Abstract 3101: Functional analysis of HDAC11 in plasma cell development and multiple myeloma survival

Abstract

Abstract Though HDAC11 expression is confirmed in B cells and plasma cells (PC), its functions in these cells remain largely unknown. In this study, we attempted a functional analysis of HDAC11 in PC development along with its pro-tumorigenic function in multiple myeloma (MM) cells. Mouse models, including a transgenic mouse strain expressing eGFP under the regulation of the HDAC11 promoter (Tg-HDAC11-eGFP), and also an HDA11-deficient mouse (B6.HDAC11-/-) were studied to establish the importance of HDAC11 in PC biology. Pharmacologic inhibition of HDAC11 in MM cell lines was accomplished by using elevenostat (ES), a new HDAC11-selective inhibitor. Post-translational modifications and subcellular localization changes induced by ES exposure were assessed by western blotting of fractionated cell lysates, while co-immunoprecipitation (Co-IP) and proximity ligation assays (in situ PLA) were used to identify a binding partner for HDAC11. Studies in Tg-HDAC11-eGFP mice revealed that HDAC11 expression in B cell lymphopoiesis was minimally detectable prior to B cell activation but demonstrated strong induction upon maturation into a PC. Consistent with this, PC development was markedly impaired in the absence of HDAC11. Furthermore, HDAC11-selective inhibitor ES showed significant cytotoxic potential, measured by CCK-8 assay, in different MM cell lines and primary MM cells that express moderate to high level of HDAC11, with IC50 values ranging 0.6-2.0 µM. Apoptotic cell death was confirmed via detection of activated caspase-3 and annexin/propidium iodide staining by flow cytometry. Consistently, MM cell lines expressing null/very low level of HDAC11 were insensitive to ES. Moreover, combining ES with proteasome inhibitors bortezomib and carfilzomib resulted in significant synergistic effects. Furthermore, ES-treated cells showed a time-dependent alteration in the subcellular localization of HDAC11, it gradually disappeared from the nuclear fractions with simultaneous upregulation in cytoplasmic fractions. Inhibition of HDAC11 also caused downstream suppression of several pro-tumorigenic factors in MM cells, including IRF4 and c-Myc. Moreover, a novel interaction between HDAC11 and IRF4, an essential regulator of PC differentiation and MM survival, was identified by using PLA and Co-IP. HDAC11 dynamically interacts with IRF4 which can be induced by LPS and IL-6 stimulation and inhibited by ES, indicating the involvement of HDAC11 in the IRF4-mediated regulatory circuit. We also identified changes in the extent of IRF4 acetylation upon genomic or pharmacological induction/inhibition of HDAC11. Overall, we observe that targeted inhibition of HDAC11 can impair MM cell survival and overcome acquired resistance to proteasome inhibitors. Furthermore, we identify IRF4 as a nuclear binding partner of HDAC11 and propose this interaction as a candidate mechanism regulating PC maturation and survival. Citation Format: A G M MOSTOFA, Jason Brayer. Functional analysis of HDAC11 in plasma cell development and multiple myeloma survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3101.