Mutant BRAF upregulates MCL-1 to confer apoptosis resistance that is reversed by MCL-1 antagonism and cobimetinib in colorectal cancer

Oncogenic BRAF^V600E mutations activate MAPK signaling and are associated with treatment resistance and poor prognosis in patients with colorectal cancer. In BRAF^V600E-mutant colorectal cancers, treatment failure may be related to BRAF^V600E-mediated apoptosis resistance that occurs by an as yet undefined mechanism. We found that BRAF^V600E can upregulate anti-Apoptotic MCL-1 in a gene dose-dependent manner using colorectal cancer cell lines isogenic for BRAF. BRAF^V600E-induced MCL-1 upregulation was confirmed by ectopic BRAF^V600E expression that activated MEK/ERK signaling to phosphorylate (MCL-1Thr163) and stabilize MCL-1.Upregulation ofMCL-1 wasmediated byMEK/ERK shown by the ability of ERK siRNA to suppress MCL-1. Stabilization of MCL-1 by phosphorylation was shown by a phosphorylationmimicking mutant and an unphosphorylated MCL-1 mutant that decreased or increasedMCL-1 protein turnover, respectively. MEK/ ERK inhibition by cobimetinib suppressed MCL-1 expression/ phosphorylation and induced proapoptotic BIMto a greater extent than did vemurafenib in BRAF^V600E cell lines. MCL-1 knockdown versus control shRNA significantly enhanced cobimetinib-induced apoptosis in vitro and in HT29 colon cancer xenografts. The smallmolecule MCL-1 inhibitor, A-1210477, also enhanced cobimetinib-induced apoptosis in vitro that was due to disruption of the interaction ofMCL-1 with proapoptoticBAK and BIM. Knockdown of BIM attenuated BAX, but not BAK, activation by cobimetinib plus A-1210477. In summary, BRAF^V600E-mediatedMEK/ERK activation can upregulate MCL-1 by phosphorylation/stabilization to confer apoptosis resistance that can be reversed by MCL-1 antagonismcombined with cobimetinib, suggesting a novel therapeutic strategy against BRAF^V600E-mutant CRCs.