Uncovering Hidden Cancer Self-Dependencies through Analysis of shRNA-Level Dependency Scores

Abstract

Large-scale short hairpin RNA (shRNA) screens on well-characterized human cancer cell lines have been widely used to identify novel cancer dependencies. However, the off-target effects of shRNA reagents pose a significant challenge in the analysis of these screens. To mitigate these off-target effects, various approaches have been proposed that aggregate different shRNA viability scores targeting a gene into a single gene-level viability score. Most computational methods for discovering cancer dependencies rely on these gene-level scores. In this paper, we propose a computational method, named NBDep, to find cancer self-dependencies by directly analyzing shRNA-level dependency scores instead of gene-level scores. The NBDep algorithm begins by removing known batch effects of the shRNAs and selecting a subset of concordant shRNAs for each gene. It then uses negative binomial random effects models to statistically assess the dependency between genetic alterations and the viabilities of cell lines by incorporating all shRNA dependency scores of each gene into the model. We applied NBDep to the shRNA dependency scores available at Project DRIVE, which covers 26 different types of cancer. The proposed method identified more well-known and putative cancer genes compared to alternative gene-level approaches in pan-cancer and cancer-specific analyses. Additionally, we demonstrated that NBDep controls type-I error and outperforms statistical tests based on gene-level scores in simulation studies. Author SummaryLarge-scale shRNA screening is increasingly being used in cancer genomics to discover genes involved in cancer by analyzing the viabilities of cell lines upon knocking down a gene using a pool of short hairpin RNAs (shRNA). However, off-target effects, which result from the knockdown of unintended genes, are a major issue in RNAi screening. To address this issue, various computational methods have been developed to aggregate shRNA viability scores into gene-level dependency scores. In this paper, we propose a method called NBDep to identify cancer gene drivers that directly addresses the challenge of off-target effects at the shRNA level. NBDep identifies cancer gene drivers in three classes: amplification, missense, and non-missense alterations. In this method, we first remove known batch effects, select a subset of the most consistent shRNAs of each gene, and then perform a negative binomial mixed-effect model. The NBDep method not only identifies well-recognized and novel cancer driver genes but also has more statistical power than gene-level-score methods while controlling type-error. In summary, NBDep presents a new technique for analyzing shRNA screens and has the potential to uncover previously unknown cancer dependencies.