Spatial multi‐omics identifies a NOTCH3‐mediated capillary–mCAF crosstalk driving immune exclusion in hepatocellular carcinoma

Abstract Fibrosis induced immune exclusion is a hepatocellular carcinoma (HCC) hallmark, underscoring the key role of cancer‐associated fibroblasts (CAFs) in immune regulation. Through HCC spatial multi‐omics data and integrating pan‐cancer scRNA‐seq profiles of CAFs under immune checkpoint blockade (ICB) treatment, we characterized a potential crosstalk between capillaries and CAFs mediated by the NOTCH signaling pathway. Specifically, endothelial DLL4‐NOTCH3 signaling appears to be associated with matrix‐producing CAFs (mCAFs) polarization, leading to extracellular matrix remodeling and the establishment of immune‐restrictive niches that hinder T cell infiltration. Perturbation of NOTCH signaling attenuated mCAF differentiation and enhanced T cell infiltration in vitro, and was associated with improved ICB response in both spontaneous and orthotopic HCC mouse models. Collectively, our findings suggest that capillary‐mCAFs communication through the NOTCH pathway, particularly NOTCH3 activation, may contribute to fibrosis‐driven immune exclusion in HCC. Targeting this axis could provide a promising strategy to alleviate stromal barriers and potentiate immunotherapy efficacy.