New Mechanisms of and Mouse Models for Medulloblastoma

描述（申请人提供）：髓母细胞瘤是一种常见的原始神经外胚层肿瘤（PNET），是儿童脑肿瘤中最恶性的肿瘤之一，平均5年生存率仅为50%。髓母细胞瘤被认为起源于小脑未分化的外颗粒层（EGL）细胞，但其机制尚不清楚。我们的初步结果提示了成神经管细胞瘤发生的一种新机制。我们发现髓母细胞瘤细胞系过表达一种转录抑制因子，re1 -沉默转录因子(REST)/神经元限制性沉默因子（NRSF），在裸鼠中对抗REST/NRSF的作用抑制细胞的致瘤潜能，并且50%的人髓母细胞瘤肿瘤标本过表达REST/NRSF。我们最近的实验表明，当正常情况下不表达REST/NRSF的神经干细胞（NSCs）被改造成过度表达REST/NRSF时，它们在裸鼠体内形成颅内pnet样肿瘤。因此，我们的工作假设是NSCs中REST/NRSF的过表达导致体内成神经管细胞瘤。在这项拨款中，提出了以下3个具体目标。首先，我们的假设将通过使用报告基因标记的NSC系C17.2进行验证，该系类似于EGL细胞，可以在颅内植入后进行追踪。REST/NRSF和另一个报告基因（GFP）将在C17.2细胞中稳定和诱导共表达，并将在新生、年轻和成年小鼠中检测其颅内致瘤潜力。REST/NRSF的两个抑制域在这一过程中的作用也将被确定。由于GFP的表达，这种原位小鼠模型也将成为通过光学成像研究肿瘤特性的非侵入性系统。其次，构建表达可诱导水平REST/NRSF的转基因小鼠，以确定该假设在生理条件下是否正确。第三，由于REST/NRSF通过调节染色质来控制转录，最近开发的一种新的染色质重构系统将用于识别这些相互作用并检查它们在成神经管细胞瘤形成中的作用。因此，通过结合发育生物学、转录生物学和癌症生物学，所提出的研究将为我们研究成神经管细胞瘤发生的长期目标提供关键信息，并产生基于机制的动物模型，这些模型可用于识别新的、生理学相关的治疗靶点，并测试现有的和新的治疗成神经管细胞瘤的药物。

DESCRIPTION (provided by applicant): Medulloblastoma, a common form of primitive neuroectodermal tumor (PNET), is among the most malignant of the pediatric brain tumors with an average 5-year survival rate of only 50%. Medulloblastoma is believed to arise from the undifferentiated external granule layer (EGL) cells in the cerebellum but the mechanism remains unknown. Our preliminary results suggested a novel mechanism for the development of medulloblastoma. We found that medulloblastoma cell lines overexpress a transcriptional repressor, RE1-Silencing Transcription Factor (REST)/Neuron Restrictive Silencer Factor (NRSF), that countering the actions of REST/NRSF inhibits the cells' tumorigenic potential in nude mice, and that 50% of human medulloblastoma tumor specimens overexpress REST/NRSF. Our more recent experiments indicated that when neuronal stem cells (NSCs), which do not normally express REST/NRSF, are engineered to overexpress REST/NRSF, they form intracranial PNET-like tumors in nude mice. Thus, our working hypothesis is that overexpression of REST/NRSF in NSCs cause medulloblastoma in vivo. In this grant, the following 3 Specific Aims are proposed. First, our hypothesis will be tested by, using the reporter gene-tagged NSC line, C17.2, which resemble EGL cells and can be traced after intracranial implantation. REST/NRSF and another reporter gene (GFP) will be stably and inducibly coexpressed in C17.2 cells and their intracranial tumorigenic potential will be examined in newborn, young, and adult mice. The role of the two repressor domains of REST/NRSF in this process will also be determined. Because of the GFP expression, this orthotopic mouse model will also be a noninvasive system for studying tumor properties by optical imaging. Second, transgenic mice expressing inducible levels of REST/NRSF will be constructed to determine whether the hypothesis is correct under physiological conditions. Third, as REST/NRSF controls transcription by modulating chromatin, a recently developed novel chromatin reconstitution system will be used to identify these interactions and examine their roles in medulloblastoma formation. Thus, by combining developmental biology, transcription biology, and cancer biology, the proposed studies will yield critical information towards our long-term goal of studying the genesis of medulloblastoma and produce mechanism-based animal models that can be used to identify new, physiologically relevant targets for therapy and to test both existing and new drugs for medulloblastoma.