Digitalis structure-activity relationship analyses. Conclusions from indirect binding studies with cardiac (Na+ + K+)-ATPase.

We have performed direct and indirect binding studies with [3H]ouabain or [3H]digitoxin on beef or guinea pig cardiac (Na+ + K+)-ATPase to measure the potencies of a broad range of cardiotonic steroids for structure-activity relationship (SAR) studies for comparison with previously determined positive inotropic potencies. The positive inotropic potencies of twelve compounds on contracting guinea pig left atria correlated well with the equilibrium dissociation constants (KD values) from the inhibition of [3H]ouabain binding to guinea pig cardiac (Na+ + K+)-ATPase (r = 0.98 for seven 5 beta-compounds, r = 0.95 for five 5 alpha-compounds). Further we calculated KD values from the inhibition of [3H]ouabain binding data for a total of 33 digitalis derivatives on the digitalis-sensitive beef cardiac (Na+ + K+)-ATPase. For the 27 compounds tested on both beef cardiac (Na+ + K+)-ATPase and guinea pig left atria, the potencies showed a significant correlation (r = 0.92 for 22 5 beta-compounds, r = 0.96 for five 5 alpha-compounds. For seven compounds, KD values were measured on beef cardiac (Na+ + K+)-ATPase using inhibition of binding of [3H]digitoxin. These values correlated well (r = 0.99) with the KD values from the [3H]ouabain studies. These results show that: (1) The significant correlation observed between KD values on guinea pig cardiac (Na+ + K+)-ATPase and positive inotropic potency in guinea pig left atria is further evidence that the pharmacological receptor for inotropy is part of the enzyme, (2) Inhibition of the binding of [3H]ouabain or [3H]digitoxin can be used to determine the relative potencies of unlabelled digitalis derivatives. The similar relative potencies on beef and guinea pig cardiac (Na+ + K+)-ATPase of a broad range of digitalis derivatives indicate that the binding site is similar for both species; and (3) SAR studies indicate that functional groups on these steroids have the same influence on potency on either the positive inotropy or cardiac (Na+ + K+)-ATPase studies.