Background Histone deacetylase inhibitors (HDACi) are currently being used in the clinic to treat a variety of cancer types. Despite their wide use, the mechanism by which they exert anti-tumor effects is largely unknown. Although originally posited to abrogate tumor proliferation via regulating tumor suppressor genes, responses to monotherapies of HDACi have been shown to be dependent on an adaptive immune system and to enhance responses to immunotherapy. However, whether this mechanism is driven by enhancing tumor immunogenicity or enhancing anti-tumor immune responses is unclear. Understanding this could help identify optimal combination regimens for augmenting immunotherapies. Given the role of epigenetics in regulating T cell differentiation upon antigen encounter into discrete subsets, these studies sought to determine whether HDACi differentially impact naïve from memory T cell subsets.
Methods PBMCs from healthy donors were stimulated with either anti-CD3/anti-CD28 or PMA/Ionomycin in the presence or absence of different HDAC inhibitors (OKI-005, 250 nM; Entinostat, 5 uM; and Vorinostat, 1 uM). Cells were evaluated at different time points by flow cytometric analysis to compare responses by T cell subsets for changes in cytokine production, protein acetylation and other functional responses. Supernatant was collected in separate experiments for comprehensive cytokine bead arrays.
Results Cytokine analysis of supernatants showed clear differences in response to HDACi as while most cytokines decreased, others were either unaffected or increased. We next performed ICS with surface markers to determine if these changes in cytokine production levels were subset specific. Comparisons of naïve and memory subsets found decreased IL-2 levels was primarily attributed to loss of production by naïve T cells. Furthermore, gain of TNFa was almost completely restricted to naïve cells. The preferential responses by naïve T cells was further verified during global changes in acetylated protein levels. Lastly, we found differences between inhibitors on their effects on T cells. As these differences remained even after controlling for potency, this suggests the specificity profiles toward individual HDACs was responsible for their unique effects.
Conclusions These studies demonstrate clear differences in the effect of HDACi on cytokine production by distinct T cell subsets. Ongoing studies are aimed at elucidating the specific HDACs responsible for regulating T cell effector functions and tumor immunogenicity when targeted. Ultimately, understanding this could help identify inhibitors with the desired specificity profile for combining with immunotherapy.
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