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

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 <i>NOTCH3</i> 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.