The Sodium Potassium ATPase, Caveolin 1 and their Interaction as Potential Anti-Breast Cancer Therapeutic Targets

Breast cancer pathogenesis is modulated by complex signalling pathways centred on cholesterol-enriched membrane domains termed lipid rafts. Caveolae, a subtype of rafts, express the protein caveolin-1, which has been suggested as both as a tumour suppressor and promoter. One protein that interacts with caveolin-1 is the sodium potassium ATPase (Na+ K+ ATPase) ion channel. Na+ K+ ATPase has recently been implicated in cell signalling via interaction with its natural inhibitors, cardiac glycosides (CGs). Although predominantly used for heart failure indications, CGs reportedly have anti-proliferative properties in tumour cells. Given the importance of caveolin-1 in malignant behaviours, we hypothesized that CGs exert anti-breast cancer effects by modulating functional associations between Na+ K+ ATPase and caveolae. We first demonstrated concentration- and time-dependent anti-proliferative responses to the CGs digoxin, ouabain and oleandrin in breast cell lines and primary cultures. These effects were most pronounced in primary cell cultures, and least in ER/PR/HER2 triple-negative cells. We next showed concentration-dependent reductions in cell migration in response to CGs, particularly in ER-positive cells. CGs arrested the cell cycle at both G1 and S phase in ER-positive cells, with partial arrest at G1 in ER-negative cells. This was paralleled by increased p21 expression at both gene and protein level in ER-positive cells, and decreased p53 expression in ERnegative cells. CGs slightly decreased Na+ K+ ATPase association with caveolae in ER-positive cells, but increased that in ER-negative cells. Knockdown of caveolin-1 (but not the planar lipid raft marker flotillin-1) potentiated the effects of CGs in ER-negative but not ER-positive cells. Pharmacological lipid raft disruption abrogated the effects of CGs in ER-positive cells, while raft augmentation did the opposite. In contrast, raft disruption did not alter CG effects in ER-negative cells, but augmentation potentiated their effects. Finally, we showed that ouabain transiently activated both Src and ERK in ER-negative cells, but induced delayed Src and ERK activation in ER-positive cells. Taken together, our results provide novel evidence that the powerful anti-proliferative effects of CGs in breast cancer cells involve modulations of the functional association between Na+ K+ ATPase and caveolin-1. We suggest that the Na+ K+ ATPase, caveolin-1 and their interaction in caveolae represent exciting targets for consideration in future breast cancer drug development strategies.