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: 9887225
• 负责人: Roel GW Verhaak
• 金额: $ 51.81万
• 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 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9887225
• 关键词: 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; 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; Genomics; Glioblastoma; Goals; Growth; Heterogeneity; Hybrids; In Vitro; Knowledge; Label; Laboratories; Location; Maintenance; Malignant Neoplasms; Mediating; Methods; Methotrexate; Mitosis; Modeling; Molecular; Monitor; Oncogene Activation; Oncogenes; Oncogenic; Outcome; Patients; Pharmacology; 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 software; Work; base; cancer cell; cancer genome; cancer therapy; cancer type; cell behavior; cell killing; dosage; endonuclease; extrachromosomal DNA; gemcitabine; genetic element; genomic locus; hydroxyurea; in vivo Model; innovation; insight; interest; neoplastic cell; novel; 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.

项目摘要/摘要 该项目的目的是确定外肌体DNA（ECDNA）对肿瘤进化的影响 耐药性并验证基于ECDNA抑制的潜在治疗策略。存在 首先通过Arthur Spriggs的开创性细胞病理学研究认识到ECDNA超过五十年 前。我们知道，可以在所有固体的一半中检测到ECDNA在很大程度上是肿瘤细胞的现象 癌症，并经常表达致癌基因，使它们成为癌基因表达的车辆 和随之而来的肿瘤发生。我们在胶质母细胞瘤方面的研究扩展了这一知识，表明ECDNA是 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在很大程度上是不存在的。推进ecdna 研究以下方面的理解水平，我们现在必须定义其起源，复制机制和 针对治疗的策略。但是，目前，此类研究有重大障碍，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步的障碍。我们假设这一点 ECDNA驱动肿瘤发生超出其作为癌基因激活剂的作用，以及ECDNA的机制 形成和复制可以用作潜在的治疗靶标。为了检验我们的假设，我们将发展 可视化和跟踪活细胞中ecdna并使用这些工具定义ecdna机制的方法 肿瘤内定位和转移。我们将在功能上评估ECDNA是否介导的顺式调节 可以调节非ECDNA基因转录以改变肿瘤细胞行为。最后，我们将探索 ECDNA的药理耗竭是否会影响细胞和动物的肿瘤生长和维持 模型系统。我们的具体目的是：1）开发基于CRISPR的跟踪工具，以实时成像 ECDNA动力学； 2）ercDNA介导的癌基因激活的发现和功能询问； 3） 通过抑制核糖核苷酸还原酶（RNR）的靶向ECDNA，该酶负责产生 脱氧核糖核苷酸三磷酸是常染色体和外染色体外DNA的基础。 影响：该项目基于概念创新，即ECDNA是癌症特异性和癌症 - 实现基因组改变。尽管几十年前认识到ecdna，但这些结构的功能 以及如何有效地瞄准其起源，实际上是未知的领域。除了开发新的 适用的，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 可以利用治疗的脆弱性为患者利益。