Alteration of the epigenetic landscape of immune cells can modify the pattern of gene expression, thus resulting in phenotypic and functional changes. Small molecule inhibitors targeting epigenetic modifiers, such as histone deacetylases (HDACs), have been shown to reduce tumor growth. Besides promoting direct anti-tumor effects, HDAC inhibitors also target immune cells and alter their gene regulation. Here, we demonstrate that the HDAC6 selective inhibitors ACY-241 and ACY-1215 (ricolinostat) decrease the function of myeloid derived suppressor cells (MDSC) and T regulatory (Treg) cells, maintain an effector phenotype by CD8+ T cells, and do not reduce viability of immune cells. First, peripheral blood mononuclear cells derived from melanoma patients were treated with ACY-241, and the phenotype of MDSCs was assessed. Expression of the suppressive molecules ARG1 (p<0.01) and NOS2 (p<0.05) was decreased in CD14+HLA DR lo CD11b+ and CD14+HLA DR lo CD33+ MDSC populations, suggesting less potent MDSCs. To gain insight into other suppressive populations, we evaluated the phenotype and function of Treg cells derived from melanoma patients. Cultures of CD3+ T-cells treated with ACY-1215 or ACY-241 resulted in decreased expression of FOXP3 and reduced frequency of Tregs (CD4+CD25+CD127-; p<0.001). ACY-1215 pre-treated Tregs exhibited less suppressive activity against responding conventional T-cells in standard assays, compared to Tregs pre-treated with DMSO control (p<0.01). CD3+ T-cells exposed to ACY-241 or ACY-1215 were activated via CD3/CD28 co-stimulation to assess effects on cytokine production. Selective HDAC6 inhibition shifted T-cell differentiation towards Th1-type (Tbet+) over Th2-type (GATA3+), compared to DMSO (p<0.05). Additionally, Th2 cytokines (e.g. IL-4, IL-5, IL-6, IL-10, IL-13) were significantly decreased (p<0.05). In accordance with decreased Th2 differentiation, mTORC signal transduction, including phosphor-SGK1, was similarly reduced (p<0.05). Targeted inhibition of mTORC signaling (i.e. SGK1 inhibitor) recapitulated decreased Th2 cytokine production (p<0.05). In order to address effector immune function, tumor infiltrating lymphocytes (TILs) were harvested from melanoma biopsies and expanded in vitro in the presence of ACY-1215 or ACY-241, and IL-2. Measuring CD45RA, CD45RO, CD62L and/or CCR7, an accumulation of central memory CD4+ and CD8+ T-cells was observed (p<0.05). Activated TILs treated with ACY-241 or ACY-1215 displayed higher expression of IFN-gamma and CD107a. Collectively, these data indicate that epigenetic reprograming of immune cells by HDAC6 selective inhibitors may decrease the function of suppressive subsets (e.g. Tregs, MDSCs), while enhancing the accumulation of anti-tumor memory and effector T-cell subsets.

Citation Format: Andressa L. Sodre, David M. Woods, Amod Sarnaik, Brian C. Betts, Steven Quayle, Simon Jones, Jeffrey Weber. Epigenetic reprograming of immune cells through selective inhibition of HDAC6 reduces suppressive phenotypes and augments anti-tumor properties of T-cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 638. doi:10.1158/1538-7445.AM2017-638