Inhibition of Notch4 with Novel Neutralizing Antibodies Reduces Tumor Growth in Multiple Tumor Types and Targets Endothelium and Tumor Associated Macrophages in a Mouse Breast Cancer Model [RiboTag RNA-seq]

Endothelial Notch signaling is critical for tumor angiogenesis. Notch1 blockade reduces tumor vessel function but causes vascular dysfunction, hypoxia, and gastrointestinal toxicity. Notch4 expression is restricted to developing vasculature, where it promotes angiogenesis. Unlike other Notch proteins, Notch4 has rarely been targeted therapeutically. We developed highly specific Notch4-blocking antibodies, 6-3-A6 and humanized E7011, allowing therapeutic targeting of Notch4 in tumor models. E7011 treatment significantly delayed tumor growth and impaired tumor vessel perfusion in mouse models of breast, skin, and lung tumors. In a mouse orthotopic breast cancer model, Notch4 expression was confined to endothelial cells, but E7011 treatment significantly increased tumor-associated macrophages and increased macrophage expression of anti-tumor genes. Liposomal clodronate-mediated macrophage depletion reversed the anti-tumor effects of E7011, demonstrating that Notch4 supports tumor growth via angiocrine signaling to tumor macrophages. We conclude that targeting Notch4 alters tumor vessel function and tumor associated macrophage phenotype, leading to tumor inhibition.