ALDH1A1 is a cytosolic enzyme upregulated in tumor cells, involved in detoxifying cells from reactive aldehydes and in acquiring resistance to chemotherapeutic drugs. Its expression correlates with poor clinical outcomes in a number of cancers, including melanoma. The present study hypothesized that the increased ALDH1A1 expression and activity upregulated the release of proangiogenic factors from melanoma cells, which regulate angiogenic features in endothelial cells (ECs) through a rearrangement of the Notch pathway. In vivo, when subcutaneously implanted in immunodeficient mice, ALDH1A1 overexpressing melanoma cells displayed a higher microvessel density. In a 3D multicellular system, obtained co‑culturing melanoma cancer cells with stromal cells, including ECs, melanoma ALDH1A1 overexpression induced the recruitment of ECs into the core of the tumorspheres. By using a genes array, overexpression of ALDH1A1 in tumor cells also promoted modulation of Notch cascade gene expression in ECs, suggesting an interaction between tumor cells and ECs mediated by enrichment of angiogenic factors in the tumor microenvironment. To confirm this hypothesis, inactivation of ALDH1A1 by the pharmacological inhibitor CM037 significantly affected the release of angiogenic factors, including IL‑8, from melanoma cells. High levels of ALDH1A1, through the retinoic acid pathway, regulated the activation of NF‑kB‑p65 and IL‑8. Further, in a 2D co‑culture system, the addition of an IL‑8 neutralizing antibody to ECs co‑cultured with melanoma cells forced to express ALDH1A1 dampened endothelial angiogenic features, both at the molecular (in terms of gene and protein expression of mediators of the Notch pathway) and at the functional level (proliferation, scratch assay, tube formation and permeability). In conclusion, these findings demonstrated the existence of a link between melanoma ALDH1A1 expression and EC Notch signaling modification that results in a pro‑angiogenic phenotype. Based on the crucial role of ALDH1A1 in melanoma control of the tumor microenvironment, the enzyme seems a promising target for the development of novel drugs able to interrupt the cross‑talk between cancer (stem) cells and endothelial cells.
Keywords: IL‑8; Notch; aldehyde dehydrogenase 1A1; angiogenesis; melanoma; multicellular 3D system; stemness.