CRISPR-Cas9 Mediated Efficient PD-1 Disruption on Human Primary T Cells From Cancer Patients

Sci Rep. 2016 Jan 28;6:20070. doi: 10.1038/srep20070.

Abstract

Strategies that enhance the function of T cells are critical for immunotherapy. One negative regulator of T-cell activity is ligand PD-L1, which is expressed on dentritic cells (DCs) or some tumor cells, and functions through binding of programmed death-1 (PD-1) receptor on activated T cells. Here we described for the first time a non-viral mediated approach to reprogram primary human T cells by disruption of PD-1. We showed that the gene knockout of PD-1 by electroporation of plasmids encoding sgRNA and Cas9 was technically feasible. The disruption of inhibitory checkpoint gene PD-1 resulted in significant reduction of PD-1 expression but didn't affect the viability of primary human T cells during the prolonged in vitro culture. Cellular immune response of the gene modified T cells was characterized by up-regulated IFN-γ production and enhanced cytotoxicity. These results suggest that we have demonstrated an approach for efficient checkpoint inhibitor disruption in T cells, providing a new strategy for targeting checkpoint inhibitors, which could potentialy be useful to improve the efficacy of T-cell based adoptive therapies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • CRISPR-Cas Systems*
  • Cells, Cultured
  • Cytokines / biosynthesis
  • Cytotoxicity, Immunologic
  • Gene Editing
  • Gene Knockout Techniques
  • Gene Order
  • Gene Targeting
  • Gene Transfer Techniques
  • Genetic Loci
  • Genetic Vectors / genetics
  • Humans
  • Lymphocyte Activation
  • Neoplasms / genetics*
  • Neoplasms / immunology
  • Neoplasms / metabolism
  • Neoplasms / pathology
  • Phenotype
  • Programmed Cell Death 1 Receptor / deficiency
  • Programmed Cell Death 1 Receptor / genetics*
  • RNA, Guide / genetics
  • T-Lymphocyte Subsets / immunology
  • T-Lymphocyte Subsets / metabolism*
  • Transfection

Substances

  • Cytokines
  • Programmed Cell Death 1 Receptor
  • RNA, Guide