Fig. 6: CD8⁺ TILs express NRP1 and are captured in SEMA3A rich areas in ccRCC tumors.

a Correlation of SEMA3A mRNA level with survival of ccRCC patients. b Schematic representing ccRCC patient cohort utilized in (c–m). c Representative flow cytometric analysis of CD8 and NRP1 expression in PBMC and TILs. d NRP1 expression on CD8⁺ T cells in PBMC, tumor and tumor-adjacent tissue (n = 12 samples from PBMC, 13 from tumor and 7 from tumor-adjacent tissue). e Representative flow cytometric analysis of PD1 and NRP1 on CD8⁺ TIL. f PD1 expression on NRP1 positive CD8⁺ T cells in PBMC, tumor and tumor-adjacent tissue (n = 8 samples from PBMC, 10 from tumor and 7 from tumor-adjacent tissue). g CDR3β diversity in NRP1 positive (⁺) and negative (-) CD8⁺ TILs (n = 4). Colored bars represent the five most abundant clonotypes, gray bars the remaining sequences. h Heatmap of TRBV usage in NRP1 positive (⁺) and negative (-) CD8⁺ TILs (n = 4). Color indicates relative usage within all of individual patients. i Representative flow cytometric analysis of TCRαβ and CT tetramer positive CD8⁺ TILs (left) and NRP1⁺ CD8⁺ TILs (right). j Percentage CT tetramer positive NRP1⁺ and NRP1- CD8⁺ TILs. k Representative CD8 and CD31 staining in ccRCC. Dashed area indicates zoom area in bottom image. Scale bar = 500 μm (top) and 250 μm (bottom). l Representative CD31, SEMA3A and CD8 staining in SEMA3A poor region (top row) and SEMA3A rich region (bottom row). Arrows indicate association between SEMA3A and CD8 staining. Scale bar = 50 μm. m Enumeration of CD8⁺ TILs in SEMA3A rich and poor regions (n = 12). Abbreviations: CT cancer testis, DI diversity indices. ns not significant. SA Shannon index, SI Simpson index. TIL tumor-infiltrating lymphocytes. TCGA The Cancer Genome Atlas, TRBV TCR beta chain variable. Error bars are means ± SD. ****P < 0.0001 by two-way ANOVA (d, f). Source data are provided as a Source Data file.