Vitamin D3, Interference with Androgen Signal and Inhibition of Prostate Cancer

维生素 D3、干扰雄激素信号和抑制前列腺癌

• 批准号: 8195918
• 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 2013-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8195918
• 关键词: Affinity; Age; Agonist; Alien; Androgen Antagonists; Androgen Receptor; Androgens; Animals; Antineoplastic Agents; Apoptosis; Bicalutamide; Binding; Binding Sites; Boxing; Breast; Cancer Etiology; Cell Cycle; Cell Proliferation; Cells; Cessation of life; Cholecalciferol; Chromatin; Chromatin Remodeling Factor; Clinical Management; Colon Carcinoma; Complex; DNA Methylation; Demography; Development; Dose; E2F Transcription Factor 1; E2F1 gene; Elderly; Elderly man; Elements; Epigenetic Process; Epithelial Cells; Gene Expression; Gene Silencing; Genomics; Glean; Growth; Health; Healthcare; Histone Deacetylase; Histones; Hormones; Human; Human body; Hypercalcemia; Immune; Knowledge; LAPC4; LNCaP; Laboratory Research; Lentivirus Infections; Life; Ligand Binding; Link; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of prostate; Mass Spectrum Analysis; Mediating; Metastatic Prostate Cancer; Methods; Mind; Mouse Protein; Mus; Normal Cell; Nucleic Acid Regulatory Sequences; PC3 cell line; Pathway interactions; Patients; Play; Population; Prevention strategy; Production; Prostate; Prostatic Neoplasms; Proteins; Recruitment Activity; Repression; Research; Resistance; Retinoblastoma Protein; Role; SCID Mice; Serum; Signal Transduction; Site; Technology; Therapeutic; Tissues; Toxic effect; Transfection; Transferase; Veterans; Vitamin D; Vitamin D Deficiency; Vitamin D3 Receptor; Xenograft procedure; analog; base; cancer cell; cancer risk; cancer therapy; cytotoxicity; design; disorder risk; drug development; gene repression; genetic regulatory protein; growth inhibitory proteins; human SLC25A5 protein; knock-down; men; neoplastic cell; novel; population based; prohibitin; promoter; protein expression; public health relevance; receptor binding; response; stem; transcription factor; tumor growth; tumor progression

DESCRIPTION (provided by applicant): Anti-proliferation activity of 1,25-(OH)2vitamin D3 (D3) in prostate cancer cells highlights the therapeutic potential of D3/D3 analogs. This potential is, however, limited by D3/D3 analog-induced hypercalcemia and hypercalciurea. Regulatory molecules and pathways, which are involved in D3-mediated proliferation inhibition of prostate cancer cells, can be targeted to inhibit tumor growth, presenting opportunities for therapeutic exploitation and development of new lines of defense against prostate cancer. Since metastatic prostate cancer is often associated with elevated levels of E2F1, which is a key cell cycle regulatory transcription factor, we designed this project to identify regulatory factors which, through cross-talk with androgen receptor (AR), vitamin D3 receptor (VDR) and E2F1, are integrally linked to the anti-proliferation action of D3. In novel findings we show that androgen-regulated E2F1 induction in human prostate cancer cells is interfered by D3 due to transcriptional repression. The corepressor Alien and anti-proliferative protein prohibitin appear to play roles in the repression. AR and VDR were present at the regulated promoter during both induction and de-induction of E2F1. Transcription factors WSTF and E47 occupied regulated chromatin in response to androgen. Prohibitin occupied this region in D3- and androgen+D3-treated cells; but Alien occupancy occurred only in androgen+D3 co-treated cells. Clinically important anti-androgen Casodex induced Alien recruitment. We hypothesize a) WSTF and E47 are part of an AR-associated coactivator complex, while D3-bound VDR promotes co-regulator switch to a prohibitin- and Alien-included corepressor assembly, which mediates E2F1 gene repression and reduced cell proliferation; b) E2F1 repression is aided by self inhibition at an auto-regulated site in the E2F1 promoter that results from enhanced E2F1 interaction with Alien, prohibitin or well-known E2F1 partner Rb (retinoblastoma protein), and with Alien/prohibitin/Rb-recruited corepressors. Four Specific Aims will examine: 1), roles of Alien, prohibitin, Rb, and associated corepressors in E2F1 activity in prostate cancer cells treated with androgen or D3 or androgen+D3; 2), interplay of Alien and prohibitin with WSTF and E47 in regulating E2F1 gene expression; 3), roles of endogenous WSTF/E47/Alien/prohibitin in androgen-stimulated and D3-inhibited prostate cancer cell proliferation and in the growth of xenograft prostate tumors in immune-deficient mice; 4), additional components of the holo corepressor complex, following affinity enrichment, for roles in E2F1 gene silencing and in the inhibition of prostate cancer cell proliferation. Methods: We will use Co-IP, chromatin IP, transfection, ShRNAs, inducible protein expression, lentivirus infection, cell proliferation, xenograft tumor growth in SCID mice, protein enrichment and mass spectrometry. Significance: Knowledge of the factors responding to the androgen-mediated proliferation stimulation, and to the D3-mediated anti-proliferation signaling will define mechanisms that may be targeted to develop novel prostate cancer therapeutics. The project has high relevance to health care issues of the Veterans population. PUBLIC HEALTH RELEVANCE: Project Narrative: Prostate cancer is a major health concern among elderly men, and metastatic prostate cancer is a leading cause of cancer death in men in the USA, second only to death from lung cancer. Risk for the disease rises from the 5th decade of life. The VA demography is steadily shifting to a late-life population, with a projection that by the year 2020 about 46% of the veteran population will reach age 65 or more. Vitamin D deficiency is a common problem among the elderly. Population based research has shown that low serum vitamin D levels in men correlate with increased prostate cancer risk. In the laboratory, research involving cells and animals has shown that the active form of vitamin D (known as vitamin D3) is very effective in reducing growth and proliferation of prostate tumor cells. Indeed, research in recent years revealed that high vitamin D levels protect human body from many cancers, such as cancer of the colon, breast and prostate. However, high-dose clinically effective D3 and synthetic compounds that act like D3 (known as D3 analogs) are highly toxic. Thus they cannot be used in prostate cancer treatment. A protein known as E2F1 plays a key role in promoting the growth and multiplication of normal cells. However, high activity of E2F1 and high levels of this protein promote excessive and uncontrolled multiplication of cells in the prostate and other tissues, causing cancer. Treatment-resistant prostate cancer tissues in a large number of patients are found to contain much higher levels of E2F1 than normal prostate tissues. We have designed a study to investigate why E2F1 activity and E2F1 protein levels increase in prostate cancer cells, and how D3 can inhibit this increase. We will undertake various analyses using cutting edge research technology to find out how specific regulatory protein molecules are influenced by D3 in such a way that they silence or reduce the activity and production of E2F1. Our study is expected to find alternative ways to reduce E2F1 activity and protein levels to circumvent use of high-dose D3. Information gleaned from our research may open up new strategies for prevention and clinical management of prostate cancer.

描述（由申请人提供）： 前列腺癌细胞中1,25-（OH）2Vitamin D3（D3）的抗增殖活性突出了D3/D3类似物的治疗潜力。然而，这种潜力受D3/D3类似物诱导的高钙血症和高钙尿素的限制。与D3介导的前列腺癌细胞增殖抑制有关的调节分子和途径可以针对抑制肿瘤的生长，为治疗性开发和开发针对前列腺癌的新型防御措施提供了机会。由于转移性前列腺癌通常与E2F1水平升高有关，这是一个关键的细胞周期调节转录因子，我们设计了该项目旨在鉴定通过与雄激素受体（AR），维生素D3受体（VDR）和E2F1进行串扰的调节因素，与D3的抗生育作用链接在一起。 在新的发现中，我们表明，由于转录抑制作用，D3干扰了人类前列腺癌细胞中雄激素调节的E2F1诱导。 Corepressor外星人和抗增殖性蛋白禁止素似乎在抑制中起着作用。在E2F1的诱导和去诱导期间，AR和VDR存在于受管制的启动子中。转录因子WSTF和E47响应雄激素占据了调节的染色质。在D3和雄激素+D3处理的细胞中占据了该区域。但是外星人占用仅发生在雄激素+D3共同处理的细胞中。临床上重要的抗雄激素Casodex诱导外星人募集。我们假设A）WSTF和E47是与AR相关的共激活剂复合物的一部分，而D3结合的VDR促进了共调节剂切换到不包括核心压力的核心压缩器组件，介导了E2F1基因抑制和减少细胞的增殖； b）E2F1抑制通过在E2F1启动子中自动调节的位点的自我抑制作用，这是由于E2F1与外星人，禁止素或众所周知的E2F1伴侣RB（视网膜细胞瘤蛋白）以及外星/普罗维蒂/Prouniditin/Rb-rb-recrunited Corepresser的相互作用而导致的。 将研究四个具体目的：1），在前列腺癌细胞中E2F1活性中外星人，禁止素，RB和相关的核心核心的作用，该细胞用雄激素或D3或D3或雄激素+D3处理； 2），在调节E2F1基因表达中，外星和禁止素与WSTF和E47的相互作用； 3），内源性WSTF/E47/Alien/provitin在雄激素刺激和D3抑制的前列腺癌细胞增殖以及免疫缺陷小鼠中异种移植前列腺肿瘤的生长中的作用； 4），在亲和力富集之后，Holo Corepressor复合物的其他成分在E2F1基因沉默和抑制前列腺癌细胞增殖中的作用。方法：我们将使用co-IP，染色质IP，转染，shRNA，诱导蛋白表达，慢病毒感染，细胞增殖，SCID小鼠中的异种移植肿瘤生长，蛋白质富集和质谱法。意义：了解响应雄激素介导的增殖刺激的因素以及D3介导的抗增殖信号传导的知识将定义可能针对开发新型前列腺癌疗法的机制。该项目与退伍军人人口的医疗保健问题有很高的相关性。 公共卫生相关性： 项目叙述：前列腺癌是老年男性的主要健康问题，转移性前列腺癌是美国男性癌症死亡的主要原因，仅次于肺癌死亡。这种疾病的风险从生命的第五个十年开始增加。 VA人口统计学正在稳步转移到晚年人口，预测，到2020年，大约46％的退伍军人人口将达到65岁或更高。 维生素D缺乏是老年人的普遍问题。基于人群的研究表明，男性血清维生素D水平较低与前列腺癌风险增加相关。在实验室中，涉及细胞和动物的研究表明，维生素D的活性形式（称为维生素D3）非常有效地减少前列腺肿瘤细胞的生长和增殖。实际上，近年来的研究表明，高维生素D水平可保护人体免受许多癌症的侵害，例如结肠癌，乳腺癌和前列腺癌。但是，像D3（称为D3类似物）一样的高剂量临床有效的D3和合成化合物是剧毒的。因此，它们不能用于前列腺癌治疗。 一种称为E2F1的蛋白质在促进正常细胞的生长和繁殖中起关键作用。但是，E2F1的高活性和高水平的蛋白质促进了前列腺和其他组织中细胞的过度和不受控制的繁殖，从而导致癌症。发现大量患者的耐治疗前列腺癌组织含有比正常前列腺组织更高的E2F1水平。 我们设计了一项研究，以研究为什么E2F1活性和E2F1蛋白水平在前列腺癌细胞中增加，以及D3如何抑制这种增加。我们将使用尖端研究技术进行各种分析，以了解特定的调节蛋白分子如何受到D3的影响，以使它们保持沉默或减少E2F1的活性和产生。我们的研究有望找到降低E2F1活性和蛋白质水平的替代方法，以规避使用高剂量D3的使用。从我们的研究中收集的信息可能会为预防和临床治疗前列腺癌开辟新的策略。