Metastatic Melanoma Progression Is Associated with Endothelial Nitric Oxide Synthase Uncoupling Induced by Loss of eNOS:BH4 Stoichiometry

• Main Authors: Fabiana Henriques Machado de Melo, Diego Assis Gonçalves, Ricardo Xisto de Sousa, Marcelo Yudi Icimoto, Denise de Castro Fernandes, Francisco R. M. Laurindo, Miriam Galvonas Jasiulionis
• Format: Article
• Language: English
• Published: MDPI AG 2021-09-01
• Series: International Journal of Molecular Sciences
• Subjects: metastatic melanoma; eNOS uncoupling; tetrahydrobiopterin; L-sepiapterin; eNOS:BH4 stoichiometry
• Online Access: https://www.mdpi.com/1422-0067/22/17/9556

Melanoma is the most aggressive type of skin cancer due to its high capability of developing metastasis and acquiring chemoresistance. Altered redox homeostasis induced by increased reactive oxygen species is associated with melanomagenesis through modulation of redox signaling pathways. Dysfunctional endothelial nitric oxide synthase (eNOS) produces superoxide anion (O₂^−•) and contributes to the establishment of a pro-oxidant environment in melanoma. Although decreased tetrahydrobiopterin (BH4) bioavailability is associated with eNOS uncoupling in endothelial and human melanoma cells, in the present work we show that eNOS uncoupling in metastatic melanoma cells expressing the genes from de novo biopterin synthesis pathway <i>Gch1</i>, <i>Pts,</i> and <i>Spr</i>, and high BH4 concentration and BH4:BH2 ratio. Western blot analysis showed increased expression of <i>Nos3</i>, altering the stoichiometry balance between eNOS and BH4, contributing to NOS uncoupling. Both treatment with L-sepiapterin and eNOS downregulation induced increased nitric oxide (NO) and decreased O₂^• levels, triggering NOS coupling and reducing cell growth and resistance to <i>anoikis</i> and dacarbazine chemotherapy. Moreover, restoration of eNOS activity impaired tumor growth in vivo. Finally, <i>NOS3</i> expression was found to be increased in human metastatic melanoma samples compared with the primary site. eNOS dysfunction may be an important mechanism supporting metastatic melanoma growth and hence a potential target for therapy.