title: Mutant BRAF Upregulates MCL-1 to Confer Apoptosis Resistance that Is Reversed by MCL-1 Antagonism and Cobimetinib in Colorectal Cancer.
abstract: Oncogenic BRAFV600Emutations activate MAPK signaling and are associated with treatment resistance and poor prognosis in patients with colorectal cancer. In BRAFV600E-mutant colorectal cancers, treatment failure may be related to BRAFV600E-mediated apoptosis resistance that occurs by an as yet undefined mechanism. We found that BRAFV600Ecan upregulate anti-apoptotic MCL-1 in a gene dose-dependent manner using colorectal cancer cell lines isogenic for BRAF BRAFV600E-induced MCL-1 upregulation was confirmed by ectopic BRAFV600Eexpression 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 proapoptotic BIM to a greater extent than did vemurafenib in BRAFV600Ecell 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, 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. Mol Cancer Ther; 15(12); 3015-27. ©2016 AACR.
authors.affiliation: Departments of Medicine and Oncology, Mayo Clinic, Rochester, Minnesota.
authors.forename: Hisato
authors.initials: H
authors.lastname: Kawakami
authors.affiliation: Departments of Medicine and Oncology, Mayo Clinic, Rochester, Minnesota.
authors.forename: Shengbing
authors.initials: S
authors.lastname: Huang
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Jacksonville, Florida.
authors.forename: Krishnendu
authors.initials: K
authors.lastname: Pal
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Jacksonville, Florida.
authors.forename: Shamit K
authors.initials: SK
authors.lastname: Dutta
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Jacksonville, Florida.
authors.forename: Debabrata
authors.initials: D
authors.lastname: Mukhopadhyay
authors.affiliation: Departments of Medicine and Oncology, Mayo Clinic, Rochester, Minnesota. sinicrope.frank@mayo.edu.
authors.forename: Frank A
authors.initials: FA
authors.lastname: Sinicrope
mesh_terms: D000483:Alleles; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics*; D001384:Azetidines / Q000494:pharmacology*; D000072224:Bcl-2-Like Protein 11 / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000378:metabolism; D004195:Disease Models, Animal; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects; D051379:Mice; D009154:Mutation; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D010880:Piperidines / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D023041:Xenograft Model Antitumor Assays
chemical_list: D001384:Azetidines; D000072224:Bcl-2-Like Protein 11; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D010880:Piperidines; D047428:Protein Kinase Inhibitors; D048493:Proto-Oncogene Proteins B-raf; C574276:cobimetinib