Single-cell genomic approaches to study the cellular origins of brain tumors

摘要 脑肿瘤由于其多样性、患病率、贫穷和缺乏营养， 对治疗的反应以及高发病率和死亡率。在美国有50多万人 患有原发性脑肿瘤或其他中枢神经系统肿瘤，约有80，000个新肿瘤 每年诊断。然而，脑肿瘤的起源-即每个脑肿瘤的特定细胞类型 肿瘤的产生-仍然是个谜，在我们对它们的生物学的理解中是一个基本的未知数。这 该项目将通过开发一种新的单细胞基因组学技术来识别细胞， 脑肿瘤的起源动物模型已经表明，“起源细胞”的表型-第一个细胞， 获得基因突变以引发肿瘤-通常在决定随后的肿瘤中起主要作用。 表型和行为。起源细胞的表观遗传遗产可能在人类脑肿瘤中起作用， 好吧，但是由于主要的技术原因， 局限性-缺乏系统追踪人体组织中细胞谱系的方法。我们将 通过三个相关的目标来解决这一研究挑战：1）开发一种新的单细胞技术，称为 TAPE首次能够从人体组织中重建高分辨率谱系树。 TAPEARTH将通过选择性地捕获数十个细胞中自发发生的体细胞突变来实现这一目标。 数千个单细胞，同时分析相同单细胞的转录组，从而 允许以可行的成本重建表型注释的谱系树; 2）应用 TAPE将正常大脑皮层中祖细胞谱系的层次分类作为参考， 随后鉴定该层次内单个脑肿瘤的起源细胞和早期谱系。 这一目标将集中在三种可能具有不同起源细胞的肿瘤类型：胶质母细胞瘤（最常见的， 最致命的恶性脑肿瘤），间变性少突胶质细胞瘤和原始神经外胚层肿瘤; 3）在 通过TAPERT-PCR鉴定的早期肿瘤谱系的原位空间定位，以了解它们是否占据了 肿瘤内的特定区域以及它们是否有助于肿瘤迁移和侵袭。总的来说，这项工作 将提供对脑肿瘤病因学中一个重要的未知因素--起源细胞--的深入了解， 假设在肿瘤生物学中起着重要但未被充分认识作用。识别起源细胞将 改变我们对脑肿瘤发生的原因、时间和地点、表型多样性的理解， 重要的是，可以促进早期检测，创造新的动物模型，以及设计谱系- 靶向治疗。此外，这里开发的技术有可能对癌症产生重大影响。 生物学和发育生物学的研究。

Summary Abstract Brain tumors are responsible for an immense burden of disease due to their diversity, prevalence, poor response to therapies, and high morbidity and mortality. More than half a million people in the United States have a primary brain tumor or other central nervous system tumor, and about 80,000 new tumors are diagnosed each year. However, the origins of brain tumors— i.e. the specific cell type from which each brain tumor arises— remains enigmatic and is a fundamental unknown in our understanding of their biology. This project will address this major gap by developing a novel single-cell genomics technology to identify the cells of origin of brain tumors. Animal models have shown that the phenotype of the “cell of origin”— the first cell that acquires the genetic mutations to initiate a tumor— often plays a major role in determining subsequent tumor phenotype and behavior. The epigenetic legacy of the cell of origin is likely at play in human brain tumors as well, but the identification of tumor cells of origin has not been possible due to a major technological limitation— the absence of a method for systematically tracing lineages of cells in human tissues. We will address this research challenge via three related aims: 1) Development of a novel single-cell technology called TAPESTRY capable of reconstructing high-resolution lineage trees from human tissues for the first time. TAPESTRY will achieve this by selectively capturing spontaneously occurring somatic mutations in tens of thousands of single cells, while simultaneously profiling the transcriptomes of the same single cells, thereby allowing reconstruction of phenotypically annotated lineage trees at feasible costs; 2) Application of TAPESTRY to catalogue the hierarchy of progenitor cell lineages in normal cerebral cortex as a reference, followed by identification of the cells of origin and early lineages of individual brain tumors within that hierarchy. This aim will focus on three tumor types with likely different cells of origin: glioblastoma (the most common and most lethal malignant brain tumor), anaplastic oligodendroglioma, and primitive neuroectodermal tumors; 3) In situ spatial mapping of the early tumor lineages identified by TAPESTRY to understand whether they occupy specific domains within tumors and whether they contribute to tumor migration and invasion. Overall, this work will provide insight into a significant unknown factor in brain tumor etiology—the cell of origin—which, we hypothesize plays a major and under-appreciated role in tumor biology. Identifying the cell of origin would transform our understanding of why brain tumors arise where and when they do, their phenotypic diversity, and importantly, could facilitate earlier detection, the creation of new animal models, and the design of lineage- targeted therapies. Moreover, the technology developed here has the potential for major impact across cancer biology and developmental biology research more generally.