Tumor-directed VTN signaling is a shared across clusters. A, CellChat results were subset to communication predicted to originate from hepatocyte-annotated cluster 7 and be received by tumor-annotated clusters 0, 3, and 4. Mean CellChat probabilities were calculated for each pathway across tumor clusters, pathways were ordered by their mean value, and the top 60 were selected for plotting. B, Ligands for the top 20 pathways in (A) were retrieved from CellChat results and annotated with tissue-level expression from The Human Protein Atlas bulk RNA-seq data. Tissue specificity was calculated for each ligand with the Tau index, and pathways were ordered by the mean Tau index of their ligands. C, CellChat-predicted ligand–receptor pairs involved in VTN signaling were retrieved from the CellChat object. CellChat probability scores and P values (permutation test) were retrieved for each pair and communicating cluster and subset to significant (P < 0.05) hepatocyte-to-tumor communication. Ligand–receptor pairs were ordered by their median CellChat probability score across tumor clusters for plotting. D, Single-cell normalized expression data were extracted for CellChat-annotated ligand–receptor pairs involved in VTN signaling. Boxplots show the distribution of cell-level expression for hepatocytes and tumor cells, and points show cluster means. Significance was calculated with DESeq2 from sample and tumor/hepatocyte-level pseudobulked counts. ns, not statistically significant. E, UMAPs colored by binary VTN, ITGB1, or ITGAV expression. F and G, Count and FPKM matrices were obtained from GSE211448 and cBioPortal for the 22Rv1 orthotopic model and SU2C Dream Team datasets, respectively. For 22Rv1, DESeq2 was applied to normalize counts and calculate FCs between 22Rv1-LMD and primary 22Rv1. For SU2C Dream Team data, FCs were calculated from pseudocount-adjusted FPKMs. Then, FCs were inputted to fgsea for GSEA, which uses a false discovery rate–adjusted permutation test to calculate significance. Enrichment results for Reactome integrin signaling (R-HSA-354192) are shown. H, Liver tissue was harvested at autopsy (Supplementary Table S8) and formalin-fixed, paraffin-embedded, and sectioned. Slides were stained with hematoxylin and eosin (H&E), relevant lineage markers, and VTN. ***, P < 0.001.