Ultrasensitive multiomic platform using epitope-targeted DNA methylation mapping

使用表位靶向 DNA 甲基化作图的超灵敏多组学平台

• 批准号: 10758061
• 负责人: Michael-Christopher Keogh
• 金额: $ 103.57万
• 依托单位: EPICYPHER, INC.
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10758061
• 关键词: Ultrasensitive; multiomic; platform; using; epitope

PROJECT SUMMARY Gene expression is regulated by the complex molecular cross-talk between DNA methylation (DNAme) and other chromatin features: e.g. histone post-translational modifications (PTMs) and chromatin associated proteins (ChAPs; transcription factors and chromatin remodelers). Significantly, changes in the chromatin landscape can have a profound impact on DNAme patterning (and vice versa), and these changes are connected to development as well as a broad range of diseases (from cancer to neurological disorders). However, our understanding of how DNAme co-occurs / coordinates with additional chromatin features to control gene expression is limited by a lack of reliable genomic tools. Here, EpiCypher is partnering with New England Biolabs (NEB) to develop Targeted Enzymatic Methylation-sequencing (TEM-seqTM), an ultra-sensitive multiomic mapping technology that delivers high resolution DNAme profiles (5mC/5hmC) at epitope-defined chromatin features. EpiCypher is leading the development of ultra-sensitive genomic mapping assays that use CUT&RUN / CUT&Tag methods (under the CUTANA® platform) to generate truly quantitative data using dramatically reduced cell input and sequencing depth (>10-fold savings on each parameter vs. ChIP-seq). CUTANA assays are supported by EpiCypher’s proprietary spike-in designer nucleosome (dNuc) technology to enable technical monitoring and quantitative normalization. The key innovation of the TEM-seq project is the development of a novel multiomic workflow that marries EpiCypher’s quantitative CUTANA CUT&RUN technology with unbiased DNAme analysis using NEB’s enzymatic methyl-seq (EM-seq) approach. EM-seq utilizes the enzymatic conversion of DNAme (5mC / 5hmC) and provides a much-needed alternative to bisulfite sequencing (BS; a chemical treatment that degrades DNA and has systemic sequence biases) to generate high resolution, unbiased DNAme profiles with ~10-fold less sample input (vs. BS). In Phase I Aim 1, we will rigorously validate our TEM-seq workflow in three cell lines, benchmark results against standard CUT&RUN and EM-seq assays, and further develop EpiCypher’s spike-in controls for compatibility with TEM-seq. We will advance to Phase II when we demonstrate that TEM-seq generates highly reliable DNAme maps associated with histone PTMs and ChAPs using <50k cells and <10M reads. In Phase II Aim 2, we will expand development of spike-in control panels and develop robust protocols for a wide panel of chromatin features (using validated antibodies) and sample processing methods (fresh, frozen, and fixed), including drug treatment time-course experiments to enable clinical applications. In Phase II Aim 3, we will develop / validate a TEM-seq beta kit, and also create a data analysis portal and automated assays to accelerate commercial adoption and enable a high-throughput service offering. TEM-seq will provide a powerful new tool to expand our understanding of complex chromatin signaling, further unlocking the potential of epigenetics-targeted drugs and diagnostics.

项目总结