Tamoxifen-induced increases in cytoplasmic free Ca2+ levels in human breast cancer cells

Tamoxifen has been shown to increase cytoplasmic free Ca2+ levels [Ca2+](i) in renal tubular cells and bladder cancer cells, and to alter Ca2+ signaling in MCF-7 breast cancer cells. The present study examined the effect of tamoxifen on [Ca2+](i) in ZR-75-1 human breast cancer cells using fura-2 as an indicator. Tamoxifen increased [Ca2+](i) at a concentration above 2 muM with an EC(5)0 of 5 muM. Removing extracellular Ca2+ reduced the response by 48 +/- 2%. In Ca2+-free medium, after tamoxifen-induced [Ca2+](i) increased had returned to baseline, adding 3 mM Ca2+ increased [Ca2+](i) in a concentration-dependent manner. Further, pretreatment with 10 muM tamoxifen abolished the [Ca2+](i) increase induced by 1 muM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor); and conversely, pretreatment with thapsigargin prevented tamoxifen from releasing more Ca2+. Tamoxifen (10 muM)-induced Ca2+ release was not changed by inhibiting phospholipase C activity with 2 muM U73122. Trypan blue exclusion assay revealed that tamoxifen (1-10 muM) did not alter viability after 1 min of incubation, but killed 10% of cells after 3-10 min of incubation. Together, this study shows that tamoxifen (>2 muM) induced a significant, immediate increase in [Ca2+](i) in ZR-75-1 breast cancer cells. Tamoxifen acted by releasing Ca2+ from the endoplasmic reticulum Ca2+ stores in a manner independent of phospholipase C activity, and by inducing Ca2+ entry from extracellular medium. Tamoxifen may be of mild cytotoxicity after acute exposure.