Inhibition of HIV transmission in human cervicovaginal explants and humanized mice using CD4 aptamer-siRNA chimeras
Lee Adam Wheeler, Radiana Trifonova, Vladimir Vrbanac, Emre Basar, Shannon McKernan, Zhan Xu, Edward Seung, Maud Deruaz, Tim Dudek, Jon Ivar Einarsson, Linda Yang, Todd M. Allen, Andrew D. Luster, Andrew M. Tager, Derek M. Dykxhoorn, Judy Lieberman
Lee Adam Wheeler, Radiana Trifonova, Vladimir Vrbanac, Emre Basar, Shannon McKernan, Zhan Xu, Edward Seung, Maud Deruaz, Tim Dudek, Jon Ivar Einarsson, Linda Yang, Todd M. Allen, Andrew D. Luster, Andrew M. Tager, Derek M. Dykxhoorn, Judy Lieberman
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Research Article AIDS/HIV

Inhibition of HIV transmission in human cervicovaginal explants and humanized mice using CD4 aptamer-siRNA chimeras

Abstract

The continued spread of the HIV epidemic underscores the need to interrupt transmission. One attractive strategy is a topical vaginal microbicide. Sexual transmission of herpes simplex virus type 2 (HSV-2) in mice can be inhibited by intravaginal siRNA application. To overcome the challenges of knocking down gene expression in immune cells susceptible to HIV infection, we used chimeric RNAs composed of an aptamer fused to an siRNA for targeted gene knockdown in cells bearing an aptamer-binding receptor. Here, we showed that CD4 aptamer-siRNA chimeras (CD4-AsiCs) specifically suppress gene expression in CD4+ T cells and macrophages in vitro, in polarized cervicovaginal tissue explants, and in the female genital tract of humanized mice. CD4-AsiCs do not activate lymphocytes or stimulate innate immunity. CD4-AsiCs that knock down HIV genes and/or CCR5 inhibited HIV infection in vitro and in tissue explants. When applied intravaginally to humanized mice, CD4-AsiCs protected against HIV vaginal transmission. Thus, CD4-AsiCs could be used as the active ingredient of a microbicide to prevent HIV sexual transmission.

Authors

Lee Adam Wheeler, Radiana Trifonova, Vladimir Vrbanac, Emre Basar, Shannon McKernan, Zhan Xu, Edward Seung, Maud Deruaz, Tim Dudek, Jon Ivar Einarsson, Linda Yang, Todd M. Allen, Andrew D. Luster, Andrew M. Tager, Derek M. Dykxhoorn, Judy Lieberman

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Figure 4

CD4-AsiCs applied IVAG inhibit vaginal HIV transmission to humanized BLT mice.

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CD4-AsiCs applied IVAG inhibit vaginal HIV transmission to humanized BLT...
(A–C) NOD/SCID-BLT mice (2 per group) were treated twice IVAG with the indicated 5–80 pmol mixture of CD4 chimeras bearing Cy3-labeled CCR5 siRNA and CD45 siRNA. 2 days later, a single-cell suspension extracted from vaginal tissue was analyzed by flow cytometry for Cy3-labeled siRNA uptake and target gene knockdown. (B) Representative histograms (blue, mock; red, 80-pmol dose). (C) MFI of CD4+ T cells in the vaginal tissue of AsiC-treated mice showed a dose-dependent decrease of both CD45 and CCR5 expression relative to mock-treated controls, but the MFI of CD8+ T cells did not change (mean ± SEM; *P < 0.01, **P < 0.001, 1-way ANOVA with Dunnett multiple-comparison test). (D) HIV challenge experiment. NSG-BLT mice were treated IVAG with CCR5 CD4-AsiCs 48 hours (80 pmol) and 24 hours (40 pmol) before, and 40 pmol each of gag and vif chimeras 24 hours before and 4 hours after, IVAG challenge with HIVJR-CSF. CD4-AsiC treatment was compared with treatment with the same concentration of CD4 aptamer or mock treatment with PBS (n = 4 per group). Mice were observed for 12 weeks after HIV challenge. Shown is the effect of treatment on (E) survival; (F) plasma HIV Ag, assessed by ELISA; (G) plasma HIV RNA, analyzed by qRT-PCR (dashed line denotes limit of detection); and (H) circulating CD4+/CD8+ T cell ratio. Data are mean ± SEM. **P < 0.001, ***P < 0.0001 vs. mock, 2-way ANOVA with Bonferroni correction.