RU 38486: potent antiglucocorticoid activity correlated with strong binding to the cytosolic glucocorticoid receptor followed by an impaired activation

Abstract

In order to explain the potent antiglucocorticoid activity of RU 38486 and the absence of agonist effect in spite of its very strong interaction with the cytoplasmic glucocorticoid receptor (GR), we investigated the compound's ability to promote GR "activation" and nuclear translocation. We have compared the dissociation-rates of the "non-activated" (molybdate stabilized) and of the "activated" (25 degrees C pre-heated) GR complexes formed either with [3H]RU 38486 or with different tritiated glucocorticoid agonists. While agonists dissociated more slowly from the "activated" than from the "non-activated" complex, RU 38486 dissociated much faster from the "activated" than from the "native" receptor. This difference of activation was confirmed in a DNA-cellulose binding assay. The affinity of the "activated" RU 38486-GR complex for DNA was much lower than that of the dexamethasone-GR complex. Finally, the in vitro nuclear uptake of [3H]RU 38486 was compared with that of [3H]dexamethasone after incubation with thymus minces at 25 or 37 degrees C. A very weak or nearly undetectable level of specific uptake of [3H]RU 38486 was observed in purified nuclei, whatever the concentration or the time of incubation used. These observations suggest that while glucocorticoid agonists form with the non-activated receptor a complex able to be activated into a more stable form (lower k-1), RU 38486 interacts strongly with the non-activated receptor (impeding the binding of DM) but the complex is "transformed" by heat to a less stable form (higher k-1), unable to translocate properly into the nucleus in order to trigger a glucocorticoid response.