The catalytic Na⁺,K⁺-ATPase subunits include four isoforms (α1-α4 isoforms). Volume-regulated anion channel (VRAC) plays an important role in cell death signaling pathway in addition to its fundamental role in the cell volume maintenance. Disruption of actin filaments causes the dysfunction of VRAC, which elicits resistance to cisplatin in the cancer cells. First, we introduce the cardiac glycosides-induced signaling pathway mediated by the crosstalk between Na⁺,K⁺-ATPase α1-isoform (α1NaK) and VRAC in the cancer cells. In this mechanism, sub-micromolar concentrations of cardiac glycosides bind to the receptor-type α1NaK localized in the membrane microdomain of the cells, and increase VRAC activities concomitantly with a deceleration of cancer cell proliferation. Second, we introduce the pathophysiological function of α3NaK, which is abnormally expressed in the intracellular vesicles of cancer cells. In general, cancer cells can survive even under loss of anchorage because they have the avoidance mechanism for anoikis. On cancer cell detachment, we found that intracellular α3NaK is translocated to the plasma membrane and this event contributes to survival of the cells. Interestingly, cardiac glycosides inhibited the α3NaK translocation. Our findings may open up new opportunities for development of cancer medicines.