Tamoxifen-induced increases in cytoplasmic free Ca ²⁺ levels in human breast cancer cells

Tamoxifen has been shown to increase cytoplasmic free Ca ²⁺ levels [Ca ²⁺]i in renal tubular cells and bladder cancer cells, and to alter Ca ²⁺ signaling in MCF-7 breast cancer cells. The present study examined the effect of tamoxifen on [Ca ²⁺]i in ZR-75-1 human breast cancer cells using fura-2 as an indicator. Tamoxifen increased [Ca ²⁺]i at a concentration above 2 μM with an EC ₅₀ of 5 μM. Removing extracellular Ca ²⁺ reduced the response by 48 ± 2%. In Ca ²⁺-free medium, after tamoxifen-induced [Ca ²⁺]i increased had returned to baseline, adding 3 mM Ca ²⁺ increased [Ca ²⁺]i in a concentration-dependent manner. Further, pretreatment with 10 μM tamoxifen abolished the [Ca ²⁺]i increase induced by 1 μM thapsigargin (an endoplasmic reticulum Ca ²⁺ pump inhibitor); and conversely, pretreatment with thapsigargin prevented tamoxifen from releasing more Ca ²⁺. Tamoxifen (10 μM)-induced Ca ²⁺ release was not changed by inhibiting phospholipase C activity with 2 μM U73122. Trypan blue exclusion assay revealed that tamoxifen (1-10 μM) 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 μM) induced a significant, immediate increase in [Ca ²⁺]i in ZR-75-1 breast cancer cells. Tamoxifen acted by releasing Ca ²⁺ from the endoplasmic reticulum Ca ²⁺ stores in a manner independent of phospholipase C activity, and by inducing Ca ²⁺ entry from extracellular medium. Tamoxifen may be of mild cytotoxicity after acute exposure.