The effect of selective inhibition of HDAC6 with ACY1215 on bortezomib activity in multiple myeloma (MM).

Abstract

e18569 Background: HDACs are being studied as novel therapeutic targets in MM. Here, we investigated the preclinical activity of an HDAC6 selective inhibitor ACY1215 in MM, alone and in combination with bortezomib (BZ). Methods: In vitro enzyme assays were used to evaluate the selectivity of ACY1215. The cytotoxicity of ACY1215 versus pan-HDAC inhibitor (e.g. SAHA) were compared in MM cell lines, MM patient cells, MM osteoblasts (OB) and osteoclasts (OC), as well as peripheral blood mononuclear cells (PBMCs) from healthy donors. Western blotting and RT-PCR were used to identify the mechanisms of cell death. The activity of ACY1215 and BZ was confirmed in vivo using two different murine xenograft models of human MM, plasmacytoma model and disseminated MM model. Results: ACY1215 showed more potent inhibitory activity against HDAC6 (EC50 0.0054 µM) than SAHA. Like SAHA, ACY1215 triggered potent MM cytotoxicity but showed less toxicity than SAHA against PBMCs and activated T-cells. Low doses of ACY1215 with BZ showed synergistic anti MM activity, resulting in apoptosis via caspase-8,-9,-3 and PARP activation. Moreover, ACY1215 increased BZ-induced ER stress in MM cells, evidenced by induction of UPR components including PERK, CHOP and p-IRE1α. ACY1215, alone and with BZ, enhanced OB differentiation (increased alkaline phosphatase enzyme activity) and increased mRNA expression of β-catenin, osteocalcin, and Runx2 (OB markers). Importantly, both treatments inhibited OC differentiation (reduced number of multinucleated cells staining for TRAP). In vivo studies confirmed decreased human MM cell growth and prolonged host survival after combination ACY1215 and BZ therapy versus controls (34 vs 22 days, n=7, p<0.0011, in plasmacytoma model and 40 vs 17 days, n=12, p<0.0001, in disseminated model), without significant adverse effects. PK/PD studies showed ACY1215 plasma levels (2133 ng/ml) at 1 hr following treatment. Immunohistochemistry on excised human MM demonstrated increased in acetylated-α tubulin, a biomarker of HDAC6 inhibition, stained cells 4 hrs after ACY1215 treatment. Conclusions: These results provide the rationale for clinical evaluation of ACY1215, alone and together with BZ, in the treatment of MM.