Fig 1: CEACAM1–TIM-3 blockade alleviates anti–PD-1 resistance in RCC.a Schematic illustration of experimental workflow using a humanized PD-1 mouse model (BALB/c-hPD1, n = 5). b Representative images of orthotopically implanted tumors across treatment groups (n = 5). c Quantification of tumor weight (top) and body weight (bottom) in each group (n = 5). P values were calculated using an unpaired two-sided t-test. Data are presented as mean ± SEM. d UMAP plots showing clustering of 124,559 single cells into 12 major cell types across four treatment groups. e Differential abundance analysis by MiloR comparing Toripalimab + ML-T7 versus control groups. Nhood size corresponds to circle size, and overlap size corresponds to line thickness. The color scale indicates the log2FC between Toripalimab + ML-T7 and control groups. f Beeswarm plot showing the enrichment (orange) or decrease (green) of neighborhoods in the Toripalimab + ML-T7 group for each indicated cell type calculated using MiloR. g Violin plots showing CD8+ T cell pathway activity scores across treatment groups (Control, n = 1031; αPD-1, n = 3071; ML-T7, n = 618; αPD-1 + ML-T7, n = 1544), including oxidative phosphorylation (OXPHOS), mitochondria-related pathways, T cell tolerance induction and exhaustion. ‘n’ represents the number of single cells that passed quality control and were included in the analysis. P values were calculated using an two-sided Wilcoxon rank-sum test. P values less than 0.01, 0.001, and 0.0001 were considered statistically significant, denoted by **, ***, and ****, respectively. h Heatmap showing expression of representative metabolic and exhaustion-associated genes in CD8+ T cells across treatment groups, color indicates the normalized expression. i GSEA comparing macrophages from Toripalimab + ML-T7 vs. control, highlighting downregulation of the NLRP3 inflammasome pathway. Statistical analysis by an adaptive multi-level split Monte-Carlo scheme. j Violin plots showing expression of Nlrp3, Il1a, and Il1b in macrophages across groups (Control, n = 4033; αPD-1, n = 4961; ML-T7, n = 7780; αPD-1 + ML-T7, n = 7465). “n” represents the number of single cells that passed quality control and were included in the analysis. P values were calculated using an two-sided Wilcoxon rank-sum test. P values less than 0.05 and 0.0001 were considered statistically significant, denoted by * and ****.