The effect of menopause on the innate antiviral activity of cervicovaginal lavage.

OBJECTIVE: Reproductive hormones are known to impact innate mucosal immune function of the lower genital tract. Our objectives were to determine the effect of hormonal status on intrinsic antiviral (herpes simplex virus [HSV]-1, HSV-2, and human immunodeficiency virus [HIV]-1) activity of cervicovaginal lavage (CVL). STUDY DESIGN: CVL was collected from 165 asymptomatic women including postmenopausal women (n = 29); women not on contraception in days 1-14 (n = 26) or days 15-28 (n = 27) of the menstrual cycle; and women using the levonorgestrel intrauterine device (n = 28), depot medroxyprogesterone acetate (n = 28), or combined oral contraceptives (n = 27). The anti-HSV-1/-2 and the anti-HIV-1 activity of the CVL were measured using plaque assays and the Jurkat-Tat-CCR5 assay, respectively. RESULTS: CVL from all of the groups had modest antiviral activity. Anti-HIV-1 activity was decreased in CVL from postmenopausal women when compared to premenopausal women (11% vs 34%, P = .002). However, there was no difference in anti-HIV-1 activity among premenopausal women regardless of phase of menstrual cycle or contraceptive use. Anti-HIV-1 activity was associated with the protein content of the CVL (r = 0.44, P < .001). There was no difference in anti-HSV-1 or -2 activity by hormonal group. CONCLUSION: Menopause is associated with decreased innate HIV-1 activity in the lower genital tract, suggesting that factors in the vaginal fluid could play a role in increased susceptibility of HIV-1 infection in postmenopausal women. Hormonal contraceptive use, menopause, and phase of menstrual cycle did not have a measurable impact on the intrinsic anti-HSV-1 or -2 activity.

Formulation and characterization of polymeric films containing combinations of antiretrovirals (ARVs) for HIV prevention.

PURPOSE: To develop polymeric films containing dual combinations of anti-HIV drug candidate tenofovir, maraviroc and dapivirine for vaginal application as topical microbicides. METHODS: A solvent casting method was used to manufacture the films. Solid phase solubility was used to identify potential polymers for use in the film formulation. Physical and chemical properties (such as water content, puncture strength and in vitro release) and product stability were determined. The bioactivity of the film products against HIV was assessed using the TZM-bl assay and a cervical explant model. RESULTS: Polymers identified from the solid phase solubility study maintained tenofovir and maraviroc in an amorphous state and prevented drug crystallization. Three combination film products were developed using cellulose polymers and polyvinyl alcohol. The residual water content in all films was <10% (w/w). All films delivered the active agents with release of >50% of film drug content within 30 min. Stability testing confirmed that the combination film products were stable for 12 months at ambient temperature and 6 months under stressed conditions. Antiviral activity was confirmed in TZM-bl and cervical explant models. CONCLUSIONS: Polymeric films can be used as a stable dosage form for the delivery of antiretroviral combinations as microbicides.

HIV-1 infection of female genital tract tissue for use in prevention studies.

OBJECTIVE: Ex vivo HIV-1 challenge has been proposed as a bioindicator of microbicide product effectiveness. The objective of this study was to establish optimal parameters for use of female genital tract tissue in this model. DESIGN: Ex vivo challenge involves in vivo product use, followed by tissue biopsy, and exposure of the tissue to HIV-1 in the laboratory. METHODS: Paired ectocervical and vaginal biopsies were collected from 42 women, and 28 women had additional biopsies from each site collected after 5% lidocaine (n = 14) or chlorhexidine (n = 14) treatment. Tissues were transported immediately to the laboratory and exposed to HIV-1. HIV-1 infection was followed by p24 enzyme-linked immunosorbent assay on culture supernatants and at study end after weighing and fixing the tissue for immunohistochemistry to detect p24 expressing cells. RESULTS: Although both tissue types were equally infected with HIV-1 based on the immunohistochemistry results, ectocervical tissues had significantly higher HIV-1 replication than vaginal tissues (P < 0.005). Lidocaine and chlorhexidine had minimal impact on HIV-1 infection and replication. Point estimates for p24 levels were defined for 95% probability of p24-positive tissues and were 3.43 log10 for ectocervical tissue and 2.50 log10 for vaginal tissue based on the weight-adjusted cumulative p24 end points. CONCLUSIONS: Although similar proportions of ectocervical and vaginal tissues support HIV-1 infection, higher levels of HIV-1 replication were observed in ectocervical tissues. Defining point estimates for HIV-1 infection in fresh ectocervical and vaginal tissues provides valuable information for the evaluation of HIV-1 preventative treatments during early clinical studies.

Is wetter better? An evaluation of over-the-counter personal lubricants for safety and anti-HIV-1 activity.

Because lubricants may decrease trauma during coitus, it is hypothesized that they could aid in the prevention of HIV acquisition. Therefore, safety and anti-HIV-1 activity of over-the-counter (OTC) aqueous- (n = 10), lipid- (n = 2), and silicone-based (n = 2) products were tested. The rheological properties of the lipid-based lubricants precluded testing with the exception of explant safety testing. Six aqueous-based gels were hyperosmolar, two were nearly iso-osmolar, and two were hypo-osmolar. Evaluation of the panel of products showed Gynol II (a spermicidal gel containing 2% nonoxynol-9), KY Jelly, and Replens were toxic to Lactobacillus. Two nearly iso-osmolar aqueous- and both silicone-based gels were not toxic toward epithelial cell lines or ectocervical or colorectal explant tissues. Hyperosmolar lubricants demonstrated reduction of tissue viability and epithelial fracture/sloughing while the nearly iso-osmolar and silicon-based lubricants showed no significant changes in tissue viability or epithelial modifications. While most of the lubricants had no measurable anti-HIV-1 activity, three lubricants which retained cell viability did demonstrate modest anti-HIV-1 activity in vitro. To determine if this would result in protection of mucosal tissue or conversely determine if the epithelial damage associated with the hyperosmolar lubricants increased HIV-1 infection ex vivo, ectocervical tissue was exposed to selected lubricants and then challenged with HIV-1. None of the lubricants that had a moderate to high therapeutic index protected the mucosal tissue. These results show hyperosmolar lubricant gels were associated with cellular toxicity and epithelial damage while showing no anti-viral activity. The two iso-osmolar lubricants, Good Clean Love and PRÉ, and both silicone-based lubricants, Female Condom 2 lubricant and Wet Platinum, were the safest in our testing algorithm.

Reformulated tenofovir gel for use as a dual compartment microbicide.

OBJECTIVES: Coital use of 1% tenofovir gel was shown to be modestly effective at preventing HIV transmission when applied vaginally in the CAPRISA 004 trial. Because the gel is hyperosmolar, which would reduce the integrity of the epithelium and induce fluid movement into the lumen, rectal use may not be acceptable. This study evaluated the pre-clinical safety and efficacy of a reformulated (reduced osmolality) tenofovir gel product. METHODS: Reduced glycerine (RG)-tenofovir gel was compared with the original tenofovir gel for physiochemical characteristics, product safety and anti-HIV-1 activity. RESULTS: The formulations were similar in all characteristics except for osmolality and spreadability/firmness. The RG-tenofovir gel had a 73% lower osmolality, a 29.6% increase in spreadability and a 27% decrease in firmness as compared with the original tenofovir gel. When applied to epithelial cell monolayers, tenofovir gel showed a transient reduction in the transepithelial resistance while the RG-tenofovir gel did not. Both gels retained ectocervical and colorectal explant viability. However, tenofovir gel treatment resulted in epithelial stripping that was absent after RG-tenofovir gel treatment of the polarized explants. Anti-HIV-1 activity was confirmed by lack of HIV-1 infection in polarized explants treated with either gel as compared with the control explants. CONCLUSIONS: Reducing the osmolality of the tenofovir gel resulted in improved epithelial integrity, which suggests better safety upon rectal use. The improved gel safety did not compromise drug release or anti-HIV-1 activity. These data support the use of this gel as a dual compartment microbicide.