LAUSR

Background: Histone Deacetylases (HDACs) have been demonstrated to modify a variety of non-histone proteins involved in oncogenic- and immune-related pathways. Recent studies in macrophages and tumor cells have implicated HDAC6 in the regulation of the immunomodulatory cytokines IL10 and IL6, and the immunosuppressive proteins PD-L1 and PD-L2. Additionally, specific inhibitors for this HDAC have shown promising results when used alone or in combination with other anticancer agents. Despite this newly characterized role of HDAC6 in immunobiology, its role in the regulation of immune-related pathways in the brain parenchymal environment is still unclear. Glia cells play a key role in a number of physiological processes in the brain, including the productions of inflammatory mediators and antigen presentation. Here, we report the first evidence of the functional effects of selective HDAC6inh in the modulation of the cytokine homeostasis and immune-related pathways in antigen presenting cells in the brain. Methods: Thioglycolate elicited peritoneal macrophages (PEM), primary murine cortical astrocytes, macrophage cell lines RAW264.7 and microglial cell line BV.2 were treated with the HDAC6 inhibitors Nexturastat and Tubastatin A, and the pan-HDACinh LBH589. Cytokine production was measured by qRT-PCR and ELISA. Viability, apoptosis, cell cycle, and HDAC activity was evaluated in parallel for all tested conditions. Results: In our comparative studies, the HDAC6 inhibition significantly lowered levels of cytokines IL6, IL10, IL1β, TNFα and IL12p70 levels in macrophages, and primary murine PEMs, indicating that both, anti- and pro-inflammatory cytokines are affected by this particular HDAC. Similar results were found in the expression of the pro-inflammatory cytokines IL1β, TNFα and IL12p70b in parallel studies using BV.2 microglia cells. In contrast, we observed an increase in the expression of IL6 and only slight variations in IL10 expression in BV.2, suggesting that HDAC6 might be controlling preferentially pro-inflammatory cytokines in glia cells. Additional control studies using the non-specific pan-HDACinh LBH589 showed similar outcomes in all analyzed cytokines in macrophages and glia cells. To further investigate the functional consequences of HDAC6inh in microglia cells, we evaluated the polarization phenotype of primary astrocytes and BV.2 cells under HDAC6inh treatment. While macrophages treated with Nexturastat and Tubastatin A were equally distributed between M1 and M2, we observed an important transition from M1 to M2 in brain cells, indicating a reduction in the pro-inflammatory phenotype of these cells when treated with HDAC6inh. Conclusion: Taken collectively, our studies demonstrate that HDAC6 plays a major role in the modulation of the functional phenotype and production of pro-inflammatory mediators in brain glia cells. These findings open the possibility for further evaluation of selective HDAC6inh as adjuvants in antitumor brain therapies. Citation Format: Jennifer Kim, Jayakumar R. Nair, Melissa M. Hadley, Alan Kozikowski, Robert Miller, Alejandro Villagra. A novel regulatory role of HDAC6 in the functional inflammatory phenotype of glia 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 LB-295. doi:10.1158/1538-7445.AM2017-LB-295