Inactivation of Cancer Mutations Utilizing CRISPR/Cas9

J Natl Cancer Inst. 2016 Aug 30;109(1):djw183. doi: 10.1093/jnci/djw183. Print 2017 Jan.

Abstract

Although whole-genome sequencing has uncovered a large number of mutations that drive tumorigenesis, functional ratification for most mutations remains sparse. Here, we present an approach to test functional relevance of tumor mutations employing CRISPR/Cas9. Combining comprehensive sgRNA design and an efficient reporter assay to nominate efficient and selective sgRNAs, we establish a pipeline to dissect roles of cancer mutations with potential applicability to personalized medicine and future therapeutic use.

MeSH terms

  • Bacterial Proteins* / genetics
  • Bacterial Proteins* / metabolism
  • CRISPR-Associated Protein 9
  • Carcinoma / genetics*
  • Clustered Regularly Interspaced Short Palindromic Repeats*
  • Colonic Neoplasms / genetics*
  • Computational Biology
  • DNA Cleavage
  • Endonucleases* / genetics
  • Endonucleases* / metabolism
  • Genes, Reporter
  • Green Fluorescent Proteins / genetics
  • HeLa Cells
  • Humans
  • Leukemia, Myeloid, Acute / genetics*
  • Mutation / genetics*
  • Nuclear Proteins / genetics
  • Nucleophosmin
  • Proto-Oncogene Proteins B-raf / genetics
  • RNA, Guide, CRISPR-Cas Systems / genetics*
  • Transfection

Substances

  • Bacterial Proteins
  • Nuclear Proteins
  • RNA, Guide, CRISPR-Cas Systems
  • Nucleophosmin
  • Green Fluorescent Proteins
  • BRAF protein, human
  • Proto-Oncogene Proteins B-raf
  • CRISPR-Associated Protein 9
  • Cas9 endonuclease Streptococcus pyogenes
  • Endonucleases