Oncogenic BRAFV600E mutations activate MAP kinase signaling and are associated with treatment resistance and poor prognosis in patients with colorectal cancer (CRC). In BRAFV600E mutant CRCs, treatment failure may be related to BRAFV600E -mediated apoptosis resistance that occurs by an as yet undefined mechanism. We found that BRAFV600E can upregulate anti-apoptotic MCL-1 in a gene dose-dependent manner using CRC cell lines isogenic for BRAF. BRAFV600E -induced MCL-1 upregulation was confirmed by ectopic BRAFV600E expression that activated MEK/ERK signaling to phosphorylate (MCL-1Thr163) 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 pro-apoptotic BIM to a greater extent than did vemurafenib in BRAFV600E cell lines. MCL-1 knockdown vs 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 pro-apoptotic BAK and BIM. Knockdown of BIM attenuated BAX, but not BAK, activation by cobimetinib plus A-1210477. In summary, BRAFV600E -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 BRAFV600E mutant CRCs.
- Received January 12, 2016.
- Revision received August 26, 2016.
- Accepted September 27, 2016.
- Copyright ©2016, American Association for Cancer Research.