Contribution of several metabolites of the vitamin D analog 20-epi-22-oxa-24a,26a,27a-tri-homo-1,25(OH)(2) vitamin D-3 (KH 1060) to the overall biological activity of KH1060 by a shared mechanism of action

The synthetic 1,25-dihydroxyvitamin D-3 (1,25-(OH)(2)D-3) analog 20-epi-22- oxa-24a,26a,27a-trihomo-1,25-(OH)(2) vitamin D-3 (KH1060) is considerably m ore potent than its cognate hormone. The mechanism of action of KH1060 incl udes interaction with the vitamin D receptor(VDR). We previously showed tha t KH1060 increases VDR stability in ROS 17/2.8 osteoblastic cells by induci ng a specific conformational change in the VDR. KH1060 is metabolized, both in vivo and in vitro, into several stable products. In the present study, we investigated whether these metabolites might contribute to the increased biological activity of KH1060. We found that the potencies of two of these metabolites, 24a-OH-KH1060 and 26-OH-KH 1060, were similar to that of 1,25 -(OH),D, in inducing osteocalcin production by the osteoblast cell line ROS 17/2.8. This report further showed that these metabolites had the same eff ects as KH1060 on VDR: they increased VDR stability in ROS 17/2.8 cells, wh ile limited proteolytic analysis revealed that they caused a conformational change in the VDR, resulting in an increased resistance against proteolyti c cleavage. Furthermore, as shown in gel mobility shift assays, both compou nds clearly induced VDR binding to vitamin D response elements. Together, t hese results show that the potent in vitro activity of KH1060 is not only d irected by the effects on the VDR conformation/stabilization of the analog itself, but also by certain of its long-lived metabolites, and emphasizes t he importance of detailed knowledge of the metabolism of synthetic hormonal analogs. (C) 2000 Elsevier Science Inc.