Development of Enhancer RNA-based Biomarkers in FFPE Tissue

 描述（由申请人提供）：增强子RNA代表了一种令人兴奋的新的转录物类别，其产生于基因组中的基因表达增强子。这些增强子衍生的RNA标记负责驱动基因表达程序的基因组的活性增强子和调控区。由于技术限制，增强子RNA尚未在临床环境中彻底探索。增强子RNA的精确定量可以通过提供整个基因组的基因调控事件的高分辨率图谱来改变癌症诊断。特别是，靶向表观遗传修饰酶的药物，在许多不同的癌症类型中表现出很大的希望，应该在全基因组范围内影响基因调控。增强子RNA的分析将提供增强子活性的直接读数，并询问表观遗传修饰药物的主要作用机制。为了探索增强子RNA定量的临床实用性，我们建议开发一种称为FFPEcap-seq的方法，该方法将能够在最常见的患者样本形式中准确地测量增强子RNA，福尔马林固定石蜡包埋（FFPE）。该项目概述了FFPEcap-seq的开发分为三个阶段：（1）在新鲜收集的RNA中进行优化，（2）在FFPE衍生的RNA中进行测试，以及（3）应用于HDAC抑制剂处理的细胞系和患者样本。在第1阶段，我们将使用定义的质量控制指标优化完全完整RNA的FFPEcap-seq。对于第2阶段，我们将从匹配的新鲜生长细胞和FFPE细胞中收集RNA。这种强大的资源将允许在来自相同样品的完全完整和FFPE降解的RNA中比较FFPEcap-seq和其他方法（最初不是为FFPE RNA设计的）。在第3阶段，FFPEcap-seq将应用于HDAC抑制剂治疗前后的细胞系和患者活检，HDAC抑制剂是一类令人兴奋的表观遗传修饰药物。该项目的总体目标是产生一种方法，该方法将允许分析FFPE患者样品中的增强子RNA，并在HDAC抑制剂处理的样品的试点研究中展示FFPEcap-seq。从长远来看，FFPEcap-seq可以作为一种用于患者选择/监测和发现基因组位点的技术得到广泛采用，这些基因组位点负责在许多癌症类型中观察到的异常基因调控。

 DESCRIPTION (provided by applicant): Enhancer RNAs represent an exciting new class of transcripts that arise from gene expression enhancers across the genome. These enhancer derived RNAs mark active enhancers and the regulatory regions of the genome that are responsible for driving gene expression programs. Due to technical limitations, enhancers RNAs have not been thoroughly explored in a clinical setting. Accurate quantification of enhancer RNAs could transform cancer diagnostics by providing a high-resolution map of gene regulation events across the genome. In particular, drugs that target epigenetic modifying enzymes, which have shown great promise across many different cancer types, should impact gene regulation on a genome-wide scale. Analysis of enhancer RNAs will provide a direct readout of enhancer activity and interrogate the main mechanism of action of epigenetic modifying drugs. To explore the clinical utility of enhancer RNA quantification we propose to develop an approach, called FFPEcap-seq, which will enable the accurate genome-wide measurement of enhancer RNAs in the most commonly available form of patient samples, formalin-fixed paraffin embedded (FFPE). This project outlines the development of FFPEcap-seq in three phases: (1) Optimization in freshly collected RNA, (2) Testing in FFPE derived RNA, and (3) Application to HDAC inhibitor treated cell lines and patient samples. In phase 1, we will optimize FFPEcap-seq on fully intact RNA using defined quality control metrics. For phase 2, we will collect RNA from matched freshly grown cells and FFPE cells. This powerful resource will allow for the comparison of FFPEcap-seq and other methods (not originally designed for FFPE RNA), in both fully intact and FFPE degraded RNA from the same sample. During phase 3, FFPEcap-seq will be applied to cell lines and patient biopsies pre- and post-treatment with HDAC inhibitors, an exciting class of epigenetic modifying drugs. The overall goal of this project is to produce a method that will permit the analysis of enhancer RNAs in FFPE patient samples as well as showcase FFPEcap-seq in pilot studies with HDAC inhibitor treated samples. Over the long term, FFPEcap-seq could find wide adoption as a technique for both patient selection/monitoring and discovering genomic loci that are responsible for aberrant gene regulation observed in many cancer types.