Small-molecule screening using a human primary cell model of HIV latency identifies compounds that reverse latency without cellular activation
Hung-Chih Yang, … , Joseph B. Margolick, Robert F. Siliciano
Hung-Chih Yang, … , Joseph B. Margolick, Robert F. Siliciano
Published October 1, 2009
Citation Information: J Clin Invest. 2009;119(11):3473-3486. https://doi.org/10.1172/JCI39199.
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Technical Advance AIDS/HIV

Small-molecule screening using a human primary cell model of HIV latency identifies compounds that reverse latency without cellular activation

Abstract

The development of highly active antiretroviral therapy (HAART) to treat individuals infected with HIV-1 has dramatically improved patient outcomes, but HAART still fails to cure the infection. The latent viral reservoir in resting CD4+ T cells is a major barrier to virus eradication. Elimination of this reservoir requires reactivation of the latent virus. However, strategies for reactivating HIV-1 through nonspecific T cell activation have clinically unacceptable toxicities. We describe here the development of what we believe to be a novel in vitro model of HIV-1 latency that we used to search for compounds that can reverse latency. Human primary CD4+ T cells were transduced with the prosurvival molecule Bcl-2, and the resulting cells were shown to recapitulate the quiescent state of resting CD4+ T cells in vivo. Using this model system, we screened small-molecule libraries and identified a compound that reactivated latent HIV-1 without inducing global T cell activation, 5-hydroxynaphthalene-1,4-dione (5HN). Unlike previously described latency-reversing agents, 5HN activated latent HIV-1 through ROS and NF-κB without affecting nuclear factor of activated T cells (NFAT) and PKC, demonstrating that TCR pathways can be dissected and utilized to purge latent virus. Our study expands the number of classes of latency-reversing therapeutics and demonstrates the utility of this in vitro model for finding strategies to eradicate HIV-1 infection.

Authors

Hung-Chih Yang, Sifei Xing, Liang Shan, Karen O’Connell, Jason Dinoso, Anding Shen, Yan Zhou, Cynthia K. Shrum, Yefei Han, Jun O. Liu, Hao Zhang, Joseph B. Margolick, Robert F. Siliciano

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Figure 1

Generation of the Bcl-2–transduced primary CD4+ T cells.

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Generation of the Bcl-2–transduced primary CD4+ T cells. (A) The str...
(A) The structure of the lentiviral vector carrying the human Bcl-2 gene (EB-FLV). The U3 region of 3′ LTR is deleted (3′ΔLTR) for self inactivation. The expression of human Bcl-2 is driven by EF1α promoter. Ψ, packaging signal; RRE, rev responsive element; cPPT, central polypurine tract; pEF1α, EF1α promoter. (B) Strategy to generate Bcl-2–transduced primary CD4+ T cells. Primary CD4+ T cells from normal donors were activated and transduced with the Bcl-2–expressing lentiviral vector. Viable cells were isolated after 3 to 4 weeks of culture in the absence of TCR stimulants or cytokines. (C) Intracellular staining for Bcl-2 with FITC-conjugated anti–Bcl-2 antibody in freshly isolated CD4+ T cells and Bcl-2–transduced cells. The Bcl-2–transduced cells were maintained in culture without cytokines for 4 weeks following transduction. Freshly isolated CD4+ T cells stained with FITC-conjugated isotype control antibodies served as a negative control (purple).