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-1^Thr163) and stabilize MCL-1. Upregulation of MCL-1 was mediated by MEK/ERK shown by the ability of ERK siRNA to suppress MCL-1. Stabilization of MCL-1 by phosphorylation was shown by a phosphorylation-mimicking mutant and an unphosphorylated MCL-1 mutant that decreased or increased MCL-1 protein turnover, respectively. MEK/ERK inhibition by cobimetinib suppressed MCL-1 expression/phosphorylation and induced proapoptotic BIM to 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 small-molecule MCL-1 inhibitor, A-1210477, also enhanced cobimetinib-induced apoptosis in vitro that was due to disruption of the interaction of MCL-1 with proapoptotic BAK and BIM. Knockdown of BIM attenuated BAX, but not BAK, activation by cobimetinib plus A-1210477. In summary, BRAF^V600E-mediated MEK/ERK activation can upregulate MCL-1 by phosphorylation/stabilization to confer apoptosis resistance that can be reversed by MCL-1 antagonism combined with cobimetinib, suggesting a novel therapeutic strategy against BRAF^V600E-mutant CRCs. Mol Cancer Ther; 15(12); 3015–27. ©2016 AACR.

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