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

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

Fingerprint

DNA Mismatch Repair
Genetic Engineering
Cultured Cells
Cells
Repair
Genetic Testing
DNA
Neoplasms
Clustered Regularly Interspaced Short Palindromic Repeats
Screening
Cell Line
Drug Discovery
Pharmaceutical Preparations
Ports and harbors
Clone Cells
Alleles
Genetics
Engineers
Molecules

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

Cite this

Engineering Forward Genetics into Cultured Cancer Cells for Chemical Target Identification. / Povedano, Juan Manuel; Liou, Joel; Wei, David; Srivatsav, Ashwin; Kim, Jiwoong; Xie, Yang; Nijhawan, Deepak; McFadden, David G.

In: Cell Chemical Biology, Vol. 26, No. 9, 19.09.2019, p. 1315-1321.e3.

Research output: Contribution to journalArticle

Povedano, Juan Manuel ; Liou, Joel ; Wei, David ; Srivatsav, Ashwin ; Kim, Jiwoong ; Xie, Yang ; Nijhawan, Deepak ; McFadden, David G. / Engineering Forward Genetics into Cultured Cancer Cells for Chemical Target Identification. In: Cell Chemical Biology. 2019 ; Vol. 26, No. 9. pp. 1315-1321.e3.
@article{ba511e0d1e2d46b28979086af1316cad,
title = "Engineering Forward Genetics into Cultured Cancer Cells for Chemical Target Identification",
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.",
keywords = "Ewing sarcoma, forward genetics, genetically-engineered mouse model (GEMM), hypermutation, mismatch repair, mismatch repair deficiency, phenotypic screening, target identification",
author = "Povedano, {Juan Manuel} and Joel Liou and David Wei and Ashwin Srivatsav and Jiwoong Kim and Yang Xie and Deepak Nijhawan and McFadden, {David G}",
year = "2019",
month = "9",
day = "19",
doi = "10.1016/j.chembiol.2019.06.006",
language = "English (US)",
volume = "26",
pages = "1315--1321.e3",
journal = "Cell Chemical Biology",
issn = "2451-9448",
publisher = "Elsevier Inc.",
number = "9",

}

TY - JOUR

T1 - Engineering Forward Genetics into Cultured Cancer Cells for Chemical Target Identification

AU - Povedano, Juan Manuel

AU - Liou, Joel

AU - Wei, David

AU - Srivatsav, Ashwin

AU - Kim, Jiwoong

AU - Xie, Yang

AU - Nijhawan, Deepak

AU - McFadden, David G

PY - 2019/9/19

Y1 - 2019/9/19

N2 - 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.

AB - 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.

KW - Ewing sarcoma

KW - forward genetics

KW - genetically-engineered mouse model (GEMM)

KW - hypermutation

KW - mismatch repair

KW - mismatch repair deficiency

KW - phenotypic screening

KW - target identification

UR - http://www.scopus.com/inward/record.url?scp=85072179584&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85072179584&partnerID=8YFLogxK

U2 - 10.1016/j.chembiol.2019.06.006

DO - 10.1016/j.chembiol.2019.06.006

M3 - Article

C2 - 31303577

AN - SCOPUS:85072179584

VL - 26

SP - 1315-1321.e3

JO - Cell Chemical Biology

JF - Cell Chemical Biology

SN - 2451-9448

IS - 9

ER -