Cardiac Glycosides Inhibit p53 Synthesis by a Mechanism Relieved by Src or MAPK Inhibition

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Abstract
P53 is regulated at multiple levels. We report here that p53, in multiple lines of human cancer cells, is down-regulated by cardiac glycoside drugs digoxin and ouabain, potent inhibitors of Na+/K+-ATPase. These drugs reduced the basal levels of p53 protein at nanomolar concentrations in a dose-, time-, and cancer cell line–dependent manner, but independent of p53 status of wild-type or mutant. The drugs also reduced the levels of p53 induced by its activators as well as p53 transfected into human cancer cells, regardless of its status. Interestingly, the drugs had no effect on endogenous p53 in two immortalized human cell lines. Mechanistically, p53 reduction occurred not at the mRNA levels but at the protein levels, as a result of reduced protein synthesis rather than enhanced degradation. The cellular sensitivity to drug-induced p53 reduction was not associated with the levels of αsubunits of Na+/K+-ATPase in different cell lines. Although lowering extracellular K+ did not reduce p53 as did ouabain and digoxin, it did potentiate both digoxin- and ouabain-induced p53 reduction in sensitive lines. Finally, p53 reduction seems to be triggered by activation of Src/mitogen-activated protein kinase (MAPK) signaling pathways upon drug binding to the Na+/K+-ATPase and can be completely blocked by the inhibitors of Src or MAP/ERK kinase. This is the first report that cardiac glycoside drugs, by initiating the Src/MAPK signaling pathways, reduce the p53 levels via inhibition of p53 protein synthesis. The drugs may be useful in the treatment of human cancers with a gain-of-function p53 mutation. [Cancer Res 2009;69(16):6556–64]