|Budget Amount *help
¥3,100,000 (Direct Cost : ¥3,100,000)
Fiscal Year 1999 : ¥900,000 (Direct Cost : ¥900,000)
Fiscal Year 1998 : ¥900,000 (Direct Cost : ¥900,000)
Fiscal Year 1997 : ¥1,300,000 (Direct Cost : ¥1,300,000)
To clarify the structure-function relationship of vitamin D analogues, we synthesized novel vitamin D analogues and evaluated their biological activities.
(1) A series of singly dehydroxylated 19-nor-1α, 25-dihydroxyvitamin DィイD23ィエD2 A-ring analogues were synthesized. The analogues could be clearly divided into two categories ; one group, bearing 1α-hydroxy or 3β-hydroxy groups in the A-ring, were potent differentiators and the second group, bearing 1β-hydroxy or 3α-hydroxy groups, were potent stimulators of apoptosis. These findings will provide useful information not only for development of therapeutic agents for treatment of leukemia and other cancers.
(2) Biological activities of a series of 2β-substituted analogues of 1α, 25(OH)ィイD22ィエD2)DィイD23ィエD2 were evaluated. Binding affinity for VDR, transactivation potency on the target gene and VDR-mediated gene regulation of the hydroxyalkyl and hydroxyalkoxy 2β-substituted analogues were almost comparable to those of 1α, 25(OH)ィイD22ィエD2Dィ
イD23ィエD2, while the alkyl and alkenyl analogues were much less active than 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2. Furthermore, we found a promising candidate for preventive and therapeutic medicine of osteoporosis among these compounds.
(3) The biological activity of two sets of ring A stereo isomers of 2-methyl-1α, 25-dihydroxyvitamin DィイD23ィエD2 and 2-methyl-20-epi-1α, 25-dihydroxyvitamin DィイD23ィエD2 were evaluated. We found that 2β-methyl-20-epi-3-epi-1β, 25(OH)ィイD22ィエD2DィイD23ィエD2 is transcriptionally more active than 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2 despite lacking the 1α-hydroxyl group, which was believed to be essential for expressing VDR-mediated gene transcription. Since the C-20 natural counterpart, 2β-methyl-3-epi-1β, 25(OH)ィイD22ィエD2DィイD23ィエD2, was almost completely biologically inactive, 20-epimerization is probably responsible for activation of gene expression.
(4) We found that binding affinity 1α, 25(OH)ィイD22ィエD2DィイD22ィエD2 and 1α, 25(OH)ィイD22ィエD2DィイD24ィエD2 were almost comparable to the effects of 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2, while 24-epi-1α, 25(OH)ィイD22ィエD2DィイD22ィエD2 and 1α, 25(OH)ィイD22ィエD2DィイD27ィエD2 were much less active than 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2 in these respects. This is the first finding concerning biological assessment of 1α, 25(OH)ィイD22ィエD2DィイD22ィエD2, 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2, 1α, 25(OH)ィイD22ィエD2DィイD24ィエD2, 24-epi-1α, 25(OH)ィイD22ィエD2DィイD22ィエD2 and 1α, 25(OH)ィイD22ィエD2DィイD27ィエD2 at the molecular level, especially with regards to the structural differences at the 24R- or 24S-methyl group and a double bond between carbons 22 and 23 in the side chain of 1α, 25(OH)ィイD22ィエD2D derivatives.
(5) The effects of 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2 and its analogues on transmembrane CaィイD12+ィエD1 influx were examined in the growth phase of rat osteosarcoma ROS17/2.8 cells.The results suggest that 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2 and its analogues modulate transmembrane CaィイD12+ィエD1 influx in osteoblast-like cells by opening L-type CaィイD12+ィエD1 channels which can recognize 1α-hydroxy analogues as agonists and 1β-hydroxy analogues as antagonists.
(6) We demonstrated that 1α, 25(OH)ィイD22ィエD2DィイD23ィエD2-mediated suppressive effects on the inducible expression of cytokine genes in human T cells may, in part, be due to diminished activity of the transcription factor NFAT. The vitamin DィイD23ィエD2 receptor (VDR) specifically bound to the distal NFAT site in the human IL-2 promoter.