vitro and ex vivo testing of tenofovir shows it is effective as an HIV-1 microbicide

Abstract Background: Tenofovir gel has entered into clinical trials for use as a topical microbicide to prevent HIV-1 infection but has no published data regarding pre-clinical testing using in vitro and ex vivo models. To validate our findings with on-going clinical trial results, we evaluated topical tenofovir gel for safety and efficacy. We also modeled systemic application of tenofovir for efficacy

Investigating the Contribution of Drug-Metabolizing Enzymes in Drug-Drug Interactions of Dapivirine and Miconazole

Dapivirine (DPV) is a potent NNRTI used to prevent the sexual transmission of HIV. In a phase 1 trial (IPM 028), the concomitant use of a DPV vaginal ring and an antifungal miconazole (MIC) vaginal capsule was found to increase the systemic exposure to DPV in women, suggesting a potential for drug-drug interactions. This study’s objective was to investigate the mechanism of DPV-MIC interactions using drug-metabolizing enzymes (DMEs; CYPs and UGTs) that are locally expressed in the female reproductive tract (FRT). In vitro studies were performed to evaluate the metabolism of DPV and its inhibition and induction potential with DMEs. In addition, the impact of MIC on DPV metabolism and the inhibitory potential of DPV with DMEs were studied. Our findings suggest that DPV is a substrate of CYP1A1 and CYP3A4 enzymes and that MIC significantly decreased the DPV metabolism by inhibiting these two enzymes. DPV demonstrated potent inhibition of CYP1A1 and moderate/weak inhibition of the six CYP and eight UGT enzymes evaluated. MIC showed potent/moderate inhibition of seven CYP enzymes and weak/no inhibition of eight UGT enzymes. The combination of DPV and MIC showed potent inhibition of seven CYP enzymes (1A1, 1A2, 1B1, 2B6, 2C8, 2C19, and 3A4) and four UGT enzymes (1A3, 1A6, 1A9, and 2B7). DPV was not an inducer of CYP1A2, CYP2B6, and CYP3A4 enzymes in primary human hepatocytes. Therefore, the increased systemic concentrations of DPV observed in IPM 028 were likely due to the reduced metabolism of DPV because of CYP1A1 and CYP3A4 enzymes inhibition by MIC in the FRT

Tenofovir releases from the formulated gel.

<p>The <i>in vitr</i>o release data show the release profile of tenofovir from the gel; the slope of the line represents the release rate of the product. The data shown represent the mean ± standard deviation of 13 replicates.</p

Viability of explant cultures after a 24 hour exposure to tenofovir or vehicle control gels.

<p>Ectocervical and colorectal explants were polarized and tenofovir or vehicle control gels were diluted 1:5 in the appropriate culture medium and applied to the apical surface. A 1:5 dilution of nonoxynol-9 (N9) was applied apically to the explants at the same time as the tenofovir and vehicle control gels as a toxicity control. Untreated explants were the negative control tissues. The explants were cultured for 24 h, washed five times, and placed in either medium containing 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT) to assess tissue viability by measuring mitochondrial activity (A) or formalin to fix the tissue for hematoxylin-eosin staining for histology (original magnification 20×; bar length 0.05 mm) (B). Explants from five tissue donors were evaluated for viability after exposure to topical gels. The % viability was determined by dividing the corrected optical density of the treated explant by the corrected optical density of the control explant. Histology shown is representative of the five tissues evaluated.</p

Efficacy of tenofovir against primary isolates of HIV-1.

<p>Peripheral blood mononuclear cells were activated and cultured with HIV-1 (BaL, laboratory-adapted CCR5-using clade B isolate; A103, primary CCR5/CXCR4-using clade A isolate; C012, primary CCR5-using clade C isolate; and C959, primary CCR5-using clade C isolate) with or without tenofovir or vehicle control gel. After 4 hours, the cultures were washed and fresh medium was added. Supernatant was collected every 3 to 4 days and stored at -80°C. HIV-1 infection was followed using a p24gag ELISA. The data shown represent the log₁₀-transformed (pg/mL) ±95% confidence interval of 4 (BaL) or 5 (A103, C012, and C959) independent experiments.</p