Extended Data Fig. 8 |. Activation, cytotoxicity, cytokine secretion, and proliferation induced by neoantigen-specific TCRs from patient 6 upon co-culture with the autologous cell line.

Healthy donor T cells genetically engineered to express the captured neoTCRs from patient 6 were co-cultured with the autologous (M490) or a mismatched cell line (M202). a, 4-1BB and OX-40 upregulation in the CD8⁺ neoTCR⁺ T cells after co-culture. Melanoma cell lines were pre-treated with regular media or media with IFNγ 24 h prior co-culture with T cells (n = 3). b, Specific target-cell killing in the autologous cell line (top panel) or a mismatched cell line (bottom panel), (P:T ratio 10:1, n = 4). c, Cytokine release at 24 h after co-culture (n = 3). Melanoma cell lines were pre-treated with IFNγ for 24 h before co-culture with T cells. d, Proliferation of CD8⁺ neoTCR⁺ T cells measured by Ki67 mean fluorescence intensity upon 24, 48 and 72 h co-culture with autologous melanoma cell line (M490, top panel) or a mismatched cell line (M202, bottom panel). Melanoma cell lines were pretreated with IFNγ 24 h prior co-culture with T cells (n = 3). *p < 0.05, **p < 0.005, ***p < 0.0005, ****p < 0.0001 vs Neo12, two-tailed unpaired t test with Holm-Sidak adjustment for multiple comparisons in figure a, b and c. *p < 0.05, **p < 0.005, ***p < 0.0005, ****p < 0.0001 vs M202, two-tailed unpaired t test with Holm-Sidak adjustment for multiple comparisons in figure d. (n) indicates the number of biological replicates. Exact p-values provided in Supplementary Information. Mean ± SD and individual values are shown in a, c and d. Mean and individual values are shown in b. All T-cell products contain CD8⁺ and CD4⁺ gene-edited T cells.