an immediate 4 days. It was predicted that the protocol might hypersensitivity reaction. There are many reports be useful to many patients. of immediate reactions, in particular anaphylaxis, to infl iximab. From FDA and other sources, over 11.1.3.3 Infl iximab 650 anaphylactic reactions to the mAb at an inci- Like cetuximab, infl iximab is a mouse-human dence of just under 0.9 % occurred in the period chimeric IgG1κ mAb. With specifi city for TNF, 1999–2012. Most reactions (nearly 70 %) occur the antibody is used to treat autoimmune diseases during the fi rst month of therapy; females account such as Crohn’s disease and rheumatoid arthritis for approximately 60 % of reactors and children (Table 11.1 ) and there are a few reports of it under 10 years of age for 7.5 % of reactors. inducing rapid recovery of lesions in several cases Cutaneous reactions may also be provoked by of toxic epidermal necrolysis (Sect. 3.6.3.7 ). A infl iximab. Three patients with rheumatoid arthri- variety of reactions, both systemic and cutaneous, tis, none of whom had a personal or family his- have been reported following administration of tory of psoriasis, developed what was described infl iximab. These include maculopapular rashes, as psoriasiform skin lesions 6–9 months after the urticaria, psoriasis, fl are-up of atopic dermatitis, initiation of infl iximab therapy. Two of the

[email protected] 11.1 Monoclonal Antibodies for Therapy 377 patients developed palmoplantar pustular lesions umab has been reported to elicit psoriasiform and scaly plaques on the extremities, while the plaques on elbows, arms, and thighs together third patient had erythematous plaques with sil- with palmoplantar pustular lesions. More severe very white scales on the scalp. Other rare adverse cutaneous reactions to the mAb also occur such reactions to infl iximab that may have an immu- as erythema multiforme-like skin reaction with nological basis include demyelinating polyneu- papulopustular exanthema at the injection site ropathies, peripheral neuropathy, drug-induced and on the palms and soles followed by skin des- lupus, and hepatitis. A combination of the latter quamation and at least two cases of Stevens– two conditions has been reported with the use of Johnson syndrome. Systemic reactions to infl iximab for psoriasis. adalimumab have been reported. In one case, a Successful desensitizations have been reported patient with spondylarthritis treated with the in adult and child patients who experienced an mAb experienced two reactions consisting of anaphylactic/anaphylactoid reaction to infl ix- generalized itching, angioedema of the lips, diz- imab. Based on the adult patient’s weight of ziness, and visual disturbances. Skin prick and 70 kg and the standard dose of 5 mg/kg, a total intradermal tests with adalimumab produced infl iximab infusion of 353 mg divided into 11 strong immediate positive reactions, but serum increments was given IV every 15 min over a 4 h IgE antibodies were not detected using a spe- period with a starting dose of 3 μg and a fi nal cially prepared adalimumab Phadia solid phase. dose of 160 mg. For a 10-year-old male child, the schedule was the same except for the lower dos- 11.1.3.5 Rituximab ages (total of 208 mg), increments ranging from Rituximab, like cetuximab and infl iximab, is a 2 μg to 80 mg and a fi nal dose of 80 mg. For both -ximab chimeric IgG1κ mAb (Table 11.1). It con- patients, IV infl iximab was tolerated without tains murine light and heavy chain variable incident and each experienced clinical improve- region sequences and human constant region ment over the subsequent 2 months of treatment. sequences. Administered for non-Hodgkin lymphoma, rituximab is targeted to CD20 (human B 11.1.3.4 Adalimumab lymphocyte-restricted differentiation antigen Like infl iximab, adalimumab is a mAb targeted Bp35), a B lymphocyte antigen involved in the at TNF and used for Crohn’s disease and rheuma- development and differentiation of B cells into toid arthritis, but, unlike infl iximab, it is a fully plasma cells. CD 20 is found on B cell lympho- human mAb of the IgG1κ class and can be mas, B cell chronic lymphocytic leukemia, hairy administered subcutaneously. Being a possible cell leukemia, and melanoma cancer stem cells substitute for infl iximab in patients intolerant to and is expressed on more than 90 % of B cell that mAb, in 2004 adalimumab was examined for non-Hodgkin lymphomas but not on normal safety and effi cacy in seven patients who had plasma cells, hematopoietic stem cells, or other experienced immediate or delayed hypersensitiv- normal tissues. In 1997 rituximab was the fi rst ity reactions to infl iximab and one patient with mAb approved specifi cally for cancer therapy. In infl iximab-induced lupus. Adalimumab proved the early years following its release, a relation- to be well tolerated leading to the conclusion that ship between cytokine release syndrome in it might prove to be a safe and effective substitute patients and high lymphocyte counts was for patients allergic or otherwise intolerant to inf- observed after treatment with the mAb. Patients liximab, In subsequent years, a range of different with lymphocyte counts greater than 50 × 10 9 /L apparent hypersensitivity reactions induced by experienced a severe cytokine release syndrome adalimumab have become apparent. These shown by peaks in release of TNF and IL-6 90 include psoriasis, exacerbation of palmoplantaris min after infusion and accompanied by fever, pustulosa psoriasis, asthma, bronchospasm, auto- chills, nausea, vomiting, hypotension, dyspnea, immune hepatitis, and a number of different skin an increase in liver enzymes, and prolongation of reactions, some severe. Like infl iximab, adalim- the prothrombin time. When used to treat B cell

[email protected] 378 11 Biologics cancers, the frequency and severity of fi rst-dose distress syndrome. Respiratory events such as reactions to rituximab were shown to be depen- cough, dyspnea, and bronchospasm are fairly dent on the initial number of circulating tumor common adverse reactions, but serious reactions cells—patients with counts exceeding 50 × 109 /L such as fatal pulmonary fi brosis also occur. A experienced more adverse reactions than patients review of 62 cases of rituximab-induced severe with lesser numbers of peripheral tumor cells. respiratory reactions revealed that 74 % suffered Recently published results of a survey of ritux- from interstitial pneumonitis and other respira- imab hypersensitivity reactions in patients at tory problems including pulmonary fi brosis, Massachusetts General Hospital between 2006 bronchiolitis obliterans, organizing pneumonia, and 2010 showed that immediate hypersensitivity hypersensitivity pneumonia, and acute respira- reactions to rituximab occurred in 8.8 % (79 of tory distress syndrome. Most patients were 901) of patients treated with the mAb and pre- elderly but two pediatric patients, both with medications. Approximately three-quarters of the refractory nephrotic syndrome, were affected. patients developed symptoms after the fi rst infu- Such fi ndings have led to the suggestion that sion and 46 % of moderate or severe reactions rituximab should never be administered to occurred on subsequent infusions. Severity of patients suffering from lung diseases such as reactions correlated with the risk of a recurrent pneumonia, pleural effusion, and atelectasis. adverse response—all patients with severe, and Other mAbs including infl iximab, gemtuzumab, 56 % of those with moderate reactions, but only OKT3, and a mAb-ozogamacin conjugate have 36 % of patients with mild reactions, experienced also been implicated in cases of acute respiratory a recurrence. Interestingly, there was an increased distress syndrome which is believed to be medi- risk of a moderate or severe hypersensitivity reac- ated by pro-infl ammatory cytokines. Cutaneous tion in those with advanced disease. Waldenstrom’s side effects of rituximab are frequent, generally macroglobulinemia accounted for 10 % of all occur from 1 to 13 weeks after exposure, and are reactions while representing only 1 % of patients not usually serious but Stevens–Johnson syn- treated with rituximab. In another retrospective drome, toxic epidermal necrolysis, lichenoid der- analysis of reactions occurring over a 2-year matitis, and vesiculobullous dermatitis have been period following infusion of rituximab in the described. treatment of multiple sclerosis, 25.7 % of patients had mostly mild to moderate reactions, most often 11.1.3.6 The Next Generation during the fi rst infusion. Most patients completed of Monoclonal Antibodies the infusion and went on to subsequent infusions Until now, FDA-approved mAbs (Table 11.1 ) are without reactions. In common with a number of usually full-length antibodies, making them large other surveys on rituximab use, it was concluded molecular weight (~150 kDa) therapeutic agents that infusion-related reactions to the therapeutic with sometimes poor tissue penetration, espe- agent in patients with multiple sclerosis are com- cially for solid tumors. As understanding of the mon, premedication with drugs including cortico- biological mechanisms in different diseases steroids dramatically reduces the incidence of increases and genetic engineering technology reactions, and reactions that do occur can be advances, attention is turning to improving the effectively managed by treatment with H 1 and H 2 performance and effi ciency of mAbs in terms of antihistamines and infusion rate adjustments. increased selectivity, improved pharmacokinet- As well as infusion-related reactions and reac- ics, higher binding affi nities, more effi cient tions related to the number of circulating target cytotoxicity, better tissue penetration, and cells, a number of post-infusion hypersensitivity increased half-life in serum. More than 50 % of or hypersensitivity-like reactions occur to ritux- mAbs in phase I clinical trials and about 40 % of imab. These reactions include serum sickness, those in phases II and III are now modifi ed anti- vasculitis, various cutaneous manifestations, bodies such as antibody–drug conjugates, bispe- interstitial pneumonitis, and acute respiratory cifi c antibodies, antibodies with modifi ed Fc

[email protected] 11.2 Etanercept 379 functions, and antibody fragments/domains. What effects these developments will have on Employment of two mAbs may sometimes pro- the incidences and nature of adverse, and in par- duce a better therapeutic outcome than one anti- ticular allergic, reactions to therapeutic mAbs body alone, for example, the combination of remains to be seen, but it is certain that the num- cetuximab with specifi city for EGFR and bevaci- ber and variety of genetically engineered mAbs zumab which recognizes VEGF, in the treatment and modifi ed mAbs will greatly increase in the of metastatic colorectal cancer. A more logical immediate future. We might expect that many and effi cient alternative to the use of more than reactions already encountered with the existing one mAb is the creation of an antibody with spec- approved antibodies will still be seen, but we ifi cities for two different targets, that is, bispe- should not be surprised if some new, unantici- cifi c antibodies. By simultaneously binding to pated adverse reactions emerge as newer genera- two different complementary sites on a cell, tion agents showing some physical, chemical, bispecifi c antibodies may enhance binding selec- and biological differences become established tivity, avidity, and tissue distribution, expand the and increasingly used. However, as summarized mAb’s disease indications, and increase antibody above, the carefully planned strategies already load on target cells. Nonspecifi c toxicity of che- being applied to modify and thereby improve the motherapy is a major limitation of much of performance and tolerability of next generation today’s drug therapy for cancers (see chapter 13). mAbs (e.g., smaller size and intrabodies) may Antibodies conjugated to carefully selected drugs also carry with them a decreased capacity to pro- or toxins delivered to specifi c tumor sites have voke adverse responses in patients. In any case, the potential to reduce systemic toxicity and there the continuing elucidation of cellular pathways in is little doubt that such mAb drug conjugates will an expanding range of diseases coupled with a be increasingly developed and applied in future deeper understanding of the myriad immunologi- mAb immunotherapies. One recent imaginative cal and infl ammatory processes and the advanc- approach in applying antibody–drug conjugates ing bioengineering expertise should give hope for cancer therapy is based on the occurrence of that such intellectual and technical ingenuity angiogenesis in virtually all types of aggressive could also be applied to minimizing the detri- cancers but the rarity of the process in healthy mental effects of these precisely engineered ther- adults. Early experiments have demonstrated that apeutic agents. Efforts expended to make mAbs strong antitumor activity in vivo can be achieved more immunologically acceptable to humans with vascular targeting of an antibody–drug con- have been impressive; equal efforts directed at jugate that does not require antibody internaliza- minimizing adverse reactions might prove just as tion. Fc engineered mAbs can be utilized for successful. improved antibody-dependent cellular and complement-dependent cytotoxicities via their Fc regions and antibody fragments and single 11.2 Etanercept domain antibodies offer the opportunity to retain or enhance the binding properties of full-length Like the mAbs infl iximab, certolizumab, adalim- antibodies by varying their size, valency, and umab, and golimumab, etanercept is specifi c for, pharmacokinetic profi les. Another potential and binds to, TNF, thus making it useful for the improvement offered by antibody fragments is treatment of some autoimmune diseases including improved tissue penetration although this may be rheumatoid arthritis, ankylosing spondylitis, pso- offset by a shorter serum half-life. Approaches riatic arthritis, plaque psoriasis, and Crohn’s dis- already employed to overcome this include the ease. TNF receptors are of two types, those on chemical addition of polyethylene glycol (PEG) nucleated cells and soluble receptors that circulate or so-called PEGylation to increase the size of and deactivate the cytokine. Etanercept is a recom- fragments. This strategy was applied to certoli- binant, engineered, fully human dimeric fusion zumab, an anti-TNF Fab fragment. protein of molecular weight 150 kDa, made up of

[email protected] 380 11 Biologics the extracellular ligand-binding portion of human IFNs, pegylated IFNα-2a and pegylated IFNα-2b, 75 kDa TNF receptor (TNFR) linked to an Fc por- have found application as antiviral agents in the tion of human IgG1. The Fc portion contains the treatment of hepatitis C virus. IFNα, now usually CH2 and CH3 domains and the hinge region but given in pegylated form, in combination with not the CH1 domain. In this form, the protein acts ribavirin (1-β- D - ribofuranosyl -1H -1,2,4,-triazole- as a “decoy” receptor for TNF, mimicking the 3-carboxyamide) is the mainstay of treatment of natural soluble receptor and improving on it by hepatitis C infection. Pegylation enhances the possessing a longer half-life (115 h; compare inf- half-life of the IFN by covalent attachment of liximab half-life 210 h). Both infl iximab and etan- polyethylene glycol (PEG) polymer chains to the ercept are sometimes used for patients with molecule, reducing both antigenicity and immu- rheumatoid arthritis when other disease-modify- nogenicity. Adverse reactions to IFNα include ing antirheumatic drugs have failed. The most fl u-like symptoms, localized infl ammation, and a common side effects reported for etanercept wide range of cutaneous reactions including mac- include mild reactions at the injection site, infec- ulopapular rash, pruritus, urticaria, angioedema, tions (mainly upper respiratory), and sinusitis. pemphigus, vasculitis (and systemic), fi xed drug U.S. FDA data on etanercept adverse events lists, eruption, and lichen planus. Autoimmune disor- in order of frequency, infections, followed by der- ders thrombocytopenia and hemolytic anema also matologic, neurologic, musculoskeletal, pulmo- occur. Cytokine release syndrome probably nary, cardiac, and vascular effects. Both etanercept accounts for many reactions with fl u-like symp- and infl iximab have been associated with cutane- toms. Recombinant IFNβ has been implicated in ous vasculitis in which symptoms coincided with anaphylactic shock, indicating that IgE-mediated their administration and symptom resolution coin- reactions are probably involved in a few immedi- cided with treatment withdrawal. Lesions associ- ate reactions. Of 51 patients given pegylated ated with etanercept-induced autoimmune skin IFNα-2b-ribavirin, 10 were found to have experi- reactions include purpuric papules and erythema- enced serious adverse drug reactions after 1 tous papules and nodules on the trunk and extrem- month and adverse reactions were the main rea- ities. In one reported case of cutaneous vasculitis son for discontinuing the therapy in eight patients associated with both etanercept and infl iximab, (15.7 %). The most common reaction to this ther- vasculitis provoked by etanercept in a patient with apeutic agent is localized skin lesions at the injec- Crohn’s disease was found to worsen signifi cantly tion site, but vesicle erythematous eruptions and after therapy was switched to infl iximab. autosensitization dermatitis away from the injection site have been described.

11.3 Interferons 11.4 Interleukin 2 Interferons (IFNs) are an important class of broad spectrum antiviral cytokines found in higher ani- The cytokine interleukin 2 (IL-2) is necessary for mals, reptiles, ﬁ sh, and birds. IFNs make up a T cell growth, proliferation, and differentiation. large, still growing list of proteins, seven of which An FDA-approved recombinant human IL-2 is occur in humans. These are further divided into used clinically as Proleukin® (Prometheus three classes types I, II, and III. Two important Laboratories) for the treatment of metastatic type I IFNs in human are IFNα and IFNγ. There melanoma and renal cell carcinoma. Erythema are at least 14 different alpha IFNs. These are pro- during IL-2 immunotherapy is common and was duced by leukocytes and dendritic cells and are well described in a French study nearly 20 years part of an innate immune response with antiviral ago with a report on generalized erythema fol- action. IFNβ, produced by ﬁ broblasts and proba- lowed by desquamation in 12 patients treated with bly many other cells, is used in the treatment of the cytokine for renal cancer. Urticaria in eight multiple sclerosis. It inhibits viral replication and renal cell cancer patients after the end of IL-2 ther- is the product of a single gene. Two synthetic apy has also been reported. Skin tests with IL-2 on

[email protected] 11.6 Anakinra 381 two of the patients proved negative. Cutaneous of acute infusion reactions decreased signifi cantly side effects in metastatic melanoma patients with only three patients experiencing reactions treated with IL-2 were observed in 53 of 78 treat- and only two patients developing vascular leak- ment cycles of 25 patients (72 %); 53 were mild age. Overall, a signifi cant improvement of 60 % reactions with a burning pruriginous erythema, occurred when premedication with steroids was and three were severe with urticaria, necrotic employed. Cutaneous reactions to denileukin lesions, and blisters. Regression proved constant diftitox include injection site reaction, erythema, without sequelae. Other observed cutaneous reac- and pruritus, and there has been one fatal case of tions to IL-2 include injection site reactions, exfo- toxic epidermal necrolysis. liative dermatitis, angioedema, reactivation of Another recombinant fusion protein, afl iber- eczema, vasculitis, pemphigus, and exacerbation cept (Zaltrap®), used to treat colorectal cancer and of psoriasis. There appear to be no reports of ana- macular degeneration, is a chimera of the Fc piece phylaxis following IL-2 immunotherapy. of IgG1 and the extracellular ligand-binding domains of human vascular endothelial growth factor receptors VEGFR1 and VEGFR2. Adverse/ 11.5 Denileukin Diftitox hypersensitivity reactions to the drug include and Afl ibercept cytopenias, hemorrhage, thromboembolism, GI perforation, acral erythema and stomatitis. Diphtheria toxin is a single polypeptide chain of 535 amino acids. For use as a targeted toxin, it has been modifi ed by deleting the 147 amino acid 11.6 Anakinra residue receptor-(cell-) binding domain to produce a protein of 388 amino acids commonly Interleukin-1 (IL-1) is a cytokine produced in referred to as DT388 or DAB389 . This remaining response to infl ammatory stimuli in conditions protein consists of the adenosine diphosphate such as rhinitis, rheumatoid arthritis, and other (ADP)-ribosyltransferase and membrane translo- immunological reactions. The IL-1 receptor (IL- cating domains of native diphtheria toxin. 1R) exists in both membrane-bound and soluble Replacing the receptor-binding domain of the forms and is expressed on many organs and tissues. native toxin by the sequences encoding the IL-2 Anakinra, a specifi c receptor antagonist for IL-1, is gene produced the recombinant fusion toxin des- a 153 amino acid non-glycosylated, molecular ignated DAB389 IL-2 or denileukin diftitox weight 17.258 kDa recombinant protein prepared (Ontak® , Eisai) which was approved by the FDA in Escherichia coli . It differs from natural IL-1R in 1999 for the treatment of cutaneous T cell lym- by the addition of a single methionine added to the phoma. Bound to the IL-2 receptor, the fusion amino terminal end. The recombinant receptor toxin undergoes endocytosis and is proteolytically antagonist competes with IL-1, blocking its access cleaved liberating the modifi ed toxin and causing to its complementary receptor, thus making it a ADP-ribosyltransferase-mediated inhibition of useful agent in the treatment of some infl ammatory protein synthesis. In phase I and III studies on conditions such as rheumatoid arthritis where it

DAB 389IL-2, infusion-related acute hypersensitiv- acts as a biological response modiﬁ er rather than a ity reactions occurred in 70 % of patients and vas- disease-modifying antirheumatic drug. The protein cular leakage occurred in 27 % forcing 29 % of has a half-life of 4–6 h and peak plasma concentra- patients to discontinue therapy. There were no tions occur 3–7 h after subcutaneous injection. In correlations between these reactions and the dose an examination of the safety proﬁ le of anakinra, or the half-life of the fusion toxin. Since only the over 1,300 patients were initially studied in a dou- antihistamine diphenhydramine and acetamino- ble blind trial comparing the drug (100 mg/day) phen were used for premedication in these stud- with placebo before proceeding to open-label ies, the steroids prednisone and dexamethasone treatment for up to 3 years. All adverse events were were examined to see if they improved tolerability similar in the anakinra and placebo groups and for of DAB 389IL-2. Results showed that the incidence each of the years of anakinra treatment. Injection

[email protected] 382 11 Biologics site reactions were the most frequent adverse event therapy to delay rejection. Two preparations are (122 events/100 patient years) and, overall, it was available, Thymoglobulin® (Genzyme), obtained concluded that anakinra is safe and well tolerated by immunizing rabbits, and Atgam ® (Pfi zer), from for up to 3 years of continuous use. Cutaneous hyperimmune horse serum. These two preparations reactions are usually at the injection site and usu- are contraindicated in patients with a history of ally well tolerated. Skin biopsy specimens from allergy and anaphylaxis to rabbits or horses, respec- rheumatoid arthritis patients treated with anakinra tively. Serious immune- mediated reactions have and with well-defi ned erythema and edema at the been reported, including anaphylaxis, severe cyto- injection sites showed marked dermal edema, an kine release syndrome, and severe acute infusion- increased number of mast cells, and a lichenoid associated reactions. The last named may occur as infi ltrate of mainly lymphocytes together with early as the fi rst or second infusion and serum sick- eosinophils and CD68 macrophages. In some ness with fever, rash, arthralgia, and myalgia may cases, cutaneous reactions were associated with appear 5–15 days after the initiation of therapy. As systemic involvement. The observed skin reactions a precaution against the possibility of anaphylaxis, were said to resemble reactions seen in patients it is recommended that before the fi rst infusion, receiving chemotherapy and colony-stimulating patients should be tested intradermally with the factors. A cutaneous reaction in one patient was diluted globulin preparation (for example, with shown to be mediated by specifi c IgE antibodies. ~5 ug of horse globulins). The most frequently After several injections of anakinra, an adult seen adverse reactions, that is, seen in more than female patient began to experience erythema and 25 % of patients, include fever, chills, headache, pruritus at the injection site within 15 min after nausea, diarrhea, abdominal pain, peripheral each injection. No systemic symptoms occurred, edema, hypertension, thrombocytopenia, infec- but 1 day later swelling, erythema, and pruritus tion, dyspnea, and tachycardia. Cutaneous reac- were apparent and these persisted for a further day tions seen include urticaria, morbilliform or two. Prick and intradermal tests together with an eruptions, and acral erythematous eruptions pre- ELISA for specifi c IgE antibodies to anakinra were ceding rash. Discontinuation of treatment with undertaken. Prick tests with concentrations of 500 anti- thymocyte globulin is recommended upon the and 2,500 μg/ml were negative, but intradermal appearance of systemic reactions such as anaphy- testing with 125 μg proved positive. The reaction laxis, generalized rash, tachycardia, dyspnea, was still positive after 48 h. Five normal control hypotension, severe thrombocytopenia, or severe subjects had no reaction to the same dose of the leukopenia. drug. The specifi c IgE test was also positive and this was confi rmed by inhibition experiments with free anakinra. 11.8 Epoetins

Human erythropoietin (or erthropoetin) (EPO), 11.7 Anti-thymocyte Globulin also called hematopoietin (or hemopoietin), is a glycoprotein hormone of 165 amino acids MW Indicated and approved for the management of 34 kDa that controls erythropoiesis (red blood cell allograft rejection in renal transplant patients, anti- production). Recombinant human EPO, introduced thymocyte globulin preparations are puriﬁ ed in 1986, is available as epoetins alfa, beta, delta, immune globulins (primarily IgG) from horses or and omega each differing from the endogenous rabbits immunized with human thymus lympho- hormone, and from each other, by the individual cytes. The resultant globulin preparations contain sugar and sialic acid residues present. Epoetins are cytotoxic antibodies to human T lymphocytes administered for renal and non-renal anemias and, which function as an immunosuppressive agent. As despite being given to a large number of patients well as its use for the treatment of renal transplant over a period of 25 years, the preparations have rejection, anti-thymocyte globulin may be adminis- proved poorly immunogenic and few side effects tered as an adjunct to other immunosuppressive have been observed. In 2001 the FDA and European

[email protected] 11.9 Human Insulin 383

Medicines Agency approved a new epoetin, darbe- of human recombinant insulin reduced the inci- poetin alfa, for treatment of anemia due to renal dence of reactions, but allergic reactions are still failure and in patients undergoing immunotherapy. occasionally seen and insulin allergy is now Darbepoetin alfa is a recombinant epoetin mole- reported to be less than 1 % of diabetic patients cule containing an extra two N-linked oligosaccha- treated with insulin. As early as 1982, IgE antibod- ride chains introduced to give greater stability and ies to human recombinant insulin that cross-reacted thus allow less frequent administrations. In recent with bovine and porcine insulins were demon- years, there appears to have been an increase in the strated in two diabetic patients previously untreated number of patients developing neutralizing anti- with insulin. Although these patients did not EPO antibodies during therapy and there are now develop any of the clinical manifestations of insu- in excess of 250 known cases of pure red cell apla- lin allergy, large local reactions in association sia. There are also reports of anaphylaxis, rashes, with IgE to human insulin were later seen in a urticaria, and angioedema to the recombinant hor- patient within 12 days of insulin initiation therapy mone and injection site reactions are well known. with the human recombinant product. The patient Cutaneous reactions at the sites of former subcuta- showed similar cutaneous reactivity with bovine neous injections of epoetins following the IV injec- and porcine insulins despite never having received tion of different epoetins proved to be the signs of those preparations, and the insulin-reactive IgE an allergic skin and systemic reaction in a patient antibodies cross- reacted in vitro with the heterolo- with pure red cell aplasia and anti-EPO antibodies. gous proteins. This strongly indicated that com- After switching the patient from epoetin alfa to fi rst mon, or markedly similar, antigenic determinants epoetin beta and then darbepoetin alfa, a systemic are present on the insulins from all three species. anaphylactic/anaphylactoid response occurred and Subsequent experience has confi rmed these fi nd- anti-EPO antibodies cross-reactive with epoetin ings. Skin testing as well as IgE antibody measure- beta and darbepoetin alfa were detected in the ments have clearly demonstrated immediate type patient’s serum. This case illustrates that continua- I reactions to recombinant human insulin and to tion of epoetin therapy in patients with anti-EPO bovine and porcine insulins in patients never previ- antibodies may carry the risk of a serious systemic ously given the heterologous insulins. In the inves- (anaphylactic or anaphylactoid) reaction and that tigation of an anaphylactic reaction to human skin reactions at the injection site may be the fi rst recombinant insulin, employment of skin testing sign of sensitization which precedes the develop- and the Novo Insulin Allergy Kit (Novo Nordisk ment of anemia. In three other rare cases, anaphy- A/S, Bagsvaerd, Denmark) showed positive intra- laxis and serum IgE antibodies, and a generalized cutaneous tests to 1–100 dilutions of human and eczematous reaction to recombinant EPO, were porcine insulins and to the genetically engineered reported in the presence of negative skin tests to the recombinant insulin analog, insulin lispro. Once glycoprotein and acute exanthematous pustulosis again, the patient had never been treated with por- was diagnosed after epoetin alfa was replaced by cine insulin in the past. A number of different darbepoetin alfa. In an unexpected EPO allergy models have been applied in attempts to desensitize “false-alarm,” the nonionic surface active agent patients allergic to recombinant human insulin. polysorbate 80, and not Eprex ® (Janssen) contain- In one successful attempt, insulin lispro was deliv- ing epoetin alfa, was shown to be responsible for a ered as a continuous infusion via an insulin pump. case involving generalized pruritus, erythema, and The delivery and dosage schedule was: 0.7 IU/h for angioedema. 2–8 h; 0.3 IU/h for 8–13 h; 0.6 IU/h for 13–18 h; 0.8 IU/h for 18–21 h; and 0.6 IU/h for 21–22 h, plus an additional bolus of 6 IU before breakfast, 5 IU 11.9 Human Insulin before lunch, and 6 IU before dinner. Following this procedure, the allergic reaction did not reoccur. Adverse reactions to insulin were not uncommon in Although the patient remained clinically asymp- the past when the administered preparations were tomatic, the skin prick test to insulin remained from bovine and porcine sources. The introduction positive 3 months later.

[email protected] 384 11 Biologics

• Some patients receiving cetuximab experi- Summary enced severe immediate hypersensitivity reactions. The antibodies involved were found to • MAbs were fi rst produced by mouse hybridoma be IgE, specifi c for α-D - galactose-(1–3)-β-d - cells prepared by fusing spleen cells from an galactose and reactive with this disaccharide immunized mouse with mouse myeloma cells. present on the Fab portion of the chimeric The hybridoma cells retain the capacity to make antibody at asparagine 88 of the heavy chains. specifi c antibody while the myeloma cells impart Some cases of anaphylaxis to the mAb appear the capacity of the cells to grow indefi nitely in to be associated with tick bites and consump- culture, continuously secreting antibody. tion of red meat. • Because mouse antigens rapidly induce an • Systemic and cutaneous reactions have been immune response in humans, methods have reported following administration of infl iximab. been developed to humanize mAbs. One These include anaphylaxis, serum sickness, approach involves the production of chimeric maculopapular rashes, urticaria, psoriasis, fl are- antibodies. Other methods now produce fully up of atopic dermatitis, and leukocytoclastic humanized antibodies. vasculitis. The overall incidence of infusion • MAbs of murine origin are designated by the reactions in one study was 6.1 %. Mild, moder- stem -omab; chimeric antibodies in which the ate, and severe reactions occurred in 3.1, 1.2, variable region is spliced into a human constant and 1 % of infl iximab infusions, respectively. region are given the -ximab stem; humanized • Patients with lymphocyte counts greater than antibodies with the murine hypervariable regions 50 × 10 9 /L experienced a severe cytokine release spliced into a human antibody have the -zumab syndrome shown by peaks in release of TNF stem and antibodies with a complete human and IL-6 90 min after infusion with rituximab. sequence are given the -mumab or -umab suffi x. • A number of post-infusion hypersensitivity or • Currently, ~28–30 mAbs are approved, or under hypersensitivity-like reactions occur to ritux- consideration for approval, as specifi c therapies imab. These reactions include serum sickness, in the USA or European Union, although about vasculitis, various cutaneous manifestations, 350 new mAbs for therapeutic application in interstitial pneumonitis, and acute respiratory humans are in the commercial pipeline. distress syndrome. • So far, the number of target antigens for the • As genetic engineering technology advances, mAbs is surprisingly small with more than attention is turning to improving the perfor- one of the approved antibodies specifi c for mance and effi ciency of mAbs in terms of TNF, HER2, CD20, EGFR, or VEGF. Other increased selectivity, improved pharmacoki- specifi cities include EpCAM, glycoprotein netics, higher binding affi nities, more effi cient IIb/IIIa, CD30, CD52, C5, α-4 integrin, IgE, cytotoxicity, better tissue penetration, and IL-6R, BLys, IL-1β, and RANK-L. increased half-life in serum. • Initial infusion reaction to some mAbs, for • Etanercept is a recombinant, engineered, fully example, rituximab, may provoke tumor lysis human dimeric fusion protein of molecular syndrome, cytokine release syndrome, and weight 150 kDa made up of the extracellular systemic infl ammatory response syndrome. ligand- binding portion of human 75 kDa TNF • Omalizumab, a humanized IgG1κ mAb with receptor linked to an Fc portion of human IgG1. specifi city for human IgE antibodies, is U.S. FDA data on etanercept adverse events approved for the treatment of severe allergic lists, in order of frequency, infections, followed asthma. It binds to free, circulating IgE anti- by dermatologic, neurologic, musculoskeletal, bodies and membrane-bound IgE molecules on pulmonary, cardiac, and vascular effects. some cells such as B lymphocytes expressing • Two synthetic IFNs, pegylated IFNα-2a and the antibody, but it does not bind to IgE already pegylated IFNα-2b, have found application as bound to mast cells. The incidence of anaphy- antiviral agents in the treatment of hepatitis C laxis to the mAb is about 0.2 %. virus.

[email protected] Further Reading 385

• Adverse reactions to IFNα include ‘fl u’-like • Allergic reactions to recombinant human symptoms, a wide range of cutaneous reac- insulin with cross-reactivity to bovine and tions, autoimmune thrombocytopenia, and porcine insulins occasionally occur. hemolytic anemia. • Reactions to IL-2 immunotherapy include erythema, urticaria, and a variety of other Further Reading cutaneous reactions. There appears to be no report of anaphylaxis. Barnes E, Salio M, Cerundolo V, et al. Impact of alpha • Replacing the receptor-binding domain of the interferon and ribaviron on the function of maturing dendritic cells. Antimicrob Agents Chemother. 2004; diphtheria toxin by the sequences encoding 48:3382–9. the IL-2 gene produced the recombinant Bernardes GJL, Casi G, Trűssel S, et al. A traceless vascular- targeting antibody-drug conjugate for cancer fusion toxin designated DAB389 IL-2 or denileukin diftitox. Cutaneous reactions to deni- therapy. Angew Chem Int Ed. 2012;51:941–4. Cheifetz A, Smedley M, Martin S, et al. The incidence leukin diftitox include injection site reaction, and management of infusion reactions to infl iximab: a erythema, and pruritus. There has been one large center experience. Am J Gastroenterol. 2003;98: fatal case of toxic epidermal necrolysis. 1315–24. • Afl ibercept, used to treat metastatic colorectal Chung CH. Managing premedications and the risk for reactions to infusional monoclonal antibody therapy. cancer and wet macular degeneration, is a Oncologist. 2008;13:725–32. fusion protein of the Fc piece of IgG1 and the Chung CH, Mirakhur B, Chan E, et al. Cetuximab- extracellular ligand-binding domains of induced anaphylaxis and IgE specifi c for galactose-α- human vascular endothelial growth factor 1,3-galactose. N Engl J Med. 2008;358:1109–17. Corren J, Casale TB, Lanier B, et al. Safety and tolerabil- receptors VEGFR1 and VEGFR2. It acts as a ity of omalizumab. Clin Exp Allergy. 2009;39: soluble decoy VEGF receptor and angiogene- 788–97. sis inhibitor. Common adverse reactions Fleischmann RM, Tesser J, Schiff MH, et al. Safety include cytopenias, hemorrhage, proteinuria, of extended treatment with anakinra in patients with rheumatoid arthritis. Ann Rheum Dis. 2006; hypertension, acral erythema, hyperpigmenta- 65:6–12. tion, and stomatitis. Foss FM, Bacha P, Osann KE, et al. Biological correlates • Anakinra, a specifi c receptor antagonist for of acute hypersensitivity events with DAB389 IL-2 (deni- ® IL-1, is a 153 amino acid non-glycosylated, leukin diftitox, Ontak ) in cutaneous T-cell lymphoma: Decreased frequency and severity with steroid premed- molecular weight 17.258 kDa recombinant ication. Clin Lymphoma. 2001;1: 298–302. protein prepared in E. coli . Cutaneous reac- Hansel TT, Kropshofer H, Singer T, et al. The safety and tions following anakinra medication are usu- side effects of monoclonal antibodies. Nat Rev Drug ally at the injection site and usually well Discov. 2010;9:325–38. Kaneko E, Niwa R. Optimizing therapeutic antibody tolerated. In some cases, cutaneous reactions function: progress with Fc domain engineering. are associated with systemic involvement. BioDrugs. 2011;25:1–11. • Serious immune-mediated reactions to anti- Li J, Zhu Z. Research and development of next generation thymocyte globulin include anaphylaxis, of antibody-based therapeutics. Acta Pharmacol Sin. 2010;31:1198–207. severe cytokine release syndrome, and severe Reichert JM. Which are the antibodies to watch in 2012? acute infusion-associated reactions. Serum MAbs. 2012;4:1–3. sickness with fever, rash, arthralgia, and myal- Scott AM, Wolchok JD, Old LJ. Antibody therapy of can- gia may appear 5–15 days after the initiation cer. Nat Rev Cancer. 2012;12:278–87. Senter PD. Potent antibody drug conjugates for cancer of therapy. Cutaneous reactions seen include therapy. Curr Opin Chem Biol. 2009;13:235–44. urticaria, morbilliform eruptions, and acral Shim H. One target, different effects: a comparison of dis- erythematous eruptions preceding rash. tinct therapeutic antibodies against the same targets. • Reactions to epoetins are rare, but anaphylaxis Exp Mol Med. 2011;43:539–49. Winkler U, Jensen M, Manzke O, et al. Cytokine-release to the recombinant forms of the hormone EPO syndrome in patients with B-cell chronic lymphocytic have been reported, one in particular after leukemia and high lymphocyte counts after treatment injection site reactions in a patient with pure with an anti-CD20 monoclonal antibody (rituximab, red cell aplasia. IDEC-C2B8). Blood. 1999;94:2217–24.

[email protected] Corticosteroids 1 2

Abstract Despite their anti-infl ammatory and immunosuppressive properties, corticosteroids (CSs) occasionally provoke immediate type I and delayed type IV allergic reactions. Frequency of reactions after topical application is 0.2–5.98 %; for systemic reactions, the incidence is 0.1–0.3 %. The reaction of steroid glyoxals with arginine is important for allergenicity of CSs. Allergic contact eczema is the most common delayed hypersensitivity reaction to CSs. Type IV reactions are mainly diagnosed by patch testing with tixocortol pivalate, budesonide, and hydrocortisone 17-buyrate being the principal drugs employed for testing. A combination of tixocortol pivalate and budesonide detects over 91.3 % of allergic patients. CSs have been classifi ed into four structural groups A, B, C, and D on the basis of clinical cross-reactivity patterns obtained from patch test results. Recently, a new classifi cation divides the CSs into three groups (1) Those that produce most of the allergic reactions—CSs that are non-methylated and usually nonhalogenated. (2) Halogenated molecules with a C16/C17 cis -ketal/ diol structure. (3) CSs that produce allergy rarely—halogenated and C16- methylated CSs. Anti-infl ammatory and immunosuppressive actions of CSs do not seem to affect immediate reactions. Skin testing, both prick and intracutaneous, together with challenge tests form the basis of the diagnosis of immediate systemic reactions to CSs.

corticosteroids (CSs) do occasionally induce 12.1 Introduction, Incidence immediate type I and delayed type IV allergic of Allergy, and Sensitization reactions. Type II cytotoxic and type III immune complex reactions do not appear to have been Although it might seem counterintuitive that reported. Leaving aside their pharmacological drugs with pronounced anti-inﬂ ammatory and activities, these drugs are so widely and heavily immunosuppressive properties can provoke both used, their relative rarity in eliciting reactions immunological and inﬂ ammatory responses, might be seen as unusual. In fact, when reactions

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 387 Relationships, DOI 10.1007/978-1-4614-7261-2_12, © Springer Science+Business Media, LLC 2013

[email protected] 388 12 Corticosteroids to CSs do occur, there is the likelihood that they foot dermatitis, and stasis dermatitis suggesting may be masked by the drugs’ anti-infl ammatory the infl uence of skin barrier breakdown and properties and diagnoses of allergic reactions involvement of antigen-presenting Langerhan’s may be missed. In cases of contact allergy, symp- cells. A surprising fi nding in the study of the 315 toms are often minor and not always obvious, European CS-allergic patients was the discovery and lesions are rarely alarming and generally that 15 of 22 sensitized by the inhalation of ste- chronic. Reports of incidences of allergy to CSs roids were not themselves treated by CS aerosols— vary, sometimes markedly. For reactions after sensitization apparently resulted from exposure topical application, frequencies from 0.2 to to CS used by others in their environment who 5.98 % are claimed. From 11,596 European they were taking care of. Sensitization by airborne patients surveyed over a period of 18.5 years, exposure to budesonide in ophthalmic prepara- patch tests revealed 315 with at least one reactions was also found in 19 patients. Forty-fi ve of tion to a CS, which is an incidence of 2.7 %. the 315 patients (14 %) had previous exposure to Eighty fi ve percent of the 315 patients reacted to systemic CSs. more than one CS. An overall incidence of 2.6 % For systemic reactions, the best estimate so far was shown in a multicenter study from 10 of the incidence of reactions is 0.1–0.3 %. European countries undertaken by the European Environmental and Contact Dermatitis Research Group. Results were variable, ranging from 0.4 12.2 Corticosteroid Haptens: to 0.6 % in Portugal and Spain to 6.4 % in Metabolism and Degradation Belgium. A retrospective study of patch testing at the Mayo Clinic, USA, of 1,188 patients over It is currently believed that the allergenic struc- a 6 year period (2000–2005) revealed 10.7 % of ture of CSs hydroxylated or esterifi ed at C21 patients reacted to at least one CS and 4.7 % (e.g., as occurs with hydrocortisone (Fig. 12.1a ) reacted to multiple CSs. Tixocortol pivalate and methylprednisolone succinates) are not the alone detected less than 50 % of the allergic unaltered parent drugs but a metabolite formed patients. within the skin when the CS degrades to a reac- Patients allergic to a CS often react to multiple tive glyoxal that binds to nucleophilic protein allergens, but it remains unclear whether atopy is residues. Steroid glyoxals react with most of the a risk factor for CS hypersensitivity. A history of amino acids but the reaction with arginine pre- atopy was found in 34 % of the 315 CS-allergic dominates and seems to be the most important patients but overall, it is diffi cult to determine since those CSs with the greatest capacity to bind whether a higher incidence is due to increased to arginine are also the strongest allergens. The susceptibility or greater exposure. Few reactions breakdown of the C21 esters is unaffected by the in asthmatics have been noted despite the length of the carbon chain. It is not clear what extremely widespread use of inhaled CSs. Risk mechanisms operate, if any, to form altered aller- factors for systemic sensitivity do not seem to genic structures from CSs that are neither hydrox- have been studied but some reports indicate a ylated nor esterifi ed but have a halogen atom at higher incidence in asthmatic subjects. C21 (e.g., clorbetasol and halcinonide). The extensive use of CSs is affected by all Substituents on the gonane nucleus of CSs also delivery routes—parenterally, via cutaneous con- affect the anti-infl ammatory activity and the rate junctival, nasal, oral and rectal mucosa, and the of degradation of the compounds. The oxygen at respiratory and gastrointestinal tracts, so there is C11 enhances degradation whereas the β-methyl ample opportunity for sensitization to these drugs group at C16 and hydrogen at C17 stabilize the to occur. The risk of sensitization via topical molecule. Halogens at C6 and C9 have been application is highest for patients with long-term claimed to both stabilize and determine aller- skin conditions such as chronic eczema, hand and genic cross-reaction patterns of CSs.

[email protected] a OH O 18 20 12 21 HO 11 17 OH 19 13 CD16 1 9 H 14 2 10 15 H 8 AB H 7 O 3 5 4 6 Hydrocortisone

HO b c S O O O O HO OH HO O H H H HH HH

O O Tixocortol pivalate Triamcinolone acetonide Group A Group B

deOH OH O O

HO OH HO O H H O FH HH

O O Betamethasone Hydrocortisone-17-butyrate Group C Group D

f O O HO O H H HH

O Budesonide (classified in group B)

Fig. 12.1 Corticosteroids from the four structural groups taneperhydrophenanthrene nucleus of 17 carbons shown A , B , C, and D classiﬁ ed on the basis of clinical cross- numbered on the hydrocortisone structure in a . Note that reactivity patterns obtained from patch test results. the A – D designation of the four different rings of the Drug examples from each of the four groups are shown. gonane nucleus should not be confused with the A , B , C , The basic structure of the CSs is the gonane or cyclopen- D classiﬁ cation of cross-reactivity

[email protected] 390 12 Corticosteroids

12.3 Delayed Reactions

12.3.1 Clinical Presentation

Reports of delayed hypersensitivity reactions to CSs include allergic contact dermatitis (Fig. 12.2 ), maculopapular rash, exanthematous, papulo- vesicular and fl exural rashes, rash with or without bullae or purpura, acute generalized exanthematous pustulosis, and erythema multiforme. Allergic contact eczema is the most common delayed hypersensitivity reaction to CSs. Affected skin sites in order of frequency are hands, legs, face, feet, a generalized reaction, arms, eyes, trunk, and neck. Signs of a reaction are often minor, but if the time to reach a diagnosis has been lengthy, effects such as rosacea, perioral Fig. 12.2 Allergic contact dermatitis to hydrocortisone (Fig. 12.3) or perinasal dermatitis, and cutaneous showing minor infl ammation (Photograph courtesy of A. atrophy may be seen. What may appear to be Goossens). From Brandão FM, in Johansen JD, Frosch PJ, chronic eczema with “edge effects,” that is, a Lepoittevin J-P, editors. Contact Dermatitis, 5th ed. Berlin: Springer-Verlag; 2011. Reprinted with permission reaction that is more pronounced at the periphery from Springer Science + Business Media of the treated area, may in fact be allergic contact dermatitis induced by CSs. Reactions due to ocular use of CSs may manifest as facial edema, eczema, and conjunctivitis while those resulting from inhalant use include eczematous eruptions around the facial orifi ces with possible mucosal involvement. A small number of patients present with generalized eruptions and, of these, an occasional patient also has systemic symptoms such as hypotension and malaise. Following systemic CS administration, generalized maculopapular or eczematous eruptions may sometimes occur. Note that CS-sensitive diseases that are exacerbated by CS treatment, respond poorly or not at all, or reoccur soon after CS treatment is discontinued should be considered as a possible allergy to CSs. Fig. 12.3 Perioral dermatits. From Zaidi Z, Lanigan SW. Dermatology in Clinical Practice. London: Springer- Verlag; 2012. Reprinted with permission from Springer Science + Business Media 12.3.2 Diagnosis of Corticosteroid Hypersensitivity most useful CSs for the detection of CS contact 12.3.2.1 Drugs for Patch Testing allergy in 2,123 patients showed that 5.98 % of Type IV reactions to CSs are mainly diagnosed patients were allergic to one or more CSs; 4.5 % by patch testing. Tixocortol pivalate, budesonide, to tixocortol pivalate, 2.4 % to hydrocortisone and hydrocortisone 17-buyrate are the principal butyrate, 2.2 % to budesonide, 0.52 % to each of drugs employed for testing. An evaluation of the betamethasone valerate and clobetasone butyrate,

[email protected] 12.3 Delayed Reactions 391 and 0.38 % to clobetasol propionate. A combina- 12.3.2.2 Concentrations of Drugs tion of tixocortol pivalate and budesonide for Patch Testing detected 91.3 % of CS-allergic patients suggest- Care should be exercised by those seeking test ing that both should be included as standard test concentrations of CSs in the literature since there agents when patch testing for CS delayed hyper- is no universal agreement, and numerous authors sensitivity. Of the 315 patients positive to at least have published different “optimum” fi gures. For one CS in the aforementioned European study, example, in a study of contact allergy to CSs pub- 61 % reacted to budesonide, 43 % to tixocortol lished in 2000, concentrations of 0.1 and 1 % in pivalate, 31 % to hydrocortisone 17-butyrate, and alcohol or petrolatum were recommended for 22 % to prednisolone caproate. Importantly, posi- budesonide and tixocortol pivalate, respectively. tive tests to other CSs occurred with 91, 81, and In a 2009 study with one author common to both 99 % of patients allergic to budesonide, tixocor- publications, budesonide 0.01–0.02 % in petrola- tol pivalate, and hydrocortisone 17-butyrate, tum and 0.002 % in ethanol and 0.1 % tixocortol respectively. Budesonide and tixocortol pivalate pivalate were said to detect most patients allergic together detected 88.5 % of the CS-allergic to CSs. This is not surprising and is perhaps patients and addition of hydrocortisone explained by the anti-infl ammatory effect of too 17-butyrate increased the detection fi gure to high a concentration of CS. This could be espe- 92.5 %. Of course, other CSs used by the patient cially important in patients mounting weak reac- should also be tested along with the appropriate tions so it might be said that in such patients, an vehicle controls. A mixture of the above three inverse relationship exists between positive patch CSs detected less than 50 % of patients allergic to tests and the concentrations of test agents. tixocortol pivalate (presumably because of mask- However, the thorough investigator should be ing of anti-infl ammatory effects by the different prepared to try different concentrations of test drugs in the mixture) and therefore each drug materials, including higher concentrations, espe- should be tested separately. cially in patients with a negative test result but a Intradermal testing has been applied to the clear clinical suspicion of allergy. Hydrocortisone diagnosis of CS allergy but the atrophy-inducing 17-butyrate and most other CSs are tested at a effect of the drugs in this presentation form, par- concentration of 1 %. ticularly for suspensions, is a practical limitation. For CSs not known to induce atrophy, intrader- 12.3.2.3 Reading Patch Tests mal tests with solutions formulated at 30, 10, and Reading and interpretation of patch tests should 1 % in saline may be considered for particular always be undertaken by a well-trained and expe- cases, that is, for patients with a suggestive his- rienced person and the need for that requirement tory of allergy but a negative patch test fi nding. In is even more obvious in the diagnosis of CS aller- a comparison of patch testing with CSs in 1 % gies. Because the anti-infl ammatory effect of CSs ethanol and intradermal tests with CSs suspended can mask allergic reactions, too high a concentra- in physiological saline, tixocortol pivalate, and tion of test agents may give false negative reac- budesonide detected all patients allergic to hydro- tions and this has led to the recommendation of cortisone and budesonide, respectively, but patch extended reading times to 6 or 7 days or even tests with hydrocortisone 17-butyrate failed to longer. The edge effect is a major factor to con- detect 30 % of the positive reactions detected by sider in reading patch test results for CSs. intradermal testing. The study also demonstrated Manifesting as a clear area of no reaction sur- that use of ethanol as a vehicle for other CSs led rounded by erythema, induration, and/or papulo- to both false positive and false negative reactions. vesicles, this phenomenon is probably due to the It was suggested that consideration should be anti-infl ammatory effect of the CS in the center given to the avoidance of hydrocortisone where the concentration is highest and the 17-butyrate testing in patients patch test positive grading into lower concentrations as the drug dif- to tixocortol pivalate and budesonide. fuses outward where the anti-infl ammatory effect

[email protected] 392 12 Corticosteroids becomes less pronounced and the infl ammatory that fresh and stored solutions gave similar results reaction correspondingly more apparent. In later in skin tests. In one comparative study of stabili- readings, this reaction pattern is less clear until ties, budesonide, tixocortol pivalate, and hydro- the test site becomes entirely eczematous. Initial cortisone 17-butyrate in ethanol and petrolatum whitening of the skin test site due to vasocon- were kept at room temperature, refrigerated and striction followed by vasodilation accompanied deep frozen, and checked for stability by high by erythema are other possible effects that can performance liquid chromatography over a 1 year lead inexperienced readers of patch test results to period. The ethanolic and petrolatum preparations interpret them as negative or positive reactions, of bunesonide and tixocortol pivalate remained respectively. stable for the entire storage period irrespective of the storage conditions. Hydrocortisone 17-butyr- 12.3.2.4 The Vehicle ate 1 % in ethanol was also stable for the same for Corticosteroids period but was stable for only 3 months at room in Patch Testing temperature. It has been suggested that retention The amount of skin penetration of CSs used in of skin test reactivity after storage may be due to testing is obviously important and this is to a the breakdown products which are, in fact, the large extent infl uenced by the vehicle employed. allergens rather than the parent compounds. As The drug needs to preferably be soluble, or at this subject remains sparingly investigated at best, least well dispersed, and stable in the vehicle to it seems prudent to prepare fresh solutions of CS ensure optimal presentation and availability. test agents at least every 4, or perhaps 6, weeks. Vehicles most often used are ethanol, petrolatum, physiological saline, and sometimes dimethyl 12.3.2.6 Other Diagnostic Tests sulfoxide. For most CSs, ethanol is usually used In the early 1990s, Wilkinson in England showed but for hydrocortisone penetration is better with that a hydrocortisone–albumin complex could an equal mixture of ethanol and dimethyl sulfox- induce a proliferative response in peripheral ide as vehicle. Matura, in tests on CS-allergic blood mononuclear cells from patients allergic to patients with a large panel of CSs, did not detect the CS. In the same period, Lauerma and col- signifi cant differences between tixocortol piva- leagues in Finland demonstrated fl are-up in skin late in ethanol and petrolatum but budesonide reactions after oral hydrocortisone in patients and hydrocortisone 17-butyrate showed more with contact allergy to the drug and proliferative positive reactions in ethanol than in petrolatum. responses to corticosteroids in T lymphocytes Controls for patch testing should include each of from patients with CS contact hypersensitivity. the vehicles used. Positive patch test reactions However, cell proliferation only occurred when have been reported for ethanol, ethanol/dimethyl epidermal Langerhans’ cells were present and sulfoxide, glutaraldehyde, and glyoxal. not when peripheral blood monocytes were pres- Preservatives and other agents added to CS for- ent as antigen-presenting cells. The lymphocyte mulations may also need to be ruled out as the transformation test, discussed in Sect. 4.7.1 , is elicitor of a positive reaction. Substances that are said to have a sensitivity of 60–70 %, but there sometimes added in formulating topical CS prepare a number of practical issues that make the test arations include neomycin, propylene glycol, problematic for many clinical departments and benzyl alcohol, sorbic acid, wool alcohols, and even research laboratories. More recently, two many others. Where possible, these added sub- cases of prednisolone-induced acute generalized stances should also be patch tested to ensure that exanthematous pustulosis were confi rmed by they themselves do not provoke reactions. positive skin and lymphocyte transformation tests to prednisolone and structurally related CSs. 12.3.2.5 Stability of Test Solutions Other techniques like ELISPOT assays for cyto- CSs have limited stability in ethanol; in one study kines such as IFN-γ (Sect. 4.7.3.2 ) appear to be the chromatographic purity of some CSs fell to potentially promising techniques to aid the diag- 75–95 %. However, several investigations showed nosis of delayed reactions to CSs.

[email protected] 12.3 Delayed Reactions 393

12.3.3 Structure–Activity group (bridged C16–C17). Cross- reaction with Relationships group D esters is thought to be due to the hydro- of Corticosteroid phobic cavity present in the S isomer. This resem- Cross-Reactions blance of different isomeric forms of budesonide to both the acetonide structure and C17 esters The basic structure of the CSs is the gonane or appears to explain why budesonide is such a use- cyclopentaneperhydrophenanthrene nucleus of ful patch test screening agent. About 90 % or 17 carbons. The CSs contain an extra two (as in more of individuals with a positive patch test testosterone) or four carbon atoms and some in reaction to budesonide are allergic to other CSs. the latter group, for example, cortisol or hydro- Despite the apparent utility of the four category cortisone, have a hydroxyl group at position 17 CS structure–activity classifi cation, cross- (Fig. 12.1a ). In one of the few comprehensive reactions between different groups do occur, par- structure–activity analyses undertaken in the ticularly between groups A and D, and fi eld of delayed drug hypersensitivities, CSs were consequently alternative explanations have been classifi ed into four structural groups A, B, C, and advanced. Substitutions of halogen atoms at car- D on the basis of clinical cross-reactivity patterns bons 6 and 9 on the B ring has been suggested as obtained from patch test results on 19 patients a basis of cross-reactivity between CSs but while and from the literature. Drug examples corre- this seems to hold for groups A, C, and D and sponding to each group are: Group A, tixocortol positive reactions to fl uorinated derivatives occur pivalate; group B, triamcinolone acetonide; less frequently, there seems to be no difference group C, betamethasone; group D, hydrocortisone- between halogenated and nonhalogenated deriva- 17-butyrate (Fig. 12.1b–e ). The D ring was iden- tives in group B. In a modifi cation of the C6/C9 tifi ed as the structural feature that distinguished cross-reaction hypothesis, a second principal the different groups, and it was suggested that immune recognition site, viz. C16/C17 was also allergic reactions occurred more often to CSs of suggested. It seems therefore that all participants the same group. (Note that the A to D designation in the study of CS cross-reactions appear to agree of the four different rings of the gonane nucleus on the importance of C16/17 substitutions on the should not be confused with the A, B, C, D clas- D ring of the steroid backbone. However, in what sifi cation of cross-reactivity.) is surely the most confusing and immunochemi- Subsequently, this classifi cation was supported cally poorly based explanation of immune cross- by conformational analysis and statistical calcula- reactivity between different member drugs with a tions on some CSs which were interpreted as common basic structure, the original A, B, C, D showing that within each group A, B, and D, CS classifi cation was recently “simplifi ed” into structures were homogeneous in volume, shape, three groups. The D ring is said to have an impor- and charge distribution. The situation with group tant role in CS cross-reactivity as a result of amino C remained unclear. The fi ndings were said to acids (such as arginine) in skin proteins binding at explain why cross-reactions between CSs from C21. After the initial subdivision of the CSs into different groups are unlikely. Clinical cross- the four groups, group D was further divided into reactivity between budesonide (Fig. 12.1f), which two subgroups D1, the “stable” esters, and D2, is classifi ed in group B, and some CSs in group D the “labile” esters. It has been hypothesized that was also explained by conformational consider- non-methylated CSs are implicated in sensitiza- ations. The clinical cross-reaction exhibited by tion to CSs after selective reaction with arginine this CS and group D compounds was said to be to form stable cyclic adducts and, with this in due to the S diastereoisomer of budesonide while mind, C16-methylated and non-methylated CSs cross-reactions with group B are said to arise were compared in patch tests on patients with from both the R and S forms. In terms of the struc- proven CS contact allergy. Results showed that tures recognized, budesonide, as an equal mixture signifi cantly more positive reactions occurred of R and S diastereoisomers, is thought to cross- with groups A and D2 CSs without C16 methyl react with group B molecules via the acetonide substituents than with groups C and D1 molecules

[email protected] 394 12 Corticosteroids carrying a C16 methyl group. This led to a new results cannot automatically be extrapolated to classifi cation of CSs into three groups (1) CSs chronic urticaria, it seems likely that CSs are that produce most of the allergic reactions—CSs unlikely to prevent or to improve this condition. that are non-methylated and usually nonhaloge- However, in the case of systemic immediate reac- nated and essentially made up of groups A and D2 tions, the situation may be different as discussed plus budesonide. (2) The halogenated molecules below (Sect. 12.4.2 ). with a C16/C17 cis-ketal/diol structure, that is, acetonide group B. (3) CSs that produce allergy rarely—halogenated and C16-methylated CSs, 12.4.1 Incidence, Sensitization, that is, groups C and D1. It remains to be seen if Clinical Presentation, this is the last word on CS cross-reaction patterns and Risk Factors and classifi cation of the different patterns. Some comments seem pertinent to this body of With an incidence estimated to be about 0.1– work on structure–activity relationships and 0.3 %, immediate reactions to CSs are fortunately cross-reactivities of topically applied CSs. In the rare given the widespread usage of these drugs in fi rst place, the conclusions should not be automat- many different diseases and because the risk of ically applied to delayed and immediate reactions anaphylaxis to CSs may not be considered by following systemic administration of CSs. some clinicians who may think that progressing Second, the fact that the above interpretations and symptoms are due to insuffi cient dosage. There conclusions resulted from work unconnected to are over 100 published reports of immediate type I and type IV systemic reactions indicates hypersensitivity reactions to CSs, including many why the data used to understand cross-reactivities life-threatening reactions, after oral and paren- at the cutaneous level are so diffi cult to explain teral administration. Most reactions have and categorize and why the presented analyses of occurred in adults with surprisingly few reactions the data are so complex and frequently, and differ- in children even though administration of CSs to ently, interpreted. Results obtained from patch children is common. Immediate reactions have testing can, at best, be semiquantitative and mean- followed after IV, intramuscular, subcutaneous, ingful, and accurate comparisons of recognition and intra-articular injection although the IV route and reactivities of individual CSs are not possible is the most frequent elicitor of reactions. on a molar basis. This is unlike the situation where Symptoms include angioedema, generalized urti- specifi c IgE antibodies or T cells (less easily) can caria, pruritus, bronchospasm, hypotension, be utilized for side-by-side comparisons of recog- dyspnea, sneezing, nausea/vomiting, decreased nition of structurally similar and dissimilar drugs. consciousness, respiratory arrest, and anaphylactic shock including death. Risk factors include asthma, particularly aspirin-sensitive 12.4 Immediate Reactions asthmatics, and renal transplant patients but it to Corticosteroids remains unclear whether atopy and heavy or prolonged exposure are risks—in fact, factors pre- Interestingly, the anti-infl ammatory and immu- disposing patients to risk of immediate reactions nosuppressive actions of CSs seen with delayed to CSs seem little investigated. cutaneous reactions to the drugs do not seem to affect immediate reactions in the skin as shown by the failure of locally injected methylpredniso- 12.4.2 Immediate reactions: lone to prevent or attenuate cutaneous reactivity Important/Interesting induced by prick tests of histamine and the hista- Findings and Observations mine liberator codeine. In fact, when injected at Reported so Far the prick test site, the locally administered CS was associated with a signifi cant increase in the Some systemic reactions to CSs are pseudoaller- fl are induced by both agents. Although these gic, and there are many cases in the literature

[email protected] 12.4 Immediate Reactions to Corticosteroids 395 where clear evidence for a true type I or type IV codeine (see above), it seems that immunosup- reaction is lacking. Almost certainly some of pressive and/or anti-infl ammatory actions of the these reactions are anaphylactoid in nature, and it methylprednisolone delayed the positive response has been pointed out that such reactions may at the skin test site initially negative at 60 min. result from infusion of high doses of CSs such as The delayed appearance of positive skin tests to methylprednisolone. Unlike true anaphylactic CSs suggests that it may be prudent to read such reactions, anaphylactoid reactions to drugs may tests after an extended period, for example, be related to both the dose of drug and its rate of 90 min or 2 h or more, if earlier readings at 20 delivery. With methylprednisolone, for example, and 60 min prove negative. In attempting to doses of 11 mg–2 g have caused systemic reac- explain the appearance of the positive skin test to tions, but most reactions occur after administra- prednisolone after the intramuscular challenge tion of more than 250 mg. In high dose therapy with the drug, the investigators speculated that, (more than 30 mg/kg), the drug should be infused unlike with the challenge test, the amount of drug over 30 min; more rapid infusion signifi cantly injected intradermally was too small to form increases the chance of an anaphylactoid response. enough hapten–protein antigenic complex by As already mentioned, there is no shortage of binding to arginine on human albumin. reports of immediate reactions to CSs but most of To overcome their poor solubility, CSs are the results add little new information relevant to often esterifi ed at position C21 and used as the improving diagnosis or elucidating underlying succinate, phosphate, or other esters. Succinate mechanisms. Much of the reported information is esters appear to be more immunogenic, but this similar, including the predominance of methyl- has not been unequivocally established and no prednisolone and hydrocortisone in provoking mechanism of increased antigenic/allergenic reactions; the spectrum of symptoms observed; potential has been convincingly argued or dem- the limited diagnostic investigations employed; onstrated. As for CSs in general, it is thought that and the frequent absence of clear conclusions on CS esters act as haptens in larger molecular cross-reactions, but occasional skin test fi ndings weight complexes with carrier molecules. The are noteworthy. Over 40 years ago, intradermal question of cross-reactivity between CSs in tests for hydrocortisone, prednisolone acetate, immediate reactions has not always been clear. and prednisone were positive in a patient who The rather complex cross-reactivity story elabo- experienced hives after oral and intra-articular rated for the topical CSs does not seem to apply CS administration but only after an interval of 3 h for systemic reactions and different patterns of or more. This “delayed” positive skin response cross-reactivity, not easily categorized, have been has been occasionally observed by others, a good observed. Cross-reactions between steroids with recent example being in a 2008 study when an similar chemical structures are seen but often intramuscular challenge with 10 mg of methyl- they belong to different groups used in the topical prednisolone hemisuccinate precipitated an CS classifi cation. Other reports describe cross- immediate reaction and a positive intradermal reactions between group A CSs, others failed to test in a prednisolone-allergic patient who was detect any cross-reactivity, and recognition of initially skin test negative to the drug when the different succinate esters has also been noted. In test was read at 20 and 60 min earlier the same the latter category, a patient who experienced an day. Subsequent intradermal tests with hydrocor- IgE-mediated anaphylactic reaction to tisone, betamethasone, triamcinolone, and para- methylprednisolone- 21-succinate sodium proved methasone and intramuscular challenges with prick test positive to this drug and to prednisolone- hydrocortisone, betamethasone, and parametha- 21-sodium succinate but not to prednisolone sone all produced negative results. Unlike the (i.e., the unesterifi ed drug) or to betamethasone- fi ndings with locally administered CSs at sites 21- dihydrogen phosphate, and oral challenges previously prick tested with histamine and with both of these drugs were well tolerated.

[email protected] 396 12 Corticosteroids

The succinate ester grouping was suspected to be Table 12.1 Skin test concentrations for some commonly allergenically important since other nonesterifi ed used corticosteroid drugs CSs and CSs with groups other than succinate at Prick test Intradermal a C21 were negative in both prick and challenge Corticosteroid (mg/ml) test (mg/ml) tests. Wide cross-reactivity between CSs could Betamethasone phosphate 4 0.04 b indicate immune recognition of the steroid back- Budesonide 0.25 0.0025 Dexamethasone phosphate 4 0.04 bone of the CS drugs whereas lack of cross-reac- Hydrocortisone succinate 100 1 tivity with other CSs is probably a consequence Methylprednisolone 40 0.4 of recognition of individual, and sometimes succinate unique, structural features. Prednisolone succinate 10c 0.1 Prednisone 30 – Triamcinolone acetonide 40 0.4 12.4.3 Identifying and Sensitizations to the excipients carboxymethylcellulose Understanding Cross- and macrogel have been observed in some patients with Reactions: Finding a Safe corticosteroid hypersensitivity aSome investigators use up to a maximum of 10× these Alternative Corticosteroid concentrations, i.e., a 1–10 dilution of the prick test concentrations At the clinical level, negative skin and challenge bCommonly used in patch tests at 0.01 % in petrolatum tests can identify safe and alternative CSs for and 0.002 % in ethanol c Up to 30 mg/ml has been used patients who have experienced a CS-mediated immediate allergic reaction. If cross-reactions, weak or strong, are identifi ed in skin tests, systematic challenge testing might be employed, but the risks of challenge testing with even weakly 12.4.4 Diagnostic Methods cross-reacting drugs must be very carefully considered. If the risk to benefi ts ratio is judged to be Skin testing, both prick and intracutaneous, with in the patient’s best interest, extreme care with all appropriate concentrations of drugs together with precautions should be followed in the provoca- challenge tests are the basis of the diagnosis of tion testing (see Sect. 4.4). In any case, for sys- immediate systemic reactions to CSs. The recom- temic reactions to CSs, further studies are needed mended skin test concentrations are set out in on the sensitivity, specifi city, and positive and Table 12.1, but the clinician should, as always, be negative predictive values of skin tests and, for a aware that smaller concentrations may sometimes better understanding of clinical cross-reactions be advisable in the occasional patient showing between CSs, these studies are a necessary pre- exquisite sensitivity. For intradermal testing, sus- lude. For an understanding of cross-reactions in pensions of drug should not be injected. Specifi c structural terms, quantitative immunochemical IgE antibodies to CSs have rarely been described, investigations are necessary to identify fi ne struc- probably partly because an appropriate assay has tural features recognized on individual CSs and not generally been available, although a Phadia to have any chance of anticipating and predicting test for the detection of IgE antibodies to possible reactions. With reactions mediated by methylprednisolone-21- succinate sodium has drug-reactive IgE antibodies, hapten inhibition been successfully applied. Note that a negative studies employing selected CSs, analogs, and fi nding in a test for CS-reactive IgE does not nec- other carefully chosen structurally relevant com- essarily rule out immediate hypersensitivity to a pounds in hapten inhibition experiments can CS. Development of IgE assays for many CSs provide a quantitative comparison of inhibitory faces the technical problem of poor solubility of potencies and thus a spectrum of cross- many of the CSs. Another potential diffi culty recognition from those CSs that are strongly may be the presentation of the CS in an appropri- cross-reactive to those that are completely unrec- ate haptenic form, which is the drug-carrier com- ognized by the sensitizing antibodies. plex form involved in the initial sensitization and/

[email protected] Further Reading 397 or immediate allergic reaction. The basophil acti- • ELISPOT assays for cytokines such as IFN-γ vation test offers another potential test for the appear to be potentially promising techniques detection of an immediate reaction, and it has to aid the diagnosis of delayed reactions been successfully employed in at least three sepa- to CSs. rate diagnostic investigations in different centers. • CSs have been classifi ed into four structural In one case, CD63 expression was measured after groups A, B, C, and D on the basis of clinical the patient’s blood was incubated for 30 min with cross-reactivity patterns obtained from patch methylprednisolone (0.1 μg/ml). The drug acti- test results. Drug examples corresponding to vated 20.2 % of the basophils compared to 6 % in each group are: Group A, tixocortol pivalate; control samples for a stimulation index of 3.4. group B, triamcinolone acetonide; group C, betamethasone; group D, hydrocortisone-17- butyrate. After the initial subdivision of the Summary CSs into the four groups, group D was further divided into two subgroups D1, the “stable” • Despite their anti-infl ammatory and immuno- esters, and D2, the “labile” esters. suppressive properties, CSs can provoke both • A recent new classifi cation divides the CSs immunological and infl ammatory responses, into three groups (1) CSs that produce most of occasionally inducing immediate type I and the allergic reactions—CSs that are non- delayed type IV allergic reactions. Type II methylated, usually nonhalogenated and cytotoxic and type III immune complex reac- essentially made up of groups A and D2 plus tions do not appear to have been reported. budesonide. (2) The halogenated molecules • For reactions after topical application, fre- with a C16/C17 cis-ketal/diol structure, that quencies of from 0.2 to 5.98 % are claimed. is, acetonide group B. (3) CSs that produce For systemic reactions, the incidence of reac- allergy rarely—halogenated and C16- tions is 0.1–0.3 %. methylated CSs, that is, groups C and D1. • The reaction of steroid glyoxals with arginine • Anti-infl ammatory and immunosuppressive seems to be important for allergenicity of CSs actions of CSs seen with delayed cutaneous since those CSs with the greatest capacity to bind reactions to the drugs do not seem to affect to arginine also make the strongest allergens. immediate reactions in the skin. • Allergic contact eczema is the most common • Wide cross-reactivity between CSs probably delayed hypersensitivity reaction to CSs. indicates immune recognition of the steroid Other delayed reactions include maculopapu- backbone of the CS drugs whereas lack of lar rash, exanthematous, papulo-vesicular and cross-reactivity with other CSs is probably a fl exural rashes, rash with or without bullae or consequence of recognition of individual, and purpura, acute generalized exanthematous sometimes unique, structural features. pustulosis, and erythema multiforme. • Skin testing, both prick and intracutaneous, with • Type IV reactions to CSs are mainly diagnosed appropriate concentrations of drugs together by patch testing. Tixocortol pivalate, budesonide, with challenge tests are the basis of the diagno- and hydrocortisone 17-buyrate are the principal sis of immediate systemic reactions to CSs. drugs employed for testing. A combination of tixocortol pivalate and budesonide detects over 91.3 % of CS-allergic patients. Further Reading • Because the anti-infl ammatory effect of CSs can mask allergic reactions, too high a con- Baeck M, Chemelle J-A, Terreux R, et al. Delayed hyper- centration of test agents may give false nega- sensitivity to corticosteroids in a series of 315 patients: tive reactions, and this has led to the clinical data and patch test results. Contact Dermatitis. recommendation of extended reading times. 2009;61:163–75. Baeck M, Goossens A. Immediate and delayed allergic The so-called ‘edge effect’ is a major factor to hypersensitivity to corticosteroids: practical guide- consider in reading patch test results for CSs. lines. Contact Dermatitis. 2011;66:38–45.

[email protected] 398 12 Corticosteroids

Boffa MJ, Wilkinson SM, Beck MH. Screening for corti- Lauerma AI. Contact hypersensitivity to glucocorticoste- costeroid contact hypersensitivity. Contact Dermatitis. roids. Dermatitis. 1992;3:112–32. 1995;33:149–51. Nancey S, Freymond N, Catelain A, et al. Effects of local Burgdorff T, Venemalm L, Vogt T, et al. IgE-mediated corticosteroids on acute experimental urticaria. Eur J anaphylactic reaction induced by succinate ester of Dermatol. 2004;14:323–6. methylprednisolone. Ann Allergy Asthma Immunol. Wilkinson SM. Corticosteroid cross-reactions: an alterna- 2002;89:425–8. tive view. Contact Dermatitis. 2000;42:59–63.

[email protected] Drugs Used for Chemotherapy 1 3

Abstract Many of the drugs used for chemotherapy have been, and still are, alkylating agents, antimetabolites, organoplatinum compounds, cytoskeletal disruptors, or anthracyclines, all agents with relatively broad rather than targeted and specifi c modes of action. The tyrosine kinase inhibitor imatinib mesylate and proteasome inhibitor bortezomib are recent examples of a more specifi c treatment strategy. Up to 30 % of patients develop acute infusion reactions to taxanes. Hypersensitive cross-sensitivity between docetaxel and paclitaxel is ~90 %. Most reactions to platinum drugs appear after multiple treatment cycles (usually at least six). Reactions are mainly type I or type IV hypersensitivity responses with a few cases of type II and type III hypersensitivities. The drug imatinib mesylate that inhibits both the ABL and BCR-ABL tyrosine kinases has been successful in treating chronic myeloid leukemia. In the chronic phase of treatment, neutropenia results in 35–45 % of cases, thrombocytopenia in 20 %, and anemia in 10 % of cases. Gefi tinib and erlotinib are EGFR inhibitors, inhibiting the receptor’s tyrosine kinase domain. Main hypersensitivities to both drugs include cutaneous reactions. GI symptoms, thrombocytopenia, peripheral neuropathy, and neuropathic pain are the most common side effects of the proteasome inhibitor bortezomib. Adverse cutaneous reactions to the drug are numerous.

Although a strictly literal meaning of “chemo- normal, healthy rapidly dividing cells (e.g., in the therapy” would include, for example, treatment bone marrow, mucosal linings, and hair follicles) of an infection with an antibiotic or administra- also undergo collateral harm. Recently, more tion of aspirin for a headache, the word has selective strategies in which drugs are targeted to become synonymous with drug treatment of cancer cells, for example, by recognition of cancers. Until fairly recently, chemotherapy with speciﬁ c cellular antigens, have begun to be antineoplastic drugs has been largely based on employed. In keeping with the universal dread of the broad, nonspeciﬁ c strategy of killing rapidly cancer, its widespread occurrence in so many dividing cancer cells even though some other different forms in humans of all ages, its high

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 399 Relationships, DOI 10.1007/978-1-4614-7261-2_13, © Springer Science+Business Media, LLC 2013

[email protected] 400 13 Drugs Used for Chemotherapy profi le in medical research, and the vast amounts of money invested in attempts for a “cure,” it is 13.1 Taxanes not surprising that numerous drugs have been employed for treatment of the many different Taxanes are diterpenes and the name taxane is cancers that occur in humans. Even a quick derived from the fact that these compounds were perusal of the medical literature reveals in excess found to be produced by plants of the yew genus of 100 drugs currently administered in signifi cant Taxus. The drug paclitaxel is so named because it quantities to cancer patients somewhere in the was originally identifi ed in the bark of the Pacifi c world. Of course, despite being less expensive, yew tree. Because of the diffi culties involved in some of these drugs therapies are older, less spe- their synthesis (paclitaxel for example, has 11 cifi c in their action, and often less effective than chiral centers), natural products remain a source some newer, more targeted therapies, and, for the for some taxanes. Docetaxel is a semi-synthetic immediate future, many long-established anticancer compound produced from a precursor found in drugs seemed destined to be used less and less. the needles of the European yew tree. The tax- We are interested here in covering some of the anes are mitotic inhibitors, disrupting microtu- most important currently used, and new, anti- bule function so adverse reactions, including cancer drugs and the adverse/hypersensitivity hypersensitivity responses, to these drugs when reactions they elicit. given as anticancer agents might be expected. In In the identifi cation and development of new a women’s cancer program retrospective study anticancer drugs over many years, many of the covering the period 1999–2004, severe hyper- compounds that have found application in the sensitivity reactions occurred in 16 of 718 clinic have cytotoxic properties arising from a patients (2.2 %) given paclitaxel and in 9 of 93 small number of different mechanisms of action. patients (9.7 %) who received docetaxel. Up to Many of the drugs used for treatment of malignant 30 % of patients have been found to develop cells have been, and still are, alkylating agents, acute infusion reactions to taxanes. Acute hyper- antimetabolites, organoplatinum compounds, cyto- sensitivity reactions are marked by urticaria, skeletal disruptors, or anthracyclines, all agents fl ushing, rashes, dyspnea, gastrointestinal symp- with relatively broad rather than targeted and spe- toms, hypo-, and hypertension and back pain, the cifi c modes of action. Table 13.1 summarizes some latter not yet well understood. Taxanes present important properties of, and adverse reactions to, a diffi culties in their formulation since they are selection of 17 anticancer drugs currently com- poorly soluble in aqueous media and their conse- monly used. Alkylating agents such as busulfan quent presentation in vehicles such as Cremophor and cyclophosphamide and platinum-based agents also contribute to adverse reactions. Multiple like cisplatin that cross-link DNA; inhibitors of mechanisms may underlie infusion reactions to mitotic cell division by taxanes such as paclitaxel; taxanes. Reactions may be IgE antibody-mediated drugs that intercalate with DNA like daunorubicin; or due to direct mast cell/basophil or comple- pyrimidine analogs including 5-fl uorouracil; and ment activation. The similarity in structure other antimetabolites such as pemetrexed represent between docetaxel and paclitaxel suggests cross- a broad group of drugs that can be compared and reactions and cross-sensitivity. Some early contrasted with some recently introduced agents results on small numbers of patients indicated where a more refi ned strategy of specifi c action on successful substitution of docetaxel for pacli- a particular cancer cell rather than nonspecifi c taxel, but this conclusion was not supported in inhibition and killing has been used. The tyrosine the later women’s cancer study mentioned above kinase inhibitor imatinib mesylate and proteosome where ten patients with severe hypersensitivity to inhibitor bortezomib are examples of the latter paclitaxel also reacted to docetaxel, giving a approach. Adverse reactions to these drugs will be cross-sensitivity rate of 90 %. The different vehi- discussed in detail together with reactions to some cles used for the two drugs indicated that the of the more important other groups of chemother- cross-reactions were probably due to the taxanes apeutic drugs currently widely used. and not the vehicles.

[email protected] Table 13.1 Important non-mAb anticancer (chemotherapeutic) agents and their adverse/hypersensitivity reactions Type of Adverse/hypersensitivity reactions chemotherapy drug Incidence/risk factors/ and generic name Chemical class Cancer indications Mechanism of action Cutaneous Systemic diagnosis/comments Alkylating agents Busulfan Alkylsulfonate Chronic myelo- Alkylation producing MER; U; hyper- Pulm. fi brosis; Tc; Periodic blood counts for bone genous leukemia guanine–adenine pigmentation; EM leukopenia; anemia; marrow suppression cross-linking seizures Chlorambucil N mustards. Chronic lymphocytic Nonspecifi c DNA EM; TEN; CD; Ex; U; St Fever; interstitial pn; Fairly well tolerated. Type III Chloroalkylamine leukemia alkylating agents immune hemolytic immune complex reactions? anemia; bms;Tc Diagnosisa b c [email protected] Cyclophosphamide N mustard. Lymphomas, brain, Forms DNA cross-links U; Angio; rash; EM; SJS; Anaph; Br; bms Some IgE-mediated reactions ? Oxazophorine leukemia TEN; Vasc Diagnosis: SPT and IDT 1 mg/ml Mitotic inhibitors Docetaxeld,e Taxane (Diterpene) Breast, prostate, ovarian, Inhibition mitotic Acral erythema; Neutropenia; anemia; Cross-reaction with paclitaxel but nonsmall cell lung cell-division scleroderma- like; Dys; Br; GI symptoms; reactions less frequent. Infusion St; alopecia back pain reactions—C′ activation, IgE, mast cell/basophil actionf ? Paclitaxeld , e , g Taxane (Diterpene) Breast, ovarian, head Inhibition mitotic Acral erythema; injection Neutropenia; hypersensitiv- Incidence 10–16 % without and neck, lung, cell division site reaction; ity pn; GI symptoms; premedication. Risks—atopy, bee bladder, etc. AGEP-like; EM; SJS back pain; Dys sting allergy Platinum-based agentsh Carboplatin Organoplatinum Ovarian, lung, head Cross-links DNA Rash; Pr; U; Angio; Anaph; Dys; Br; CV Incidence 12 % (1 % < 6, 27 % > 7 and neck, breast, erythema; fl ushing and GI symptoms courses). Deaths reported. Diag. testicular, brain SPT and IDT. Some reactions (children) IgE-mediated? Cisplatin Metal coordination Sarcomas, some carcinomas Cross-links DNA Rash; fl ushing; Pr; U Anaph; Dys; Br Incidence 5–20 %. Repeated complex (small cell lung, ovarian), exposure before onset of symp- lymphomas, germ cell toms. Diagnosis: SPT and IDTi Oxaliplatin Organoplatinum Colorectal (admin. Cross-links DNA Pr; U; Angio; rash; Anaph; fever; Dys; GI Incidence 10–19 %. Reactions after with fl uorouracil erythema; fl ushing; symptoms; types II 7–9 courses. Diag. SPT and IDT. and leucovorin) acral erythema and III hypersensitivities Some IgE-med. reactions? (continued) Table 13.1 (continued) Type of Adverse/hypersensitivity reactions chemotherapy drug Incidence/risk factors/ and generic name Chemical class Cancer indications Mechanism of action Cutaneous Systemic diagnosis/comments Anthracycline antibiotics Daunorubicin Anthracycline Acute myeloid and Intercalates Rash; U; Angio Anaph; fever; cardiac Reactions uncommon (1–2 %). lymphocytic leukemias, with DNA toxicity; bms Cross-reaction with doxorubicin? neuroblastoma Doxorubicinj Anthracycline Hematological, ovarian, Intercalates Pr; U; Angio; acral Anaph; hypersens. infusion Reactions uncommon (U, 0.6–3 %). Kaposi’s with DNA erythema; rash; St; reaction; Br; cardiotox- Hypersensitivity reaction 8 % injection site reactions icity Tyrosine kinase inhibitorsk Imatinib mesylatel Phenylaminopyrimidine Chronic myeloid leukemia, Inhibits BCR- ABL Rash; Angio; Pr; SJS; Hypersensitivity pn; Incidence 31–44 % cutaneous derivative GI stromal tumors, tyrosine kinase AGEP; DRESS; Vasc neutropenia; tumor reactions. 21 % mild-moderate and other cancers lysis syndr; anemia; reactions. Severe exfoliative rash [email protected] hepatotoxicity 1–500 Proteosome inhibitor Bortezomib m Boronic acid derivative Multiple myeloma, B atom binds to 26S Erythematous plaques; Peripheral neuropathy; Rash 8–18 % of N -protected non-Hodgkin, mantle proteasome rashes; Vasc; TEN Neutropenia; Tc; Dys; dipeptide cell lymphoma cardiac disorders; Tc Antimetabolites: Pyrimidine analogs n : Capecitabine Derivative of Colorectal, pancreatic, Prodrug form of Acral erythema; CV; GI; hematologic; Not to be used with leucovorin 5-fl uorouracil metastatic breast, 5-fl uorouracil erythema; palmar- cytopenias stomach plantar keratoderma; exfol. dermatitis; St 5-Fluorouracil Pyrimidine Colorectal, pancreatic, Thymidylate Acral erythema; Anaph; diarrhea; Palmar-plantar dermatitis risk breast synthase maculopap. rash; inj. myelosuppression; GI; factor. Diag.—IDT and patch inhibitor site reaction; CD; St mucositis (with care) IgE-mediated? Direct cytotoxic effect Gemcitabine o Nucleoside analog Pancreatic, lung, ovarian, Replaces cytidine during Maculopap. rash; Fever; fl u-like symptoms; − breast, bladder DNA replication acral erythema; Pr; hypersensitivity pn; bullous dermatitis; SJS; diarrhea; cytopenias TEN; Vasc; St Antimetabolites: Anti-folates : Methotrexate p An amino- Breast, head, neck, lung, Inhibits dihydro Rash; acneiform papular Anaph; low white cell count; Types I q , II (hemolytic anemia), pteroylglutamic bladder, leukemia, folate reductase and nodular eruptions; Br; hem anemia; and III (cut. Vasc.) Diagnosis: SPT acid lymphoma acral erythema; EM; hepatotoxicity; GI and IDTq SJS; TEN; Vasc; St; U Pemetrexed Pyrrolopyrimidine Lung, mesothelioma Inhibits folate- dependent Rash; Pr; TEN; U; Vasc; Low blood count; diarrhea; Incidence of cutaneous reactions enzymes St; mucositis nausea and vomiting; Dys 14% r AGEP acute generalized exanthematous pustulosis; Anaph anaphylaxis, Angio angioedema, bms bone marrow suppression, Br bronchospasm, CD contact dermatitis, CV cardiovascular, DRESS drug reaction with eosinophilia and systemic symptoms; Dys , dyspnea; EM erythema multiform, Ex exanthema, GI gastrointestinal, MER macular erythematous rash, pn pneumonitis, Pr pruritus, SJS Stevens–Johnson syndrome, St stomatitis, Tc thrombocytopenia, TEN toxic epidermal necrolysis, U urticaria, Vasc vasculitis a Lymphocyte stimulation test positive in a few cases. Challenge test often positive but may be harmful b Potential cross-reactivity with other N mustards due to common bischloroethylamine group c Can form antigenic complexes with protein d Desensitization generally well tolerated and successful within 6–7 h (see text) e Premedication with oral corticosteroids often used (see Sect. 13.1.1 ) f Vehicle polysorbate 80 implicated in some reactions g Cremophor vehicle associated with some reactions involving C′ activation h Premedication and desensitization employed (see Sect. 13.2.4 ) i See Sect. 13.2.3 j Premedication with corticosteroids and antihistamines. Slow infusion rate (0.1–0.2 mg/min) required k See also gefi tinib and erlotinib in Sect. 13.3.3 [email protected] l For medication with prednisolone to achieve tolerance of therapeutic dosage and desensitization see Sect. 13.3.2 m Premedication with corticosteroids n Some patients unable to metabolize pyrimidine drugs (~8 % of population) may have dihydropyrimidine dehydrogenase (DPD) defi ciency o Structurally related to cytarabine p Often symptoms despite premedication. Desensitization sometimes employed q Over 20 reported cases of anaphylaxis. Skin prick test conc. 10 mg/ml r To prevent skin rashes, dexamethasone 4 mg b.i.d. the day before, the day of, and the day after administration 404 13 Drugs Used for Chemotherapy

13.1.1 Premedication for Taxanes

There is a signifi cant risk of hypersensitivity reactions following taxane administration and to minimize this, patients require premedication. Both docetaxel and paclitaxel are usually administered once every 3 weeks with docetaxel being infused over 1 h and paclitaxel over periods of from 1 to 96 h. The premedication regimen for patients receiving docetaxel consists of oral dexamethasone 8 mg twice daily for 3 days starting 24 h prior to the commencement of infusion. For paclitaxel infused over 1, 3, and 24 h, antihis- Fig. 13.1 Hand-foot skin reaction in a patient given sorafenib, a targeted inhibitor of some tyrosine kinases tamines are administered as well as the steroid. (including VEGFR and PDGFR) and Raf kinases The H1 antagonist diphenhydramine (50 mg) and approved for treatment of renal cell and hepatocellular carcinomas. Note that hand-foot skin reaction is a distinct an H2 antagonist (cimetidine 300 mg, ranitidine 50 mg, or famotidine 20 mg) are given IV prior to entity from hand-foot syndrome or acral erythema which manifests as erythema, swelling, and desquamation of the infusion beginning while oral dexamethasone palms and soles in cancer patients mainly during non- 20 mg is administered 12 and 6 h prior. There is targeted chemotherapy. Drugs most commonly implicated some evidence that premedication is not required in the latter reaction include 5-fl uorouracil, capecitabine, for infusions extending over 96 or more hours. cytarabine, pegylated doxorubicin, and also sorafenib and sunitinib, but a range of other chemotherapeutic agents The corticosteroid is used to prevent hypersensi- including docetaxel, paclitaxel, oxaliplatin, gemcitabine, tivity reactions, decrease fl uid retention, and and methotrexate are also known causes. Both sorafenib decrease skin and nail adverse effects. Note that and sunitinib provoke a high incidence of hand-foot skin the dose-limiting toxicity of docetaxel and pacli- reaction in patients (Reproduced with permission from Hauschild A, Kähler KC, Egberts F in Leong, SPL, editor. −2 taxel is febrile neutropenia; docetaxel 100 mg m From Local Invasion to Metastatic Cancer, 1st ed. New administered over 1 h every 3 weeks is associated York: Springer, 2009) with neutropenia in ~75 % of patients; for paclitaxel 135–300 mg m −2 over 24 h the fi gure is whether or not doses of dexamethasone can be 52 %. Another published premedication protocol reduced is important for patients experiencing, or that claimed to limit the development of acute at high risk of, steroid-induced side effects. Some hypersensitivity to docetaxel consists of oral have claimed that the optimal schedules remain methylprednisolone 32 mg, cetirizine 10 mg, and to be determined and larger prospective clinical ketotifen 1 mg, 12 and 3 h before infusion. For trials are needed. taxanes given weekly, the effi cacy and safety of A recent multi-institution survey in Japan premedication were assessed in the treatment of drew attention to an important effect of histamine nonsmall cell lung cancer. Weekly administra- H2 antagonists on docetaxel-induced skin toxic- tions of docetaxel and paclitaxel were assessed as ity. Analyses revealed that administration of H2 safe and active protocols. For docetaxel, oral blockers was associated with a signifi cantly dexamethasone 4.5–7.5 mg twice daily for the higher incidence of acral erythema (hand-foot day before, the day of, and the day after, together syndrome; palmar-plantar erythrodysesthesia; with IM promethazine 25 mg and IV cimetidine compare with hand-foot skin reaction, 600 mg 30 min before docetaxel are recom- Sects. 13.3.2 and 13.3.3 and see Fig. 13.1 ) and mended. For paclitaxel the recommended proto- facial erythema. Steroids and H 2 blockers affect col is: dexamethasone 2.25–7.5 mg orally 12 and the metabolism of docetaxel by cytochrome

2 h before, and promethazine 25 mg IM and P450 3A4 (CYP3A4), but dexamethasone dosage cimetidine 600 mg IV 30 min before the taxane. did not change the incidence of hand-foot For weekly taxane dosage, the question of syndrome or facial edema.

[email protected] 13.2 Organoplatinum Chemotherapeutic Drugs 405

13.1.2 Desensitization and almost all of the common tumors except for Hypersensitivity breast and prostate tumors. Treatments with plat- Reactions to Taxanes inum drugs is often effected in combination with other anticancer agents and, overall, it is esti- In the rapid desensitization protocol for pacli- mated that up to 50–70 % of cancer patients are taxel published by Sullivan (Sullivan TJ. Protocols treated with platinum drugs. Cisplatin, the fi rst of for rapid and slow drug allergy desensitization, the so-called organoplatinum drugs, contains no Atlanta, Georgia, 2009), three concentrations of organic component and is, in fact, a metal coordi- drug are employed—full strength solution, that is nation compound with a square-planar platinum 300 mg paclitaxel in 500 ml in physiological (II) center coordinated to two ammonia and two saline (0.6 mg/ml); a 1 in 10 dilution (0.06 mg/ml); chlorine ligands in a cis -ligand conformation and a 1 in 100 dilution (0.006 mg/ml). Except for (Fig. 13.2 ). The success of cisplatin in the clinic the last dosage step, each of the 11 preceding focused research interest on other possible plati- steps is infused for 15 min before changing to num chemotherapeutic drugs and led to the gen- the next dose. Beginning with the 1 in 100 dilu- eral rule that a neutral square-planar platinum (II) tion (0.006 mg paclitaxel/ml) and an infusion rate center containing two cis -amines and two leaving of 2 ml/h, the solution is infused for 15 min groups are required for good anticancer activity. followed by infusion rates of 5, 10, and 20 ml/h, The fi rst follow-up drug to gain worldwide respectively for steps two, three, and four. For the clinical acceptance was carboplatin that contains next four steps (numbers fi ve to eight, again at the cis -Pt(NH3 )2 active group of cisplatin with the 15 min intervals), the one in ten dilution of drug chloride leaving groups replaced by a bidentate (0.06 mg/ml paclitaxel) is infused at rates of 5, 10, dicarboxylate (Fig. 13.2 ). Carboplatin has a 20, and 40 ml/h, respectively. At steps 9, 10, and similar anticancer profi le as cisplatin but, 11, full strength solution (0.6 mg paclitaxel/ml) is although it is just as effective for ovarian cancer, infused at 10, 20, and 40 ml/h, respectively. For its potencies against head, neck, and testicular the last step (step 12), full strength solution is cancers are less. On the other hand, carboplatin infused at 80 ml/h until the remaining full strength evokes fewer side effects, and this has generally solution has been given. made it the drug of choice over cisplatin. In 2004 The Castells group treated 17 consecutive oxaliplatin became the third widely accepted and patients with hypersensitivity to taxanes using a used platinum drug. Oxaliplatin, unlike the other standard 6–7 h desensitization protocol. The two platinum chemotherapeutics, is effective patients underwent a total of 77 rapid desensitiza- against colorectal cancer and, in addition, it is tions to docetaxel or paclitexal. Seventy two of active against some cisplatin-resistant cancers. the procedures were tolerated without reactions, In the structure of oxaliplatin, the amines are four patients responded with hypersensitivity part of the 1,2-diaminocyclohexane framework reactions that were milder than their initial reac- (Fig. 13.2 ). tions, and these patients tolerated re-administration of infusions. Five patients rechallenged before desensitization experienced recurrent reactions 13.2.1 Symptoms even though they were given additional premedi- of Hypersensitivity cation, and the infusion rate was reduced. to Platinum Drugs

Exposure to platinum salts, especially in miners 13.2 Organoplatinum and other industrial workers, has been known to Chemotherapeutic Drugs provoke hypersensitivity reactions since, at least, the 1940s while reactions to platinum therapeutic Platinum-based cytotoxic compounds that cross- agents, viz. cisplatin, were ﬁ rst described in the link DNA are some of the most active and effec- 1970s. Symptoms to the drugs may develop tive cytotoxic drugs for the treatment of ovarian during infusion within minutes or after hours or

[email protected] 406 13 Drugs Used for Chemotherapy

H3N Cl

Cisplatin Pt

H3NCl

O H3N O Carboplatin Pt

H3N O O

H2 N O O Oxaliplatin Pt

N O O H2

Fig. 13.2 Two- and three-dimensional structures of plati- of cisplatin with the chloride leaving groups replaced by a num-based chemotherapeutic drugs. Cisplatin, the fi rst of bidentate dicarboxylate. Oxaliplatin also has the square the so-called organoplatinum drugs, contains no organic planar platinum center but, unlike cisplatin and carbopla- component. It is a metal coordination compound with a tin, it has the bidentate ligand 1,2-diaminocyclohexane square-planar platinum (II) center coordinated to two instead of two monodentate ammine ligands as well as a ammonia and two chlorine ligands in a cis -ligand confor- bidentate oxalate group mation. Carboplatin contains the cis -Pt(NH 3 ) 2 active group days with a mild rash being the fi rst manifesta- (Table 13.1). Other more severe symptoms tion. The clinical features of the reactions are sometimes reported are bronchospasm, chest highly variable—in one survey of carboplatin- pain, seizures, and systemic anaphylaxis that induced hypersensitivity, 100 % of patients had may be life-threatening. Reactions to oxaliplatin cutaneous symptoms (mainly palmar or facial are similar to those seen in response to cispla- fl ushing), 57 % had cardiovascular symptoms, tin and carboplatin, but the responses to oxali- 42 % gastrointestinal disturbances, and 40 % platin tend to be more heterogeneous and respiratory symptoms. Commonly seen mild unpredictable with fewer cutaneous reactions; reactions are rash, urticaria, fl ushing, palmar idiosyncratic reactions like cytokine release syn- itching, a burning feeling, hand and facial edema, drome and pulmonary fi brosis; fewer reports of pruritus, back pain, abdominal cramping, and severe anaphylaxis; and a higher incidence of diarrhea. These symptoms usually resolve quickly respiratory symptoms including laryngeal spasms with antihistamines and steroids. Moderate to and hypoxemia. A few cases of type II thrombo- severe symptoms include diffuse erythroderma, cytopenia and type III immune complex-mediated tachycardia, chest tightness, wheezing, facial urticaria, joint pain, and proteinuria associated swelling, dyspnea, hypertension, or hypotension with oxaliplatin have also been reported.

[email protected] 13.2 Organoplatinum Chemotherapeutic Drugs 407

13.2.2 Incidences of, and Risk 13.2.3 Mechanisms and Diagnosis Factors for, Hypersensitivity of Platinum Drug-Induced Reactions to Platinum Drugs Hypersensitivity Reactions

For cisplatin, the overall incidence of reactions Reactions are thought to be mainly type I or type is 5–20 %, reactions occur within minutes of IV hypersensitivity responses but, as mentioned the commencement of infusion, most reactions above, a few cases of type II and type III hyper- occur between the fourth and eighth course, and sensitivities have been reported. Nevertheless, reactions increase with concomitant radiation. the complexity and unpredictable nature of many The overall incidence of reactions to carbopl- responses suggests that a number of mechanisms, atin has been reported as 1–44 % and, in another both immune and nonimmune, may be opera- study, 9–27 %. Reactions occur within minutes tive in many reactions. A number of authors are or days of infusion; less than 1 % of hypersen- sure that many reactions to the platinum drugs sitivity reactions result during cycles 1–5; are consistent with type I, IgE antibody-mediated 6.5 % occur during cycle six; 27 % are seen in hypersensitivity. Although there has been no cycle seven or more; and 44 % occur in third- clear and unequivocal demonstration of the exis- line treatment. Approximately half of all reac- tence of IgE antibodies specifi cally directed at tions to carboplatin are moderate to severe. cisplatin, carboplatin, or oxaliplatin (including Reactions to oxaliplatin occur with an inci- specifi c inhibition of antibody binding by the dence of 10–19 % and manifest within minutes drug and structural analogs), previous fi ndings of or hours of infusion. Again, most reactions IgE antibodies to platinum salts in platinum- appear after a number of treatment cycles exposed workers, the appearance of sensitization (usually at least six). only after multiple infusions, positive skin tests Risk factors for the platinum chemotherapeu- to carboplatin, and anaphylactic reactions are all tic drugs have not yet been thoroughly studied taken as evidence of a type I hypersensitivity and well defi ned. Most information so far has mechanism. Skin testing with the three platinum been obtained for carboplatin. Apart from the drugs has been employed to identify at-risk already clearly established risks of the number of patients and predict platinum hypersensitivity but prior treatments with platinum drugs and a high as yet it has not found widespread acceptance and rate of drug infusion, other suggested risk factors application as a routine diagnostic procedure. In a so far include a history of drug allergy, a study of patients with recurrent ovarian or perito- carboplatin-free interval of greater than 13 neal carcinoma who had received more than months, patients with ovarian cancer, children seven courses of carboplatin, intradermal testing receiving weekly carboplatin infusion rather than with 100–240 μg of carboplatin revealed 13 of 47 monthly infusions, and the female gender. The patients (28 %) with a positive test. A negative antineoplastic drug used in combination with the skin test correctly predicted the absence of a platinum drug can also infl uence the incidence of hypersensitivity reaction in 166 of 168 courses of hypersensitivity reactions to the platinum agent. chemotherapy (98.8 %). Two patients experienced For example, the CALYPSO study of the a reaction after showing a negative skin test. Gynecologic Cancer Intergroup showed that A follow-up study on 126 patients confi rmed the carboplatin with pegylated liposomal doxorubicin association between a negative carboplatin skin produced fewer reactions than the combination of test and no resultant severe hypersensitivity reac- the platinum drug with paclitaxel. Risk factors tion after the next infusion, but the implications identifi ed so far for oxaliplatin reactions include of a positive test remained less certain. Patch, prick, a young age, female gender, and use of the drug and intradermal tests with cisplatin, carboplatin, as salvage therapy. and oxaliplatin on 21 patients produced negative

[email protected] 408 13 Drugs Used for Chemotherapy patch test results in all 21 patients, fi ve positive tive patients. Skin testing may also help in ruling reactions in the prick test, and 12 positives in the out cross-reacting drugs when substituting one intradermal test. Cross-reactions were observed platinum drug for another. Nevertheless, it has in four cases, and delayed reactions occurred in been argued that it is not practical to employ skin three patients. It was concluded that the intra- testing as a routine test in everyday clinical prac- dermal test was superior to the other two tests tice since prior experience in its execution is and its good negative predictive value allows safe required, and hypersensitivity reactions might re-treatment by detecting potential cross-reactions. still occur even in the case of a negative skin test. Results from some other studies also support the The fi rst of these objections cannot be sus- superiority of the intradermal test. In response to tained—if the simple skills of skin testing are the report of the three delayed reactions, it was lacking, the situation can, and should, be readily pointed out that intradermal tests with platinum rectifi ed by professionals who are charged with drugs have a good negative predictive value in the responsibility of seeking and maintaining the immediate reactions, but the test can induce false best available diagnostic procedures. The possi- positives with these drugs. Most skin test investi- bility of reactions in skin test-negative patients is gations have been on patients with carboplatin, indeed likely but that, again, should always be but results and conclusions have not always been seen as a possibility to be anticipated and man- in agreement. Fifty three of sixty patients referred aged and such a possibility is not a suffi cient for carboplatin hypersensitivity were skin test reason to forego the possible advantages that skin positive to the drug, one patient showed a delayed testing can provide. positive reaction, two became positive after further infusions, and the remaining four skin test- negative patients experienced hypersensitivity 13.2.4 Desensitization during a subsequent infusion. Skin tests on 54 patients receiving re-treatment with carboplatin Attempts to readminister the same drug or change predicted hypersensitivity reactions in only 64 % to a different platinum drug can be dangerous, of affected patients leaving the authors to con- and desensitization to the drug is sometimes clude that skin testing did not reliably predict considered to be the best option. Desensitization carboplatin-induced reactions. Skin test concen- protocols to the platinum drugs are not standard- trations generally employed for carboplatin are ized, and a number of different protocols are 10 mg/ml in the prick test and 0.02 ml of 0.1 mg/ currently employed in different institutions. ml in the intradermal test increasing in tenfold Table 13.2 sets out a 12 step, ~ 6 h desensitization concentration steps to a maximum of 10 mg/ml. protocol for carboplatin in skin test-positive In some studies, a maximum skin test concentra- patients hypersensitive to the drug and receiving tion of 3 mg/ml has been used for carboplatin. desensitization for the fi rst time. Conversion of a For oxaliplatin, a concentration of 1 mg/ml is positive skin test to carboplatin to a negative used in the prick test and a maximum of 0.1 mg/ response after desensitization supports the ml in the intradermal test. existence of a specifi c IgE antibody-mediated In summary, it has been claimed that skin tests response in patients. As well as carboplatin, a are positive in more than 80 % of the platinum number of different desensitization protocols for drug-treated patients tested and, when the test is oxaliplatin have been published, but the number negative, the risk of a hypersensitivity reaction is of patients treated so far is small. The essential reduced sevenfold or even eliminated. This, in procedures of premedication, escalating dosage, turn, has led to the recommendation that skin infusion times, and duration of the procedure are testing should be performed on every patient similar to those employed for carboplatin. Drugs before the eighth drug infusion. Skin testing for used for premedication are generally diphen- oxaliplatin sensitivity has been claimed to be hydramine or hydroxyzine as histamine H1 inhib- positive in 75–100 % of oxaliplatin hypersensi- itors; famotidine, cimetidine, or ranitidine as H2

[email protected] 13.3 Tyrosine Kinase Inhibitors 409

Table 13.2 Desensitization protocola for carboplatin hypersensitivity in patients undergoing desensitization for the ﬁ rst time Step Infusion rate (ml/h)b Time infused (min) Administered dose (mg) Cumulative dose infused (mg) 1 2 15 0.010 0.010 2 5 15 0.025 0.035 3 10 15 0.050 0.085 4 20 15 0.100 0.185 5 5 15 0.250 0.435 6 10 15 0.500 0.935 7 20 15 1.000 1.935 8 40 15 2.000 3.935 9 10 15 5.000 8.935 10 20 15 10.000 18.935 11 40 15 20.000 38.935 12c 75c 184.4 461.065 500.00 Totals: 5 h 49.4 min 500 mg Data from Lee C-W et al. Gynecol Oncol 2004;95:370 a Conducted in intensive care unit, Brigham and Women’s Hospital. β-Blockers withheld 1 day before. Informed consent obtained. All rescue medications on hand. Patients premedicated with diphenhydramine 25 mg, famotidine 20 mg IV 30 min before initiation of infusion b Using appropriate concentrations to deliver the required dose in the required time c A constant rate of infusion maintained to deliver the remainder of the total carboplatin dose antagonists; and corticosteroids such as dexa- desired, but their arrival (in the form of tyrosine methasone, prednisolone, and hydrocortisone. kinase inhibition strategies), has only recently Oxaliplatin dilutions employed cover the range seemed likely. Tyrosine kinases have become from 1:100,000 to 1:1 in from 5 to 13 steps over recognized as important targets for cancer a total time range of from 5.8 to 8 h. Some proto- therapy because they have an important role in cols employ continuous ﬁ xed rate infusion the modulation of growth factor signaling, and extending over 24–48 h. In some procedures, some tyrosine kinase inhibitors have been found magnesium sulfate and calcium carbonate have to have antitumor activity and to be effective in been added and claimed to increase the success treating some cancers in the clinic. This targeted rate of desensitization. strategy for killing tumor cells and some of the important drugs developed to achieve it will be discussed. 13.3 Tyrosine Kinase Inhibitors

Until relatively recently, chemotherapy has not 13.3.1 The Philadelphia discriminated effectively between rapidly divid- Chromosome and Tyrosine ing tumor cells and normal cells, in particular Kinases cells in the bone marrow and gastrointestinal tract. This has led to toxic effects for patients The Philadelphia translocation t(9;22)(q34;q11) including an array of severe nonimmune adverse or Philadelphia chromosome is a chromosomal reactions as well as hypersensitivities ranging defect resulting in gene fusion of the BCR and from mild skin manifestations to life-threatening ABL genes. The BCR (b reakpoint c luster r egion) cutaneous and systemic reactions. Effective gene is on chromosome 22 (region q11) and the targeted therapies with high speciﬁ city toward ABL (so named because the Ab e l son leukemia tumor cells but broad therapeutic application virus has a similar protein) tyrosine kinase gene and absence of toxicity have always been is on chromosome 9 (region q34). The resultant

[email protected] 410 13 Drugs Used for Chemotherapy fusion gene is the BCR-ABL oncogene. The N Philadelphia chromosome is a cytogenetic abnor- HN N N mality seen in 95 % of chronic myeloid leukemia H patients and 15–30 % of adults with acute lym- N O N phoblastic leukemia. N Tyrosine kinases can be classifi ed as receptor Imatinib and non-receptor kinases. They are enzymes that F catalyze the transfer of the γ-phosphate group from adenosine triphosphate to the hydroxyl O HN Cl group of tyrosine on signal transduction mole- NO N cules. This is an important activating step that leads to increases in tumor cell proliferation and ON growth. The oncogene BCR-ABL results in the Gefitinib expression of two forms of tyrosine kinases and a large increase in myeloid cell numbers. In sum- O N mary, the BCR-ABL mutation is present in the O O N great majority of chronic myeloid leukemia O H patients, the Bcr-abl fusion protein is unique to C HN C leukemic cells, it is expressed in high levels, and its tyrosine kinase activity is essential in the Erlotinib induction of leukemia. However, although tyrosine kinases were implicated as oncogenes in Fig. 13.3 Structures of the tyrosine kinase inhibitors: some animal tumors induced by retroviruses imatinib, a phenylaminopyrimidine derivative, and the 4-aminoquinazoline derivatives, gefi tinib and erlotinib. more than 30 years ago, it was not until the intro- Imatinib is marketed as the mesylate salt duction of the drug imatinib that inhibits both the ABL and BCR-ABL tyrosine kinases and which has been successful in treating chronic myeloid 33–81, colorectal 25–100, pancreatic 30–50, leukemia that tyrosine kinases were regarded as ovarian 35–70, breast 15–37, prostate 40–90, and good targets for cancer chemotherapy. In addition glioma 40–92 % to imatinib, some inhibitors of receptor tyrosine kinases targeting epidermal growth factor receptor (EGFR; ErbB1; HER1; a member of the ErbB 13.3.2 Imatinib Mesylate family of receptors), vascular endothelial growth factor receptors (VEGFR), and platelet-derived Used for the treatment of chronic myeloid leuke- growth factor receptors (PDGFR) have been mia, unresectable or metastatic gastrointestinal found to have antitumor and/or other activities. stromal tumors, and some other cancers, imatinib These receptor tyrosine kinase inhibitors include mesylate, a phenylaminopyrimidine derivative gefi tinib, erlotinib (both inhibitors of EGFR), (Fig. 13.3 ), and some other targeted tyrosine lapatinib (inhibits ErbB1 and ErbB2), vatalanib kinase inhibitors are generally well tolerated with less (inhibits VEGFR-1 and VEGFR-2), semaxinib severe systemic effects, especially when compared (inhibits VEGFR-2), and lefl unomide (inhibits to most cytotoxic chemotherapies. Dermatologic PDGFR-mediated cell signaling, i.e., phosphory- reactions such as papulopustular rash, hand-foot lation). Elevated EGFR tyrosine kinase activity is skin reaction (Fig. 13.1 ), xerosis, pruritus, and found in most solid tumors; following is a list of mouth, hair, scalp, and nail abnormalities are the the percentage expression of EGFR by some of main adverse responses to many of the targeted the most common human cancers: Nonsmall cell drugs including gefi tinib, lapatinib, erlotinib, lung cancer 40–80, head and neck 80–100, gastric regorafenib, pazopanib, sorafenib, and sunitinib.

[email protected] 13.3 Tyrosine Kinase Inhibitors 411

Imatinib is marketed as the mesylate, that is, the 13.3.3 Gefi tinib and Erlotinib salt of methanesulfonic acid (CH3 SO 3 H). The most common non-hematologic adverse reactions Gefi tinib and erlotinib are derivatives of to the drug include superfi cial edema, especially 4-aminoquinazoline (Fig. 13.3 ). Both drugs are periorbital edema, nausea, vomiting, diarrhea, EGFR inhibitors, inhibiting the receptor’s tyro- muscle cramps, myalgia, arthralgia, fatigue, sine kinase domain by binding to the ATP- abdominal pain, headache, and, most common of binding site of the enzyme. Approved by the all, cutaneous reactions. Patients receiving stan- FDA in 2003, gefi tinib is indicated for locally dard dose imatinib therapy in the chronic phase of advanced or metastatic nonsmall cell lung cancer chronic myeloid leukemia experience neutropenia with activated mutations of EGFR tyrosine in 35–45 % of cases, thrombocytopenia in 20 % kinase. EGFR is over-expressed by the cells of of cases, and anemia in 10 % of cases. Although some cancers such as lung and breast leading to most cutaneous reactions are mild and dose uncontrolled cell proliferation, the blocking of dependent, severe reactions such as Stevens– apoptosis, and increased production of angio- Johnson syndrome (SJS), exfoliative dermatitis, genic factors and metastasis. The mutations also toxic epidermal necrolysis (TEN), and drug reac- incur increased sensitivity to tyrosine kinase tion with eosinophilia and systemic symptoms inhibitors like gefi tinib, but no clinically benefi - (DRESS) have been reported. Other severe reac- cial activity of the drug is shown in patients with tions to the drug include rare cases of acute gen- EGFR-negative tumors. The most frequent eralized exanthematous pustulosis (AGEP), adverse reactions to gefi tinib, that is reactions nearly 20 cases of lichenoid drug eruptions of occurring in more than 20 % of patients, are diar- the skin or oral mucosa, vasculitis, pityriasis- rhea and skin reactions. Reactions may be cate- rosea-like eruption, palmoplantar hyperkerato- gorized by the affected organ: skin reactions like sis, and exacerbation of psoriasis. For mild to hand-foot skin reaction, pruritus, erythema, and moderate rashes, antihistamines or, if necessary, papulopustular rash are common as are nail dis- topical or short course corticosteroids can be orders while bullous eruptions (erythema multi- used. For severe skin reactions, the following forme, SJS, and TEN) are rare. Note that management schedule has proved effective for hand-foot skin reaction should not be confused achieving tolerance of therapeutic dosages of with hand-foot syndrome or acral erythema seen imatinib even after severe cutaneous reactions to during the administration of some cytotoxic the drug: prednisolone 1 mg/kg per day, tapered anticancer drugs such as 5-fl uorouracil and doxo- to 20 mg per day over several weeks along with rubicin. The former reaction is distinguished by the gradual reintroduction of imatinib (100 mg localized blisters or hyperkeratosis whereas per day increased by 100 mg per week) given as hand-foot syndrome shows diffuse, symmetrical the prednisolone dose is tapered off. This, of erythematous, and edematous lesions on the course, is only continued with if the skin mani- palms and soles. Eye disorders (conjuctivitis, festations do not recur. Oral desensitization to blepharitis), gastrointestinal disorders, vascular imatinib in patients with recurrent rash induced effects (hemorrhage), and renal and urinary by the drug has been reported. Ten patients were disorders are also common. Interstitial lung dis- subjected to a 4 h dosage procedure beginning ease has been found to occur in 1.3 % of patients, with a dose of 10 ng followed by increases every often of severe grade and occasionally fatal. 15 min. Four patients experienced no recurrence Erlotinib is an EGFR type 1 receptor (HER1/ of rash after desensitization, four had recurrent EFGR) tyrosine kinase inhibitor. These receptors rash that resolved after corticosteroid/antihis- are involved in the control of cell divisions and tamine administration, and two patients each proliferation and by inhibiting their functions; developed a rash and were unable to resume erlotinib limits tumor cell division and metastasis imatinib therapy. and may even help in initiating apoptitic cell death.

[email protected] 412 13 Drugs Used for Chemotherapy

A randomized, placebo-controlled, double-blind proteasome that are mediated by three β-subunits trial carried out by a National Cancer Institute of in the core: β2 trypsin-like, β5 chymotrypsin- Canada Clinical Trials Group revealed that the like, and β1 caspase-like activities. Proteolytic main adverse responses to erlotinib were rash, activity of each of these subunits is associated fatigue, anorexia, diarrhea, nausea, ocular effects, with the N -terminal threonine residues in peptide infection, vomiting, and stomatitis. Rash and bond hydrolysis. Cancer cells show higher pro- diarrhea were the main reasons for dose reduc- teasome activity than normal cells. Inhibition of tion and interruption of treatment. In a 2009 proteasome function leads to intracellular accu- warning, the FDA referred to rare serious gastro- mulation of unwanted proteins and ultimately intestinal, skin, and ocular disorders in some cell death and, here too, cancer cells are more patients taking erlotinib. As with other tyrosine sensitive to the apoptosis-promoting effects of kinase inhibitors, papulopustular rash, hand-foot inhibition than normal cells. Proteasome inhibi- skin reaction, and pigmentary changes are tors can induce apoptosis in leukemia- and commonly seen. Serious eye conditions include lymphoma-derived cells without causing the corneal lesions, some patients develop gastroin- death of non-transformed cells. Multiple testinal perforations, and rare bullous and exfoli- myeloma cells synthesize and secrete large ative skin reactions, some leading to death, have amounts of immunoglobulin, and this high rate of occurred. biosynthesis is thought to increase the sensitivity of the synthesized proteins to proteasome inhibitors by, for example, inducing the immunoglobu- 13.4 Proteasome Inhibitors lins into the unfolded state.

13.4.1 The Proteasome 13.4.2 Bortezomib The ubiquitin-proteasome system has a central role in the turnover of proteins—in the regulation Most proteasome inhibitors are short peptides of cellular proteins involved in growth and sur- that serve as protein substrates in the proteasome vival and in the destruction of defective proteins. 20S core where they target the active site threo- The proteasome consists of a hollow cylindrical nine residues. Bortezomib, an N -protected dipep- or barrel-like 20S (0.7 MDa) proteolytic core tide that contains a boron atom (Fig. 13.5 ), was capped at one or both ends by a 19S (0.9 MDa) the fi rst proteasome inhibitor to be introduced regulatory particle or activator. These structures into the clinic and is approved for treating make up the single-capped proteasome complex relapsed multiple myeloma and mantle cell lym- or 30S double-capped form (Fig. 13.4 ). Note that phoma. The drug inhibits proteasomes by bind- the enzymically active double-capped protea- ing with high affi nity via the boron atom to the some complex, which is thought to be the func- β-subunit chymotrypsin- and caspase-like pro- tional unit in the cell, is usually referred to as the teolytic catalytic sites. It has little effect on the 26S (2.5 MDa) proteasome even though physico- trypsin-like activity. Apoptosis is normally sup- chemical analysis has revealed that the correct pressed in mantle cell lines and myeloma cells, sedimentation coeffi cient is ~30S. The ~26S but proteasome inhibition may overcome this form probably represents the single-capped pro- suppression and activate cell death. Bortezomib teolytic core. Proteins that are defective in some suppresses tumor growth and spread and angio- way, for example, due to aging, incorrect folding, genesis through multiple mechanisms and, in etc. are tagged by ubiquitin and directed to the addition to directly inducing apoptosis of tumor proteasome for degradation via the endoplasmic cells, it mediates a myriad of biological effects reticulum degradation pathway. Cell-cycle pro- including reduced adherence of myeloma cells to gression is dependent on the ubiquitin-proteasome bone marrow cells, prevention of IL-6 production pathway with its three proteolytic activities in the and signaling in myeloma cells, interference with

[email protected] 13.4 Proteasome Inhibitors 413

Ubiquitinated proteins

Lid Recognizes ubiquitinated proteins & Arm transfers them to the 20S catalytic core Base PARTICLE 0.9MDa 0.9MDa PARTICLE 19S REGULATORY 19S REGULATORY Gated channel

Proteins enter α-ring the barrel though gate

β-ring Region of proteolytic activity Chymotrypsin-like β-ring Trypsin-like Caspase-like 20S HOLLOW 20S HOLLOW CORE 0.7 MDa 0.7 CORE PROTEOLYTIC PROTEOLYTIC

SINGLE-CAPPED PROTEASOME ~1.7 MDa α-ring DOUBLE-CAPPED PROTEASOME ~2.6 MDa ~2.6 PROTEASOME DOUBLE-CAPPED PARTICLE 0.9MDa PARTICLE 0.9MDa 19S REGULATORY 19S REGULATORY

Free ubiquitin Degraded proteins

Fig. 13.4 Diagrammatic representation of the proteasome composed of a base (dark blue ), lid (yellow-brown ), and so- and its role in protein degradation via the ubiquitin- called arm (pink ). The 20S proteolytic core may be capped at proteasome pathway. After being tagged with ubiquitin and one or both ends by a 19S regulatory particle. Proteasomes unfolded for degradation on the 19S regulatory particles are thus referred to as single-capped (sedimentation coefﬁ - which aid the opening of a proteolytic gate in the 20S core, cient ~26S) or double-capped (~30S). In the literature, the proteins are degraded into small peptides in the barrel- term 26S proteasome is often used incorrectly when refer- shaped core where β1 caspase-, β2 trypsin-, and β5 chymo- ring to the double-capped form. The double-capped complex trypsin-like activities reside. Regulatory particles are is thought to be the functional proteasome unit in the cell the production of pro-angiogenic factors, and a stimulant in attempts to understand the molecu- suppression of nuclear-factor-κ-light chain- lar mechanisms underlying its clinical effective- enhancer (NF-κB). Bortezomib gained regula- ness and identifying new drugs acting on the tory approval in 2003, and its success has proved same pathway.

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O OH H N N B N OH H O N Bortezomib O O OH Cl O O H N B N O H OH O O Cl Fig. 13.6 An example of Sweet’s syndrome, sometimes MLN9708 termed acute febrile neutrophilic dermatosis. The erythematous plaques that are characteristic of the condition and associated with fever and neutrophilic leukocytosis, O O are painful, and may occur on almost any part of the body. O H H Most cases of Sweet’s syndrome are idiopathic and N N N N N although drugs rarely induce the reaction, besides bort- H H ezomib, at least 25 other drugs have been associated with O O O O the induction of the syndrome. (Reproduced with permission from Fam AG in Klippel JH, Stone JH, Crofford LeJ, White PH, editors. Primer on the Rheumatic Diseases, Carfilzomib 13th ed. New York: Springer, 2008)

OH OH numerous. Rash (often pruritic) is frequently H O H O reported in more than 10 % of patients (an inci- N N O O dence of 8–18 % has been stated) and pruritus, O O erythema, urticaria, periorbital edema, and eczema are commonly seen. Bortezomib has been associated with a few cases of drug-induced Sweet’s Cl syndrome (Fig. 13.6) or acute febrile neutrophilic Clastolactacystin b-lactone Salinosporamide A (Omuralide) (Marizomib) dermatosis, a rare variant of this uncommon skin disease characterized by fever, an elevated neutro- Fig. 13.5 Proteasome inhibitors used (or intended) for phil count, and erythematous lesions infi ltrated by the treatment of relapsed multiple myeloma, mantle cell neutrophils. Histological examination of a bort- lymphoma, and some other tumors. Structures of the pep- ezomib-induced skin eruption showed a clinical tide boronates bortezomib and the orally active MLN9708, the peptide epoxyketone carfi lzomib, and the γ-lactam-β- picture similar to Sweet’s syndrome but which lactone bicyclics salinosporamide A (marizomib) and differed by the presence of a signifi cant number of omuralide (β-clastolactacystin), both derived from natural CD30+ lymphocytes. The presence of these cells, sources. The drugs inhibit normal proteasome action by which are seen during some treatments of blood binding to the β-subunit proteolytic sites in the 20S core (see Fig. 13.4 ) malignancies, is not understood. Subcutaneous infusion of bortezomib as an alternative to intravenous administration was recently approved by the Gastrointestinal symptoms, thrombocytopenia, US Food and Drug Administration (FDA). This peripheral neuropathy, and neuropathic pain are has proved a more convenient and less toxic route the most common side effects of bortezomib of administration and seems likely to become the and adverse cutaneous reactions to the drug are standard form of the drug’s delivery.

[email protected] Summary 415

13.4.3 Second Generation lactacystin isolated from Streptomyces spp. Proteasome Inhibitors Lactacystin inhibits trypsin- and chymotrypsin- like activity of the 20S proteasome. Omuralide Like bortezomib, MLN9708 is also a peptide inhibits cell cycle progression in several tumor boronate (Fig. 13.5 ) but it is orally active, shows cell lines. greater tissue penetration, and has a shorter half- life. The drug is primarily an inhibitor of the chymotrypsin-like activity of the 20S proteasome 13.5 Cytokine-release and tumor core and, like bortezomib, it inhibits NF-κB acti- lysis syndromes vation and has antitumor activity in multiple myeloma and some other hematologic malignan- Cytokine-release syndrome (CRS), also called cies. Besides peptide boronates like bortezomib, acute infusion reaction, is not a true hypersensi- other synthetic compounds tested as proteasome tivity reaction but the two share some signs and inhibitors include peptide aldehydes, peptide symptoms such as nausea, fever, cough, dyspnea, epoxyketones, and peptide vinyl sulfones. bronchospasm, hypotension, rash, itching, and Carfi lzomib , a peptide epoxyketone (Fig. 13.5 ) urticaria. CRS is usually short term, developing irreversibly binds to and inhibits chymotryptase during or soon after drug infusion followed by activity but has less activity toward the other two resolution within 24 hours. Drug-induced ezymatic actions. The drug was approved by the destruction of cells is thought to release cyto- FDA in 2012 for patients with relapsed and kines such as TNF and interleukins producing refractory multiple myeloma. It leads to cell symptoms similar to type I hypersensitivity. cycle arrest and induces apoptosis in multiple Within 48-72 hours of initiating cancer ther- myeloma, other hematologic malignancies, and apy, large numbers of tumor cells may be some solid tumors. A potentially very important destroyed in a short time releasing intracellular property is the drug’s activity against primary contents and producing ionic imbalances in cal- multiple myeloma cells and cell lines resistant to cium, phosphate, potassium, and uric acid. This bortezomib. condition, termed tumor lysis syndrome (TLS), Adverse reactions noted so far include pulmo- may progress to acute renal failure, cardiac nary hypertension, dyspnea, cardiac disorders, arrhythmias, seizures, and death. Unlike CRS, cytopenias, infusion reactions, tumor lysis syn- TLS is easy to distinguish from drug-induced drome, hepatotoxicity, peripheral neuropathy, hypersensitivity. Drugs causing TLS include rash, and urticaria. etoposide, fl udarabine, hydroxyurea, paclitaxel, The proteasome inhibitors salinosporamide A thalidomide, and bortezomib. and omuralide are both γ-lactam-β-lactone bicyclic compounds (Fig. 13.5 ) derived from natural sources. Salinosporamide A, also known as Summary marizomib, is obtained from Salinospora tropica, a bacterium found in ocean sediments. The drug • In excess of 100 drugs are currently adminis- is an irreversible proteasome inhibitor that tered in signifi cant quantities to cancer patients shows little effect on the caspase-like activity but somewhere in the world. Many of the drugs inhibits chymotrypsin- and trypsin-like protease used have been, and still are, alkylating agents, activities. Preclinical studies have demonstrated antimetabolites, organoplatinum compounds, antitumor activity in models for multiple cytoskeletal disruptors, or anthracyclines, all myeloma, hematologic malignancies, and solid agents with relatively broad rather than tar- tumors and, importantly, marizomib does not geted and specifi c modes of action. show cross-resistance with other proteasome • A more refi ned strategy of specifi c action on inhibitors. Omuralide (clasto-lactacystin β-lactone; a particular cancer cell rather than nonspe- β-clastolactacystin) is the active metabolite of cifi c inhibition and killing has been used in

[email protected] 416 13 Drugs Used for Chemotherapy

developing some of the more recently intro- • Skin testing with the three platinum drugs has duced agents. The tyrosine kinase inhibitor been employed to identify at-risk patients and imatinib mesylate and proteasome inhibitor predict platinum hypersensitivity. As yet, skin bortezomib are examples of the more speciﬁ c testing with the drugs has not found wide- approach. spread acceptance and application as a routine • Taxanes such as docetaxel and paclitaxel are diagnostic procedure. mitotic inhibitors, disrupting microtubule • A 12 step, ~ 6 h desensitization protocol for function so adverse reactions, including carboplatin has been successfully employed in hypersensitivity responses, to these drugs skin test-positive patients hypersensitive to the might be expected. drug. • Up to 30 % of patients develop acute infusion • The Philadelphia translocation t(9;22) reactions to taxanes. Acute hypersensitivity (q34;q11) or Philadelphia chromosome is a reactions are marked by urticaria, ﬂ ushing, chromosomal defect resulting in gene fusion rashes, dyspnea, gastrointestinal symptoms, of the BCR and ABL genes. The resultant hypo- and hypertension, and back pain. fusion gene is the BCR-ABL oncogene. The • Patients with severe hypersensitivity to pacli- Philadelphia chromosome is a cytogenetic taxel also react to docetaxel, giving a cross- abnormality seen in 95 % of chronic myeloid sensitivity rate of ~90 %. leukemia patients and 15–30 % of adults with • Premedication regimen for docetaxel: oral acute lymphoblastic leukemia. dexamethasone 8 mg twice daily for 3 days • The drug imatinib mesylate that inhibits both starting 24 h prior to the commencement of the ABL and BCR-ABL tyrosine kinases has infusion. For paclitaxel infused over 1, 3, and been successful in treating chronic myeloid

24 h: H1 antagonist diphenhydramine (50 mg) leukemia.

and an H2 antagonist (cimetidine 300 mg, • In addition to imatinib, some inhibitors of ranitidine 50 mg, or famotidine 20 mg) given receptor tyrosine kinases targeting epidermal IV prior to infusion beginning. Oral dexameth- growth factor receptor (EGFR), vascular asone 20 mg administered 12 and 6 h prior. endothelial growth factor receptors (VEGFR) • Up to 50–70 % of cancer patients are treated and platelet-derived growth factor receptors with platinum drugs. Treatment with platinum (PDGFR) have been found to have antitumor drugs is often effected in combination with and/or other activities. other anticancer agents. • For most of the tyrosine kinase inhibitors • Mild reactions to platinum drugs are rash, targeting EGFRs, papulopustular rash; hand- urticaria, ﬂ ushing, palmar itching, a burning foot skin reaction; pigmentary changes; xero- feeling, hand and facial edema, pruritus, back sis; pruritus and mouth, hair, scalp, and nail pain, abdominal cramping, and diarrhea. abnormalities are the primary adverse events. Moderate to severe symptoms include diffuse • Imatinib is generally well tolerated especially erythroderma, tachycardia, chest tightness, when compared to most cytotoxic chemother- wheezing, facial swelling, dyspnea, hyper- apies. The most common non-hematologic tension, or hypotension. Severe symptoms adverse reactions include superﬁ cial edema, include bronchospasm, chest pain, seizures, nausea, diarrhea, muscle cramps, and vomit- and systemic anaphylaxis that may be ing and, most common of all, cutaneous reac- life-threatening. tions. Patients receiving standard dose • Most reactions to platinum drugs appear after imatinib therapy in the chronic phase of multiple treatment cycles (usually at least six). chronic myeloid leukemia experience neutro- • Reactions to the platinum drugs are mainly penia in 35–45 % of cases, thrombocytopenia type I or type IV hypersensitivity responses in 20 %, and anemia in 10 % of cases. with a few cases of type II and type III • Although most cutaneous reactions to ima- hypersensitivities. tinib are mild and dose dependent, severe

[email protected] Further Reading 417

reactions such as SJS, exfoliative dermatitis, Common terminology criteria for adverse events v3.0, TEN, AGEP, DRESS, and lichenoid eruptions DTCD, NCI, NIH, DHHS. Publish date: August 9, 2006 (http:/cstep.cancer.gov). have been reported. Crawford J, Dale DC, Lyman GH. Chemotherapy-induced • Gefi tinib and erlotinib are EGFR inhibitors, neutropenia. Risks, consequences, and new directions inhibiting the receptor’s tyrosine kinase for its management. Cancer. 2004;100:228–37. domain by binding to the ATP-binding site of Curtis BR, Kaliszewski J, Marques MB, Wasif Saif M, Nabelle L, Blank J, et al. Immune-mediated thrombo- the enzyme. EGFR is over-expressed by the cytopenia resulting from sensitivity to oxaliplatin. Am cells of some cancers such as lung and breast J Hematol. 2006;81:193–8. leading to uncontrolled cell proliferation. Druker BJ. Translation of the Philadelphia chromosome • The most frequent adverse reactions to gefi - into therapy for CML. Blood. 2008;112:4808–17. Guastalla JP, Diéras V. The taxanes: toxicity and quality tinib are diarrhea and skin reactions. The main of life considerations in advanced ovarian cancer. Br J adverse responses to erlotinib are rash, fatigue, Cancer. 2003;89 Suppl 3:S16–22. anorexia, diarrhea, nausea, ocular effects, Hannon MJ. Metal-based anticancer drugs: from a past infection, vomiting, and stomatitis. The FDA anchored in platinum chemistry to a post-genomic future of diverse chemistry and biology. Pure Appl has referred to rare serious gastrointestinal, Chem. 2007;79:2243–61. skin, and ocular disorders in some patients Howard SC, Jones DP, Pui C-H. The tumor lysis syn- taking erlotinib. drome. N Engl J Med. 2011;364:1844–54. • Bortezomib, an N -protected dipeptide that Kantarjian H, Sawyers C, Hochhaus A, et al. Hematologic and cytogenetic responses to imatinib mesylate in contains a boron atom, inhibits proteasomes chronic myelogenous leukemia. N Engl J Med. 2002; by binding via the boron atom to the catalytic 346:645–52. site of the 26S proteasome with high affi nity. Kisselev AF, van der Linden WA, Overkleeft HS. This ultimately leads to the killing of multiple Proteasome inhibitors: an expanding army attacking a unique target. Chem Biol. 2012;19:99–115. myeloma cells. Lenz H-J. Management and preparedness for infusion and • Gastrointestinal symptoms, thrombocytopenia, hypersensitivity reactions. Oncologist. 2007;12:601–9. peripheral neuropathy, and neuropathic pain Levitzki A, Klein S. Signal transduction therapy of cancer. are the most common side effects of bortezo- Mol Aspects Med. 2010;31:287–329. Limper AH. Chemotherapy-induced lung disease. Clin mib, and adverse cutaneous reactions to the Chest Med. 2004;25:53–64. drug are numerous—rash is frequently Lynch TJ, Kim ES, Eaby B, et al. Epidermal growth factor reported in more than 10 % of patients. receptor inhibitor – associated cutaneous toxicities: an • Newer proteasome inhibitors include the evolving paradigm in clinical management. Oncologist. 2007;12:610–21. orally active peptide boronate MLN9708 and Markman M, Kennedy A, Webster K, et al. Clinical carfi lzomib. Adverse reactions to carfi lzomib features of hypersensitivity reactions to carboplatin. include pulmonary hypertension, dyspnea, J Clin Oncol. 1999;17:1141–5. infusion reactions, tumor lysis syndrome, Moreau P, Richardson PG, Cavo M, et al. Proteasome inhibitors in multiple myeloma: ten years later. Blood. hepatotoxicity, rash, and urticaria. 2012;120:947–59. Nelson Jr RP, Cornetta K, Ward KE, et al. Desensitization to imatinib in patients with leukemia. Ann Allergy Further Reading Asthma Immunol. 2006;97:216–22. Pagani M. The complex clinical picture of presumably allergic side effects to cytostatic drugs: symptoms, Arora A, Scholar EM. Role of tyrosine kinase inhibitors pathomechanism, reexposure, and desensitization. in cancer therapy. J Pharmacol Exp Ther. 2005;315: Med Clin North Am. 2010;94:835–52. 971–9. Tanaka K. The proteasome: overview of structure and Balagula Y, Lacouture ME, Cotliar A. Dermatologic tox- functions. Proc Jpn Acad Ser B Phys Biol Sci. icities of targeted anticancer therapies. J Support 2009;85:12–36. Oncol. 2010;8:149–61. Wang D, Lippard SJ. Cellular processing of platinum Chen HX, Cleck JN. Adverse effects of anticancer agents anticancer drugs. Nat Rev Drug Discov. 2005;4: that target the VEGF pathway. Nat Rev Clin Oncol. 307–20. 2009;6:465–77.

[email protected] 418 13 Drugs Used for Chemotherapy

Wong M, Grossman J, Hahn BH, La Cava A. Cutaneous targeted molecular therapeutics. Annu Rev Pharmacol vasculitis in breast cancer treated with chemotherapy. Toxicol. 2012;52:549–73. Clin Immunol. 2008;129:3–9. Zanotti KM, Rybicki LA, Kennedy AW, et al. Carboplatin Yap TA, Workman P. Exploiting the cancer genome: strat- skin testing: a skin-testing protocol for predicting egies for the discovery and clinical development of hypersensitivity to carboplatin chemotherapy. Clin Oncol. 2001;19:3126–9.

[email protected] Proton Pump Inhibitors 1 4

Abstract Proton pump inhibitors (PPIs), omeprazole, lansoprazole, pantoprazole, rabeprazole, esomeprazole, and dexlansoprazole bind irreversibly to the H+ , K+ - ATPase (the “proton pump”) inhibiting its activity and decreasing gastric acid production. Systemic reactions to PPIs include anaphylaxis, urticaria, angioedema, interstitial nephritis, and thrombocytopenia. Cutaneous reactions include contact dermatitis, maculopapular and lichenoid eruptions, vasculitis, exfoliative erythrodermia, AGEP, DRESS, and SJS/TEN. Autoimmune reactions, including cutaneous lupus erythematosus, have been described. Cross-reactions between PPIs may be limited to one or two drugs or all drugs may be recognized. Cross-reaction studies so far have been based on skin testing, but the interpretations lack a quantitative basis. Successful oral desensitization following anaphylaxis to a PPI has been achieved in a few hours. Skin testing and challenge testing have been the only procedures employed to diagnose immediate reactions to PPIs. A suitable test for the detection of PPI-speciﬁ c IgE antibodies is not yet available, and application of the positive basophil activation test has been limited.

Proton pump inhibitors (PPIs) reduce gastric with the timoprazole backbone structure acid production in a pronounced and sustained (Table 14.1 ). Esomeprazole is the S -enantiomer manner. They are the most potent of the drugs of omeprazole and dexlansoprazole the that inhibit gastric acid secretion and are now R -enantiomer of lansoprazole. The structures of widely used, essentially replacing the formerly each of these PPIs consist of substituted pyridine heavily used histamine H 2 -receptor antagonists. and a benzimidazole heterocyclic center linked by a methylsulﬁ nyl group. Some new PPIs being developed, for example, tenatoprazole 14.1 Chemistry (Table 14.1 ), have an imidazopyridine instead of the benzimidazole ring structure. The imidaz- All marketed PPIs, omeprazole, lansoprazole, opyridine drugs have a longer half-life than the pantoprazole, rabeprazole, esomeprazole, and existing PPIs. dexlansoprazole are benzimidazole derivatives

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 419 Relationships, DOI 10.1007/978-1-4614-7261-2_14, © Springer Science+Business Media, LLC 2013

[email protected] 420 14 Proton Pump Inhibitors

Table 14.1 Chemical structures of benzimidazole proton pump inhibitors (PPIs) showing the timoprazole backbone structure and structure of tenatoprazole, a new generation imidazopyridine PPI

R * 2 N O R3 R4 S N R R1 H 5 N Substituted benzimidazole (C at ) Methyl Substituted * or sulfinyl pyridine imidazopyridine (N at *) group group group

General structure of PPI Proton pump Atom Enantiomorph inhibitor (PPI) at pos. * at S sulﬁ nyl –R1 –R2 –R3 –R 4 –R 5 Backbone structure Timoprazole C – –H –H –H –H –H Benzimidazole group a a Omeprazole C RS –OCH3 –H –CH 3 –OCH3 –CH3 b b Esomeprazole C S –OCH 3 –H –CH 3 –OCH3 –CH3 c c Lansoprazole C RS –H –H –CH3 –OCH2 CF3 –H d d Dexlansoprazole C R –H –H –CH 3 –OCH2 CF3 –H

Rabeprazole C RS –H –H –CH 3 –O(CH2 )3 OCH3 –H

Pantoprazole C RS –OCHF 2 –H –OCH3 –OCH3 –H Imidazopyridine group

Tenatoprazole N RS –H –OCH3 –CH 3 –OCH3 –CH 3 a Omeprazole is a 1:1 racemic mixture of the R - and S -enantiomers b Esomeprazole is the S -enantiomer of omeprazole c Lansoprazole is a 1:1 racemic mixture of the R - and S -enantiomers d Dexlansoprazole is the R -enantiomer of lansoprazole

14.2 Mechanism of Action Systemic reactions include anaphylaxis, urticaria, angioedema, acute interstitial nephritis, cytopenia, The PPIs are prodrugs, activated by exposure to thrombocytopenia, and vasculitis. Cutaneous pHs less than 5. Once activated, the drugs bind reactions include occupational contact dermatitis, irreversibly to the H + , K +- ATPase (the “proton photoallergic dermatitis, lichenoid eruption, pump”) in the parietal cell apical membrane, erythema nodosum, pytiriasis rosea, exfoliative inhibiting its activity and decreasing gastric acid erythrodermia, acute generalized exanthematous production by more than 95 %. The process is pustulosis (AGEP), fi xed drug eruption, maculo- irreversible in that new enzyme needs to be pro- papular eruption, drug reaction with eosinophilia duced to overcome the inhibition. PPIs have little and systemic symptoms (DRESS), and Stevens- effect on gastric acid volume and do not affect Johnson syndrome/toxic epidermal necrolysis gastric motility. (SJS/TEN). Autoimmune reactions, including cutaneous lupus erythematosus, have been described. 14.3 Hypersensitivity Reactions There are a number of reports of anaphylaxis to Proton Pump Inhibitors to PPIs, particularly omeprazole and pantoprazole. This may refl ect usage. Recent fi gures on Hypersensitivity reactions to PPIs may be mild the incidence of anaphylaxis to PPIs are hard to but the spectrum of possible reactions is wide fi nd but as of May 1999, the Uppsala Monitoring and some may be severe and life-threatening. Centre database contained 42 reports of anaphy-

[email protected] 14.3 Hypersensitivity Reactions to Proton Pump Inhibitors 421 lactic reactions to the drugs with omeprazole, the pyridine ring. Lansoprazole and rabeprazole lansoprazole, and pantoprazole showing inci- differ only at position 4 on the pyridine ring where dences (percentages of all reported adverse reac- the former has a trifl uoroethoxy and the latter a tions) of 0.2, 0.2, and 0.4 %, respectively. Judging methoxypropoxy group (Table 14.1 ). It seems by the number of reports in the literature, it seems likely that structures of the two drugs are suffi - certain that the number of cases of anaphylaxis to ciently similar to be recognized by some IgE PPIs since 1999 is considerably more than 42. antibodies. While these fi ndings demonstrate Investigations of immediate reactions to PPIs limited cross-reactivities, other investigations have generally been carried out by skin testing, have detected cross-reactivity covering all of the sometimes yielding results that provide informa- PPIs in current use. This area of PPI hypersensi- tion on cross-recognition between the different tivity research has essentially been based on drugs as well as the drug(s) provoking the reac- clinical studies principally using skin testing to tion. In one example, a patient with a severe demonstrate cross-recognitions, and the interpre- immediate reaction to lansoprazole confi rmed by tations lack a quantitative basis. Application of skin prick testing and challenge with the drug also quantitative hapten inhibition experiments along- reacted to a 5 mg challenge with rabeprazole, side skin test results are sorely needed, but this despite showing negative skin reactions to that will be diffi cult without a suitable method for the drug, omeprazole, and pantoprazole and negative detection of PPI-reactive IgE antibodies in aller- challenge tests to omeprazole and pantoprazole. gic patients’ sera. Possible cross-reactivity between lansoprazole Successful oral desensitization of a patient and rabeprazole was also demonstrated in a sepa- who experienced anaphylaxis to omeprazole was rate study by intradermal tests on a patient aller- achieved after 5.6 h, starting with an initial dose gic to the former drug. Investigations of immediate of 1 μg of drug and ending with a full dose of reactions to PPIs have revealed allergic recogni- 16 mg for a total cumulative dose of 32.6 mg. tion of omeprazole and lansoprazole in the same After the desensitization, the patient was able to patient, and one case of hypersensitivity to tolerate the full dose uneventfully and the wheal omeprazole showed a prick test-positive response size of the intradermal response to omeprazole to lansoprazole. Other observed patterns of was signifi cantly reduced. limited cross-reactivity include recognition of There appears to be fewer reports of delayed omeprazole, lansoprazole, and pantoprazole and reactions to PPIs, but the range of adverse skin cross-recognition between omeprazole and reactions seen is wide. Pantoprazole, for exam- pantoprazole. There is also a report of a patient ple, has been implicated in severe cutaneous with anaphylaxis and a positive skin test to responses including SJS/TEN, lichenoid erup- omeprazole and a negative skin test to pantopra- tion, exfoliative erythrodermia, and vasculitis. zole, lansoprazole, esomeprazole, and rabepra- At least one fatal reaction has occurred following zole. Another case report describes a patient TEN induced by a PPI. Maculopapular eruptions allergic to omeprazole but tolerant to both pan- and pruritus are frequently seen and mild in toprazole and lansoprazole (esomeprazole and intensity. Erythrodermic reactions to omeprazole rabeprazole were not tested). The clinical features and lansoprazole and allergic contact dermatitis of immediate reactions to PPIs suggest an IgE to lansoprazole have also been reported. A case antibody-mediated mechanism and the observed of DRESS induced by esomeprazole is notewor- cross-reactions likely refl ect antibody cross- thy since it involved co-sensitivity to other PPIs recognition of fi ne structural features on the and suggested caution in skin testing PPIs in different drugs. Omeprazole and pantoprazole patients with severe reactions. Patch testing are structurally fairly similar; the former has a using esomeprazole as a 10 % solution gave a methoxy substituent on the benzimidazole group positive reaction at 48 and 72 h. A second series while the latter has a difl uoromethoxy group at of patch tests proved positive to omeprazole and the equivalent position and an extra methoxy on pantoprazole as well as to esomeprazole, but no

[email protected] 422 14 Proton Pump Inhibitors reaction was seen with rabeprazole. Histological reaction to a PPI. Three doses of lansoprazole, examination of the esomeprazole-positive test 7.5, 15, and 30 mg, were given at 60 min showed typical signs of a delayed hypersensi- intervals. Twenty minutes after the third and tivity response. At 60 h after the second tests, fi nal dose, that is after a total dose of 52.5 mg, the patient experienced a mild erythroderma the patient reacted with erythema of palms, with facial edema and desquamation, indicating itching, rash, and malaise. induction of a fl are of DRESS. A recently published (2012) European multicenter study compared the diagnostic accuracy of skin and oral provocation tests in patients with 14.4 Diagnosis of Hypersensitivity immediate hypersensitivity to PPIs. Patients to Proton Pump Inhibitors with reactions that were not immediate were excluded. Skin prick tests were performed with Skin testing and to a lesser extent challenge solutions of omeprazole, esomeprazole, panto- testing have almost invariably been the only prazole, and rabeprazole at 40 mg/ml and lanso- clinical or laboratory test procedures employed prazole, 30 mg/ml. Omeprazole, esomeprazole, in the diagnosis of immediate reactions to PPIs. and pantoprazole were used in intradermal tests Prick test concentrations used have shown up to at 0.4 and 4 mg/ml. Oral provocation tests a tenfold variation: for omeprazole and panto- carried out on some patients after skin testing prazole, 4–40 mg/ml; esomeprazole, 20–40 mg/ consisted of the administration of four talc ml; lansoprazole, 3–30 mg/ml; and rabeprazole, capsules on day one followed on day two by 10–20 mg/ml. Solutions of omeprazole and pan- lansoprazole (5, 10, 15 mg) or one of the other toprazole have been prepared by dissolving four drugs (5, 5, 10, 20 mg) at 30 min intervals. lyophilized drug in physiological saline while Skin tests were positive in 12 of 53 patients; four solutions of the other three PPIs are usually for- of these underwent provocation testing with mulated from crushed and powdered tablets. the suspected PPI and in each case a positive Reported investigations have not always included response was obtained. Provocation tests on the results of skin tests on nonallergic controls. For 41 patients with a negative skin test showed intradermal testing, the concentration ranges three more positive reactors. For the skin tests, used have been more consistent—omeprazole specifi city and the positive predictive value were and pantoprazole, 0.04–8 mg/ml; lansoprazole, both 100 %. The negative predictive value was 0.015–3 mg/ml; esomeprazole, 0.02–2 mg/ml; 91.9 %. A higher frequency of skin test positiv- rabeprazole, 0.01–2 mg/ml. These concentra- ity occurred in patients with severe reactions tions generally represent 1:10, 1:100, and and cross-reactions consistent with previous 1:1,000 serial dilutions of the prick test concen- observations were observed. The study’s trations with testing starting at the lowest con- authors concluded that skin testing with PPIs centration and stepping up until a positive on patients with immediate hypersensitivity to reaction results. So far, there is limited informa- these drugs is a useful diagnostic test, and the tion available on patch testing with PPIs. Test test has the additional advantage of allowing the concentrations employed are in the range clinician to avoid oral challenges. 10–30 % in petrolatum or aqueous medium, So far a suitable test for the detection of PPI- and tests are generally read at least twice after specifi c IgE antibodies does not appear to be avail- 48–96 h. If cutaneous reactions are severe, great able. There are at least two reports of a positive caution should be exercised. basophil activation test on patients allergic to Oral challenge with lansoprazole of a patient omeprazole—one utilizing the CD63 basophil who experienced an anaphylactic-type reaction marker that was positive to omeprazole but negative to the drug provides an example of the use of this to pantoprazole and the other detected by the fl ow- test in confi rming a diagnosis of an immediate cytometric cellular allergen simulation test (FAST).

[email protected] Summary 423

possibility that long-term PPI use might lead to 14.5 Proton Pump Inhibitors, hypomagnesemia has led the FDA to suggest that Gastroesophageal Refl ux serum magnesium levels of patients taking PPIs Disease, and Asthma should be monitored. Again, the mechanism of such an effect of the PPIs is unclear. More clini- There appears to be a higher incidence of asthma cal data are needed in the case of each of these in children with gastroesophageal refl ux disease three concerns and until that is the situation, (prevalence estimated to be 34–89 %), and this clinicians and researchers should remain aware has prompted the suggestion, and the belief and keep abreast of developments in each area. seemingly supported by some studies, that PPI Lastly, the use of PPIs has been shown to be a treatment of these children may lead to an signifi cant risk for both community- and hospital- improvement in asthma symptoms. Three ran- acquired pneumonia. domized trials showed that PPIs had a benefi cial effect on asthma symptoms but one randomized, double-blind, placebo-controlled trial failed to Summary show that omeprazole improved symptoms. At present, there is not enough data from well • All marketed PPIs, omeprazole, lansoprazole, constructed and controlled clinical trials to reach pantoprazole, rabeprazole, esomeprazole, and a confi dent and conclusive decision on this dexlansoprazole are benzimidazole deriva- question. tives. Some new PPIs being developed have an imidazopyridine instead of the benzimidazole ring structure. 14.6 Other Safety Concerns with • The PPIs are prodrugs, activated by exposure Proton Pump Inhibitors to pHs less than fi ve. Once activated, the drugs bind irreversibly to the H + , K + - ATPase (the Besides hypersensitivity responses to PPIs, there “proton pump”) in the parietal cell apical are at least four other specifi c concerns related to membrane, inhibiting its activity and decreas- the interactions and/or direct effects of PPIs in ing gastric acid production by more than 95 %. humans. The oral antiplatelet drug clopidogrel is • Systemic reactions to PPIs include anaphy- used to inhibit blood clots. PPIs inhibit the bioac- laxis, urticaria, angioedema, interstitial tivation of clopidogrel to its active metabolite and nephritis, cytopenia, thrombocytopenia, and reduce the antiplatelet effects of the drug. It has vasculitis. been suggested that this may lead to an increased • Cutaneous reactions include occupational risk of vascular events. The results of a recent contact dermatitis, photoallergic dermatitis, randomized control trial with clopidogrel and pruritus, maculopapular eruptions, vasculitis, omeprazole do not add support to this belief, but lichenoid eruption, erythema nodosum, pyt- the makers of PPIs have agreed to work with the iriasis rosea, exfoliative erythrodermia, AGEP, FDA to conduct studies to obtain additional fi xed drug eruption, DRESS, and SJS/TEN. information that will allow a better understand- Autoimmune reactions, including cutaneous ing of the effects of PPIs on clopidogrel. A sec- lupus erythematosus, have been described. ond concern associated with PPIs is a suggested • Patterns of limited cross-reactivity include link between the drugs and fractures. Some recognition of omeprazole, lansoprazole, and believe that this could be related to altered pantoprazole and cross-recognition between absorption of calcium, vitamin B12 , or iron. Clear omeprazole and pantoprazole. Other investi- evidence to support an association with bone gations have detected cross-reactivity cover- fractures is, at present, lacking and no convincing ing all of the PPIs in current use. mechanism has been suggested, so the alleged • Cross-reaction studies so far have been based association remains to be resolved. Thirdly, the on skin testing, and the interpretations lack a

[email protected] 424 14 Proton Pump Inhibitors

quantitative basis. Development of IgE tests • Other safety concerns with PPIs include the and application of quantitative hapten inhibi- suggested inhibition of the antiplatelet effects tion experiments alongside skin test results are of clopidogrel leading to an increased risk of needed. vascular events, and associations with bone • Successful oral desensitization of a patient fractures, hypomagnesemia, and pneumonia. who experienced anaphylaxis to omeprazole was achieved after 5.6 h. • Skin testing and to a lesser extent challenge Further Reading testing have almost invariably been the only clinical or laboratory test procedures Bonadonna P, Lombardo C, Bortolami O, et al. employed in the diagnosis of immediate reac- Hypersensitivity to proton pump inhibitors: diagnostic accuracy of skin tests compared to oral provocation tions to PPIs. test. J Allergy Clin Immunol. 2012;130:547–9. • A recently published multicenter study Chang YS. Hypersensitivity reactions to proton pump compared the diagnostic accuracy of skin and inhibitors. Curr Opin Allergy Clin Immunol. 2012;12: oral provocation tests in patients with immedi- 348–53. Drepper MD, Spahr L, Frossard JL. Clopidogrel and pro- ate hypersensitivity to PPIs. For the skin tests, ton pump inhibitors – where do we stand in 2012? speciﬁ city and the positive predictive value World J Gastroenterol. 2012;18:2161–71. were both 100 %. The negative predictive Eurich DT, Sadowski CA, Simpson SH, et al. Recurrent value was 91.9 %. community-acquired pneumonia in patients starting acid-suppressing drugs. Am J Med. 2010;123:47–53. • A suitable test for the detection of PPI-speciﬁ c Roche VF. The chemically elegant proton pump inhibi- IgE antibodies is not yet available, and appli- tors. Am J Pharm Educ. 2006;70:101. cation of the positive basophil activation test Sachs G, Shin JM, Howden CW. Review article: the clinical has been limited. pharmacology of proton pump inhibitors. Aliment Pharmacol Ther. 2006;23 Suppl 2:2–8.

[email protected] Postface – Concluding Remarks

• Improved phenotyping of patients to clearly Drug Allergy Research in the identify associations with individual drug- Immediate Future: The Promise induced skin hypersensitivities. of Elucidating Mechanisms of • HLA–drug hypersensitivity associations (as the Immune Response to already seen, for example, with carbamaze- Small Molecules pine, allopurinol, and abacavir). Should prior genotyping be employed for some drugs? The Work on, and progress in, answering outstanding fascinating recent demonstration of modifi ca- questions in drug allergy today has the potential tion by abacavir of the HLA–peptide reper- not only to contribute signifi cantly to the diagno- toire and its effect on immune self-reactivity sis and treatment of poorly understood, diffi cult already promises to add a new dimension to to manage, and sometimes life-threatening the way we look at, and seek to understand, hypersensitivities but also to provide insights autoimmunity. into some of the most fundamental and impor- • The relationship between cutaneous and sys- tant areas of immunological investigation. These temic reactions in diseases such as drug reac- include cellular recognition and interactive pro- tion with eosinophilia and systemic symptoms cesses, relationships between receptors and (DRESS or hypersensitivity syndrome) and effector and signaling pathways, mechanisms of why the same drug produces both reactions in mediator action, and the genetic basis of drug some patients but only cutaneous reactions in reactions. More specifi cally, insights into the others. mechanisms of hypersensitivity and some other • Insights into cellular recognition of antigens intolerant reactions to drugs are likely to expand in the skin, resultant cell interactions, and sub- knowledge of and/or lead to: sequent cell-mediated toxic effects. • The immune response’s capacity (both • Identifi cation and mechanisms of action of humoral and cellular) to distinguish fi ne struc- important toxic mediators of cell damage in tural differences on structurally closely related local reactions. “small” molecules. • Further elucidation of drug receptor (hista- • Which drugs are recognized as hapten–pro- mine, leukotriene, PAF, etc.)-activated signal- tein complexes requiring processing by ing pathways. antigen- presenting cells and which drugs • Implication of the involvement of new media- activate an immune response without cova- tors in drug-induced anaphylaxis and cutane- lent binding. In each case, the mechanisms ous reactions. involved are likely to be amenable to investi- • Possible different pathways of anaphylaxis in gation and have the prospect of offering fasci- humans. nating insights into the allergic recognition of • The in vivo cellular effects and clinical conse- different structures. quences of sudden and complete removal of the

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 425 Relationships, DOI 10.1007/978-1-4614-7261-2, © Springer Science+Business Media, LLC 2013

[email protected] 426 Postface – Concluding Remarks

culprit drug in allergic patients by selective case, relatively recent insights have advanced to sequestration as seen, for example, with the stage where increasingly relevant questions sugammadex in anaphylaxis induced by the can be asked with the expectation that further sig- neuromuscular blocker rocuronium. nifi cant progress will result in the not-too-distant • The origin of preexisting allergic sensitivity in future. As mentioned immediately above and dis- patients who react to a drug without previous cussed earlier in this book, the recent elucidation exposure. of the mechanism underlying abacavir-induced • Detailed mechanisms involved in drug- hypersensitivity looks likely to prove a turning induced redirection of synthetic metabolic point in the way we look at small molecules and pathways thought to be operative, for exam- the immune response and the possible effects ple, in sensitivities to nonsteroidal anti- drugs, environmental toxins, and other chemicals infl ammatory drugs. might have in altering T cell immune responses. • The interrelationship between the anti- After being overlooked for so long on the fringe infl ammatory and immunosuppressive actions of mainstream immunology and virtually ignored of corticosteroids on the one hand and, on the in its textbooks, drug allergy research now other, the body’s allergic responses to the drugs. appears to offer paths that might lead to previ- • The molecular basis of cutaneous and systemic ously unsuspected advances in understanding allergic cross-recognition of corticosteroids. mechanisms of diseases where the interrelation- • Genetic engineering advances in improving ship between genetics, biochemical interactions, the selectivity of therapeutic monoclonal anti- and immune recognition processes had previ- bodies (mAbs) in terms of their improved ously appeared too daunting in terms of both the pharmacokinetics, binding affi nities, specifi c- biological complexities and the seemingly illogi- ities, toxicities, and increased half-life. cal nature of the drug-induced outcomes. For • The development of more, and improved, example, how could small amounts of some recombinant proteins (like, for example, etan- chemically unrelated drugs, often apparently ercept and denileukin diftitox), specifi cally chemically unreactive, provoke a catastrophic targeted at selected ligands or receptors cutaneous and systemic reaction such as toxic involved in disease. epidermal necrolysis or Stevens–Johnson syn- • The development of novel and therapeutically drome or DRESS or a range of other severe reac- more effective inhibitors of the ubiquitin– tions that seem to have an immune basis? Could proteasome pathway for chemotherapy of it turn out, as already suggested, that research cancer cells. into the immunopathogenesis of such poorly • Development of improved forms of drug understood drug-induced hypersensitivities also delivery, dosage schedules, and optimum leads us to a better understanding of autoimmune routes of administration for some important disease and perhaps even infectious diseases and drugs known to cause infusion and other cancer? severe reactions but not able to be substituted Finally, at this exciting time in the expansion (e.g., some specifi c anticancer drugs). of knowledge of the many known and suspected • Development of effective desensitization prodrug allergies and intolerances, it seems impor- cedures for important and increasingly used tant to keep a realistic perspective of the every- biologic agents and anticancer drugs com- day medical importance of reactions to different monly provoking hypersensitivity and other drugs or groups of drugs. Such a perspective adverse reactions. should relate to the frequencies and severities of Progress varies in the continuing efforts to reactions, short- and long-term morbidities, understand different mechanisms in the above- mortalities, economic costs, and the conse- listed, far from exhaustive, summary of impor- quences for continuing drug therapy with the tant research categories. Each is a major and offending drug and/or other drugs. A quick often diffi cult research area in itself, but, in every perusal of this volume reveals that what have

[email protected] Postface – Concluding Remarks 427 long been regarded by many as “true” allergies is ~0.5 per 106 person–years. By comparison, (e.g., immediate reactions such as drug-induced urticaria (the second most common drug-induced urticaria/angioedema/anaphylaxis, many drug- cutaneous reaction after erythematous reactions) induced exanthematous reactions such as macu- and anaphylaxis, for example, are far more lopapular exanthema, allergic contact dermatitis) commonly encountered and there is still much still constitute a large proportion of allergic drug to understand about both conditions. Comparing reactions throughout the world, and this is even the above frequencies for the severe, toxic more apparent if the whole spectrum of “hyper- bullous and other delayed cutaneous reactions to sensitivities” to the nonsteroidal anti-infl ammatory those for the most common offending drugs drugs is included. Indeed, signifi cant increases in causing adverse drug reactions (both true hyper- the incidences of both urticaria and anaphylaxis sensitivities and drug intolerances), namely, non- to drugs have recently been reported in a number steroidal anti-infl ammatory drugs, the penicillins, of countries. However, in recent years, the allergy cephalosporins, other antibiotics, the numerous literature at times gives the impression that drugs used in anesthesia and surgery, therapeutic drug-induced delayed hypersensitivity reactions mAbs, antineoplastic drugs, contrast media, and constitute almost the sole area of clinical and opioid analgesics, provides a truer perspective research interest in drug allergy and for patient of drug “allergy” than that gleaned from many importance. There is no doubting the advances current research priorities. made and the severe consequences for patients of In summary, while there is, and should be, a some drug- induced delayed reactions, but it research emphasis on the still poorly understood should be remembered that a high proportion of aspects of a number of cutaneous and cutane- patients with rashes and urticaria-like reactions ous/systemic drug-induced syndromes, and the are eventually shown to tolerate the previously considerable benefi ts of this research both at suspected drug and many of the severe responses the clinical and fundamental scientifi c levels remain extremely rare events. While research on are likely to extend beyond the confi nes of drug reactions to drugs such as sulfamethoxazole, allergy, the majority of “allergic” drug reactions carbamazepine, allopurinol, and abacavir has in everyday clinical practice usually fall outside yielded important scientifi c and clinical insights the currently favored areas of the most intense and provided knowledge that may be useful in research interest. In this volume we have preventing these and other potential drug toxici- attempted to cover the full range of different ties, the numbers of treated patients are often drug hypersensitivities and intolerances, from relatively small and the drug reactions sometimes common hives to life-threatening anaphylaxis predominate in often small groups of patients and simple erythematous rash to toxic epider- where special circumstances are involved. For mal necrolysis, with the aim of providing infor- example, apart from the treatment of HIV mation on the immune and some nonimmune patients, drugs such as sulfamethoxazole and drug reactions and their diagnoses. In addition, abacavir are not widely and heavily used and and with clinicians, researchers, teachers, and cases of drug- induced severe reactions like ery- students in mind, efforts have been made to theme multiforme, Stevens–Johnson syndrome, present a critical but balanced perspective of the and toxic epidermal necrolysis are infrequent importance and incidences of reactions, our with collective incidences of all three estimated current understanding of mechanisms under- to be 7, 1.8, and 9 per 106 person-years for lying the various drug hypersensitivities and patients <20, 20–64, and 65 years of age and intolerances, the remaining important gaps in older, respectively. For toxic epidermal necroly- our knowledge, and some likely important areas sis alone, the overall incidence of hospitalization for future research.

[email protected] Index

A fenamic acid derivatives , 323 Abacavir-HLA association , 72, 73, 89 immediate allergy , 336 Abacavir hypersensitivity syndrome (AHS) , 10 phenylpyrazolones , 323 α-Bungarotoxin (α-BT) , 272 pseudoallergic , 338 ACE inhibitors , 65 respiratory and cutaneous , 329 Acral erythema , 401–404 worldwide , 341 ACS. See Acute coronary syndromes (ACS) systemic toxicity , 283 Acute coronary syndromes (ACS) , 308 terminology , 2–5 Acute generalized exanthematous pustulosis (AGEP) , vasovagal reactions , 283 29–30, 81, 156, 157 Adverse reactions Acyl (R1) side chain, cephalosporin anti-cancer drugs , 400, 411, 416, 417 clinical hypersensitivity chemotherapeutics , 400, 411, 416 allergenic determinant investigations , 166 A ﬂ ibercept , 381 2-aminothiazol-4-yl R1 group , 168 Agent-induced anaphylaxis antigenic determinants , 167 Australian survey , 237–238 benzylpenicilloyl and amoxicilloyl conjugates , 168 culprit drugs , 238 dihydrothiazine ring , 166 E-mediated reactions , 237 IgE antibody inhibition , 164, 165 in EU , 238 methoxyimino group , 168, 169 in France , 237 n-butylamine , 166 latex , 237, 238

OH and O(CH3 )2 at C-3 , 167 NMBDs , 237 Sepharose radioimmunoassay , 164 penicillin and cephalosporin antibiotics , 238 in vitro and in vivo cross-reactivity , 168 responsibility, type I allergic reactions , 237 penaldate-and penamaldate-like structures , 164 AGEP. See Acute generalized exanthematous pustulosis Acyl (R2) side chain, cephalosporin (AGEP) anaphylactic reaction , 172 Agonist-biased signaling , 50, 52 4-carboxyl group retain stable complexes , 172–173 Agranulocytosis , 85, 193 challenges , 171 AHS. See Abacavir hypersensitivity syndrome (AHS) EDAC , 170 Alcuronium , 240–242, 258, 259, 261–263

and R1 side chains IgE , 170, 172 Allergen-induced late phase reaction serum IgE antibodies , 170, 171 delayed-type hypersensitivity response , 69–70 weak inhibitory activity, sera , 170 IgE antibodies , 69 Adalimumab , 377 immediate wheal and fl are cutaneous reaction , 68, 69 Adverse drug reactions (ADRs) lung function result , 68 “allergy” , 5–6 PEFR , 68 amide , 281 Allergic contact dermatitis , 25, 79 books, drug allergy (see Drug allergy) Allergic reactions, drugs defi nition , 1–2 categories , 2 delayed , 283–284 classifi cation , 26 dental treatment , 282–283 hypersensitivity epinephrine , 282 Chase’s experiments , 16 erythema/edema , 281 immune mechanisms , 16 hypersensitivity responses , 281–282 Mantoux test , 16 immediate , 283 Mycobacterium tuberculosis , 16 NSAIDs type I ( see Type I hypersensitivity drug reactions) aspirin-induced urticaria-angioedema , 334 type II , 23

B.A. Baldo and N.H. Pham, Drug Allergy: Clinical Aspects, Diagnosis, Mechanisms, Structure-Activity 429 Relationships, DOI 10.1007/978-1-4614-7261-2, © Springer Science+Business Media, LLC 2013

[email protected] 430 Index

Allergic reactions, drugs (cont.) hypersensitivity (see Hypersensitivities) type III , 23–24 IgE ( see Immunoglobulin E (IgE)) type IV ( see Type IV hypersensitivity drug IgE cross-reactivity with morphine , 241, 243, reactions) 252, 255 Allergy, defi nition , 5 leukocyte histamine release , 255 Allergy CSs mediators , 245 anti-infl ammatory effect , 397 morphine , 252–254 budesonide , 391, 393 persistent IgE antibodies , 246–247 contact eczema , 397 serum tryptase , 247 delayed type IV , 387, 397 skin tests (see Skin tests) hydrocortisone , 390 surgery, modern era , 239 immediate type I , 387 Anaphylaxis to proton pump inhibitors , 420–421, 424 incidences , 388 Anesthesia and surgery intradermal testing , 391 anaphylactic and anaphylactoid reactions , patients , 388 238–239 structure , 388 anaphylaxis tixocortol pivalate , 388 agent-induced anaphylaxis (see Agent-induced Allergy to steroids. See Corticosteroids (CSs) anaphylaxis) Allopurinol , 74 clinical features , 238 Amoxicillin allergenic determinants , 144 colloids ( see Anaphylaxis to colloids) Anakinra , 381–382 hypnotic drugs (see Anaphylaxis to hypnotic Anaphylactoid and anaphylactic reactions drugs) histamine release , 352–353 NMBDs (see Neuromuscular blocking drugs IgE antibodies , 353–354 (NMBDs)) infl ammatory mediators , 352 patent blue (see Patent blue anaphylaxis) iodinated contrast media , 360 drug-induced reactions kinin system and bradykinin , 354–355 administration , 235–236 NMBDs , 236 adverse reactions , 236 opioids , 303, 306, 314 education, clinicians , 236 Anaphylaxis risk , 235 clinical features , 19 surveys , 236 FcεRI-mediated mast cell activation , 59–60 symptoms, pseudoallergic , 236 grading system , 20 heparin ( see Heparin allergy) IgG involvement , 62 local anesthetics (see Local anesthetic allergy) incidence , 18 polypeptides ( see Polypeptides, anesthesia and nitric oxide , 62 surgery) PAF , 62 Angioedema , 21, 62, 65, 66 regulatory systems and signaling pathways , 58 Angiotensin-converting enzyme (ACE) inhibitors S1P ( see Sphingosine-1-phosphate (S1P)) and angioedema Anaphylaxis to colloids ACE2 , 66 description , 278 actions , 65 dextrans (see Dextran allergy) ARB , 66, 68 gelatin , 279 black Americans and Afro-Caribbeans , 65 HES ( see Hydroxyethyl starch (HES) allergy) bradykinin , 66 Anaphylaxis to CSs , 394 drugs implication , 64 Anaphylaxis to hypnotic drugs kallikrein-kinin system , 66, 67 cremophor-based induction agents , 273 Leydig cells , 65 ketamine and midazolam , 273 nitric oxide (NO) , 66 propofol ( see Propofol) NSAIDs , 63 surveys , 276 patients , 65–66 thiopentone ( see Thiopentone anaphylaxis) plasma kallikrein–kinin system , 66 Anaphylaxis to neuromuscular blockers renin–angiotensin system , 66 analogs , 250–252 somatic enzyme , 65 basophil activation test (BAT) , 254–255 Angiotensin II receptor blockers (ARBs) determination, serum tryptase , 247 and angioedema, 66, 68 diagnosis, allergic sensitization , 246 Antibiotics diagnosis with morphine solid phase , 255 aminoglycosides , 193, 196, 197 drug-induced muscle relaxation , 239 anaphylaxis , 184 epidemiology , 239 calculations , 184 HMT , 245–246 chloramphenicol , 198

[email protected] Index 431

clindamycin , 198–200 sensitivity , 194 defi nition , 184 skin test , 196 fosfomycin , 200 streptomycins , 196, 197 macrolides (see Macrolide allergy) Basophil activation markers neomycin , 193–194 CCR3 and CD63 , 114 non-β-lactams, 184 CD203c , 114–115 pristinamycin , 200 CD63 vs. CD203c , 115 ribostamycin , 197–198 E-NPPS3 and LAMP-3 , 114 rifamycins , 187–188 monoclonal antibody 97A6 , 114 tetracyclines (see Tetracycline allergy) phosphorylation, p38 MAPK , 115 vancomycin and teicoplanin PTPRC/CD45 , 114 adverse reactions , 191–192 Basophil activation test (BAT) desensitization , 192–193 diagnosis drug allergies red man syndrome , 190–191 application , 117 skin tests , 192 basophil activation markers , 114–115 structures , 188–190 CD203c and CD63 , 116 Antigen-presenting cells (APC) , 42, 71, 74 cell-damaging manipulations , 118 Antimicrobials controls and evaluating results , 116 chlorhexidine (see Chlorhexidine allergy) donor cells, sensitizing , 116 povidone-iodine , 230 fl ow cytometry (see Flow cytometry) quinolones ( see Quinolone allergy) granulocytes , 113–114 sulfonamides (see Sulfonamide allergy) HistaFlow method , 118 trimethoprim ( see Trimethoprim allergy) IgE antibody-mediated activation , Anti-thymocyte globulin , 382 115–116 APC. See Antigen-presenting cells (APC) isolated leukocytes and blood , 115 Aprotinin allergy mast cells and measurement , 114 adverse reactions , 287–288 technical aspects , 115 cardiopulmonary bypass surgery , 286 passive sensitization , 116, 118 description , 286 NMBDs , 254–255 fi brin glues, hemostasis , 286 B A T . See Basophil activation test (BAT) risk factors , 286 Biologics Aspirin challenge tests , 331 afl ibercept , 381 Aspirin desensitization , 333, 334 anakinra , 381–382 Aspirin-induced asthma anti-thymocyte globulin , 382 diagnosis and therapy , 341 autoimmune diseases , 369 Korean populations , 340 backbone treatments , 1990s, 369 leukotriene receptor genes , 341 composition , 369–370 5-lipoxygenase gene involvement (ALOX5) , 341 denileukin diftitox , 381

LTC4 synthase , 340–341 epoetins , 382–383 and urticaria/angioedema , 341 etanercept , 379–380 Azithromycin , 185 human insulin , 383 Aztreonam allergy IFNs , 380 ceftazidime , 175, 176 IL-2 , 380–381 cross-reaction , 175 mAbs ( see Monoclonal antibodies (mAbs)) monobactam structures , 175 Bortezomib adverse reactions , 400, 415, 417 patients allergic to β-lactam antibiotics, 176 Bortezomib cutaneous reactions , 414, 417 synthetic compound , 175 Bortezomib proteasome inhibition gastrointestinal symptoms and thrombocytopenia , 414 B molecular mechanisms , 413 Bacitracin allergy NF-κB , 412–413 and aminoglycoside antibiotics , N -protected dipeptide, boron atom , 412, 414 195–196 peripheral neuropathy and neuropathic pain , 414 antibiotics , 194 rash and subcutaneous infusion , 414 delayed and immediate reactions , 195 suppresses tumor growth and spread and dermatitis , 195 angiogenesis, 412 ear diseases , 196 Sweet’s syndrome/acute febrile neutrophilic kanamycin and tobramycin , 196 dermatosis, 414 patch test , 196 Bradykinin , 65, 67 penicillins and cephalosporins , 197 Budesonide , 389–393, 396

[email protected] 432 Index

C Cephamycins , 130 cAMP response element-binding (CREB) , 51 Cetuximab , 375–376 Carbamazepine-HLA association Challenge tests, drug allergy abacavir hypersensitivity and fl ucloxacillin-induced anaphylactic shock , 107 liver injury , 73 antihistamines , 106 allopurinol , 74 aspirin and β-lactams, 107 HLA-A30 , 74 contrast media , 359 HLA-B22 , 74 description , 105 HLA-B*58:01 allele , 74 doses , 106 and HLA-B*15:02 interaction , 73–74, 89 histamine-releasing drugs and index reaction , 105 HLA-DRw4 , 74 NSAIDs , 331 NSAID, phenylbutazone derivative and feprazone , 74 penicillins , 153, 154, 157 SJS and TEN , 73 placebo-controlled , 106 Carbapenem allergy , 176–177 PPIs , 442 Case history , 92 provocation , 106, 107 CAST. See Cellular allergy stimulation test (CAST) quinolones , 221 CCDs. See Cross-reactive carbohydrate determinants risk-to-benefi t ratio , 107 (CCDs) skin test/serum IgE antibody , 105 Cell surface activation markers , 121–122 Chemokines Cellular allergy stimulation test (CAST) cysteine–cysteine chemokine receptor 3 , 114 basophils and cysteinyl leukotrienes , 112 and skin-homing , 123–124 cysteinyl leukotrienes , 126 Chemotherapy and ELISA , 112, 126, 152–153 anticancer drugs , 400 NSAIDs and β-lactam drugs, 113 description , 399 positive CAST and negative BAT results , 113 non-mAb anticancer , 400–403 Cephalosporins allergy organoplatinum (see Organoplatinum drugs) acyl (R1) side chain (see Acyl (R1) side chain, proteasome inhibitors , 412–415 cephalosporin) taxanes (see Taxane hypersensitivity) allergic patients , 160 tyrosine kinase inhibitors , 409–412 aminolysis , 163 Chinese hamster ovary (CHO) cells , 49, 373 antigens and allergenic determinants , 161 Chloramphenicol allergy , 198 Cephalosporium acremonium , 159 Chlorhexidine allergy clinical use , 159–160 anaphylaxis , 226, 227 cross-reactions , 160–161 catheters , 226–227 and decomposition products , 163–164 defi nition , 225 delayed hypersensitivity reactions , 174 delayed reactions , 227 delayed reactions , 174–175 hypersensitivities description , 159 anaphylaxis , 227 determinants IgE antibody tests , 228–230 anti-cephalosporin IgE antibodies , 173 skin tests , 227–228 and IgE antibody tests , 170 immediate hypersensitivity reactions R2 side chain (see Acyl (R2) side chain, (see Hypersensitivities) cephalosporin) structure , 226 hypersensitivity , 160 survey , 227 imipenem , 176 CHO cells. See Chinese hamster ovary (CHO) cells monobactam aztreonam , 175 Choline , 250–252 and penicillin cross-reactivity Cilastatin , 176 anaphylaxis , 161 CLA. See Cutaneous lymphocyte-associated antigen children, anaphylactic reactions , 161 (CLA) patients history , 160 Classifi cation, ADRs risk factors , 161 adverse drug reactions , 2, 3 the United Kingdom General Practice Research dose-related and non-dose-related reactions , 2 Database, 160–161 DRESS , 5 penicillins , 159 EAACI , 3 prick and intradermal testing , 173 hypersensitivity reactions , 2 side chain determinants (see Acyl (R1) side chain, “immediate hypersensitivity” , 4 cephalosporin) immune and nonimmune sensitivities , 2, 4 skin testing , 173–174 “nonallergic hypersensitivity” , 5 structures and classifi cation , 161, 162 NSAIDs , 4, 5 therapeutic use , 160 terminology , 5

[email protected] Index 433

type B reactions , 2 metabolism and degradation , 388 type I and type IV hypersensitivities , 4 patch testing , 390–392 unpredictable reactions , 2 risk factors , 39 Clavams , 177 sensitization , 394 Clavulanic acid allergy , 177–178 structure-activity relationships , 393 Clindamycin allergy structural groups , 388, 389 compound structure , 198–199 COX isoenzymes , 324 delayed reactions , 199–200 CPK models. See Corey, Pauling and Koltun (CPK) diagnostic tests , 199–200 models hypersensitivity , 199 CREB. See cAMP response element-binding (CREB) COADEX classifi cation , 99, 126 Cross-reactions Contact dermatitis , 25, 26, 79 CSs Contrast media C6/C9 and C16/C17 , 393–394 adverse reactions (see Reactions to contrast media) classifi cation, structure–activity analyses , 393 anatomical structures , 343 clinical, budesonide , 393 challenge test , 105, 359–360 delayed and immediate reactions , 394 gadolinium (see Gadolinium contrast media) gonane/cyclopentaneperhydrophenanthrene investigative techniques and diagnostic equipment, 343 nucleus , 393 iodinated ( see Iodinated contrast media) group D esters , 393 nephrogenic systemic fi brosis , 361 IgE antibodies or T cells , 394 premedication (see Pre-medication for contrast media) patch testing , 394 skin tests of sulfonamides delayed , 356 allergic reaction , 211–212 diagnosis , 356 and arylamines , 212–213 gadolinium , 364–365 compound structure , 209, 210 hypersensitivity , 356 heterocyclic ring , 209, 211 intradermal and patch , 356 IgE antibody-mediated cross-reactivity , 211 limitations , 356–357 pharmacological drugs , 209 nephrogenic systemic fi brosis , 361 risk factor , 211 prick , 356 skin test , 211 procedures , 357 “sulfa allergy” , 211 tools , 357 sulfonyl group , 209, 210 usage and safety , 346–347 Cross-reactive carbohydrate determinants (CCDs) , 39 X-ray imaging techniques , 343–344 CSs. See Corticosteroids (CSs) Corey, Pauling and Koltun (CPK) models Cutaneous lymphocyte-associated antigen (CLA) allergenic determinants, ampicillin , 145 chemokines , 127 amoxicillanyl-and amoxicilloyllysine conjugates , 146 β-lactams, 124 ball-and-stick three-dimensional molecular , 270 l -fucose-containing carbohydrate epitope , 124 gadopentetic acid , 362 positive cells , 124 IgE antibody-binding regions , 143 Cutaneous reactions ionic and nonionic iodinated contrast media , 347 chemotherapeutic drugs , 399 l -methadone , 300 NSAIDs morphine , 253 aspirin-induced asthma , 333 PAF , 56 chemotherapeutic drugs , 400, 405–409 quaternary allylammonium and allylpyrrolidinium desensitization , 334 ions , 263 management , 334 rocuronium , 270 urticaria angioedema (see Urticaria angioedema, sulfamethoxazole, sulfamerazine and sulfamethazine , NSAIDs) 205 to proton pump inhibitors , 420, 422, 423 thiopentone , 274 Cyclooxygenase isoforms , 324 trimethoprim with IgE antibody-binding Cyclooxygenases , 324–328 determinants, 217 Cysteinyl leukotrienes Corticosteroids (CSs) allergen-induced release , 112–113 allergy and sensitization , 387–388 basophils , 112 delayed hypersensitivity reactions , 390–394 biosynthesis , 52–54 diagnosis of hypersensitivity , 390–392, 396 CAST ( see Cellular allergy stimulation test (CAST)) fi nding safe alternative drug , 396 description , 52 haptens , 388–389 histamine and snake venom , 52 immediate reactions (see Immediate hypersensitivity isolation , 52

reactions, CSs) LTC4 , LTE4 and LTD4 , 52

[email protected] 434 Index

Cysteinyl leukotrienes (cont.) Delayed-type hypersensitivity reactions receptors , 54–55 CSs structures , 111 allergic contact dermatitis , 390 sulfi doleukotrienes , 111–112 cell proliferation , 392 Cytokine release syndrome , 415 diagnosis , 390–392 Cytokines edge effects , 390 ELISPOT assay , 122–123 hydrocortisone–albumin complex , 392 fl ow cytometry and ELISA assay , 122 lymphocyte transformation test , 392 in vitro tests , 126 perioral dermatits , 390 T cells , 122 structure–activity relationships , 393–394 test solutions stability , 392 description , 156 D diagnostic tests , 156–158 DAG. See 1,2-diacylglycerol (DAG) T cells , 158–159 Danaparoid , 290 Denileukin diftitox , 381 DAO. See Diamine oxidase (DAO) Dermatitis , 25, 26, 79 Darbepoetin , 383 Desensitization , 75 Defi nition of hypersensitivity aspirin , 333 adverse drug reactions and allergy cetuximab , 375 ( see also Classifi cation, ADRs) imatinib , 411 “immediate hypersensitivity” , 4 infl iximab , 377 “nonallergic hypersensitivity” , 5 insulin , 383 type I and type IV hypersensitivities , 4 NSAIDs, 333, 334 AHS with HLA-B*5701 , 13 penicillins , 154 “allergy” , 5 platinum drugs , 408 animal model , 7 PPIs , 421 delayed type , 7 taxanes , 405 drug allergies and sensitivities vancomycin , 192 ADRs, hospital inpatients , 7 Dextran allergy defi nition , 7 adverse reactions , 281 immune system , 7 anaphylactoid reactions , 280 reactions , 6–7 chains , 280 type B reactions , 2 defi nition , 280 type II hypersensitivity reactions , 11 DIAR , 280 Delayed cutaneous ADRs IgG antibodies , 280 acute generalized exanthematous pustulosis , 81 molecular weight , 280 allergic contact dermatitis , 79–80 neurological impairment and deaths, neonates , 280 cytokine actions , 79 skin testing , 281 fi xed drug eruption , 81 1,2-diacylglycerol (DAG) , 13, 49 hypersensitivity reactions , 83–84 Diagnosis corticosteroid hypersensitivity maculopapular exanthema , 81 immediate systemic reactions , 396 psoriasis , 80 patch testing (see Patch tests) rash with eosinophilia and systemic symptoms , 81 rosacea and perioral , 390 T cell lines and clones , 79 Diagnosis delayed drug allergy toxic epidermal necrolysis and Stevens–Johnson description , 118, 120 syndrome , 82–83 in vitro Assays , 121–124 Delayed drug reactions local lymph node assay , 121 vs. late phase reaction , 69–70 lymphocyte transformation test , 120–121 NSAIDs T cells , 118 cutaneous and systemic , 337, 338 type iv cell-mediated mechanisms , 118 diagnosis , 337 Diagnosis drug allergy drug provocation , 337 avoidance strategy , 92 immune-based type IV reactions , 337 BAT ( see Basophil activation test (BAT)) patch and lymphocyte transformation test , 337 causal relationship, drug and patient reactions , Stevens–Johnson syndrome , 338 124–125 toxic epidermal necrolysis , 338 challenge (provocation) testing (see Challenge tests, type II hypersensitivity , 84–86 drug allergy) type III hypersensitivity , 86–87 culprit drug , 93 type IV hypersensitivity (see Type IV hypersensitivity cysteinyl leukotrienes (see Cysteinyl leukotrienes) drug reactions) delayed type hypersensitivity reactions , 118–124

[email protected] Index 435

description , 91–92 classifi cation , 62–63 histamine (see Histamine) genetic mechanisms , 63 IgE antibody tests , 100–105 heterogeneous disease , 63 patient’s clinical history , 92 immune mechanisms , 63–64 physician/anesthetist , 93 nonimmune ( see Nonimmune-mediated urticaria skin testing ( see Skin tests, drug allergy) and angioedema) T cell-mediated mechanism , 93 NSAIDs , 62 tryptases (see Tryptases) Drug-induced cutaneous reactions Diagnosis immediate drug allergy and ADRs (see Delayed cutaneous ADRs) BAT , 114-117 CD4+ and CD8+ CD3+ T cells , 76 challenge test , 105-107 type IV hypersensitivity , 76–77 cysteinyl leukotrienes , 111–112 Drug-induced delayed reactions. See Type IV cytokines , 122 hypersensitivity drug reactions ELISPOT assay , 122–123, 126–127 Drug-induced histamine release histamine , 109–111 smooth muscle , 304 IgE tests , 100–105 triple response in human skin , 304–305 skin tests , 93–97 urticaria pigmentosa , 305 tryptase test , 107–109 vascular reactions, skin , 304 Diamine oxidase (DAO) , 47 Drug-induced hypersensitivity syndrome (DIHS) , 81 Diaveridine , 215 Drug-induced immune hemolytic anemia (DIIHA) DIHS. See Drug-induced hypersensitivity syndrome antibodies implication , 84 (DIHS) drug participates , 84 DIIHA. See Drug-induced immune hemolytic anemia proposed mechanism , 84 (DIIHA) red cell autoantibodies , 84 DILI. See Drug-induced liver injury (DILI) Drug-induced liver injury (DILI) , 81, 178–190 DILD. See Drug-induced lung disease (DILD) Drug-induced lung disease (DILD) , 87, 178, 179 Dimethyl sulfoxide (DMSO) , 94 Drug-induced serum sickness DMSO. See Dimethyl sulfoxide (DMSO) amoxicillin , 24 DRESS. See Drug reaction (rash) with eosinophilia and and anaphylaxis , 21 systemic symptoms (DRESS) classic , 24 Drug allergy , 5 discovery , 16 ADR , 5 and penicillin-induced hemolytic anemia , 23 drugs, haptens and prior exposure , 6–7 Gell and Coombs classifi cation , 26 hypersensitivities and sensitivities , 7–8 lymphadenopathy , 24 immunology , 2, 11 type II and III hypersensitivity , 23 nonantigenic substances , 6 Drug-induced type II hypersensitivity. See Type II opioids hypersensitivity drug reactions methadone , 312 Drug-induced type III hypersensitivity. See Type III sIgE antibodies , 313 hypersensitivity drug reactions quinine , 6 Drug-induced urticaria, type I hypersensitivity reference book , 11 angioedema , 22–23 risk factors , 8–9 chronic , 21 scientifi c chemical framework , 6 cutaneous reaction , 21 single text book , 12 erythematous patches , 21–22 Drug anaphylaxis types II and III reactions , 23 administration, new drugs , 17 Drug reaction (rash) with eosinophilia and systemic clinical features , 19–20 symptoms (DRESS), 5, 30–31, 97, 107, grading system , 20–21 120–121, 126, 154, 156, 157, 179 incidence , 18–19 signs and symptoms , 17 terminology , 17–18 E Drug delivery and allergenicity , 11 EAACI. See The European Academy of Allergology and Drug hypersensitivity , 3, 7 Clinical Immunology (EAACI) Drug-induced angioedema Ecto-nucleotide pyrophosphatase phosphodiesterase 3 Gell and Coombs classifi cation , 25 (E-NPPS3), 114 type I hypersensitivity , 16–23 EDAC. See 1-Ethyl-3-(3-dimethylaminopropyl) type III hypersensitivity , 24 carbodiimide (EDAC) and urticaria EGFR. See Epidermal growth factor receptor (EGFR) autoimmune basis , 64 ELISA. See Enzyme-linked immuno sorbent assay basophil activation , 64 (ELISA)

[email protected] 436 Index

ELISPOT assay , 123 FDA , 361 E-NPPS3. See Ecto-nucleotide pyrophosphatase gadopentetate dimeglumine , 364 phosphodiesterase 3 (E-NPPS3) histamine release test , 364 Enzyme-linked immuno sorbent assay (ELISA) life-threatening reactions , 364 alkaline phosphatase , 112 magnetic resonance angiography (MRA) , 361 CAST® , 126 molecular structures cysteinyl leukotrienes , 112 acyclic agent , 361 fl ow cytometry , 121, 122 2D structure , 361, 362 RIA , 110 three macrocyclic , 361, 363 Enzyme-linked immunospot (ELISPOT) assay NSF (see Nephrogenic systemic fi brosis (NSF)) cytokines , 122–123 risks, acute reactions , 361, 364 drug allergy diagnosis , 122 skin tests , 364–365 granzyme B , 123 survey , 364 IFN-γ , 123 Gefi tinib and erlotinib , 411–412 Eoxins , 54 Gelatin allergy , 279–280 Epidermal growth factor receptor (EGFR) , 410, 411, 416 GM-CSF. See Granulocyte-macrophage Epoetins , 382–383 colony-stimulating factor (GM-CSF) Erlotinib , 412 GPCR. See G protein-coupled receptor (GPCR) Erythema multiforme , 32–33 G protein-coupled receptor (GPCR) , 47, 52, 54 Erythromycin allergy. See Macrolide allergy Granulocyte-macrophage colony-stimulating factor Etanercept , 379–380 (GM-CSF), 45, 70, 81 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide Granulysin , 82 (EDAC) , 170 Granzyme B ELISPOT assay , 123 The European Academy of Allergology and Clinical Immunology (EAACI) , 3 H Hand-foot skin reaction , 404, 412 F Heparin allergy FAST. See Flow-cytometric cellular allergen simulation adverse reactions , 289–290 test (FAST) cardiac surgery , 289 FDA. See Food and Drug Administration (FDA) danaparoid and hirudins , 290 FDE. See Fixed drug eruption (FDE) diagnostic skin testing , 290 Fenamic acid derivatives , 323 fondaparinux , 291 Fentanyl , 299, 303, 312 low molecular weight (LMW) , 289 Feprazone , 74 meaning , 289 Fixed drug eruption (FDE) , 31–32 patch tests , 290 Flow-CAST , 112, 152 polymers , 289 Flow-cytometric cellular allergen simulation prick testing , 290 test (FAST) , 422 HES allergy. See Hydroxyethyl starch (HES) allergy Flow cytometry 5-HETE. See 5-Hydroxyeicosatetraenoic acid (5-HETE) and BAT’s , 118 Hirudins, 290 CD203c and CD63 , 117 Histamine development , 114 anaphylactic patients and in vitro , 111 HistaFlow , 117–118 anti-IgE and spontaneous release , 110 histamine and PMA , 117 concentrations in blood , 110 intracellular histamine and release , 118, 119 control for skin tests , 113 single cell level and anaphylactic degranulation , 117 cross-reactivity ratios , 110 Flucloxacillin , 140–142, 159 in drug allergy diagnosis , 111 Fondaparinux , 290 fl ow cytometry , 117–118 Food and Drug Administration (FDA) , 2, 10 human blood leukocytes and blood sampling , 109 Fosfomycin , 200 in vivo half-life , 110–111 opioid drug-induced release , 303–305 receptors G allergic processes , 304 Gadolinium contrast media Ca2+ triggers , 51–52 “allergic-type” reactions , 362 chemotactic response , 304 anaphylactic/anaphylactoid reactions , 362 1-[(chloro-1-H -indol-2-yl)carbonyl]-4-methyl- clinical safety and diagnostic value , 364 piperazine, 52

detection, MRI , 361 G α i/o proteins , 51 exposure products , 364–365 GPCRs , 52

[email protected] Index 437

G protein-coupled receptor , 303–304 risk factor , 388

H1 receptor , 49 defi nition , 4 H2 receptor , 49 drug-allergic patients desensitization H3 receptor , 49–50 adverse reaction , 75 H4 receptor , 50–51 long-term tolerance , 75–76 human T-and dendritic cells , 304 mast cells , 75 2-(imidazol-4-yl)ethylamine , 46–47 RDD , 75 and l -histidine , 47, 48 drug-induced and immune receptors mast cells and basophils , 47, 304 abacavir and MHC-presented altered peptide signaling pathways , 49 model , 72–73 types , 47–48 carbamazepine and HLA , 73–74 RIA/ELISA , 110 immunological dogma , 70–71 Histamine-N -methyltransferase (HMT) , 47, 245–246 MHC-peptide complex , 74–75 HIV-1. See Human immunodefi ciency virus-1 (HIV-1) p–i concept , 74 HLA. See Human leukocyte antigens (HLA) T cell recognition and the immune response , 71 HMT. See Histamine-N -methyltransferase (HMT) drug-induced delayed-type cutaneous hypersensitivity 5-HPETE. See 5-Hydroxyperoxyeicosatetraenoic acid reactions, 79 (5-HPETE) early years , 16 Human immunodefi ciency virus-1 (HIV-1) , 10, 208 granulysin , 82

Human insulin , 383 H1 and H2 receptor , 49 Human leukocyte antigens (HLA) , 10 H 3 and H4 receptor , 49, 50 Hydrocortisone , 389–391, 396 histamine , 46 5-Hydroxyeicosatetraenoic acid (5-HETE) , 53, 54 human cysteinyl leukotriene receptors , 52 Hydroxyethyl starch (HES) allergy IgE antibodies and IgE-mediated drug administration , 278 (see IgE antibodies) chemistry , 278 “inert” parent drugs , 37

diagnosis , 279 LTC 4 and LTD4 , 52 property , 278 macrolide allergy (see Macrolide allergy) reaction mechanisms , 279 mast cells , 46 risk and adverse reactions , 278–279 NSAIDs 5-Hydroxyperoxyeicosatetraenoic acid (5-HPETE) , 53, 54 acute angioedema , 328–329 Hypersensitivities aspirin , 329 abacavir–HLA binding studies , 72, 88 and BAT , 340 allergen-induced late phase reaction classifi cation , 330 delayed-type hypersensitivity response , 69–70 cutaneous reactions (see Cutaneous reactions, IgE antibodies , 69 NSAIDs) immediate wheal and fl are cutaneous reaction, 68, 69 defi nition and epidemiology , 329 lung function result , 68 delayed reactions , 337–338 PEFR , 68 mixed pattern , 338–339 allergic sensitization respiratory reactions (see Respiratory reactions, immunogenicity ( see Immunogenicity) NSAID) immunological recognition , 41–42 systemic pattern , 339 mediator release , 40–41 opioids anaphylactic reactions , 37 adverse skin reactions , 317 anaphylaxis to neuromuscular blockers phenanthrene drugs , 316 IgE antibodies , 256 pneumonitis and maculopapulous toxic skin , investigative procedures , 256 312–313 life-threatening response , 255 PAF , 55 skin testing , 256 pathophysiological effects , 88 tryptase release , 255–256 pneumonitis , 69 angioedema , 21, 62 quinolone allergy (see Quinolone allergy) carbamazepine , 89 sphingosine-1-phosphate , 60 CD23 , 87–88 steroids ( see Corticosteroids (CSs)) to chemotherapeutics (see Organoplatinum drugs) Syk kinase and subsequent phosphorylation chlorhexidine allergy (see Chlorhexidine allergy) activation , 59 classifi cation , 16 trimethoprim allergy , 213–214 CSs type II cytotoxic antibody-mediated drug reactions , 84 delayed reactions (see Delayed-type type III reactions (see Type III hypersensitivity drug hypersensitivity reactions) reactions) immediate , 394

[email protected] 438 Index

type II reactions (see Type II hypersensitivity drug antisera , 141 reactions) benzylpenicilloyl-protein conjugate , 142 Hypersensitivities (cont.) clinical relevance , 147 type IV reactions (see Type IV hypersensitivity drug complementary immune receptors , 143 reactions) haptenic determinants, allergenic drugs , 141 urticaria , 21, 62 immediate allergic reactions , 142 vasculitis , 87 patients sera , 144 reagins mediating skin , 141 thiazolidine ring , 144 I quinolone allergy ICOS , 78 administration , 224 Idiosyncratic reactions , 2–5 bis-oxirane coupling drug , 221 IFNs. See Interferons (IFNs) histamine release , 223 IgE. See Immunoglobulin E (IgE) pipemidic acid , 222–223 IgE and contrast media preparations , 221–224 anaphylactoid and anaphylactic reactions , 353–354 radioimmunoassay , 223–224 basophil activation , 358 sulfonamide-reactive , 202 Prausnitz–Kustner , 357–358 tests serum , 358 automation , 101 IgE antibodies calibration system , 105 adverse reactions to contrast media class/scale system range , 104 basophil activation test , 358 clinician-diagnosed positive and negative , 104–105 Prausnitz–Kustner test , 357–358 detection, drug-speciﬁ c , 104 serum tests , 358 eczema/psoriasis , 100 ampliﬁ cation and production , 45 in vitro detection , 101–103 cephalosporin reactive ImmunoCAP® assay , 103, 105 allergic patients , 170, 171 “in-house” tests , 103 anti-Cephalosporin , 173 liquid phase systems and solid phase technology , benzylpenicillin , 168 100–101 cephalothin Sepharose , 165 Prausnitz–Kűstner test, 100 HSA solid phases , 170 preparation, drug solid phases and procedures , human anti-benzylpenicilloyl , 176 103–104 in vitro studies , 166, 167 radioallergosorbent test (RAST) , 100 non-IgE-mediated , 160 semiquantitative scale and ROC , 104 paradoxical situation , 170 skin test-negative/equivocal reactors , 100 patients allergic sera , 164 trimethoprim-reactive , 214

R1 and R2 side chains , 171, 172 IgE to drugs side chain (R1) , 164 detection , 104 chlorhexidine-reactive , 228 in vitro detection , 101–103 clinical relevance , 147 radioallergosorbent test (RAST) , 100 and IgE-mediated drug hypersensitivities IgG antibodies anaphylaxis (see Anaphylaxis) anaphylaxis , 61–62 B cells , 45, 46 anti-cefotetan , 84 FcεRI receptor , 44 anti-immunogloulin E autoantibodies , 64 IgG, PAF and nitric oxide , 61–62 chimeric mouse–human IgG1 , 39 low-afﬁ nity IgE receptor FcεRII (CD23), 45–46 and IgM , 63, 84–86 mast cells , 43–45 receptor FcγRIII , 58, 88 T cells , 42–43 IgM antibodies type I immediate allergic response (see Type I IgE-bearing cells , 42 immediate allergic response) and IgG , 63, 84, 86 urticaria and angioedema (see Drug-induced IL-2. See Interleukin 2 (IL-2) angioedema) Imatinib adverse reactions , 410 neuromuscular blocking drug-reactive , 241 Imatinib mesylate opioid-reactive , 308 dermatologic reactions , 410–411 penicillins reactive mild and dose dependent, severe reactions , 411 amoxicillanyl-and amoxicilloyllysine conjugates , oral desensitization , 411 144, 146 tyrosine kinase inhibitor structures and chronic amoxicillin reaction , 144 myeloid leukemia , 410 ampicillin molecule , 144, 145 Imipenem , 176 anti-penicillin , 144 Immediate hypersensitivity reactions, CSs

[email protected] Index 439

anaphylactic and anaphylactoid reactions , 395 and sugammadex , 271–272 anti-infl ammatory and immunosuppressive actions , 394 pharmacological activities , 241 basophil activation test , 397 pholcodine and anaphylaxis , 264–266 clinical presentation and risk factors , 394 phosphorylcholine , 265, 267 cross-reactions , 395–396 quaternary ammonium compounds , 241, 242 “delayed” positive skin response , 395 radioimmunoassay , 240 incidence and sensitization , 394 recognition, quaternary alkyl ammonium salts , 241–242 methylprednisolone , 395 serological cross-reactivity , 240–241 prick test site , 394 skin testing , 240 pseudoallergic , 394–395 speculations and evidence , 263–264 skin test concentrations , 396 Infl iximab , 376–377 succinate esters , 395 Infl iximab and TEN , 82, 83, 376 Immune reactions, mAbs Infusion reactions, mAb acute , 371 and anaphylaxis , 374 adalimumab , 377 anti-T cell , 374 anaphylaxis and anaphylactoid , 371 anti-thymocyte globulin , 382 antitumor activity , 379 cetuximab , 375 cetuximab , 375–376 denileukin diftitox , 381 chemotherapy , 379 infl iximab , 376–377 Fc engineered , 379 initial , 371 FDA-approval , 378 insulin, human , 383 genetic engineering technology , 378 rituximab , 377–378 infl iximab , 376–377 Insulin , 383 infusions , 371 Interferons (IFNs) , 380 metastatic colorectal cancer , 379 Interleukin 2 (IL-2) , 380–381 mild skin rashes , 374 International Union of Basic and clinical Pharmacology omalizumab , 374–375 (IUPHAR), 296 PEGylation , 379 Intolerances , 2–5 rituximab , 377–378 Intradermal skin tests skin , 371 description , 95, 96 syndromes , 371, 374 development , 95 Immunogenicity positive wheal and fl are reactions , 96–97 allergenic drugs , 38 small blister/bleb , 95–96 allergic cross-sensitivity , 40 sterile physiological/sterile saline , 95 allergic reaction , 39 In vitro assays antigenic heterogeneity , 38–39 cell surface activation markers , 121–122 antigenicity , 38 chemokines and skin-homing, T cells , 123–124 bioisosteres , 40 CLA ( see Cutaneous lymphocyte-associated antigen CCDs , 39 (CLA)) cephalosporin , 39 cytokines , 122–123 chemicals , 38 delayed type drug hypersensitivity reactions , 121 IgE antibody-binding determinants , 38, 39 granzyme B ELISPOT assay , 123 neuromuscular blocking drugs , 39 In vitro detection, IgE antibody tests potentially allergenic and hapten–protein complexes , binding assay in inhibition format , 102 38, 39 biotin-avidin and nucleophilic addition reactions , 101 rational drug design , 40 carbodiimides , 101–102 sensitizing drug , 39–40 immunoassay inhibition procedure , 102, 103 type I immediate hypersensitivity , 40 inhibition assays and chemical strategies , 102 Immunoglobulin E (IgE) solid phase immunoassay procedure , 101 drugs and chemicals , 242, 243 In vitro tests, penicillins allergy NMBDs basophil activation test , 153 adjacent structures, ammonium groups , 244, 245 CAST-ELISA and Flow CAST® assay , 152–153 ammonium group(s) , 243, 244 challenge testing , 153–154 ammonium ions , 244, 245 description , 151 cross-reaction with morphine , 241, 243, 252, 255 IgE antibodies , 151–152 detection , 250–254 Iodinated contrast media high cross-reactive , 244, 245 adverse reactions nitrogens , 243, 244 anaphylactoid and anaphylactic (see persistence of sensitivity , 246–247 Anaphylactoid and anaphylactic reactions) structural recognition patterns , 232, 244 delayed , 355–356

[email protected] 440 Index

range and diversity , 344 Lysosomal-associated membrane glycoprotein-3 type I allergic , 344–345 (LAMP-3), 114 Iodinated contrast media (cont.) attenuation, X-rays , 344 compounds , 344 M developments , 344 mAbs. See Monoclonal antibodies (mAbs) 2D structures and CPK models , 346, 347 mAbs therapy molecules , 345–346 approval , 371 side effects and low toxicity , 345–346 autoimmune diseases , 371–373 structures , 345 cardiovascular diseases , 371–373 water-soluble form , 344 chimeric antibodies , 370 Ionic contrast media , 344 human B cell hybridomas , 370 Isosulfan blue , 291 hybridoma technology , 370 IUPHAR. See International Union of Basic and clinical immunoglobulin , 370–371 Pharmacology (IUPHAR) mouse antigens , 370 nomenclature , 370 production, hybridoma cells , 370 K scientiﬁ c literature , 370 Kallikrein-kinin system , 67 Macrolide allergy erythromycin , 184 hypersensitivities L diagnosis , 186 β-Lactam antibiotics Ige antibodies , 185 allergenic structures , 131 Prausnitz–Kustner test , 185 antibacterials , 129–131 skin test , 185 carbapenems , 176–177 symptoms , 185 cephalosporins (see Cephalosporins) structures , 185 clavams , 177–178 Maculopapular exanthema , 29, 81 drugs classes , 129, 178 Major histocampatibility complex (MHC) HLA-B*57:01 genotype , 179 and abacavir , 72–73 intradermal tests , 179 drug interaction , 41 monobactams , 175–176 MHC class I and II molecules , 76–77 penicillins (see Penicillins) MHC-II-bound peptide , 71 LAMP-3. See Lysosomal-associated membrane Ni-peptide complex , 79 glycoprotein-3 (LAMP-3) and peptide complex , 74–75 Landsteiner , 6 Major histocompatibility complex (MHC) , 8–10 Late phase reaction , 68 MAPK. See Mitogen-activated protein kinase (MAPK) Latex , 288 Mechanisms of anaphylaxis. See Anaphylaxis Leukotriene receptors, cysteinyl , 54–55 Mediator release , 40, 43, 46 Local anesthetic allergy Meperidine , 298, 303, 312 adverse reactions (see Adverse drug reactions Methylene blue , 291 (ADRs)) Meropenem , 176 application , 281 MHC. See Major histocampatibility complex (MHC) chemical structures, esters and amides , Mitogen-activated protein kinase (MAPK) , 49 281, 282 Monobactam allergy , 175–176 diagnosis reactions , 284 Monoclonal antibodies (mAbs) penicillins , 281 adverse reactions , 372, 373 public’s perception , 281 allergic reactions , 372, 373 Local lymph node assay , 121 immune reactions (see Immune reactions, mAbs) Low-afﬁ nity IgE receptor FcεRII (CD23), 45–46 mouse antigens , 375 Lyell’s syndromes , 150, 153, 154, 156, 289, 313. next generation , 378 See also Toxic epidermal necrolysis secretion , 378 Lymphocyte transformation test systemic and cutaneous reactions , 372 anti-epileptics and β-lactam antibiotics, therapy , 370, 372, 373 120–121 Morphine allergy application and in vitro procedure , 120 antibody-mediated type I , 308 causative drug and DRESS , 120 placebo-controlled challenge , 308 delayed hypersensitivity reactions , 121 semisynthetic opioid drugs , 308–311 peripheral blood mononuclear cells , 123 synthetic opioid drugs , 311–313 T and B cells , 120 Morphine anaphylactoid reactions

[email protected] Index 441

anaphylactic to OAD and diagnostic conclusions , relationship , 257, 261 313–315 rocuronium ( see Rocuronium anaphylaxis) bronchospasm , 307 serum antibody test , 256–257 cutaneous symptoms , 316 cross-sensitivity , 249–250 “immediate anaphylactoid reactions” , 315 IgE conundrum ( see Immunoglobulin E (IgE)) tramadol , 308 skin testing , 247–250 Morphine cross-reactivity with NMBDs to diagnose NFAT. See Nuclear factor of activated T cells (NFAT) anaphylaxis to NMBDs , 252, 255 Nickel , 71 Morphine and histamine release Nitric oxide , 62 animal tissues , 303 nitric oxide synthase , 62 antibody-mediated type I allergic hypersensitivity , 303 NMBDs. See Neuromuscular blocking drugs (NMBDs) drug-induced release , 304–305 Nonimmune-mediated urticaria and angioedema hemodynamic and cutaneous changes in humans ACE , 65–66 ACS and tramadol , 308 ARBs ( see Angiotensin II receptor blockers (ARBs)) anaphylactoid reaction , 307 C1 esterase inhibitor , 65 bronchospasm and fentanyl , 307 mast cell release , 65 human heart mast cells , 307–308 Nonsteroidal anti-infl ammatory drugs (NSAIDs) in vivo and in vitro studies , 305–307 and ACE , 63 intra-arterial infusion , 308 adverse reactions , 12 mast cell degranulation in vivo , 305 angioedema , 62 meperidine and skin tests , 307 aspirin and oxicams provoke sensitivity , 9 venous blood plasma , 305 aspirin-induced asthma (see Aspirin-induced asthma) receptors , 303–304 biosynthesis, prostanoids , 324, 325 Morphine hypersensitivity birth and evolution, pharmaceutical industry , 319 adverse skin reactions , 317 classifi cation histamine , 308 anilides , 323 tramadol , 312–313 anthranilates , 323 MRP-1. See Multidrug resistance-associated protein 1 aryl and heteroaryl acetic acids , 321 (MRP-1) COX-2 selective inhibitors , 323–324 Multidrug resistance-associated protein 1 (MRP-1) , 53 oxicams , 323 Muscle relaxant anaphylaxis pharmacological and chemical properties , 321, 322 early years , 239 phenylpyrazolones , 323 neuromuscular blocking activity , 240–242 propionic acid derivatives , 321 NMBD , 253, 259, 260, 266 salicylates , 321 properties , 240 clinical classifi cation of sensitivities , 330–339 COX selectivities , 326, 328 cyclooxygenase isoforms N acetaminophen , 327 Neomycin allergy active site , 324, 326 exfoliative dermatitis and erythroderma , 194 canine COX-3 , 327–328 hydrolysis, neomycin B and C , 193–194 classifi cation , 326–327 patch test , 194 COX-1 and COX-2 , 326 sensitization , 194 fatty acid metabolism , 324 Streptomyces fradiae , 193 hypersensitivity (see Hypersensitivities) structure , 194 IgE antibodies, aspirin , 339–340 Nephrogenic systemic fi brosis , 361 over-the-counter usage , 320–321 Neuromuscular blocking drugs (NMBDs) phenylbutazone derivative and feprazone , 74 anaphylaxis (see Anaphylaxis to neuromuscular skin reactions , 4 blockers) therapeutic applications , 320 and α-Bungarotoxin (α-BT) , 272 twentieth century , 320 combining sites , 243–245 Western medicine , 319–320 cross-reactions NSAIDs. See Nonsteroidal anti-infl ammatory drugs alcuronium , 257, 260 (NSAIDs) antibody binding, ammonium ions , 257, 259 Nuclear factor of activated T cells (NFAT) , 60 clinical level , 261 decamethonium , 257 IgE recognition , 240 O in vitro inhibition, IgE antibody , 257, 258 OADs. See Opioid analgesic drugs (OADs) morphine , 241, 243, 252, 255 Omalizumab , 374–375 N -methyl determinants , 257, 258 Opioid analgesic drugs (OADs)

[email protected] 442 Index

alkaloids and codeine , 296 Pancuronium , 260, 262 allergenicity , 308–313 Patch tests classifi cation , 301–303 CSs clinical implications , 316–317 drug concentrations , 391 Opioid analgesic drugs (OADs) (cont.) intradermal testing , 391 defi nition , 296 reading and interpretation , 391–392 diagnosing reactions , 313–315 tixocortol pivalate, budesonide and hydrocortisone and histamine release (see Morphine and histamine 17-buyrate , 390–391 release) type IV reactions , 390 and IUPHAR and NC-IUPHAR , 296 vehicles , 392 morphine and narcotic , 296 drug allergy nalorphine (N -allylnormorphine) , 296 adverse drug reactions and clinical features , 97 opioid ligands binding to opioid receptors , 296, 297 COADEX classifi cation , 99 opioid peptides , 296 concentrations , 97–98 receptors , 296, 303 description , 97 structure–activity relationships healthy volunteers , 100 comparison, morphine , 298, 299 hormonal effects and upper back , 98 conformational similarities , 298–301 hypersensitivity and provocation , 97 l -methadone and enol tautomer , 300 immunomodulating drugs , 98 meperidine , 298 photopatch testing , 99–100 methadone , 299 polysensitization and scoring , 99 methoxy , 297 positive reactions , 98–100 morphine , 296–298 Stevens–Johnson syndrome , 97 nalbuphine and nitrogen , 297 Patent blue anaphylaxis oxymorphone and oxycodone , 297 adverse reactions , 291 phenylpropylamine group , 298–299 chemical structures, dyes , 291, 292 tramadol and O -desmethyltramadol , 301, 302 description , 291 two-dimensional , 297–298 immediate hypersensitivity , 292 Opioids immunoassay methods , 292 analgesics drugs (see Opioid analgesic drugs (OADs)) isosulfan blue, lymph nodes , 292 anaphylaxis methylene blue , 292–293 Administration of Papaveretum , 309 sentinel lymph node localization , 291, 292 drug-induced release of histamine , 304–305 Peak expiratory fl ow rate (PEFR) , 68 fentanyl , 312 PEFR. See Peak expiratory fl ow rate (PEFR) heroin’s , 310–311 Pegylated IFNα-2a and 2b , 380 drug allergy (see Drug allergy) Penicillins allergy hypersensitivity (see Hypersensitivities) allergic patients sera , 178 naturally occurring , 296, 308 anaphylaxis , 147 peptides , 296 antigens and determinants synthetic , 299, 311 acyl side chains , 140–143 Organoplatinum drugs 6-aminopenicillanic acid and penicoyl , 137 carboplatin and cisplatin , 405 hapten-carrier complexes , 132 cross-reactions , 408 “minor” determinants , 137–139 desensitization , 408–409 penicillanyl determinant , 139–140 IgE antibody-mediated hypersensitivity , 407 penicilloic acid, penicillamine and penamaldate , incidences of and risk factors , 407 135–137 ovarian treatment and oxaliplatin , 405 penicilloyl and benzylpenicillenic acid , patch, prick and intradermal tests , 407–408 132–133, 135 skin testing , 407, 408 side-by-side 2D and 3D structure , 132 symptoms , 405–406 side chain (R) groups , 132, 134 two-and three-dimensional structures , 405, 406 benzylpenicillin-specifi c T cell clone , 179 Osteopontin , 80 clinical reactions , 131 Oxacephems , 130 delayed reactions and lymphocyte transformation test , 157 delayed-type hypersensitivity reactions , 156–159 P desensitization , 154–156 Paclitaxel hypersensitivity hypersensitivity docetaxel , 404, 416 clinical adverse reactions , 131–132 platinum drug , 407 clinical manifestations , 132 taxanes , 400 erythematous rash , 132 P A F . See Platelet-activating factor (PAF) types , 131

[email protected] Index 443

urticarial/maculopapular rashes , 132 Pholcodine and anaphylaxis , 264–266

IgE antibodies Phospholipase A 2 (PLA2 ) , 49, 60, 62 amoxicillanyl-and amoxicilloyllysine conjugates , Phosphorylcholine , 265 144, 146 Photopatch testing , 99–100 ampicillin molecule , 144, 145 Piperacillin , 135, 158 anti-penicillin , 144 PKC. See Protein kinase C (PKC)

antisera , 141 PLA2 . See Phospholipase A2 (PLA2 ) benzylpenicilloyl-protein conjugate , 142 Platelet-activating factor (PAF) clinical relevance , 147 biological actions and pathogenic involvements , 55 complementary immune receptors , 143 biosynthesis and cellular sources , 55–57 half life of antibodies , 147 chemistry and structure–activity relationships , 55, 56 haptenic determinants, allergenic drugs , 141 health and disease , 57–58 immediate allergic reactions , 142 measurement in laboratory , 58 immune mechanisms , 131 receptor , 58 patients sera , 144 Platinum drug hypersensitivity reagins mediating skin , 141 cross-reactions , 408 thiazolidine ring , 144 desensitization , 408 IgE-based sensitivity , 147 IgE antibody-mediated hypersensitivity , 407 in vitro tests incidences of and risk factors , 407 basophil activation test , 153 mechanisms and diagnosis , 407 CAST-ELISA and Flow CAST® assay , 152–153 patch, prick and intradermal tests , 407–408 challenge testing , 153–154 skin testing , 407, 408 description , 151 symptoms , 405–406 IgE antibodies , 151–152 Polypeptides, anesthesia and surgery immunological reactions , 178 aprotinin ( see Aprotinin allergy) β-lactam antibiotics, 178 latex , 288–289 liver injury , 159 protamine (see Protamine allergy) negative history , 149 Povidone-iodine allergy Phadia ImmunoCAP® , 152 hypersensitivity reactions , 230 Pre-Pen® , 148 IgE antibodies , 230 risk factors , 148 polyvinyl-pyrrolidone (PVP) , 230 skin testing PPIs. See Proton pump inhibitors (PPIs) allergic sensitivity , 150–151 PPIs allergy historical perspective , 148–149 former drug , 421 persistence of reactivity , 151 IgE antibodies , 421 reading tests , 150 PPIs anaphylaxis reagents , 149–150 omeprazole and pantoprazole , 420 T cell recognition , 71 oral desensitization , 421, 424 type I IgE-mediated , 178 skin testing , 421 Peri-operative anaphylaxis , 236, 238, 270, 276, 278 PPIs cross-reactions Pharmacogenomics and drug allergy, ADRs lansoprazole and rabeprazole , 421, 423 abacavir , 10 skin testing , 424 ADR studies , 10 PPIs hypersensitivity AHS , 10 anaphylaxis , 420 clinical response , 9 autoimmune reactions , 420 cytochrome P450 2D6/CYP2D6 , 9–10 cutaneous reactions , 420 drug therapy , 9 desensitization , 421 genetic polymorphisms , 9 diagnosis , 422 genomics , 9 DRESS case , 421 HLA-B*1502 allele , 10 erythrodermic reactions , 421 HLA-B*5701 screening , 10 immediate reactions , 421 MHC ancestral haplotype , 10 lansoprazole and rabeprazole cross-reaction , 421 myelotoxicity , 10 maculopapular eruptions and pruritus , 421 single genes , 11 omeprazole and pantoprazole , 421 SLCO1B1 gene , 10–11 patient allergic to omeprazole , 421 Stevens–Johnson syndrome , 10 quantitative hapten inhibition application , 421 Philadelphia chromosome skin testing , 421 BCR and ABL genes , 409–410 Premedication for chemotherapeutic drugs cytogenetic abnormality , 410 platinum drugs , 408, 409 EGFR and imatinib , 410 taxanes , 404 receptor and non-receptor kinases , 410 Premedication for contrast media

[email protected] 444 Index

corticosteroids , 360 management , 337 effectiveness , 360 NSAIDs , 336 histamine H1 antagonists , 360 risks , 337 injection , 360 skin testing , 336, 337 Premedication for contrast media (cont.) lower osmolality , 360 nonionic agent , 360–361 Q prednisone and diphenhydramine , 360 Quinolone allergy risk , 361 challenge tests , 221 Pristinamycin , 200 cross-reactions , 224–225 Propofol delayed reactions , 225 administration , 276 hypersensitivities allergy adverse reactions , 220 chemical structure , 277 antibacterials developments , 220 egg and soybean oil , 277–278 cutaneous reactions , 220 and anaphylaxis , 276–277 immediate , 220 formulation , 276 prevalences , 220 skin tests , 277 IgE antibodies (see IgE antibodies) Protamine allergy skin tests , 220–221 adverse reactions , 285 structure , 218–219 arginine-rich nuclear proteins , 284 cardiorespiratory effects , 284 diagnosis , 286 R guanidine groups , 284 RAST , 100 immune-mediated hypersensitivity reaction, Reactions to contrast media 285–286 adverse reactions Proteasome inhibitors activation of kinin system , 354 bortezomib , 412–414 acute (immediate) reactions , 347–348 cancer cells and β-subunits , 412 anaphylactoid and anaphylactic reactions (see cell-cycle progression and cellular Anaphylactoid and anaphylactic reactions) proteins , 412 biphasic reactions , 350–351 leukemia-and lymphoma-derived cells , 412 delayed reactions , 355–356 second generation , 415 histamine release , 352 single-capped proteasome complex/30S incidence of reactions , 349–350 double-capped, 412, 413 involvement of IgE , 353 Protein kinase C (PKC) , 49, 60 late reactions , 348–349 Protein tyrosine phosphatase receptor type C mechanisms of reactions , 351 (PTPRC) , 114 risk factors , 350 Proton pump inhibitors (PPIs) diagnosis and study activation , 422 challenge tests , 359–360 anaphylaxis (see PPIs anaphylaxis) IgE antibody tests (see IgE antibodies, tests) benzimidazole chemical structures , 419, 420 mediators release , 358–359 cross-reactions (see PPIs cross-reactions) skin tests , 356–357 esomeprazole , 419 gadolinium-based contrast agents (see Gadolinium gastric acid production , 419 contrast media) gastroesophageal reﬂ ux disease and asthma , 423 iodinated ( see Iodinated contrast media) hypersensitivity (see PPIs hypersensitivity) non-renal adverse , 348 imidazopyridine drugs , 419 usage and safety , 346–347 patterns, cross-reactivity , 423 Reading and interpretation patch testing , 391–392 safety , 423 Receiver-operating characteristic (ROC) , Provocation testing. See Challenge tests 104, 116, 126 Pseudoallergy , 2–5 Renin-Angiotensin system , 66 Psoriasis , 26–29, 80 Respiratory reactions, NSAID PTPRC. See Protein tyrosine phosphatase receptor asthmatic attacks , 330 type C (PTPRC) basophil activation test , 332 Pyrazolone hypersensitivity challenge tests , 331 anaphylactic shock reactions , 335–336 cysteinyl leukotrioenes release , 332 BAT and CAST test , 337 desensitization , 333 IgE antibodies , 336 genetic polymorphisms , 331

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15-HETE generation , 332 aminoglycoside antibiotics and positive control , 94 management , 332–333 forearm/back , 94 observations , 330–331 glycerin and penicillins , 95 pharmacologic mechanism , 330 intradermal and tendency , 95 symptoms and exposure , 331 lancet/fi ne-gauge needle , 94 Ribostamycin allergy , 197–198 late-phase reactions , 95 Rifamycin allergy mean diameter and false positives , 95 adverse reactions , 188 wheal and erythema (fl are) reaction , 94–95 bacterium products , 187–188 Skin tests biosynthesis , 187–188 CSs IgE antibodies , 188 concentrations , 396 structure , 188 identifi cation , 396 Risk factors, drug allergy methylprednisolone delayed , 395 atopic patients , 8 positive, prednisolone , 395 cross-reactivity , 9 prednisolone-allergic patient , 395 epidemiological data , 8 prick and intracutaneous , 396, 397 genetics and ethnicity , 8 retention , 392 HIV infection , 8 vasoconstriction , 392 hypersensitivity reactions , 8 drug allergy immune responses , 9 antihistamines and high-risk patients , 94 infectious agent , 8 dextran allergy , 281 intramuscular administration , 9 diagnosis , 247 patient-related and drug-related , 8 DMSO , 94 sensitization and severe reactions , 9 heparin allergy , 289–290 Rituximab , 377–378 hypersensitivity , 93, 256 ROC. See Receiver-operating characteristic (ROC) intradermal , 95–97 Rocuronium anaphylaxis late and delayed reactions , 94 and alcuronium , 262 methodology , 247–249 ammonium ions structures , 262 NMBDs (see Neuromuscular blocking drugs cross-reactivity detection, IgE antibodies , 261 (NMBDs)) immunochemical investigation , 261 patch tests (see Patch tests, drug allergy) Norwegian Medicines Agency , 261–262 prick and intradermal , 248–249 safety , 261 propofol , 277 structural features , 263 skin prick test method , 94–95 sugammadex (see Sugammadex) tablet/capsule form , 94 thiopentone anaphylaxis , 275 penicillins allergy S allergic sensitivity , 150–151 Safety of PPIs historical perspective , 148–149 drugs and fractures , 423 reagents , 149–150 FDA , 423 S1P. See Sphingosine-1-phosphate (S1P) oral antiplatelet drug clopidogrel , 423 Sphingosine-1-phosphate (S1P) Second generation proteasome inhibitors bioactive signaling phospholipid , 60, 61 carfi lzomib , 415 Fyn protein activation , 60 MLN9708 , 415 Sphk1 and Sphk2 , 61

salinosporamide A and omuralide , 415 S1P2 receptor , 61 Semisynthetic opioid drugs Stevens–Johnson syndrome (SJS) AMA drug evaluations , 310 adverse cutaneous reactions , 82 cyclohexenyl ring, antibody binding , 309 allopurinol-induced , 73 direct binding immunoassays , 309 carbamazepine-induced , 73 heroin , 310–311 granulysin , 82 IgE antibodies , 308–309 keratinocyte necrosis , 82 intradermal tests and N -cyclopropylmethyl , 309 management , 83 quantitative hapten inhibition studies , 309, 310 mucosal oriﬁ ces , 82 skin testing , 309 necrotic epidermis/skin detachment greater , 82 two-dimensional molecular models, morphine , operative mechanisms , 82 309, 311 pathogenetic mechanism , 82 Serum sickness , 24, 85–87 type IV drug hypersensitivity , 33–34 SJS. See Stevens–Johnson syndrome (SJS) Streptomycin , 197 Skin prick test method Succinylcholine , 250–252, 257–259

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Sugammadex diagnosis delayed drug allergy , 118 allergy , 272 drug-reaction , 123 ammonium group , 271 and ELISPOT assay , 122–123 association constant , 269 helper cell responses , 78 Sugammadex (cont.) lymphocyte transformation test , 120–121 binding to rocuronium , 267, 269 penicillin antigens , 158–159 cyclodextrins and two-dimensional structure , skin testing, aminopenicillins , 156 267–271 TCR. See T cell receptor (TCR) IgE antibodies , 271–272 Teicoplanin , 188 oligosaccharide , 267 TEN. See Toxic epidermal necrolysis (TEN) space-fi lling CPK molecular model , 269, 270 Tetracycline allergy thiol-ether group , 269 adverse reactions , 186–187 treatment , 271 structure , 187 Sulfamethoxazole , 201–209, 215, 216 Th cell subsets , 78 Sulfonamide allergy Thiopentone anaphylaxis adverse reactions , 202 administration , 274 chemotherapeutic and rheumatologic agents , 200 clinical features and epidemology , 273 cross-reactions (see Cross-reactions, of sulfonamides) diagnosis , 273–274 2D and 3D CPK space-fi lling molecular models , histamine release, leukocyte , 274–276 204, 205 hypersensitivity reactions , 273 delayed reactions serum IgE antibody tests , 274 bioactivation , 204–205 skin testing , 274 hematological malignancies , 206 ultrashort acting barbiturate , 273 interpretations , 206–207 Thrombocytopenia , 23, 84, 85, 289, 291, 401, 414 life-threatening reactions , 204 TNF and TEN , 82, 83, 376 metabolism , 204, 206 TNF-like weak inducer of apoptosis (TWEAK) T cells , 206 pathway , 82 toxicity , 204 TNF-related apoptosis inducing ligand (TRAIL) HIV , 202, 208, 209 pathway , 82 IgE antibody-mediated reactions , 202–204 Toxic epidermal necrolysis (TEN). See also Lyell’s lethal toxidermias , 202 syndrome structure , 200–201 carbamazepine-induced , 73 type I , 202 Fas-FasL killing , 82 Sweet’s syndrome , 414 granulysin , 82 Synthetic opioid drugs immune mechanisms and biomarkers , 82 fentanyl and methadone , 312 keratinocyte necrosis , 82 immunological cross-reactivity , 312, 313 management , 83 meperidine , 311–312 necrotic epidermis/skin detachment greater , 82 “suspected narcotic allergy” , 312 operative mechanisms , 83 tramadol , 312–313 pathogenetic mechanism , 82 type IV drug hypersensitivity , 33–34 TRAIL pathway. See TNF-related apoptosis inducing T ligand (TRAIL) pathway Taxane hypersensitivity Tramadol hypersensitivity , 312–313 acute reactions and cremophor , 400 Trimethoprim allergy description , 400 hypersensitivites , 213–214 desensitization , 405 T-cells , 218 docetaxel and paclitaxel , 400 type I IgE antibody-mediated reactions premedication , 404 CPK sapce-fi lling models , 217 T cell receptor (TCR) diagnostic procedure , 214 CTLs/NK cells , 83 inhibition of , 215 MHC–peptide complex , 74–75 skin testing , 214 Ni-reactive T cells , 71 structure , 215 Th2 cell , 42 Tryptases T-cells α and β , 108 antigen-specifi c , 158 and drug allergy , 109 benzylpenicillin-specifi c , 179 genes , 107–108 chemokines and skin-homing , 123–124 mature and total , 108–109 cytokines , 122 Tumor lysis syndrome , 371, 415 delayed-type hypersensitivity , 143–144 Tumor necrosis factor (TNF)

[email protected] Index 447

abacavir-specifi c CD8+ T cells secrete , 72 biosynthesis and metabolism , 47 anti-TNF monoclonal antibody infl iximab , 62 granules , 47 central role , 77 humans—enterochromaffi n-like cells , 47 naive T cells , 77, 78 2-(imidazol-4-yl)ethylamine , 46–47 psoriasis , 80 PAF ( see Platelet-activating factor (PAF)) treatment , 80 Type IV hypersensitivity drug reactions TWEAK and TRAIL , 82 AGEP , 29–30 TWEAK pathway. See TNF-like weak inducer allergic contact dermatitis of apoptosis (TWEAK) pathway acute , 25, 27 Type I hypersensitivity drug reactions chemicals , 25 anaphylaxis (see Drug anaphylaxis) nickel , 25, 27 causes , 17 classifi cations , 25 description , 16 consequences , 24 IgE antibodies , 17 delayed cutaneous ADRs (see Delayed cutaneous late phase reactions , 16 ADRs) urticaria and angioedema , 21–23 description , 24 Type I IgE antibody-mediated reactions DRESS , 30–31 mast cells and basophils , 107 erythema multiforme , 32–33 trimethoprim allergy FDE , 31–32 CPK space-fi lling models , 217 maculopapular exanthema , 29 diagnostic procedure , 214 prototypic tuberculin test , 24 inhibition of , 215 psoriasis , 26–29 skin testing , 214 “recall” antigens , 76 structure , 215 SJS and TEN , 33–34 Type II hypersensitivity drug reactions T helper cell responses and Th17 , 78–79 acute agranulocytosis , 85 Tyrosine kinase inhibitors aminopyrine-induced agranulocytosis , 85 description , 409 antibody–antigen complexes , 84 gefi tinib and erlotinib , 411–412 cefotetan , 84 imatinib mesylate , 410–411 cephalosporins , 84 philadelphia chromosome , 409–410 chimeric human–mouse monoclonal antibody , 85 clozapine-induced agranulocytosis , 85–86 U DIIHA , 84 Urticaria , 21, 62 drug-dependent antibodies , 84 Urticaria angioedema, NSAIDs drug-induced thrombocytopenia , 84 aspirin-induced asthma , 333 erythrocytes , 84 cross-reaction heparin–PF4 complex , 85 chronic idiopathic , 333 in vitro hapten inhibition experiments , 84 COX-2 inhibitors , 333 platelet antigens , 85 cross-sensitivity , 333 thrombocytopenia , 84 measurements , 333–334 “unifying hypothesis” , 84 oral provocation , 333, 334 Type III hypersensitivity drug reactions skin testing , 334 amoxicillin–clavulanic acid , 24 multiple reactions , 335 antigen, subsequent immunological events single reactions and time , 86 description , 335 β-lactam drugs, 86 pyrazolones , 335–337 DNA/anti-DNA/complement , 23 drug immune complexes , 87 drug-induced reaction , 24 V intravascular and extravascular equilibration , 87 Vancomycin red man syndrome , 190–191 Jarisch–Herxheimer reaction , 23–24 Vasculitis , 24, 87, 121, 192, 376, 401 penicillin-induced reaction , 24 Vecuronium , 260, 262 reticuloendothelial cells , 87 Vesicular monoamine transporter 2 (VMAT2) , 47 serum sickness , 24, 86 VMAT2. See Vesicular monoamine transporter 2 small-vessel vasculitis patients , 87 (VMAT2) vasculitis induced , 87 vasoactive amines liberation , 86 Type I immediate allergic response W cysteinyl leukotrienes (see Cysteinyl leukotrienes) WAO. See The World Allergy Organization histamine (see also Histamine, receptors) (WAO) anionic proteoglycans—heparin , 47 The World Allergy Organization (WAO) , 3, 5

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