Epi-seq: Multiplexed ChIP-seq for personalized medicine and drug discovery

Epi-seq：用于个性化医疗和药物发现的多重 ChIP-seq

• 批准号: 10021703
• 负责人: Mary Anne Jelinek
• 金额: $ 64.77万
• 依托单位: ACTIVE MOTIF, INC.
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-24 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10021703
• 关键词: Adipose tissue; Adoption; Affect; Algorithms; Antibodies; Autoimmune Process; Automation; Bar Codes; Benchmarking; Bioinformatics; Biological Assay; Biopsy; Cell Line; Cells; Characteristics; Childhood; Chromatin; Clinical; Complex; Contract Services; Coupled; Cultured Cells; DNA Polymerase II; DNA Sequence; DNA mapping; DNA sequencing; DNA-Binding Proteins; Data; Data Set; Dependence; Development; Diagnostic; Disease; Drug Screening; Epigenetic Process; Evaluation; Feedback; Freezing; Frequencies; Genetic Transcription; Genome; Genomics; Genotype; Hand; Heart; HepG2; Heritability; Histones; Human; Human Pathology; Individual; Inflammatory; K-562; Kidney; Laboratories; Libraries; Liquid substance; Malignant Neoplasms; Maps; Measures; Mediating; Methods; Mind; Morphologic artifacts; Mus; Mutation; Neurologic; Oligonucleotides; Pathology; Phase; Phenotype; Play; Preparation; Procedures; Prosencephalon; Proteins; Protocols documentation; RNA; Reagent; Recombinants; Recommendation; Resolution; Role; Sample Size; Sampling; Site; Small Business Innovation Research Grant; Sonication; System; Systems Integration; Technology; Teleconferences; Testing; Therapeutic; Time; Tissue Sample; Tissues; Transposase; Work; chromatin immunoprecipitation; commercialization; disorder subtype; drug discovery; epigenome; epigenomics; genetic profiling; genetic regulatory protein; genome sequencing; genome-wide; histone modification; human tissue; innovation; instrument; instrumentation; neuro-oncology; next generation; next generation sequencing; novel; novel diagnostics; novel therapeutics; personalized medicine; precision drugs; product development; prognostic; success; whole genome

SUMMARY Dysregulation of epigenetic mechanisms is known to play important roles in the development and progression several disease states including cancer, autoimmune, neurological and inflammatory disorders. In cancer alone, high frequency mutations in sixty epigenetic regulatory proteins have been identified. Like genome sequencing, epigenetic information can be elucidated across the entire genome and profiling can be used to find epigenetic abnormalities that cause or correlate with disease. Computational integration of genome-wide genetic and epigenetic profiles (epi-genotyping) has enabled identification of novel disease subgroups, with pediatric neuro-oncology being a notable example. Developing a systems solution that facilitates integration of epigenetic data sets is key to enabling personalized medicine solutions for pathologies with epigenetic dependencies. Chromatin immunoprecipitation (ChIP) is the primary method for genome-wide mapping of DNA associated proteins. In contrast to the tremendous advances in DNA sequencing technologies, which are approaching the $1,000 genome, ChIP has only had incremental improvements in work flow, reduction of background and sample size requirements. This SBIR Fast Track proposal intends to develop Epi-Seq, a high throughput multiplexing ChIP-Seq platform to map functional alterations in the epigenome on a scale and time frame suitable for clinical and drug screening applications. With automation compatibility in mind, Epi-Seq combines sample preparation and solubilization of fixed chromatin using PIXUL, a 96-well plate sonicator capable of shearing 96 chromatin samples in less than 30 minutes, with Transposase- Assisted Multiplex ChIP-Seq (TAM-ChIP) to functionally annotate the entire genome in a high throughput format. Multiplexing is achieved with a panel of recombinant AbFlex® ChIP-qualified antibodies targeting various hallmark features of the epigenetic landscape, each of which has been conjugated to a transposase- compatible barcoded oligonucleotide via site-directed conjugation. Phase I feasibility will use PIXUL from human and mouse tissues in Aim1 and compare TAM-ChIP enabled AbFlex® antibodies for key DNA binding proteins and histone PTMs in traditional and TAM-ChIP-seq methods. Phase II product development efforts will optimize TAM-ChIP target multiplexing in Aim 3 and Aim 4 efforts will adapt current workflows for automation compatibility such that a hands-free “sample-to-sequence” workflow is achieved. Phase II efforts will conclude with alpha-testing by independent laboratories whose feedback will be integrated into subsequent commercialization efforts by Active Motif. Epi-Seq is an enabling technology with potential accelerate development of epigenetic diagnostics, prognostic and therapeutics for diverse human pathologies and will be offered both as a contract service and as a product, sold independently or bundled with the PIXUL instrument.

总结 已知表观遗传机制的失调在肿瘤的发生和发展中起重要作用 几种疾病状态，包括癌症、自身免疫性疾病、神经系统疾病和炎性疾病。 仅在癌症中，已经确定了60种表观遗传调节蛋白的高频突变。像 通过基因组测序，可以在整个基因组中阐明表观遗传信息，并且可以 用于发现导致疾病或与疾病相关的表观遗传异常。计算积分 全基因组遗传和表观遗传谱（表观基因分型）已经能够识别新的疾病 亚组，儿科神经肿瘤学是一个值得注意的例子。开发系统解决方案， 促进表观遗传数据集的整合是实现病理学个性化医学解决方案的关键 表观遗传依赖性染色质免疫沉淀（ChIP）是全基因组检测的主要方法。 DNA相关蛋白质图谱。与DNA测序的巨大进步相比， 技术，这是接近1,000美元的基因组，ChIP只有渐进的改进， 工作流程，减少背景和样本量的要求。SBIR快速通道提案旨在 开发Epi-Seq，这是一种高通量多路复用ChIP-Seq平台，用于映射 表观基因组的规模和时间框架适合于临床和药物筛选应用。与自动化 考虑到兼容性，Epi-Seq结合了样品制备和使用PIXUL溶解固定的染色质， 96孔板超声波仪，能够在30分钟内剪切96个染色质样品，使用转座酶- 辅助多重ChIP-Seq（TAM-ChIP）以高通量功能性注释整个基因组 格式.使用一组重组AbFlex® ChIP合格抗体实现多重化， 表观遗传景观的各种标志性特征，其中每一个都与转座酶结合， 通过位点定向缀合将相容的条形码化寡核苷酸结合。第一阶段可行性研究将使用PIXUL， 并比较TAM-ChIP使能AbFlex®抗体的关键DNA结合 蛋白质和组蛋白PTM在传统和TAM-ChIP-seq方法。第二阶段产品开发工作将 在目标3和目标4中优化TAM-ChIP目标多路复用的努力将适应当前的自动化工作流程 因此，该系统具有良好的兼容性，从而实现了无需手动的“样品到序列”工作流程。第二阶段工作将结束 独立实验室的阿尔法测试，其反馈将被整合到后续的 积极的商业化努力。Epi-Seq是一种具有潜在加速 发展表观遗传学诊断，预后和治疗各种人类病理学，并将 作为合同服务和产品提供，独立销售或与PIXUL仪器捆绑销售。