Synergistic drug combinations for cancer identified in a CRISPR screen for pairwise genetic interactions

Abstract

Identification of effective combination therapies is critical to address the emergence of drug-resistant cancers, but direct screening of all possible drug combinations is infeasible. Here we introduce a CRISPR-based double knockout (CDKO) system that improves the efficiency of combinatorial genetic screening using an effective strategy for cloning and sequencing paired single-guide RNA libraries and a robust statistical scoring method for calculating genetic interactions (GIs) from CRISPR-deleted gene pairs. We applied CDKO to generate a large-scale human GI map, comprising 490,000 double-sgRNAs directed against 21,321 pairs of drug targets in K562 leukemia cells and identified synthetic lethal drug target pairs for which corresponding drugs exhibit synergistic killing. These included the BCL2L1 and MCL1 combination, which was also effective in imatinib-resistant cells. We further validated this system by identifying known and previously unidentified GIs between modifiers of ricin toxicity. This work provides an effective strategy to screen synergistic drug combinations at high-throughput and a CRISPR-based tool to dissect functional GI networks.

Randomized CRISPR-Cas Transcriptional Perturbation Screening Reveals Protective Genes against Alpha-Synuclein Toxicity

“…In future work, assessing how different expression levels of identified genes modulate the phenotypes identified through PRISM screening will also be of interest. Finally, we envision that genetic interactions between genes identified by PRISM screening can be further mapped through combinatorial CRISPR technologies (Han et al, 2017; Shen et al, 2017; Wong et al, 2016) and both gene activation and inhibition technologies. Thus, high-throughput randomized crisprTF screens should provide access to a broader range of biological phenotypes across a wide range of organisms in the future.…”

Interrogation of Mammalian Protein Complex Structure, Function, and Membership Using Genome-Scale Fitness Screens

“…However, the generation of such datasets faces major limitations. In addition to the roughly 16-fold increase in combinatorial space of all possible double knockouts in human cells compared to yeast (due to the four-fold increase in genes), human screening libraries for simultaneously knocking out multiple genes of interest are relatively new and still face unsolved technical challenges (Boettcher et al, 2018; Du et al, 2017; Han et al, 2017; Najm et al, 2018; Shen et al, 2017). In contrast, genome-scale single-gene knockout libraries are technically more advanced and have been used extensively in pooled fitness screens (Doench, 2018).…”

CRISPR-based genetic interaction maps inform therapeutic strategies in cancer

“…In a recently published paper (19), Han and colleagues design a CRISPR-cutting based double knockout (CDKO) system to generate a large-scale mammalian GI map comprising of 21,321 drug combinations and successfully identified synthetic lethal drug target pairs in K562 chronic myeloid leukemia (CML) cells. The authors rationally designed their gene-interaction library based on the following criteria: (I) druggable genes; (II) expression level in K562 cells; (III) growth phenotype from previous screens.…”