Abstract
The RAF–mitogen-activated protein kinase kinase 1/2–extracellular signal-regulated kinase 1/2 (RAF–MEK1/2–ERK1/2) pathway is activated in many human tumours and can protect cells against growth factor deprivation; however, most such studies have relied upon overexpression of RAF or MEK constructs that are not found in tumours. Here we show that expression of the endogenous BRAFV600E allele in mouse embryonic fibroblasts from conditional knock-in transgenic mice activates ERK1/2, represses the BH3-only protein BIM and protects cells from growth factor withdrawal. Human colorectal cancer (CRC) cell lines harbouring BRAFV600E are growth factor independent for the activation of ERK1/2 and survival. However, treatment with the MEK1/2 inhibitors U0126, PD184352 or the novel clinical candidate AZD6244 (ARRY-142886) overcomes growth factor independence, causing CRC cell death. BIM is de-phosphorylated and upregulated following MEK1/2 inhibition in all CRC cell lines studied and knockdown of BIM reduces cell death, indicating that repression of BIM is a major part of the ability of BRAFV600E to confer growth factor-independent survival. We conclude that a single endogenous BRAFV600E allele is sufficient to repress BIM and prevent death arising from growth factor withdrawal, and CRC cells with BRAFV600E mutations are addicted to the ERK1/2 pathway for repression of BIM and growth factor-independent survival.
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Acknowledgements
We thank members of the SJC and CAP labs for discussions and Richard Marais for his advice and encouragement. We particularly thank the Division of Biomedical Services at Leicester for help with breeding and Susan Giblett for isolation of MEFs. We are grateful to the Trono lab for provision of lentiviral expression systems, Paul Smith (AstraZeneca) for provision of AZD6244 and discussions and Richard Hamelin for provision of CO115 cells. Work in the CAP lab was funded by a CRUK programme grant number C1362/A6969. Work in the SJC lab was supported by the Association for International Cancer Research (AICR), AstraZeneca, BBSRC (BB/E02162X/1) and the Babraham Institute.
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Wickenden, J., Jin, H., Johnson, M. et al. Colorectal cancer cells with the BRAFV600E mutation are addicted to the ERK1/2 pathway for growth factor-independent survival and repression of BIM. Oncogene 27, 7150–7161 (2008). https://doi.org/10.1038/onc.2008.335
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DOI: https://doi.org/10.1038/onc.2008.335
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