Expression of latent HIV induced by the potent HDAC inhibitor suberoylanilide hydroxamic acid

Abstract

Histone deacetylases (HDACs) act on histones within the nucleosome-bound promoter of human immunodeficiency virus type 1 (HIV-1) to maintain proviral latency. HDAC inhibition leads to promoter expression and the escape of HIV from latency. We evaluated the ability of the potent inhibitor recently licensed for use in oncology, suberoylanilide hydroxamic acid (SAHA; Vorinostat), selective for Class I HDACs, to induce HIV promoter expression in cell lines and virus production from the resting CD4(+) T cells of antiretroviral-treated, aviremic HIV-infected patients. In J89, a Jurkat T cell line infected with a single HIV genome encoding the enhanced green fluorescence protein (EGFP) within the HIV genome, SAHA induced changes at nucleosome 1 of the HIV promoter in chromatin immunoprecipitation (ChIP) assays in concert with EGFP expression. In the resting CD4(+) T cells of antiretroviral-treated, aviremic HIV-infected patients clinically achievable exposures to SAHA induced virus outgrowth ex vivo. These results suggest that potent, selective HDAC inhibitors may allow improved targeting of persistent proviral HIV infection, and define parameters for in vivo studies using SAHA.

Figures

FIG. 1.

Like VPA, SAHA increases acetylation of Nuc 1 and decreases HDAC-1 occupancy at the HIV-1 LTR. J89 cell treated with media, 3 mM VPA, or 500 nM SAHA for 4 h were assayed by chromatin immunoprecipitation with anti-acetylated H3, anti-HDAC-1. To demonstrate that the apparent quantitative changes are significant in this semi-quantitative assay, DNA products of ChIP were quantitated in triplicate by real-time PCR. Assays are representative of 3 independent experiments, and real-time quantitation of the fold change relative to untreated control is shown.

FIG. 2.

SAHA activates HIV LTR transcription. J89 cells were treated with the indicated concentrations of SAHA or VPA for 4hrs. Semi-quantitative RT PCR was performed on total RNA isolated from treated and untreated cells as described in methods. The data presented is the mean ± SE of 3 independent experiments.

FIG. 3.

Toxicity of SAHA in seronegative donor PBMC and J89 cells. PBMC or J89 cells were cultured in the absence or presence of SAHA or VPA at the indicated concentrations. MTT assays were performed as described in methods. The percentage of proliferating cells was calculated compared to cells cultured in standard media. The data represent the mean ± SE of 3 independent experiments.