Miura D, Manabe K, Ozono K, Saito M, Gao Q, Norman A W, Ishizuka S
Safety Research Department, Teijin Institute for Bio-Medical Research, 4-3-2 Asahigaoka, Hino, Tokyo 191-8512, Japan.
J Biol Chem. 1999 Jun 4;274(23):16392-9. doi: 10.1074/jbc.274.23.16392.
We examined the effects of two novel 1alpha,25-dihydroxyvitamin D3-26,23-lactone (1alpha,25-lactone) analogues on human promyelocytic leukemia cell (HL-60) differentiation using the evaluation system of the vitamin D nuclear receptor (VDR)/vitamin D-responsive element (DRE)-mediated genomic action stimulated by 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3) and its analogues. We found that the 1alpha,25-lactone analogues (23S)-25-dehydro-1alpha-hydroxyvitamin-D3-26,23-lactone (TEI-9647), and (23R)-25-dehydro-1alpha-hydroxyvitamin-D3-26,23-lactone (TEI-9648) bound much more strongly to the VDR than the natural (23S, 25R)-1alpha,25(OH)2D3-26,23-lactone, but did not induce cell differentiation even at high concentrations (10(-6) M). Intriguingly, the differentiation of HL-60 cells induced by 1alpha,25(OH)2D3 was inhibited by either TEI-9647 or TEI-9648 but not by the natural lactone. In contrast, retinoic acid or 12-O-tetradecanoylphorbol-13-acetate-induced HL-60 cell differentiation was not blocked by TEI-9647 or TEI-9648. In separate studies, TEI-9647 (10(-7) M) was found to be an effective antagonist of both 1alpha,25(OH)2D3 (10(-8) M) mediated induction of p21(WAF1, CIP1) in HL-60 cells and activation of the luciferase reporter assay in COS-7 cells transfected with cDNA containing the DRE of the rat 25(OH)D3-24-hydroxylase gene and cDNA of the human VDR. Collectively the results strongly suggest that our novel 1alpha,25-lactone analogues, TEI-9647 and TEI-9648, are specific antagonists of 1alpha, 25(OH)2D3 action, specifically VDR/DRE-mediated genomic action. As such, they represent the first examples of antagonists, which act on the nuclear VDR.
我们使用由1α,25 - 二羟基维生素D3（1α,25(OH)2D3）及其类似物刺激的维生素D核受体（VDR）/维生素D反应元件（DRE）介导的基因组作用评估系统，研究了两种新型1α,25 - 二羟基维生素D3 - 26,23 - 内酯（1α,25 - 内酯）类似物对人早幼粒细胞白血病细胞（HL - 60）分化的影响。我们发现，1α,25 - 内酯类似物（23S）-25 - 脱氢 - 1α - 羟基维生素 - D3 - 26,23 - 内酯（TEI - 9647）和（23R）-25 - 脱氢 - 1α - 羟基维生素 - D3 - 26,23 - 内酯（TEI - 9648）与VDR的结合比天然的（23S, 25R）-1α,25(OH)2D3 - 26,23 - 内酯强得多，但即使在高浓度（10^(-6) M）下也不诱导细胞分化。有趣的是，TEI - 9647或TEI - 9648可抑制1α,25(OH)2D3诱导的HL - 60细胞分化，但天然内酯则无此作用。相反，维甲酸或12 - O - 十四烷酰佛波醇 - 13 - 乙酸酯诱导的HL - 60细胞分化未被TEI - 9647或TEI - 9648阻断。在单独的研究中，发现TEI - 9,647（10^(-7) M）是1α,25(OH)2D3（10^(-8) M）介导的HL - 60细胞中p21（WAF1, CIP1）诱导以及在转染了含大鼠25(OH)D3 - 24 - 羟化酶基因DRE的cDNA和人VDR cDNA的COS - 7细胞中荧光素酶报告基因测定激活的有效拮抗剂。总体而言，结果强烈表明我们的新型1α,25 - 内酯类似物TEI - 9647和TEI - 9648是1α, 25(OH)2D3作用的特异性拮抗剂，特别是VDR/DRE介导的基因组作用。因此，它们代表了作用于核VDR的拮抗剂的首个实例。
