PROFILE OF LIGAND SPECIFICITY OF THE VITAMIN-D-BINDING PROTEIN FOR 1-ALPHA,25-DIHYDROXYVITAMIN-D-3 AND ITS ANALOGS

The profile of structural preference for the ligand binding domain of the human vitamin D binding protein (DBP) was determined by steroid co mpetition assay of 71 analogs of 1 alpha,25-dihydroxyvitamin D-3 [1 al pha,25(OH)(2)D-3]. The following categories of structural modification were evaluated [values represent fold change; R = reduction, I = incr ease in binding to the DBP from the reference 1 alpha,25(OH)(2)D-3]: ( 1) deletion in the A ring of the 1 alpha-hydroxyl (20-1600I); (2) conv ersion of the triene system to the previtamin form (6-40R); (3) additi on of substituents to carbon 11 of the C ring (4-14R); (4) inversion o f the C/D ring junction (8-20R); (5) unsaturation of the D ring (16-en e; 4-140R); (6) replacement of hydrogen with deuterium atoms (no effec t); alteration of the side chain by (7) adding or deleting carbon atom s (5-12R); (8) addition of fluorines (0.2-10R); (9) presence of unsatu ration (22-ene, 0-5R; 23-ene, 3R-10I; 23-yne, 5-20R); (10) addition of hydroxyls (2-100R); and (11) addition of an aromatic ring (0-20I). Th us the DBP ligand binding domain could tolerate only modest changes to the structure of 1 alpha,25(OH)(2)D-3 without a reduction in binding of the analog. The increases in binding seen in the aromatic side chai n and with a triple bond at carbon-23 may be indicative of a preferred conformation of the flexible 1 alpha 25(OH)(2)D-3 side chain. In addi tion, a comparison was made of the DBP ligand binding domain with that of the human HL-60 cell 1 alpha,25(OH)(2)D-3 nuclear receptor. Both l igand binding domains could equivalently accommodate to the presence o f (1) a side-chain cyclopropyl group, (2) 22-ene or 23-yne, (3) length ening the side chain by two carbons, (4) presence of four to six fluor ine atoms, (5) substitution of an oxygen for carbon 22, and (6) presen ce of a 22-[m-(dimethylhydroxymethyl)phenyl] aromatic group in the sid e chain. The DPB could tolerate better than the HL-60 cell receptor th e presence of a 22-(p-hydroxyphenyl) aromatic group in the side chain and the absence of the 1 alpha-hydroxyl. In contrast, the HL-60 cell r eceptor could tolerate better than the DBP the following structural mo difications: presence of a 16-ene, or 16-ene plus 23-yne unsaturation, and presence of an 11 beta-hydroxyl.