Different molecular mechanisms of vitamin D-3 receptor antagonists

Two structurally different antagonists of the nuclear hormone 1 alpha ,25-d ihydroxyvitamin D-3 [1 alpha ,25(OH)(2)D-3], the 25-carboxylic ester ZK1592 22 and the 26,23-lactone TEI-9647, have recently been described. In this st udy, the molecular mechanisms and the efficacy of both antagonists were com pared. ZK159222 showed similar potency and sensitivity to 1 alpha ,25(OH)(2 )D-3 in ligand-dependent gel shift assays using the vitamin D receptor (VDR ), the retinoid X receptor, and specific DNA binding sites, whereas TEI-964 7 displayed reduced potency and >10-fold lower sensitivity in this assay sy stem. Limited protease digestion and gel shift clipping assays showed that the two antagonists stabilized individual patterns of VDR conformations. Bo th antagonists prevented the interaction of the VDR with coactivator protei ns, as demonstrated by GST-pull-down and supershift assays; like the natura l hormone, however, they were able to induce a dissociation of corepressor proteins. Interestingly, ZK159222 demonstrated functional antagonism in rep orter gene assays both in HeLa and MCF-7 cells, whereas TEI-9647 functioned as a less sensitive antagonist only in MCF-7 cells. In conclusion, the two 1 alpha ,25(OH)(2)D-3 analogs act in part via different molecular mechanis ms, which allows us to speculate that ZK159222 is a more complete antagonis t and TEI-9647 a more selective antagonist.