Para-coumaric acid suppresses VEGF-induced DLL4–NOTCH1 signaling, tumor angiogenesis, and tumor growth

Introduction: DLL4–NOTCH1 signaling mediates juxtacrine communication between tip- and stalk-like endothelial cells during vascular sprouting, contributing to complementary aspects of endothelial function. Thus, simultaneous modulation of DLL4 and NOTCH1 may provide a strategy to broadly interfere with the endothelial functions involved in tumor angiogenesis. Accordingly, this study aims to identify a novel natural compound capable of modulating DLL4–NOTCH1-mediated endothelial communication and to investigate how such modulation influences tumor angiogenesis and growth. Methods: We screened 66 natural compounds to identify those that suppress DLL4 and NOTCH1 signaling in endothelial cells. Using a dual-reporter system that combines a DLL4 promoter-driven reporter and a CSL luciferase reporter, we simultaneously assessed ligand-associated activity and downstream transcriptional output of the DLL4–NOTCH1 axis. We confirmed VEGF-induced target gene expression via western blotting and quantitative real-time polymerase chain reaction. We further performed in vitro angiogenesis assays and in vivo Lewis lung carcinoma (LLC) allograft tumor experiments. Results: Among the 66 screened natural compounds, para-coumaric acid (p-CA) dose-dependently suppressed DLL4 transcription and NOTCH1 signaling activity in endothelial cells, with IC50 values of 4.57 and 8.34 µM, respectively. Furthermore, p-CA significantly suppressed VEGF-induced endothelial cell proliferation, migration, and tube formation. In the LLC allograft model, p-CA treatment suppressed tumor growth and vascular density, increased pericyte coverage, reduced tumor hypoxia, and enhanced CD3+ T-cell infiltration and tumor cell apoptosis. Conclusion: In summary, p-CA is a natural compound that functionally suppresses VEGF-driven DLL4–NOTCH1 signaling in endothelial cells, exhibiting strong anti-angiogenic effects and promoting functional vascular normalization, resulting in marked tumor suppression and enhanced antitumor immunity in vivo. These findings suggest that p-CA warrants further investigation as an anti-angiogenic compound for cancer therapy.