Development of in vitro and in vivo tools to evaluate the antiangiogenic potential of melatonin to neutralize the angiogenic effects of VEGF and breast cancer cells: CAM assay and 3D endothelial cell spheroids

Melatonin is a molecule with different antitumor actions in breast cancer and has been described as an inhibitor of vascular endothelial growth factor (VEGF). Despite the recognition of the key role exerted by VEGF in tumor angiogenesis, limitations arise when developing models to test new antiangiogenic molecules. Thus, the aim of this study was to develop rapid, economic, high capacity and easy handling angiogenesis assays to test the antiangiogenic effects of melatonin and demonstrate its most effective dose to neutralize and interfere with the angiogenic sprouting effect induced by VEGF and MCF-7. To perform this, 3D endothelial cell (HUVEC) spheroids and a chicken embryo chorioallantoic membrane (CAM) assay were used. The results showed that VEGF and MCF-7 were able to stimulate the sprouting of the new vessels in 3D endothelial spheroids and the CAM assay, and that melatonin had an inhibitory effect on angiogenesis. Specifically, as the 1 mM pharmacological dose was the only effective dose able to inhibit the formation of ramifications around the alginate in the CAM assay model, this inhibition was shown to occur in a dose-dependent manner. Taken together, these techniques represent novel tools for the development of antiangiogenic molecules such as melatonin, with possible implications for the therapy of breast cancer. ; Funding: This work was funded in part by PE-0106–2019 from the Consejería de Salud de la Junta de Andalucía, C19047–2018 from Fundación Unicaja and UMA18-FEDERJA-042 from UMA-FEDER & ALIANZA MIXTA ANDALUCÍA-ROCHE. Alicia González González is a recipient of a postdoctoral grant Margarita Salas (RMS-08) from European Union-NextGenerationEU, Spanish Ministry of Universities and Recovery Transformation and Resilience Plan, through a call from University of Cantabria. Aurora Laborda Illanes is a recipient of a predoctoral grant, PFIS-ISCIII (FI19–00112), co-funded by the Fondo Social Europeo (FSE). Lidia Sanchez Alcoholado is a recipient of a postdoctoral grant (RH-0026–2021) from the ...