Identifying and targeting collateral lethal vulnerabilities in cancers

识别和定位癌症中的侧枝致死弱点 摘要/摘要 针对主要抑癌基因的基因组缺失通常包括相邻的乘客基因， 编码细胞基本的内务功能。这些癌细胞由于功能上的共表达而存活下来。 驻留在基因组的非缺失区域的多余的副猪。因此，肿瘤中的这些“侧枝缺失” 抑制子基因可以通过靶向消除剩余的基因来赋予癌细胞特异性的脆弱性 副对数。我们的“侧枝致命性”的概念首先在GBM中被证明，并删除了1p36肿瘤。 围绕ENO1的抑制子基因座，导致对ENO2耗竭或药理学的高度敏感性 抑制(穆勒等人2012年)。随后，我们证明了线粒体苹果酸酶2的缺失 Smad4基因的(Me2)在shRNA介导的剩余线粒体耗尽时产生致死性 由ME3 Paralog编码的苹果酸酶活性(Dey等人2017年)。系统而全面地 确定癌症的侧枝致死靶点，我们首先分析了布罗德研究所的癌症依赖图 (Depmap)，这是一个公众可访问的数据库，存储着来自一个集合基因组的17386个基因的重要性分数- 规模CRISPR基因敲除研究在1054个不同谱系的癌细胞系中进行(Dempster等人。2019年； 甘地等人。2019年)。计算工作产生了多个抵押品致命候选对，包括 REEP3/4、PTDSS1/2、INTS6/INTS6L、PRPS1/2、LDHA/B和CSTF2/CSTF2T 一种回归细胞系敏感载体对全基因组CCLE表达和拷贝数的方法 用于预测基于平行对数耗尽的灵敏度的数据。而对DepMap数据的计算分析可以识别 一些候选抵押品致命对，许多抵押品缺失的小样本数量阻碍了健壮性 结论。在这些计算方法的基础上，这里我们将应用CRISPR/Cas12a多基因 基因敲除平台，用于系统地识别锚定在针对共同基因的缺失事件中的附属致死对 抑癌基因TP53、CDKN2A/B、ARID1A、PTEN、Smad4、RB1和NF1在肿瘤中的表达 类型。这一概念和实验框架，再加上细胞/类器官/肿瘤验证平台， 寻求识别并严格验证可随后导入药物的附带致命靶点集 发现流水线，目标是扩大癌症的精确治疗。

Identifying and targeting collateral lethal vulnerabilities in cancers Abstract/Summary Genomic deletions targeting major tumor suppressor genes frequently include adjacent passenger genes, encoding cell essential housekeeping functions. These cancer cells survive due to co-expressing functionally redundant paralogs residing in non-deleted regions of the genome. As such, these “collateral deletions” in tumor suppressor loci can confer cancer cell-specific vulnerabilities through targeted extinction of the remaining paralog. Our “collateral lethality” concept was first demonstrated in GBM with deletion of the 1p36 tumor suppressor locus encompassing ENO1, resulting in profound sensitivity to ENO2 depletion or pharmacologic inhibition (Muller et al. 2012). Subsequently, we demonstrated that deletion of mitochondrial malic enzyme 2 (ME2) in the SMAD4 locus engendered lethality upon shRNA-mediated depletion of the remaining mitochondrial malic enzyme activity encoded by the ME3 paralog (Dey et al. 2017). To systematically and comprehensively identify collateral lethal targets in cancer, we first analyzed the Broad Institute’s Cancer Dependency Map (DepMap), a publicly accessible database hosting essentiality scores of 17386 genes from a pooled genome- scale CRISPR knockout study conducted in 1054 cancer cell lines of diverse lineages (Dempster et al. 2019; Ghandi et al. 2019). The computational efforts yielded multiple collateral lethal candidate pairs including REEP3/4, PTDSS1/2, INTS6/INTS6L, PRPS1/2, LDHA/B, and CSTF2/CSTF2T via a two-class comparison method that regressed cell line sensitivity vectors against whole-genome CCLE expression and copy number data to predict paralog-depletion based sensitivity. While computational analysis of DepMap data can identify some candidate collateral lethal pairs, small sample sizes for many collateral deletions have stymied robust conclusions. Building on these computational methods, here we will apply the CRISPR/Cas12a polygenic knockout platform to systematically identify collateral lethal pairs anchored in deletion events targeting common tumor suppressor gene loci such as TP53, CDKN2A/B, ARID1A, PTEN, SMAD4, RB1 and NF1 across cancer types. This conceptual and experimental framework, coupled with a cell/organoid/tumor validation platform, seeks to identify and stringently validate collateral lethal target sets that can then be channeled into a drug discovery pipeline with the goal of expanding precision cancer treatments.