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Candidate polyanion microbicides inhibit HIV-1 infection and dissemination pathways in human cervical explants.

Fletcher PS, Wallace GS, Mesquita PM, Shattock RJ - Retrovirology (2006)

Bottom Line: While both compounds were virucidal for X4 HIV-1, neither was virucidal for R5 virus.Our results demonstrate that PRO 2000 is a potent inhibitor of R5 HIV-1 infection and dissemination pathways in human cervical explants.DxS, while demonstrating significant inhibition of R5 infection, was less active against DC mediated dissemination pathways.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Infection, Department of Cellular and Molecular Medicine, St George's, University of London, London, UK. pfletche@sgul.ac.uk

ABSTRACT

Background: Heterosexual intercourse remains the major route of HIV-1 transmission worldwide, with almost 5 million new infections occurring each year. Women increasingly bear a disproportionate burden of the pandemic, thus there is an urgent need to develop new strategies to reduce HIV-1 transmission that could be controlled by women themselves. The potential of topical microbicides to reduce HIV transmission across mucosal surfaces has been clearly identified, and some agents are currently under evaluation in clinical trials. Many of these "first generation" microbicides consist of polyanionic compounds designed to interfere with viral attachment. Here we have evaluated two candidate polyanion compounds in clinical trials, PRO 2000 and dextrin sulphate (DxS) to determine their safety and efficacy against in vitro HIV-1 and HSV-2 infection using cellular and tissue explant models.

Results: PRO 2000 and DxS potently inhibited infection by HIV-1 X4 and R5 isolates when present during viral exposure. However PRO 2000 required 10-fold and DxS 2000-fold more compound to block infection with R5 virus than X4. While both compounds were virucidal for X4 HIV-1, neither was virucidal for R5 virus. PRO 2000 efficiently inhibited infection of cervical explants and dissemination of virus by migratory DC. DxS was less active, able to completely inhibit cervical explant infection, but providing only partial reduction of virus dissemination by DC. PRO 2000, but not DxS, also inhibited HIV-1 binding to DC-SIGN+ cells and trans infection of co-cultured target cells. The inflammatory potential of both compounds was screened by measurement of cytokine production from cervical explants, and statistically significant increases were only observed for IL-1beta and RANTES following treatment with PRO 2000. Both compounds also demonstrated potent activity against HSV-2 infection of cervical epithelial cells.

Conclusion: Our results demonstrate that PRO 2000 is a potent inhibitor of R5 HIV-1 infection and dissemination pathways in human cervical explants. DxS, while demonstrating significant inhibition of R5 infection, was less active against DC mediated dissemination pathways. PRO 2000 has now entered human phase III efficacy trials.

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Inhibitory effect of polyanionic compounds against HSV-2 infection of epithelial cells. ME180 cells (seeded at 1.5 × 104 cells/well and cultured overnight), were exposed to HSV-2 (~5 × 104 Pfu/well) in the presence of compound for 1 hour, or alternatively, exposed to compound alone. Following compound/virus removal by washing, cells were cultured for a further 48 hours when viability was determined by MTT dye reduction. Cell viability (○, dotted line) following compound treatment was calculated as a percentage of the viability of cells exposed to culture medium alone. The effect of compound treatment on the infectivity of HSV-2 (●, solid line) was calculated as a percentage of infection observed in cells exposed to virus alone. Compounds tested were: A) PRO 2000; and B) Dextrin sulphate. Data represent the mean ± SEM of 3 independent experiments where each condition was tested in triplicate. Inserted figures represent the mean ± SEM concentration inhibiting 50% HSV-2 infection (IC50) or concentration causing 50% toxicity (TD50) towards ME180 cells.
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Figure 6: Inhibitory effect of polyanionic compounds against HSV-2 infection of epithelial cells. ME180 cells (seeded at 1.5 × 104 cells/well and cultured overnight), were exposed to HSV-2 (~5 × 104 Pfu/well) in the presence of compound for 1 hour, or alternatively, exposed to compound alone. Following compound/virus removal by washing, cells were cultured for a further 48 hours when viability was determined by MTT dye reduction. Cell viability (○, dotted line) following compound treatment was calculated as a percentage of the viability of cells exposed to culture medium alone. The effect of compound treatment on the infectivity of HSV-2 (●, solid line) was calculated as a percentage of infection observed in cells exposed to virus alone. Compounds tested were: A) PRO 2000; and B) Dextrin sulphate. Data represent the mean ± SEM of 3 independent experiments where each condition was tested in triplicate. Inserted figures represent the mean ± SEM concentration inhibiting 50% HSV-2 infection (IC50) or concentration causing 50% toxicity (TD50) towards ME180 cells.

Mentions: Due to the strong correlation reported between the presence of genital herpes and HIV-1 transmission [24], the effect of both PRO 2000 and DxS on the ability of HSV-2 to infect vaginal epithelial cells was investigated using the ME180 cell line. ME180 cells were exposed to HSV-2 (1 hour) in the presence of test compound and, following compound removal, cells were cultured for 48 hours in the absence of compound and virus, and viability determined by the principle of MTT dye reduction (see Methods). PRO 2000 and DxS demonstrated no significant toxicity towards ME180 cells, and both compounds demonstrated potent anti-HSV-2 activity, with IC50 values of 11.5 (± 1.4) μg/ml (PRO 2000) and 5.2 (± 1.4) μg/ml (DxS) (Figure 6).


Candidate polyanion microbicides inhibit HIV-1 infection and dissemination pathways in human cervical explants.

Fletcher PS, Wallace GS, Mesquita PM, Shattock RJ - Retrovirology (2006)

Inhibitory effect of polyanionic compounds against HSV-2 infection of epithelial cells. ME180 cells (seeded at 1.5 × 104 cells/well and cultured overnight), were exposed to HSV-2 (~5 × 104 Pfu/well) in the presence of compound for 1 hour, or alternatively, exposed to compound alone. Following compound/virus removal by washing, cells were cultured for a further 48 hours when viability was determined by MTT dye reduction. Cell viability (○, dotted line) following compound treatment was calculated as a percentage of the viability of cells exposed to culture medium alone. The effect of compound treatment on the infectivity of HSV-2 (●, solid line) was calculated as a percentage of infection observed in cells exposed to virus alone. Compounds tested were: A) PRO 2000; and B) Dextrin sulphate. Data represent the mean ± SEM of 3 independent experiments where each condition was tested in triplicate. Inserted figures represent the mean ± SEM concentration inhibiting 50% HSV-2 infection (IC50) or concentration causing 50% toxicity (TD50) towards ME180 cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 6: Inhibitory effect of polyanionic compounds against HSV-2 infection of epithelial cells. ME180 cells (seeded at 1.5 × 104 cells/well and cultured overnight), were exposed to HSV-2 (~5 × 104 Pfu/well) in the presence of compound for 1 hour, or alternatively, exposed to compound alone. Following compound/virus removal by washing, cells were cultured for a further 48 hours when viability was determined by MTT dye reduction. Cell viability (○, dotted line) following compound treatment was calculated as a percentage of the viability of cells exposed to culture medium alone. The effect of compound treatment on the infectivity of HSV-2 (●, solid line) was calculated as a percentage of infection observed in cells exposed to virus alone. Compounds tested were: A) PRO 2000; and B) Dextrin sulphate. Data represent the mean ± SEM of 3 independent experiments where each condition was tested in triplicate. Inserted figures represent the mean ± SEM concentration inhibiting 50% HSV-2 infection (IC50) or concentration causing 50% toxicity (TD50) towards ME180 cells.
Mentions: Due to the strong correlation reported between the presence of genital herpes and HIV-1 transmission [24], the effect of both PRO 2000 and DxS on the ability of HSV-2 to infect vaginal epithelial cells was investigated using the ME180 cell line. ME180 cells were exposed to HSV-2 (1 hour) in the presence of test compound and, following compound removal, cells were cultured for 48 hours in the absence of compound and virus, and viability determined by the principle of MTT dye reduction (see Methods). PRO 2000 and DxS demonstrated no significant toxicity towards ME180 cells, and both compounds demonstrated potent anti-HSV-2 activity, with IC50 values of 11.5 (± 1.4) μg/ml (PRO 2000) and 5.2 (± 1.4) μg/ml (DxS) (Figure 6).

Bottom Line: While both compounds were virucidal for X4 HIV-1, neither was virucidal for R5 virus.Our results demonstrate that PRO 2000 is a potent inhibitor of R5 HIV-1 infection and dissemination pathways in human cervical explants.DxS, while demonstrating significant inhibition of R5 infection, was less active against DC mediated dissemination pathways.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Infection, Department of Cellular and Molecular Medicine, St George's, University of London, London, UK. pfletche@sgul.ac.uk

ABSTRACT

Background: Heterosexual intercourse remains the major route of HIV-1 transmission worldwide, with almost 5 million new infections occurring each year. Women increasingly bear a disproportionate burden of the pandemic, thus there is an urgent need to develop new strategies to reduce HIV-1 transmission that could be controlled by women themselves. The potential of topical microbicides to reduce HIV transmission across mucosal surfaces has been clearly identified, and some agents are currently under evaluation in clinical trials. Many of these "first generation" microbicides consist of polyanionic compounds designed to interfere with viral attachment. Here we have evaluated two candidate polyanion compounds in clinical trials, PRO 2000 and dextrin sulphate (DxS) to determine their safety and efficacy against in vitro HIV-1 and HSV-2 infection using cellular and tissue explant models.

Results: PRO 2000 and DxS potently inhibited infection by HIV-1 X4 and R5 isolates when present during viral exposure. However PRO 2000 required 10-fold and DxS 2000-fold more compound to block infection with R5 virus than X4. While both compounds were virucidal for X4 HIV-1, neither was virucidal for R5 virus. PRO 2000 efficiently inhibited infection of cervical explants and dissemination of virus by migratory DC. DxS was less active, able to completely inhibit cervical explant infection, but providing only partial reduction of virus dissemination by DC. PRO 2000, but not DxS, also inhibited HIV-1 binding to DC-SIGN+ cells and trans infection of co-cultured target cells. The inflammatory potential of both compounds was screened by measurement of cytokine production from cervical explants, and statistically significant increases were only observed for IL-1beta and RANTES following treatment with PRO 2000. Both compounds also demonstrated potent activity against HSV-2 infection of cervical epithelial cells.

Conclusion: Our results demonstrate that PRO 2000 is a potent inhibitor of R5 HIV-1 infection and dissemination pathways in human cervical explants. DxS, while demonstrating significant inhibition of R5 infection, was less active against DC mediated dissemination pathways. PRO 2000 has now entered human phase III efficacy trials.

Show MeSH
Related in: MedlinePlus