Mechanisms triggered by calcitriol and menadione on breast cancer cells.

GUIZZARDI S; PICOTTO G; RODRIGUEZ V; BOHL L; TOLOSA DE TALAMONI N

Mar del Plata

Congreso; LXI Reunión Científica Anual de la Sociedad Argentina de Investigación Clínica; 2016

SAIC

Calcitriol regulates proliferation, differentiation, angiogenesis and cell death of breast cancer cells. Since calcitriol produces hypercalcemic effects in vivo as a side effect, our working hypothesis was that the combination of calcitriol with menadione (MEN), a glutathione (GSH) depleting drug, would enhance 1,25 (OH)2D3 (D) antiproliferative effects without undesirable side elevations of serum calcium. The aim of the present study was to investigate about the mechanisms involved in the antiproliferative action of the combined treatment. MCF-7 cells were treated with 100 nM D, 10 µM MEN, both drugs or vehicle (ethanol) for 96 hours. Intracellular calcium concentration and mitochondrial membrane potential were evaluated by flow cytometry. Superoxide anion and nitric oxide (NO) levels were measured by spectrophotometry. The expression of TRPV6 and PMCA1b mRNA levels were determined by quantitative real-time PCR and acidic vesicular organelles (AVOs) formation, by fluorescence microscopy. Statistical analyses were performed by ANOVA / Bonferroni. Both D and the combined treatment increased NO production. Superoxide anion levels, mitochondrial membrane permeability, intracellular calcium concentration and TRPV6 and PMCA1b mRNA levels were enhanced only by the combined treatment, as compared to controls. The increment of AVOs formation suggests activation of a cell death process. In conclusion, MEN increases the effect of calcitriol on MCF-7 cells through oxidative and nitrosative stress, alteration in intracellular Ca2+concentration and changes of the expression of molecules closely related to calcium regulation, thus inducing cell death pathways.