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Vitamin D in Prostate Cancer: Tumor Vasculature Effects

维生素 D 在前列腺癌中的作用：肿瘤脉管系统的影响

基本信息

批准号：
7882288
负责人：
CANDACE S JOHNSON
金额：
$ 27.72万
依托单位：
ROSWELL PARK CANCER INSTITUTE CORP
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-09-01 至 2013-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7882288
关键词：
25-hydroxycholecalciferol-24-hydroxylase Address Affect Apoptosis Apoptotic Binding Proteins Biological Availability Biological Models Breast Carcinoma Calcitriol Catabolism Cell Cycle Arrest Cell Death Cells Characteristics Conditioned Culture Media CpG Islands Cytotoxic agent DNA DNA Methylation DNA Methyltransferase DNA Methyltransferase Inhibitor DNA Modification Methylases Data Defect Deoxycytidine Dexamethasone Dihydroxycholecalciferols Endothelial Cells Endothelium Enzymes Epigenetic Process Event Fibroblasts G1 Arrest Gene Expression Gene Silencing Gene Targeting Genes Genome Scan Glucocorticoids Human Incubated Kinetics Laboratories Lasers Lead Malignant Neoplasms Malignant neoplasm of prostate Mammary Gland Parenchyma Mediating Messenger RNA Metabolism Methods Methylation Microscopy Minerals Mus Normal Cell Normal tissue morphology Pattern Pharmaceutical Preparations Phase II Clinical Trials Phosphotransferases Population Prostatectomy Proteins Resistance Role Screening procedure Signal Pathway Signal Transduction Signaling Molecule System Therapeutic Time Tissues Tumor-Derived Tumor-Suppressor Gene Inactivation Vitamin D Vitamin D3 Receptor androgen independent prostate cancer angiogenesis base bone enzyme activity histone modification in vitro activity in vivo matrigel men neoplastic cell phase 1 study promoter receptor tumor tumor xenograft

项目摘要

DESCRIPTION (provided by applicant): Calcitriol (vitamin D or 1,25 dihydroxycholecalciferol), a central factor in bone and mineral metabolism, has significant antitumor activity in vitro and in vivo. Induction of CYP24, the enzyme primarily responsible for calcitriol catabolism, may be a factor in the anti-proliferative activity pre-clinically and clinically. In addition, the endothelial cells in tumors are sensitive to calcitriol and uniquely modulate CYP24 expression through epigenetic changes. Epigenetic events affect gene expression without alteration in DNA gene sequence and lead to transcriptional gene silencing and inactivation of tumor suppressor genes in human cancer. While many studies document epigenetic changes in tumor cells, only limited data support a role for epigenetic changes in the "normal" cells found in the tumor microenvironment. Calcitriol also inhibits proliferation of endothelial cells and can inhibit angiogenesis in a number of tumor model systems. We have established a method for the isolation of fresh, tumor-derived endothelial cells (TDEC) that maintain phenotypic characteristics which are distinct from endothelial cells isolated from normal tissues and from Matrigel plugs (MDEC). In TDEC, calcitriol induces G0/G1 arrest, modulates p27 and p21, and induces apoptotic cell death and decreases P-Erk and P-Akt. In contrast, endothelial cells isolated from normal tissues and MDEC are unresponsive to calcitriol-mediated anti-proliferative effects despite intact signaling through the vitamin D receptor (VDR). Differences may be due to the over-expression of CYP24 in MDEC where mRNA, protein and enzymatic activity for CYP24 are markedly increased. In TDEC, which is sensitive to calcitriol, the CYP24 promoter is hypermethylated in two CpG island regions located at the 5' end, which may contribute to gene silencing of CYP24 in TDEC. The extent of methylation in these two regions is significantly less in MDEC. Treatment of TDEC with the DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine, restores calcitriol-mediated induction of CYP24 and resistance to calcitriol. Lastly, when MDEC are incubated with conditioned media from tumor cells for 14-21 days, these cells regain sensitivity to the antiproliferative effects of calcitriol, CYP24 induction is silenced and regions 1 and 2 of the CYP24 promoter are hypermethylated. The characteristics of these cells are similar to the endothelial cells found in tumors or TDEC. These unique differences in the epigenetic silencing of calcitriol-induced CYP24 gene expression in endothelial cells from the tumor microenvironment directly impact on calcitriol-mediated signaling pathways and ultimately on therapeutic application. Therefore, we propose 1) to elucidate the epigenetic signal transduction mechanism(s) that lead to the epigenetic silencing of CYP24; 2) to determine whether epigenetic silencing of CYP24 occurs in human endothelial cell and tumor cell populations isolated from human prostate cancer; and 3) To identify other potential targets of DNA methylation in TDEC vs. MDEC by investigating the scope of the epigenetic defect from the tumor microenvironment on the endothelial cells. We developed a model system for isolation of endothelial cells freshly from tumors and demonstrate that CYP24, the catabolic enzyme involved in vitamin D (1,25 dihydroxycholecalciferol or calcitriol) signaling, becomes epigenetically silenced selectively in tumor-derived endothelial cells. These unique differences in the epigenetic silencing of calcitriol-induced CYP24 gene expression in endothelial cells from the tumor microenvironment directly impact on calcitriol-mediated signaling pathways and ultimately on therapeutic application.

描述（由申请人提供）：骨化三醇（维生素D或1，25二羟基胆钙化醇）是骨和矿物质代谢的中心因子，在体外和体内具有显著的抗肿瘤活性。诱导CYP 24（主要负责骨化三醇催化的酶）可能是临床前和临床抗增殖活性的一个因素。此外，肿瘤中的内皮细胞对骨化三醇敏感，并通过表观遗传变化独特地调节CYP 24表达。表观遗传事件影响基因表达而不改变DNA基因序列，并导致人类癌症中转录基因沉默和肿瘤抑制基因失活。虽然许多研究记录了肿瘤细胞中的表观遗传变化，但只有有限的数据支持肿瘤微环境中发现的“正常”细胞中的表观遗传变化的作用。骨化三醇还抑制内皮细胞的增殖，并且可以抑制许多肿瘤模型系统中的血管生成。我们已经建立了一种分离新鲜的、肿瘤衍生的内皮细胞（TDEC）的方法，所述TDEC保持与从正常组织和基质胶塞（MDEC）分离的内皮细胞不同的表型特征。在TDEC中，骨化三醇诱导G 0/G1期阻滞，调节p27和p21，诱导凋亡性细胞死亡并降低P-Erk和P-Akt。相反，从正常组织和MDEC分离的内皮细胞对骨化三醇介导的抗增殖作用无反应，尽管通过维生素D受体（VDR）的信号传导完整。差异可能是由于MDEC中CYP 24的过度表达，其中CYP 24的mRNA、蛋白质和酶活性显著增加。在对骨化三醇敏感的TDEC中，CYP 24启动子在位于5'端的两个CpG岛区域高甲基化，这可能有助于TDEC中CYP 24的基因沉默。 MDEC中这两个区域的甲基化程度显著较低。用DNA甲基转移酶抑制剂5-氮杂-2 '-脱氧胞苷治疗TDEC，可恢复骨化三醇介导的CYP 24诱导和对骨化三醇的耐药性。最后，当MDEC与来自肿瘤细胞的条件培养基一起孵育14-21天时，这些细胞重新获得对骨化三醇的抗增殖作用的敏感性，CYP 24诱导被沉默，并且CYP 24启动子的区域1和2被高甲基化。这些细胞的特征与肿瘤或TDEC中发现的内皮细胞相似。肿瘤微环境中内皮细胞中骨化三醇诱导的CYP 24基因表达的表观遗传沉默的这些独特差异直接影响骨化三醇介导的信号传导途径，并最终影响治疗应用。因此，我们提出：1）阐明导致CYP 24表观遗传沉默的表观遗传信号转导机制; 2）确定CYP 24表观遗传沉默是否发生在从人前列腺癌分离的人内皮细胞和肿瘤细胞群体中;和3）通过研究肿瘤微环境中表观遗传缺陷的范围，确定TDEC与MDEC中DNA甲基化的其他潜在靶点，内皮细胞。我们开发了一种模型系统，用于从肿瘤中分离新鲜的内皮细胞，并证明CYP 24，参与维生素D（1，25二羟基胆钙化醇或骨化三醇）信号传导的分解代谢酶，在肿瘤衍生的内皮细胞中选择性地表观遗传沉默。肿瘤微环境中内皮细胞中骨化三醇诱导的CYP 24基因表达的表观遗传沉默的这些独特差异直接影响骨化三醇介导的信号传导途径，并最终影响治疗应用。