Theory of partial agonist activity of steroid hormones

Abstract

The different amounts of residual partial agonist activity (PAA) of antisteroids under assorted conditions have long been useful in clinical applications but remain largely unexplained. Not only does a given antagonist often afford unequal induction for multiple genes in the same cell but also the activity of the same antisteroid with the same gene changes with variations in concentration of numerous cofactors. Using glucocorticoid receptors as a model system, we have recently succeeded in constructing from first principles a theory that accurately describes how cofactors can modulate the ability of agonist steroids to regulate both gene induction and gene repression. We now extend this framework to the actions of antisteroids in gene induction. The theory shows why changes in PAA cannot be explained simply by differences in ligand affinity for receptor and requires action at a second step or site in the overall sequence of reactions. The theory also provides a method for locating the position of this second site, relative to a concentration limited step (CLS), which is a previously identified step in glucocorticoid-regulated transactivation that always occurs at the same position in the overall sequence of events of gene induction. Finally, the theory predicts that classes of antagonist ligands may be grouped on the basis of their maximal PAA with excess added cofactor and that the members of each class differ by how they act at the same step in the overall gene induction process. Thus, this theory now makes it possible to predict how different cofactors modulate antisteroid PAA, which should be invaluable in developing more selective antagonists.

Keywords: antagonist; antiestrogen; antisteroid; gene expression; gene transcription; glucocorticoid receptor; ligand; nuclear receptor; raloxifene; steroid receptor; tamoxifen.

Figure 1. Dose-response curves of GR induction of GREtkLUC ± Ubc9 are first order

Triplicate wells of CV-1 cells were transiently transfected with 100 ng of GREtkLUC reporter, 10 ng of Renilla-TS as an internal control, and the indicated amounts of pSG5/GR plasmid ± 135 ng of pSG5/Ubc9 with enough pSG5/hSA to have a constant amount of pSG5 plasmid. After 18 hr of induction by the indicated Dex concentrations, the amount of Luciferase activity in lysed cells was determined and expressed as percent of the maximal activity in each set. Error bars are S.D. of the triplicate samples. Curves are best fits for a first-order Hill plot. Similar results were obtained in three additional independent experiments.

Figure 2

Structures of steroidal ligands used in this study.

Figure 3. Effect of changing concentrations of GR and Ubc9 on PAA of antiglucocorticoids

Experiments were conducted as in Figure 1 with the indicated amounts of pSG5/GR ± 135 ng pSG5/Ubc9 except that the steroid treatments were 1 μM antisteroid. Luciferase activities were determined and the PAA of each steroid was calculated relative to 1 μM Dex under the same conditions. The values of 4 independent experiments were averaged and plotted ± S.E.M. The thin horizontal line at 50% is only for reference.