Synthesis, Biological Evaluation, and Conformational Analysis of A-Ring Diastereomers of 2-Methyl-1,25-dihydroxyvitamin D₃ and Their 20-Epimers:  Unique Activity Profiles Depending on the Stereochemistry of the A-Ring and at C-20

Abstract

All eight possible A-ring diastereomers of 2-methyl-1, 25-dihydroxyvitamin D(3) (2) and 2-methyl-20-epi-1, 25-dihydroxyvitamin D(3) (3) were convergently synthesized. The A-ring enyne synthons 19 were synthesized starting with methyl (S)-(+)- or (R)-(-)-3-hydroxy-2-methylpropionate (8). This was converted to the alcohol 14 as a 1:1 epimeric mixture in several steps. After having been separated by column chromatography, each isomer led to the requisite A-ring enyne synthons 19 again as 1:1 mixtures at C-1. Coupling of the resulting A-ring enynes 20a-h with the CD-ring portions 5a,b in the presence of a Pd catalyst afforded the 2-methyl analogues 2a-h and 3a-h in good yield. In this way, all possible A-ring diastereomers were synthesized. The synthesized analogues were biologically evaluated both in vitro and in vivo. The potency was highly dependent on the stereochemistry of each isomer. In particular, the alpha alpha beta-isomer 2g exhibited 4-fold higher potency than 1 alpha,25-dihydroxyvitamin D(3) (1) both in bovine thymus VDR binding and in elevation of rat serum calcium concentration and was twice as potent as the parent compound in HL-60 cell differentiation. Furthermore, its 20-epimer, that is, 20-epi-alpha alpha beta 3g, exhibited exceptionally high activities: 12-fold higher in VDR binding affinity, 7-fold higher in calcium mobilization, and 590-fold higher in HL-60 cell differentiation, as compared to 1 alpha,25-dihydroxyvitamin D(3) (1). Accordingly, the double modification of 2-methyl substitution and 20-epimerization resulted in unique activity profiles. Conformational analysis of the A-ring by (1)H NMR and an X-ray crystallographic analysis of the alpha alpha beta-isomer 2g are also described.

New insights into Vitamin D sterol-VDR proteolysis, allostery, structure–function from the perspective of a conformational ensemble model

“…1A) [41,42]. For example the 2α-CH 3 modification enhances 1,25D's VDR RCI value 4-fold (RCI = 400%), the potency of MK antagonism of 1,25D mediated gene transcription [42]; however, 2α-CH 3 -1,25D fails to stimulate apoptosis [43].…”

Toward the synthesis of the carbacylic ansa antibiotic kendomycin

“…Unfortunately, the opening of epoxide 20 (17) with aryl bromide 19 failed completely, as did the Pd(0)-catalyzed coupling of aryl bromide 5 with alkene 21 and the corresponding alkyne, respectively. Eventually, a moderate yield could be obtained in the opening of epoxide 20 with aryl bromide 22 to give coupling product 23.…”

Metabolism of 2-Methyl Analogs of 1α,25-Dihydroxyvitamin D₃ in Rat Osteosarcoma Cells (UMR 106)

“…Cell culture medium was changed every 3-4 d. The cells were subcultured when approximately 80% confluent and were not subcultured beyond five passages. For the metabolism studies, 3ϫ10 6 cells were seeded in T150 tissue culture bottles and grown to confluence. The incubations were carried out at 37°C in a humidified atmosphere under 5% CO 2 .…”

“…We recently synthesized all eight possible A-ring diastereomers of 2-methyl-1a,25(OH) 2 D 3 (these analogs contain a methyl group on C-2 of the A-ring, and differ from each other in the stereochemistry of the methyl group on C-2 and the hydroxyl groups on C-1 and C-3) in order to investigate the structure-function relationship of the natural hormone and demonstrated that in addition to the C-1 and C-3 hydroxyl groups, the configuration of the 2-methyl group also considerably alters the activity. [5][6][7] The 2-methyl analogs show unique biological properties. For example, the VDR binding affinities exhibited by the 1a-isomers are significantly higher than those exhibited by the 1b-isomers.…”