Truvada ® for prevention Tenofovir disoproxil fumarate (TDF) + Emtricitabine (FTC)

1. Extracellular and Intracellular Tenofovir DF and Emtricitabine Exposurein Mucosal Tissue after a Single Dose of Fixed-Dose TDF/FTC: Implications for Pre-exposure Prophylaxis Kristine B Patterson, Heather A Prince, Eric Kraft, Amanda Jones, Sunita Paul, Nicholas J Shaheen, Melissa Spacek, Paris E Heidt, Sunila Reddy, Jim Rooney, Julie B Dumond, Myron S Cohen, and Angela DM Kashuba University of North Carolina, Chapel Hill, NC USA Gilead Sciences, Foster City, CA USA

2. Truvada® for preventionTenofovir disoproxil fumarate (TDF) + Emtricitabine (FTC) Daily oral dosing of TDF +/- FTC being evaluated. Clinical trials evaluating EPISODIC ORAL dosing are being planned. There are no PK data in mucosal tissue after a single oral dose of TDF and FTC.

3. Objectives Primary • To characterize tenofovir and emtricitabine (TFV/FTC and TFV-DP/FTC-TP) exposure in multiple biological compartments in both men and women after a single oral dose of Truvada®. Secondary • To analyze the decay characteristics (t1/2) of TFV/FTC and TFV-DP/FTC-TP concentrations in multiple biological compartments in both men and womenafter a single oral dose of Truvada®.

4. Methods Single site, open-label trial Healthy HIV-negative men and women (ages 18-40) • Comprehensive STD screening • Sexually abstinent • Using contraception Single observed dose of Truvada®

5. Pharmacokinetic Sampling Blood Plasma (BP), PBMC, Cerv-Vag Fluid (CVF) Cervical (CT) and Vaginal Tissue (VT) Rectal Tissue (RT) 1 2 5 7 10 14 Sampling Time (Days)

6. Analyses Sample Analyses: • LC/MS/MS • TFV and FTC: LLOQ 0.1 ng/mL • TFV-DP and FTC-TP: LLOQ 2-10 fmol Data Analyses: • Non-compartmental PK (WinNonlin 6.0) (composite) • Summary statistics (SAS 9.1.3) • AUC TFV and FTC: ng*days/mL or gm • AUC TFV-DP and FTC-TP: fmol*days/106 cells or 0.2uL • Penetration ratios (AUC Ratios); • CVF or tissue AUC 0-14d ÷ BP AUC 0-14d

7. Subject Demographics

8. AUC (ng*days/mL) t½(hr) AUC (ng*days/mL) t½(hr) BP 89 56 BP 60 67 Median Blood Plasma Concentrations Can Be Quantified Up to 14 Days Post-Dose Tenofovir Emtricitabine

9. Median Rectal Tissue Concentrations Are Higher Than Blood Plasma Up to 14 Days Post-Dose AUC AUCRT:BP t½ AUC AUCRT:BP t½ 42 RT 266 BP 60 87 67 33 4.3 BP 89 56 Tenofovir Emtricitabine rectal tissue rectal tissue RT 2981

10. Median Intracellular Rectal Tissue Concentrations Can Be Quantified 2-14 Days Post-Dose AUC AUC Ratio t½ AUC AUC Ratio t½ RT 6,495 PBMC 10,813 45 56 RT 199 PBMC 10,832 67 87 NA Tenofovir Diphosphate Emtricitabine Triphosphate NA

11. Median Cervicovaginal Fluid Concentrations Are Higher Than Blood Plasma Up to 14 Days Post-Dose AUC AUCCVF:BP t½ AUC AUCCVF:BP t½ CVF 233 BP 89 53 56 CVF 2520 BP 60 43 67 2.6 42 Tenofovir Emtricitabine cervicovaginal fluid cervicovaginal fluid

12. Vaginal and Cervical Tissue Concentrations Are Similar To, or Higher Than, Blood Plasma Up to 14 Days Post-Dose Tenofovir Emtricitabine cervical tissue cervical tissue vaginal tissue vaginal tissue AUC AUC Ratio t½ AUC AUC Ratio t½ 5.8 42 CT 510 24 CT 2496 57 0.6 7 VT 419 BP 60 VT 50 BP 89 110 56 32 67

13. Median Intracellular Vaginal and Cervical Tissue Concentrations Can Be Detected 1-14 Days Post-Dose Tenofovir Diphosphate Emtricitabine Triphosphate AUC AUC Ratio t½ AUC AUC Ratio t½ CT 132 CT 167 NA NA NA VT 1171 PBMC 10,813 VT 1492 PBMC 10,832 NA 148 NA 43 86

14. Preferential penetration of TFV/FTC seen in this study supports quantification of other ARVs in all mucosal tissues as part of early development strategies for oral PrEP. Differential drug terminal elimination (“Tail”) emphasizes consideration for combination therapy esp. episodic dosing. Summary and Implications

15. Acknowledgements Study Volunteers Gilead Sciences UNC CFAR Clinical Pharmacology and Analytical Chemistry Laboratory (P30 AI50410) National Institute of Health (K23 AI077355) UNC Clinical Translational Research Center (RR025747)