Novel genomic mechanism for ligand-dependent transcription by androgen receptor

雄激素受体配体依赖性转录的新基因组机制 项目概要/摘要 雄激素受体（Androgen receptor，AR）是核激素受体（nuclear hormone receptor，NR）超家族的成员， 激素应答元件（HRE）并调节内源性配体中靶基因表达 （激动剂）诱导的方式在不同的组织。由于AR在前列腺疾病的发生和发展中起着关键作用， 在癌症中，已经开发了许多合成的AR拮抗剂来抑制内源性AR的作用， 配体。一个突出的例子是恩杂鲁胺（Xtandi®），一种第二代AR拮抗剂，其显示出强的AR抑制作用。 前列腺癌的抗癌活性。然而，对enzalutamide的固有或获得性耐药，以及所有 可用的AR拮抗剂，导致治疗失败。因此，当前AR的治疗功效 需要改进拮抗剂。阐明拮抗剂配体AR的基因组机制 功能对于改善AR靶向治疗至关重要。在初步研究中，我们定义了 前列腺癌中第一个高分辨率（基序分辨率）激动剂和拮抗剂配体的AR顺反序列 细胞通过使用一种新的染色质免疫沉淀-核酸外切酶（ChIP-exo）的方法。出乎意料的是，我们 发现AR与天然激动剂（双氢睾酮，DHT）和拮抗剂（恩杂鲁胺）结合， 染色质上明显不同的DNA基序（称为“DNA基序切换”）。令人惊讶的是，集成ChIP-exo 和RNA-seq分析发现，与新的AR结合基序结合的Enzalutamide配体AR，显著地 影响整体癌症相关转录。通过将我们新的ChIP-exo基因组方法与其他方法相结合， 通过表观基因组学、蛋白质组学和生物化学方法，我们进一步发现Enzalutamide配体AR与 与特异性协作转录因子（如FoxA 1）和非DNA结合辅助调节因子（如Hsp 90）共同作用， 特异性活性顺式调节区。重要的是，药理学Hsp 90抑制显著降低了 Enzalutamide配体AR靶基因（例如癌症促进基因GR和CD 55）的表达，以及 增强恩杂鲁胺的细胞生长抑制作用。基于这些令人信服的数据，我们假设， DNA基序转换是一种新的基因组机制，它依赖于拮抗剂，与癌症相关， 通过拮抗剂配体的AR转录复合物进行转录。我们的具体目标是：1）确定是否 特异性转录因子和表观遗传特征在全局上促进AR DNA基序转换;和2） 研究拮抗剂配体AR结合如何调节癌症相关基因的表达。通过显著 增强了我们对拮抗剂如何调控AR转录的理解， 本研究将为进一步开发AR靶向治疗奠定基础。

Novel genomic mechanism for ligand-dependent transcription by androgen receptor Project Summary/Abstract Androgen receptor (AR) is a member of nuclear hormone receptor (NR) superfamily that binds to cognate hormone responsive elements (HREs) and regulates target gene expression in an endogenous ligand (agonist)-inducible manner in diverse tissues. As AR plays a key role in the onset and progression of prostate cancer, numerous synthetic AR antagonists have been developed to inhibit the action of endogenous AR ligands. A prominent example is enzalutamide (Xtandi®), a second-generation AR antagonist showing strong anti-cancer activity for prostate cancer. However, intrinsic or acquired resistance to enzalutamide, and all available AR antagonists, occurs leading to treatment failure. Thus, therapeutic efficacy of current AR antagonists needs to be improved. Elucidation of genomic mechanisms underlying antagonist-liganded AR function is critically important in order to improve AR-targeted therapy. In preliminary studies, we have defined the first high-resolution (motif-resolution) agonist- and antagonist-liganded AR cistromes in prostate cancer cells by using a novel chromatin immunoprecipitation-exonuclease (ChIP-exo) approach. Unexpectedly, we found that AR bound to natural agonist (dihydrotestosterone, DHT) and antagonist (enzalutamide) recognizes distinctly different DNA motifs on chromatin (termed “DNA motif switching”). Surprisingly, integrated ChIP-exo and RNA-seq analysis found that enzalutamide-liganded AR, bound to a novel AR binding motif, significantly affects global, cancer-relevant transcription. By combining our novel ChIP-exo genomic approach with other epigenomic, proteomic and biochemical approaches, we further found that enzalutamide-liganded AR interacts with specific collaborating transcription factors (e.g. FoxA1) and non-DNA binding coregulators (e.g. Hsp90) on specific active cis-regulatory regions. Importantly, pharmacological Hsp90 inhibition significantly decreases expression of enzalutamide-liganded AR target genes (e.g. cancer promoting genes GR and CD55) and enhances cell growth inhibitory effect of enzalutamide. Based on these compelling data, we hypothesize that DNA motif switching is a novel genomic mechanism underlying antagonist-dependent, cancer-relevant transcription by antagonist-liganded AR transcription complex. Our specific aims are to: 1) determine whether specific transcription factors and epigenetic features globally facilitate AR DNA motif switching; and 2) investigate how antagonist-liganded AR binding regulates expression of cancer-relevant genes. By significantly enhancing our understanding of how antagonist-regulated transcription by AR is controlled at the genomic level, this study will lay the foundation for future development of improved AR-targeted therapy.