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Nat Chem Biol. 2016 May;12(5):361-6. doi: 10.1038/nchembio.2050. Epub 2016 Mar 28.

Parallel shRNA and CRISPR-Cas9 screens enable antiviral drug target identification.

Author information

1
Department of Chemistry, Stanford University, Stanford, California, USA.
2
Department of Genetics, Stanford University, Stanford, California, USA.
3
Department of Microbiology and Immunology, Stanford University, Stanford, California, USA.
4
Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biomedical Research and Howard Hughes Medical Institute, San Francisco, California, USA.
5
Stanford University Chemistry, Engineering, and Medicine for Human Health (ChEM-H), Stanford, California, USA.
6
Division of Gastroenterology and Hepatology, Stanford University School of Medicine, Stanford, California, USA.
7
Department of Chemical Engineering, Stanford University, Stanford, California, USA.
8
Department of Biochemistry, Stanford University, Stanford, California, USA.

Abstract

Broad-spectrum antiviral drugs targeting host processes could potentially treat a wide range of viruses while reducing the likelihood of emergent resistance. Despite great promise as therapeutics, such drugs remain largely elusive. Here we used parallel genome-wide high-coverage short hairpin RNA (shRNA) and clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 screens to identify the cellular target and mechanism of action of GSK983, a potent broad-spectrum antiviral with unexplained cytotoxicity. We found that GSK983 blocked cell proliferation and dengue virus replication by inhibiting the pyrimidine biosynthesis enzyme dihydroorotate dehydrogenase (DHODH). Guided by mechanistic insights from both genomic screens, we found that exogenous deoxycytidine markedly reduced GSK983 cytotoxicity but not antiviral activity, providing an attractive new approach to improve the therapeutic window of DHODH inhibitors against RNA viruses. Our results highlight the distinct advantages and limitations of each screening method for identifying drug targets, and demonstrate the utility of parallel knockdown and knockout screens for comprehensive probing of drug activity.

PMID:
27018887
PMCID:
PMC4836973
DOI:
10.1038/nchembio.2050
[Indexed for MEDLINE]
Free PMC Article

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