Elucidating the role of the noncoding genome in neuroblastoma

阐明非编码基因组在神经母细胞瘤中的作用

基本信息

批准号：
10596528
负责人：
Sharon Diskin
金额：
$ 32.78万
依托单位：
CHILDREN'S HOSP OF PHILADELPHIA
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-03-12 至 2026-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10596528
关键词：
3-Dimensional ATAC-seq Affect Architecture Automobile Driving Binding Binding Sites Biological Biological Assay Cell Line Cell model Cells ChIP-seq Child Chromatin Chromatin Loop Chromosomal translocation Chromosome Mapping Clinical Clustered Regularly Interspaced Short Palindromic Repeats Code Correlative Study Coupled DNA DNA Methylation Data Diagnostic Disease Progression Distal Enhancers Epigenetic Process Etiology Event Evolution Exhibits Gene Expression Regulation Genes Genetic Transcription Genome Genomics Germ-Line Mutation Histones Human Invaded Knowledge MYC gene MYCN gene Malignant Childhood Neoplasm Malignant Neoplasms Maps Mediating Mission Mutagenesis Mutation Nature Neural Crest Cell Neuroblastoma Nucleic Acid Regulatory Sequences Oncogenic Outcome Proteins Public Health RNA-Directed DNA Polymerase Recurrence Recurrent disease Relapse Reporter Reporting Research Research Project Grants Resolution Resources Role Shapes Somatic Mutation Structural Protein Susceptibility Gene Sympathetic Nervous System Telomerase Testing Transcriptional Activation United States National Institutes of Health Untranslated RNA Variant Work cancer predisposition causal variant cell type clinical biomarkers clinical development epigenetic regulation epigenome epigenomics evidence base experimental study genetic manipulation genome sequencing genome wide association study genome-wide analysis high risk improved outcome in silico insight multiple omics neuroblastoma cell precursor cell programs promoter transcription factor transcriptome sequencing tumor tumor initiation tumorigenesis ultra high resolution whole genome

项目摘要

Project Summary Neuroblastoma (NB) remains one of the deadliest childhood cancers. NB exhibits a paucity of recurrent protein coding mutations and few targetable mutations (2-5), providing the rationale for this proposal. Noncoding variants can disrupt regulatory and/or structural DNA leading to dysregulated transcriptional programs that promote tumorigenesis. Our objective here is to identify noncoding variants and mechanisms that drive NB. Our central hypothesis is that germline variants and somatic mutations within noncoding regulatory regions of DNA potently influence NB initiation, progression and/or disease relapse. We will test our hypothesis in three specific aims: 1) Define and evaluate differences in the epigenomic landscape of NB and NB precursor cells. First, a panel of genetically diverse human-derived NB cell lines and neural crest cells (NCC; NB precursor cells) will be characterized. 3D chromatin architecture at all promoters will be ascertained using an ultra-high-resolution promoter-focused Capture C approach. Cells will be further profiled by whole genome sequencing (WGS), RNA- seq, ATAC-Seq, and ChIP-seq for histone marks and key structural proteins. Evolution of the epigenomic landscape from NB precursor to NB cells will be assessed. Data will be integrated with transcription factor binding site functional predictions to provide a comprehensive resource for the interpretation of noncoding variants. 2) Identify germline noncoding variants influencing NB tumorigenesis. We will perform variant-to-gene mapping at NB susceptibility loci identified by GWAS and identify putative causal variants mapping to open chromatin and involved in chromatin interactions in NB precursor or NB cells. Next, rare germline noncoding mutations from WGS will be assessed in a similar manner to identify variants interacting with known cancer predisposition genes. Further in silico prioritization will be accomplished via clinical correlative and integrative host-tumor analyses. The mechanism by which top prioritized noncoding variants promote NB will be determined using genetic manipulation in cell models in combination with Capture C, ChIP-seq, RNA-seq and/or functional studies. 3) Discover and assess biological relevance of somatic noncoding drivers of NB. We will integrate WGS of diagnostic and relapsed NB tumors to identify noncoding mutations affecting regulatory DNA and chromatin interactions in NB genomes. Recurrent variants will be further characterized through integration with matched RNA-seq (n=443), DNA methylation (n=223) and clinical correlative studies. We will elucidate biological relevance of prioritized mutations via massively parallel reporter assay (MPRA) coupled with CRISPR-based genetic manipulation in cell models in combination with Capture C, ChIP-seq, RNA-seq and/or functional assays. This work will have a sustained and positive impact on the field by providing substantial insights into the role of the noncoding genome in this important childhood cancer, and has the potential to inform development of clinical biomarkers and/or evidence-based therapies to improve outcomes of children with NB.

项目概要神经母细胞瘤（NB）仍然是最致命的儿童癌症之一。 NB 表现出缺乏循环蛋白编码突变和少数可靶向突变 (2-5)，为该提案提供了基本原理。非编码变体可以破坏调控和/或结构 DNA，导致转录程序失调，从而促进肿瘤发生。我们的目标是识别驱动 NB 的非编码变体和机制。我们的中央假设是DNA非编码调控区域内的种系变异和体细胞突变有效地影响 NB 的发生、进展和/或疾病复发。我们将在三个具体目标上检验我们的假设：1）定义并评估 NB 和 NB 前体细胞表观基因组景观的差异。首先是一个面板遗传多样性的人源 NB 细胞系和神经嵴细胞（NCC；NB 前体细胞）将被特点。所有启动子的 3D 染色质结构将使用超高分辨率确定以启动子为中心的 Capture C 方法。细胞将通过全基因组测序（WGS）、RNA- 用于组蛋白标记和关键结构蛋白的 seq、ATAC-Seq 和 ChIP-seq。表观基因组的进化将评估从 NB 前体到 NB 细胞的景观。数据将与转录因子结合整合位点功能预测为非编码变体的解释提供全面的资源。 2）鉴定影响 NB 肿瘤发生的种系非编码变异。我们将进行变体到基因映射由 GWAS 识别的 NB 易感位点，并识别推定的因果变异映射至开放染色质并参与 NB 前体或 NB 细胞中的染色质相互作用。接下来，罕见种系非编码将以类似的方式评估全基因组测序的突变，以识别与已知癌症相互作用的变异体易感基因。进一步的计算机优先级将通过临床相关性和综合性来完成宿主-肿瘤分析。将确定最高优先级非编码变体促进 NB 的机制在细胞模型中结合使用 Capture C、ChIP-seq、RNA-seq 和/或功能性基因操作研究。 3) 发现并评估 NB 体细胞非编码驱动因素的生物学相关性。我们将整合对诊断性和复发性 NB 肿瘤进行全基因组测序，以识别影响调节性 DNA 和 NB 基因组中的染色质相互作用。反复出现的变异将通过整合进一步表征匹配的 RNA-seq (n=443)、DNA 甲基化 (n=223) 和临床相关研究。我们将阐明生物学通过大规模并行报告分析 (MPRA) 结合基于 CRISPR 的优先突变的相关性结合 Capture C、ChIP-seq、RNA-seq 和/或功能测定在细胞模型中进行遗传操作。这项工作将通过提供对以下角色的深入见解，对该领域产生持续和积极的影响：这种重要的儿童癌症中的非编码基因组，并有可能为临床的发展提供信息生物标志物和/或循证疗法可改善 NB 儿童的预后。