title: Stable, Nonviral Expression of Mutated Tumor Neoantigen-specific T-cell Receptors Using the Sleeping Beauty Transposon/Transposase System.
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.
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Drew C
authors.initials: DC
authors.lastname: Deniger
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Anna
authors.initials: A
authors.lastname: Pasetto
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Tran
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Maria R
authors.initials: MR
authors.lastname: Parkhurst
authors.affiliation: Tumor Immunology and Immunotherapy, Bar-Ilan University, Ramat Gan, Israel.
authors.forename: Cyrille J
authors.initials: CJ
authors.lastname: Cohen
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Paul F
authors.initials: PF
authors.lastname: Robbins
authors.affiliation: Division of Pediatrics, University of Texas M.D. Anderson Cancer Center, Houston, Texas, USA; ZIOPHARM Oncology, Inc., Boston, Massachusetts, USA.
authors.forename: Laurence Jn
authors.initials: LJ
authors.lastname: Cooper
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA. Electronic address: sar@nih.gov.
authors.forename: Steven A
authors.initials: SA
authors.lastname: Rosenberg
mesh_terms: D000818:Animals; D004251:DNA Transposable Elements; D005818:Genetic Engineering; D015789:HLA-A2 Antigen / Q000378:metabolism; D059866:HLA-DQ beta-Chains / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000276:immunology; D051379:Mice; D009369:Neoplasms / Q000276:immunology*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D013601:T-Lymphocytes / Q000276:immunology / Q000473:pathology*; D019895:Transposases / Q000378:metabolism*
chemical_list: D004251:DNA Transposable Elements; D015789:HLA-A2 Antigen; D059866:HLA-DQ beta-Chains; D008565:Membrane Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases; D011948:Receptors, Antigen, T-Cell; D019895:Transposases; C435939:HLA-A*02:01 antigen; C057615:HLA-DQbeta antigen; C494471:sleeping beauty transposase, human