Elucidation of the mechanism of cancer cell metastasis and permeation using active vitamin D as a molecular base and development of therapeutic agents

在本研究中，我们通过体外和体内实验证明了1α，25-二羟基维生素D_3 （1α,25-D_3）及其衍生物具有预防和治疗癌症的作用。结果如下：(1)研究发现22-oxa-骨化三醇是一种临床用于治疗银屑病和继发性甲状旁腺功能亢进的低钙维生素D类似物，通过侧链脱水或a环C-3羟基异构化途径失活。此外，我们还发现上述途径中可能涉及的酶协同调节la，25-D_3的代谢。(2)我们证实，由于我们的结果缺乏对物种差异（从动物到人类）的适当解释，使用已知的维生素D代谢酶表达系统对维生素D化合物的生物活性筛选是有用的。(3)我们明确了la，25-D_3通过诱导G1阻滞抑制LLC-GFP细胞的生长，通过抑制MMP-2和-9的表达抑制癌细胞的转移，通过抑制血管生成诱导因子的表达抑制癌细胞的血管生成。(4)我们明确了利用VDR基因靶向小鼠作为评估癌细胞生长的动物模型，α，25-D_3可以抑制VDR阳性的LLC-GFP细胞的肿瘤生长，并且α，25-D_3的抗肿瘤作用不需要其钙化作用的参与，并且在超生理和正常血清浓度下，α，25-D_3都能有效抑制肿瘤生长。(5)在动物实验中发现22-oxa-骨化三醇抑制了LLC-GFP细胞的生长而不引起任何钙血症症状。通过本研究，我们建立了在基因、细胞和动物水平上筛选活性维生素D抗癌活性的特异性实验模型体系。利用这些方法，生物学组和化学组之间的相互合作将有望创造出新的维生素D类似物，特别是具有优异的抗癌活性。在本研究中，我们通过体外和体内实验证明了1α，25-二羟基维生素D_3 （1α,25-D_3）及其衍生物具有预防和治疗癌症的作用。结果如下：(1)研究发现22-oxa-骨化三醇是一种临床用于治疗银屑病和继发性甲状旁腺功能亢进的低钙维生素D类似物，通过侧链脱水或a环C-3羟基异构化途径失活。此外，我们还发现上述途径中可能涉及的酶协同调节1α，25-D_3的代谢。(2)我们证实，由于我们的结果缺乏对物种差异（从动物到人类）的适当解释，使用已知的维生素D代谢酶表达系统对维生素D化合物的生物活性筛选是有用的。(3)阐明了1α，25-D_3通过诱导G1阻滞抑制LLC-GFP细胞生长，通过抑制MMP-2和-9表达抑制癌细胞转移，通过抑制血管生成诱导因子表达抑制癌细胞血管生成。(4)我们明确利用VDR基因靶向小鼠作为评估癌细胞生长的动物模型，1α，25-D_3抑制VDR阳性的lc - gfp细胞的肿瘤生长，且1α，25-D_3的抗肿瘤作用不需要其钙化作用的参与，在超生理和正常血清浓度下，1α，25-D_3都能有效抑制肿瘤生长。(5)在动物实验中发现22-oxa-骨化三醇抑制了LLC-GFP细胞的生长而不引起任何钙血症症状。通过本研究，我们建立了在基因、细胞和动物水平上筛选活性维生素D抗癌活性的特异性实验模型体系。利用这些方法，生物学组和化学组之间的相互合作将有望创造出新的维生素D类似物，特别是具有优异的抗癌活性。少

In the present study, we demonstrated from the in vitro and in vivo studies that 1α,25-dihydroxyvitamin D_3 (1α,25-D_3) and its derivatives are effective for the prevention and treatment of cancer. The results were follows. (1) It was found that 22-oxa-calcitriol, a low calcemic vitamin D analogue used clinically for the treatment of psoriasis and secondary hyperparathyroidism, is inactivated via through the pathway of either side-chain dehydration or isomerization of a hydroxy group at C-3 of the A-ring. In addition, it was found that the putative enzymes involved in the above pathways regulate the metabolism of la,25-D_3 cooperatively. (2) We confirmed that due to the lack of appropriate explanation for species difference (from animals to humans) of our results, employments of known-vitamin D metabolic enzymes expression systems were useful for the screening of the biological activity of vitamin D compounds. (3) We clarified that la,25-D_3 inhibits the growth of LLC-GFP cells by thro … More ugh the induction of G1 arrest, and inhibits the metastasis of cancer cells via the inhibition of MMP-2 and -9 expression, and suppress the angiogenesis of cancer cells via the inhibition of the expression of angiogenesis-inducing factors. (4) We clarified that using VDR gene targeting mice as a an animal model for the evaluation of cancer cell growth, l a,25-D_3 suppressed the tumor growth of LLC-GFP cells, which are VDR-positive, and the anti-tumorigenesis of l a,25-D_3 did not require the involvement of its calcemic action, and at both supraphysiological and normal serum concentrations, 1α,25-D_3 effectively suppressed the tumor growth. (5) I was found that 22-oxa-calcitriol suppressed the LLC-GFP cell growth without inducing any calcemic symptoms in animals. By the present study, we established the distinguished experimental model system for the screening of the anti-cancer activity of active vitamin D at gene, cellular and animal levels. Using these methods, it can be expected that the inter-cooperation between biology group and chemistry group will creates novel vitamin D analogues, especially having superior anti-cancer activity.In the present study, we demonstrated from the in vitro and in vivo studies that 1α,25-dihydroxyvitamin D_3 (1α,25-D_3) and its derivatives are effective for the prevention and treatment of cancer. The results were follows. (1)It was found that 22-oxa-calcitriol, a low calcemic vitamin D analogue used clinically for the treatment of psoriasis and secondary hyperparathyroidism, is inactivated via through the pathway of either side-chain dehydration or isomerization of a hydroxy group at C-3 of the A-ring. In addition, it was found that the putative enzymes involved in the above pathways regulate the metabolism of 1α,25-D_3 cooperatively. (2)We confirmed that due to the lack of appropriate explanation for species difference (from animals to humans) of our results, employments of known-vitamin D metabolic enzymes expression systems were useful for the screening of the biological activity of vitamin D compounds. (3)We clarified that 1α,25-D_3 inhibits the growth of LLC-GFP cells by through the induction of G1 arrest, and inhibits the metastasis of cancer cells via the inhibition of MMP-2 and -9 expression, and suppress the angiogenesis of cancer cells via the inhibition of the expression of angiogenesis-inducing factors. (4)We clarified that using VDR gene targeting mice as a an animal model for the evaluation of cancer cell growth, 1α,25-D_3 suppressed the tumor growth of LLC-GFP cells, which are VDR-positive, and the anti-tumorigenesis of 1α,25-D_3 did not require the involvement of its calcemic action, and at both supraphysiological and normal serum concentrations, 1α,25-D_3 effectively suppressed the tumor growth. (5)I was found that 22-oxa-calcitriol suppressed the LLC-GFP cell growth without inducing any calcemic symptoms in animals. By the present study, we established the distinguished experimental model system for the screening of the anti-cancer activity of active vitamin D at gene, cellular and animal levels. Using these methods, it can be expected that the inter-cooperation between biology group and chemistry group will creates novel vitamin D analogues, especially having superior anti-cancer activity. Less