Engineering Forward Genetics into Cultured Cancer Cells for Chemical Target Identification

Juan Manuel Povedano, Joel Liou, David Wei, Ashwin Srivatsav, Jiwoong Kim, Yang Xie, Deepak Nijhawan, David G McFadden

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

Target identification for biologically active small molecules remains a major barrier for drug discovery. Cancer cells exhibiting defective DNA mismatch repair (dMMR) have been used as a forward genetics system to uncover compound targets. However, this approach has been limited by the dearth of cancer cell lines that harbor naturally arising dMMR. Here, we establish a platform for forward genetic screening using CRISPR/Cas9 to engineer dMMR into mammalian cells. We demonstrate the utility of this approach to identify mechanisms of drug action in mouse and human cancer cell lines using in vitro selections against three cellular toxins. In each screen, compound-resistant alleles emerged in drug-resistant clones, supporting the notion that engineered dMMR enables forward genetic screening in mammalian cells.

Original languageEnglish (US)
Pages (from-to)1315-1321.e3
JournalCell Chemical Biology
Volume26
Issue number9
DOIs
StatePublished - Sep 19 2019

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Keywords

  • Ewing sarcoma
  • forward genetics
  • genetically-engineered mouse model (GEMM)
  • hypermutation
  • mismatch repair
  • mismatch repair deficiency
  • phenotypic screening
  • target identification

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmacology
  • Drug Discovery
  • Clinical Biochemistry

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