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Seminar

Graft-versus-

James L M Ferrara, John E Levine, Pavan Reddy, Ernst Holler

Lancet 2009; 373: 1550–61 Haemopoietic-cell transplantation (HCT) is an intensive used to treat high-risk haematological malignant Published Online disorders and other life-threatening haematological and genetic . The main complication of HCT is graft- March 12, 2009 versus-host disease (GVHD), an immunological disorder that aff ects many organ systems, including the DOI:10.1016/S0140- , liver, , and . The number of patients with this complication continues to grow, and 6736(09)60237-3 many return home from transplant centres after HCT requiring continued treatment with immunosuppressive drugs University of Michigan, and Internal that increases their risks for serious and other complications. In this Seminar, we review our understanding , and Marrow of the risk factors and causes of GHVD, the cellular and networks implicated in its pathophysiology, and Transplantation Program, current strategies to prevent and treat the disease. We also summarise supportive-care measures that are essential for Ann Arbor, MI, USA management of this medically fragile population. (Prof J L M Ferrara MD, Prof J E Levine MD, P Reddy MD); and and Introduction of mounting an eff ective response to eliminate the Department of Haematology/ The number of allogeneic haemopoietic-cell trans- transplanted cells.3 We know now that the immuno- , University , plantations (HCTs) continues to rise, with more than logically competent cells are T cells and that GVHD can Regensburg, Germany (Prof E Holler MD) 25 000 procedures undertaken annually. The graft-versus- develop in various clinical settings when tissues Correspondence to: leukaemia or graft-versus-tumour eff ect during this containing T cells (blood products, bone marrow, and Prof James L M Ferrara, procedure eff ectively eradicates many haema tological solid organs) are transferred from one person to another University of Michigan, malignant diseases.1 Development of novel strategies who is not able to eliminate those cells.4,5 Patients whose 1500 East Medical Center Drive, that use donor leucocyte infusions, non-myeloablative immune systems are suppressed and who receive white 6303 CCC, Ann Arbor, MI 48109-5942, USA conditioning, and umbilical-cord blood transplantation blood cells from another individual are at especially high [email protected] has helped expand the indications for allogeneic HCT risk for the disease. over the past few years, especially for older patients.2 GVHD arises when donor T cells respond to Improvements in infectious prophylaxis, immuno- genetically defi ned on host cells. The most suppressive treatments, suppor tive care, and DNA-based important proteins are human leucocyte tissue typing have also contributed to enhanced outcomes (HLAs),2,6,7 which are highly polymorphic and are after the technique.1 Yet, the major complication of encoded by the major histocompatibility complex allogeneic HCT—graft-versus-host disease (GVHD)— (MHC). Class I HLA (A, B, and C) proteins are expressed remains lethal and limits use of this important procedure.2 on almost all nucleated cells of the body at various In view of current trends, the number of transplants densities. Class II proteins (DR, DQ, and DP) are mainly from unrelated donors is expected to double within the expressed on haemopoietic cells (B cells, dendritic cells, next 5 years, substantially increasing the population of and ), but their expression can be induced on patients with GVHD. In this Seminar, we review advances many other cell types after infl ammation or injury. made in identifi cation of genetic risk factors and High-resolution DNA typing of HLA genes with pathophysiology of this major HCT complication and its PCR-based techniques has now largely replaced earlier prevention, diagnosis, and treatment. methods. The frequency of acute GVHD is directly related to the degree of mismatch between HLA Cause and clinical features proteins,8,9 and thus ideally, donors and recipients are 50 years ago, Billingham formulated three requirements matched at HLA A, B, C, and DRB1 (referred to as for development of GVHD: (1) the graft must contain 8/8 matches), but mismatches can be tolerated for immunologically competent cells; (2) the recipient must umbilical-cord blood grafts (see Clinical features of express tissue antigens that are not present in the acute GVHD).10–12 transplant donor; and (3) the patient must be incapable Despite HLA identity between a patient and donor, about 40% of recipients of HLA-identical grafts develop systemic acute GVHD that needs treatment with Search strategy and selection criteria high-dose . This disorder is due to genetic We searched PubMed and Medline with the term ‘‘GVHD’’ diff erences that lie outside the HLA loci and that encode and cross-referenced it with the following words: ‘‘clinical’’, proteins referred to as minor histocompatibility antigens. ‘‘’’, ‘‘MHC’’, ‘‘HLA antigens’’, ‘‘biology’’, and Some minor histocompatibility antigens, such as HY ‘‘’’. We included mostly peer-reviewed original and HA-3, are expressed on all tissues and are targets for 13 and review journal articles published within the past both GVHD and graft-versus-leukaemia. Others, such decade, except for seminal articles that initially described as HA-1 and HA-2, are expressed most abundantly on the clinical features. All non-peer-reviewed manuscripts, haemopoietic cells (including leukaemic cells) and could, supplements, and textbooks were excluded. therefore, induce an enhanced graft-versus-leukaemia eff ect with diminished GVHD.13,14

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Panel 1: Acute GVHD symptoms Skin • Maculopapular skin Upper gastrointestinal tract • Nausea, anorexia, or both, and positive histological fi ndings Lower gastrointestinal tract • Watery diarrhoea (≥500 mL) • Severe • Bloody diarrhoea or ileus (after exclusion of infectious causes) Liver • Cholestatic hyperbilirubinaemia

Polymorphisms in both donors and recipients of cytokines that have a role in the classic cytokine storm of GVHD (see Pathophysiology of acute GVHD) have been implicated as risk factors for the disorder.15 Tumour necrosis factor (TNF) α, 10, and interferon γ variants have correlated with GVHD in some, but not all, studies.16–18 Genetic polymorphisms of proteins connected with innate , such as nucleotide oligomerisation Figure 1: Acute GVHD of the skin (grade I) domain 2 and keratin 18 receptors, have also been Photograph courtesy of J Levine. associated with the disorder.19–22 Future strategies to Gastrointestinal-tract involvement of acute GVHD identify the best possible transplant donor will probably usually presents as diarrhoea but can also include incorporate both HLA and non-HLA genetic factors. , anorexia, abdominal pain, or a combination when severe.29 Diarrhoea in GVHD is secretory and Clinical features of acute GVHD usually voluminous (>2 L per day). , which has On the basis of early work, acute GVHD was defi ned as poor prognosis, happens as a result of mucosal arising before day 100 post-transplant, whereas chronic ulceration,31 but patchy involvement of mucosa generally disease happened after that time.23–25 This defi nition is leads to a normal appearance on endoscopy.32 Radiological far from satisfactory, and a National Institutes of Health fi ndings of the gastrointestinal tract include luminal classifi cation includes late-onset acute GVHD (after dilatation with thickening of the wall of the small bowel day 100) and an overlap syndrome with features of both (ribbon sign on CT) and air or fl uid levels suggestive of acute and chronic disorder.26,27 Late-onset acute GVHD an ileus.28 Histological features include patchy ulcerations, and the overlap syndrome arise with greater frequency apoptotic bodies in the base of crypts, crypt abscesses, after reduced-intensity conditioning, an increasingly and loss and fl attening of surface epithelium.33 widespread technique (see Prevention of GVHD). Panel 1 caused by GVHD can be diffi cult to shows the clinical manifestations of acute GVHD. In a distinguish from other causes of liver dysfunction after comprehensive review, Martin and colleagues noted that bone-marrow transplantation, such as veno-occlusive at onset of acute GVHD, aff ected regions included skin disease, toxic drug eff ects, viral , , or iron (81% of patients), gastrointestinal tract (54%), and overload. The histological features of hepatic GVHD are liver (50%).23 endothelialitis, lymphocytic infi ltration of the portal Skin is most frequently aff ected and is usually the fi rst areas, pericholangitis, and bile-duct destruction.34,35 organ involved, generally coinciding with engraftment of However, biopsy specimens of liver are taken rarely donor cells. The characteristic maculopapular rash is because early after transplantation pruritic and can spread throughout the body, sparing the greatly increases the risks of the biopsy procedure, scalp (fi gure 1). In severe cases the skin can and making the diagnosis of GVHD one of exclusion. ulcerate.28 Apoptosis at the base of epidermal rete pegs is Severity of acute GVHD is ascertained by the extent of a characteristic pathological fi nding. Other features involvement of the three main target organs. Overall include dyskeratosis, exocytosis of , satellite grades are I (mild), II (moderate), III (severe), and lymphocytes adjacent to dyskeratotic epidermal IV (very severe). Severe GVHD has poor prognosis, with keratinocytes, and perivascular lymphocytic infi ltration 25% long-term survival (5 years) for grade III disease and in the .29,30 5% for grade IV.36 www.thelancet.com Vol 373 May 2, 2009 1551 Seminar

somewhat protean and typically of an autoimmune Panel 2: Chronic GVHD symptoms nature. Clinical signs are generally seen fi rst in the buccal Skin mucosa (fi gure 2). New consensus criteria for diagnosis 26 Dyspigmentation, new-onset alopecia, poikiloderma, lichen and staging of chronic GVHD have been developed. planus-like eruptions, or sclerotic features Pathophysiology of acute GVHD Nails Two important principles should be considered with Nail dystrophy or loss respect to the pathophysiology of acute GVHD. First, Mouth the disease is indicative of exaggerated but typical Xerostomia, ulcers, lichen-type features, restrictions of infl ammatory mechanisms mediated by donor mouth opening from sclerosis lymphocytes infused into the recipient, in whom they function appropriately in view of the foreign Eyes environment they encounter. Second, the recipient’s Dry eyes, sicca syndrome, cicatricial conjunctivitis tissues that stimulate donor lymphocytes have usually Muscles, fascia, joints been damaged by underlying disease, previous 29 Fasciitis, myositis, or joint stiff ness from contractures infections, and the transplant conditioning regimen. As a result, these tissues produce molecules such as Female genitalia proinfl ammatory cytokines and chemokines, which Vaginal sclerosis, ulcerations increase expression of key receptors on Gastrointestinal tract -presenting cells (APCs), thereby enhancing Anorexia, weight loss, oesophageal web or strictures cross-presentation of polypeptide proteins (eg, minor histocompatibility antigens) to the donor immune cells Liver that mediate GVHD.42–45 Jaundice, transaminitis Mouse models have been central to identifi cation and Lungs understanding of pathophysiological mechanisms of Restrictive or obstructive defects on pulmonary function GVHD, and work undertaken in dogs has been vital for tests, obliterans, pleural eff usions development of clinically useful strategies for GVHD prophylaxis and treatment advances in donor leucocyte Kidneys infusions.36,46,47 Largely on the basis of these experimental Nephrotic syndrome (rare) data, progression of acute GVHD can be summarised in Heart three sequential steps or phases: (1) activation of APCs; (2) donor T-cell activation, proliferation, diff erentiation, Pericarditis and migration; and (3) target tissue destruction Marrow (fi gure 3). Thrombocytopenia, anaemia, The fi rst step entails activation of APCs by the underlying disease and the HCT conditioning regimen. Damaged host tissues respond by producing so-called Prevalence of acute GVHD is directly related to the danger signals, including proinfl ammatory cytokines (eg, degree of mismatch between HLA proteins. It ranges TNFα and 1 and 6), chemokines, and from 35–45% in recipients of full-matched sibling donor amplifi ed expression of adhesion molecules, MHC grafts8,9 to 60–80% in people receiving one-antigen antigens, and costimulatory molecules on host APCs.42,48–50 HLA-mismatched unrelated-donor grafts.6,37–39 The same Findings of a report showed that 1 week after HCT, amount of mismatch causes diminished GVHD with increased amounts of TNFα 1—a surrogate umbilical-cord blood grafts, and frequency of acute marker for TNFα—correlated strongly with later GVHD is low after transplantation of partly matched development of GVHD.51 Injury to the gastrointestinal umbilical-cord blood units (35–65%).12 tract from conditioning is especially important because it allows for systemic translocation of additional Clinical features of chronic GVHD infl ammatory stimuli, such as microbial products Chronic GVHD is the major cause of late non-relapse including or other pathogen-associated after HCT.40 Its presentation can be progressive molecular patterns, that further enhance activation of (active or acute GVHD merging into chronic), quiescent host APCs.49 (acute disease that resolves completely but is followed The secondary lymphoid tissue in the gastrointestinal later by chronic), or de novo. Older recipient age and a tract is probably the initial site of interaction between history of acute GVHD are the greatest risk factors for activated APCs and donor T cells.52 These observations chronic disease.41 Therefore, strategies for acute GVHD have led to an important clinical strategy to diminish prevention could help to prevent chronic disease. Panel 2 acute GVHD by reducing the intensity of the conditioning shows that manifestations of chronic GVHD are regimen.53,54 Experimental GVHD can also be decreased

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by manipulation of distinct subsets of APCs.55,56 Furthermore, non-haemopoietic stem cells, such as mesenchymal stromal cells, can reduce allogeneic T-cell responses and ameliorate GVHD, although the mechanism for such inhibition remains unclear.57 The idea that amplifi ed activation of host APCs increases the risk for acute GVHD unifi es several seemingly disparate clinical associations with that risk, such as advanced stages of malignant disease, more intense transplant conditioning regimens, and history of viral infection. APCs detect infections with receptors on their cell surfaces, such as Toll-like receptors, which recognise conserved molecular patterns of microbes.27,58 Toll-like receptors specifi c for viral DNA or RNA activate APCs and could boost GVHD, providing a potential mechanistic basis for enhanced disease associated with viral infections such as cytomegalovirus.59 The core of the graft-versus-host reaction is the second step, in which donor T cells proliferate and diff erentiate in response to host APCs (fi gure 3). The danger signals generated in the fi rst phase augment this activation, at least in part, by increasing expression of costimulatory molecules.60 Blockade of costimulatory pathways to prevent GVHD is successful in animal models, but this approach has not yet been tested in large clinical trials.2 In mouse models, in which genetic diff erences between donor and recipient strains can be tightly controlled, CD4+ cells induce acute GVHD to MHC class II Figure 2: Lichenoid changes of buccal mucosa in chronic GVHD diff erences and CD8+ cells induce acute disease to class I Photograph courtesy of J Ferrara and J Levine. diff erences.61,62 In most HLA-identical HCTs, both CD4+ and CD8+ subsets respond to minor histocompatibility with this cytokine could unintentionally stop development antigens and can cause GVHD in HLA-identical of long-term tolerance after allogeneic HCT.66 procedures. Interferon γ has many functions and can either amplify Regulatory T cells can suppress proliferation of or reduce GVHD.67,68 It could boost disease by increasing conventional T cells and prevent GVHD in animal expression of molecules such as chemokine receptors, models when added to donor grafts containing con- MHC proteins, and adhesion molecules; it also raises ventional T cells,63 but use of regulatory T cells in clinical the sensitivity of monocytes and to stimuli acute GVHD will need enhanced techniques to identify such as lipopolysaccharide and accelerates intracellular and expand them. Natural killer T-cell 1.1+ subsets from cascades in response to these stimuli.69 Early polarisation the host and donors have also been shown to modulate of donor T cells so that they secrete less interferon γ and acute GVHD.64 In a clinical trial of total lymphoid more can also attenuate experimental acute irradiation (as conditioning), GVHD was reduced GVHD.70 Interferon γ might amplify GVHD by direct signifi cantly and host natural killer T-cell function was damage to epithelium in the gastrointestinal tract and amplifi ed.65 skin and by induction of by Activation of immune cells results in rapid intracellular generation of nitric oxide.71 By contrast, this cytokine biochemical cascades that induce transcription of genes could suppress GVHD by hastening apoptosis of for many proteins, including cytokines and their activated donor T cells.68,72 This complexity means receptors. T-helper 1 cytokines (interferon γ, , manipulation of interferon γ could have diverse eff ects and TNFα) are released in large amounts during acute in vivo, making the cytokine a challenging target with GVHD. Production of interleukin 2 by donor T cells respect to therapeutic intervention. remains the main target of many current clinical has a key role in suppression of immune therapeutic and prophylactic approaches to GVHD, such responses, and clinical data suggest it might regulate as cyclosporine, tacrolimus, and monoclonal acute GVHD.17 Transforming growth factor β, another directed against this cytokine and its receptor.9 However, suppressive cytokine, can subdue acute GVHD but emerging data indicate an important role for interleukin 2 exacerbate chronic disease.73 Thus, timing and duration in the generation and maintenance of CD4+CD25+ of secretion of any given cytokine could establish the regulatory T cells, suggesting that prolonged interference specifi c eff ects of that molecule on GVHD severity. www.thelancet.com Vol 373 May 2, 2009 1553 Seminar

Conditioning: tissue damage

Small (1) Host APC Host intestine activation tissues

TNFα IL1 LPS LPS

IFNγ TNFα IL1 Treg

Target cell apoptosis Treg CD4 TNFα CTL IL1 Th1 (2) Donor T-cell CD8 activation CTL CD8 (3) Cellular and CTL inflammatory effectors

Figure 3: Pathophysiology of acute GVHD IL 1=interleukin 1. IFN γ=interferon γ. LPS=lipopolysaccharide. Treg=. Th1=T-helper 1 cell. CTL=cytotoxic T .

The third eff ector phase of the graft-versus-host process for homing of donor T cells to Peyer’s patches during (fi gure 3) is a complex cascade of cellular mediators (such intestinal GVHD.52,76,77 as cytotoxic T lymphocytes and natural killer cells) and Microbial products such as lipopolysaccharide, which soluble infl ammatory agents (eg, TNFα, interferon γ, leak through damaged intestinal mucosa or skin, can interleukin 1, and nitric oxide).2,29 These molecules work stimulate secretion of infl ammatory cytokines through synergetically to amplify local tissue injury and further Toll-like receptors.49,78 The gastrointestinal tract is promote infl ammation and target tissue destruction. especially susceptible to damage from TNFα, and the The cellular eff ectors of acute GVHD are mainly gastrointestinal tract has a major role in amplifi cation cytotoxic T lymphocytes and natural killer cells.49 Cytotoxic and propagation of the cytokine storm characteristic of T lymphocytes that prefer to use the Fas and FasL pathway acute GVHD.49 TNFα can be produced by both donor and of target lysis seem to predominate in GVHD liver host cells and it acts in three diff erent ways: (1) it activates damage (hepatocytes express large amounts of Fas) APCs and enhances alloantigen presentation; (2) it whereas cells that use the perforin and granzyme recruits eff ector cells to target organs via induction of pathways are more important in the gastrointestinal tract infl ammatory chemokines; and (3) it directly causes and skin.2,74 Chemokines direct migration of donor T cells tissue necrosis (as its name suggests).79–81 from lymphoid tissues to the target organs in which they cause damage. infl ammatory 1α and Prevention of GVHD other chemokines (such as CCL2–CCL5, CXCL2, CXCL9, On the basis of evidence from animal models for the CXCL10, CXCL11, CCL17, and CCL27) are overexpressed central role of T cells in initiation of GVHD, many clinical and enhance homing of cellular eff ectors to target organs studies of T-cell depletion as prophylaxis for the disease during experimental GVHD.75 Expression of integrins, were undertaken in the 1980s and 1990s. Three main such as α4β7 and its ligand MADCAM1, is also important depletion strategies were studied: (1) negative selection

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of T cells ex vivo; (2) positive selection of CD34+ stem In retrospective studies, rabbit anti- globulin cells ex vivo; and (3) antibodies against T cells in vivo.82 reduced the frequency of GVHD in related-donor Most approaches showed substantial limitation of both haemopoietic stem-cell transplant recipients without acute and chronic GVHD.83–85 Unfortunately, the lowest seeming to enhance survival.98,99 In patients receiving frequency of severe GVHD was off set by high rates of unrelated-donor haemopoietic stem cells, addition of graft failure, relapse of malignant disease, infections, anti-lymphocyte globulin to standard GVHD prophylaxis and Epstein-Barr -associated lymphoproliferative prevented severe GVHD eff ectively but did not result in disorders. Negative-selection purging strategies with better survival because of increased infections.86 In a various antibodies against T cells achieved similar long-term follow-up study, however, pretransplant long-term results irrespective of the breadth of anti-thymocyte globulin provided signifi cant protection specifi city.86–88 Findings of one large registry study showed against extensive chronic GVHD and chronic that purging techniques that used antibodies with broad dysfunction.100 specifi cities produced inferior leukaemia-free survival The primary pharmacological strategy to prevent GVHD than standard immunosuppression in patients receiving is inhibition of the cytoplasmic calcineurin, unrelated donor transplants.89 which is important for activation of T cells. The calcineurin Several research groups have investigated partial T-cell inhibitors cyclosporine and tacrolimus have similar depletion, either by elimination of specifi c T-cell subsets mechanisms of action, clinical eff ectiveness, and toxic (eg, CD8+) or by of the dose of T cells present in eff ects, including hypomagnesaemia, hyperkalaemia, the inoculum.90–92 None of these approaches, however, hypertension, and nephrotoxicity.9,101 Serious side-eff ects has been shown convincingly to be the best strategy that include transplant-associated thrombotic microangio- enhances long-term survival. pathy and neurotoxic eff ects that can lead to premature Alemtuzumab is a that binds discontinuation. Although clinically similar to thrombotic CD52, a protein expressed on a broad range of leukocytes thrombocytopenic purpura, transplant-associated including lymphocytes, monocytes, and dendritic cells. thrombotic micro angiopathy does not respond reliably to Its use in a phase II trial of GVHD prophylaxis lowered therapeutic plasmapheresis, carries a high mortality rate, incidence of acute and chronic GVHD after and removal of the off ending agent does not always result reduced-intensity transplant.93 In two prospective studies, in improvement.102 Posterior reversible encephalopathy patients who received alemtuzumab rather than syndrome includes mental status changes, seizures, methotrexate showed signifi cantly lower rates of acute neurological defi cits, and characteristic fi ndings on MRI; and chronic GVHD,94 but they had more infectious this syndrome has been seen in 1–2% of patients complications and higher rates of relapse, so no overall undergoing HCT and taking calcineurin inhibitors.103 survival benefi t was recorded. Alemtuzumab might also Side-eff ects of these drugs fall as the dose is tapered, contribute to graft failure when used with minimum- usually 2–4 months after transplantation. intensity conditioning regimens.95 Calcineurin inhibitors are usually administered in An alternative strategy to T-cell depletion attempted to combination with other immunosuppressants, such as induce anergy in donor T cells by ex-vivo antibody methotrexate, which is given at low doses in the early blockade of costimulatory pathways before transplantation. post-transplant period.9,101 The toxic eff ects of methotrexate Findings of a small study of this approach in patients (neutropenia and mucositis) have led some investigators undergoing haploidentical HCT was quite encouraging, to replace it with mycophenolate mofetil. In a prospective but they have not yet been replicated.96 Thus, the focus of randomised trial, patients who received mycophenolate most preventive strategies remains pharmacological mofetil as part of GVHD prophylaxis had signifi cantly less manipulation of T cells after transplant. severe mucositis and more rapid engraftment Administration of antibodies against T cells in vivo as than did those who received methotrexate.104 Frequency GVHD prophylaxis has also been tested extensively. The and severity of acute GVHD was similar between the two best studied drugs are anti-thymocyte globulin or groups, but the study closed early because of superiority of anti-lymphocyte globulin preparations. These serum the mycophenolate mofetil arm with respect to reduced samples, which have high titres of , mucositis and speed of haemopoietic engraftment. A are made by immunisation of horses or rabbits to desire for faster neutrophil engraftment has led to use of or lymphocytes, respectively. A complicating mycophenolate mofetil in umbilical-cord blood transplants factor in establishing the role of these polyclonal serum for which graft failure is a major concern.105 This drug is samples in transplantation is the observation that even also sometimes used after reduced-intensity conditioning diff erent brands of the same class exert diverse biological regimens for similar reasons.106,107 eff ects.97 However, the side-eff ects of anti-thymocyte Sirolimus is an immunosuppressant that is structurally globulin and anti-lymphocyte globulin infusions are similar to tacrolimus but does not inhibit calcineurin. In similar across diff erent preparations and include fever, phase II trials, sirolimus was very eff ective in combination chills, , thrombocytopenia (from cross-reactivity with tacrolimus;108,109 the drug damages endothelial cells, to platelets), and, infrequently, . however, and it might enhance transplant-associated www.thelancet.com Vol 373 May 2, 2009 1555 Seminar

thrombotic microangiopathy, which is associated with -dependent or steroid-refractory GVHD showed calcineurin inhibitors.110 The combination of tacrolimus resolution of disease in most patients, with 50% long-term and sirolimus is currently being compared in a large survival in this very-high-risk group.120 Randomised randomised multicentre trial. multicentre studies of this approach are needed to Reduced-intensity conditioning regimens attempt to establish its place in management of acute GVHD. suppress the host suffi ciently so Another strategy to treat GVHD is blockade of the that donor T cells can engraft and then ablate the infl ammatory cytokine TNFα. TNFα can activate APCs, lymphohaemopoietic compartment of the recipient. recruit eff ector cells, and cause direct tissue damage (see The term non-myeloablative is therefore somewhat Pathophysiology of acute GVHD).121 Data from a phase II misleading. Reduced-intensity conditioning regimens trial of etanercept (solubilised TNFα receptor 2) showed diminish tissue damage and lead to decreased amounts signifi cant eff ectiveness of the drug when added to of infl ammatory cytokines, which are important in the systemic steroids as primary treatment for acute GVHD. initiation of GVHD pathophysiology; this eff ect could 70% of patients had complete resolution of all GVHD account for the reduced frequency of severe GVHD after symptoms within 1 month, with 80% complete responses reduced-intensity conditioning versus full-intensity in the gastrointestinal tract and skin. The researchers also conditioning used in historical controls.53,54,93,111 Onset of reported that concentrations in plasma of TNFα receptor 1 acute GVHD can be delayed after reduced-intensity were a signifi cant biomarker for clinical GVHD.122 conditioning until after day 100, however, and acute disease could present simultaneously with elements of Treatment of chronic GVHD chronic GVHD (known as overlap syndrome).111–113 By contrast with acute GVHD, the pathophysiology of chronic GVHD remains poorly understood and the Treatment of acute GVHD disease is treated with various immunosuppressive GVHD fi rst develops, generally, in the second month agents. The response of chronic GVHD to treatment is after HCT during calcineurin-based prophylaxis.114 unpredictable, and mixed responses in diff erent organs Steroids, with their potent anti-lymphocyte and can take place in the same patient. Confounding variables anti-infl ammatory activity, are the gold standard for such as infection and comorbidities also make responses treatment of GVHD. Many centres treat mild GVHD of hard to measure. Use of (with or without the skin (grade I) with topical steroids alone, but for more a calcineurin inhibitor) is the standard of care, but severe disease and any degree of visceral GVHD fi ndings of a randomised trial of more than 300 patients involvement high-dose systemic steroids are usually with chronic GVHD noted no diff erence between initiated. Administration of steroids results in complete cyclosporine plus versus prednisone alone.123 remission in less than half of patients,115 and more severe Chronic immunosuppressants, especially those con- GVHD is less likely to respond to treatment.116 In a taining steroids, are highly toxic and result in from prospective randomised study, addition of anti-thymocyte infection. Many second-line treatments have been studied, globulin to steroids as primary treatment did not increase but none has achieved widespread acceptance. As the response rate.116 In a retrospective study, use of mentioned in the Treatment of acute GVHD section, anti-thymocyte globulin in patients who showed early extracorporeal photopheresis shows some promise, with signs of steroid resistance was benefi cial,115 but not all relevant response rates in high-risk patients. The best study fi ndings show such benefi t, and this antibody responses were seen in skin, liver, oral mucosa, eye, and preparation is not used as standard because of raised lung.124 This observation is especially pertinent because infection risks.100,117 Infusion of mesenchymal stromal lung GVHD has the potential to be a particularly cells—expanded in culture either from the original HCT devastating complication of chronic GVHD. Inhaled donor or from a third party—is a promising approach, high-dose steroids, when added to existing immuno- which produced 55% complete responses in a phase II suppressant regimens, have stabilised the pulmonary study of patients with steroid-resistant GVHD.57 function of patients with bronchiolitis obliterans in a An increasingly frequent treatment for GVHD is small trial.125 If other treatments fail, extracorporeal photopheresis. During this procedure, the might be the only remaining therapeutic option.126 patient’s white blood cells are gathered by apheresis, incubated with the DNA-intercalating agent Essential supportive care in GVHD patients 8-methoxypsoralen, exposed to ultraviolet light, and Meticulous supportive care is vital for patients with both returned to the patient. Extracorporeal photopheresis is acute and chronic GVHD owing to the extended duration known to induce cellular apoptosis, which has strong of immunosuppressive regimens and because the many anti-infl ammatory eff ects in several systems, including drugs administered could have synergistic toxic eff ects. prevention of rejection of solid organ grafts.118 Work done Such care includes extensive infectious prophylaxis, early in animals shows that extracorporeal photopheresis interventions in cases of suspected infections, and reverses acute GVHD by increasing the number of prophylaxis against non-infectious side-eff ects of drugs regulatory T cells.119 Data from a phase II clinical study of (table). These complications usually need rapid responses

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Clinical symptoms Routine Prophylaxis Recommended intervention Bacterial infections Catheter-related Fever, chills, pain, Assessment of catheter sites Sterile dressing, regular line maintenance Obtain cultures, immediate intravenous treatment, remove line Other Fever, chills, sepsis Clinical signs, chest radiograph in high-risk patients (high-dose Obtain cultures, immediate intravenous symptoms or CT scan for possible corticosteroids or asplenia), intravenous broad-spectrum antibiotics, because of risk of pneumonia immunoglobulin if IgG level <400 g/L overwhelming sepsis within hours Viral infections Cytomegalovirus Gastroenteritis, Blood cytomegalovirus PCR or Pre-emptive treatment in patients with reactivation Antiviral treatment (ganciclovir, valganciclovir, interstitial pneumonia pp65 antigen levels or foscarnet) Respiratory Symptoms of upper or Clinical monitoring Annual infl uenza vaccination (starting 6 months post Early treatment with neuraminidase inhibitors lower respiratory-tract HCT), vaccination of caregivers (infl uenza), other antivirals infections Varicella-zoster virus Vesicular skin lesions Clinical monitoring prophylaxis Treatment doses of antivirals Fungal and other infections , other Pulmonary lesions, Galactomannan, assays in Voriconazole, or posaconazole prophylaxis in high-risk treatment emerging fungal , skin nodules high-risk patients, CT scan if patients (eg, high-dose steroids) infections signs of infection Candida Thrush, pulmonary Clinical examination, CT scan if ( prophylaxis protects against Antifungal treatment lesions signs of infection candida too) Pneumocystis Fever, , Clinical assessment Cotrimoxazole, or pentamadine until 1 month off Treatment doses of anti-PCP drugs respiratory distress immunosuppression Other toxic eff ects of immunosuppressive agents Calcineurin inhibitors Clinical assessment, drug Adjust dose to desired trough levels ·· concentrations Calcineurin inhibitors Neurotoxic eff ects Assess mental status ·· Stop calcineurin inhibitors Calcineurin inhibitors Renal impairment Creatinine levels and glomerular Adequate fl uid uptake (about 3 L per day) Intravenous fl uids fi ltration rate Calcineurin inhibitors Hypertension monitoring ·· Antihypertensive treatment (angiotensin-converting-enzyme inhibitors, β-blocking agents) Calcineurin inhibitors Transplant-associated Assess blood smear for ·· Stop calcineurin inhibitors, plasmapheresis microangiopathy haemolysis, schistocytes Corticosteroids Cushing’s disease Clinical assessment Taper as recommended ·· symptoms Corticosteroids Diabetes Blood glucose levels Nutritional guidance treatment Corticosteroids Osteoporosis Assessment of bone density Calcium or vitamin D supplementation Bisphosphonate treatment for osteoporosis Corticosteroids Myopathy ·· Physiotherapy ·· Late graft failure Blood disorders Bleeding symptoms, Blood counts Stop or switch drugs Growth factors ( colony-stimulating anaemia factor), erythropoetin, transfusions

PCP=Pneumocystis jirovecii.

Table: Recommendations for supportive care to prevent serious or irreversible damage and are best Viral infections are frequent in people with GVHD. handled by close collaboration between the primary Cytomegalovirus causes interstitial pneumonia and doctor and the transplant specialist. gastritis. Patients who are at risk should have their All patients should receive at least fl uconazole as blood monitored several times a month. Techniques prophylaxis against fungal infections. Invasive moulds, that directly detect virus should be undertaken, such as especially aspergillus, are typical with prolonged steroid cytomegalovirus PCR or pp65 antigen, and evidence of use.127 Prophylaxis with voriconazole or posaconazole increased viral load should prompt pre-emptive should be considered for these individuals. Usual sites treatment with ganciclovir or foscarnet before clinical of infection are the lungs, sinuses, brain, and skin,128 and manifestations of disease. Shingles is not uncommon serial galactomannan assays could aid in early detection.129 and aciclovir prophylaxis could be benefi cial.131 Patients Candida can cause lesions in the lung, liver, and spleen, and caregivers should receive vaccinations against which might need screening with ultrasonography. infl uenza, and treatment with neuraminidase inhibi- Pneumocystis is another opportunistic infection that tors is recommended in the event of infl uenza should receive cotrimoxazole (bactrim) prophylaxis.130 infection.132,133 www.thelancet.com Vol 373 May 2, 2009 1557 Seminar

Patients with GVHD sometimes have IgG2 and IgG4 develop (many new cases present up to day 200), and need subclass defi ciencies despite usual amounts of lgG, for close monitoring of individuals in this period has making them susceptible to infections with encapsulated risen. Patients have typically returned to the care of their organisms. Treatment of severe hypo gammaglobu lin- primary haematologists by this time, increasing the need aemia with intravenous immunoglobulin is standard in for these doctors to collaborate with transplant specialists many centres,134 but the level that triggers replacement in the management of GVHD and its complications. varies considerably between transplant specialists. Identifi cation of biomarkers for GVHD with diagnostic Supporting evidence for routine use of intravenous (and possibly prognostic) signifi cance might eventually immunoglobulin as prophylaxis is scarce,135 but patients make treatment of GVHD pre-emptive rather than should receive routine prophylaxis ( or its prophylactic. Cellular component —such as equivalent) because of increased risk for streptococcal regulatory T cells that have been expanded ex vivo—will sepsis.136 Pneumococcal conjugate and Haemophilus also enter clinical trials in the near future, but the extensive infl uenzae might provide additional protection infrastructure needed for such approaches will probably and are recommended for all patients, including those restrict their use initially to large academic centres, with chronic GVHD.130,137 intensifying need for close communication between The sites of any indwelling catheters should be assessed transplant specialists and referring haematologists. regularly and early treatment of a suspected infection Confl ict of interest statement initiated. Early signs or symptoms of septic , such as We declare that we have no confl ict of interest. shaking chills or low blood pressure, need prompt Acknowledgments assessment with chest , CT scan, or both, blood This work was supported in part by grants from the National Institutes culture, and treatment with broad-spectrum antibiotics of Health (Bethesda, MD, USA), P01 CA 39542, and the Jose Carreras Foundation. JLMF is a Doris Duke distinguished clinical scientist and an because shock can progress rapidly in these patients. American Cancer Society clinical research professor. Chronic immunosuppressant treatment has many References toxic eff ects. Diabetes (which further increases risks for 1 Appelbaum FR. Haematopoietic cell transplantation as infection), muscle weakness, osteoporosis, avascular . Nature 2001; 411: 385–89. necrosis (usually requiring joint replacement), and 2 Welniak LA, Blazar BR, Murphy WJ. Immunobiology of allogeneic hematopoietic stem cell transplantation. 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