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Principal Investigator: Michael-Christopher Keogh
Organization: EPICYPHER, INC.
Fiscal Year: 2024
Award: $1,032,776
Funding agency: National Human Genome Research Institute
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.
Terms: <Acceleration><Acetylation><Adoption><Antibodies><Antigenic Determinants><Assay><BS-seq><Bar Codes><Basal Transcription Factor><Basal transcription factor genes><Benchmarking><Best Practice Analysis><Binding><Binding Determinants><Bio-Informatics><Bioassay><Bioinformatics><Biologic Sciences><Biological Assay><Biological Markers><Biological Sciences><Bioscience><Bisulfite-based sequencing><Body Tissues><CUT&RUN><Cancers><Cell Body><Cell Communication and Signaling><Cell Line><Cell Signaling><CellLine><Cells><ChIP assay><Chemicals><Chromatin><Cleavage Targets and Release Using Nuclease><Cleavage Under Targets and Release Using Nuclease><Clinical><Complex><DNA><DNA Integration><DNA Methylation><DNA analysis><DNA methylation profiling><Data><Deoxyribonucleic Acid><Development><Diagnostic><Disease><Disorder><Dose><Drug Targeting><Drug Therapy><Epigenetic><Epigenetic Change><Epigenetic Mechanism><Epigenetic Process><Epitopes><Exhibits><Freezing><Gene Expression><General Transcription Factor Gene><General Transcription Factors><Genomics><High Throughput Assay><Histones><Intracellular Communication and Signaling><K-562><K562><K562 blasts><L-Lysine><Letters><Life Sciences><Lysine><MCF-7><MCF-7 Cell><MCF-7DR><MCF-7WT><MCF7><MCF7 cell><Malignant Neoplasms><Malignant Tumor><Manuals><Maps><Methods><Methyl-Seq><MethylSeq><Methylation><Methylation sequencing><Molecular><Molecular Interaction><Monitor><Nervous System Diseases><Nervous System Disorder><Neurologic Disorders><Neurological Disorders><New England><Noise><Northeastern United States><Nucleosomes><Pharmacotherapy><Phase><Post-Translational Modification Protein/Amino Acid Biochemistry><Post-Translational Modifications><Post-Translational Protein Modification><Post-Translational Protein Processing><Posttranslational Modifications><Posttranslational Protein Processing><Protein Modification><Proteins><Protocol><Protocols documentation><Regulation><Resolution><Sampling><Savings><Services><Signal Transduction><Signal Transduction Systems><Signaling><Strains Cell Lines><Technology><Time><Tissues><Titrations><Transcription Factor Proto-Oncogene><Transcription factor genes><Ubiquitilation><Ubiquitination><Ubiquitinoylation><analyze DNA><barcode><benchmark><bio-markers><biologic marker><biological signal transduction><biomarker><bisulfite><bisulfite sequencing><bisulfite-seq><chromatin immunoprecipitation><clinical applicability><clinical application><cultured cell line><data and analysis portal><developmental><drug treatment><epigenetically><experiment><experimental research><experimental study><experiments><genomic tools><hESC><high throughput screening><human ES cell><human ESC><human embryonic stem cell><hydrogen sulfite><hydrosulfite><inhibitor><innovate><innovation><innovative><lab development><laboratory development><malignancy><methylation pattern><multiomics><multiple omics><neoplasm/cancer><neurological disease><new approaches><next generation><novel><novel approaches><novel strategies><novel strategy><panomics><resolutions><response><targeted drug therapy><targeted drug treatments><targeted therapeutic><targeted therapeutic agents><targeted therapy><targeted treatment><tool><transcription factor><ubiquination><ubiquitin conjugation><user-friendly>