Fig. 1: Identification of neoTCRs via scTCR-seq of CD8⁺ T cells from melanoma patient Mel15.

a Schematic experiment setting. b Increase in total number of cells of known (KIF-P1 and -P2, SYT-T1, -T2, -P1, -P2) and additionally identified (KIF-sc1 and -sc2) TCRs upon antigen-specific stimulation and CD137-enrichment with dominance of KIF-P1 and -P2 harboring high precursor frequency. The two additionally identified KIF2C-TCRs were selected based on fold change of TCR frequency and highest absolute frequency in the stimulated sample. c Assessment of antigen-specific IFN-γ-secretion for the two identified TCRs KIF-sc1 and -sc2 in comparison to the known TCR KIF-P2. Cytokine secretion was measured by IFN-γ-ELISA upon 24 h of co-culture of TCR-tg T cells from one representative donor with Mel15-LCL transgenic for the mutated KIF2C^P13L minigene (mut mg) and the wildtype KIF2C minigene (wt mg) as well as pulsed for 2 h at 37 °C with the mutated and wildtype peptide (mut pep and wt pep). An irrelevant peptide (irr peptide), target cells (LCL only) or T cells alone (T cell only) served as negative controls. Mean and SD of technical triplicates depicted. Data representative for two different donors. d Frequency of KIF-sc1 and -sc2 in relation to the previously identified TCR-sequences³¹ identified by deep sequencing of the TCR-β-chain in intestinal (M_Int) and lung metastases (M_Lung) as well as corresponding non-malignant draining lymph nodes (M_Int-LN1, M_Int-LN2 and M_Lung-LN) of patient Mel15. Non-td non-transduced.