HIV pre-exposure prophylaxis (PrEP) is dominated by clinical therapeutic antiretroviral (ARV) drugs. Griffithsin (GRFT) is a non-ARV lectin with potent anti-HIV activity. GRFT's preclinical safety, lack of systemic absorption after vaginal administration in animal studies, and lack of cross-resistance with existing ARV drugs prompted its development for topical HIV PrEP. We investigated safety, pharmacokinetics (PK), pharmacodynamics (PD), and immunogenicity of PC-6500 (0.1% GRFT in a carrageenan (CG) gel) in healthy women after vaginal administration. This randomized, placebo-controlled, parallel group, double-blind first-in-human phase 1 study enrolled healthy, HIV-negative, non-pregnant women aged 24-45 years. In the open label period, all participants (n = 7) received single dose of PC-6500. In the randomized period, participants (n = 13) were instructed to self-administer 14 doses of PC-6500 or its matching CG placebo (PC-535) once daily for 14 days. The primary outcomes were safety and PK after single dose, and then after 14 days of dosing. Exploratory outcomes were GRFT concentrations in cervicovaginal fluids, PD, inflammatory mediators and gene expression in ectocervical biopsies. This trial is registered with ClinicalTrials.gov, number NCT02875119. No significant adverse events were recorded in clinical or laboratory results or histopathological evaluations in cervicovaginal mucosa, and no anti-drug (GRFT) antibodies were detected in serum. No cervicovaginal proinflammatory responses and no changes in the ectocervical transcriptome were evident. Decreased levels of proinflammatory chemokines (CXCL8, CCL5 and CCL20) were observed. GRFT was not detected in plasma. GRFT and GRFT/CG in cervicovaginal lavage samples inhibited HIV and HPV, respectively, in vitro in a dose-dependent fashion. These data suggest GRFT formulated in a CG gel is a safe and promising on-demand multipurpose prevention technology product that warrants further investigation.

To extend our observations that single or repeated application of a gel containing the NNRTI MIV-150 (M) and zinc acetate dihydrate (ZA) in carrageenan (CG) (MZC) inhibits vaginal transmission of simian/human immunodeficiency virus (SHIV)-RT in macaques, we evaluated safety and anti-SHIV-RT activity of MZC and related gel formulations ex vivo in macaque mucosal explants. In addition, safety was further evaluated in human ectocervical explants. The gels did not induce mucosal toxicity. A single ex vivo exposure to diluted MZC (1∶30, 1∶100) and MC (1∶30, the only dilution tested), but not to ZC gel, up to 4 days prior to viral challenge, significantly inhibited SHIV-RT infection in macaque vaginal mucosa. MZC's activity was not affected by seminal plasma. The antiviral activity of unformulated MIV-150 was not enhanced in the presence of ZA, suggesting that the antiviral activity of MZC was mediated predominantly by MIV-150. In vivo administration of MZC and CG significantly inhibited ex vivo SHIV-RT infection (51-62% inhibition relative to baselines) of vaginal (but not cervical) mucosa collected 24 h post last gel exposure, indicating barrier effect of CG. Although the inhibitory effect of MZC (65-74%) did not significantly differ from CG (32-45%), it was within the range of protection (∼75%) against vaginal SHIV-RT challenge 24 h after gel dosing. Overall, the data suggest that evaluation of candidate microbicides in macaque explants can inform macaque efficacy and clinical studies design. The data support advancing MZC gel for clinical evaluation.

Prevalent infection with human herpes simplex 2 (HSV-2) or human papillomavirus (HPV) is associated with increased human immunodeficiency virus (HIV) acquisition. Microbicides that target HIV as well as these sexually transmitted infections (STIs) may more effectively limit HIV incidence. Previously, we showed that a microbicide gel (MZC) containing MIV-150, zinc acetate (ZA) and carrageenan (CG) protected macaques against simian-human immunodeficiency virus (SHIV-RT) infection and that a ZC gel protected mice against HSV-2 infection. Here we evaluated a modified MZC gel (containing different buffers, co-solvents, and preservatives suitable for clinical testing) against both vaginal and rectal challenge of animals with SHIV-RT, HSV-2 or HPV. MZC was stable and safe in vitro (cell viability and monolayer integrity) and in vivo (histology). MZC protected macaques against vaginal (p<0.0001) SHIV-RT infection when applied up to 8 hours (h) prior to challenge. When used close to the time of challenge, MZC prevented rectal SHIV-RT infection of macaques similar to the CG control. MZC significantly reduced vaginal (p<0.0001) and anorectal (p = 0.0187) infection of mice when 10(6) pfu HSV-2 were applied immediately after vaginal challenge and also when 5×10(3) pfu were applied between 8 h before and 4 h after vaginal challenge (p<0.0248). Protection of mice against 8×10(6) HPV16 pseudovirus particles (HPV16 PsV) was significant for MZC applied up to 24 h before and 2 h after vaginal challenge (p<0.0001) and also if applied 2 h before or after anorectal challenge (p<0.0006). MZC provides a durable window of protection against vaginal infection with these three viruses and, against HSV-2 and HPV making it an excellent candidate microbicide for clinical use.

Women need multipurpose prevention products (MPTs) that protect against sexually transmitted infections (STIs) and provide contraception. The Population Council has developed a prototype intravaginal ring (IVR) releasing the non-nucleoside reverse transcriptase inhibitor (NNRTI) MIV-150 (M), zinc acetate (ZA), carrageenan (CG) and levonorgestrel (LNG) (MZCL IVR) to protect against HIV, HSV-2, HPV and unintended pregnancy. Our objective was to evaluate the anti-SHIV-RT activity of MZCL IVR in genital mucosa. First, macaque vaginal tissues were challenged with SHIV-RT in the presence of (i) MIV-150 ± LNG or (ii) vaginal fluids (VF); available from studies completed earlier) collected at various time points post insertion of MZCL and MZC IVRs. Then, (iii) MZCL IVRs (vs. LNG IVRs) were inserted in non-Depo Provera-treated macaques for 24h and VF, genital biopsies, and blood were collected and tissues were challenged with SHIV-RT. Infection was monitored with one step SIV gag qRT-PCR or p27 ELISA. MIV-150 (LCMS/MS, RIA), LNG (RIA) and CG (ELISA) were measured in different compartments. Log-normal generalized mixed linear models were used for analysis. LNG did not affect the anti-SHIV-RT activity of MIV-150 in vitro. MIV-150 in VF from MZC/MZCL IVR-treated macaques inhibited SHIV-RT in vaginal mucosa in a dose-dependent manner (p<0.05). MIV-150 in vaginal tissue from MZCL IVR-treated animals inhibited ex vivo infection relative to baseline (96%; p<0.0001) and post LNG IVR group (90%, p<0.001). No MIV-150 dose-dependent protection was observed, likely because of high MIV-150 concentrations in all vaginal tissue samples. In cervical tissue, MIV-150 inhibited infection vs. baseline (99%; p<0.05). No cervical tissue was available for MIV-150 measurement. Exposure to LNG IVR did not change tissue infection level. These observations support further development of MZCL IVR as a multipurpose prevention technology to improve women's sexual and reproductive health.