Figure 4.

Characterization of KRAS-specific TCRs on common alleles

(A–D) TCRs isolated from healthy donor T cells stimulated against different KRAS neoantigens were transduced into Jurkat^CD8 cells. TCR avidity was evaluated by titration of each cognate peptide presented on A375 cells transduced to express the cognate HLA-I molecule. TCR stimulation was assessed by IL-2 secretion or CD69 expression, and half-maximal effective concentration was calculated. (A) TCR specific for KRAS^G12V on HLA-A∗11:01. (B) TCR specific for KRAS^G12V on HLA-A∗03:01. (C) TCR specific for KRAS^G12D on HLA-A∗11:01. (D) TCR specific for KRAS^G12D on HLA-A∗03:01. Data are presented as mean ± SD and are representative of at least two independent experiments. See also Figure S5.

(E) The TCR from (A) was transduced into PBMCs and used in a cytotoxicity assay against SW620^A∗11:01 across a range of specific effector to target (E:T) ratios (red). The ratio was calculated using the number of TCR-transduced CD8+ T cells in the well. As a control, an equivalent number of PBMCs transduced with an irrelevant TCR was co-cultured with SW620^A11:01 cells (blue). Both target cell growth (top) and target cell death (bottom) were measured. Data are presented as mean ± SD and are representative of three independent experiments.

(F) The TCR from (B) was transduced into PBMCs and used in a cytotoxicity assay against SW620^A∗03:01 (red), and cell death of the target cell was measured. As a negative control, PBMCs transduced with the TCR from (A) were co-cultured with SW620^A∗03:01 (blue). Data are presented as mean ± SD and are representative of two independent experiments.

(G) The TCR from (B) was transduced into PBMCs and used in a cytotoxicity assay against NCI-H441 (red), and cell death of the target cell was measured. As a negative control, PBMCs transduced with the TCR from (A) were co-cultured with NCI-H441 (blue). Data are presented as mean ± SD and are representative of two independent experiments.