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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 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.Formulation assessment of tenofovir gel included osmolality, viscosity, in vitro release, and permeability testing. Safety was evaluated by measuring the effect on the viability of vaginal flora, PBMCs, epithelial cells, and ectocervical and colorectal explant tissues. For efficacy testing, PBMCs were cultured with tenofovir or vehicle control gels and HIV-1 representing subtypes A, B, and C. Additionally, polarized ectocervical and colorectal explant cultures were treated apically with either gel. Tenofovir was added basolaterally to simulate systemic application. All tissues were challenged with HIV-1 applied apically. Infection was assessed by measuring p24 by ELISA on collected supernatants and immunohistochemistry for ectocervical explants. Formulation testing showed the tenofovir and vehicle control gels were >10 times isosmolar. Permeability through ectocervical tissue was variable but in all cases the receptor compartment drug concentration reached levels that inhibit HIV-1 infection in vitro. The gels were non-toxic toward vaginal flora, PBMCs, or epithelial cells. A transient reduction in epithelial monolayer integrity and epithelial fracture for ectocervical and colorectal explants was noted and likely due to the hyperosmolar nature of the formulation. Tenofovir gel prevented HIV-1 infection of PBMCs regardless of HIV-1 subtype. Topical and systemic tenofovir were effective at preventing HIV-1 infection of explant cultures.These studies provide a mechanism for pre-clinical prediction of safety and efficacy of formulated microbicides. Tenofovir was effective against HIV-1 infection in our algorithm. These data support the use of tenofovir for pre-exposure prophylaxis.

An abstract of the article "Defining Pharmacokinetic and Pharmacodynamic linkages between Genital Tissue and Lumen Compartments" by Charlene S. Dezzutti and colleagues is presented.

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.

Human immunodeficiency virus (HIV) infection and unintended pregnancy, which can lead to life-threatening complications, are two major burdens for female reproductive health. To address these pressing health issues, multipurpose prevention technologies (MPTs) are proposed to deliver two or more drugs simultaneously. MPTs could offer several benefits for users such as improved convenience, increased effectiveness, reduced cost, and decreased environmental burden. Here, we report the development, and in vitro and in vivo assessment of a bioadhesive vaginal film as a coitally-independent MPT dosage form for delivering dapivirine (DPV) and levonorgestrel (LNG) to prevent HIV infection and unintended pregnancy, respectively. After confirming the feasibility of bioadhesive film use for weekly drug delivery in vivo through colpophotography and MRI evaluation, the pharmacokinetics (PK) of DPV/LNG single entity and combination bioadhesive films was investigated in pigtailed macaques ( n = 5). Both drugs from single entity or combination films were able to provide sustained drug release in vivo. The combination film showed lower local tissue clearance for DPV and exhibited significantly increased plasma concentration for LNG as compared to the single entity film. This proof-of-concept study demonstrates the ability of this novel bioadhesive film platform to deliver LNG and DPV simultaneously as an MPT product for the prevention of HIV infection and unintended pregnancy., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

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., Methods and Findings: Formulation assessment of tenofovir gel included osmolality, viscosity, in vitro release, and permeability testing. Safety was evaluated by measuring the effect on the viability of vaginal flora, PBMCs, epithelial cells, and ectocervical and colorectal explant tissues. For efficacy testing, PBMCs were cultured with tenofovir or vehicle control gels and HIV-1 representing subtypes A, B, and C. Additionally, polarized ectocervical and colorectal explant cultures were treated apically with either gel. Tenofovir was added basolaterally to simulate systemic application. All tissues were challenged with HIV-1 applied apically. Infection was assessed by measuring p24 by ELISA on collected supernatants and immunohistochemistry for ectocervical explants. Formulation testing showed the tenofovir and vehicle control gels were >10 times isosmolar. Permeability through ectocervical tissue was variable but in all cases the receptor compartment drug concentration reached levels that inhibit HIV-1 infection in vitro. The gels were non-toxic toward vaginal flora, PBMCs, or epithelial cells. A transient reduction in epithelial monolayer integrity and epithelial fracture for ectocervical and colorectal explants was noted and likely due to the hyperosmolar nature of the formulation. Tenofovir gel prevented HIV-1 infection of PBMCs regardless of HIV-1 subtype. Topical and systemic tenofovir were effective at preventing HIV-1 infection of explant cultures., Conclusions: These studies provide a mechanism for pre-clinical prediction of safety and efficacy of formulated microbicides. Tenofovir was effective against HIV-1 infection in our algorithm. These data support the use of tenofovir for pre-exposure prophylaxis.

The photolytic degradations of imidacloprid, carbofuran, diazinon, chlorpyrifos, pyridaben, propoxur, and esfenvalerate were independently compared in both moist (75% field moisture capacity at 0.33 bar) and air-dry microbially viable soils at 5 microg/g. All compounds were applied to sandy soil except for propoxur, which was applied to sandy loam soil. Diazinon was applied to both sandy soil and sandy loam soil. The samples were exposed for up to 360 h, depending on the half-life of the compound. Moisture and temperature were maintained through the use of a specially designed soil photolysis apparatus. Corresponding dark control studies were performed concurrently. With the exception of esfenvalerate, the other compounds exhibited significantly shorter half-lives in moist soils, attributed to the increased hydrolysis and microbial activity of the moist soil. The esfenvalerate metabolism was not first order due to limited mobility in the soil because of its very low water solubility. The overall half-life for esfenvalerate was 740 h, as the percent remaining did not drop below 60%. The imidacloprid half-life in irradiated moist soil was 1.8 times shorter than in air-dry soils. However, on dry soil the photodegradation showed poor first-order kinetics after 24 h of exposure. The metabolism of carbofuran and diazinon was highly dependent on soil moisture. Carbofuran exhibited 2.2 times longer half-lives when less moisture was available in the soil. Diazinon in moist sandy soil degraded rapidly, but slowed significantly in irradiated and dark control air-dry sandy soil. Diazinon photolysis on sandy loam soil was not first order, as it attained a constant concentration of 54.9%, attributed to decreased mobility in this soil. Chlorpyrifos photolysis was 30% shorter on moist sand than on air-dry sand. Pyridaben photolyzed rapidly throughout the first 72 h of irradiation but maintained 48% through 168 h. Propoxur metabolism in moist sandy loam soil was not first order and did not degrade below 50% after 360 h of exposure, but the overall half-life was still nearly half of that on irradiated air-dry soil. Three of the compounds showed differences in metabolism patterns during exposure on moist or air-dry soil. Typically, the moist soils produced a more linear decline than that seen in the dry soils, corresponding to the susceptibility of the particular chemical to hydrolysis and/or biodegradation. Four of the eight experiments had shorter half-lives in dark control moist soils than in irradiated dry soils.

The problem of maintaining the moisture content of samples throughout the course of a soil photolysis study is addressed. The photolytic degradations of asulam, triclopyr, acifluorfen, and atrazine were independently compared in air-dried soils and in moist (75% field moisture capacity at 0.33 bar) soils maintained at initial conditions through the use of a specially designed soil photolysis apparatus. Each pesticide was applied at 5 microg/g. The exposure phase extended from 144 to 360 h, depending on the half-life of the compound. A dark control study, also using moist and air-dried soils, was performed concurrently at 25 degrees C. The results showed significant differences in half-life. The dissipations generally demonstrated a strong dependence on moisture. In most cases, photolytic degradation on air-dried soil was longer than in the moist dark control soils. Half-lives in dry soil were 2-7 times longer, and in the case of atrazine, the absence of moisture precluded significant degradation. Moist soil experiments also tended to correlate more strongly with linear first-order degradations. The dark control experiments also demonstrated shorter half-lives in moist soil. Moisture was also observed to affect the amount of degradate formed in the soils.

An environmental fate study was performed analyzing the effects of soil composition on the soil photolysis of a chemical. The study was conducted in two phases in which both moist and air-dried soils were fortified with either the common fertilizer sodium nitrate or the natural soil components iron or humic acid and dosed with niclosamide. The soils were photolyzed under a xenon lamp for 7 days. Increasing concentration of sodium nitrate did not affect the degradation pattern but did produce a lower concentration of aminoniclosamide. Soils fortified with iron displayed an unknown, which was not observed in other experiments, and the degradation of niclosamide from these soils was slower than from the sodium nitrate-fortified soils. There were no extractable degradates from any of the soils fortified with humic acid. In irradiated moist soils, the half-life of niclosamide increased when sodium nitrate was present at 20 ppm, and the half-lives of niclosamide in iron- and humic acid-fortified soil were increased slightly over that in unfortified soil. The effect of the nitrate and iron on the half-lives in dark control moist soils was minimal, but humic acid increased the dark control half-life from 420 to 611 h. No transformation of niclosamide was observed in the dark control air-dried soils. Soils with higher organic or iron contents or exposed to fertilizers do not affect as dramatically the half-life of pesticides as does the presence of moisture in the soil. Soil photolysis samples that were not maintained with moisture exhibited differences in half-life and degradation pattern. The maintenance of moisture was found to be more crucial to the reliability of soil photolysis studies than soil composition.

The effects of soil depth and moisture on pesticide photolysis were studied. Moist soil at depths of 3, 2.5, 2, 1.5, 1, and 0.5 mm were each dosed at 2.5 microg/g with (14)C-niclosamide and photolyzed under a xenon lamp at constant temperature. Samples were removed after 20, 40, 110, and 153 h of continuous irradiation. The decrease in percent of niclosamide and the appearance of degradates were followed by analyzing the soil extracts by HPLC. A corresponding set of experiments used air-dried soil. An experiment was also performed using initially moist soil which was permitted to dry during photolysis but returned to moist conditions at each sampling. Qualitative and quantitative differences were found in the rate and route of degradation of niclosamide under these conditions. These differences have resulted from a combination of reduced photochemical activity and microbial population in dry soil. The half-lives of niclosamide in the dry soils were 2 to 5 times longer than those in the moisture-maintained soil. There was also a noticeable difference in the half-lives in soil of different depths. Moisture-maintained soil showed a uniform linear increase in half-life from 95 to 195 h as soil depth increased from 0.5 mm to 3.0 mm. With air-dried soil the half-lives were greatly dependent on soil depth, showing a much broader range of 199 h at 0.5-mm to 1064 h in 3.0-mm soil. An experimental design is described which maintains soil temperature and moisture to preset conditions.