Paradoxical differences in the receptor binding of two new antimineralocorticoids

Two synthetic derivatives of spironolactone were used to examine various aspects of the mineralocorticoid receptor structure and function. Introduction of a propyl residue in the 7-position of spironolactone produced a molecule (RU 26752) that saturated the aldosterone specific receptor in the 1-10 nM range, and another, more abundant species in the 10-100 nM range which had little affinity for the natural hormone. The specificity for both sites was increased when the methoxycarbonyl group was introduced in the 7-position (ZK 91587). Neither antagonist exhibited affinity for blood serum transcortin or receptors in non-target organs like the lung and the liver. RU 26752-receptor complex was more unstable than the hormone-receptor complex at 35 degrees C but underwent comparable thermal activation as evidenced by binding to DNA cellulose and the 7 S to 4 S shift on sucrose gradients. In contrast, ZK 91587 did not permit thermal activation and greatly labilized the receptor at 35 degrees C. In ion exchange chromatography, two peaks were observed with unactivated ZK 91587-receptor complex, but RU 26752 was bound exclusively to the component eluted with high salt. Molecular filtration revealed two peaks of bound radioactivity with both antimineralocorticoids. These studies reveal important differences in the mechanism of action of two antagonists differing solely in the residue in position 7 of the spironolactone molecule. Such differences could be exploited to purify the mineralocorticoid receptor and clinically to prescribe the appropriate drug with greater precision.