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. Author manuscript; available in PMC: 2024 Dec 11.
Published in final edited form as: Cancer Cell. 2023 Nov 30;41(12):2154–2165.e5. doi: 10.1016/j.ccell.2023.11.005

Figure 1. Tetramer enrichment and scRNAseq of circulating neoantigen-reactive CD8+ T cells.

Figure 1.

(A) Graphical pipeline summary of discovery, tetramer-enrichment and phenotypic analysis of neoantigen-reactive circulating CD8+ cells (prepared using BioRender.com). (B) Known neoantigen-reactive TCR CDR3β sequences, specificities, HLA of restriction, minimal epitope, and their frequencies in the pre-surgery blood of metastatic colon cancer patient (Pt. 4246). (C) CD137 expression of health-donor PBL TCR-transduced PBLs following overnight co-culture with 4246 dendritic cells (DCs) pulsed with various dilutions of their cognate minimal mutated or wild-type peptides. (D) Tetramer-enrichment sort of circulating neoantigen-reactive T cells spiked back into bulk CD8+ population at a 1:10 ratio. (E) UMAP projection of the single-cell transcriptome of Pt.4246 PBL. (F) Previously known neoantigen-reactive T cells (against ARMC9L146F or MYO5BK1410Q) highlighted in Pt.4246 PBL (G) Public viral-targeting TCRs highlighted (H) Healthy donor PBLs virally-transduced with candidate TCRs from cluster 7 (C7) stained with ARMC9L146F or MYO5BK1410Q fluorescent tetramers. (I) Highlighting all T cell clones expressing neoantigen-reactive TCRs. (J) Summary of enrichment of neoantigen-reactive clones in the pre-enrichment PBL, scRNA after enrichment, and specifically in cluster 7. (K) UMAP projection of scRNAseq of PIK3CAP449T:HLA-A*02:01-tetramer-enriched T cells from the peripheral blood sample of a metastatic cancer patient (Pt. 4317). (L) Expression of cell surface feature-barcoded CITE-Seq antibody staining intensities of protein markers. Also see Fig. S1–3 and Table S1.