Abstract B09: Nitric oxide stimulates PI3K-AKT pathway activation by S-nitrosylation of PTEN in human melanoma cells

Abstract

Abstract Based on our previous finding that inflammation associated nitric oxide synthase (NOS) expression in melanoma and its association with poor prognosis in melanoma patients, we investigated the role of nitric oxide (NO) on the PI3K-Akt pathway. Human melanoma cell lines were exposed in vitro to the NO donor (DETA NONOate), and increased Akt Serine 473 phosphorylation (pAkt S473) was detected after addition of 50-500μM DETA NONOate at as early as 2 hours. Significant increases of pAkt S473 were observed mostly in cell lines with wild-type PTEN, such as A375 and MeWo, which under our conditions do not have elevated basal Akt phosphorylation. A375 cells were further examined for DETA NONOate-induced functional consequences of Akt phosphorylation and activation; enhanced kinase activity was found to increase phosphorylation of 12 of 16 AKT signaling proteins, which include GSK, MAPK, Bad, and others. By analyzing melanoma data in The Cancer Genome Atlas (TCGA), we found a similar result that patients with positive iNOS mRNA expression have higher pAkt S473 levels compared with those without iNOS. When both PTEN and iNOS expression status were analyzed, pAkt S473 is only significantly (P<0.05) increased in patients with positive iNOS and intact PTEN, supporting our hypothesis that NO may inhibit the role of PTEN as the PI3K antagonist. Further examination in A375 cells expressing iNOS confirmed the endogenous formation of PTEN S-nitrosylation (SNO), a NO-dependent post-translational modification. PTEN in cells treated with NO showed attenuated phosphatase activity. Consistent with this view, an analysis from stage III melanoma patient tissue microarray also revealed that patients with PTEN expression had a lower survival rate when they also had positive iNOS expression. In summary, our study revealed NO can stimulate the PI3K-Akt pathway in human melanoma cells, which could be promoted by NO-mediated PTEN SNO. These experimental discoveries are supported by clinical association data and have provided one plausible mechanistic insight into the possible functional consequences of inflammatory NO produced in human melanoma cells and microenvironment. Citation Format: Zhen Ding, Dai Ogata, Jason Roszik, Yong Qin, Sun-Hee Kim, Suhendan Ekmekcioglu, Elizabeth A. Grimm. Nitric oxide stimulates PI3K-AKT pathway activation by S-nitrosylation of PTEN in human melanoma cells [abstract]. In: Proceedings of the AACR Special Conference on Melanoma: From Biology to Target; 2019 Jan 15-18; Houston, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(19 Suppl):Abstract nr B09.