Fig. 2: The immune transcriptomic landscape of immune checkpoint blockade (ICB) resistant melanoma.

A Heatmap of melanoma module- and immune pathway transcriptomic scores^40,41,86,87. Patient samples are divided according to anti-CTLA4 resistant (CTLA4res, n = 17), anti-PD1 resistant (PD1res, n = 10) and anti-PD1 resistant biopsies taken at day 7 during BRAFi treatment (PD1res*, n = 10). *P < 0.05 between anti-PD1 without and under BRAFi treatment. ^P < 0.05 between anti-PD1 without BRAFi and anti-CTLA4 resistant lesions. T.accum – accumulated T cell score, T.exhaust – exhausted T cell score, T.regulatory – regulatory T cell score, Ipi.resist – signature score associated with anti-CTLA4 resistance, T.exhaust.fixed – exhausted/fixed T cell score. P-values from t-test. All tests were two-sided. Exact p-values are provided in Source data. Three mucosal melanomas are displayed in heatmap but excluded from statistical analyses. B Top ten pathways from gene set enrichment analysis of genes differentiating anti-CTLA4 from anti-PD1 resistant melanomas. This analysis was conducted on genes derived from the DESeq2 analysis. P-values from gene set enrichment analysis and adjusted for multiple testing. Red = enriched in anti-CTLA4 group, blue = enriched in anti-PD1 group. C Fraction of CD3⁺ cells as determined by multiplex immunofluorescence in n = 10 CTLA4res, n = 8 PD1res and n = 6 PD1res* tumors. Representative images are shown. Scalebar is indicated by white line and corresponds to 100 μm. P-value from Wilcoxon test. Test was two-sided. D Boxplots of T cell receptor (TCR) clonality data in n = 9 CTLA4res, n = 9 PD1res and n = 11 PD1res* tumors. Left boxplot shows the clonotype count (Efron Thisted). P-value from Kruskal-Wallis text. Right boxplot shows the evenness according to normalized Shannon Wiener index. P-value from Wilcoxon test. E Spatial gene expression data of tumor cell regions using Nanostring GeoMx, normalized for SOX10 expression. Volcano plot showing differentially expressed genes between n = 10 anti-CTLA4 and n = 7 anti-PD1 resistant tumors. Boxplot of PMEL expression between the two groups. P-values from t-test. Test was two-sided. Boxplots are displayed with the center-line as median, the box limits as lower and upper quartiles, and with whiskers covering the most extreme values within 1.5 x Interquartile-Range. Source data and exact p values are provided as a Source Data file.