CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer

Fabrizio Simeoni; Ioannis Loukas; Thomas Stuart Wilson; Paola Scaffidi

doi:10.1016/j.xpro.2023.102097

CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer

STAR Protoc. 2023 Mar 17;4(1):102097. doi: 10.1016/j.xpro.2023.102097. Epub 2023 Feb 11.

Authors

Fabrizio Simeoni¹, Ioannis Loukas², Thomas Stuart Wilson², Paola Scaffidi³

Affiliations

¹ Cancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UK. Electronic address: fabrizio.simeoni@crick.ac.uk.
² Cancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UK.
³ Cancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UK; Department of Experimental Oncology, IEO, European Institute of Oncology IRCCS, 20139 Milan, Italy. Electronic address: paola.scaffidi@crick.ac.uk.

Abstract

Dissecting mechanisms driving subclone expansion in primary cancers has been challenging. Here, we present a protocol to systematically disrupt entire gene networks and assess the functional impact of this perturbation on cancer cell fitness. By combining arrayed CRISPR libraries and high-content microscopy, we describe steps to identify classes of genes whose inactivation promotes resistance to environmental challenges faced by cancer cells during tumor growth or upon therapy. A proof-of-principle interrogation of the epigenetic regulatory network is described. For complete details on the use and execution of this protocol, please refer to Loukas et al. (2022).¹.

Keywords: CRISPR; Cancer; Cell Biology; Cell-based Assays; Genetics; High-throughput Screening; Molecular Biology.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Epigenomics
Gene Regulatory Networks
Humans
Mutation / genetics
Neoplasms* / genetics

Abstract

Publication types

MeSH terms

Grants and funding