Calcium signaling in cancer and vitamin D

Calcium signals induced by the Ca2+ regulatory hormone 1,25(OH)2D3 may determine the fate of the cancer cell. We have shown that, in breast cancer cell lines, 1,25(OH)2D3 induces a sustained increase in concentration of intracellular Ca2+ ([Ca2+]i) by depleting the endoplasmic reticulum (ER) Ca2+ stores via inositol 1,4,5-trisphosphate receptor/Ca2+ release channel and activating Ca2+ entry from the extracellular space via voltage-insensitive Ca2+ channels. In normal cells, 1,25(OH)2D3 triggered a transient Ca2+ response via activation of voltage-dependent Ca2+ channels, which were absent in breast cancer cells. The normal cells, but not breast cancer cells, expressed the Ca2+ binding/buffering protein calbindin-D28k and were capable of buffering [Ca2+]i increases induced by a mobilizer of the ER Ca2+ stores, thapsigargin, or a Ca2+ ionophore, ionomycin. The 1,25(OH)2D3-induced sustained increase in [Ca2+]i in breast cancer cells was associated with induction of apoptotic cell death, whereas the transient [Ca2+]i increase in normal cells was not. The forced expression of calbindin-D28k in cytosol or increase in the cytosolic Ca2+ buffering capacity with the cell-permeant Ca2+ buffer BAPTA prevented induction of apoptosis with 1,25(OH)2D3 in cancer cells. The sustained increase in [Ca2+]i in breast cancer cells was associated with activation of the Ca2+-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 Ca2+ binding sites of μ-calpain decreased apoptotic indices in the cancer cells treated with 1,25(OH)2D3, 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 [Ca2+]i increase, activated Ca2+-dependent apoptotic proteases, and induced apoptosis in breast cancer cells in a fashion similar to that of 1,25(OH)2D3. The 1,25(OH)2D3-induced transient Ca2+ 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)2D3: increase in [Ca2+]i → μ-calpain activation → caspase-12 activation → apoptosis. Our results support the hypothesis that 1,25(OH)2D3 directly activates this apoptotic pathway by inducing a sustained increase in [Ca2+]i. Differences of Ca2+ regulatory mechanisms in cancer versus normal cells seem to allow 1,25(OH)2D3 and vitamin D analogs to induce Ca2+-mediated apoptosis selectively in breast cancer cells. Thus, deltanoids may prove to be useful in the treatment of tumors susceptible to induction of Ca2+-mediated apoptosis.