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Inhibitors of Histone Deacetylases

CORRELATION BETWEEN ISOFORM SPECIFICITY AND REACTIVATION OF HIV TYPE 1 (HIV-1) FROM LATENTLY INFECTED CELLS
Open AccessPublished:April 29, 2011DOI:https://doi.org/10.1074/jbc.M110.180224
      Deacetylation of histone proteins at the HIV type 1 (HIV-1) long terminal repeat (LTR) by histone deactylases (HDACs) can promote transcriptional repression and virus latency. As such, HDAC inhibitors (HDACI) could be used to deplete reservoirs of persistent, quiescent HIV-1 proviral infection. However, the development of HDACI to purge latent HIV-1 requires knowledge of the HDAC isoforms contributing to viral latency and the development of inhibitors specific to these isoforms. In this study, we identify the HDACs responsible for HIV-1 latency in Jurkat J89GFP cells using a chemical approach that correlates HDACI isoform specificity with their ability to reactivate latent HIV-1 expression. We demonstrate that potent inhibition or knockdown of HDAC1, an HDAC isoform reported to drive HIV-1 into latency, was not sufficient to de-repress the viral LTR. Instead, we found that inhibition of HDAC3 was necessary to activate latent HIV-1. Consistent with this finding, we identified HDAC3 at the HIV-1 LTR by chromatin immunoprecipitation. Interestingly, we show that valproic acid is a weak inhibitor of HDAC3 (IC50 = 5.5 mm) relative to HDAC1 (IC50 = 170 μm). Because the total therapeutic concentration of valproic acid ranges from 275 to 700 μm in adults, these data may explain why this inhibitor has no effect on the decay of latent HIV reservoirs in patients. Taken together, our study suggests an important role for HDAC3 in HIV-1 latency and, importantly, describes a chemical approach that can readily be used to identify the HDAC isoforms that contribute to HIV-1 latency in other cell types.

      Introduction

      Combination antiretroviral therapy (cART)
      The abbreviations used are: cART
      combination antiretroviral therapy
      HDAC
      histone deacetylase
      HDACI
      HDAC inhibitor
      SAHA
      suberoylanilide hydroxamic acid
      EGFP
      enhanced green fluorescent protein
      FW
      forward
      REV
      reverse
      HIV-1
      HIV type 1.
      can effectively reduce plasma HIV-1 to undetectable levels. However, upon its interruption, there is usually a rapid rebound of viremia (
      • Davey Jr., R.T.
      • Bhat N.
      • Yoder C.
      • Chun T.W.
      • Metcalf J.A.
      • Dewar R.
      • Natarajan V.
      • Lempicki R.A.
      • Adelsberger J.W.
      • Miller K.D.
      • Kovacs J.A.
      • Polis M.A.
      • Walker R.E.
      • Falloon J.
      • Masur H.
      • Gee D.
      • Baseler M.
      • Dimitrov D.S.
      • Fauci A.S.
      • Lane H.C.
      ). This viremia is thought to arise from latently infected reservoirs such as memory CD4(+) T cells or CD34(+) multipotent hematopoietic progenitor cells (
      • Chun T.W.
      • Stuyver L.
      • Mizell S.B.
      • Ehler L.A.
      • Mican J.A.
      • Baseler M.
      • Lloyd A.L.
      • Nowak M.A.
      • Fauci A.S.
      ,
      • Finzi D.
      • Hermankova M.
      • Pierson T.
      • Carruth L.M.
      • Buck C.
      • Chaisson R.E.
      • Quinn T.C.
      • Chadwick K.
      • Margolick J.
      • Brookmeyer R.
      • Gallant J.
      • Markowitz M.
      • Ho D.D.
      • Richman D.D.
      • Siliciano R.F.
      ,
      • Wong J.K.
      • Hezareh M.
      • Günthard H.F.
      • Havlir D.V.
      • Ignacio C.C.
      • Spina C.A.
      • Richman D.D.
      ,
      • Carter C.C.
      • Onafuwa-Nuga A.
      • McNamara L.A.
      • Riddell 4th, J.
      • Bixby D.
      • Savona M.R.
      • Collins K.L.
      ). Therefore, any long term therapeutic strategy targeted toward eliminating HIV-1 infection must include compounds that purge the latent viral reservoirs thereby rendering them susceptible to cART.
      HIV-1 can be maintained in a latent state by multiple different mechanisms that inhibit virus gene expression after integration into the cellular DNA (
      • Colin L.
      • Van Lint C.
      ,
      • Margolis D.M.
      ,
      • Trono D.
      • Van Lint C.
      • Rouzioux C.
      • Verdin E.
      • Barré-Sinoussi F.
      • Chun T.W.
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      ). For example, epigenetic modifications at or near the HIV-1 5′-long terminal repeat (LTR) can induce chromatin condensation that diminishes the accessibility of the HIV-1 promoter to transcription factors. In this regard, it has been well documented that different transcription factors can recruit histone deacetylase (HDAC) enzymes to the HIV-1 LTR where they promote chromatin condensation by deacetylating the ∈-amino groups of lysine residues in histone tails (
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      ,
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      ,
      • Imai K.
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      ,
      • Marban C.
      • Suzanne S.
      • Dequiedt F.
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      • Redel L.
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      • Rohr O.
      ,
      • Jiang G.
      • Espeseth A.
      • Hazuda D.J.
      • Margolis D.M.
      ,
      • Tyagi M.
      • Karn J.
      ). Eleven distinct zinc-dependent HDAC isoforms have been identified in humans. These can be classified into four families, namely class I (HDAC1–3 and -8), IIa (HDAC4, -5, -7, and -9), IIb (HDAC6 and -10), and IV (HDAC11), which differ in structure, enzymatic function, subcellular localization, and expression patterns (
      • Haberland M.
      • Montgomery R.L.
      • Olson E.N.
      ). To date, multiple studies have demonstrated that recruitment of HDAC1 to the HIV-1 LTR by different DNA-binding complexes is sufficient to induce viral latency (
      • Coull J.J.
      • Romerio F.
      • Sun J.M.
      • Volker J.L.
      • Galvin K.M.
      • Davie J.R.
      • Shi Y.
      • Hansen U.
      • Margolis D.M.
      ,
      • Williams S.A.
      • Chen L.F.
      • Kwon H.
      • Ruiz-Jarabo C.M.
      • Verdin E.
      • Greene W.C.
      ,
      • Imai K.
      • Okamoto T.
      ,
      • Marban C.
      • Suzanne S.
      • Dequiedt F.
      • de Walque S.
      • Redel L.
      • Van Lint C.
      • Aunis D.
      • Rohr O.
      ,
      • Jiang G.
      • Espeseth A.
      • Hazuda D.J.
      • Margolis D.M.
      ,
      • Tyagi M.
      • Karn J.
      ). However, HDAC2 and HDAC3 can also bind to the HIV-1 LTR and may also play an important role in viral latency (
      • Marban C.
      • Suzanne S.
      • Dequiedt F.
      • de Walque S.
      • Redel L.
      • Van Lint C.
      • Aunis D.
      • Rohr O.
      ,
      • Malcolm T.
      • Chen J.
      • Chang C.
      • Sadowski I.
      ,
      • Keedy K.S.
      • Archin N.M.
      • Gates A.T.
      • Espeseth A.
      • Hazuda D.J.
      • Margolis D.M.
      ).
      Treatment of latently infected HIV-1 cell lines and/or CD4(+) T cells from aviremic HIV-1-infected individuals on cART with HDACI can lead to chromatin relaxation and induction of viral transcription (reviewed in Ref.
      • Colin L.
      • Van Lint C.
      ). Therefore, HDACIs are considered as potential therapeutic agents for purging the latent viral reservoir in HIV-1-infected individuals. However, the active site structures of the HDAC family are largely conserved, and many HDACIs exhibit activity against multiple HDAC isoforms. For example, suberoylanilide hydroxamic acid (SAHA, vorinostat), an activator of latent HIV-1 expression (
      • Edelstein L.C.
      • Micheva-Viteva S.
      • Phelan B.D.
      • Dougherty J.P.
      ,
      • Archin N.M.
      • Espeseth A.
      • Parker D.
      • Cheema M.
      • Hazuda D.
      • Margolis D.M.
      ,
      • Contreras X.
      • Schweneker M.
      • Chen C.S.
      • McCune J.M.
      • Deeks S.G.
      • Martin J.
      • Peterlin B.M.
      ), is a nonselective HDACI that inhibits both class I and class II HDAC isoforms (
      • Savarino A.
      • Mai A.
      • Norelli S.
      • El Daker S.
      • Valente S.
      • Rotili D.
      • Altucci L.
      • Palamara A.T.
      • Garaci E.
      ). Because HDACs exert crucial roles in numerous biological processes, including cell cycle, cell differentiation, and survival (
      • Haberland M.
      • Montgomery R.L.
      • Olson E.N.
      ), simultaneous inhibition of multiple HDAC isoforms will likely reduce their therapeutic window by promoting undesirable side effects and/or toxicity. Accordingly, the development of HDACI for an HIV-1 curative strategy requires knowledge of the HDAC isoforms contributing to viral latency and the development of inhibitors targeting these isoforms. In this study, we use a chemical approach that correlates the isoform specificities of HDACI with their abilities to reactivate latent HIV-1 expression to identify the HDAC isoforms responsible for HIV-1 latency in Jurkat J89GFP cells. The results from this study suggest that potent inhibition of HDAC3 may be important for reactivation of latent HIV-1.

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