Digitoxin enhances the growth inhibitory effects of thapsigargin and simvastatin on ER negative human breast cancer cells

Background: The cardiac glycoside digitoxin preferentially inhibits the growth of breast cancer cells and targets the Erk pathway. Digitoxin alters the expression of genes that mediate calcium metabolism and IAP genes. Purpose: Since the optimal treatment for cancer involves the use of agents in combination, we assessed the growth inhibitory effects of digitoxin combined with agents that alter calcium metabolism, thapsigargin, a sarcoplasmic/ER Ca2+-ATPase inhibitor, and the statin simvastatin, as well as digitoxin's effect on the IAP pathway of apoptosis. Methods: To reveal signaling pathways, we treated human cancer cells with digitoxin, alone or combined with thapsigargin or simvastatin, and measured cell growth using the MTT and colony formation assays. We used histology and Western blot analysis of HEK293 cells to assay effects on IAPs. Results: Digitoxin inhibited the growth of breast, colon and ovarian cancer cells. Consistent with an effect on calcium metabolism, digitoxin exhibited synergy with thapsigargin and simvastatin on ER-negative breast cancer cells. Digitoxin activates expression of Erk pathway genes and suppresses expression of IAP genes. The growth inhibitory effects on HEK293 cells are not blocked by the pancaspase inhibitor zVAD-FMK, indicating that digitoxin may act by a caspase independent pathway of apoptosis. Furthermore; digitoxin does not have an effect on XIAP protein, a major anti-apoptotic protein. Conclusion: Digitoxin appears to act through the Erk and stress response pathways and is worthwhile to study to prevent and treat cancer. Our findings warn of possible safety issues for cardiac patients who take a combination of digitoxin and statins. (C) 2015 Elsevier B.V. All rights reserved.