International AIDS Conference IMPACT OF VAGINAL MICROBIOTA ON GENITAL TISSUE AND PLASMA CONCENTRATIONS OF DAPIVIRINE Sharon L Hillier, Leslie A Meyn, Katherine Bunge, Michele Austin, Bernard J Moncla, Charlene S. Dezzutti, Brid Devlin, Mark Marzinke, Craig Hendrix and Lisa Rohan for the FAME Team University of Pittsburgh and Magee-Womens Research Institute, Pittsburgh, PA; the International Partnership for Microbicides, Silver Springs, MD and Johns Hopkins University, Baltimore, MD International AIDS Conference Paris, France July 25, 2017

Tenofovir gel effective against HIV with Lactobacillus dominance Using adjusted Cox proportion hazard models, the HIV incidence rate in women with Lactobacillus dominance was 2.7 and 6.9 per 100 women-years in the tenofovir and placebo arms, respectively, translating to relative reduction of the rate of HIV-1 infection of 61% offered by tenofovir. [As the analysis compares HIV outcomes based on LD status, which neither the trial participants nor the investigators were aware of at randomization, overall random allocation in the CAPRISA 004 trial is expected to lead to comparable groups of women assigned to tenofovir or placebo gel in each of the LD and non-LD strata ] Klatt, et al. Science Volume 356(6341):938-945. June 2, 2017

What does the microbiota do to tenofovir in vitro? Klatt, et al. Science Volume 356(6341):938-945. June 2, 2017

Limitations of Published Data Women having greater numbers of partners and more frequent sexual activity at greater risk of BV and HIV; association between BV and apparent decreased efficacy in CAPRISA-04 trial could be attributed to unmeasured differences in behavior Degradation of tenofovir in presence of G vaginalis was demonstrated within hours in vitro, but this may not replicate in vivo conditions. Klatt, et al. Science Volume 356(6341):938-945. June 2, 2017

Connecting the Dots In vitro studies of microbes and drugs Results from the trial FAME trials: Timed in vivo exposure with direct measurement of drug in plasma and tissues after accounting for differences in the microbiome Hillier et al, CROI 2017 Klatt, et al. Science Volume 356(6341):938-945. June 2, 2017 Klatt, et al. Science Volume 356(6341):938-945. June 2, 2017

Objective and Hypothesis Objective: To evaluate whether vaginal microbiota associated with BV impact dapivirine concentrations in genital tract tissues and plasma following vaginal application. Hypothesis: Vaginal microbiota associated with BV will not decrease genital tract tissue and plasma levels of dapivirine

Study Population 66 healthy, nonpregnant women from Pittsburgh, PA, USA using effective contraception Asymptomatic, HIV-, negative for chlamydia, gonorrhea, trichomoniasis Characteristics: 76% unmarried 67% White, 27% Black

Study Design drug applied Tenofovir or dapivirine (film or gel) applied in clinic Collect: vaginal swabs for qPCR for microbiota drug applied daily at home Collect plasma, CVL and vaginal fluid for drug levels 2 hours Collect: Plasma Cervical biopsy Insert 7th dose in clinic Bunge, et al, CROI 2016 and Hillier et al, CROI 2017

Methods qPCR performed from vaginal swab samples collected at baseline Gardnerella vaginalis Lactobacillus crispatus, L jensenii, L gasseri, Bacterial vaginosis detected using Nugent criteria from a Gram stained vaginal smear collected at baseline Statistics: Relationship between vaginal microbiota and TFV or dapivirine concentrations was assessed using linear regression models A quadratic term was included in the tenofovir models with G. vaginalis to improve model fit. Reported P-values are from the global F-test. 

Tenofovir and Dapivirine: Tissue and Plasma LB dominant Genital epithelium Plasma Vaginal administration of tenofovir or dapivirine BV

Drug Level in Plasma vs Nugent Score (Score of 0-3 Lactobacillus dominant) Hillier et al, CROI 2017

Drug Level in Plasma vs Concentration of G vaginalis Hillier et al, CROI 2017

Drug Level in Plasma vs quantity of L crispatus, L jensenii or L gasseri Hillier et al, CROI 2017

Drug Levels in Cervical Tissue vs Nugent Score

Drug Levels in Cervical Tissue vs Concentration of G vaginalis Hillier et al, CROI 2017

Drug Levels in Cervical Tissue vs L crispatus, L jensenii or L gasseri Hillier et al, CROI 2017

Conclusions In contrast to tenofovir, genital and plasma concentrations of dapivirine were not decreased by bacteria associated with BV. These data suggest that the levels of dapivirine following vaginal application should not be impacted by the microbiota associated with bacterial vaginosis. This is supportive of the ASPIRE study results 40% of women had bacterial vaginosis at baseline. No difference in the efficacy of the dapivirine ring in women with bacterial vaginosis and in those without bacterial vaginosis at baseline (39% vs. 21%, P=0.4 for interaction). (NEJM 2017 Mar 9;376: 995-6 This emphasizes the need for HIV-1 prevention products that work in women with vaginal dysbiosis.

Acknowledgements Study participants National Institute of Allergy and Infectious Diseases, National Institutes of Health, Grant AI082639 and UL1 TR000005. CONRAD was the regulatory sponsor of the tenofovir trial and donated tenofovir drug substance and some study product. IPM was the regulatory sponsor for the dapivirine trial and donated drug substance and study product.