New Mechanisms of and Mouse Models for Medulloblastoma

描述（由申请人提供）：髓母细胞瘤是原始神经外胚层肿瘤（PNET）的一种常见形式，是最恶性的小儿脑肿瘤之一，平均5年生存率仅为50%。髓母细胞瘤被认为起源于小脑的未分化外颗粒层（EGL）细胞，但其机制尚不清楚。我们的初步结果提示了髓母细胞瘤发生发展的新机制。我们发现髓母细胞瘤细胞系过表达一种转录抑制因子，RE 1-沉默转录因子（REST）/神经元限制性沉默因子（NRSF），其对抗REST/NRSF的作用，抑制细胞在裸鼠中的致瘤潜力，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，当神经元干细胞（NSC），通常不表达REST/NRSF，被改造成过表达REST/NRSF，他们形成颅内PNET样肿瘤在裸鼠。因此，我们的工作假设是，过度表达REST/NRSF在神经干细胞引起髓母细胞瘤在体内。在这一补助金中，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来检验，C17.2类似于EGL细胞，并且可以在颅内植入后追踪。REST/NRSF和另一种报告基因（GFP）将在C17.2细胞中稳定且可诱导地共表达，并将在新生小鼠、幼龄小鼠和成年小鼠中检查其颅内致瘤潜力。REST/NRSF的两个阻遏物结构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是一种通过光学成像研究肿瘤特性的非侵入性系统。其次，将构建表达可诱导水平的REST/NRSF的转基因小鼠，以确定在生理条件下该假设是否正确。第三，由于REST/NRSF通过调节染色质来控制转录，因此最近开发的新型染色质重建系统将用于识别这些相互作用并检查它们在髓母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，拟议的研究将产生关键信息，对我们的长期目标，研究髓母细胞瘤的起源，并产生机制为基础的动物模型，可用于确定新的，生理相关的治疗目标，并测试现有的和新的药物的髓母细胞瘤。

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.