Fig. 3

Dysfunctional T cells correlate with Macro-2. A UMAP embedding demonstrating T cell subpopulations. B Visualization of the average cell density across Bone Met (n = 9) and multiple control conditions (Healthy n = 12, Benign n = 7, Involved n = 4, Distal n = 4), using embedding density estimates. Brighter colors correspond to denser regions (see the “Methods” section). C Expression of representative T cell exhaustion markers on UMAP embedding. D Box plots showing T cell exhaustion score within CTL-3 across Bone Met (n = 9) and control conditions (Healthy n = 12, Benign n = 7, Involved n = 4, Distal n = 4). Statistics are accessed with two-sided Wilcoxon rank sum test and BH multiple testing correction (*p < 0.05). For box plots, center line represents the median and box limits represent upper and lower quartiles, and whiskers depict 1.5 × the interquartile range (IQR). E Comparison of PDCD1 expression (MFI) in Distal (n = 4) and Bone Met (n = 4) samples. Statistical significance determined using two-sided t-test (*p < 0.05). F ICOS, CTLA4, TNFRSF4, and TNFRSF18 expression in Tregs shown as violin plot. G Bar plot showing CTL-3 (top) and Treg abundance (bottom) comparing RCC Bone Met (n = 9) with RCC Primary (n = 14) and adjacent normal (n = 9) fractions. Statistics are accessed with two-sided Wilcoxon rank sum test (*p < 0.05, **p < 0.01). H Violin plot showing representative exhausted T cell signature gene expression in CTL-3 comparing RCC Bone Met with RCC Primary and adjacent normal fractions. I Correlations of the cell abundance between myeloid and T cell subsets shown as heatmap. Significance was assessed using Pearson correlation test and BH multiple testing correction. Color represents correlation coefficient and star presents the significance. (*p < 0.05). J Heatmap showing scaled average expression of CCL18 and CCR8 in major cell populations. K Circle plots showing the inferred CCL18-CCR8 signaling between Macro-2 and Treg. L Box plot showing CCL18 and CCR8 abundance in tumor (n = 72) compared to adjacent normal (n = 533) tissue in TCGA KIRC. Statistics are accessed with two-sided Wilcoxon rank sum test (****p < 0.0001). M Correlation of CCR8 expression in Tregs and CTL-3 exhaustion score in CTL-3 is shown as a scatter plot. Pearson linear correlation estimate, and p-values are shown. The error band indicates 95% confidence interval. N Correlation of CCR8 expression and CTL-3 exhaustion score is shown as a scatter plot for TCGA KIRC data (n = 533). Pearson linear correlation estimate, and p-values are shown