We examined the effects of three cardiac glycosides, ouabain, digoxin and proscillaridin A, on the proliferation of estrogen independent MDA-MB-231 breast cancer cells. In terms of reduction in cell viability, the compounds rank for both 24 hand 48 h of incubation in MDA-MB-231 cells in the order proscillaridin A > digoxin > ouabain. Digoxin for 24 hand 48 h of incubation in MDA-MB-231 cells proved to be only slightly more potent than ouabain, with $IC_{50}$ values of $122{pm}2;and;70{pm}2nM$, respectively, compared to $150{pm}2;and;90{pm}2nM$ for ouabain. In contrast, proscillaridin A, was much more active and showed a high level of cytotoxic potency, $IC_{50};51{pm}2;and;15{pm}2nM$ for 24 hand 48 h of incubation, respectively. The concentrations of digoxin, ouabain and proscillaridin A needed to inhibit [$^3H$]thymidine incorporation into DNA by 50% ($IC_{50}$) in MDA-MB-231 cells for 24 h of incubation were found to be $124{pm}2nM,;142{pm}2nM,;and;48{pm}2nM$, respectively. In the present study, we demonstrated that ouabain, digoxin, and proscillaridin A induce apoptosis in MDA-MB-231 cells by increasing free calcium concentration and by activating caspase-3.