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Androgen Receptor Coregulators in Prostate Cancer

前列腺癌中的雄激素受体共调节因子

基本信息

批准号：
6778267
负责人：
JOSEPH D FONDELL
金额：
$ 27.21万
依托单位：
UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
依托单位国家：
美国
项目类别：
财政年份：
2001
资助国家：
美国
起止时间：
2001-09-30 至 2006-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (Provided by the applicant) Regulation of gene expression by the androgen receptor (AR) involves the association and action of transcriptional coregulatory proteins. Although a plethora of AR-interacting proteins have been identified, the physiological and pathological roles fulfilled by these factors in AR-mediated signaling pathways remain poorly understood. The goal of this application is to investigate and characterize the involvement of specific AR-coregulatory factors during normal prostate growth and the progression of prostate tumorigenesis. Given that prostate cancer cells are initially androgen-dependent and eventually progress into androgen-independent cells, we hypothesize that neoplastic changes in AR signaling pathways and/or the AR protein itself can abnormally affect the specific types of coregulatory protein complexes that bind to the receptor. To address these issues, we will generate stable FLAG epitope-tagged AR (f:AR) expressing cell lines from immortalized primary prostate cells (normal and malignant) and from metastatic prostate tumor cells. The lines will serve as biological tools with which we will immunoaffinity purify f:AR from hormone treated (and untreated) cells and subsequently examine and characterize the AR-associated proteins using a number of biochemical techniques. Our specific goals are to: (1) Determine whether distinct types of transcriptional coregulatory proteins are differentially associated with f:AR in normal versus malignant prostate cells. Stable f:AR-expressing prostate lines will be cultured in the presence (or absence) of distinct androgens and anti-androgens; f:AR-cofactor complexes will be purified and characterized by silver stain, Western blotting and mass spectrometry. (2) Determine whether androgen-independent signaling pathways induce f:AR-cofactor complex assembly in normal and malignant prostate cells. These studies will examine whether activation of specific receptor tyrosine kinases (previously implicated in prostate cancer and androgen-independent growth) can trigger specific f: AR-cofactor complex formation in the absence of AR ligands. (3) Determine whether pathologically associated mutations/polymorphisms in the AR gene affect f:AR-cofactor assembly. The f:AR cDNA will be subjected to site-directed mutagenesis and subsequently stably introduced into prostate cells. The mutated f:AR will be purified from ligand-treated cells and the associated cofactors identified and characterized. In summary, the studies outlined here should increase our fundamental understanding of the role of coregulatory factors in AR-mediated signaling pathways and potentially identify and define new targets for therapeutic agents in the treatment of prostate cancer.

描述（由申请人提供）雄激素受体（AR）对基因表达的调节涉及到关联和转录共调节蛋白的作用。虽然已经鉴定了过多的AR相互作用蛋白，生理和这些因子在AR介导的信号通路中所起的病理作用仍然知之甚少。该应用程序的目标是调查和表征正常过程中特定AR共调节因子的参与，前列腺生长和前列腺肿瘤发生的进展。鉴于前列腺癌细胞最初是雄激素依赖性的，雄激素非依赖性细胞，我们假设AR中的肿瘤性变化信号通路和/或AR蛋白本身可以异常影响与受体结合的特定类型的共调节蛋白复合物。到为了解决这些问题，我们将产生稳定的FLAG表位标记的AR（f：AR）表达来自永生化原代前列腺细胞的细胞系（正常和恶性）和转移性前列腺肿瘤细胞。这些线路将作为我们将用免疫亲和法从激素中纯化f：AR的生物工具，处理（和未处理）的细胞，随后检查和表征 AR相关蛋白的研究。我们的具体目标是：（1）确定不同类型的转录辅调节蛋白与正常对照组和正常对照组中的f：AR差异相关。恶性前列腺细胞稳定的表达f：AR的前列腺系将是在存在（或不存在）不同雄激素和抗雄激素的情况下培养; f：AR-辅因子复合物将被纯化并通过银染色表征，蛋白质印迹和质谱。(2)确定是否雄激素非依赖性信号通路诱导f：AR-辅因子复合物组装在正常和恶性前列腺细胞中。这些研究将探讨是否特异性受体酪氨酸激酶的激活（先前与前列腺癌和雄激素非依赖性生长）可以触发特异性F：在不存在AR配体的情况下AR-辅因子复合物的形成。(3)确定 AR基因中的病理相关突变/多态性是否影响 f：AR辅因子组装。将对f：AR cDNA进行定点扩增，诱变并随后稳定地引入前列腺细胞中。突变的 f：从配体处理的细胞和相关辅因子中纯化AR 识别和表征。总之，这里概述的研究应该增加我们对共调节因子在以下方面的作用的基本理解： AR介导的信号通路，并可能识别和定义新的靶点用于治疗前列腺癌的治疗剂。