Synergistic Interactions between Vorinostat and Sorafenib in Chronic Myelogenous Leukemia Cells Involve Mcl-1 and p21CIP1 Down-Regulation

Abstract

Interactions between the multikinase inhibitor sorafenib (Bay 43-9006) and the histone deacetylase inhibitor vorinostat were examined in chronic myelogenous leukemia (CML) cells sensitive and resistant to imatinib mesylate. K562, LAMA 84, and primary CML patient-derived CD34(+) mononuclear cells were exposed to vorinostat followed by sorafenib, after which effects on cell viability and various survival signaling pathways were monitored by flow cytometry, clonogenic assays, and Western blotting. Real-time reverse transcription-PCR was used to monitor gene expression, and the functional contribution of p21(CIP1) and Mcl-1 down-regulation were determined in cells transfected with corresponding constructs. Pretreatment (24 h) with vorinostat followed by sorafenib optimally induced mitochondrial injury and cell death in Bcr/Abl(+) cells (e.g., K562 and LAMA 84). Similar results were obtained in imatinib mesylate-resistant cells expressing activated Lyn as well as in primary CD34(+) bone marrow cells obtained from CML patients. This regimen also markedly inhibited CML cell colony formation. Combined but not individual treatment of CML cells with vorinostat and sorafenib triggered pronounced mitochondrial dysfunction (i.e., cytochrome c, Smac, and AIF release), caspase activation, poly(ADP-ribose) polymerase cleavage, and down-regulation of Mcl-1. Sorafenib also blocked vorinostat-mediated induction of p21(CIP1). Down-regulation of Mcl-1 was caspase and transcription independent, whereas p21(CIP1) down-regulation was partially caspase and transcription dependent. Enforced expression of p21(CIP1) and particularly Mcl-1 significantly attenuated vorinostat/sorafenib-mediated lethality. These findings suggest that combined treatment with vorinostat and sorafenib synergistically induces apoptosis in CML cells through a process that involves Mcl-1 down-regulation and inhibition of p21(CIP1) induction.