Defining the epigenetic landscape in human prostate cancer

 描述（由申请人提供）：雄激素受体（AR）是前列腺癌发生、进展和耐药性的核心。AR是一种核转录因子（TF），与DNA结合并调节基因活性。基因组范围的AR-DNA结合位点的集合被称为AR顺反子组。AR及其共调节因子之间复杂的相互作用决定了转录调节的靶基因。使用新开发的AR染色质免疫沉淀技术，然后在人前列腺标本中进行高通量测序（ChIP-seq），我们明确表明AR程序是高度动态的，至少部分取决于可用的辅助调节剂的存在。特别是 先驱TFFOXA 1和前列腺谱系特异性TFHOXB 13似乎共定位于肿瘤组织特异性AR位点。本提案的总体目标是表征肿瘤发生期间AR重编程的机制，并首次表征从局限性前列腺癌进展为转移性耐药疾病期间人体组织中的AR程序。在第一个目标中，将在AR表达正常前列腺上皮（LHSAR）和前列腺癌（LNCaP和VCaP）的细胞系模型中进行ChIP-seq。将测量AR、FOXA 1和HOXB 13的顺式组的变化，因为每个TF通过shRNA和基因组编辑敲低或通过慢病毒转导过表达。还将对每种细胞系条件进行RNA-seq，以确定受表观遗传重编程影响的基因。在第二个和第三个目标中，将通过ChIP-seq在人类标本中系统地绘制AR的表观遗传景观：局部组织，未治疗的转移组织，去势抵抗性转移和恩杂鲁胺耐药疾病。完成本提案中概述的新实验将为主转录因子如何驱动前列腺癌进展提供无与伦比的视角。具体而言，我们将发现：（i）在前列腺癌发生和进展期间AR如何重编程，以及哪些共调节因子促进了这一过程，（ii）在获得Enzalutamide耐药性期间AR如何重编程，如果是，哪些共调节因子促进了这一过程，以及（iii）其他驱动前列腺癌进展的非AR主调节因子。拟议的实验还将使我们能够识别受AR重编程影响的靶基因。该项目将导致一个地图集的表观遗传景观的疾病进展到去势抵抗转移状态，这是一致致命的。遗传基因座 并且包含该数据集的靶基因将刺激用于治疗干预的新靶点。

 DESCRIPTION (provided by applicant): The androgen receptor (AR) is central to prostate cancer development, progression, and drug resistance. The AR is a nuclear transcription factor (TF) that binds to DNA and regulates gene activity. The set of genome-wide AR-DNA binding sites is termed the AR cistrome. A complex interplay between AR and its co-regulators determines the genes targeted for transcriptional regulation. Using newly developed techniques for AR chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) in human prostate specimens, we unambiguously show that the AR program is highly dynamic and is determined, at least in part, by the presence of available co-regulators. In particular, the pioneer TF FOXA1 and the prostate lineage- specific TF HOXB13 appear to co-localize at tumor tissue-specific AR sites. The overall objectives of the present proposal are to characterize the mechanisms underlying AR reprogramming during tumorigenesis and - for the first time - to characterize the AR program in human tissue during progression from localized prostate cancer to metastatic, drug-resistant disease. In the first aim, ChIP-seq will be performed in cell line models for AR-expressing normal prostate epithelium (LHSAR) and prostate cancer (LNCaP and VCaP). Changes in the cistromes of AR, FOXA1 and HOXB13 will be measured as each TF is knocked down via shRNA and genome editing or overexpressed via lentiviral transduction. RNA-seq will also be performed for each cell line condition to determine the genes affected by epigenetic reprogramming. In the second and third aims, the epigenetic landscape of the AR will be systematically charted via ChIP-seq in human specimens: localized tissue, untreated metastatic tissue, castration-resistant metastases and enzalutamide-resistant disease. Completion of the novel experiments outlined in this proposal will provide an unparalleled look at how master transcription factors drive prostate cancer progression. Specifically, we will discover: (i) how the AR is reprogrammed during prostate cancer development and progression, and which co-regulators are facilitating this process, (ii) how the AR is reprogrammed during the acquisition of resistance to enzalutamide and, if so, which co-regulators are facilitating this process, and (iii) other non-AR master regulators that are driving prostate cancer progression. The proposed experiments will also enable us to identify the target genes that are affected by AR reprogramming. The project will result in an atlas of the epigenetic landscape as the disease progresses to the castration-resistant metastatic state, which is uniformly fatal. The genetic loci and target genes comprising this dataset will stimulate new targets for therapeutic intervention.