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The biological activity of 16-epoxy side-chain analogs of 1 alpha,25-dihydroxyvitamin D3, (1 alpha,25(OH)2D3) was evaluated in vitro and in vivo. Compared to 1 alpha,25(0H)2D3, all analogs had lower affinities for the pig duodenal vitamin D receptor and also for the human serum vitamin D binding protein. The in vitro effects on cell proliferation or differentiation of human promyeloid leukemia (induction of superoxide production in HL-60 cells), human osteosarcoma MG-63 cells (osteocalcin secretion), or human breast cancer cells (incorporation of thymidine in MCF-7 cells), was markedly inhibited by several epoxy analogs, compared to 1 alpha,25(OH)2D3, but the rank order of their activity widely varied among different cancer cells. The most potent analogs (24S,25S-24-hydroxy-25,26-epoxy-22-ene-1 alpha-OHD3, 25,26-epoxy-23-yne-1 alpha-OHD3, and 25,26-epoxy-23-yne-20-epi-1 alpha-OHD3 or compounds, 16, 5, and 7, respectively) were equipotent (16 and 5) or 30-fold (compound 7 on MG-63 cells) to 40-fold (compound 7 on MCF-7 cells) more active than 1 alpha,25-(OH)2D3. These analogs were nevertheless poorly antirachitic (3%) when tested in vitamin D-deficient chicks (using serum and bone calcium, serum osteocalcin and duodenal calbindin D-28K, as end points). Compound 7 was also 100-fold more active than 1 alpha,25-(OH)2D3 in inhibition of proliferation of human foreskin keratinocytes. Some epoxy analogs of 1 alpha,25-(OH)2D3 thus display interesting dissociations between their receptor affinity and their potency to induce cell differentiation, whereas their effect on cell proliferation/differentiation exceed their calcemic effects more than 100- to 1000-fold.

Maternal and fetal 1,25-dihydroxyvitamin D 3 (l,25(OH) 2 D 3 ) and osteocalcin were measured in guinea pigs, to examine their potential use as animal models for fetal bone development and calcium homeostasis. Measurements were performed on days 42, 57 and 63 of gestation. Maternal serum total l,25(OH) 2 D 3 concentrations were increased only at the end of gestation (day 63). However, because the vitamin D binding protein (DBP) and albumin levels were decreased by 35–50% from day 42 onwards, the unbound l,25(OH) 2 D 3 , calculated as the l,25(OH) 2 D 3 /DBP molar ratio, was increased before day 63. Osteocalcin concentrations during gestation were 50–54% of levels found in nongravid animals. Fetal serum total l,25(OH) 2 D 3 concentrations were 20% of those in maternal guinea pigs. Since DBP levels were only 9–15% of maternal levels, the unbound l,25(OH) 2 D 3 was consistently higher in fetuses, from day 42 onwards. There was a rise in total and unbound l,25(OH) 2 D 3 between days 57 and 63 of fetal life. Osteocalcin concentrations were higher in fetal than in adult guinea pigs, and reached peak values on day 57 (1023 μ g/l, i.e. 4.2 times higher than in adult female guinea pigs). Fetuses of guinea pigs that had received a restricted food supply for 14 days (days 49–63) had normal l,25(OH) 2 D 3 concentrations, but decreased osteocalcin concentrations compared with normal fetuses. The data obtained in fetal guinea pigs are comparable with those found in human fetuses, and suggest that the guinea pig may be a suitable model for studies on fetal bone and mineral development.

The synthesis includes the side-chain construction starting from the Inhoffen-Lythgoe diol and coupling with the A ring. Both 23-oxa- and 23-thia-analogues showed a decreased cell differentiating effect but even a more decreased calcemic effect compared with 1α,25-(OH) 2 D 3 .

The hormone 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) generates biological responses via both genomic and nongenomic mechanisms. This article addresses activity differences between the 6-s-trans (extended) and the 6-s-cis (steroid-like) conformation of 1,25-(OH)2D3 to initiate these responses. Because of facile interconversion of the 6-s-trans and 6-s-cis conformers of 1,25-(OH)2D3, kinetically competent amounts of both conformers exist to interact with any potential receptors for 1,25-(OH)2D3. We have chemically synthesized 1,25-(OH)2-9,14,19,19,19-pentadeuterio-pre-D3 (1,25-(OH)2-d5-pre-D3), an analog of the 6-s-cis conformation of 1,25-(OH)2D3. We found that 1,25-(OH)2-d5-pre-D3 and 1,25-(OH)2D3 were equivalently active in two nongenomic systems (transcaltachia as measured in the perfused chick intestine and 45Ca2+ uptake through voltage-gated Ca2+ channels in ROS 17/2.8 cells). 1,25-(OH)2-d5-pre-D3 was significantly less active both in binding in vitro to the plasma vitamin D-binding protein (7%) and to the chick (10%) and pig (4%) intestinal nuclear 1,25-(OH)2D3 receptors generating genomic biological responses in vivo (induction of plasma levels of osteocalcin, < 5%) or in cultured cells (inhibition of HL-60 cell differentiation, < 5%; inhibition of MG-63 proliferation, < 2%; and induction of osteocalcin, < 2%). These results suggest that the genomic and nongenomic responses are mediated by separate receptors. Further, the 6-s-cis form (steroid-like conformation) of the natural hormone, 1,25-(OH)2D3, may be selectively responsible for its nongenomic function(s).