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. Author manuscript; available in PMC: 2021 Jul 12.
Published in final edited form as: Nat Med. 2021 Jan 21;27(3):515–525. doi: 10.1038/s41591-020-01206-4

Extended Data Figure 2. Vaccination induces strong multi-functional CD4+ T cell responses in two additional patients with high-risk melanoma.

Extended Data Figure 2.

a. Exemplary schema of immunizing (IMP), assay (ASP), and epitope (EPT) peptides. Mutated amino acid is shaded. Each predicted epitope peptide is in colored text and its location is underlined below the IMP peptide. b. PBMCs collected pre- and post- vaccination were tested with individual peptides after one round of in vitro stimulation by IFN-γ ELISpot using week 16 post-vaccination PBMCs, tested in triplicate wells per peptide (error bars, s.e.m.). Only results from positive peptides are shown. c. Deconvolution of T cell reactivity against individual ASP after one round of in vitro stimulation by IFN-γ ELISpot using week 16 post-vaccination PBMCs, tested in triplicate wells per peptide (error bars, s.e.m.). *Responses were scored positive if spot-forming cells (SFC) were at least 2.5-fold over the DMSO control. d. Dual chromogenic immunohistochemical staining of FFPE tumor samples from Pts. 11 and 12 (see Methods for details) for HLA class I and HLA class II. Red: SOX10 (melanoma transcription factor); brown: HLA class I or class II. Representative images of 5 to 11 fields of view (median 7) are shown. e. Summary of immunohistochemical results of seven patients with available FFPE tissue. Semi-quantitative scoring was performed for the intensity of positive staining of melanoma cell membranes for class I or II (0, negative; 1, weak; 2, moderate; 3, strong) and for the percentage of positive staining malignant cells (0–100%). A cumulative H score was obtained by multiplying intensity score by the percentage of malignant cells with positive staining.