Synthesis and metabolic studies of 1α,2α,25-, 1α,4α,25- and 1α,4β,25-trihydroxyvitamin D₃

Three different A-ring perhydroxylated trihydroxyvitamin D₃ metabolites were synthesized from their appropriate A-ring precursors and CD-ring for their potential therapeutic applications. We first chemically synthesized 1α,2α,25-trihydroxyvitamin D₃ [1α,2α,25(OH)₃D₃] to study its VDR binding affinity because this metabolite is a product of recombinant human CYP3A4 catalysis when 2α-(3-hydroxypropoxy)-1α,25-dihydroxyvitamin D₃ (O2C3), a more potent vitamin D receptor (VDR) binder than 1α,25-dihydroxyvitamin D₃ [1α,25(OH)₂D₃], is used as the substrate. We found that this metabolite retained 27.3% of the VDR binding affinity compared to 1α,25(OH)₂D₃. The k_cat/K_m value of CYP24A1 for 1α,2α,25(OH)₃D₃ is 60% of that for 1α,25(OH)₂D₃. Since the biological activity and the metabolic fate of a naturally occurring C4-hydroxylated vitamin D₂ metabolite found in the serum of rats treated with pharmacological doses of vitamin D₂ have never been described, we next synthesized 1α,4α,25-trihydroxyvitamin D₃ and its diastereoisomer, 1α,4β,25-trihydroxyvitamin D₃, to study their metabolism and biological activities. Both 4-hydroxylated isomers showed weaker VDR binding affinity than 1α,25(OH)₂D₃. Although either 4-hydroxylated isomer can be metabolized by CYP24A1 almost at the same level as 1α,25(OH)₂D₃, their metabolic patterns catalyzed by uridine 5′-diphosphoglucuronosyltransferase (UGT) are different; only the 4α-hydroxylated analog can be metabolized by UGT to produce a glucuronate conjugate. The results provide important information for the synthesis of new novel chemotherapeutic vitamin D analogs which would be less subjective to degradation and therefore more bioavailable than 1α,25(OH)₂D₃. This article is part of a Special Issue entitled '17th Vitamin D Workshop'.