Harnessing Single Cell Technology to Define Self-Renewal in Normal and Malignant Stem Cells

利用单细胞技术定义正常和恶性干细胞的自我更新

• 批准号: 9350788
• 负责人: Jessica S. Blackburn
• 金额: $ 229.5万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-22 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9350788
• 关键词: Acute Lymphocytic Leukemia; Automobile Driving; Biology; Cancer Biology; Cancer Model; Cancer Relapse; Cell surface; Cells; Clinical; Data; Disease; Drug Design; Frequencies; Gene Expression Profile; Genomics; Goals; Hematopoietic stem cells; Human; Impairment; Label; Leukemic Cell; Maintenance; Malignant Neoplasms; Modeling; Normal tissue morphology; Organ; Patients; Pharmaceutical Preparations; Population; Preclinical Drug Evaluation; Relapse; Research Personnel; Role; Sampling; Stem cells; Technology; Tissues; Translating; Transplantation; Zebrafish; base; biomarker panel; cancer stem cell; cell type; chemotherapy; in vivo; leukemia; mouse model; neoplastic cell; outcome forecast; patient subsets; prevent; self renewing cell; self-renewal; single cell sequencing; single cell technology; stem cell niche; success; transcriptome sequencing; tumor; tumor progression

! Project Summary A major clinical issue in most cancers is relapse. Patients often respond very well to chemotherapy, and can go years without any sign of disease, but a subset of patients will invariably re- develop their cancer with a poor final prognosis. Relapse occurs because our current chemotherapies are unable to reliably and completely eliminate tumor propagating cells, also known as cancer stem cells. These cells are unique among the tumor cell population in that they can self-renew, meaning that they can replenish a tumor cell population indefinitely, similar to the role of the normal tissue stem cells in tissue and organ maintenance. Preventing self-renewal in tumor propagating cells would cause them to terminally differentiate, thereby blocking their ability to form relapse. Unfortunately, self-renewal of tumor propagating cells is not well understood, precluding rational drug design. A major issue in regards to studying these cells is their rarity; they often comprise 1 in every 105-107 cells within the total tumor cell population in human cancers and mouse models, and culture ex vivo alters their self-renewal capability. Researchers necessarily rely on FACS enrichment based on certain cell surface markers, but this biases towards the cells expressing the markers and excludes some subsets of self-renewing cells. The goal of this project is to have a major impact in the biomedical field by defining self-renewal in tumor propagating cells in a completely unbiased manner. We will use leukemia propagating cells as a model and determine how these cells differ from normal hematopoeitic stem cells, which can also self- renew, and how are they unique from other leukemic cells that cannot self-renew. Based on these data, we will find ways to detect leukemia propagating cells in patients, and identify drugs that can inhibit their self-renewal ability. Initially, we will use a panel of high self-renewing acute lymphoblastic leukemias and normal hematopoietic stem cells isolated from zebrafish models, which will allow us to use single cell RNA sequencing to identify the unique gene expression profile of self-renewing leukemia propagating cells without the need for FACS enrichment. We will translate our findings to human cells to build a biomarker panel that can detect the frequency of self-renewing cells in patient samples, and tell us whether chemotherapy has successfully eliminated them. We will also characterize the stem cell niche in leukemias with high and low self-renewal rates, to identify how the niche is regulating self-renewal rate. Finally, we will use transplantation approaches in zebrafish, as well as new zebrafish models in which the leukemia propagating cells are fluorescently labeled, for high-throughput, in vivo drug screens to identify compounds that impair self-renewal. In total, this project will provide an unbiased genomic and functional analysis of tumor propagating cells, allowing us to answer fundamental questions about the biology of this important tumor cell type.

呢 项目摘要 在大多数癌症中，一个主要的临床问题是复发。病人经常对 化学疗法，并且可能没有任何疾病的迹象，但一部分患者将总是会重新 最终预后不良发展癌症。复发是因为我们当前的化学疗法 无法可靠，完全消除传播细胞的肿瘤，也称为癌细胞。 这些细胞在肿瘤细胞群中是独一无二的，因为它们可以自我更新，这意味着它们 可以无限期地补充肿瘤细胞群体，类似于正常组织干细胞在 组织和器官维护。防止在肿瘤传播细胞中自我更新会导致它们 最终区分，从而阻止了它们形成复发的能力。不幸的是，肿瘤的自我更新 传播细胞尚不清楚，排除了理性的药物设计。关于 研究这些细胞是它们的稀有性。它们通常在总肿瘤细胞中每105-107个细胞中构成1个 人类癌症和小鼠模型中的种群，并且培养在体内改变了他们的自我更新能力。 研究人员必然依靠基于某些细胞表面标记的FACS富集，但这有偏见 朝向表达标记的细胞，排除一些自我更新细胞的子集。 该项目的目的是通过定义自我更新对生物医学领域产生重大影响 在肿瘤中以完全无偏的方式传播细胞。我们将使用白血病传播细胞作为 模型并确定这些细胞与正常造血干细胞的不同，这也可以自我自我 更新，以及它们与无法自我更新的其他白血病细胞的独特之处。基于这些数据， 我们将找到方法来检测患者中传播细胞的白血病，并确定可以抑制其其的药物 自我更新能力。最初，我们将使用一组高自我更新急性淋巴细胞白血病和 正常的造血干细胞从斑马鱼模型中分离出来，这将使我们能够使用单细胞 RNA测序以识别自我更新白血病的独特基因表达谱传播 细胞无需FACS富集。我们将把发现转化为人类细胞以建立 可以检测患者样品中自我更新细胞的频率并告诉我们的生物标志物面板 化学疗法是否成功消除了它们。我们还将表征干细胞小众 具有高自我更新率的白血病，以确定利基市场如何调节自我更新率。 最后，我们将在斑马鱼中使用移植方法，以及新的斑马鱼模型，其中 白血病传播细胞被荧光标记，用于高通量的体内药物筛选 损害自我更新的化合物。总体而言，该项目将提供公正的基因组和功能 分析肿瘤传播细胞，使我们能够回答有关生物学的基本问题 这种重要的肿瘤细胞类型。