Figure 3.

Identification of neoantigen-specific TCRs from a metastatic tumor resected from patient M2 with melanoma. (A) T cells were stimulated by PP-pulsed autologous DCs for 4 hours, and then subjected to single-cell analysis. The expression of IFN-γ in each single cell was obtained by bioinformatic analysis. The detailed information of each IFN-γ⁺ single cell is listed in online supplemental tables 2 and 3. The TCR identified from individual single cells are labeled by numbers (online supplemental tables 1-3). Unique TCRs are not labeled. (B, C) Individual TCRs isolated from tumor M2 were transduced into autologous T cells. Purified 25-mer WT or mutated peptide at various concentrations was pulsed on autologous DCs for 24 hours, and peptide-pulsed DCs were cocultured with TCR-transduced T cells for 16 hours. The secretion of IFN-γ from T cells was determined by ELISA. The gene names of neoantigen corresponding to individual neoantigen-specific TCRs are shown in each panel. A, autoreactive TCRs; DCs, dendritic cells; IFN-γ, interferon-γ; PPs, peptide pools; TCRs, T-cell receptors; UMI, unique molecular identifier; WT, wild-type.