A Novel Synthetic Androgen Receptor Antagonist

一种新型合成雄激素受体拮抗剂

基本信息

批准号：
8301019
负责人：
Zhongyun Dong
金额：
$ 31.4万
依托单位：
UNIVERSITY OF CINCINNATI
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-09-09 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8301019
关键词：
Ablation Agonist Amino Acids Androgen Analogues Androgen Antagonists Androgen Receptor Androgens Animal Model Animals Bicalutamide Binding Binding Sites Biochemical Biological Assay Breast Cancer Cell Castration Cells Clinic Competitive Binding Complex DU145 Development Disease Docking Down-Regulation Estradiol Flutamide Gene Expression Gene Targeting Genes Genetic Transcription Goals Green Fluorescent Proteins Growth Health Heat-Shock Proteins 70 Hormonal Human In Vitro LNCaP Laboratories Letters Ligand Binding Ligand Binding Domain MCF7 cell Malignant neoplasm of prostate Mediating Messenger RNA Molecular Chaperones Mus Mutation Names Nilutamide Nuclear PC3 cell line Pathway interactions Pharmaceutical Preparations Promoter Regions Property Prostate-Specific Antigen Proteins Receptor Signaling Refractory Reporter Research Resistance Running Stanolone Structure Testing Therapeutic Therapeutic Effect Translating Ubiquitination base cancer cell cell growth cellular engineering chromatin immunoprecipitation clinical application cofactor deprivation expression vector interest multicatalytic endopeptidase complex mutant novel overexpression palliative promoter protein expression receptor receptor binding receptor expression tumor tumor growth tumor progression ubiquitin-protein ligase

项目摘要

DESCRIPTION (provided by applicant): Current hormonal ablation therapy, the mainstay treatment for advance prostate cancer, is palliative, due the development of androgen-independent growth. Androgen receptor (AR) is expressed in most prostate cancer cells and AR overexpression of AR is sufficient and necessary for androgen-independent growth, which provides strong rationale for developing novel therapies against advanced prostate cancer through downregulation of AR. Dr. Dong's laboratory has identified a novel synthetic compound 6-amino-2-[2-(4-tert- butyl-pnenoxy)-ethylsulfanyl]-1H-pyrimidin-4-one, named DL3 for simplicity, as a potent AR antagonist. DL3 inhibits dihydrotestosterone (DHT)-stimulated cell growth and gene expression in all prostate cancer cell lines tested, including androgen-independent cells and cells resistant to antiandrogens, flutamide (Flut) or nilutamide (Nilut). The inhibitory effects of DL3 are more potent than bicalutamide (Bical), Flut, and Nilut. It inhibits neither AR nuclear localization nor DHT-induced AR NH2-terminus and COOH-terminus interaction and has no detectable AR agonist activity. DL3 reduces AR stability and downregulates of AR protein expression. It competes with DHT but not estradiol for the binding to cells. Docking analysis using protein crystal structure of AR ligand-binding domain (LBD) implies that DL3 can bind to the LBD. These observations prompted Dr. Dong to hypothesize that DL3 is a novel AR antagonist that binds to AR and induces the formation of inactive AR transcription machinery and AR degradation, and, hence, interrupts AR signaling. Three specific aims are proposed to test the hypothesis and to investigate efficacy of DL3 therapy against human prostate cancer cells in animals. In the specific aim 1, he will characterize the binding of DL3 to AR and identify amino acid residues of the DL3 binding site. He will determine the biochemical properties of the binding, investigate whether DL3 competes with DHT for AR binding, and identify amino acid residues of AR that interact with DL3. In the specific aim 2, he will investigate effects of DL3 on proteosome-mediated degradation of AR and assembly of AR transcription complex. He will determine effects of DL3 and Bical on AR stability, ubiquitination, and association with E3 ubiquitin ligase. By using the chromatin immunoprecipitation (ChIP) assay, he will investigate whether treatment with DL3 and Bical induce formation of transcription-inactive AR complex at the promoter region of prostate-specific antigen gene and identify cofactors in the complex. In the specific aim 3, Dr. Dong propose to investigate therapeutic effects of DL3 against human prostate cancer cells in mice. He will determine and compare effects of DL3 and Bical, alone or in combination with castration, on growth of tumors formed by both androgen-dependent and -independent cells and by cells refractory to Flut. He will determine DL3 distribution in tumor-bearing mice and correlate therapeutic effects with expression of AR and AR-target genes in tumors. These studies will firmly establish that DL3 is a novel AR antagonist and will enrich the understanding of mechanisms by which Bical interrupts AR signaling. Project Narrative Our preliminary studies have identified a novel synthetic compound 6-amino-2-[2-(4-tert- butyl-pnenoxy)-ethylsulfanyl]-1H-pyrimidin-4-one, named DL3 for simplicity. DL3 competitively inhibits the binding of DHT to human prostate cancer cells, but not estradiol to human breast cancer cells. It downregulates androgen receptor expression and suppresses androgen-induced cell growth and gene expression in prostate cancer cells. The proposed research will establish that DL3 is a novel AR antagonist, will elucidate mechanisms by which DL3 interrupts AR signaling, and will determine therapeutic effects of DL3 against human prostate cancer cells in mice. PUBLIC HEALTH RELEVANCE: Our preliminary studies have identified a novel synthetic compound 6-amino-2-[2-(4-tertbutyl-pnenoxy)-ethylsulfanyl]-1H-pyrimidin-4-one, named DL3 for simplicity. DL3 competitively inhibits the binding of DHT to human prostate cancer cells, but not estradiol to human breast cancer cells. It downregulates androgen receptor expression and suppresses androgen-induced cell growth and gene expression in prostate cancer cells. The proposed research will establish that DL3 is a novel AR antagonist, will elucidate mechanisms by which DL3 interrupts AR signaling, and will determine therapeutic effects of DL3 against human prostate cancer cells in mice.

描述（由申请人提供）：由于雄激素非依赖性生长的发展，当前的荷尔蒙消融疗法是预防前列腺癌的主要治疗方法。雄激素受体（AR）在大多数前列腺癌细胞中表达，AR的AR过表达对于雄激素独立的生长来说是足够且必需的，这为通过下调AR提供了针对晚期前列腺癌的新型疗法提供了强有力的理由。 Dong博士的实验室已经确定了一种新型的合成化合物6-氨基-2-- [2-（4-TERT-丁基 - 苯氧基） - 乙基磺胺基] -1H-吡啶蛋白-4-替代，命名为DL3，以简单性为DL3，作为一种有效的AR拮抗剂。 DL3在所有测试的前列腺癌细胞系中抑制二氢睾丸激素（DHT）刺激的细胞生长和基因表达，包括雄激素独立的细胞和对抗雄激素，氟丁酰胺（Flut）或尼罗二酰胺（Nilutamide）抗性的细胞。 DL3的抑制作用比Bicalutamide（Bical），Flut和Nilut更有效。它既不抑制AR核定位化，也不抑制DHT诱导的AR NH2末端和COOH-terminus相互作用，也没有可检测到的AR激动剂活性。 DL3降低了AR稳定性和AR蛋白表达的下调。它与DHT竞争，但没有雌二醇与细胞结合。使用AR配体结合结构域（LBD）的蛋白质晶体结构的对接分析意味着DL3可以与LBD结合。这些观察结果促使Dong博士假设DL3是一种新型的AR拮抗剂，它与AR结合并诱导了非活性AR转录机制和AR降解的形成，因此，中断AR信号传导。提出了三个具体目的，以检验假设并研究DL3治疗对动物中前列腺癌细胞的功效。在特定目标1中，他将表征DL3与AR的结合并鉴定DL3结合位点的氨基酸残基。他将确定结合的生化特性，研究DL3是否与DHT竞争AR结合，并鉴定与DL3相互作用的AR的氨基酸残基。在特定目标2中，他将研究DL3对AR蛋白体介导的AR和AR转录复合物组装的降解的影响。他将确定DL3和Bical对AR稳定性，泛素化和与E3泛素连接酶的关联的影响。通过使用染色质免疫沉淀（CHIP）测定，他将研究在前列腺特异性抗原基因的启动子区域的DL3和Bical诱导转录 - 活性AR复合物的形成，并鉴定综合体中的辅助因子。在特定的目标3中，董博士建议研究DL3对小鼠人类前列腺癌细胞的治疗作用。他将确定和比较DL3和Bical，单独或与cast割结合的影响，对雄激素依赖性和非依赖性细胞形成的肿瘤的生长以及对Flut难治性的细胞的生长。他将确定含有肿瘤小鼠的DL3分布，并将治疗作用与肿瘤中AR和AR-TARGET基因的表达相关。这些研究将牢固地确定DL3是一种新型的AR拮抗剂，并将丰富对Bical中断AR信号传导的机制的理解。项目叙述我们的初步研究已经确定了一种新型的合成化合物6-氨基-2-- [2-（4-TERT-丁基 - 苯甲酰氧基） - 乙基磺胺] -1H-羟基胺-4-one，称为DL3，以简单性为DL3。 DL3有竞争力抑制DHT与人前列腺癌细胞的结合，但不能抑制雌二醇与人类乳腺癌细胞的结合。它下调了雄激素受体的表达，并抑制了前列腺癌细胞中雄激素诱导的细胞生长和基因表达。拟议的研究将确定DL3是一种新型的AR拮抗剂，将阐明DL3中断AR信号的机制，并确定DL3对小鼠人类前列腺癌细胞的治疗作用。公共卫生相关性：我们的初步研究已经确定了一种新型的合成化合物6-氨基-2-- [2-（4- terttbutyl-pnenoxy） - 乙基磺胺] -1H-羟基酰胺素-4-one，称为DL3，称为DL3。 DL3有竞争力抑制DHT与人前列腺癌细胞的结合，但不能抑制雌二醇与人类乳腺癌细胞的结合。它下调了雄激素受体的表达，并抑制了前列腺癌细胞中雄激素诱导的细胞生长和基因表达。拟议的研究将确定DL3是一种新型的AR拮抗剂，将阐明DL3中断AR信号的机制，并确定DL3对小鼠人类前列腺癌细胞的治疗作用。