Preclinical Testing of Candidate Topical Microbicides for Anti-Human Immunodeficiency Virus Type 1 Activity and Tissue Toxicity in a Human Cervical Explant Culture

ABSTRACT A human cervical explant culture was utilized for the preclinical assessment of anti-human immunodeficiency virus type 1 (HIV-1) activity and tissue toxicity of formulated, candidate topical microbicides. Products tested included cellulose acetate 1,2-benzene dicarboxylate (CAP), a carrageenan-based product (PC-515), a naphthalene sulfonate polymer (PRO 2000), a lysine dendrimer (SPL7013), a nonnucleoside reverse transcriptase inhibitor (UC781), and an antimicrobial peptide (D2A21), along with their placebos. Cervical explants were cultured overnight with HIV-1 with or without product, washed, and monitored for signs of HIV-1 infection. HIV-1 infection was determined by p24gag levels in the basolateral medium and by immunohistochemical analysis of the explant. Product toxicity was measured by the MTT [1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan] assay and histology. CAP, PRO 2000, SPL7013, and UC781 consistently prevented HIV-1 infection in all explants tested. PC-515 and D2A21 prevented HIV-1 infection in 50% or fewer of the explants tested. Placebos did not prevent infection in any of the explants tested. With the exception of PRO 2000 (4%), the MTT assay and histological analysis of the other products and placebos showed minimal toxicity to the epithelium and submucosa. Collectively, these data suggest that this culture system can be used for evaluating the safety and efficacy of topical microbicides designed for vaginal use.

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Human Immunodeficiency Virus Type 1 Nucleocapsid Inhibitors Impede trans Infection in Cellular and Explant Models and Protect Nonhuman Primates from Infection

Journal of Virology ◽

10.1128/jvi.00820-09 ◽

2009 ◽

Vol 83 (18) ◽

pp. 9175-9182 ◽

Cited By ~ 31

Author(s):

Gregory S. Wallace ◽

Cecilia Cheng-Mayer ◽

Marco L. Schito ◽

Patricia Fletcher ◽

Lisa M. Miller Jenkins ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Drug Candidates ◽

Trans Infection ◽

Immunodeficiency Virus ◽

Cervical Explant ◽

Infected Cells ◽

Hiv 1

ABSTRACT Here, we report that the S-acyl-2-mercaptobenzamide thioester (SAMT) class of human immunodeficiency virus type 1 (HIV-1) nucleocapsid protein (NCp7) inhibitors was able to prevent transmission of HIV-1 from infected cells, including primary cells. Furthermore, when SAMTs were introduced during an HIV-1 challenge of cervical explant tissue, inhibition of dissemination of infectious virus by cells emigrating from the tissue explants was observed. Preliminary studies using a rhesus macaque vaginal challenge model with mixed R5 and X4 simian-human immunodeficiency virus infection found that five of six monkeys were completely protected, with the remaining animal being partially protected, infected only by the R5 virus. These data suggest that SAMTs may be promising new drug candidates for further development in anti-HIV-1 topical microbicide applications.

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Effect of Topical Microbicides on Infectious Human Immunodeficiency Virus Type 1 Binding to Epithelial Cells

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01358-06 ◽

2007 ◽

Vol 51 (6) ◽

pp. 1972-1978 ◽

Cited By ~ 9

Author(s):

Susan Roth ◽

Michael Monsour ◽

Amanda Dowland ◽

Patricia C. Guenthner ◽

Kelly Hancock ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Epithelial Cell ◽

Epithelial Cells ◽

Cell Line ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Immunodeficiency Virus ◽

Topical Microbicides ◽

Hiv 1

ABSTRACT Topical microbicides (cellulose acetate 1,2 benzene dicarboxylate [CAP], PRO 2000, SPL7013, and UC781) are being investigated to reduce the sexual transmission of human immunodeficiency virus type 1 (HIV-1). These products were shown to prevent the transfer of infectious HIV-1 from urogenital and colorectal epithelial cell lines to peripheral blood mononuclear cells. However, it was unclear if the topical microbicides rendered the virus noninfectious and/or reduced the binding to the epithelial cells. To test this, epithelial cells were cultured with HIV-1 in the presence or absence of topical microbicides or their placebos. The cells were washed, RNA lysates were made, and real-time PCR was performed for HIV-1. PRO 2000 and SPL7013 significantly (P < 0.0001) reduced the amount of bound HIV-1 to the colorectal epithelial cell line across clades A, B, C, and CRF01-AE. While none of the products reduced the binding of HIV-1 clades A and C to the urogenital cell line, CAP, PRO 2000, and SPL7013 significantly (P ≤ 0.002) reduced the binding of clades B and CRF01-AE. In general, PRO 2000 and SPL7013 placebos significantly (P < 0.0001) reduced the amount of bound HIV-1 but were less than the active products. UC781, its placebo, and hydroxyethyl cellulose (placebo for CAP) minimally affected the amount of bound HIV-1. These results suggest that rendering HIV-1 noninfectious may not correlate to the amount of HIV-1 bound to epithelial cells and possible shedding into mucosal secretions. Therefore, functional virological assays in addition to measuring viral RNA should be included when clinically evaluating topical microbicide use by infected persons.

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Development of a Comprehensive Human Immunodeficiency Virus Type 1 Screening Algorithm for Discovery and Preclinical Testing of Topical Microbicides

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01328-07 ◽

2008 ◽

Vol 52 (5) ◽

pp. 1768-1781 ◽

Cited By ~ 78

Author(s):

Carol Lackman-Smith ◽

Clay Osterling ◽

Katherine Luckenbaugh ◽

Marie Mankowski ◽

Beth Snyder ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Inhibitory Activity ◽

Screening Algorithm ◽

Immunodeficiency Virus ◽

Topical Microbicides ◽

Hiv 1

ABSTRACT Topical microbicides are self-administered, prophylactic products for protection against sexually transmitted pathogens. A large number of compounds with known anti-human immunodeficiency virus type 1 (HIV-1) inhibitory activity have been proposed as candidate topical microbicides. To identify potential leads, an in vitro screening algorithm was developed to evaluate candidate microbicides in assays that assess inhibition of cell-associated and cell-free HIV-1 transmission, entry, and fusion. The algorithm advances compounds by evaluation in a series of defined assays that generate measurements of relative antiviral potency to determine advancement or failure. Initial testing consists of a dual determination of inhibitory activity in the CD4-dependent CCR5-tropic cell-associated transmission inhibition assay and in the CD4/CCR5-mediated HIV-1 entry assay. The activity is confirmed by repeat testing, and identified actives are advanced to secondary screens to determine their effect on transmission of CXCR4-tropic viruses in the presence or absence of CD4 and their ability to inhibit CXCR4- and CCR5-tropic envelope-mediated cell-to-cell fusion. In addition, confirmed active compounds are also evaluated in the presence of human seminal plasma, in assays incorporating a pH 4 to 7 transition, and for growth inhibition of relevant strains of lactobacilli. Leads may then be advanced for specialized testing, including determinations in human cervical explants and in peripheral blood mononuclear cells against primary HIV subtypes, combination testing with other inhibitors, and additional cytotoxicity assays. PRO 2000 and SPL7013 (the active component of VivaGel), two microbicide products currently being evaluated in human clinical trials, were tested in this in vitro algorithm and were shown to be highly active against CCR5- and CXCR4-tropic HIV-1 infection.

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