Vaginal lactic acid elicits an anti-inflammatory response from human cervicovaginal epithelial cells and inhibits production of pro-inflammatory mediators associated with HIV acquisition.

Inflammation in the female reproductive tract (FRT) is associated with increased HIV transmission. Lactobacillus spp. dominate the vaginal microbiota of many women and their presence is associated with reduced HIV acquisition. Here we demonstrate that lactic acid (LA), a major organic acid metabolite produced by lactobacilli, mediates anti-inflammatory effects on human cervicovaginal epithelial cells. Treatment of human vaginal and cervical epithelial cell lines with LA (pH 3.9) elicited significant increases in the production of the anti-inflammatory cytokine IL-1RA. When added simultaneously or prior to stimulation, LA inhibited the Toll-like receptor agonist-elicited production of inflammatory mediators IL-6, IL-8, TNFα, RANTES, and MIP3α from epithelial cell lines and prevented IL-6 and IL-8 production by seminal plasma. The anti-inflammatory effect of LA was mediated by the protonated form present at pH≤3.86 and was observed with both L- and D-isomers. A similar anti-inflammatory effect of LA was observed in primary cervicovaginal cells and in an organotypic epithelial tissue model. These findings identify a novel property of LA that acts directly on epithelial cells to inhibit FRT inflammation and highlights the potential use of LA-containing agents in the lower FRT as adjuncts to female-initiated strategies to reduce HIV acquisition.

Synthesis, stability, antiviral activity, and protease-bound structures of substrate-mimicking constrained macrocyclic inhibitors of HIV-1 protease.

Three new peptidomimetics (1-3) have been developed with highly stable and conformationally constrained macrocyclic components that replace tripeptide segments of protease substrates. Each compound inhibits both HIV-1 protease and viral replication (HIV-1, HIV-2) at nanomolar concentrations without cytotoxicity to uninfected cells below 10 microM. Their activities against HIV-1 protease (K(i) 1.7 nM (1), 0.6 nM (2), 0.3 nM (3)) are 1-2 orders of magnitude greater than their antiviral potencies against HIV-1-infected primary peripheral blood mononuclear cells (IC(50) 45 nM (1), 56 nM (2), 95 nM (3)) or HIV-1-infected MT2 cells (IC(50) 90 nM (1), 60 nM (2)), suggesting suboptimal cellular uptake. However their antiviral potencies are similar to those of indinavir and amprenavir under identical conditions. There were significant differences in their capacities to inhibit the replication of HIV-1 and HIV-2 in infected MT2 cells, 1 being ineffective against HIV-2 while 2 was equally effective against both virus types. Evidence is presented that 1 and 2 inhibit cleavage of the HIV-1 structural protein precursor Pr55(gag) to p24 in virions derived from chronically infected cells, consistent with inhibition of the viral protease in cells. Crystal structures refined to 1.75 A (1) and 1.85 A (2) for two of the macrocyclic inhibitors bound to HIV-1 protease establish structural mimicry of the tripeptides that the cycles were designed to imitate. Structural comparisons between protease-bound macrocyclic inhibitors, VX478 (amprenavir), and L-735,524 (indinavir) show that their common acyclic components share the same space in the active site of the enzyme and make identical interactions with enzyme residues. This substrate-mimicking minimalist approach to drug design could have benefits in the context of viral resistance, since mutations which induce inhibitor resistance may also be those which prevent substrate processing.

Effect of protease inhibitors on HIV-1 maturation and infectivity.

The effects of HIV-1 protease inhibitors on proteolytic processing and infectivity of virions produced from lymphocytes chronically infected with the virus were studied. Protease inhibition was detected by the accumulation of the polyprotein precursors Pr55gag and Pr160gag-pol and their cleavage intermediates. Immunoblot analysis demonstrated that while the processing of Pr55gag was largely irreversible, cleavage of Pr160gag-pol proceeded once the inhibitor was removed, although it was not completed during 96 h of subsequent observation. Virions produced during exposure of cells to protease inhibitors regained some degree of infectivity post-withdrawal of the inhibitor, suggesting that the processing of Pr160gag-pol following drug withdrawal resulted in the production of those enzymes necessary to enable at least limited viral replication. When cells were exposed to a protease inhibitor for 72 h then the inhibitor withdrawn, a lag phase of up to 24 h occurred before these cells produced virions with equivalent infectivity to virus produced from cells not exposed to drug. These observations may reflect a clinical situation likely to occur as trough plasma concentrations of protease inhibitors fall below the IC100 for HIV, highlighting the need for adherence to drug regimens containing these inhibitors.

Clinical effects and in vitro studies of trifluorothymidine combined with interferon-alpha for treatment of drug-resistant and -sensitive herpes simplex virus infections.

Three AIDS patients with severe cutaneous herpes simplex virus (HSV) infection refractory to therapy with acyclovir and foscarnet (2 patients) were treated with a topical preparation of trifluorothymidine (TFT) and interferon-alpha. Complete healing of lesions occurred in 1 patient; a second had significant regression of the infected area. In the third, the lesion was stabilized twice after application of the preparation and reduced in size after a subsequent treatment. In vitro studies confirmed that isolates from these patients were acyclovir- or acyclovir/foscarnet-resistant. In addition, they revealed strong synergy between TFT and interferon-alpha for these isolates and for strains with wild-type drug sensitivity profiles. Topical TFT/interferon-alpha may be of benefit in the therapy of mucocutaneous HSV infections, especially when they are resistant to treatment with systemic antiviral agents.