Calcium signaling in cancer and vitamin D

Abstract

Calcium signals induced by the Ca 2+ regulatory hormone 1,25(OH) 2D 3 may determine the fate of the cancer cell. We have shown that, in breast cancer cell lines, 1,25(OH) 2D 3 induces a sustained increase in concentration of intracellular Ca 2+ ([Ca 2+] i ) by depleting the endoplasmic reticulum (ER) Ca 2+ stores via inositol 1,4,5-trisphosphate receptor/Ca 2+ release channel and activating Ca 2+ entry from the extracellular space via voltage-insensitive Ca 2+ channels. In normal cells, 1,25(OH) 2D 3 triggered a transient Ca 2+ response via activation of voltage-dependent Ca 2+ channels, which were absent in breast cancer cells. The normal cells, but not breast cancer cells, expressed the Ca 2+ binding/buffering protein calbindin-D 28k and were capable of buffering [Ca 2+] i increases induced by a mobilizer of the ER Ca 2+ stores, thapsigargin, or a Ca 2+ ionophore, ionomycin. The 1,25(OH) 2D 3-induced sustained increase in [Ca 2+] i in breast cancer cells was associated with induction of apoptotic cell death, whereas the transient [Ca 2+] i increase in normal cells was not. The forced expression of calbindin-D 28k in cytosol or increase in the cytosolic Ca 2+ buffering capacity with the cell-permeant Ca 2+ buffer BAPTA prevented induction of apoptosis with 1,25(OH) 2D 3 in cancer cells. The sustained increase in [Ca 2+] i in breast cancer cells was associated with activation of the Ca 2+-dependent apoptotic proteases, μ-calpain and caspase-12, as evaluated with antibodies to active (cleaved) forms of the enzymes and the fluorogenic peptide substrates. Selective inhibition of the Ca 2+ binding sites of μ-calpain decreased apoptotic indices in the cancer cells treated with 1,25(OH) 2D 3, thapsigargin, or ionomycin. The μ-calpain activation preceded expression/activation of caspase-12, and calpain was required for activation/cleavage of caspase-12. Certain non-calcemic vitamin D analogs (e.g., EB 1089) triggered a sustained [Ca 2+] i increase, activated Ca 2+-dependent apoptotic proteases, and induced apoptosis in breast cancer cells in a fashion similar to that of 1,25(OH) 2D 3. The 1,25(OH) 2D 3-induced transient Ca 2+ response in normal mammary epithelial cells was not accompanied by activation of μ-calpain and caspase-12. In conclusion, we have identified the novel apoptotic pathway in breast carcinoma cells treated with 1,25(OH) 2D 3: increase in [Ca 2+] i → μ-calpain activation → caspase-12 activation → apoptosis. Our results support the hypothesis that 1,25(OH) 2D 3 directly activates this apoptotic pathway by inducing a sustained increase in [Ca 2+] i . Differences of Ca 2+ regulatory mechanisms in cancer versus normal cells seem to allow 1,25(OH) 2D 3 and vitamin D analogs to induce Ca 2+-mediated apoptosis selectively in breast cancer cells. Thus, deltanoids may prove to be useful in the treatment of tumors susceptible to induction of Ca 2+-mediated apoptosis.