PMID- 26041287 OWN - NLM STAT- MEDLINE DCOM- 20160122 LR - 20210102 IS - 1098-5514 (Electronic) IS - 0022-538X (Print) IS - 0022-538X (Linking) VI - 89 IP - 16 DP - 2015 Aug TI - H3K27 Demethylation at the Proviral Promoter Sensitizes Latent HIV to the Effects of Vorinostat in Ex Vivo Cultures of Resting CD4+ T Cells. PG - 8392-405 LID - 10.1128/JVI.00572-15 [doi] AB - Histone methyltransferase inhibitors (HMTis) and histone deacetylase inhibitors (HDACis) are reported to synergistically induce the expression of latent human immunodeficiency virus type 1 (HIV-1), but studies have largely been performed with cell lines. As specific and potent HMTis directed at EZH1 (enhancer of zeste 2 Polycomb repressive complex 2 subunit 1)/EZH2 are now in human testing, we wished to rigorously test such an inhibitor in a primary resting T-cell model of HIV latency. We found that GSK343, a potent and selective EZH2/EZH1 inhibitor, reduced trimethylation of histone 3 at lysine 27 (H3K27) of the HIV provirus in resting cells. Remarkably, this epigenetic change was not associated with increased proviral expression in latently infected resting cells. However, following the reduction in H3K27 at the HIV long terminal repeat (LTR), subsequent exposure to the HDACi suberoylanilide hydroxamic acid or vorinostat (VOR) resulted in increases in HIV gag RNA and HIV p24 antigen production that were up to 2.5-fold greater than those induced by VOR alone. Therefore, in primary resting CD4(+) T cells, true mechanistic synergy in the reversal of HIV latency may be achieved by the combination of HMTis and HDACis. Although other cellular effects of EZH2 inhibition may contribute to the sensitization of the HIV LTR to subsequent exposure to VOR, and to increase viral antigen production, this synergistic effect is directly associated with H3K27 demethylation at nucleosome 1 (Nuc-1). Based upon our findings, the combination of HMTis and HDACis should be considered for testing in animal models or clinical trials. IMPORTANCE: Demethylation of H3K27 mediated by the histone methyltransferase inhibitor GSK343 in primary resting T cells is slow, occurring over 96 h, but by itself does not result in a significant upregulation of cell-associated HIV RNA expression or viral antigen production. However, following H3K27 demethylation, latent viral expression within infected primary resting CD4(+) T cells is synergistically increased upon exposure to the histone deacetylase inhibitor vorinostat. Demethylation at H3K27 sensitizes the HIV promoter to the effects of an HDACi and provides a proof-of-concept for the testing of combination epigenetic approaches to disrupt latent HIV infection, a necessary step toward the eradication of HIV infection. CI - Copyright (c) 2015, American Society for Microbiology. All Rights Reserved. FAU - Tripathy, Manoj K AU - Tripathy MK AD - Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA. FAU - McManamy, Mary E M AU - McManamy ME AD - Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA. FAU - Burch, Brandon D AU - Burch BD AD - Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA. FAU - Archin, Nancie M AU - Archin NM AD - Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA. FAU - Margolis, David M AU - Margolis DM AUID- ORCID: 0000-0001-5714-0002 AD - Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA Department of Epidemiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA dmargo@med.unc.edu. LA - eng GR - RR024383/RR/NCRR NIH HHS/United States GR - P30 CA016086/CA/NCI NIH HHS/United States GR - P30CA016086/CA/NCI NIH HHS/United States GR - UL1 RR025747/RR/NCRR NIH HHS/United States GR - U19 AI096113/AI/NIAID NIH HHS/United States GR - AI50410/AI/NIAID NIH HHS/United States GR - P30 AI050410/AI/NIAID NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, Non-U.S. Gov't DEP - 20150603 PL - United States TA - J Virol JT - Journal of virology JID - 0113724 RN - 0 (GSK343) RN - 0 (Histones) RN - 0 (Hydroxamic Acids) RN - 0 (Indazoles) RN - 0 (Pyridones) RN - 0 (RNA, Small Interfering) RN - 58IFB293JI (Vorinostat) RN - EC 2.1.1.43 (EZH1 protein, human) RN - EC 2.1.1.43 (EZH2 protein, human) RN - EC 2.1.1.43 (Enhancer of Zeste Homolog 2 Protein) RN - EC 2.1.1.43 (Polycomb Repressive Complex 2) SB - IM MH - Analysis of Variance MH - CD4-Positive T-Lymphocytes/*virology MH - Chromatin Immunoprecipitation MH - Enhancer of Zeste Homolog 2 Protein MH - Enzyme-Linked Immunosorbent Assay MH - HIV/*drug effects/physiology MH - Histones/*metabolism MH - Humans MH - Hydroxamic Acids/*pharmacology MH - Immunoblotting MH - Indazoles/*pharmacology MH - Methylation/drug effects MH - Polycomb Repressive Complex 2/antagonists & inhibitors MH - Promoter Regions, Genetic/genetics MH - Proviruses/genetics MH - Pyridones/*pharmacology MH - RNA, Small Interfering/genetics MH - Vorinostat PMC - PMC4524215 EDAT- 2015/06/05 06:00 MHDA- 2016/01/23 06:00 PMCR- 2016/02/15 CRDT- 2015/06/05 06:00 PHST- 2015/03/03 00:00 [received] PHST- 2015/05/26 00:00 [accepted] PHST- 2015/06/05 06:00 [entrez] PHST- 2015/06/05 06:00 [pubmed] PHST- 2016/01/23 06:00 [medline] PHST- 2016/02/15 00:00 [pmc-release] AID - JVI.00572-15 [pii] AID - 00572-15 [pii] AID - 10.1128/JVI.00572-15 [doi] PST - ppublish SO - J Virol. 2015 Aug;89(16):8392-405. doi: 10.1128/JVI.00572-15. Epub 2015 Jun 3.