(PA-523) HDAC and CDK Inhibitor Combinations Synergize in Limiting Myeloma

Human MM cell lines treated with dinaciclib (CDK inhibitor), entinostat (HDAC inhibitor), or the combination, were assessed for viability (MTS assay), apoptosis (Annexin V/PI staining), protein expression (Western blotting), and proliferation (Incucyte). Primary CD138⁺ MM cells were isolated from patient bone marrow aspirates and tested ex vivo with single agents and the combination. In vivo efficacy was evaluated in NSG xenografts and Bcl-xL spontaneous tumor models (caliper, bioluminescense). RNA-seq was performed on treated L363 cells, followed by differential expression analysis (GSEA and DAVID).

Results:   Combined CDK/HDAC inhibition with dinaciclib and entinostat in MM cells reduced MYC expression, increased p16, induced apoptosis, and suppressed RNA Pol II phosphorylation more effectively than single agents. Target engagement was confirmed by downregulation of CDK1/2/5 and HDAC1/3 in both parental and proteasome inhibitor–resistant LP1 cells. Viability, proliferation, and synergy analyses revealed that the combination is more effective than either drug alone and was potent in drug-resistant MM models. Combined CDK/HDAC inhibition was well tolerated in NSG and Bcl-xL mice at optimized dosing regimens without significant toxicity. In NSG mice bearing L363 MM xenografts, the combination reduced MYC, increased p16, delayed tumor growth, and significantly improved survival compared to single agents. In the Bcl-xL immunocompetent model, survival improvement was achieved, though combination therapy was not superior to single agents. Combined CDK/HDAC inhibition with dinaciclib and entinostat selectively reduced the viability of CD138⁺ SMM patient cells more effectively than single agents, while sparing CD138⁻ non-neoplastic cells. RNA-seq analysis of L363 myeloma cells treated with dinaciclib and/or entinostat revealed potential biomarkers and master regulators of CDKi/HDACi synergy, with downregulation of replication proteins suggesting a mechanism involving replication stress, DNA damage, and apoptosis. To address translational limitations of dinaciclib and entinostat, we tested next-generation agents: the selective oral CDK9 inhibitor KB0742 combined with the HDAC inhibitors quisinostat or zabadinostat, which have improved pharmacokinetics and safety. The synergistic action of these combinations was validated.

Conclusions:   

Further investigation into a combined genetic signature will aid in discovering mechanisms of drug synergy and provide biomarkers for a combined response that may translate to the clinic.