1. Safety, tolerability and early bactericidal activity in sputum of PNU-100480 (sutezolid) in patients with pulmonary tuberculosis RS Wallis 1, AH Diacon 2, R Dawson 3, A Venter 2, SO Friedrich 2, D Paige 1, T Zhu 1, A Silvia 1, J Gobey 1, C Ellery 1, Y Zhang 1, E Kadyszewski 1 1 Pfizer, USA; 2 Stellenbosch University and 3 University of Cape Town, South Africa

2. PNU-100480 (sutezolid) Oxazolidinone antimicrobial – Sulfur-containing linezolid analog with an active sulfoxide metabolite – Bind 23S RNA and inhibit microbial protein synthesis – No cross resistance, neither inhibits nor induces CYP3A4 Preclinical findings – Superior bactericidal activity vs. linezolid in mouse and whole blood TB models regardless of LZD dose or concentration – Earlier sterilization (1-2 months) when combined with standard TB drugs Phase 1 findings – Doses to 600 mg BID generally safe and reasonably well tolerated to 28d – No significant safety signals, incl. hematology, biochemistry, QT Potentially can address 4 major unmet medical needs in TB – DR, HIV, DS, suspected DR-LTBI The present phase 2a study is its first in TB patients

3. Study inclusion criteria Men and women aged 18-65 years Pulmonary tuberculosis – CXR consistent with pulmonary tuberculosis – Positive sputum acid-fast smears – Culture or molecular confirmation of drug-susceptible Mtb Either HIV-1 uninfected, or HIV-1 infected with CD4 T cell counts >350/mm 3 and not currently receiving ART Reasonably normal renal, hepatic, metabolic function Willing to provide written informed consent according to ICH guidelines 3

4. Study exclusion criteria Significant hemoptysis TB treatment within the preceding 6 months – Or positive test for urinary isoniazid metabolite at the time of screening Treatment with MAO inhibitors, tricyclic antidepressants, or adrenergic agonists such as pseudoephedrine or phenylpropanolamine within the preceding 7 days – Due to potential MAO-B activity 4

5. Randomization and treatment Subjects were recruited at 2 sites in South Africa, and were randomly assigned in blocks of 7 to: – PNU-100480 600 mg BID – PNU-100480 1200 mg QD – Fixed dose combination tablets consisting of isoniazid, rifampin, ethambutol, and pyrazinamide (Rifafour© e275) Neither subjects nor investigators were blinded to assigned treatment Treatment duration was 14 days – After which all patients commenced standard TB treatment – Final safety evaluation was on day 42 5

6. Subject characteristics Treatment arm 600 mg BID1200 mg QDHREZ Number of subjects25 9 Age (years, mean±SD)32.3±9.034.1±11.733.8±11.8 Sex (male/female)20/5 7/3 Race (Black/other)11/148/173/6 Weight (kg, mean±SD)54.6±6.551.1±6.751.3±7.5 Height (cm, mean±SD)167.4±8.2167.0±6.1166.5±11.8 BMI (kg/m 2, mean±SD)19.6±2.918.3±1.818.4±0.5 Baseline log CFU/ml (mean±SD)6.88±1.116.91±1.207.22±0.71

7. Safety and tolerability 600 mg BID 1200 mg QD HREZ Subjects evaluable for adverse events25 9 Number of adverse events23 (44%)17 (33%)12 (23%) Subjects with adverse events15 (60%)12 (48%)5 (56%) Subjects with serious or severe adverse events 100 Subjects discontinued due to adverse events 000 Subjects with dose reduced or temporary discontinuation due to adverse events 000 7

8. Adverse events (all causes) PNU 600mg BID (n=25)PNU 1200mg QD (n=25)HREZ (n=9) MildModSevMildModSevMildModSev CARDIAC000100000 GASTROINTESTINAL210000200 GENERAL000110100 INFECTIONS110000100 INVESTIGATIONS040030000 METABOLISM200000000 MUSCULOSKELETAL200100200 NERVOUS SYSTEM200200000 PSYCHIATRIC000110000 RENAL AND URINARY000000010 REPRODUCTIVE000000110 RESPIRATORY001110100 SKIN620210100 TOTAL158111701020 Hemoptysis Day 28 ALT increase 2-3x ULN No effect on QTc interval BID: -4.2±14 ms QD: -3.1±12 ms

9. Early Bactericidal Activity Shading indicates 90% confidence interval by mixed effects model repeated measures analysis Both PNU dosing schedules resulted in significant log CFU reductions from baseline over the 14 day period of treatment. –600 mg BID: -0.09 log/d, 90% CI -0.06 to -0.11 –1200 mg QD: -0.07 log/d, 90% CI -0.04 to -0.09 A trend was apparent toward superior responses with BID dosing

10. Treatment-emergent ALT increases 7/50 PNU-treated TB patients (14%) No apparent pre- disposing factors ALT increased to 2-3x ULN, accom- panied by smaller AST increases No changes in bilirubin or AP No cases met Hy’s Law criteria All were asymptomatic and resolved quickly 10

11. Glutathione (GSH) depletion hypothesis Drug-induced liver injury is common in TB (10-20%) – HRZ all are implicated GSH, which ordinarily protects against oxidative injury, is decreased in TB – GSH is decreased in TB animal models and in TB patients – Palanisamy, PLoS ONE 2011, and Venketaraman, Microb Path 2008 – Apparently is consumed to protect host cells from the cellular antimicrobial host response (peroxides, superoxides, NO) – Lowest levels are in TB patients with drug-induced liver injury – Chowdhury, Indian J Gastro 2001 Liver injury due to HRZ can be prevented by supplementation with N-acetylcysteine (NAC) – (Baniasadi, Eur J Gastro Hepatol 2010) 11

12. Summary Treatment with PNU-100480 at doses of 600 mg twice daily or 1200 mg once daily for 14 days resulted in significant reductions in sputum bacillary burden – A trend toward superior bactericidal effect was apparent when given twice daily Both dosing schedules were generally safe and relatively well tolerated New TB regimens containing PNU-100480 can potentially address major unmet medical needs in TB treatment

13. Contact: robert.wallis@pfizer.com

14. Questions? LZD NAC MPS Parent – metabolite Proposed clinical plan

16. EBA comparison with linezolid DrugDose and duration* mean log/d (90% CI) Study Linezolid600 mg BID and QD days 0-7 -0.082 (-0.054 to -0.110) Dietze, AJRCCM 2008 Sutezolid600 mg BID days 0-6 -0.146 ( ‑ 0.102 to -0.191) (current study) *Both LZD doses appeared equivalent, and here are combined to increase precision. LZD data indicate effects over the entire treatment period (7 days). Sutezolid data were limited to days 0-6 for comparison, as log CFU data were not collected on day 7. Caution is warranted when comparing with historical data.

17. Human Pulmonary Tuberculosis Extracellular infection Cavities contain large numbers of log phase bacilli that can give rise to resistance Inadequate treatment results in failure Eradication is termed bactericidal activity Intracellular infection Granulomas contain small numbers of semi-dormant bacilli that can give rise to persistence Inadequate treatment results in relapse Eradication is termed sterilizing activity

18. Plasma concentration/MPS IC50 PNU-100480Linezolid Wallis, AAC 2011

19. Intracellular bactericidal activity ex vivo whole blood culture

20. Linezolid, sutezolid & metabolites MIC (  g/ml) MPS IC50 (  g/ml) Plasma conc vs. parent Critical activity Linezolid0.55.5- Sutezolid0.2515.5- PNU-1016030.54.45X PNU-1012440.550.1X Linezolid Sutezolid PNU-101603 PNU-101244 MIC (  g/ml) MPS IC50 (  g/ml) Plasma conc vs. parent Critical activity Linezolid0.55.5- Sutezolid0.2515.5-Intracellular PNU-1016030.54.45XExtracellular PNU-1012440.550.1X-

21. 2-mo culture status highly related to relapse-free cure Wallis, Lancet ID 2010

22. Sutezolid Clinical Plan Adaptive licensing / Accelerated approval Confirmatory trial: Novel DR regimen Novel DS regimen (UJRbZ) vs. SOC Selection of regimens & durations Ph1EBA* Compound 1 Ph1EBA* Compound 2 Ph1EBA* Compound 3 Ph1EBA* Compound 4 Non- relapsing cure 2-mo status *or alternative 2-mo status Outcomes Registry 3 yrs 2-mo regimen selection trial: PNU-100480 (U) Bedaquiline (J) plus one of: SQ109 PA-824 Clofazimine Imatinib Rifabutin/PZA XDR MDRDS

23. NAC Supplementation Prevents TB Drug-Induced Liver Injury Baniasdai, Eur J Gastro Hepatol 2010 23