Abstract B39: Genomic copy number alterations introduce a gene-independent viability bias in CRISPR-Cas9 knock-out screens of cancer cell lines

Recent studies have demonstrated the power of CRISPR-Cas9 screening methods for identifying genetic vulnerabilities in cancer cells. As part of a larger effort to generate a comprehensive catalog of vulnerabilities, we performed CRISPR-Cas9 genome-scale loss-of-function screens in 33 cancer cell lines to identify genes essential for proliferation and survival. We found a strong correlation between gene copy number and cell viability after Cas9-targeting. Copy number alterations are extremely prevalent in human cancers and frequently lead to overexpression of driver oncogenes and potential vulnerabilities. Therefore, we sought to identify such genes by investigating the relationship of genomic copy number with essentiality from our screening data. As expected, known oncogenes scored as essential in cell lines harboring amplifications of these genes. However, the scores of all other genes in these amplified regions were also strongly enriched for apparent essentiality, even among unexpressed genes. Furthermore, the infection of cells with sgRNAs targeting Cas9 to non-coding intergenic sequences within regions of high copy number gain also induced this negative effect on cell viability. We observed this effect across multiple different chromosomal structural alterations, including tandem duplications, breakage-fusion-bridge structures, and arm-level gains. More broadly, we found a striking global correlation between cell viability in response to Cas9-targeting and the genomic copy number of the targeted site, even among low-level copy number gain and loss. For example, Cas9-targeting of genes with two copies resulted in, on average, decreased viability relative to Cas9-targeting of genes with only one copy. By examining sgRNAs that target multiple genomic sites, but not within any amplified loci, we found that this cell response to Cas9-targeting correlated strongly with the total number of target sites. Together, these observations indicate that genome targeting by CRISPR-Cas9 elicits a gene-independent anti-proliferative cell response with a severity proportional to the total number of discrete genomic loci targeted. This effect has important practical implications for interpretation of CRISPR-Cas9 screening data and confounds the use of this technology for identification of essential genes in amplified regions. This result illustrates the sensitivity of cancer cells to site-specific DNA damage, which may provide a path to novel therapeutic strategies. Targeting non-essential genes or non-coding intergenic sequences within regions of copy number amplification may reveal cancer-specific vulnerabilities. Citation Format: Robin M. Meyers, Andrew J. Aguirre, Barbara A. Weir, Francisca Vazquez, Cheng-Zhong Zhang, Uri Ben-David, April Cook, Gavin Ha, William F. Harrington, Mihir Doshi, Stanley Gill, Han Xu, Levi D. Ali, Guozhi Jiang, Sasha Pantel, Yenarae Lee, Amy Goodale, Andrew D. Cherniack, Coyin Oh, Gregory Kryukov, Glenn S. Cowley, Levi A. Garraway, Kimberly Stegmaier, Charles W. Roberts, Todd R. Golub, Matthew Meyerson, David E. Root, Aviad Tsherniak, William C. Hahn. Genomic copy number alterations introduce a gene-independent viability bias in CRISPR-Cas9 knock-out screens of cancer cell lines. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Targeting the Vulnerabilities of Cancer; May 16-19, 2016; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(1_Suppl):Abstract nr B39.