Employing the enzyme cofactor function of ascorbic acid to affect oncogenic pathways in human melanoma: modulating Hypoxia Inducible Factor-1α and DNA demethylation to reduce malignant potential

Dioxygenase enzymes such as the HIF hydroxylases (PHD1-3, FIH) and the Ten-eleven translocation (TET1-3) enzymes regulate the activity of the hypoxia inducible factor-1a (HIF1a) transcription factor and the DNA methylation status of cells, respectively. Aberrant accumulation and activation of HIF-1a can allow malignant cells to acquire attributes that promote progression, chemotherapy resistance, and survival, while aberrant hypermethylation of gene promoters can silence the expression of tumor suppressor genes essential to preventing tumorigenesis. Inadequate levels of intracellular ascorbic acid (AA), a necessary cofactor for optimal dioxygenase enzyme function, could potentiate these tumorigenic conditions. In fact, plasma levels of AA have been found to be below normal physiological levels in individuals with melanoma, as well as other cancer types. Interestingly, melanoma tumors frequently demonstrate both high expression of HIF-1a and increased promoter methylation. AA therefore may present a valuable adjuvant therapy option for melanoma patients by regulating the HIF-1 and DNA demethylation pathways. In these studies, we investigated the ability of AA supplementation to decrease aberrant HIF-1a accumulation and activity in melanoma cell lines and to increase global 5-hydroxymethylcytosine (5hmC) content of DNA in metastatic melanoma to ultimately reduce the malignant potential. Our studies showed that treatment with AA (reduced vitamin C) significantly increased intracellular vitamin C content of cultured melanoma cells and significantly decreased the stability, accumulation, and transcriptional activity of HIF-1α. Supplementation with AA also significantly increased global 5hmC content and ultimately resulted in the significant decrease of malignant potential of metastatic melanoma cells. Additionally, we discovered novel evidence that HIF-1α may regulate the DNA methylation status of melanoma cells by influencing the expression of TET2. Taken together, these results suggest that deficiencies in AA levels, commonly observed in malignant tissue and blood plasma of cancer patients, may be a contributing factor to the deregulation of the HIF-1α and DNA demethylation pathways. Deregulation of these pathways is likely due to the loss of its cofactor function for the dioxygenase enzymes that tightly regulate various oncogenic factors. Furthermore, AA has the potential to be a powerful adjuvant therapy for metastatic melanoma, and further investigations in multiple cancer types are warranted to advance the use of AA to the clinical setting.