Stable, Nonviral Expression of Mutated Tumor Neoantigen-specific T-cell Receptors Using the Sleeping Beauty Transposon/Transposase System

Mol Ther. 2016 Jun;24(6):1078-1089. doi: 10.1038/mt.2016.51. Epub 2016 Mar 5.

Abstract

Neoantigens unique to each patient's tumor can be recognized by autologous T cells through their T-cell receptor (TCR) but the low frequency and/or terminal differentiation of mutation-specific T cells in tumors can limit their utility as adoptive T-cell therapies. Transfer of TCR genes into younger T cells from peripheral blood with a high proliferative potential could obviate this problem. We generated a rapid, cost-effective strategy to genetically engineer cancer patient T cells with TCRs using the clinical Sleeping Beauty transposon/transposase system. Patient-specific TCRs reactive against HLA-A*0201-restriced neoantigens AHNAK(S2580F) or ERBB2(H473Y) or the HLA-DQB*0601-restricted neoantigen ERBB2IP(E805G) were assembled with murine constant chains and cloned into Sleeping Beauty transposons. Patient peripheral blood lymphocytes were coelectroporated with SB11 transposase and Sleeping Beauty transposon, and transposed T cells were enriched by sorting on murine TCRβ (mTCRβ) expression. Rapid expansion of mTCRβ(+) T cells with irradiated allogeneic peripheral blood lymphocytes feeders, OKT3, interleukin-2 (IL-2), IL-15, and IL-21 resulted in a preponderance of effector (CD27(-)CD45RA(-)) and less-differentiated (CD27(+)CD45RA(+)) T cells. Transposed T cells specifically mounted a polyfunctional response against cognate mutated neoantigens and tumor cell lines. Thus, Sleeping Beauty transposition of mutation-specific TCRs can facilitate the use of personalized T-cell therapy targeting unique neoantigens.

Publication types

  • Research Support, N.I.H., Intramural

MeSH terms

  • Animals
  • DNA Transposable Elements
  • Genetic Engineering
  • HLA-A2 Antigen / metabolism
  • HLA-DQ beta-Chains / metabolism
  • Humans
  • Membrane Proteins / immunology
  • Mice
  • Neoplasms / immunology*
  • Receptor, ErbB-2 / immunology
  • Receptors, Antigen, T-Cell / metabolism*
  • T-Lymphocytes / immunology
  • T-Lymphocytes / pathology*
  • Transposases / metabolism*

Substances

  • DNA Transposable Elements
  • HLA-A*02:01 antigen
  • HLA-A2 Antigen
  • HLA-DQ beta-Chains
  • HLA-DQbeta antigen
  • Membrane Proteins
  • Receptors, Antigen, T-Cell
  • Receptor, ErbB-2
  • Transposases
  • sleeping beauty transposase, human