BackgroundRepeated use, coitus-independent microbicide gels that do not contain antiretroviral agents also used as first line HIV therapy are urgently needed to curb HIV spread. Current formulations require high doses (millimolar range) of antiretroviral drugs and typically only provide short-term protection in macaques. We used the macaque model to test the efficacy of a novel combination microbicide gel containing zinc acetate and micromolar doses of the novel non-nucleoside reverse transcriptase inhibitor MIV-150 for up to 24 h after repeated gel application.Methods and FindingsRhesus macaques were vaginally challenged with SHIV-RT up to 24 h after repeated administration of microbicide versus placebo gels. Infection status was determined by measuring virologic and immunologic parameters. Combination microbicide gels containing 14 mM zinc acetate dihydrate and 50 µM MIV-150 afforded full protection (21 of 21 animals) for up to 24 h after 2 weeks of daily application. Partial protection was achieved with the MIV-150 gel (56% of control at 8 h after last application, 11% at 24 h), while the zinc acetate gel afforded more pronounced protection (67% at 8–24 h). Marked protection persisted when the zinc acetate or MIV-150/zinc acetate gels were applied every other day for 4 weeks prior to challenge 24 h after the last gel was administered (11 of 14 protected). More MIV-150 was associated with cervical tissue 8 h after daily dosing of MIV-150/zinc acetate versus MIV-150, while comparable MIV-150 levels were associated with vaginal tissues and at 24 h.ConclusionsA combination MIV-150/zinc acetate gel and a zinc acetate gel provide significant protection against SHIV-RT infection for up to 24 h. This represents a novel advancement, identifying microbicides that do not contain anti-viral agents used to treat HIV infection and which can be used repeatedly and independently of coitus, and underscores the need for future clinical testing of their safety and ability to prevent HIV transmission in humans.

Anti-HIV microbicides are being investigated in clinical trials and understanding how promising strategies work, coincident with demonstrating efficacy in vivo, is central to advancing new generation microbicides. We evaluated Carraguard® and a new generation Carraguard-based formulation containing the non-nucleoside reverse transcriptase inhibitor (NNRTI) MIV-150 (PC-817). Since dendritic cells (DCs) are believed to be important in HIV transmission, the formulations were tested for the ability to limit DC-driven infection in vitro versus vaginal infection of macaques with RT-SHIV (SIVmac239 bearing HIV reverse transcriptase). Carraguard showed limited activity against cell-free and mature DC-driven RT-SHIV infections and, surprisingly, low doses of Carraguard enhanced infection. However, nanomolar amounts of MIV-150 overcame enhancement and blocked DC-transmitted infection. In contrast, Carraguard impeded infection of immature DCs coincident with DC maturation. Despite this variable activity in vitro, Carraguard and PC-817 prevented vaginal transmission of RT-SHIV when applied 30 min prior to challenge. PC-817 appeared no more effective than Carraguard in vivo, due to the limited activity of a single dose of MIV-150 and the dominant barrier effect of Carraguard. However, 3 doses of MIV-150 in placebo gel at and around challenge limited vaginal infection, demonstrating the potential activity of a topically applied NNRTI. These data demonstrate discordant observations when comparing in vitro and in vivo efficacy of Carraguard-based microbicides, highlighting the difficulties in testing putative anti-viral strategies in vitro to predict in vivo activity. This work also underscores the potential of Carraguard-based formulations for the delivery of anti-viral drugs to prevent vaginal HIV infection.

There are two principle subsets of dendritic cells (DCs); CD11c+CD123− myeloid DCs (MDCs) and CD11c−CD123+ plasmacytoid DCs (PDCs). DC activation via TNF-TNFRs (e.g., CD40L) and TLRs (e.g., immunostimulatory oligodeoxyribonucleotides (ISS-ODNs)) is crucial for maximal stimulation of innate and adaptive immunity. Macaque DC biology is being studied to improve HIV vaccines using the SIV macaque model. Using lineage (Lin) markers to exclude non-DCs, Lin−HLA-DR+CD11c+CD123− MDCs and Lin−HLA-DR+CD11c−CD123+ PDCs were identified in the blood of uninfected macaques and healthy macaques infected with SIV or simian-human immunodeficiency virus. Overnight culture of DC-enriched Lin-depleted cells increased CD80 and CD86 expression. IL-12 production and CD80/CD86 expression by MDC/PDC mixtures was further enhanced by CD40L and ISS-ODN treatment. A CpG-B ISS-ODN increased CD80/CD86 expression by PDCs, but resulted in little IFN-α secretion unless IL-3 was added. In contrast, a CpG-C ISS-ODN and aldrithiol-2-inactivated (AT-2) SIV induced considerable PDC activation and IFN-α release without needing exogenous IL-3. The CpG-C ISS-ODN also stimulated IL-12 release (unlike AT-2 SIV) and augmented DC immunostimulatory activity, increasing SIV-specific T cell IFN-γ production induced by AT-2 SIV-presenting MDC/PDC-enriched mixtures. These data highlight the functional capacities of MDCs and PDCs in naive as well as healthy, infected macaques, revealing a promising CpG-C ISS-ODN-driven DC activation strategy that boosts immune function to augment preventative and therapeutic vaccine efficacy.

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 106 pfu HSV-2 were applied immediately after vaginal challenge and also when 5×103 pfu were applied between 8 h before and 4 h after vaginal challenge (p<0.0248). Protection of mice against 8×106 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.

BackgroundHerpes simplex virus type-2 (HSV-2) infection enhances the transmission and acquisition of human immunodeficiency virus (HIV). This occurs in symptomatic and asymptomatic stages of HSV-2 infection, suggesting that obvious herpetic lesions are not required to increase HIV spread. An animal model to investigate the underlying causes of the synergistic action of the two viruses and where preventative strategies can be tested under such complex physiological conditions is currently unavailable.Methodology/Principal FindingsWe set out to establish a rhesus macaque model in which HSV-2 infection increases the susceptibility to vaginal infection with a model immunodeficiency virus (simian-human immunodeficiency virus, SHIV-RT), and to more stringently test promising microbicides. HSV-2 exposure significantly increased the frequency of vaginal SHIV-RT infection (n = 6). Although cervical lesions were detected in only ∼10% of the animals, long term HSV-2 DNA shedding was detected (in 50% of animals followed for 2 years). Vaginal HSV-2 exposure elicited local cytokine/chemokine (n = 12) and systemic low-level HSV-2-specific adaptive responses in all animals (n = 8), involving CD4+ and CD8+ HSV-specific T cells (n = 5). Local cytokine/chemokine responses were lower in co-infected animals, while simian immunodeficiency virus (SIV)-specific adaptive responses were comparable in naïve and HSV-2-infected animals (n = 6). Despite the increased frequency of SHIV-RT infection, a new generation microbicide gel, comprised of Carraguard® and a non-nucleoside reverse transcriptase inhibitor MIV-150 (PC-817), blocked vaginal SHIV-RT infection in HSV-2-exposed animals (n = 8), just as in naïve animals.Conclusions/SignificanceWe established a unique HSV-2 macaque model that will likely facilitate research to define how HSV-2 increases HIV transmission, and enable more rigorous evaluation of candidate anti-viral approaches in vivo.