Advertisement

Histone Deacetylase Inhibitors Modulate Interleukin 6-dependent CD4+ T Cell Polarization in Vitro and in Vivo*

Open AccessPublished:January 13, 2014DOI:https://doi.org/10.1074/jbc.M113.517599
      Histone deacetylase (HDAC) inhibitors have been associated primarily with an anti-proliferative effect in vitro and in vivo. Recent data provide evidence for an anti-inflammatory potency of HDAC inhibitors in models of experimental colitis. Because the balance of T cell subpopulations is critical for the balance of the mucosal immune system, this study explores the regulatory potency of HDAC inhibitors on T cell polarization as a mechanistic explanation for the observed anti-inflammatory effects. Although HDAC inhibition suppressed the polarization toward the pro-inflammatory T helper 17 (Th17) cells, it enhanced forkhead box P3 (FoxP3)+ regulatory T cell polarization in vitro and in vivo at the site of inflammation in the lamina propria. This was paralleled by a down-regulation of the interleukin 6 receptor (IL-6R) on naïve CD4+ T cells on the mRNA as well as on the protein level and changes in the chromatin acetylation at the IL6R gene and its promoter. Downstream of the IL-6R, HDAC inhibition was followed by a decrease in STAT3 phosphorylation as well as retinoic acid receptor-related orphan receptor γT (RORγT) expression, thus identifying the IL-6/STAT3/IL-17 pathway as an important target of HDAC inhibitors. These results directly translated to experimental colitis, where IL-6R expression was suppressed in naïve T cells, paralleled by a significant reduction of Th17 cells in the lamina propria of ITF2357-treated animals, resulting in the amelioration of disease. This study indicates that, in experimental colitis, inhibition of HDAC exerts an anti-inflammatory potency by directing T helper cell polarization via targeting the IL-6 pathway.

      Introduction

      Epigenetic modifications represent an additional regulatory level for gene expression and, thus, comprise potential therapeutic targets for autoinflammatory diseases or cancer. This work focuses on the modification of histone acetylation that is under a physiological state regulated by the balance of histone acetyltransferases and histone deacetylases (HDACs).
      The abbreviations used are:
      HDAC
      histone deacetylase
      SAHA
      suberoylanilide hydroxamid acid
      Th17 cell
      T helper 17 cell
      Treg cell
      regulatory T cell
      IL-6R
      IL-6 receptor
      DSS
      dextran sulfate sodium
      MLN
      mesenteric lymph node
      LPMC
      lamina propria mononuclear cell
      RORγT
      retinoic acid receptor-related orphan receptor γT.
      HDAC inhibitors have been primarily described for their antiproliferative potency, as shown in animal models for colon, mammary, prostate, and bladder cancer (
      • Kelly W.K.
      • Marks P.A.
      Drug insight. Histone deacetylase inhibitors. Development of the new targeted anticancer agent suberoylanilide hydroxamic acid.
      ,
      • Marks P.
      • Rifkind R.A.
      • Richon V.M.
      • Breslow R.
      • Miller T.
      • Kelly W.K.
      Histone deacetylases and cancer. Causes and therapies.
      ). Subsequent clinical trials resulted in Food and Drug Administration approval of the HDAC inhibitor suberoylanilide hydroxamid acid (SAHA) for the treatment of cutaneous T cell lymphoma (
      • Marks P.A.
      • Breslow R.
      Dimethyl sulfoxide to vorinostat. Development of this histone deacetylase inhibitor as an anticancer drug.
      ). Remarkably, these antiproliferative effects have been partially attributed to the inhibition of the nuclear factor-κB pathway (
      • Krämer O.H.
      • Baus D.
      • Knauer S.K.
      • Stein S.
      • Jäger E.
      • Stauber R.H.
      • Grez M.
      • Pfitzner E.
      • Heinzel T.
      Acetylation of Stat1 modulates NF-κB activity.
      ,
      • Glauben R.
      • Batra A.
      • Stroh T.
      • Erben U.
      • Fedke I.
      • Lehr H.A.
      • Leoni F.
      • Mascagni P.
      • Dinarello C.A.
      • Zeitz M.
      • Siegmund B.
      Histone deacetylases. Novel targets for prevention of colitis-associated cancer in mice.
      ). At the same time, a strong anti-inflammatory capacity of this compound class was identified. Initial in vitro studies revealed a dose-dependent suppression of proinflammatory cytokine production by SAHA at concentrations 10- to 100-fold lower than required for apoptosis induction (
      • Glauben R.
      • Batra A.
      • Fedke I.
      • Zeitz M.
      • Lehr H.A.
      • Leoni F.
      • Mascagni P.
      • Fantuzzi G.
      • Dinarello C.A.
      • Siegmund B.
      Histone hyperacetylation is associated with amelioration of experimental colitis in mice.
      ,
      • Leoni F.
      • Zaliani A.
      • Bertolini G.
      • Porro G.
      • Pagani P.
      • Pozzi P.
      • Donà G.
      • Fossati G.
      • Sozzani S.
      • Azam T.
      • Bufler P.
      • Fantuzzi G.
      • Goncharov I.
      • Kim S.H.
      • Pomerantz B.J.
      • Reznikov L.L.
      • Siegmund B.
      • Dinarello C.A.
      • Mascagni P.
      The antitumor histone deacetylase inhibitor suberoylanilide hydroxamic acid exhibits antiinflammatory properties via suppression of cytokines.
      ). These results were complemented in vivo by investigating inflammatory models such as concanavalin A-induced hepatitis or LPS-induced shock in mice (
      • Leoni F.
      • Zaliani A.
      • Bertolini G.
      • Porro G.
      • Pagani P.
      • Pozzi P.
      • Donà G.
      • Fossati G.
      • Sozzani S.
      • Azam T.
      • Bufler P.
      • Fantuzzi G.
      • Goncharov I.
      • Kim S.H.
      • Pomerantz B.J.
      • Reznikov L.L.
      • Siegmund B.
      • Dinarello C.A.
      • Mascagni P.
      The antitumor histone deacetylase inhibitor suberoylanilide hydroxamic acid exhibits antiinflammatory properties via suppression of cytokines.
      ). Following these initial data, the anti-inflammatory potency could be confirmed in models of nephritis (
      • Mishra N.
      • Reilly C.M.
      • Brown D.R.
      • Ruiz P.
      • Gilkeson G.S.
      Histone deacetylase inhibitors modulate renal disease in the MRL-lpr/lpr mouse.
      ), arthritis (
      • Nishida K.
      • Komiyama T.
      • Miyazawa S.
      • Shen Z.N.
      • Furumatsu T.
      • Doi H.
      • Yoshida A.
      • Yamana J.
      • Yamamura M.
      • Ninomiya Y.
      • Inoue H.
      • Asahara H.
      Histone deacetylase inhibitor suppression of autoantibody-mediated arthritis in mice via regulation of p16INK4a and p21(WAF1/Cip1) expression.
      ), intestinal inflammation (
      • Glauben R.
      • Batra A.
      • Fedke I.
      • Zeitz M.
      • Lehr H.A.
      • Leoni F.
      • Mascagni P.
      • Fantuzzi G.
      • Dinarello C.A.
      • Siegmund B.
      Histone hyperacetylation is associated with amelioration of experimental colitis in mice.
      ,
      • Tao R.
      • de Zoeten E.F.
      • Ozkaynak E.
      • Chen C.
      • Wang L.
      • Porrett P.M.
      • Li B.
      • Turka L.A.
      • Olson E.N.
      • Greene M.I.
      • Wells A.D.
      • Hancock W.W.
      Deacetylase inhibition promotes the generation and function of regulatory T cells.
      ), graft versus host disease (
      • Reddy P.
      • Sun Y.
      • Toubai T.
      • Duran-Struuck R.
      • Clouthier S.G.
      • Weisiger E.
      • Maeda Y.
      • Tawara I.
      • Krijanovski O.
      • Gatza E.
      • Liu C.
      • Malter C.
      • Mascagni P.
      • Dinarello C.A.
      • Ferrara J.L.
      Histone deacetylase inhibition modulates indoleamine 2,3-dioxygenase-dependent DC functions and regulates experimental graft-versus-host disease in mice.
      ), and asthma (
      • Choi J.H.
      • Oh S.W.
      • Kang M.S.
      • Kwon H.J.
      • Oh G.T.
      • Kim D.Y.
      Trichostatin A attenuates airway inflammation in mouse asthma model.
      ). In addition, the HDAC inhibitor ITF2357 has shown clinical efficacy in clinical trials for juvenile arthritis (
      • Vojinovic J.
      • Damjanov N.
      • D'Urzo C.
      • Furlan A.
      • Susic G.
      • Pasic S.
      • Iagaru N.
      • Stefan M.
      • Dinarello C.A.
      Safety and efficacy of an oral histone deacetylase inhibitor in systemic-onset juvenile idiopathic arthritis.
      ).
      The maintenance of the mucosal homeostasis requires the balance of CD4+ T cell subsets within the compartment of the lamina propria. In particular, proinflammatory T helper 17 (Th17) cells and anti-inflammatory regulatory T (Treg) cells have been identified as critical factors that define the course and severity of intestinal inflammation (
      • Mucida D.
      • Park Y.
      • Kim G.
      • Turovskaya O.
      • Scott I.
      • Kronenberg M.
      • Cheroutre H.
      Reciprocal TH17 and regulatory T cell differentiation mediated by retinoic acid.
      ,
      • Fantini M.C.
      • Rizzo A.
      • Fina D.
      • Caruso R.
      • Becker C.
      • Neurath M.F.
      • Macdonald T.T.
      • Pallone F.
      • Monteleone G.
      IL-21 regulates experimental colitis by modulating the balance between Treg and Th17 cells.
      ). This concept is supported by studies indicating a beneficial effect by targeting the Th17-inducing cytokine IL-23 in experimental colitis (
      • Elson C.O.
      • Cong Y.
      • Weaver C.T.
      • Schoeb T.R.
      • McClanahan T.K.
      • Fick R.B.
      • Kastelein R.A.
      Monoclonal anti-interleukin 23 reverses active colitis in a T cell-mediated model in mice.
      ). This strategy has been proven successful in clinical trials for inflammatory bowel disease (
      • Sands B.E.
      • Jacobson E.W.
      • Sylwestrowicz T.
      • Younes Z.
      • Dryden G.
      • Fedorak R.
      • Greenbloom S.
      Randomized, double-blind, placebo-controlled trial of the oral interleukin-12/23 inhibitor apilimod mesylate for treatment of active Crohn's disease.
      ,
      • Sandborn W.J.
      • Gasink C.
      • Gao L.L.
      • Blank M.A.
      • Johanns J.
      • Guzzo C.
      • Sands B.E.
      • Hanauer S.B.
      • Targan S.
      • Rutgeerts P.
      • Ghosh S.
      • de Villiers W.J.
      • Panaccione R.
      • Greenberg G.
      • Schreiber S.
      • Lichtiger S.
      • Feagan B.G.
      Ustekinumab induction and maintenance therapy in refractory Crohn's disease.
      ). In addition, transfer of regulatory T cells resulted in the amelioration in various models of experimental colitis (
      • Izcue A.
      • Coombes J.L.
      • Powrie F.
      Regulatory T cells suppress systemic and mucosal immune activation to control intestinal inflammation.
      ). Thus, for inflammatory bowel disease as well as for other chronic inflammatory conditions, a therapeutic approach targeting both T cell subsets would be intriguing.
      For the in vitro generation of both Th17 and inducible Treg cells from naïve CD4+ cells, TGFβ has been identified as a mandatory cytokine (
      • Chen W.
      • Jin W.
      • Hardegen N.
      • Lei K.J.
      • Li L.
      • Marinos N.
      • McGrady G.
      • Wahl S.M.
      Conversion of peripheral CD4+CD25 naive T cells to CD4+CD25+ regulatory T cells by TGF-β induction of transcription factor Foxp3.
      ,
      • Volpe E.
      • Servant N.
      • Zollinger R.
      • Bogiatzi S.I.
      • Hupé P.
      • Barillot E.
      • Soumelis V.
      A critical function for transforming growth factor-β, interleukin 23 and proinflammatory cytokines in driving and modulating human T(H)-17 responses.
      ). Adding IL-6 to low doses of TGFβ results in the differentiation of Th17 cells in vitro (
      • Bettelli E.
      • Carrier Y.
      • Gao W.
      • Korn T.
      • Strom T.B.
      • Oukka M.
      • Weiner H.L.
      • Kuchroo V.K.
      Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells.
      ,
      • Mangan P.R.
      • Harrington L.E.
      • O'Quinn D.B.
      • Helms W.S.
      • Bullard D.C.
      • Elson C.O.
      • Hatton R.D.
      • Wahl S.M.
      • Schoeb T.R.
      • Weaver C.T.
      Transforming growth factor-β induces development of the T(H)17 lineage.
      ,
      • Veldhoen M.
      • Hocking R.J.
      • Atkins C.J.
      • Locksley R.M.
      • Stockinger B.
      TGFβ in the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells.
      ), thus identifying IL-6 as the key cytokine defining the polarization of these two cell types. IL-6 is a pleiotropic cytokine and was first described to function as a B cell differentiation factor being produced by several cell types, such as macrophages, monocytes, and T cells (
      • Hirano T.
      • Yasukawa K.
      • Harada H.
      • Taga T.
      • Watanabe Y.
      • Matsuda T.
      • Kashiwamura S.
      • Nakajima K.
      • Koyama K.
      • Iwamatsu A.
      Complementary DNA for a novel human interleukin (BSF-2) that induces B lymphocytes to produce immunoglobulin.
      ). In acute inflammation such as sepsis, IL-6 is critical for the induction of acute phase reactions (
      • Hideshima T.
      • Mitsiades C.
      • Tonon G.
      • Richardson P.G.
      • Anderson K.C.
      Understanding multiple myeloma pathogenesis in the bone marrow to identify new therapeutic targets.
      ).
      IL-6 deploys its effects through binding to the IL-6 receptor (IL-6R). The IL-6·IL-6R complex then signals through the ubiquitously expressed subunit gp130, leading to activation of the JAK/STAT pathway (classic signaling) (
      • Hibi M.
      • Murakami M.
      • Saito M.
      • Hirano T.
      • Taga T.
      • Kishimoto T.
      Molecular cloning and expression of an IL-6 signal transducer, gp130.
      ). Remarkably, the expression of the membrane-bound IL-6R is restricted to naïve T cells and hepatocytes, thus emphasizing the significance of this cytokine for these cell types. However, many other cell types can be activated by IL-6. To enable signaling in cells not expressing the IL-6R, the soluble IL-6R forms a complex with IL-6 that binds to gp130 and, thereby, activates cells that would be naturally unresponsive to IL-6 (
      • Rose-John S.
      • Heinrich P.C.
      Soluble receptors for cytokines and growth factors. Generation and biological function.
      ).
      The aim of this study was to define the effect of HDAC inhibition on CD4+ T cell polarization and to provide an explanation for the anti-inflammatory effects observed in vivo. In fact, we were able to link HDAC inhibition to decreased IL-6R expression, resulting in suppression of the associated signaling pathway and, ultimately, the suppression of an inflammatory T cell response.

      DISCUSSION

      Inhibition of HDAC results in the amelioration of colitis in a variety of models. However, the underlying mechanism remains largely to be defined. In these models, the severity, but even more the chronicity, of disease depends on the presence of CD4+ T cells and the balance of distinct subpopulations at the site of inflammation (
      • Powrie F.
      Immune regulation in the intestine. A balancing act between effector and regulatory T cell responses.
      ). This study serves to identify the regulatory capacity of HDAC inhibition on T cell polarization and, consecutively, on this critical balance of T cell subpopulations. A regulatory role of T cell function was indicated by the suppression of IL-6 and IFNγ synthesis in stimulated CD4+ T cells in the presence of an HDAC inhibitor. More important, HDAC inhibition enhanced Treg cell generation and suppressed polarization toward Th17 cells in vitro as well as in vivo at the site of inflammation. Because Th1 and Th2 polarization was unaffected, the IL-6 pathway came into focus as potential target because IL-6 represents the key cytokine distinguishing polarization toward Th17 cells from inducible Treg cells. The subsequent studies revealed a unique anti-inflammatory mode of action via regulation of the IL-6R and downstream signaling.
      Genetic as well as environmental factors contribute to the dysregulation of the mucosal immune system in Crohn disease and ulcerative colitis (
      • Siegmund B.
      • Zeitz M.
      Clinical aspects of inflammatory bowel disease.
      ). Although the balance of T helper cell subpopulations has been identified to play a critical role in inflammatory bowel disease, recent studies provide evidence that Th17 cells, in particular, add to the inflammatory process in the lamina propria (
      • Annunziato F.
      • Cosmi L.
      • Santarlasci V.
      • Maggi L.
      • Liotta F.
      • Mazzinghi B.
      • Parente E.
      • Filì L.
      • Ferri S.
      • Frosali F.
      • Giudici F.
      • Romagnani P.
      • Parronchi P.
      • Tonelli F.
      • Maggi E.
      • Romagnani S.
      Phenotypic and functional features of human Th17 cells.
      ,
      • Fujino S.
      • Andoh A.
      • Bamba S.
      • Ogawa A.
      • Hata K.
      • Araki Y.
      • Bamba T.
      • Fujiyama Y.
      Increased expression of interleukin 17 in inflammatory bowel disease.
      ). Several studies identified IL-6 to be essential for the development of Th17 cells (
      • Bettelli E.
      • Carrier Y.
      • Gao W.
      • Korn T.
      • Strom T.B.
      • Oukka M.
      • Weiner H.L.
      • Kuchroo V.K.
      Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells.
      ,
      • Mangan P.R.
      • Harrington L.E.
      • O'Quinn D.B.
      • Helms W.S.
      • Bullard D.C.
      • Elson C.O.
      • Hatton R.D.
      • Wahl S.M.
      • Schoeb T.R.
      • Weaver C.T.
      Transforming growth factor-β induces development of the T(H)17 lineage.
      ,
      • Veldhoen M.
      • Hocking R.J.
      • Atkins C.J.
      • Locksley R.M.
      • Stockinger B.
      TGFβ in the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells.
      ). Noguchi et al. (
      • Noguchi D.
      • Wakita D.
      • Ohkuri T.
      • Tajima M.
      • Chamoto K.
      • Kitamura H.
      • Nishimura T.
      Blockade of IL-6-signaling inhibits the pathogenesis of CD4+ T cell-mediated lethal graft-versus-host reaction against minor histocompatibility antigen.
      ) were able to demonstrate that blocking the IL-6 pathway through a specific IL-6R antibody resulted not only in the amelioration of experimental colitis but, furthermore, was paralleled by a significant decrease in Th17 cells in the lamina propria.
      Treatment with pan-HDAC inhibitors ameliorates experimental colitis in mice and inhibits the production of proinflammatory cytokines at the site of inflammation (
      • Glauben R.
      • Batra A.
      • Fedke I.
      • Zeitz M.
      • Lehr H.A.
      • Leoni F.
      • Mascagni P.
      • Fantuzzi G.
      • Dinarello C.A.
      • Siegmund B.
      Histone hyperacetylation is associated with amelioration of experimental colitis in mice.
      ). Via stimulation of LPMCs in the presence of HDAC inhibitors, the local effect of these compounds on effector cells could be proven. A similar dose-dependent suppression of IFNγ and IL-6 synthesis was observed when isolated CD4+ cells were stimulated in the presence of HDAC inhibitors. Both experiments suggest that the local T helper cell population within the lamina propria represents a critical target for HDAC inhibition. In the majority of experimental colitis models, Th1 as well as Th17 cells have been shown to represent a key cell population in the disease course (
      • Dohi T.
      • Fujihashi K.
      Type 1 and 2 T helper cell-mediated colitis.
      ). The absence of IL-17-producing T cells in the transfer colitis model almost completely abolished the characteristic inflammation (
      • Leppkes M.
      • Becker C.
      • Ivanov I.I.
      • Hirth S.
      • Wirtz S.
      • Neufert C.
      • Pouly S.
      • Murphy A.J.
      • Valenzuela D.M.
      • Yancopoulos G.D.
      • Becher B.
      • Littman D.R.
      • Neurath M.F.
      RORγ-expressing Th17 cells induce murine chronic intestinal inflammation via redundant effects of IL-17A and IL-17F.
      ). In contrast, the presence of Treg cells has been demonstrated to exert an ameliorating effect in experimental colitis (
      • Mottet C.
      • Uhlig H.H.
      • Powrie F.
      Cutting edge. Cure of colitis by CD4+CD25+ regulatory T cells.
      ,
      • Kjellev S.
      • Lundsgaard D.
      • Poulsen S.S.
      • Markholst H.
      Reconstitution of Scid mice with CD4+CD25 T cells leads to rapid colitis. An improved model for pharmacologic testing.
      ).
      There are several options regarding how HDAC inhibition might modulate the T cell response toward an anti-inflammatory direction. Studying the effects of HDAC inhibition on T helper cell polarization, we were able to demonstrate, in vitro, that the release of Th1 cytokines was suppressed significantly by treatment with HDAC inhibitors (Fig. 1 and Refs.
      • Glauben R.
      • Batra A.
      • Fedke I.
      • Zeitz M.
      • Lehr H.A.
      • Leoni F.
      • Mascagni P.
      • Fantuzzi G.
      • Dinarello C.A.
      • Siegmund B.
      Histone hyperacetylation is associated with amelioration of experimental colitis in mice.
      ,
      • Leoni F.
      • Zaliani A.
      • Bertolini G.
      • Porro G.
      • Pagani P.
      • Pozzi P.
      • Donà G.
      • Fossati G.
      • Sozzani S.
      • Azam T.
      • Bufler P.
      • Fantuzzi G.
      • Goncharov I.
      • Kim S.H.
      • Pomerantz B.J.
      • Reznikov L.L.
      • Siegmund B.
      • Dinarello C.A.
      • Mascagni P.
      The antitumor histone deacetylase inhibitor suberoylanilide hydroxamic acid exhibits antiinflammatory properties via suppression of cytokines.
      ,
      • Leoni F.
      • Fossati G.
      • Lewis E.C.
      • Lee J.K.
      • Porro G.
      • Pagani P.
      • Modena D.
      • Moras M.L.
      • Pozzi P.
      • Reznikov L.L.
      • Siegmund B.
      • Fantuzzi G.
      • Dinarello C.A.
      • Mascagni P.
      The histone deacetylase inhibitor ITF2357 reduces production of pro-inflammatory cytokines in vitro and systemic inflammation in vivo.
      ). Although the polarization toward either Th1 or Th2 cells was not affected by the presence of an HDAC inhibitor, the generation of Treg cells was enhanced significantly, and the polarization of Th17 cells was suppressed dose-dependently. In parallel to the in vitro findings, the analysis of T cell subsets in the lamina propria and MLN from mice with experimental colitis treated with ITF2357 showed a reduction in the frequency of Th17 cells within the CD4+ T cell fraction, and, vice versa, the Treg cell population was increased. Hence, the anti-inflammatory shift in the Treg/Th17 cell balance demonstrated in vitro could be confirmed directly in the animal model at the site of inflammation, delivering an explicit mode of action for these compounds regarding the anti-inflammatory effect exerted in a variety of inflammation models (
      • Glauben R.
      • Batra A.
      • Fedke I.
      • Zeitz M.
      • Lehr H.A.
      • Leoni F.
      • Mascagni P.
      • Fantuzzi G.
      • Dinarello C.A.
      • Siegmund B.
      Histone hyperacetylation is associated with amelioration of experimental colitis in mice.
      ,
      • Leoni F.
      • Zaliani A.
      • Bertolini G.
      • Porro G.
      • Pagani P.
      • Pozzi P.
      • Donà G.
      • Fossati G.
      • Sozzani S.
      • Azam T.
      • Bufler P.
      • Fantuzzi G.
      • Goncharov I.
      • Kim S.H.
      • Pomerantz B.J.
      • Reznikov L.L.
      • Siegmund B.
      • Dinarello C.A.
      • Mascagni P.
      The antitumor histone deacetylase inhibitor suberoylanilide hydroxamic acid exhibits antiinflammatory properties via suppression of cytokines.
      ,
      • Mishra N.
      • Reilly C.M.
      • Brown D.R.
      • Ruiz P.
      • Gilkeson G.S.
      Histone deacetylase inhibitors modulate renal disease in the MRL-lpr/lpr mouse.
      ,
      • Nishida K.
      • Komiyama T.
      • Miyazawa S.
      • Shen Z.N.
      • Furumatsu T.
      • Doi H.
      • Yoshida A.
      • Yamana J.
      • Yamamura M.
      • Ninomiya Y.
      • Inoue H.
      • Asahara H.
      Histone deacetylase inhibitor suppression of autoantibody-mediated arthritis in mice via regulation of p16INK4a and p21(WAF1/Cip1) expression.
      ,
      • Tao R.
      • de Zoeten E.F.
      • Ozkaynak E.
      • Chen C.
      • Wang L.
      • Porrett P.M.
      • Li B.
      • Turka L.A.
      • Olson E.N.
      • Greene M.I.
      • Wells A.D.
      • Hancock W.W.
      Deacetylase inhibition promotes the generation and function of regulatory T cells.
      ,
      • Reddy P.
      • Sun Y.
      • Toubai T.
      • Duran-Struuck R.
      • Clouthier S.G.
      • Weisiger E.
      • Maeda Y.
      • Tawara I.
      • Krijanovski O.
      • Gatza E.
      • Liu C.
      • Malter C.
      • Mascagni P.
      • Dinarello C.A.
      • Ferrara J.L.
      Histone deacetylase inhibition modulates indoleamine 2,3-dioxygenase-dependent DC functions and regulates experimental graft-versus-host disease in mice.
      ,
      • Choi J.H.
      • Oh S.W.
      • Kang M.S.
      • Kwon H.J.
      • Oh G.T.
      • Kim D.Y.
      Trichostatin A attenuates airway inflammation in mouse asthma model.
      ).
      Complementary to our findings, Wayne Hancock and co-workers (
      • Tao R.
      • de Zoeten E.F.
      • Ozkaynak E.
      • Chen C.
      • Wang L.
      • Porrett P.M.
      • Li B.
      • Turka L.A.
      • Olson E.N.
      • Greene M.I.
      • Wells A.D.
      • Hancock W.W.
      Deacetylase inhibition promotes the generation and function of regulatory T cells.
      ,
      • de Zoeten E.F.
      • Wang L.
      • Sai H.
      • Dillmann W.H.
      • Hancock W.W.
      Inhibition of HDAC9 increases T regulatory cell function and prevents colitis in mice.
      ) showed an HDAC inhibitor-dependent increase in Treg cells that mediated the anti-inflammatory effect. Because experimental colitis was associated with an increased local expression of HDAC9, HDAC9 knockout mice were subsequently investigated in models of experimental colitis where they proved to be protected (
      • Tao R.
      • de Zoeten E.F.
      • Ozkaynak E.
      • Chen C.
      • Wang L.
      • Porrett P.M.
      • Li B.
      • Turka L.A.
      • Olson E.N.
      • Greene M.I.
      • Wells A.D.
      • Hancock W.W.
      Deacetylase inhibition promotes the generation and function of regulatory T cells.
      ,
      • de Zoeten E.F.
      • Wang L.
      • Sai H.
      • Dillmann W.H.
      • Hancock W.W.
      Inhibition of HDAC9 increases T regulatory cell function and prevents colitis in mice.
      ). For the IL-17-producing T cells, Koenen et al. (
      • Koenen H.J.
      • Smeets R.L.
      • Vink P.M.
      • van Rijssen E.
      • Boots A.M.
      • Joosten I.
      Human CD25highFoxp3pos regulatory T cells differentiate into IL-17-producing cells.
      ) described a reduced in vitro differentiation of Treg to Th17 cells in the presence of pan-HDAC inhibitors. Furthermore, Bosisio et al. (
      • Bosisio D.
      • Vulcano M.
      • Del Prete A.
      • Sironi M.
      • Salvi V.
      • Salogni L.
      • Riboldi E.
      • Leoni F.
      • Dinarello C.A.
      • Girolomoni G.
      • Sozzani S.
      Blocking TH17-polarizing cytokines by histone deacetylase inhibitors in vitro and in vivo.
      ) suggest that the production of Th1- and Th17-polarizing cytokines by dendritic cells is suppressed by HDAC inhibition. Together with our data regarding the lack of Th1 polarizing cytokines and the decrease in Th17 cells, an explanation for the anti-inflammatory potency in various models of inflammation is provided.
      Considering the effect of HDAC inhibition on IL-6 production and the critical role of IL-6 in the polarization of naïve CD4+ T cells toward Th17 cells, the suppression of IL-6 signaling could result in a consecutive decrease of Th17 cells. Thus, our following studies targeted the IL-6/IL-6R pathway. IL-6 exerts its function by binding to the IL-6R, starting a cascade, with STAT3 being one of the key signal transduction molecules downstream (
      • Stephanou A.
      • Isenberg D.A.
      • Akira S.
      • Kishimoto T.
      • Latchman D.S.
      The nuclear factor interleukin-6 (NF-IL6) and signal transducer and activator of transcription-3 (STAT-3) signalling pathways co-operate to mediate the activation of the hsp90β gene by interleukin-6 but have opposite effects on its inducibility by heat shock.
      ). HDAC inhibition not only resulted in a down-regulation of the IL-6R but, consecutively, lead to a dose-dependent reduction of IL-6-induced STAT3 phosphorylation in naïve CD4+ T cells. Modifications of this pathway lead directly to RORγT, the key transcription factor for IL17 production, and, therefore, to the polarization of Th17 cells (
      • Yang X.O.
      • Panopoulos A.D.
      • Nurieva R.
      • Chang S.H.
      • Wang D.
      • Watowich S.S.
      • Dong C.
      STAT3 regulates cytokine-mediated generation of inflammatory helper T cells.
      ). Consequently, RORγT is also down-regulated in ITF2357-treated cells. To prove a direct impact of the HDAC inhibitor on the IL-6R expression on the chromatin level, we assessed changes in the histone acetylation pattern on the respective loci. Although our ChIP data revealed a hyperacetylation for the IL-6R gene locus, which is mostly associated with an increased transcription of the respective region, the IL-6R promoter locus was, in fact, deacetylated, indicating reduced gene transcription and, thus, confirming the proposed mechanism.
      Furthermore, our data demonstrate that, also in macrophages, the expression of the IL-6R is diminished by ITF2357, even in its soluble form. Given that the secretion of IL-6 itself is reduced in macrophages, one of the main sources of this cytokine in the lamina propria, we were able to describe a massive down-modulation of the IL-6 signaling pathway in our models.
      In view of these data and the literature discussed, we propose the IL-6/STAT3/IL-17 pathway as critical in the anti-inflammatory effect achieved by HDAC inhibition. Other studies have described IL-6 as the determining factor in the polarization of Th17 from naïve CD4+ T cells, thus being crucial for obtaining the balance between Treg and Th17 cells (
      • Ichiyama K.
      • Yoshida H.
      • Wakabayashi Y.
      • Chinen T.
      • Saeki K.
      • Nakaya M.
      • Takaesu G.
      • Hori S.
      • Yoshimura A.
      • Kobayashi T.
      Foxp3 inhibits RORγt-mediated IL-17A mRNA transcription through direct interaction with RORγt.
      ). Because naïve CD4+ T cells are one of the few cell types in possession of the membrane-bound IL-6R, they are offering a possible target for the indicated regulation of T cell polarization (
      • Jones G.W.
      • McLoughlin R.M.
      • Hammond V.J.
      • Parker C.R.
      • Williams J.D.
      • Malhotra R.
      • Scheller J.
      • Williams A.S.
      • Rose-John S.
      • Topley N.
      • Jones S.A.
      Loss of CD4+ T cell IL-6R expression during inflammation underlines a role for IL-6 trans signaling in the local maintenance of Th17 cells.
      ). Complementary to the results presented here, it has been described that treatment of multiple myeloma cells with HDAC inhibitors resulted in down-modulation of IL-6R signaling (
      • Mitsiades C.S.
      • Mitsiades N.S.
      • McMullan C.J.
      • Poulaki V.
      • Shringarpure R.
      • Hideshima T.
      • Akiyama M.
      • Chauhan D.
      • Munshi N.
      • Gu X.
      • Bailey C.
      • Joseph M.
      • Libermann T.A.
      • Richon V.M.
      • Marks P.A.
      • Anderson K.C.
      Transcriptional signature of histone deacetylase inhibition in multiple myeloma. Biological and clinical implications.
      ,
      • Todoerti K.
      • Barbui V.
      • Pedrini O.
      • Lionetti M.
      • Fossati G.
      • Mascagni P.
      • Rambaldi A.
      • Neri A.
      • Introna M.
      • Lombardi L.
      • Golay J.
      Pleiotropic anti-myeloma activity of ITF2357. Inhibition of interleukin-6 receptor signaling and repression of miR-19a and miR-19b.
      ). Here, the authors emphasized the functional consequences achieved by down-modulation of the IL-6R because the response to IL-6 was equally reduced, as determined by STAT3 phosphorylation (
      • Todoerti K.
      • Barbui V.
      • Pedrini O.
      • Lionetti M.
      • Fossati G.
      • Mascagni P.
      • Rambaldi A.
      • Neri A.
      • Introna M.
      • Lombardi L.
      • Golay J.
      Pleiotropic anti-myeloma activity of ITF2357. Inhibition of interleukin-6 receptor signaling and repression of miR-19a and miR-19b.
      ). Clinical studies are underway to evaluate HDAC inhibitors as a treatment option for relapsed multiple myeloma (
      • Weber D.M.
      • Graef T.
      • Hussein M.
      • Sobecks R.M.
      • Schiller G.J.
      • Lupinacci L.
      • Hardwick J.S.
      • Jagannath S.
      Phase I trial of vorinostat combined with bortezomib for the treatment of relapsing and/or refractory multiple myeloma.
      ), adding to the variety of other substances targeting the IL-6R in pharmacotherapy for different diseases, such as rheumatoid arthritis or Crohn disease, with remarkable clinical effects (
      • Ichiyama K.
      • Yoshida H.
      • Wakabayashi Y.
      • Chinen T.
      • Saeki K.
      • Nakaya M.
      • Takaesu G.
      • Hori S.
      • Yoshimura A.
      • Kobayashi T.
      Foxp3 inhibits RORγt-mediated IL-17A mRNA transcription through direct interaction with RORγt.
      ,
      • Ito H.
      • Takazoe M.
      • Fukuda Y.
      • Hibi T.
      • Kusugami K.
      • Andoh A.
      • Matsumoto T.
      • Yamamura T.
      • Azuma J.
      • Nishimoto N.
      • Yoshizaki K.
      • Shimoyama T.
      • Kishimoto T.
      A pilot randomized trial of a human anti-interleukin-6 receptor monoclonal antibody in active Crohn's disease.
      ,
      • Maini R.N.
      • Taylor P.C.
      • Szechinski J.
      • Pavelka K.
      • Bröll J.
      • Balint G.
      • Emery P.
      • Raemen F.
      • Petersen J.
      • Smolen J.
      • Thomson D.
      • Kishimoto T.
      • CHARISMA Study Group
      Double-blind randomized controlled clinical trial of the interleukin-6 receptor antagonist, tocilizumab, in European patients with rheumatoid arthritis who had an incomplete response to methotrexate.
      ). Hence, HDAC inhibitors might provide a novel therapeutic approach for the treatment of autoimmune diseases and other diseases by targeting the IL-6 receptor pathway and, thereby, influencing the critical balance of T cell subpopulations at the site of inflammation.

      Acknowledgments

      We thank Inka Freise for technical assistance and Ulrike Erben for critical reading of the manuscript.

      References

        • Kelly W.K.
        • Marks P.A.
        Drug insight. Histone deacetylase inhibitors. Development of the new targeted anticancer agent suberoylanilide hydroxamic acid.
        Nat. Clin. Pract. Oncol. 2005; 2: 150-157
        • Marks P.
        • Rifkind R.A.
        • Richon V.M.
        • Breslow R.
        • Miller T.
        • Kelly W.K.
        Histone deacetylases and cancer. Causes and therapies.
        Nat. Rev. Cancer. 2001; 1: 194-202
        • Marks P.A.
        • Breslow R.
        Dimethyl sulfoxide to vorinostat. Development of this histone deacetylase inhibitor as an anticancer drug.
        Nat. Biotechnol. 2007; 25: 84-90
        • Krämer O.H.
        • Baus D.
        • Knauer S.K.
        • Stein S.
        • Jäger E.
        • Stauber R.H.
        • Grez M.
        • Pfitzner E.
        • Heinzel T.
        Acetylation of Stat1 modulates NF-κB activity.
        Genes Dev. 2006; 20: 473-485
        • Glauben R.
        • Batra A.
        • Stroh T.
        • Erben U.
        • Fedke I.
        • Lehr H.A.
        • Leoni F.
        • Mascagni P.
        • Dinarello C.A.
        • Zeitz M.
        • Siegmund B.
        Histone deacetylases. Novel targets for prevention of colitis-associated cancer in mice.
        Gut. 2008; 57: 613-622
        • Glauben R.
        • Batra A.
        • Fedke I.
        • Zeitz M.
        • Lehr H.A.
        • Leoni F.
        • Mascagni P.
        • Fantuzzi G.
        • Dinarello C.A.
        • Siegmund B.
        Histone hyperacetylation is associated with amelioration of experimental colitis in mice.
        J. Immunol. 2006; 176: 5015-5022
        • Leoni F.
        • Zaliani A.
        • Bertolini G.
        • Porro G.
        • Pagani P.
        • Pozzi P.
        • Donà G.
        • Fossati G.
        • Sozzani S.
        • Azam T.
        • Bufler P.
        • Fantuzzi G.
        • Goncharov I.
        • Kim S.H.
        • Pomerantz B.J.
        • Reznikov L.L.
        • Siegmund B.
        • Dinarello C.A.
        • Mascagni P.
        The antitumor histone deacetylase inhibitor suberoylanilide hydroxamic acid exhibits antiinflammatory properties via suppression of cytokines.
        Proc. Natl. Acad. Sci. U.S.A. 2002; 99: 2995-3000
        • Mishra N.
        • Reilly C.M.
        • Brown D.R.
        • Ruiz P.
        • Gilkeson G.S.
        Histone deacetylase inhibitors modulate renal disease in the MRL-lpr/lpr mouse.
        J. Clin. Invest. 2003; 111: 539-552
        • Nishida K.
        • Komiyama T.
        • Miyazawa S.
        • Shen Z.N.
        • Furumatsu T.
        • Doi H.
        • Yoshida A.
        • Yamana J.
        • Yamamura M.
        • Ninomiya Y.
        • Inoue H.
        • Asahara H.
        Histone deacetylase inhibitor suppression of autoantibody-mediated arthritis in mice via regulation of p16INK4a and p21(WAF1/Cip1) expression.
        Arthritis Rheum. 2004; 50: 3365-3376
        • Tao R.
        • de Zoeten E.F.
        • Ozkaynak E.
        • Chen C.
        • Wang L.
        • Porrett P.M.
        • Li B.
        • Turka L.A.
        • Olson E.N.
        • Greene M.I.
        • Wells A.D.
        • Hancock W.W.
        Deacetylase inhibition promotes the generation and function of regulatory T cells.
        Nat. Med. 2007; 13: 1299-1307
        • Reddy P.
        • Sun Y.
        • Toubai T.
        • Duran-Struuck R.
        • Clouthier S.G.
        • Weisiger E.
        • Maeda Y.
        • Tawara I.
        • Krijanovski O.
        • Gatza E.
        • Liu C.
        • Malter C.
        • Mascagni P.
        • Dinarello C.A.
        • Ferrara J.L.
        Histone deacetylase inhibition modulates indoleamine 2,3-dioxygenase-dependent DC functions and regulates experimental graft-versus-host disease in mice.
        J. Clin. Invest. 2008; 118: 2562-2573
        • Choi J.H.
        • Oh S.W.
        • Kang M.S.
        • Kwon H.J.
        • Oh G.T.
        • Kim D.Y.
        Trichostatin A attenuates airway inflammation in mouse asthma model.
        Clin. Exp. Allergy. 2005; 35: 89-96
        • Vojinovic J.
        • Damjanov N.
        • D'Urzo C.
        • Furlan A.
        • Susic G.
        • Pasic S.
        • Iagaru N.
        • Stefan M.
        • Dinarello C.A.
        Safety and efficacy of an oral histone deacetylase inhibitor in systemic-onset juvenile idiopathic arthritis.
        Arthritis Rheum. 2011; 63: 1452-1458
        • Mucida D.
        • Park Y.
        • Kim G.
        • Turovskaya O.
        • Scott I.
        • Kronenberg M.
        • Cheroutre H.
        Reciprocal TH17 and regulatory T cell differentiation mediated by retinoic acid.
        Science. 2007; 317: 256-260
        • Fantini M.C.
        • Rizzo A.
        • Fina D.
        • Caruso R.
        • Becker C.
        • Neurath M.F.
        • Macdonald T.T.
        • Pallone F.
        • Monteleone G.
        IL-21 regulates experimental colitis by modulating the balance between Treg and Th17 cells.
        Eur. J. Immunol. 2007; 37: 3155-3163
        • Elson C.O.
        • Cong Y.
        • Weaver C.T.
        • Schoeb T.R.
        • McClanahan T.K.
        • Fick R.B.
        • Kastelein R.A.
        Monoclonal anti-interleukin 23 reverses active colitis in a T cell-mediated model in mice.
        Gastroenterology. 2007; 132: 2359-2370
        • Sands B.E.
        • Jacobson E.W.
        • Sylwestrowicz T.
        • Younes Z.
        • Dryden G.
        • Fedorak R.
        • Greenbloom S.
        Randomized, double-blind, placebo-controlled trial of the oral interleukin-12/23 inhibitor apilimod mesylate for treatment of active Crohn's disease.
        Inflamm. Bowel Dis. 2010; 16: 1209-1218
        • Sandborn W.J.
        • Gasink C.
        • Gao L.L.
        • Blank M.A.
        • Johanns J.
        • Guzzo C.
        • Sands B.E.
        • Hanauer S.B.
        • Targan S.
        • Rutgeerts P.
        • Ghosh S.
        • de Villiers W.J.
        • Panaccione R.
        • Greenberg G.
        • Schreiber S.
        • Lichtiger S.
        • Feagan B.G.
        Ustekinumab induction and maintenance therapy in refractory Crohn's disease.
        N. Engl. J. Med. 2012; 367: 1519-1528
        • Izcue A.
        • Coombes J.L.
        • Powrie F.
        Regulatory T cells suppress systemic and mucosal immune activation to control intestinal inflammation.
        Immunol. Rev. 2006; 212: 256-271
        • Chen W.
        • Jin W.
        • Hardegen N.
        • Lei K.J.
        • Li L.
        • Marinos N.
        • McGrady G.
        • Wahl S.M.
        Conversion of peripheral CD4+CD25 naive T cells to CD4+CD25+ regulatory T cells by TGF-β induction of transcription factor Foxp3.
        J. Exp. Med. 2003; 198: 1875-1886
        • Volpe E.
        • Servant N.
        • Zollinger R.
        • Bogiatzi S.I.
        • Hupé P.
        • Barillot E.
        • Soumelis V.
        A critical function for transforming growth factor-β, interleukin 23 and proinflammatory cytokines in driving and modulating human T(H)-17 responses.
        Nat. Immunol. 2008; 9: 650-657
        • Bettelli E.
        • Carrier Y.
        • Gao W.
        • Korn T.
        • Strom T.B.
        • Oukka M.
        • Weiner H.L.
        • Kuchroo V.K.
        Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells.
        Nature. 2006; 441: 235-238
        • Mangan P.R.
        • Harrington L.E.
        • O'Quinn D.B.
        • Helms W.S.
        • Bullard D.C.
        • Elson C.O.
        • Hatton R.D.
        • Wahl S.M.
        • Schoeb T.R.
        • Weaver C.T.
        Transforming growth factor-β induces development of the T(H)17 lineage.
        Nature. 2006; 441: 231-234
        • Veldhoen M.
        • Hocking R.J.
        • Atkins C.J.
        • Locksley R.M.
        • Stockinger B.
        TGFβ in the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells.
        Immunity. 2006; 24: 179-189
        • Hirano T.
        • Yasukawa K.
        • Harada H.
        • Taga T.
        • Watanabe Y.
        • Matsuda T.
        • Kashiwamura S.
        • Nakajima K.
        • Koyama K.
        • Iwamatsu A.
        Complementary DNA for a novel human interleukin (BSF-2) that induces B lymphocytes to produce immunoglobulin.
        Nature. 1986; 324: 73-76
        • Hideshima T.
        • Mitsiades C.
        • Tonon G.
        • Richardson P.G.
        • Anderson K.C.
        Understanding multiple myeloma pathogenesis in the bone marrow to identify new therapeutic targets.
        Nat. Rev. Cancer. 2007; 7: 585-598
        • Hibi M.
        • Murakami M.
        • Saito M.
        • Hirano T.
        • Taga T.
        • Kishimoto T.
        Molecular cloning and expression of an IL-6 signal transducer, gp130.
        Cell. 1990; 63: 1149-1157
        • Rose-John S.
        • Heinrich P.C.
        Soluble receptors for cytokines and growth factors. Generation and biological function.
        Biochem. J. 1994; 300: 281-290
        • Murphy K.M.
        • Heimberger A.B.
        • Loh D.Y.
        Induction by antigen of intrathymic apoptosis of CD4+CD8+TCRlo thymocytes in vivo.
        Science. 1990; 250: 1720-1723
        • Stephanou A.
        • Isenberg D.A.
        • Akira S.
        • Kishimoto T.
        • Latchman D.S.
        The nuclear factor interleukin-6 (NF-IL6) and signal transducer and activator of transcription-3 (STAT-3) signalling pathways co-operate to mediate the activation of the hsp90β gene by interleukin-6 but have opposite effects on its inducibility by heat shock.
        Biochem. J. 1998; 330: 189-195
        • Farache J.
        • Zigmond E.
        • Shakhar G.
        • Jung S.
        Contributions of dendritic cells and macrophages to intestinal homeostasis and immune defense.
        Immunol. Cell Biol. 2013; 91: 232-239
        • Hosokawa T.
        • Kusugami K.
        • Ina K.
        • Ando T.
        • Shinoda M.
        • Imada A.
        • Ohsuga M.
        • Sakai T.
        • Matsuura T.
        • Ito K.
        • Kaneshiro K.
        Interleukin-6 and soluble interleukin-6 receptor in the colonic mucosa of inflammatory bowel disease.
        J. Gastroenterol. Hepatol. 1999; 14: 987-996
        • Powrie F.
        Immune regulation in the intestine. A balancing act between effector and regulatory T cell responses.
        Ann. N.Y. Acad. Sci. 2004; 1029: 132-141
        • Siegmund B.
        • Zeitz M.
        Clinical aspects of inflammatory bowel disease.
        Eur. J. Immunol. 2009; 39: 2026-2030
        • Annunziato F.
        • Cosmi L.
        • Santarlasci V.
        • Maggi L.
        • Liotta F.
        • Mazzinghi B.
        • Parente E.
        • Filì L.
        • Ferri S.
        • Frosali F.
        • Giudici F.
        • Romagnani P.
        • Parronchi P.
        • Tonelli F.
        • Maggi E.
        • Romagnani S.
        Phenotypic and functional features of human Th17 cells.
        J. Exp. Med. 2007; 204: 1849-1861
        • Fujino S.
        • Andoh A.
        • Bamba S.
        • Ogawa A.
        • Hata K.
        • Araki Y.
        • Bamba T.
        • Fujiyama Y.
        Increased expression of interleukin 17 in inflammatory bowel disease.
        Gut. 2003; 52: 65-70
        • Noguchi D.
        • Wakita D.
        • Ohkuri T.
        • Tajima M.
        • Chamoto K.
        • Kitamura H.
        • Nishimura T.
        Blockade of IL-6-signaling inhibits the pathogenesis of CD4+ T cell-mediated lethal graft-versus-host reaction against minor histocompatibility antigen.
        Immunol. Lett. 2011; 136: 146-155
        • Dohi T.
        • Fujihashi K.
        Type 1 and 2 T helper cell-mediated colitis.
        Curr. Opin. Gastroenterol. 2006; 22: 651-657
        • Leppkes M.
        • Becker C.
        • Ivanov I.I.
        • Hirth S.
        • Wirtz S.
        • Neufert C.
        • Pouly S.
        • Murphy A.J.
        • Valenzuela D.M.
        • Yancopoulos G.D.
        • Becher B.
        • Littman D.R.
        • Neurath M.F.
        RORγ-expressing Th17 cells induce murine chronic intestinal inflammation via redundant effects of IL-17A and IL-17F.
        Gastroenterology. 2009; 136: 257-267
        • Mottet C.
        • Uhlig H.H.
        • Powrie F.
        Cutting edge. Cure of colitis by CD4+CD25+ regulatory T cells.
        J. Immunol. 2003; 170: 3939-3943
        • Kjellev S.
        • Lundsgaard D.
        • Poulsen S.S.
        • Markholst H.
        Reconstitution of Scid mice with CD4+CD25 T cells leads to rapid colitis. An improved model for pharmacologic testing.
        Int. Immunopharmacol. 2006; 6: 1341-1354
        • Leoni F.
        • Fossati G.
        • Lewis E.C.
        • Lee J.K.
        • Porro G.
        • Pagani P.
        • Modena D.
        • Moras M.L.
        • Pozzi P.
        • Reznikov L.L.
        • Siegmund B.
        • Fantuzzi G.
        • Dinarello C.A.
        • Mascagni P.
        The histone deacetylase inhibitor ITF2357 reduces production of pro-inflammatory cytokines in vitro and systemic inflammation in vivo.
        Mol. Med. 2005; 11: 1-15
        • de Zoeten E.F.
        • Wang L.
        • Sai H.
        • Dillmann W.H.
        • Hancock W.W.
        Inhibition of HDAC9 increases T regulatory cell function and prevents colitis in mice.
        Gastroenterology. 2010; 138: 583-594
        • Koenen H.J.
        • Smeets R.L.
        • Vink P.M.
        • van Rijssen E.
        • Boots A.M.
        • Joosten I.
        Human CD25highFoxp3pos regulatory T cells differentiate into IL-17-producing cells.
        Blood. 2008; 112: 2340-2352
        • Bosisio D.
        • Vulcano M.
        • Del Prete A.
        • Sironi M.
        • Salvi V.
        • Salogni L.
        • Riboldi E.
        • Leoni F.
        • Dinarello C.A.
        • Girolomoni G.
        • Sozzani S.
        Blocking TH17-polarizing cytokines by histone deacetylase inhibitors in vitro and in vivo.
        J. Leukocyte Biol. 2008; 84: 1540-1548
        • Yang X.O.
        • Panopoulos A.D.
        • Nurieva R.
        • Chang S.H.
        • Wang D.
        • Watowich S.S.
        • Dong C.
        STAT3 regulates cytokine-mediated generation of inflammatory helper T cells.
        J. Biol. Chem. 2007; 282: 9358-9363
        • Ichiyama K.
        • Yoshida H.
        • Wakabayashi Y.
        • Chinen T.
        • Saeki K.
        • Nakaya M.
        • Takaesu G.
        • Hori S.
        • Yoshimura A.
        • Kobayashi T.
        Foxp3 inhibits RORγt-mediated IL-17A mRNA transcription through direct interaction with RORγt.
        J. Biol. Chem. 2008; 283: 17003-17008
        • Jones G.W.
        • McLoughlin R.M.
        • Hammond V.J.
        • Parker C.R.
        • Williams J.D.
        • Malhotra R.
        • Scheller J.
        • Williams A.S.
        • Rose-John S.
        • Topley N.
        • Jones S.A.
        Loss of CD4+ T cell IL-6R expression during inflammation underlines a role for IL-6 trans signaling in the local maintenance of Th17 cells.
        J. Immunol. 2010; 184: 2130-2139
        • Mitsiades C.S.
        • Mitsiades N.S.
        • McMullan C.J.
        • Poulaki V.
        • Shringarpure R.
        • Hideshima T.
        • Akiyama M.
        • Chauhan D.
        • Munshi N.
        • Gu X.
        • Bailey C.
        • Joseph M.
        • Libermann T.A.
        • Richon V.M.
        • Marks P.A.
        • Anderson K.C.
        Transcriptional signature of histone deacetylase inhibition in multiple myeloma. Biological and clinical implications.
        Proc. Natl. Acad. Sci. U.S.A. 2004; 101: 540-545
        • Todoerti K.
        • Barbui V.
        • Pedrini O.
        • Lionetti M.
        • Fossati G.
        • Mascagni P.
        • Rambaldi A.
        • Neri A.
        • Introna M.
        • Lombardi L.
        • Golay J.
        Pleiotropic anti-myeloma activity of ITF2357. Inhibition of interleukin-6 receptor signaling and repression of miR-19a and miR-19b.
        Haematologica. 2010; 95: 260-269
        • Weber D.M.
        • Graef T.
        • Hussein M.
        • Sobecks R.M.
        • Schiller G.J.
        • Lupinacci L.
        • Hardwick J.S.
        • Jagannath S.
        Phase I trial of vorinostat combined with bortezomib for the treatment of relapsing and/or refractory multiple myeloma.
        Clin. Lymphoma Myeloma Leuk. 2012; 12: 319-324
        • Ito H.
        • Takazoe M.
        • Fukuda Y.
        • Hibi T.
        • Kusugami K.
        • Andoh A.
        • Matsumoto T.
        • Yamamura T.
        • Azuma J.
        • Nishimoto N.
        • Yoshizaki K.
        • Shimoyama T.
        • Kishimoto T.
        A pilot randomized trial of a human anti-interleukin-6 receptor monoclonal antibody in active Crohn's disease.
        Gastroenterology. 2004; 126: 989-996
        • Maini R.N.
        • Taylor P.C.
        • Szechinski J.
        • Pavelka K.
        • Bröll J.
        • Balint G.
        • Emery P.
        • Raemen F.
        • Petersen J.
        • Smolen J.
        • Thomson D.
        • Kishimoto T.
        • CHARISMA Study Group
        Double-blind randomized controlled clinical trial of the interleukin-6 receptor antagonist, tocilizumab, in European patients with rheumatoid arthritis who had an incomplete response to methotrexate.
        Arthritis Rheum. 2006; 54: 2817-2829