To study the contribution of T-cell receptors (TCR) to resulting T-cell responses, we studied three different human aß TCRs, reactive to the same gp100-derived peptide presented in the context of HLA-A*0201. When expressed in primary CD8 T cells, all receptors elicited classic antigen-induced IFN-? responses, which correlated with TCR affinity for peptide-MHC in the order T4H2 > R6C12 > SILv44. However, SILv44 elicited superior IL-17A release. Importantly, in vivo, SILv44-transgenic T cells mediated superior antitumor responses to 888-A2 + human melanoma tumor cells upon adoptive transfer into tumor-challenged mice while maintaining IL-17 expression. Modeling of the TCR ternary complexes suggested architectural differences between SILv44 and the other complexes, providing a potential structural basis for the observed differences. Overall, the data reveal a more prominent role for the T-cell receptor in defining host T-cell physiology than traditionally assumed, while parameters beyond IFN-? secretion and TCR affinity ultimately determine the reactivity of tumor-reactive T cells.
Neoepitope-specific T-cell responses have been shown to induce durable clinical responses in patients with advanced cancers. We explored the recognition patterns of tumor-infiltrating T lymphocytes (TILs) from patients with glioblastoma multiforme (GBM), the most fatal form of tumors of the central nervous system. Whole-genome sequencing was used for generating DNA sequences representing the entire spectrum of 'private' somatic mutations in GBM tumors from five patients, followed by 15-mer peptide prediction and subsequent peptide synthesis. For each mutated peptide sequence, the wildtype sequence was also synthesized and individually co-cultured with autologous GBM TILs, which had been expanded in vitro with a combination of interleukin (IL)-2, IL-15 and IL-21. After seven days of culture, interferon gamma (IFN-?), tumor necrosis factor alpha (TNF-a) and/or IL-17A production was measured by ELISA in culture supernatants, and used as an epitope-specific immune response readout. Mutated peptides that induced a strong cytokine response were considered to contain legitimate neoepitopes. TILs from 5/5 patients with GBM exhibited specific immune reactivity profiles to the nominal target peptides, defined by IFN-? and/or TNF-a production, as well as IL-17A. Neoepitopes, defined by mutated peptides inducing IFN-? and/or TNF-a production without or only minimal reactivity to the wildtype sequences, were found for each individual patient. CD8+ TILs dominated the patients' responses to private neoepitopes. The present study shows that neoepitope-specific TIL reactivity constitutes an important arm of anti-tumor immune responses in patients with GBM, and thus a powerful tool for developing next-generation personalized immunotherapies.
