Regulation of Intratumor Androgen/Androgen Receptor signaling in Prostrate Cancer

前列腺癌瘤内雄激素/雄激素受体信号传导的调节

• 批准号: 8811322
• 负责人: BANDANA CHATTERJEE
• 金额: --
• 依托单位: SOUTH TEXAS VETERANS HEALTH CARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2017-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8811322
• 关键词: Accounting; African American; Agonist; Anabolism; Androgen Antagonists; Androgen Receptor; Androgens; Antibodies; Attenuated; Automobile Driving; Beryllium; Biochemical; Biological Assay; Biopsy; Calcitriol; Chemotherapy-Oncologic Procedure; Cholesterol; Chromatin; Clinical; Computer Simulation; CpG Islands; DNA; DNA Methylation; Diagnosis; Drug Targeting; Elements; Enzymes; Epithelial Cell Proliferation; Foundations; Gene Expression; Genes; Genomic DNA; Goals; Growth; Histones; Homeostasis; Hormonal; Human; Hypermethylation; In Vitro; Indium; Indolent; Intervention; Knockout Mice; Laboratories; Lead; Ligand Binding; Link; Liver; Locally Advanced Malignant Neoplasm; Lysine; Malignant Neoplasms; Malignant neoplasm of prostate; Mediating; Medicine; Messenger RNA; Methods; Methylation; Methyltransferase; Modeling; Molecular; Mus; Neoplasm Metastasis; Nuclear Receptors; Pathologist; Pathway interactions; Patients; Pharmaceutical Preparations; Production; Prostate; Prostatic Neoplasms; Proteins; Race; Receptor Signaling; Recurrence; Regulation; Regulator Genes; Relative (related person); Reporting; Risk; Role; SULT2B1; Sampling; Serinus; Signal Pathway; Site; Specimen; Staining method; Stains; Steroids; Sterols; Steryl-sulfatase; Subfamily lentivirinae; TNFRSF10A gene; Tissue Microarray; Tissues; Transactivation; Tumor Tissue; Vitamin D3 Receptor; Western Blotting; base; cancer cell; castration resistant prostate cancer; college; dehydroepiandrosterone; dehydroepiandrosterone sulfotransferase; design; in vivo; knock-down; mutant; novel; novel strategies; outcome forecast; oxysterol binding protein; predictive marker; prevent; promoter; prostate cancer cell; public health relevance; pyrosequencing; receptor; receptor binding; response; stem; sulfation; sulfotransferase; synergism; tumor; tumor growth; tumor progression; tumor xenograft

DESCRIPTION (provided by applicant): Challenges in prostate cancer management stem from 1) inability to distinguish indolent prostate cancer from aggressive malignancy with metastatic potential; 2) limited treatment options for recurrent, castration-resistant prostate cancer (CRPC). Reactivated androgen receptor (AR) in CRPC results from elevated expression and activity of AR, and increased de novo androgen biosynthesis from DHEA. Anti-AR/anti-androgen biosynthesis drugs inhibit post chemotherapy cancer for a few months, but response is not universal. Finding new predictors for cancer progression and new drug targets are high-priority goals. We will examine a novel aspect of androgen-activated AR signaling that was revealed from our study. This entails i) regulation of androgen homeostasis by the prostate-expressed SULT2B, a sulfotransferase which mediates sulfation of cholesterol and DHEA, and ii) enhanced activity of methylated AR. We show 1) SULT2B mRNA and protein levels are markedly reduced in clinical specimens and IHC staining of tissue microarrays revealed highly statistically significant (p<0.001) loss of SULT2B in cancer tissue; 2) SULT2B silencing caused increased proliferation of prostate cancer cells; 3) Vitamin D receptor (VDR) and the oxysterol-inducible liver X receptor (LXR) can induce the SULT2B gene (SULT2B1); 4) AR is monomethylated at a lysine residue in its hinge domain by the SET9 lysine methyltransferase. SET9 depletion reduced wild type but not methylation-site-mutant AR activity; 5) LSD1, a histone lysine demethylase can convert methylated AR to unmethylated AR; 6) LXR-¿ is a potential regulator of genes encoding SET9 and LSD1, since LXR-responsive cis elements are present in these two genes based on in silico analysis. We have developed an antibody that can specifically recognize lysine- methylated AR by western blotting. We hypothesize that advanced prostate cancer associates with reduced SULT2B and elevated methyl-modified AR, LXR-¿ inhibits prostate cancer in part by preventing loss of SULT2B and elevation of methylated AR, and VDR synergizes LXR effects. The hypothesis is built from our own results and from reports that i) LXR- -/- mice show heightened androgen sensitivity in ventral prostates, but no change in AR levels; ii) steroid sulfatase (Sts) which counters DHEA sulfation, was induced in prostates of Lxr- -/- mice and suppressed in LXR- -activated mice; iii) LXR inhibited prostate cancer in xenograft tumors. We propose to determine: Aim 1) significance of SULT2B loss in prostate cancer, and a role for DNA methylation in this loss. We will examine i) diverse specimens for association of low SULT2B in cancer tissue with i) DHT levels and AR target gene expression, ii) levels of the enzymes AKR1C3 & SRD5A1, and iii) methylation status of the SULT2B1 gene. Statistical association of clinical variables (including role of race) with above parameters will be explored. Aim 2) the dynamics of LXR- signaling with SULT2B-regulated androgen homeostasis in prostate, possible synergy with VDR and their relevance in prostate cancer. We will investigate i) LXR-responsive DNA cis element(s) in SULT2B1 and its interaction with LXR- , coregulators, chromatin modifiers; ii) Synergy of LXR- with VDR in SULT2B induction and prostate cancer inhibition; iii) DHT levels in LXR- -silenced prostate cancer cells in vitro & in vivo, and in Lxr- -null mouse prostates and reversal of the effect of LXR silencing by calcitriol; iv) a link in clinical specimes between low SULT2B and attenuated expression of LXR- and LXR- -targeted lipogenic genes. Aim 3) significance of AR methylation in prostate cancer and its interplay with LXR-¿. We will examine i) methylated AR levels in LXR- -knocked down cancer cells in vitro & in vivo; ii) a role for LXR- in SET9 & LSD1 expression and in their association with AR iii) clinical specimens for elevated methylated AR levels (relative to total AR). Methods: IHC, tissue microarray, LCM, western blot, pyrosequencing, bisulfate-modified DNA, promoter assay, ChIP,, lentivirus, si RNAs, LC- MS/MS. Molecular biologists, pathologists and biostatisticians will collaborate. Multiple cores will be used. Significance: This study may reveal new predictive markers and new intervention targets to prevent CRPC.

描述（由申请人提供）： 前列腺癌管理的挑战源于1）无法区分惰性前列腺癌与具有转移潜力的侵袭性恶性肿瘤; 2）复发性去势抵抗性前列腺癌（CRPC）的治疗选择有限。CRPC中雄激素受体（AR）的再活化是由于AR的表达和活性升高以及DHEA从头雄激素生物合成增加所致。抗AR/抗雄激素生物合成药物抑制化疗后癌症几个月，但反应不是普遍的。寻找癌症进展的新预测因子和新的药物靶点是高度优先的目标。我们将研究从我们的研究中揭示的雄激素激活的AR信号的一个新方面。这需要i）通过前列腺表达的SULT 2B（一种介导胆固醇和DHEA硫酸化的磺基转移酶）调节雄激素稳态，和ii）增强甲基化AR的活性。我们发现：1）SULT 2B mRNA和蛋白水平在临床标本中显著降低，组织微阵列的IHC染色显示高度统计学显著性（p<0.001）SULT 2B在癌组织中的缺失; 2）SULT 2B沉默引起前列腺癌细胞增殖增加; 3）维生素D受体（VDR）和氧化固醇诱导的肝X受体（LXR）可以诱导SULT 2B基因（SULT 2B 1）; 4）AR在其铰链结构域中的赖氨酸残基处被SET 9赖氨酸甲基转移酶单甲基化。SET 9缺失降低野生型但不降低甲基化位点突变体AR活性; 5）LSD 1，一种组蛋白赖氨酸脱甲基酶，可将甲基化AR转化为未甲基化AR; 6）LXR-Δ是编码SET 9和LSD 1的基因的潜在调节因子，因为基于计算机模拟分析，LXR响应性顺式元件存在于这两个基因中。我们已经开发了一种抗体，可以特异性地识别赖氨酸甲基化AR的蛋白质印迹。我们假设晚期前列腺癌与SULT 2B减少和甲基化AR升高有关，LXR-1通过阻止SULT 2B的丢失和甲基化AR的升高部分抑制前列腺癌，VDR协同LXR效应。这一假设是根据我们自己的结果和以下报告建立的：i）LXR- -/-小鼠腹侧前列腺显示出增强的雄激素敏感性，但AR水平没有变化; ii）对抗DHEA硫酸化的类固醇硫酸酯酶（Sts）在LXR- -/-小鼠的前列腺中被诱导，在LXR-激活的小鼠中被抑制; iii）LXR抑制异种移植肿瘤中的前列腺癌。我们建议确定：目的1）SULT 2B丢失在前列腺癌中的意义，以及DNA甲基化在这种丢失中的作用。我们将检查i）癌症组织中低SULT 2B与i）DHT水平和AR靶基因表达，ii）酶AKR 1C 3和SRD 5A 1的水平，以及iii）SULT 2B 1基因的甲基化状态的关联的不同标本。将探索临床变量（包括人种的作用）与上述参数的统计学关联。目的2）LXR信号转导与SULT 2B调节前列腺雄激素稳态的动力学，与VDR的可能协同作用及其在前列腺癌中的相关性。我们将研究i）SULT 2B 1中的LXR响应性DNA顺式元件及其与LXR-、辅助调节剂、染色质修饰剂的相互作用; ii）LXR-与VDR在SULT 2B诱导和前列腺癌抑制中的协同作用; iii）LXR-沉默的前列腺癌细胞中的DHT水平（体外和体内）以及LXR-缺失小鼠前列腺中的DHT水平和骨化三醇对LXR沉默效应的逆转; iv）低SULT 2B与LXR-和LXR-靶向的脂肪生成基因的减弱表达之间的临床关联。目的3）AR甲基化在前列腺癌中的意义及其与LXR-γ的相互作用。我们将检查i）LXR中的甲基化AR水平-在体外和体内敲低癌细胞; ii）LXR在SET 9和LSD 1表达中的作用及其与AR的关联iii）临床标本中甲基化AR水平升高（相对于总AR）。研究方法：IHC、组织微阵列、LCM、蛋白质印迹、焦磷酸测序、硫酸氢盐修饰的DNA、启动子测定、ChIP、慢病毒、siRNA、LC-MS/MS。分子生物学家、病理学家和生物统计学家将合作。将使用多个核心。意义：这项研究可能揭示新的预测标志物和新的干预靶点，以预防CRPC。