Genetic Dissection of Context-Dependent and Opposing Roles of Androgen Receptor

雄激素受体的背景依赖性和对立作用的基因剖析

• 批准号: 8067922
• 负责人: DIANE M. ROBINS
• 金额: $ 29.59万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8067922
• 关键词: AR gene; Ablation; Affect; Age; Alleles; Androgen Receptor; Androgens; Bioinformatics; Biological; Biological Assay; Castration; Cell model; Cells; Combined Modality Therapy; Data; Development; Disease; Dissection; Engineering; Epithelial; Epithelium; Event; Future; Gene Expression; Gene Expression Regulation; Gene Targeting; Genetic; Genetic Polymorphism; Genetic Recombination; Genetic Variation; Genomics; Germ Lines; Glutamine; Growth; Health; Hormonal; Hormones; Human; In Vitro; Length; Lentivirus Vector; Ligands; Malignant Neoplasms; Malignant neoplasm of prostate; Maps; Methods; Modeling; Molecular; Molecular Chaperones; Molecular Genetics; Mus; Mutation; N-terminal; Nature; Normal Cell; Nuclear; Oncogenes; Oncogenic; Outcome; PTEN gene; Pathologic; Pathology; Pathway interactions; Patients; Pattern; Phenotype; Physiology; Pre-Clinical Model; Prostate; Prostate Cancer therapy; RNA; RNA Sequences; Receptor Gene; Regulation; Reproduction; Resistance; Resistance development; Role; Sampling; Selection for Treatments; Series; Signal Pathway; Signal Transduction; Somatic Mutation; Staging; Steroids; Testing; Time; Transcript; Transgenes; Transgenic Mice; Transgenic Model; Transgenic Organisms; Tumor Expansion; VCaP; Variant; Xenograft procedure; base; cancer cell; cancer initiation; cancer risk; cancer therapy; cell growth; cell type; chromatin immunoprecipitation; combat; genetic risk factor; high throughput screening; homologous recombination; human disease; in vivo; male; molecular marker; mouse model; mutant; neoplastic cell; new therapeutic target; next generation; novel; novel strategies; programs; promoter; public health relevance; receptor; receptor expression; receptor function; response; small hairpin RNA; tool; trait; tumor; tumor growth; tumor progression; tumor xenograft; tumorigenesis

DESCRIPTION (provided by applicant): The androgen receptor (AR) regulates male physiology and is central to the ontogeny and progression of prostate cancer. Understanding AR's role strongly impacts human health by identifying genetic risk factors and new therapeutic targets. In normal as well as pathologic cell growth, AR directs divergent actions, such as proliferation or differentiation, through gene targets that vary with promoter, cell type and stage. These context- dependent and opposing functions of AR are evident in prostate cancer where AR initially suppresses tumor development but ultimately promotes oncogenesis. AR's dual nature is highlighted by germ-line and somatic mutations of the AR gene. We hypothesize that mechanisms underlying differential gene regulation by variant ARs may reveal more effective ways to combat pathological actions of the wild type AR. In order to use genetic tools, we converted the mouse AR gene to the human sequence by homologous recombination. We first tested the effect of a polymorphic glutamine (Q) tract associated with male phenotypic variation. Q tract length in humanized AR mice impacts the androgen axis, and affects initiation, progression and therapy response of prostate cancer in a transgenic model. Somatic AR mutations in these tumors evince selection by treatment. Moreover, this was substantiated in a select set of human patients. Mutations chosen for further study affecting the N-terminal domain influence ligand-dependent and -independent activation. The mutant ARs use multiple mechanisms that ultimately alter target gene selectivity to promote disease. We will use a multipronged genetic and genomic analysis to define upstream and downstream mechanisms by which a subset of Q tract variants and mutant ARs ("the variants") confer differential gene regulation. Our aims will define variant AR activity in novel cell and mouse models and identify the downstream effectors and upstream activators of aberrant function by Next Generation deep sequencing. In Aim I, a small set of variant ARs will be stably expressed in normal and cancer cells to view their distinct biological programs in vitro and in xenograft tumors. In Aim II, pathologic activity of variant ARs will be tested in mice with prostate- targeted transgenes, in mice predisposed to cancer by deletion of one PTEN allele, and in mice with varying androgen axis strength. In Aim III, gene expression programs directed by the variant ARs will be defined by high-throughput sequencing of transcripts (RNA-tag-Seq) to identify direct and indirect AR targets. Chromatin immunoprecipitation and high-throughput sequencing (ChIP-Seq) will probe differential promoter recognition and highlight coregulators directing different biological outcomes. Differences shared amongst AR variants, in addition to unique actions, will highlight critical downstream targets and upstream regulators of wild type AR. Combined genetic and genomic study of these AR variants will reveal aspects of wild type AR function so that in future we may selectively block disease-promoting while retaining disease-inhibiting effects of AR in diverse contexts. These results will have immediate applications and broad significance for future studies. PUBLIC HEALTH RELEVANCE: As demonstrated by recent advances in prostate cancer therapy aimed at the steroid signaling pathway, the androgen receptor remains the high impact target. Our humanized mouse model provides a powerful means to compare the functions of a set of variant androgen receptors, using molecular, genetic and genomic approaches. This will identify common features that drive cell pathways towards proliferation versus differentiation and provide a comprehensive basis for targeting the functions of wild type androgen receptor, rather than the receptor itself, for more effective cancer treatments that are tailored to specific contexts.

描述（由申请人提供）：雄激素受体（AR）调节男性生理学，是前列腺癌个体发生和进展的核心。了解AR的作用通过识别遗传风险因素和新的治疗靶点对人类健康产生重大影响。在正常和病理性细胞生长中，AR通过随启动子、细胞类型和阶段而变化的基因靶点指导不同的作用，例如增殖或分化。AR的这些背景依赖性和相反的功能在前列腺癌中是明显的，其中AR最初抑制肿瘤发展，但最终促进肿瘤发生。AR基因的生殖系和体细胞突变突出了AR的双重性质。我们推测，变异AR的差异基因调控机制可能揭示了更有效的方法来对抗野生型AR的病理作用。 为了使用遗传工具，我们通过同源重组将小鼠AR基因转化为人类序列。我们首先测试了与男性表型变异相关的多态性谷氨酰胺（Q）道的影响。人源化AR小鼠的Q束长度影响雄激素轴，并影响转基因模型中前列腺癌的发生、进展和治疗反应。这些肿瘤中的体细胞AR突变表明通过治疗进行选择。此外，这在一组选定的人类患者中得到了证实。选择用于进一步研究的影响N-末端结构域的突变影响配体依赖性和非依赖性激活。突变AR使用多种机制，最终改变靶基因选择性以促进疾病。我们将使用多管齐下的遗传和基因组分析来定义上游和下游机制，通过该机制，Q束变体和突变AR（“变体”）的子集赋予差异基因调控。 我们的目标是在新的细胞和小鼠模型中定义变体AR活性，并通过下一代深度测序鉴定异常功能的下游效应子和上游激活子。在目标I中，一小组变体AR将在正常细胞和癌细胞中稳定表达，以观察它们在体外和异种移植肿瘤中的不同生物学程序。在目的II中，将在具有前列腺靶向转基因的小鼠中、在通过缺失一个PTEN等位基因而易患癌症的小鼠中以及在具有不同雄激素轴强度的小鼠中测试变体AR的病理活性。在目标III中，将通过转录物的高通量测序（RNA-tag-Seq）来定义由变体AR指导的基因表达程序，以鉴定直接和间接AR靶标。染色质免疫沉淀和高通量测序（ChIP-Seq）将探测差异启动子识别，并突出显示指导不同生物学结果的辅助调节因子。除了独特的作用外，AR变体之间共享的差异将突出野生型AR的关键下游靶标和上游调节因子。 这些AR变体的遗传和基因组研究将揭示野生型AR功能的各个方面，以便将来我们可以选择性地阻止疾病促进，同时保留AR在不同背景下的疾病抑制作用。这些结果将有直接的应用和广泛的意义，为今后的研究。 公共卫生关系：正如针对类固醇信号通路的前列腺癌治疗的最新进展所证明的那样，雄激素受体仍然是高影响力的目标。我们的人源化小鼠模型提供了一个强大的手段，比较一组变异雄激素受体的功能，使用分子，遗传和基因组的方法。这将确定驱动细胞增殖与分化途径的共同特征，并为靶向野生型雄激素受体而不是受体本身的功能提供全面的基础，以实现针对特定环境的更有效的癌症治疗。