For example, if gel is applied shortly prior to intercourse, a substantial proportion may be lost due to leakage before or during sex. This may have little impact in the context of consistent daily gel application, but could result in insufficient drug in the setting of intermittent use and frequent sexual exposure. In addition, given that the volume of semen could be as large as the volume of gel within the vagina, the addition of semen may have a substantial dilutional effect on TFV concentrations within the lumen, even if there is no effect of semen on TFV activity, per se. Thus, additional postcoital sampling studies would provide important insights into optimal dosing frequency and may support the postcoital dosing design used in the CAPRISA 004 trial. It is noteworthy that while the anti-HIV activity in CVL obtained from women in the TFV group persisted when virus was introduced in human semen, the endogenous anti-HIV activity observed in women who received placebo was significantly reduced. Similar results were observed with postcoital CVL obtained in the absence of PRO 2000 gel application. Possibly, specific enzymes and/or proteases or the high pH of semen inactivate, degrade, or interfere with protective immune mediators AMN107 Src-bcr-Abl inhibitor present in the female genital secretions resulting in a reduction in endogenous antimicrobial activity. In addition to identifying potential biomarkers of microbicide PD and adherence, the current study also suggests that Tubulin Acetylation Inducer HDAC inhibitor measurements of endogenous antimicrobial activity and immune mediators in CVL may provide additional insights into microbicide safety. We observed no increase in proinflammatory cytokines or chemokines or loss in protective immune mediators or endogenous antimicrobial activity. Although our relatively small sample size may limit our ability to detect subtle changes in concentrations of immune mediators, our results are consistent with the safety of TFV gel demonstrated in the CAPRISA 004 study. While chemokines may block viral entry by binding to CCR5 and contribute to endogenous anti-HIV activity, increases in their expression may paradoxically facilitate infection by increasing target cell availability. This notion is supported by primate studies in which SIV induces the release of MIP-3a in the epithelium, which triggers the release of MIP-1b by plasmacytoid dendritic cells, resulting in subsequent recruitment of CD4+T cellsand the generation of a microenvironment conducive to transmission. In summary, this work supports the inclusion of quantified anti-HIV activity and drug levels in CVL as biomarkers of PD and PK and soluble mucosal immune mediators and endogenous activity as surrogate markers of safety in early clinical studies.