Abstract 251: Kinome-wide RNAi screen identification of synthetic lethal interactions in BRAF-mutant melanoma

Abstract

Abstract BACKGROUND. Mutations in the BRAF oncogene are prevalent in many types of solid cancers, where they promote cell proliferation and transformation by constitutively activating the RAF-MEK-ERK signaling pathway. Inhibitors of BRAF have been developed as potential cancer therapies but unfortunately many have performed poorly in clinical trials. As an alternative approach, we performed RNA interference (RNAi) screening to identify genes that are synthetically lethal with the BRAF-mutant oncogene in malignant melanoma cell lines. Recent advances in RNAi and deep sequencing technologies have made it possible to systematically interrogate the genome for synthetic lethal interactions. METHODS. Two BRAF-mutant melanoma cell lines (SKMEL28 and Malme3M) and one wild-type BRAF cell line (MeWo) were infected with a subset of The RNAi Consortium (TRC) shRNA library comprising all the kinases in the human genome as well as the genes involved in the tyrosine kinase>RAS>RAF>MEK>ERK pathway. Cells were collected at 4 and 19 days post-infection, genomic DNA was isolated, shRNAs were PCR amplified and subjected to massively parallel sequencing. Statistical analysis identified ∼200 shRNAs that were selectively depleted in BRAF-mutant but not wild-type BRAF cell lines; individual canidate shRNA were tested in a panel of 6 melanoma cell lines 3 BRAF-mutant (UACC62, UACC257, Malme3M) and 3 wild-type BRAF (WM3912, WM3918, MeWo). SUMMARY. The screen identified a diverse number of genes that are synthetically lethal with the BRAF-mutant oncogene in human malignant melanoma. Not surprisingly, a number of the identified synthetic lethal genes are known downstream effectors of BRAF or known scaffold proteins involved in MAPK signaling. Paradoxically, epiregulin (EPGN), a known ligand of the epidermal growth factor receptor (EGFR) and EGFR itself, which are upstream of the BRAF oncogene were also identified in the screen as synthetically lethal with mutant BRAF. CONCLUSIONS. The BRAF oncogene is synthetically lethal with genes involved in diverse cellular processes such as kinase signaling, cell division, nuclear shuttling and cell metabolism. Many of the genes identified in the RNAi screen are known effectors of the MAPK cascade. Paradoxically, two members of the MAPK cascade, EGFR and its ligand EPGN, both of which are upstream of BRAF, were identified in the screen and validated to be synthetically lethal with mutant BRAF. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 251. doi:10.1158/1538-7445.AM2011-251