Vitamin D is a prooxidant in breast cancer cells

The anticancer activity of the hormonal form of vitamin D, 1,25-dihydroxyvitamin D [1,25(OH)₂D], is associated with inhibition of cell cycle progression, induction of differentiation, and apoptosis. In addition, 1,25(OH)₂D₃augments the activity of anticancer agents that induce excessive reactive oxygen species generation in their target cells. This study aimed to find out whether 1,25(OH)₂D_3.acting as a single agent, is a prooxidant in cancer cells. The ratio between oxidized and reduced glutathione and the oxidation-dependent inactivation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) are considered independent markers of cellular reactive oxygen species homeostasis and redox state. Treatment of MCF-7 breast cancer cells with 1,25(OH)₂D₃(10-100 nM for 24-48 h) brought about a maximal increase of 41 ± 13% (mean ± SE) in the oxidized/reduced glutathione ratio without affecting total glutathione levels. The in situ activity of glutathione peroxidase and catalase were not affected by 1,25(OH)₂D₃as assessed by the rate of H₂O₂degradation by MCF-7 cell cultures. Neither did treatment with 1,25(OH)₂D₃affect the levels of glutathione reductase or glutathione S-transferase as assayed in cell extracts. The hormone did not affect overall glutathione consumption and efflux as reflected in the rate of decline of total cellular glutathione after inhibition of its synthesis by buthionine sulfoximine. The extent of reversible oxidation-dependent inactivation of GAPDH in situ was determined by comparing the enzyme activity before and after reduction of cell extracts with DTT. The oxidized fraction was 0.13 ± 0.02 of total GAPDH in control cultures and increased by 56 ± 5.3% after treatment with 1,25(OH)₂D₃which did not affect the total reduced enzyme activity. Treatment with 1,25(OH)₂D₃resulted in a ∼40% increase in glucose-6-phosphate dehydrogenase, the rate-limiting enzyme in the generation of NADPH. This enzyme is induced in response to various modes of oxidative challenge in mammalian cells. Taken together, these findings indicate that 1,25(OH)₂D₃causes an increase in the overall cellular redox potential that could translate into modulation of redox-sensitive enzymes and transcription factors that regulate cell cycle progression, differentiation, and apoptosis.