Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes

Unraveling the mechanism of action and molecular target of small molecules remains a major challenge in drug discovery. While many cancer drugs target genetic vulnerabilities, loss-of-function screens fail to identify essential genes in drug mechanism of action. Here, we report CRISPRres, a CRISPR-C...

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Bibliographic Details
Published in:	Nature communications Vol. 9; no. 1; pp. 502 - 14
Main Authors:	Neggers, Jasper Edgar, Kwanten, Bert, Dierckx, Tim, Noguchi, Hiroki, Voet, Arnout, Bral, Lotte, Minner, Kristien, Massant, Bob, Kint, Nicolas, Delforge, Michel, Vercruysse, Thomas, Baloglu, Erkan, Senapedis, William, Jacquemyn, Maarten, Daelemans, Dirk
Format:	Journal Article
Language:	English
Published:	London Nature Publishing Group UK 05-02-2018 Nature Publishing Group Nature Portfolio
Subjects:	631/154/555 631/337/4041/3196 631/92/556 Acrylamides - pharmacology Aminopyridines - pharmacology Anticancer properties Antineoplastic Agents - pharmacology Cancer Cell Line Cell Survival - drug effects Cell Survival - genetics CRISPR CRISPR-Cas Systems Drug discovery Drug resistance Drug Resistance - genetics Drugs Genes Genes, Essential - genetics Genetic diversity Genetic screening HCT116 Cells HL-60 Cells Homology Humanities and Social Sciences Humans K562 Cells Molecular Targeted Therapy - methods multidisciplinary Mutagenesis Mutagenesis, Site-Directed - methods Mutation Nicotinamide Nicotinamide phosphoribosyltransferase Nicotinamide Phosphoribosyltransferase - antagonists & inhibitors Nicotinamide Phosphoribosyltransferase - genetics Non-homologous end joining Phosphoribosyltransferase Positive selection Science Science (multidisciplinary) Small Molecule Libraries - pharmacology Target recognition Tiling
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Summary:	Unraveling the mechanism of action and molecular target of small molecules remains a major challenge in drug discovery. While many cancer drugs target genetic vulnerabilities, loss-of-function screens fail to identify essential genes in drug mechanism of action. Here, we report CRISPRres, a CRISPR-Cas-based genetic screening approach to rapidly derive and identify drug resistance mutations in essential genes. It exploits the local genetic variation created by CRISPR-Cas-induced non-homologous end-joining (NHEJ) repair to generate a wide variety of functional in-frame mutations. Using large sgRNA tiling libraries and known drug–target pairs, we validate it as a target identification approach. We apply CRISPRres to the anticancer agent KPT-9274 and identify nicotinamide phosphoribosyltransferase (NAMPT) as its main target. These results present a powerful and simple genetic approach to create many protein variants that, in combination with positive selection, can be applied to reveal the cellular target of small-molecule inhibitors. Cancer therapy drugs are designed to target genetic vulnerabilities, but loss-of-function screens often fail to identify essential genes in drug mechanism studies. Here the authors demonstrate CRISPRres, which exploits in-frame variation generated by indel formation to discover gene-drug interactions.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	2041-1723 2041-1723
DOI:	10.1038/s41467-017-02349-8