Human cell mutants affected in the interaction of the 12β-OH group of cardiac glycosides with the digitalis receptor

Abstract

The cross-resistance patterns of two different types of mutants of HeLa cells selected for resistance to the digoxin analog SC4453 (SC R mutants) in which the Na +/K +-ATPase is affected [A. Chopra and R.S. Gupta, J. biol. Chem. 261, 2034 (1986)], and towards numerous cardiac glycosides (CGs) and genins, were examined. One type of SC R mutant (designated as group C) was highly resistant to all CGs and genins investigated. In contrast, the other type of SC R mutant (group D) showed a high degree of cross-resistance towards selected CG derivatives (viz. digoxin, SC4453, digoxigenin, lanatoside C, α- and β-methyldigoxin, dihydrodigoxin, α- and β-acetyldigoxin, α,β-diacetyldigoxin), all of which contained a free 12β-OH group in the steroid structure. Slight cross-resistance of the group D mutants was also observed for other compounds (viz. ouabain, ouabagenin, dihydroouabain) that contain a free 11α-OH group in the molecule. However, these mutants exhibited no cross-resistance to other CG derivatives, which either lacked the above groups (viz. digitoxin, digitoxigenin, dihydrodigitoxin, digitoxigenin mono- and bisdigitoside, nerifolin, gitoxigenin, gitoxin. 16-acetylgitoxin, lanatosides A and B, cymarin, convallatoxin, oleandrin, strophanthidin, actodigin and bufalin) or in which the 12β- OH group was acetylated (viz. as in the case of 12-acetyldigoxin). Since the 12β-OH group is not required for CG-like activity, to account for these observations it is suggested that the genetic lesion in the group D mutant leads to the creation of a new binding site in the digitalis receptor, which specifically interacts with the 12β-OH group (the site presumably also interacts weakly with the 11α-OH group) and either prevents or distorts the binding of the compounds to the drug binding site on the receptor. Further investigations with the different classes of CG-resistant mutants at the molecular level should prove very useful in identifying the drug receptor site and in understanding how these drugs interact with it.