Person: Wang, Belinda
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Wang
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Belinda
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Wang, Belinda
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Publication Copy-number and gene dependency analysis reveals partial copy loss of wild-type SF3B1 as a novel cancer vulnerability(eLife Sciences Publications, Ltd, 2017) Paolella, Brenton R.; Gibson, William; Urbanski, Laura M; Alberta, John; Zack, Travis Ian; Bandopadhayay, Pratiti; Nichols, Caitlin; Agarwalla, Pankaj Kumar; Brown, Meredith S; Lamothe, Rebecca; Yu, Yong; Choi, Peter; Obeng, Esther A; Heckl, Dirk; Wei, Guo; Wang, Belinda; Tsherniak, Aviad; Vazquez, Francisca; Weir, Barbara Ann; Root, David E; Cowley, Glenn S; Buhrlage, Sara; Stiles, Charles; Ebert, Benjamin; Hahn, William; Reed, Robin; Beroukhim, RameenGenomic instability is a hallmark of human cancer, and results in widespread somatic copy number alterations. We used a genome-scale shRNA viability screen in human cancer cell lines to systematically identify genes that are essential in the context of particular copy-number alterations (copy-number associated gene dependencies). The most enriched class of copy-number associated gene dependencies was CYCLOPS (Copy-number alterations Yielding Cancer Liabilities Owing to Partial losS) genes, and spliceosome components were the most prevalent. One of these, the pre-mRNA splicing factor SF3B1, is also frequently mutated in cancer. We validated SF3B1 as a CYCLOPS gene and found that human cancer cells harboring partial SF3B1 copy-loss lack a reservoir of SF3b complex that protects cells with normal SF3B1 copy number from cell death upon partial SF3B1 suppression. These data provide a catalog of copy-number associated gene dependencies and identify partial copy-loss of wild-type SF3B1 as a novel, non-driver cancer gene dependency. DOI: http://dx.doi.org/10.7554/eLife.23268.001Publication KEAP1 loss modulates sensitivity to kinase targeted therapy in lung cancer(eLife Sciences Publications, Ltd, 2017) Krall, Elsa B; Wang, Belinda; Munoz, Diana M; Ilic, Nina; Raghavan, Srivatsan; Niederst, Matthew J; Yu, Kristine; Ruddy, David A; Aguirre, Andrew; Kim, Jong Wook; Redig, Amanda J; Gainor, Justin; Williams, Juliet A; Asara, John; Doench, John G; Janne, Pasi; Shaw, Alice; McDonald III, Robert E; Engelman, Jeffrey A; Stegmeier, Frank; Schlabach, Michael R; Hahn, WilliamInhibitors that target the receptor tyrosine kinase (RTK)/Ras/mitogen-activated protein kinase (MAPK) pathway have led to clinical responses in lung and other cancers, but some patients fail to respond and in those that do resistance inevitably occurs (Balak et al., 2006; Kosaka et al., 2006; Rudin et al., 2013; Wagle et al., 2011). To understand intrinsic and acquired resistance to inhibition of MAPK signaling, we performed CRISPR-Cas9 gene deletion screens in the setting of BRAF, MEK, EGFR, and ALK inhibition. Loss of KEAP1, a negative regulator of NFE2L2/NRF2, modulated the response to BRAF, MEK, EGFR, and ALK inhibition in BRAF-, NRAS-, KRAS-, EGFR-, and ALK-mutant lung cancer cells. Treatment with inhibitors targeting the RTK/MAPK pathway increased reactive oxygen species (ROS) in cells with intact KEAP1, and loss of KEAP1 abrogated this increase. In addition, loss of KEAP1 altered cell metabolism to allow cells to proliferate in the absence of MAPK signaling. These observations suggest that alterations in the KEAP1/NRF2 pathway may promote survival in the presence of multiple inhibitors targeting the RTK/Ras/MAPK pathway. DOI: http://dx.doi.org/10.7554/eLife.18970.001Publication Mechanisms of Resistance to MAPK Pathway Inhibition in RAS-Mutant Cancers(2016-05-03) Wang, Belinda; Garraway, Levi; Haigis, Kevin; Hammerman, Peter; Weinberg, RobertThe RAS family of genes are among the most frequently mutated genes in human cancers, including nearly all pancreatic cancers, ~40% of colorectal cancers, and ~30% of lung cancers. Although most RAS-mutant cancers depend on RAS signaling for proliferation and survival, direct RAS inhibitors have not yet been developed for clinical use. An alternative approach to treating RAS-mutant cancers is to inhibit RAS effector pathways, such as the RAS-RAF-MEK-ERK (MAPK) pathway. However, while some patients with RAS-mutant cancers benefit clinically from MAPK pathway inhibition (MAPKi), most do not respond to this regimen. Moreover, of the patients who do respond, nearly all progress to secondary therapeutic resistance. In this work, we applied systematic gain- and loss-of-function (GOF and LOF) screening approaches to identify modifiers of MAPK pathway dependence. Using RAS- or BRAF-mutant pancreatic and lung cancer cell lines, we performed a genome scale open reading frame (ORF) screen and six genome scale CRISPR-Cas9 knockout screens to investigate mechanisms of resistance to MEK or BRAF inhibition. We found that the most potent GOF mediators of resistance were overexpression of components of the RTK-RAS-MAPK pathway, which restored MAPK signaling. Contrastingly, the majority of LOF events that mediated resistance to MAPKi were not direct regulators or effectors of the MAPK pathway. From our LOF screens, we identified KEAP1 and CIC as generalizable modulators of resistance to MAPKi in RAS-mutant cancers. We found that KEAP1 deletion mediates resistance through mechanisms orthogonal to the MAPK pathway, such as reducing oxidative stress and promoting anabolic metabolism. Conversely, CIC loss promotes resistance by partially restoring the transcriptional pathway downstream of ERK. Understanding mechanisms of intrinsic resistance can enable the identification of predictive biomarkers that improve patient selection for targeted therapy. In addition, identifying mechanisms of acquired resistance can inform the development of novel agents or combination therapy strategies. Our studies highlight the ability of systematic and comprehensive in vitro functional screens to identify clinically relevant mediators of resistance and to provide novel insights into well-studied pathways. While we selected specific genes for detailed mechanistic studies, other genes that were identified from our screens may contribute additional biological insights.Publication Synthetic Lethal Interactions With Oncogenic KRAS(2018-05-15) Wang, BelindaKRAS is one of the most frequently mutated genes across human cancers, including 96% of pancreatic cancers, 40% of colorectal cancers, and 35% of lung cancers. The majority of human cancer cell lines and tumors from genetically engineered mouse models harboring an oncogenic mutant KRAS allele demonstrate a strong dependence on KRAS for proliferation and survival. This KRAS dependency is a type of ‘oncogene addiction,’ a state in which cancer cells depend on signaling from a single oncogene for survival. Unfortunately, the development of clinically effective KRAS-directed cancer therapies has been unsuccessful, and KRAS-mutant cancers are refractory to standard and targeted therapies. Alternative approaches to combatting KRAS-mutant cancers are clearly needed. We postulate that oncogenic KRAS signaling induces changes in cell signaling networks that cause cells to become dependent on certain genes, termed a ‘synthetic lethal’ interaction. Identifying these selective vulnerabilities would lend insight to the pathways altered in KRAS-mutant cancers and may inform novel strategies to target KRAS-addicted cancers. In this thesis, we systematically identify candidate co-dependencies of oncogenic KRAS by analyzing genetic dependencies revealed by genome-scale RNAi screens across a large panel of cell lines. We highlight methods to facilitate candidate selection/validation and integrate analyses of gene-expression data and genome-scale CRISPR/Cas9 screens to nominate candidate co-dependencies for further study. In addition, we examine CRISPR-Cas9 screens to identify genes that modify sensitivity to small molecule MAPK pathway inhibition (MAPKi) in RAS-mutant cancers. We propose that suppression of the DOCK5-RAC1 pathway demonstrates a drug-conditional lethal interaction with small molecule MAPK pathway inhibitors in RAS-mutant cancers. We believe that these data provide a foundation for further examination of genetic co-dependencies of oncogenic KRAS and the potential synthetic lethal interaction between DOCK5-RAC1 pathway suppression and MAPKi in RAS mutant cancers.