Fig. 3 |. Increase in T cell infiltration evident in patients with circulating neoantigen-specific T cell responses.

a, Multiplex immunofluorescence staining of five pairs of initial and relapse tumour specimens reveals increased T cell infiltration in patients who did not receive dexamethasone during vaccine priming (red) compared to those who did (blue), with representative sections shown. Sections show SOX2 (magenta) to stain tumour cells, CD4 (yellow) and CD8 (green). Infiltrates were determined by enumerating the mean number of CD4⁺ and CD8⁺ cells in 20× fields. The number of fields evaluated per sample was: 4 fields for initial and relapse samples of patients 3 and 8, initial samples of patient 4 and relapse samples of patient 7; and 5 fields for relapse samples of patient 4, initial and relapse samples of patient 5 and initial samples of patient 7. Data are mean ± s.e.m. P values reported are two-sided and based on model F-tests. See Methods for statistical analysis. b, Each dot represents a single T cell, and cells with identical TCRs are clustered together. Two dominant SHANK2 neoantigen-reactive TCRαβ clones (dark and light blue) were identified in T cell lines that were generated from Week 16 post-vaccination PBMCs from patient 8 using an IFNγ secretion assay. These clones were detected among post-vaccination (relapse) but not pre-vaccination (initial) tumour-associated TCRαβ sequences using bulk RNA-seq. Two dominant SVEP1-reactive TCRαβ (red and orange) clones were observed in post-vaccination PBMCs but not in brain tumour biopsies. c, Enumeration of the numbers of single CD4⁺ and CD8⁺ tumour-associated T cells that expressed effector cytokines in patient 7, analysed using scRNA-seq. d, CD4⁺ and CD8⁺ tumour-associated T cells in patient 7 expressed multiple co-inhibitory receptors based on scRNA-seq.