A role for notch in self renewal in embryonal rhabdomyosarcoma

缺口在胚胎横纹肌肉瘤自我更新中的作用

基本信息

批准号：
9485912
负责人：
Myron Steve Ignatius
金额：
$ 24.9万
依托单位：
UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-05-17 至 2020-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9485912
关键词：
Animals Biological Assay Candidate Disease Gene Cell Fraction Cell Line Cell Transplantation Cell division Cells Child Childhood Soft Tissue Sarcoma Clinic Clinical DNA Binding Data Disease Embryonal Rhabdomyosarcoma Family member Fishes Frequencies Gene Chips Genes Genetic Epistasis Goals Growth Human Image Immune In Vitro Label Laboratories Malignant Childhood Neoplasm Methods Modeling Molecular Mus Muscle Mutation NOTCH1 gene PAX7 gene Pathologic Pathway interactions Patients Population Process Regulatory Element Relapse Role Sampling Self-Direction Signal Transduction Therapeutic Time Tissues Transgenic Model Transgenic Organisms Transplantation Tumor Expansion Update Zebrafish base clinically relevant diagnostic biomarker experimental study human disease in vivo in vivo imaging inhibitor/antagonist knock-down neoplastic cell notch protein novel outcome forecast overexpression relapse patients satellite cell self renewing cell self-renewal small hairpin RNA tumor tumor growth tumor heterogeneity tumor microenvironment

项目摘要

Self-renewing tumor-propagating cells drive continued tumor growth and are responsible for relapse. If the process by which tumor cells self-renew could be turned off, then tumors would regress and patients would remain relapse free. The goal of this updated proposal is to define the cellular and molecular mechanisms by which Notch regulates tumor-propagating potential and plasticity of the tumor propagating cell state in embryonal rhadomyosarcoma (ERMS), a devastating pediatric malignancy of the muscle. Relapse is the major clinical problem facing patients with ERMS, with less than 40% of relapse patients surviving their disease. Progress on this project using a combination of in vivo experiments in the zebrafish ERMS model and in in vitro experiments using ERMS cell lines and primary tissues has validated the hypothesis that Notch pathway activation increases the pool of tumor- propagating cells (TPCs), but rather surprisingly in vivo cell transplantation experiments finds that Notch enables the dedifferentiation of non-TPCs into TPCs. Using human patient samples, ERMS cell lines and correlative data in zebrafish has identified critical Notch regulated targets in human ERMS including SNAI1, MEF2C, PAX7 and MYF5. Preliminary data within my proposal shows that RAS-driven ERMS contain a molecularly distinct population of ERMS-propagating cells that express high levels of myf5 but lack differentiated muscle marker expression. These cells can be directly visualized in live, fluorescent- transgenic zebrafish, allowing unprecedented access to visualize self-renewal in live animals. Building on these observations, my proposal will determine the cellular and molecular mechanisms by which Notch alters tumor-propagating potential in both zebrafish and human ERMS. Specifically, Aim 1 will assess if Notch pathway activation alters symmetric vs. asymmetric divisions in the ERMS-propagating cell subfraction by dynamic real-time imaging of live, fluorescent transgenic fish. A sub aim will use lineage tracing methods to define the frequency and dynamics of dedifferentiation to make TPCs in ERMS. Aim 2 Will show that NOTCH1 expands TPCs in vivo in human ERMS by utilizing limiting dilution cell transplantation of low passage human primary ERMS cells into immune compromised mice. Aim 3 will assess the molecular mechanisms by which downstream NOTCH1 effector genes SNAI1, PAX7, MYF5 and MEF2C expands self-renewal, drives dedifferentiation and blocks terminal differentiation. In total, my proposal provides a comprehensive strategy to interrogate how the Notch pathway regulates ERMS self- renewal and will likely have immense therapeutic significance as clinically-relevant Notch pathway inhibitors would likely reduce tumor propagating cell frequency, dedifferentiation and ultimately relapse.

自我更新的肿瘤传播细胞驱动肿瘤的持续生长，并负责复发。如果过程通过哪些肿瘤细胞可以自我更新，然后肿瘤会消退，并且患者将继续复发自由的。该更新建议的目的是定义细胞和分子机制 Notch调节肿瘤传播的潜力和可塑性，使肿瘤传播细胞状态胚胎性乳腺癌（ERMS），肌肉的毁灭性小儿恶性肿瘤。复发是主要的 ERMS患者面临的临床问题，不到40％的复发患者幸存下来。进步在该项目上，使用斑马鱼ERMS模型中的体内实验和体外实验的组合使用ERMS细胞系和原发性组织已验证了Notch途径激活增加的假设肿瘤传播细胞（TPC），但令人惊讶的是体内细胞移植实验发现 Notch可以将非TPC的去分化为TPC。使用人类患者样品，ERMS细胞系和斑马鱼中的相关数据已经确定了包括snai1的人类ERM中的关键缺口调节靶标 MEF2C，PAX7和MYF5。我的提案中的初步数据表明，RAS驱动的ERMS包含一个表达高水平MyF5但缺乏分化的ERMS传播细胞的分子群体种群肌肉标记表达。这些细胞可以直接在现场荧光转基因斑马鱼中观察到前所未有的访问可视化活体动物中的自我更新。基于这些观察，我的建议将确定斑马鱼两种肿瘤传播潜力的细胞和分子机制和人类。具体而言，AIM 1将评估Notch途径激活是否改变对称与不对称通过活荧光转基因鱼类的动态实时成像，在ERMS传播细胞折叠的分裂。子目标将使用谱系跟踪方法来定义去分化的频率和动力学以使TPCS 在Erms。 AIM 2将表明Notch1通过利用限制稀释单元，在人类ERM中扩展了TPC 将低通道的人类原代ERMS细胞移植到免疫受损的小鼠中。 AIM 3将评估下游Notch1效应基因SNAI1，PAX7，MYF5和MEF2C的分子机制扩展自我更新，驱动去分化并阻止终端分化。总的来说，我的建议提供了综合策略来询问Notch途径如何调节ERMS自我续签，并可能有由于临床上与诊断途径抑制剂可能会减少肿瘤，因此具有巨大的治疗意义传播细胞频率，去分化并最终复发。