Extrachromosomal DNA as a Targetable Mechanism in Glioblastoma

染色体外 DNA 作为胶质母细胞瘤的靶向机制

• 批准号: 10533330
• 负责人: Roel GW Verhaak
• 金额: $ 21.06万
• 依托单位: JACKSON LABORATORY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10533330
• 关键词: Acceleration; Affect; Amplifiers; Animal Model; Behavior; Binding; Biological Models; Biology; CRISPR/Cas technology; Cancer Cell Growth; Cell model; Cells; Characteristics; Chromatin; Chromatin Interaction Analysis by Paired-End Tag Sequencing; Chromosomal Duplication; Clustered Regularly Interspaced Short Palindromic Repeats; Cytopathology; Cytoplasm; DNA; DNA Sequence Alteration; DNA amplification; DNA biosynthesis; Data; Data Set; Deoxyribonucleotides; Development; Double Minutes; Elements; Enhancers; Enzymes; Esophageal carcinoma; Evolution; Frequencies; Gene Amplification; Genetic Transcription; Genome; Genomics; Glioblastoma; Goals; Growth; Heterogeneity; In Vitro; Knowledge; Label; Laboratories; Location; Maintenance; Malignant Neoplasms; Mediating; Methods; Methotrexate; Mitosis; Modeling; Molecular; Monitor; Neurosphere; Oncogene Activation; Oncogenes; Oncogenic; Outcome; Patients; Proteins; RNA Polymerase II; RRM1 gene; RRM2 gene; Regulation; Reporting; Research; Resistance; Ribonucleotide Reductase; Ribonucleotide Reductase Subunit; Role; S phase; Solid; Structure; System; Technology; Testing; Therapeutic; Time; Visualization; Visualization software; Work; autosome; cancer cell; cancer therapy; cancer type; cell behavior; cell killing; dosage; endonuclease; extrachromosomal DNA; gemcitabine; genetic element; genomic locus; hydroxyurea; in vivo Model; innovation; insight; interest; micronucleus; neoplastic cell; novel; patient derived xenograft model; pharmacologic; real-time images; sarcoma; therapeutic target; tool; tripolyphosphate; tumor; tumor DNA; tumor growth; tumor heterogeneity; tumorigenesis; virtual

PROJECT SUMMARY/ABSTRACT The goal of this project is to determine the influence of extrachromosomal DNA (ecDNA) on tumor evolution and resistance and to validate a potential therapeutic strategy based on ecDNA inhibition. The existence of ecDNA was first recognized through pioneering cytopathology studies by Arthur Spriggs over five decades ago. We know that ecDNAs are largely a phenomenon of tumor cells, can be detected in nearly half of all solid cancers, and frequently express oncogenes, making them vehicles for amplification of oncogene expression and consequent tumorigenesis. Our studies in glioblastoma extend this knowledge, showing that ecDNAs are highly pervasive and significantly increase intra-tumoral genomic heterogeneity. Despite this knowledge, research into the mechanisms of ecDNA-driven tumorigenesis is largely non-existent. To advance ecDNA research to the next level of understanding, we must now define its origins, replication mechanisms, and strategies to target it for treatment. At this time, however, there are significant barriers to such studies, such as the absence of profiling technologies and molecular tools to accurately detect and characterize these oncogenic elements. The work proposed here will remove these barriers to progress. We hypothesize that ecDNAs drive tumorigenesis beyond their role as oncogene activators, and that mechanisms of ecDNA formation and replication could serve as potential therapeutic targets. To test our hypothesis, we will develop methods to visualize and trace ecDNA in live cells and use these tools to define the mechanisms of ecDNA localization and transfer within a tumor. We will functionally evaluate whether ecDNA mediated cis-regulation of non-ecDNA gene transcription can be modulated to change tumor cell behavior. Finally, we will explore whether pharmacological depletion of ecDNA affects tumor growth and maintenance in cellular and animal model systems. Our Specific Aims are to: 1) Develop CRISPR-based tracing tools for real-time imaging of ecDNA dynamics; 2) Discovery and functional interrogation of ecDNA mediated oncogene activation; and 3) Target ecDNA through inhibition of ribonucleotide reductase (RNR), the enzyme responsible for generating the deoxyribonucleotide triphosphates that are the building blocks of autosomal and extrachromosomal DNA. Impact: This project is based on the conceptual innovation that ecDNAs are cancer-specific and cancer- enabling genomic alterations. Despite the recognition of ecDNA decades ago, the function of these structures and how to effectively target their genesis, is virtually uncharted territory. In addition to developing new, broadly applicable, tools for ecDNA studies, the results of this work will provide novel insights into the characteristics, behavior and replicative mechanism of ecDNA, and will assess whether ecDNA is an unexploited cancer cell vulnerability that can be therapeutically exploited for patient benefit.

项目概要/摘要 该项目的目标是确定染色体外 DNA (ecDNA) 对肿瘤进化的影响 和耐药性，并验证基于 ecDNA 抑制的潜在治疗策略。的存在 ecDNA 最初是由 Arthur Spriggs 在 5 年来通过开创性的细胞病理学研究得到认可的 前。我们知道ecDNA很大程度上是肿瘤细胞的一种现象，可以在近一半的固体中检测到。 癌症，并经常表达癌基因，使其成为癌基因表达放大的载体 以及随后的肿瘤发生。我们对胶质母细胞瘤的研究扩展了这一知识，表明 ecDNA 高度普遍并显着增加肿瘤内基因组异质性。尽管有这些知识， 对 ecDNA 驱动的肿瘤发生机制的研究基本上不存在。推进 ecDNA 研究到一个新的理解水平，我们现在必须定义它的起源、复制机制和 靶向治疗的策略。然而，目前此类研究存在重大障碍，例如 缺乏分析技术和分子工具来准确检测和表征这些 致癌元素。这里提出的工作将消除这些进展障碍。我们假设 ecDNA 驱动肿瘤发生的作用超出了其作为癌基因激活剂的作用，并且 ecDNA 的机制 形成和复制可以作为潜在的治疗靶点。为了检验我们的假设，我们将开发 可视化和追踪活细胞中 ecDNA 的方法，并使用这些工具来定义 ecDNA 的机制 肿瘤内的定位和转移。我们将从功能上评估 ecDNA 是否介导顺式调节 可以调节非ecDNA基因的转录来改变肿瘤细胞的行为。最后我们将探讨 ecDNA 的药理学消耗是否会影响细胞和动物中的肿瘤生长和维持 模型系统。我们的具体目标是： 1) 开发基于 CRISPR 的追踪工具，用于实时成像 ecDNA 动力学； 2) ecDNA介导的癌基因激活的发现和功能探究；和 3) 通过抑制核糖核苷酸还原酶 (RNR) 来靶向 ecDNA，该酶负责生成 脱氧核糖核苷酸三磷酸是常染色体和染色体外 DNA 的组成部分。 影响：该项目基于 ecDNA 具有癌症特异性且癌症- 实现基因组改变。尽管几十年前人们就认识到了 ecDNA，但这些结构的功能 以及如何有效地瞄准它们的起源，实际上是一个未知的领域。除了开发新的、广泛的 适用于 ecDNA 研究的工具，这项工作的结果将为特征提供新的见解， ecDNA 的行为和复制机制，并将评估 ecDNA 是否是未开发的癌细胞 可以在治疗上利用这些脆弱性来为患者谋福利。