Lineage tracing and clonal analysis of oral cancer initiating cells

口腔癌起始细胞的谱系追踪和克隆分析

• 批准号: 8889248
• 负责人: Zhe Li
• 金额: $ 21.9万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-10 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8889248
• 关键词: Accounting; Adenoviruses; Affect; Behavior; Biological Assay; Breast Epithelial Cells; Cancer Model; Carcinogens; Cell Lineage; Cells; Chimeric Proteins; Collaborations; Daughter; Development; Diagnosis; Disease; Disease Resistance; Distant Metastasis; Ensure; Environment; Epithelial Cells; Erinaceidae; Estrogen Receptors; Exhibits; Fusion Protein Expression; Generations; Genetic Recombination; Goals; Habitats; Head and Neck Squamous Cell Carcinoma; Health; Human; Injection of therapeutic agent; Knowledge; Label; Life; Link; Malignant Epithelial Cell; Malignant Neoplasms; Malignant neoplasm of prostate; Mediating; Modeling; Molecular; Mus; Natural regeneration; Neoplasm Metastasis; Oral; Outcome; Pathway interactions; Pharmaceutical Preparations; Physiological; Population; Property; Recurrence; Reporter; Reporter Genes; Research; Resistance; Signal Pathway; Squamous Cell Neoplasms; Staging; Stem cells; Survival Rate; Tamoxifen; Testing; Therapeutic Intervention; Tissues; Translating; Transplantation; Tumor Expansion; Ursidae Family; Xenograft procedure; base; cancer cell; cancer stem cell; cancer type; cell type; chemotherapy; daughter cell; experience; genetic approach; in vitro Assay; in vivo; innovation; interest; leukemia; malignant breast neoplasm; malignant mouth neoplasm; medical schools; mouse model; mouth squamous cell carcinoma; new therapeutic target; notch protein; novel; novel strategies; novel therapeutic intervention; promoter; recombinase; reconstitution; research study; self-renewal; smoothened signaling pathway; stem cell fate; therapy resistant; tumor; tumor progression

DESCRIPTION (provided by applicant): Oral Squamous Cell Carcinomas (OSCCs) represent the vast majority cases of squamous cell carcinomas of the head and neck. Currently the 5-year survival rate for OSCCs remains at ~50-60%, largely due to their therapy resistance, recurrence, distant metastasis, and late diagnosis. It has been proposed that in many cancer types, including OSCCs, cancer cells are organized into aberrant cell hierarchies, including cancer stem cells [CSCs, also known as cancer initiating cells (CICs)] and their more differentiated daughter cancer cells. Several recent studies in murine cancer models have used lineage tracing to identify cancer cells that bear markers of tissue stem cells, and which give ris to long-lived clones that drive tumor expansion. This in vivo lineage tracing approach typically involves tamoxifen-induced transient activation of a Cre recombinase-estrogen receptor fusion protein (CreERT), which leads to activation of a conditional Cre reporter (e.g., the Rosa26-Stop-YFP conditional reporter). CreERT expression can be controlled by a cell type-specific promoter and CreERT-expressing cells and their daughter cells can be genetically labeled by the Cre reporter (e.g., YFP) upon Cre-mediated recombination (thus forming a cell lineage). Although these recent lineage tracing studies in murine models have provided compelling evidence to support the existence of CSCs in intact tumors, they give few clues about the molecular mechanisms required to ensure CSC self- renewal, nor about whether different signaling pathways might control distinct CSC fates. It is also unclear whether CSCs form a homogeneous population or whether different subpopulations of CSCs might exhibit different behaviors (e.g., differential survival) upon chemotherapy. In OSCCs, although it is thought that CSCs are responsible for their metastasis, recurrence and therapy resistance, the identity of OSCC CSCs remains elusive. In this R21 project, an innovative approach is proposed to study CSCs during OSCC development and upon therapy in a well-established carcinogen-induced murine OSCC models by a novel pathway-based in vivo lineage tracing strategy. This approach involves genetically marking subsets of cancer cells based on activation of a panel of pathways known to be involved in regulating stem cells (e.g., Axin2-CreERT or Gli1-CreERT for labeling Wnt or Hedgehog signaling-responsive cells, respectively) (Aim 1). By quantitatively comparing clonal outcomes between different pathway-labeled cell populations, one can reveal which pathways are active in CSCs and how cell fate choices are affected by pathway activity. In Aim 2, how oral cancer-initiating cells/CSCs in this murine model (i.e., defined based on their activit for stem cell-related pathway) respond to therapeutic interventions at the clonal level will be further determined by lineage tracing. The long-term goal of this project is to use this pathway-based lineage tracing approach to identify which pathways correlate with generation and regeneration (upon therapy) of CSCs or with therapeutic resistance, and through this to develop and test novel therapy to target OSCC CSCs.

描述（由申请人提供）：口腔鳞状细胞癌（oscc）代表了头颈部鳞状细胞癌的绝大多数病例。目前，OSCCs的5年生存率维持在~50-60%，主要原因是其耐药、复发、远处转移和诊断较晚。有人提出，在包括OSCCs在内的许多癌症类型中，癌细胞被组织成异常的细胞等级，包括癌症干细胞[CSCs，也称为癌症起始细胞（CICs）]及其分化程度更高的子癌细胞。最近在小鼠癌症模型中进行的几项研究使用谱系追踪来识别携带组织干细胞标记的癌细胞，这些标记会导致长寿命克隆的风险，从而导致肿瘤扩张。这种体内系系追踪方法通常涉及他莫昔芬诱导的Cre重组酶-雌激素受体融合蛋白（CreERT）的短暂激活，从而导致条件Cre报告基因的激活（例如，Rosa26-Stop-YFP条件报告基因）。CreERT的表达可以由细胞类型特异性启动子控制，表达CreERT的细胞及其子细胞可以通过Cre介导的重组（从而形成细胞谱系）被Cre报告基因（例如，YFP）标记。尽管最近在小鼠模型中的谱系追踪研究提供了令人信服的证据来支持完整肿瘤中CSC的存在，但它们几乎没有提供关于确保CSC自我更新所需的分子机制的线索，也没有提供关于不同信号通路是否可能控制不同CSC命运的线索。同样不清楚的是，CSCs是否形成一个同质群体，或者不同的CSCs亚群在化疗后是否会表现出不同的行为（例如，差异生存）。在OSCC中，尽管人们认为csc与OSCC的转移、复发和治疗抵抗有关，但OSCC csc的身份仍然难以捉摸。在这个R21项目中，我们提出了一种创新的方法，通过一种新的基于途径的体内谱系追踪策略，在一种成熟的致癌物质诱导的小鼠OSCC模型中，研究OSCC发展过程中的csc和治疗后的csc。这种方法涉及基于激活一组已知参与调节干细胞的途径（例如，分别标记Wnt或Hedgehog信号应答细胞的Axin2-CreERT或Gli1-CreERT）对癌细胞亚群进行遗传标记（Aim 1）。通过定量比较不同途径标记的细胞群之间的克隆结果，可以揭示哪些途径在CSCs中是活跃的，以及途径活性如何影响细胞命运的选择。在Aim 2中，该小鼠模型中的口腔癌起始细胞/CSCs（即基于其干细胞相关途径的活性定义）如何在克隆水平上对治疗干预作出反应将通过谱系追踪进一步确定。该项目的长期目标是使用这种基于途径的谱系追踪方法来确定哪些途径与干细胞的产生和再生（治疗后）或治疗抗性相关，并通过此开发和测试针对OSCC csc的新疗法。