Neoantigen-specific T cell reactivity in MMR-p CRC. a IFN-γ production of expanded TIL in response to synthetic long peptides (SLP) and synthetic short peptides (SSP), potential neo-epitopes in red and non-recognized peptides in black. SEB (gray) and DMSO (white) were taken along as positive and negative controls, respectively. Peptide IDs are included for neo-epitope responses that were judged positive and selected for validation. SSP and SLP with the same ID number correspond to the same mutation per patient. b IFN-γ production of TIL upon co-culture with mutant (red) and corresponding wild type (gray) peptides, and a DMSO control (dashed), at different peptide concentrations. The mean ± standard deviation of the biological duplicates in the same experiment are depicted. An asterisk indicates a significant difference (α = 0.0026) between wild type and mutant peptides. c Granzyme B production by TIL upon stimulation with autologous tumor fragments (red). TIL only (white) and tumor only (blue) conditions were taken along as negative controls, and SEB (gray) as positive control. Differential production between TIL + tumor and TIL or tumor only is analyzed by ANOVA; the asterisks indicate significant differences. d Gene expression measured by qPCR upon co-culture of different target/effector combinations of NIC3 (red), NIC4 (blue), and NIC15 (green). Differential gene expression upon co-culture with wild type and mutant peptides is indicated with an asterisk.