Correspondence: Abstract 2338 B. Johnson In Vitro Activity of Retapamulin (SB-275833), a Novel Topical Antimicrobial, against 3721 Gram-Positive IHMA (F-2050) 2122 Palmer Drive Schaumburg Isolates Associated with Skin and Skin Structure Infections (SSSIs) From 17 Sites in North America IL 60173 USA 1 1 1 1 2 2 1 1 Tel: +1 847 303 5003 B. Johnson, A. Jordan, S. Bouchillon, D. Hoban, N. Scangarella, R. Shawar, J. Johnson & R. Badal Fax: +1 847 303 5601 1International Health Management Associates, Schaumburg, IL, USA; 2GlaxoSmithKline, Collegeville, PA, USA E-mail: [email protected]

Table 3. MIC (µg/mL) Summary for Retapamulin and Comparators against S. aureus from Table 6. MIC (µg/mL) Summary For Retapamulin and Comparators against Viridans Revised Abstract Materials and Methods North America (n = 1182) Streptococci from North America (n = 471) Background: Retapamulin is a novel pleuromutilin currently in development as a topical G MIC endpoints were determined by broth microdilution according to MIC (µg/mL) MIC (µg/mL) antimicrobial for the treatment of skin and skin structure infections (SSSIs) that shows no Clinical and Laboratory Standards Institute (CLSI) guidelines3 for retapamulin a a Compound %Sus %Int %Res Range MIC50 MIC90 Compound %Sus %Int %Res Range MIC50 MIC90 target specific cross-resistance to other classes of antibiotics and exhibits a unique mode of and 14 comparators in customized dried broth microdilution panels (Trek action. Methods: A total of 3721 clinical isolates of staphylococci (Staphylococcus aureus and Retapamulin NA NA NA 0.015–0.5 0.06 0.12 Retapamulin NA NA NA 0.004–0.5 0.06 0.25 Diagnostic Systems Ltd, West Sussex, UK). Comparator antimicrobial agents coagulase-negative staphylococci, including Staphylococcus epidermidis), Streptococcus pyogenes, Amoxicillin- 61.1 0 38.9 0.12–>32 1 16 Amoxicillin- NA NA NA ≤0.015–8 0.12 1 Clavulanic acidb Streptococcus agalactiae, and viridans streptococci were collected from 17 sites in North America included: amoxicillin-clavulanic acid, bacitracin, ceftriaxone, cephalothin, Clavulanic acidb Bacitracin NA NA NA 0.25–>128 32 128 during 2004–2005. A total of 14 antimicrobials were compared with retapamulin. All isolates clindamycin, cloxacillin, erythromycin, fusidic acid, gentamicin, linezolid, Bacitracin NA NA NA ≤0.06–>128 >128 >128 were sent to the central laboratory for confirmation of identification and testing. Susceptibility mupirocin, neomycin, penicillin, and tetracycline. Ceftriaxone 92.8 2.3 4.9 ≤0.015–>64 0.12 1 Ceftriaxone 58.2 4.3 37.5 1–>64 4 32 Cephalothin NA NA NA ≤0.03–128 0.5 2 testing was performed by the central laboratory using broth microdilution panels and quality G All study organisms were clinical isolates collected and frozen at -70°C from Cephalothin 62.7 0.3 37.1 0.12–256 1 8 Clindamycin 86.0 1.1 13.0 ≤0.015–>32 0.03 8 controls were performed each day of testing following Clinical and Laboratory Standards March 2004 to March 2005 from Canada, Mexico, and the USA (Table 1). Clindamycin 75.1 2.4 22.5 0.03–>32 0.12 >32 Cloxacillin NA NA NA 0.03–>32 2 8 Institute (CLSI) guidelines. Results: Results were as follows: All isolates were obtained from SSSIs. Isolates were obtained from both adult Cloxacillin NA NA NA 0.12–>32 0.5 2 Erythromycin 42.3 2.3 55.4 ≤0.015–>32 1 >32 and paediatric patients with one isolate per patient. Erythromycin 35.3 6.9 57.8 0.06–>32 32 >32 Fusidic acid NA NA NA 0.06–>32 16 32 G Organism collection, transport, confirmation of organism identification, Fusidic acid 93.2 2.5 4.2 0.03–16 0.12 1 Gentamicin NA NA NA 0.06–32 4 16 Antimicrobial MIC90 (µg/mL) Linezolid 100 0 0 0.25–2 1 1 antimicrobial susceptibility testing, as well as construction and management Gentamicin 93.3 2.8 3.9 ≤0.03–>64 0.25 2 Mupirocin NA NA NA 0.06–>256 0.5 2 S. aureus Coagulase- S. pyogenes S. agalactiae viridans of a centralized database, were coordinated by International Health (n = 1182) negative (n = 1164) (n = 462) streptococci Linezolid 100 0 0 0.5–4 2 4 Neomycin NA NA NA 0.12–>64 32 >64 staphylococci (n = 471) Management Associates, Inc. (IHMA, Schaumburg, IL, USA). Mupirocin 90.9 0 9.1 0.06–>256 0.25 4 Penicillin 59.4 38.6 1.9 ≤0.015–>32 0.12 0.5 (n = 442) G Mueller–Hinton broth (Sensititre®, Cleveland, OH, USA) was used for all Neomycin NA NA NA ≤0.03–>64 2 64 Tetracycline 48.4 3.6 48.0 0.12–>32 4 >32 Staphylococcus species. Fresh lysed horse blood (Hemostat Laboratories, Penicillin 7.4 0 92.6 ≤0.015–>32 8 32 Retapamulin 0.12 0.06 0.06 0.06 0.25 aInterpretive criteria of compounds defined in the CLSI document M100-S15 where available. Amoxicillin- 16 8 0.12 0.25 1 Dixon, CA, USA) was added to the Mueller–Hinton broth to make the 3% Tetracycline 82.7 3.6 13.7 0.12–>32 0.5 16 bAmoxicillin/clavulanic acid was tested in a 2:1 ratio; MICs are reported based on the amoxicillin concentration. lysed horse blood broth used for MIC testing of all Streptococcus species. Sus, susceptible; Int, intermediate; Res, resistant; NA, not available. Clavulanic acid aInterpretive criteria of compounds defined in the CLSI document M100-S15 (2005) where available; mupirocin Bacitracin >128 >128 64 128 128 G The trays were incubated at 35°C in ambient air for 16–20 h before reading susceptibility defined by Finlay et al.;6 fusidic acid susceptibility defined by Toma and Barriault;7 Note:β-lactams are Ceftriaxone 32 32 0.12 0.12 1 the MIC endpoints. reported as resistant for MRSA in accordance with current CLSI guidelines. Table 7. MIC (µg/mL) Summary for Retapamulin and Comparators against S. agalactiae Cephalothin 8 4 0.25 0.5 2 bAmoxicillin/clavulanic acid was tested in a 2:1 ratio; MICs are reported based on the amoxicillin concentration. G Quality control testing was performed each day of testing as specified by the Sus, susceptible; Int, intermediate; Res, resistant; NA, not available. From North America (n = 462) Clindamycin >32 >32 0.25 32 8 CLSI using the following ATCC strains: S. aureus ATCC 29213, S. aureus ATCC Cloxacillin 2 16 0.5 2 8 MIC (µg/mL) 25923, Enterococcus faecalis ATCC 29212, Streptococcus pneumoniae ATCC Erythromycin >32 >32 2 32 >32 a Table 4. MIC (µg/mL) Summary for Retapamulin and Comparators against Compound %Sus %Int %Res Range MIC50 MIC90 Fusidic acid 1 8 8 16 32 49619, and Escherichia coli ATCC 25922. In addition, quality control ranges Coagulase-negative Staphylococci from North America (n = 442) Retapamulin NA NA NA 0.008–0.25 0.03 0.06 Gentamicin 2 16 16 32 16 previously determined for retapamulin were used as a control.4 Amoxicillin- NA NA NA ≤0.015–1 0.12 0.25 Linezolid 4 2 1 1 1 The total number of isolates, MIC (µg/mL), MIC (µg/mL) and MIC ranges MIC (µg/mL) G 50 90 Clavulanic acidb Mupirocin 4 >256 0.5 1 2 a were determined for all antimicrobial agents tested. Interpretive criteria and Compound %Sus %Int %Res Range MIC50 MIC90 Bacitracin NA NA NA 2–>128 128 128 Neomycin 64 16 64 >64 >64 resistant phenotypes to the corresponding antimicrobial agent were defined Retapamulin NA NA NA ≤0.002–0.5 0.06 0.06 Ceftriaxone 99.8 0 0.2 ≤0.015–1 0.06 0.12 Penicillin 32 16 0.06 0.12 0.5 5 Cephalothin NA NA NA ≤0.03–2 0.25 0.5 according to CLSI breakpoints or the literature (mupirocin and fusidic acid). Amoxicillin- 40.3 0 59.7 ≤0.015–32 1 8 Tetracycline 16 >32 16 >32 >32 Clindamycin 83.3 1.7 14.9 ≤0.015–>32 0.06 32 Methicillin-resistance was based upon oxacillin screening agar. Clavulanic acidb Cloxacillin NA NA NA 0.12–4 2 2 Bacitracin NA NA NA 0.25–>128 128 >128 Erythromycin 59.7 1.9 38.3 ≤0.015–>32 0.06 32 Conclusion: Retapamulin demonstrated excellent in vitro activity against staphylococci, Ceftriaxone 37.1 5.0 57.9 0.5–>64 8 32 Fusidic acid NA NA NA 0.5–>32 8 16 S. pyogenes, S. agalactiae, and viridans streptococci isolates as demonstrated by the lowest Results Cephalothin 42.1 0 57.9 ≤0.03–64 1 4 Gentamicin NA NA NA 1–>64 16 32 MIC compared with that of topical and oral antimicrobial agents commonly used in the Clindamycin 61.1 2.7 36.2 ≤0.015–>32 0.12 >32 Linezolid 100 0 0 0.5–2 1 1 90 The results are shown in Tables 2–7. treatment of uncomplicated SSSIs. Cloxacillin NA NA NA 0.06–>32 1 16 Mupirocin NA NA NA 0.12–256 1 1 Neomycin NA NA NA 0.06–>64 64 >64 Erythromycin 24.4 3.8 71.7 ≤0.015–>32 >32 >32 Penicillin 98.9 0 1.1 ≤0.015–4 0.06 0.12 Fusidic acid 82.8 1.8 15.4 ≤0.015–>32 0.12 8 Table 1. Geographic Distribution of Isolates Tetracycline 13.9 1.3 84.8 0.12–>32 32 >32 Introduction Gentamicin 73.5 13.8 12.7 ≤0.03–>64 0.12 16 Country Sites Total Isolates (n) % Total Linezolid 100 0 0 0.25–4 1 2 aInterpretive criteria of compounds defined in the CLSI document M100-S15 where available. b Gram-positive bacteria are an increasingly common cause of community- Canada 5 589 15.8 Amoxicillin/clavulanic acid was tested in a 2:1 ratio; MICs are reported based on the amoxicillin concentration. Mupirocin 61.8 0 38.2 0.06–>256 0.25 >256 Sus, susceptible; Int, intermediate; Res, resistant; NA, not available. and hospital-acquired infections, and their resistance to antibiotics is increasing. Mexico 3 228 6.1 Neomycin NA NA NA ≤0.03–64 0.25 16 USA 9 2904 78.1 Retapamulin (SB-275833; Figure 1) is a novel, semi-synthetic derivative of the Penicillin 16.5 0 83.5 ≤0.015–>32 2 16 Total North America 17 3721 100 pleuromutilins, a new class of antimicrobials, and is currently in development Tetracycline 76.7 3.6 19.7 ≤0.06–>32 1 >32 for the topical treatment of secondarily-infected traumatic lesions and Conclusions a secondarily-infected dermatoses. The pleuromutilins are potent inhibitors of Interpretive criteria of compounds defined in the CLSI document M100-S15 (2005) where available; mupirocin susceptibility defined by Finlay et al.;6 fusidic acid susceptibility defined by Toma and Barriault;7 Note: β-lactams G Retapamulin demonstrated excellent in vitro activity against all S. aureus, protein synthesis in bacteria through the interference of peptide bond formation Table 2. MIC (µg/mL) Summary for Retapamulin Activity Against 3721 Skin and Skin are reported as resistant for methicillin-resistant staphylococci in accordance with current CLSI guidelines. coagulase-negative staphylococci, S. pyogenes, S. agalactiae, and viridans by binding to the peptidyl transferase center of the 50S ribosomal subunit.1 Structure Pathogens in North America bAmoxicillin/clavulanic acid was tested in a 2:1 ratio; MICs are reported based on the amoxicillin concentration. Sus, susceptible; Int, intermediate; Res, resistant; NA, not available. streptococci isolates tested in this study, including strains resistant to Due to the unique retapamulin mode of action, retapamulin shows no target MIC (µg/mL) methicillin, erythromycin, mupirocin, or fusidic acid. specific cross-resistance with other classes of antibacterials. a Organism/Phenotype n Range MIC50 MIC90 G The in vitro activity of retapamulin was 4- to >256-fold more active than Pleuromutilins have demonstrated significant in vitro activity against a variety of S. aureus 1182 0.015–0.5 0.06 0.12 Table 5. MIC (µg/mL) Summary for Retapamulin and Comparators against S. pyogenes fusidic acid, linezolid and mupirocin against all staphylococci and streptococci fastidious and non-fastidious bacterial pathogens, especially methicillin-resistant Methicillin-susceptible 746 0.015–0.5 0.06 0.12 from North America (n = 1164) strains and equivalent to penicillin against all streptococci. Staphylococcus aureus and coagulase-negative staphylococci.2 A comprehensive Methicillin-resistant 436 0.03–0.5 0.06 0.12 G Retapamulin inhibited all study strains at MIC values of ≤0.5 µg/mL. Macrolide-susceptible 417 0.015–0.5 0.06 0.12 MIC (µg/mL) G Clinical data is necessary to assess the clinical significance of retapamulin’s study was conducted to evaluate the in vitro activity of retapamulin and Macrolide-intermediate 82 0.03–0.25 0.06 0.12 a Compound %Sus %Int %Res Range MIC50 MIC90 comparative antimicrobial agents against recently isolated clinical isolates of Macrolide-resistant 683 0.015–0.5 0.06 0.12 potent in vitro activity against strains causing SSSI, including those resistant to S. aureus, coagulase-negative staphylococci, Streptococcus pyogenes, Streptococcus Mupirocin-susceptibleb 1074 0.015–0.5 0.06 0.12 Retapamulin NA NA NA 0.008–0.25 0.03 0.06 one or more of the agents commonly used in treatment of these infections. agalactiae and viridans streptococci from skin and skin structure infections (SSSIs), Mupirocin-resistantb 108 0.03–0.5 0.06 0.12 Amoxicillin- NA NA NA ≤0.015–0.5 ≤0.015 0.12 G Overall, against 3721 Gram-positive isolates associated with SSSIs from North c b including multi-drug resistant strains, during the course of a multi-center Fusidic acid-susceptible 1102 0.015–0.5 0.06 0.12 Clavulanic acid America, retapamulin was the most potent agent in vitro and inhibited all Fusidic acid-resistantc 50 0.03–0.5 0.06 0.12 surveillance program in North America. Bacitracin NA NA NA 0.25–>128 264 isolates at a MIC of ≤0.5 µg/mL. Coagulase-negative staphylococci 442 ≤0.002–0.5 0.06 0.06 Ceftriaxone 99.9 0 0.1 ≤0.015–2 ≤0.015 0.12 Methicillin-susceptible 186 ≤0.002–0.5 0.06 0.06 Cephalothin NA NA NA ≤0.03–32 0.12 0.25 Methicillin-resistant 256 0.015–0.5 0.06 0.12 Clindamycin 96.6 1.5 1.9 ≤0.015–>32 0.03 0.25 Mupirocin-susceptibleb 273 ≤0.002–0.5 0.06 0.06 Mupirocin-resistantb 169 0.015–0.5 0.06 0.12 Cloxacillin NA NA NA ≤0.015–32 0.12 0.5 References Erythromycin 78.4 1.4 20.3 ≤0.015–>32 0.03 2 1 Schlunzen F, Pyetan E, Fucini P, et al. Mol Microbiol 2004; 54: 1287–1294. O OH S. pyogenes 1164 0.008–0.25 0.03 0.06 2 Jones RN, Pfaller MA, Rhomberg PR, Walter DH. J Clin Microbiol 2002; 40: 461–465. MeN Macrolide-susceptible 912 0.008–0.12 0.03 0.03 Fusidic acid NA NA NA 0.5–32 4 8 3 CLSI. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved S Macrolide-intermediate 16 0.06–0.06 0.06 0.06 Gentamicin NA NA NA 0.25–64 4 16 Standard 6th Edition. Document M7-A6. Wayne, PA, USA; CLSI, 2005. O Macrolide-resistant 236 0.015–0.25 0.03 0.06 Linezolid 100 0 0 0.25–2 1 1 4 Jones RN. SB-275833, An Investigational Topical Agent: Quality Control Studies for the MIC (M7-A6) Method. H Viridans streptococci 471 0.004–0.5 0.06 0.25 Document on file, UH2004/00009/00, GlaxoSmithKline, Collegeville, PA, USA, 2005. Mupirocin NA NA NA 0.06–>256 0.12 0.5 5 CLSI. Performance Standards for Antimicrobial Susceptibility Testing, in Document M100-S15. Wayne, PA, USA: S. agalactiae 462 0.008–0.25 0.03 0.06 Neomycin NA NA NA 1–>64 32 64 CLSI, 2005. Total Isolates Tested 3721 ≤0.002–0.5 0.06 0.12 Penicillin 100 0 0 ≤0.015–0.12 ≤0.015 0.06 6 Finlay JE, Miller LA, Poupard JA. Antimicrob Agents Chemother 1997; 41: 1137–1139. 7 Toma E, Barriault D. J Clin Microbiol 1995; 33: 1712–1715. aPhenotypes were determined by the in vitro susceptibility of the respective antimicrobial agent against the Tetracycline 79.4 1.5 19.2 ≤0.06–>32 0.25 16 O corresponding organism as defined in CLSI document M100-S15 unless otherwise noted; macrolide = erythromycin activity; methicillin = oxacillin activity. aInterpretive criteria of compounds defined in the CLSI document M100-S15 (2005) where available. bMupirocin susceptibility (≤4 µg/mL susceptible; ≥8 µg/mL resistant) as defined by Finlay et al.6 bAmoxicillin/clavulanic acid was tested in a 2:1 ratio; MICs are reported based on the amoxicillin concentration. Acknowledgements Figure 1. Chemical Structure of Retapamulin cFusidic acid susceptibility (≤1 µg/mL susceptible; ≥4 µg/mL resistant) as defined by Toma and Barriault.7 Sus, susceptible; Int, intermediate; Res, resistant; NA, not available. This study was sponsored by a grant from GlaxoSmithKline Pharmaceuticals.

Presented at the 45th Interscience Conference on Antimicrobial Agents and Chemotherapy, 16–19 December 2005, Washington DC, USA