Abstract
To develop a practically applicable method for T-cell receptor (TCR)-engineered T cell immunotherapy targeting neoantigens, we have been attempting to identify neoantigen-specific T cell receptors (TCRs) and establish TCR-engineered T cells in a 3-4-month period. In this study, we report the characterization of T cell repertoires in tumor microenvironment (TME) and identification of neoantigen-specific TCRs after stimulation of patient-derived T cells. We screened 15 potential neoantigen peptides and successfully identified two CD8+HLA-dextramer+ T cells, which recognized MAGOHBG17A and ZCCHC14P368L. All three dominant TCR clonotypes from MAGOHBG17A-HLA dextramer-sorted CD8+ T cells were also found in T cells in TME, while none of dominant TCR clonotypes from ZCCHC14P368L-HLA dextramer-sorted CD8+ T cells was found in the corresponding TME. The most dominant TCRA/TCRB pairs for these two neoantigens were cloned into HLA-matched healthy donors' T lymphocytes to generate TCR-engineered T cells. The functional assay showed MAGOHBG17A TCR-engineered T cells could be significantly activated in a mutation-specific, HLA-restricted and peptide-dose-dependent manner while ZCCHC14P368L TCR-engineered T cells could not. Our data showed neoantigen-reactive T cell clonotypes that were identified in the patient's peripheral blood could be present in the corresponding TME and might be good TCRs targeting neoantigens.