Abstract
Bendamustine (Cephalon, Inc., West Chester, PA), a hybrid molecule of purine analog and alkylator, induces cell death by activation of DNA damage response and apoptosis, inhibition of mitotic checkpoints, and induction of mitotic catastrophe. Entinostat (also known as MS-275 or SNDX-275, Syndax Pharmaceuticals, Inc., Waltham, MA), a selective class I inhibitor of histone deacetylase (HDAC), induces apoptosis in multiple myeloma (MM) cells. We sought to determine whether the combination of bendamustine with entinostat enhances cell death of MM cells, and the potential mechanisms. Cell growth assay showed that bendamustine inhibited proliferation of MM cells in a dose-dependent manner. The IC50 of bendamustine for RPMI8226, U266 and MM1.S cells was approximately 662.6, 295.5, and 159.6 umol/L. Apoptotic-ELISA indicated that either bendamustine or entinostat was able to induce apoptosis in all three myeloma cells, and their combinations exhibited a much more potent activity to promote the MM cells undergoing apoptosis. Further studies with western blot analyses revealed that bendamustine in combination with entinostat dramatically induced PARP cleavage and activation of caspase-3 in all three MM cell lines, and also reduced the expression levels of cyclin D1, phospho-Stat3 (P-Stat3) and Stat3 in U266 cells. Moreover, studies on DNA damage response showed that phosphohistone H2A.X, a hall marker of DNA double strand break, was significantly induced by the combinations of bendamustine and entinostat as compared to either agent alone in all three MM cell lines. Our data suggest that bendamustine and its combination with entinostat possess anti-MM activity in vitro. Regimens consisting of bendamustine and/or entinostat may represent novel therapeutic strategies against MM.
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