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Performing an In Vitro Genome-Wide CRISPR Knockout Screen in Chimeric Antigen Receptor T Cells
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In vivo profiling of metastatic double knockouts through CRISPR-Cpf1 screens

Ryan D Chow1,2,3,4, Guangchuan Wang1,2,3, Lupeng Ye1,2,3

  • 1Department of Genetics, Yale University School of Medicine, New Haven, CT, USA.

Nature Methods
|April 10, 2019
PubMed

Insights

Investigating genetic drivers of cancer metastasis is complex. We developed a new CRISPR screening method (MCAP) to efficiently map gene interactions influencing metastatic potential in vivo, revealing key genetic networks.

Area of Science:

  • Genetics
  • Cancer Biology
  • Molecular Biology

Background:

  • Understanding genetic interactions driving cancer metastasis is crucial but challenging.
  • Previous methods limited systematic, in vivo investigation of double gene knockouts.

Purpose of the Study:

  • To develop and apply a novel high-throughput screening method for interrogating genetic interactions in metastasis.
  • To create a quantitative map of gene interactions influencing metastatic potential.

Main Methods:

  • Developed massively parallel CRISPR-Cpf1/Cas12a crRNA array profiling (MCAP) for combinatorial double knockout screening.
  • Designed an MCAP library targeting 325 pairwise combinations of metastasis-implicated genes.
  • Assessed metastatic potential of double knockouts in vivo in mice.

Main Results:

  • Successfully implemented MCAP for large-scale genetic interaction profiling.
  • Unveiled a quantitative landscape of genetic interactions that significantly impact metastatic potential.
  • Identified specific gene pairs whose combined disruption alters cancer spread.

Conclusions:

  • MCAP is a powerful tool for dissecting complex genetic architectures of metastasis.
  • The identified genetic interactions provide new insights into metastasis mechanisms.
  • This approach can accelerate the discovery of therapeutic targets for metastatic cancer.

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