Comparison of in vivo activation of triamcinolone acetonide- and RU 38486-receptor complexes in the CEM-C7 and IM-9 human leukemic cell lines.

RU 38486 functions as a pure antiglucocorticoid in the human leukemic cell lines CEM-C7 and IM-9. Despite the fact that RU 38486 has been reported to promote considerable in vitro receptor activation to a DNA-binding form, this steroid may be relatively incapable of promoting this physiologically relevant conformational change in vivo. In the experiments reported here the potential ability of RU 38486 to promote in vivo activation has been compared with that of a potent agonist, triamcinolone acetonide. In vivo activation was evaluated by DEAE-cellulose chromatographic analyses of cytosolic extracts prepared from lysed cells which had previously been labeled with either 30 nM [3H]triamcinolone acetonide or [3H]RU 38486 and subsequently incubated at 37 degrees C. Using this anion-exchange procedure, in vivo activation of the agonist-receptor complexes was shown to occur in both a time- and temperature-dependent fashion in both cell lines. This in vivo activation was reflected by progressive decreases in the bound [3H]triamcinolone acetonide associated with the unactivated (high salt-eluting) peaks eluted from DEAE-cellulose, small yet detectable increases in the bound radioactivity eluted in the activated (low-salt eluting) peaks, and a significant increase in the ability of the cytoplasmic [3H]triamcinolone acetonide-receptor complexes to bind to DNA-cellulose. Additional experiments employing DEAE-cellulose chromatography demonstrated that after incubation at 37 degrees C for 1 h, at least some in vivo activation of cytosolic [3H]RU 38486-receptor complexes could be detected in CEM-C7 cells, although the antagonist was less effective than the agonist in facilitating this conformational change. In IM-9 lymphocytes essentially no in vivo activation could be detected using this same protocol. Nuclear translocation assays were also independently performed to evaluate in vivo receptor activation. Incubation of intact cells with 30 nM [3H]triamcinolone acetonide or [3H]RU 38486 for 1 h at 37 degrees C (but not at 0-4 degrees C) revealed that both steroids facilitated translocation in CEM-C7 as well as IM-9 cells, although again the antagonist was less effective than the agonist in this regard. Taken collectively these data indicate that the antagonist RU 38486 is capable of mediating detectable in vivo activation of CEM-C7 cytoplasmic glucocorticoid receptors. These data suggest that complete in vivo stabilization of unactivated receptors by RU 38486 in this cell line may not be sufficient to explain the lack of any agonist activity associated with this synthetic antagonist.(ABSTRACT TRUNCATED AT 400 WORDS)