Metabolism of 2α-[2-(tetrazol-2-yl)ethyl]-1α,25-dihydroxyvitamin D by CYP24A1 and biological activity of its 24R-hydroxylated metabolite.

Our previous study revealed that the 2α-substituted vitamin D analog 2α-[2-(tetrazol-2-yl)ethyl]-1α,25(OH)D (AH-1) exhibited a higher osteocalcin promoter transactivation activity in human osteosarcoma cells and a greater effect on bone mineral density in a rat model of osteoporosis than 1α,25(OH)D　without increasing blood calcium concentration. Thus, we hypothesized that AH-1 could be a promising therapeutic agent for osteoporosis without any serious side effects. In this study, we compared the CYP24A1-dependent metabolism of AH-1 with that of 1α,25(OH)D.

Synthesis of the CYP24A1 major metabolite of 2α-[2-(tetrazol-2-yl)ethyl]-1α,25-dihydroxyvitamin D.

Previously, we found that 2α-[2-(tetrazol-2-yl)ethyl]-1α,25-dihydroxyvitamin D (AH-1) showed higher osteocalcin promoter transactivation activity in human osteosarcoma (HOS) cells and a greater therapeutic effect on ovariectomized (OVX) rats for enhancing bone mineral density than those of 1α,25(OH)D without hypercalcemic side effects in vivo. Although CYP24A1 catalyzes multi-step oxidations toward the CD-ring side chain of the active vitamin D [1α,25(OH)D], the CYP24A1-dependent metabolism of AH-1 tended to stop at the first step hydroxylation at the C24-position of AH-1. Interestingly, the metabolite 24-hydroxy-AH-1 [24(OH)AH-1] showed potent VDR binding affinity, and the new chiral center of the 24-position might be the 24R configuration compared with the process of the natural 1α,25(OH)D catabolism.

Synthesis, metabolism, and biological activity of 2-[3-(tetrazolyl)propyl]-1α,25-dihydroxy-19-norvitamin D.

Recently, we found that 2α-[2-(tetrazol-2-yl)ethyl]-1α,25-dihydroxyvitamin D showed higher osteocalcin promoter transactivation activity in human osteosarcoma (HOS) cells and a greater therapeutic effect in ovariectomized (OVX) rats in vivo than those of active vitamin D, 1α,25(OH)D. We were interested in introducing a heterocyclic ring to the C2 position of the seco-steroidal structure via an alkyl linker, and four novel C2-(3-tetrazolylpropyl) substituted 1α,25-dihydroxy-19-norvitamin D analogs, 2α-[3-(tetrazol-1-yl)propyl]-, 2β-[3-(tetrazol-1-yl)propyl]-, 2α-[3-(tetrazol-2-yl)propyl]-, and 2β-[3-(tetrazol-2-yl)propyl]-19-nor-1α,25(OH)D were synthesized. Among them, 2α-[3-(tetrazol-1-yl)propyl]-19-nor-1α,25(OH)D showed weak binding affinity for human vitamin D receptor (hVDR) (2.