CD160 is an inducible endothelial checkpoint that suppresses sprouting angiogenesis through LRP1-dependent autophagy
Pathological angiogenesis is a hallmark of cancer and other chronic diseases, yet therapeutic strategies remain largely focused on blocking pro-angiogenic signals rather than restoring endogenous mechanisms that restrain endothelial sprouting. The identification of endothelial checkpoints capable of limiting pathological vessel growth therefore remains an important unmet need. CD160 is a glycosylphosphatidylinositol-anchored immunoglobulin-superfamily receptor of cytotoxic lymphocytes and an inhibitory checkpoint receptor on CD4 T cells. It is also present on endothelium and has been targeted therapeutically. Here, we identify CD160 as an inducible endothelial checkpoint that emerges in response to pro-angiogenic cues and limits pathological angiogenesis. CD160 is absent in resting endothelium. It is induced by basic fibroblast growth factor (bFGF) and most strongly by combining VEGF-A and bFGF, and it is detected in the sprouting region of ex vivo mouse aortic rings. Targeting of CD160 using monoclonal antibody reduces tubulogenesis, scratch closure and aortic-ring sprouting, and softens the cell cortex, while viability and membrane integrity remain intact. Mechanistically, CD160 associates with the endocytic receptor LRP1 as demonstrated by proximity ligation assay, lipid-raft co-immunoprecipitation and super-resolution fluorescence microscopy. Both receptors appear in a common complex, and two independent computational methods converge on the first ligand-binding cluster of LRP1 as the interface. Silencing CD160 or LRP1 abolishes the anti-angiogenic response mediated by the antibody. Under a chloroquine clamp, engagement raises autophagic flux in a CD160- and LRP1-dependent manner, and lysosomal or LDL-receptor-family blockade removes the anti-sprouting effect. CD160 therefore acts as an inducible endothelial checkpoint that suppresses sprouting through autophagic flux. Thus, CD160 represents an inducible endothelial checkpoint that is activated in a pro-angiogenic environment, and targeting CD160 restrains pathological angiogensis.