OPHTHALMIC ANTIMICROBIALS Alison Clode, DVM, DACVO Port City Veterinary Referral Hospital Portsmouth, New Hampshire New England Equine Medical and Surgical Center Dover, New Hampshire Overview • Interpretation of efficacy • Mechanisms of resistance • Antibacterial agents • Mechanism of action • Applications in ophtho • Antifungal agents • Mechanism of action • Applications in ophtho Interpretation of Efficacy – in vitro 1. MIC = minimum inhibitory concentration • Lowest concentration of antibiotic that inhibits growth of a specific organism 2. MBC = minimum bactericidal concentration • Lowest concentration of an antibiotic at which bacteria are killed 3. Breakpoint • Antibiotic concentration dividing susceptible and resistant • MIC < breakpoint à S • MIC ≥ breakpoint à I, R Interpretation of Efficacy – in vivo 4. PK/PD = pharmacokinetics (what the body does to the drug) pharmacodynamics (what the drug does to the body) 5. Susceptible = bacteria inhibited by usually achievable concentrations of antibiotic when recommended dose used for particular site of infection 6. Intermediate = bacteria inhibited in sites were antibiotic is physiologically concentrated or when higher-than-normal dosage can be used 7. Resistant = bacteria not inhibited by usually achievable concentrations of antibiotic with normal dosing schedules or when microbial resistance mechanisms are likely Interpretation of Efficacy • Indices utilized: • T > MIC = % time plasma concentration is above MIC • Cmax/MIC = max plasma concentration relative to MIC • AUC/MIC = plasma concentration time curve (duration of drug exposure) relative to MIC • Determined by various animal models www.rxkinetics.com Time- versus Concentration-Dependent www.slideshare.net Mechanisms of Resistance • Intrinsic to the bacteria • Acquired by the bacteria Acquired Mechanisms of Resistance 1. Modification of the antibiotic 2. Preventing antibiotic from reaching target 3. Modification of the target Acquired Mechanisms of Resistance 1. Modification of the antibiotic • Enzyme-induced damage to antibiotic à inactive antibiotic • Enzyme-induced acetylation, adenylation, phosphorylation of antibiotic à alter affinity of antibiotic for target Acquired Mechanisms of Resistance 2. Prevent antibiotic from reaching target • Preventing intracellular drug accumulation • Alteration of porin channels à reduced drug entry • Production of active efflux pumps à reduced drug retention Acquired Mechanisms of Resistance 3. Modification of target by altering: • Binding proteins • Ribosomes • Chromosomes • Cell physiology Acquired Mechanisms of Resistance • Vertical gene transfer = transfer of R-conferring gene to progeny • Horizontal gene transfer = sharing of R-conferring DNA among bacteria • Same or different strains • Transformation = DNA uptake from environment • Transduction = DNA transfer by viruses • Conjugation = plasmid exchange via cell-to-cell contact Antibacterial Agents Antibacterial Agents • Mechanisms of action = disruption of: 1. Cell wall synthesis 2. Cell membrane integrity 3. Protein synthesis 4. Folate metabolism 5. DNA synthesis Bacterial Cell Wall • Main component = peptidoglycan • PS + peptide crosslinks • Formed by transpeptidases (penicillin binding proteins) • Gram positive: • Thick cell wall with greater peptidoglycan content and teichoic acid • Cytoplasmic membrane Bacterial Cell Wall • Main component = peptidoglycan • PS + peptide crosslinks • Formed by transpeptidases (penicillin binding proteins) • Gram negative: • Outer membrane of LPS and phospholipids • Thinner cell wall with lesser peptidoglycan content • Cytoplasmic membrane 1. Cell Wall Synthesis Inhibitors • Penicillins • Cephalosporins • Bacitracin • Glycopeptides Penicillins – Structure and function thiazolidine ring • Side chain: • Spectrum side chain • Susceptibility to destruction • Pharmacokinetic properties • β-lactam: • Function • Bind transpeptidase à inhibit formation of peptide linkages between polysaccharides à inhibit formation of peptidoglycan β-lactam Penicillins – Resistance 1. β-lactamase production • à hydrolysis of β-lactam ring • Occurs extracellularly in G+ • Occurs between cell membrane and wall in G- • Induced by drug binding to bacterial cell wall or • Constitutively produced by bacteria www.wiley.com Penicillins – Resistance 2. Alter transpeptidases • Penicillins unable to bind to and inactivate transpeptidase • ‘MRSA’ www.wiley.com Penicillins – Classes • Effective versus G+ • Resistant to penicillinase • Extended spectrum • Anti-pseudomonal Penicillins Effective versus G+ Penicillin G (parenteral) Penicillin V 1. Highly susceptible to β-lactamases à poor activity versus Staph aureus and Staph epidermidis 2. Ineffective versus altered transpeptidases à poor activity versus Streptococcus pneumoniae, viridans streptococci Penicillins Resistant to penicillinases Methicillin Oxacillin Cloxacillin Dicloxacillin Nafcillin 1. Structural modifications à increased efficacy versus β- lactamase-producing Staph aureus, Staph epidermidis 2. Resistance now due to altered transpeptidases Penicillins Extended spectrum Ampicillin (+/- sulbactam) Amoxicillin (+/- clavulanate) 1. Penicillins inactivated by β-lactamases when not in combo 2. Irreversible inactivation of β-lactamases by sulbactam and clavulanate 3. Ineffective versus altered transpeptidases Penicillins Anti-pseudomonal activity Carbenicillin Ticarcillin (+/- clavulanate) Piperacillin (+/- tazobactam) Mezlocillin Also effective versus Proteus and Enterobacter Cephalosporins – Structure and Function dihydrothiazine ring • Side chains: side chain • Spectrum/classification • Susceptibility to destruction • Pharmacokinetic properties • β-lactam: • Function • Bind transpeptidase à inhibit formation of peptide linkages between polysaccharides à inhibition of peptidoglycan formation β-lactam side chain Cephalosporins – Resistance 1. Destruction by β-lactamases • Cephalosporins less susceptible than penicillins • S. aureus produces penicillinases • G- bacteria produce β-lactamases • Extended spectrum β-lactamases (E. coli, Pseudomonas, etc.) * Zapun A, et al., FEMS Microbiol Rev 2008 Cephalosporins – Resistance 2. Alteration of transpeptidases • Cephalosporins unable to bind to and inactivate enzyme • Less common for cephalosporins than for penicillins • ‘MRSA’ * Zapun A, et al., FEMS Microbiol Rev 2008 Cephalosporins First generation Second Third generation Fourth generation generation Drugs Cephalexin Cefuroxime Ceftazidime Cefepime Cefazolin Cefoxitin Cefotaxime Cefadroxil Cefaclor Ceftriaxone Cephradine Cefprozil Cefixime Cefotetan Cefdinir Other Good G+ activity Good G+ activity Modest G+ Good G+ activity activity Modest G- activity Improved G- Good G- activity activity Improved enteric Increasing G- activity resistance of Streptococcus Ceftazidime has pneumoniae to excellent activity cefazolin versus Pseudomonas aeruginosa Penicillins and Cephalosporins in Ophtho • No commercially available topical ophthalmic preparations • Systemic administration: • Orbital disease • Adnexal disease • Limited use in ocular surface disease • Staph and Strep resistance (penicillins) • Strep resistance (cephalosporins) • Limited use in endophthalmitis Bacitracin • Interrupts transporter molecule à inhibits movement of peptidoglycan precursor from cytoplasm to cell wall • G+ • Staphylococcus • Streptococcus pyogenes • Administered topically (ointment) • Nephrotoxicity • May be administered IM in very few approved situations • Poor transcorneal penetration • “Allergen of the Year” 2003 www.ccbcmd.edu Glycopeptides • Bind D-Ala-D-Ala terminal portion of peptidoglycan precursor à peptidoglycan precursor unavailable for cell wall formation à decreased cell wall growth + decreased cell wall rigidity vancomycin Glycopeptides • Strong activity vs G+ • Drug of choice for MRSA, penicillin- resistant Strep pneumoniae • Most G- are resistant • Vancomycin • Teicoplanin vancomycin Glycopeptides – Resistance 1. Alterations of the antibiotic target • VanA resistance: • Reduced affinity via alteration of terminal amino acid residues of peptidoglycan precursor (D-Ala-D-Ala à D-Ala-D-Lac) • VanC resistance: • Steric hindrance caused by substitution (D-Ala-D-Ala à D-Ala-D- Ser) Glycopeptides – Resistance 2. Altered antibiotic penetration • Inability to penetrate bacterial membrane (G- organisms) • Intrinsic resistance Glycopeptides – Resistance Glycopeptides – Resistance Enterococcal spp that are resistant to vancomycin but require vancomycin presence to grow have been isolated… Vancomycin presence induces resistance mechanisms…. This is VERY BAD… Vancomycin – Ocular Application • Reaches therapeutic AH levels when applied topically (50 mg/ml) • Effective versus corneal infections with MRSA and MRSE • Associated with cystoid macular edema when used intracamerally during cataract surgery • Non-toxic to the retina at 1 mg dose • Intravitreal injection in combination with amikacin or ceftazadime for endophthalmitis * Alster Y, et al., BJO 2000 ** Sotozono C, et al., Cornea 2002 *** Penha FM, et al., Ophthalmic Res 2010 Teicoplanin – Ocular Application • Alternative therapy for MRSA infections • No vitreal penetration when administered topically • Poor vitreal penetration when administered IV 2. Cell Membrane Disruptors • Polymyxin B • Gramicidin • Similarities between bacterial and human cell membranes limit use Polymyxin B • Detergent/surfactant • Disrupts cell membrane phospholipids à increased permeability à cell death • Positively charged drug binds negatively charged LPS layer • Binds