Scalable and quantitative chromatin profiling from formalin-fixed paraffin-embedded samples

项目摘要 组蛋白翻译后修饰（PTM）基因组定位工具的开发- wide开创了表观遗传学研究的新时代，将组蛋白PTM和相关机制置于 作为新的生物标志物和治疗靶点而备受关注。尽管取得了这一进展，但利用表观基因组学进行临床 研究一直受到缺乏高性能基因组作图测定的阻碍， 福尔马林固定石蜡包埋（FFPE）组织，由于在FFPE过程中发生的DNA损伤。 事实上，现有的FFPE样品相容的基因组作图测定是低通量和不敏感的 （需要大的样本输入），或者不能分辨特定的染色质特征（例如，启动子、增强子等）。 因此，下一代染色质谱分析测定能够使用 库存FFPE样本是解锁大量库存临床样本的关键， 疾病相关组蛋白PTM靶点的新生物标志物发现/验证。为了满足这一需求，EpiCypher 正在开发CUTANA-FFPETM，这是一种一流的低输入基因组作图解决方案，用于FFPE临床研究 样品该技术的灵感来自EpiCypher的商用CUTANA® CUT&Tag（目标下切割 和标签化）分析，并利用创新的体外转录为基础的工作流程，以减轻 FFPE样本处理引起的DNA损伤所带来的限制。本I期研究的目标是 证明了CUTANA-FFPE检测试剂盒能够可靠地定位FFPE组织样本中的组蛋白PTM。在Aim中 1，我们将开发CUTANA-FFPE工作流程，以定位固定培养细胞中的组蛋白PTM。在目标2中，我们 将在临床FFPE组织样本中使用CUTANA-FFPE检测，并将我们的结果与基因组 图来自供体匹配的新鲜冷冻组织。本研究将证明CUTANA-FFPE的可行性 成为一个强大的新工具，在以前无法进入的银行进行高价值的表观基因组研究 临床样本。这项技术将引起基础研究人员和药物开发人员的广泛兴趣， 表观基因组学在临床应用中的全部潜力。在第二阶段，我们将开发耐用的CUTANA-FFPE套件 以及使用超低样品输入针对FFPE组织优化的自动化测定方案， CUTANA-FFPE商业化的关键，使FFPE临床样本的表观基因组研究成为可能。

PROJECT SUMMARY The development of tools to map the location of histone post-translational modifications (PTMs) genome- wide has ushered in a new era of epigenetic research, placing histone PTMs and associated mechanisms in the spotlight as novel biomarkers and therapeutic targets. Despite this progress, leveraging epigenomics for clinical research has been hindered by a lack of high-performance genomic mapping assays that are compatible with formalin-fixed paraffin-embedded (FFPE) tissue, due to DNA damage that is incurred during the FFPE process. Indeed, existing FFPE sample-compatible genomic mapping assays are either low-throughput and insensitive (requiring large sample inputs), or do not resolve specific chromatin features (e.g., promoters, enhancers, etc.). Therefore, next-generation chromatin profiling assays that are able to resolve specific chromatin features using banked FFPE samples are key to unlock access to the massive collection of banked clinical samples and enable novel biomarker discovery / validation for disease-relevant histone PTM targets. To meet this need, EpiCypher is developing CUTANA-FFPETM, a first-in-class low input genomic mapping solution for the study of FFPE clinical samples. This technology is inspired by EpiCypher’s commercial CUTANA® CUT&Tag (Cleavage Under Targets & Tagmentation) assays, and leverages an innovative in vitro transcription-based workflow to mitigate the limitations imposed by FFPE sample processing-induced DNA damage. The goal of this Phase I study is to demonstrate the ability of CUTANA-FFPE assays to reliably map histone PTMs in FFPE tissue samples. In Aim 1, we will develop the CUTANA-FFPE workflow to map histone PTMs in fixed cultured cells. Then, in Aim 2, we will employ CUTANA-FFPE assays in clinical FFPE tissue samples and benchmark our results with genomic maps derived from donor-matched fresh frozen tissue. This study will demonstrate feasibility for CUTANA-FFPE to become a powerful new tool to perform high-value epigenomic research in previously inaccessible banked clinical samples. This technology will be of broad interest to basic researchers and drug developers, unlocking the full potential of epigenomics for clinical applications. In Phase II, we will develop robust CUTANA-FFPE kits and automated assay protocols that are optimized for FFPE tissue using ultra-low sample inputs, which will be key for commercializing CUTANA-FFPE to enable epigenomic studies of FFPE clinical samples.