MCL-1 as a buffer for proapoptotic BCL-2 family members during TRAIL-induced apoptosis - A mechanistic basis for sorafenib (bay 43-9006)-induced trail sensitization

Previous studies have suggested that Mcl- 1, an antiapoptotic Bcl- 2 homolog that does not exhibit appreciable affinity for the caspase 8- generated C- terminal Bid fragment ( tBid), diminishes sensitivity to tumor necrosis factor- alpha related apoptosis- inducing ligand ( TRAIL). This study was performed to determine the mechanism by which Mcl- 1 confers TRAIL resistance and to evaluate methods for overcoming this resistance. Affinity purification/ immunoblotting assays using K562 human leukemia cells, which contain Mcl- 1 and Bcl- x(L) as the predominant antiapoptotic Bcl- 2 homologs, demonstrated that TRAIL treatment resulted in binding of tBid to Bcl- xL but not Mcl- 1. In contrast, TRAIL caused increased binding between Mcl- 1 and Bak that was diminished by treatment with the caspase 8 inhibitor N-( N-alpha- acetylisoleucylglutamylthreonyl) asparticacid( O- methylester)- fluoromethylketone ( IETD( OMe)- fmk) or the c- Jun N- terminal kinase inhibitor SP600125. In addition, TRAIL caused increased binding of Bim and Puma to Mcl- 1 that was inhibited by IETD( OMe)- fmk but not SP600125. Further experiments demonstrated that down- regulation of Mcl- 1 by short hairpin RNA or the kinase inhibitor sorafenib increased TRAIL- induced Bak activation and death ligand- induced apoptosis in a wide variety of neoplastic cell lines as well as clinical acute myelogenous leukemia specimens. Collectively, these observations not only suggest a model in which Mcl- 1 confers TRAIL resistance by serving as a buffer for Bak, Bim, and Puma, but also identify sorafenib as a potential modulator of TRAIL sensitivity.