Neural Progenitor Genes and Brain Tumors

描述（由申请人提供）：多形性胶质母细胞瘤（GBM）几乎是致命的。具有自我更新能力的肿瘤起始细胞（有时被称为“癌症干细胞”）的发现，为开发新的治疗途径创造了巨大的热情。这些细胞利用熟悉的途径进行增殖，如PI3激酶途径。尽管脑肿瘤干细胞样细胞的发现带来了希望，但在治疗发展的道路上仍存在许多障碍。一个并发症是这些细胞对常规疗法和通路抑制有明显的抵抗力。另一个是存在于不同患者肿瘤中的脑肿瘤干细胞之间存在差异。本研究的目的是批判性地研究脑肿瘤干细胞样细胞生物学，以便开发攻击它们的手段并克服它们的抵抗机制。首先，我们将通过使用癌症基因组图谱（TCGA）的最新进展来检查GBM干细胞样细胞的异质性。我们将从患者中获取样本，并根据基因表达分析确定的分子亚类对其进行分组。我们将评估这些细胞在体外产生神经球的能力以及在异种移植物中形成肿瘤的能力。然后，我们将使用药理学和基因操作策略来确定GBM干细胞样细胞对PI3K通路不同节点的依赖性。我们将确定TCGA定义的四个亚组（神经、前神经、间充质和经典）是否对这些淋巴结的增殖和肿瘤发生具有不同程度的依赖性。接下来，我们将评估PI3K通路在脑肿瘤干细胞样细胞中介导增强辐射抗性中的作用。我们将在体外测试这一假设，即激活该途径会增强对辐射的抵抗力，并确定我们是否可以通过抑制特定途径成分来逆转这种抵抗力。然后，我们将验证这一假设，即途径激活促进辐射抗性的机制之一是通过激活Nrf2氧化应激反应机制。然后，我们将探索GBM干细胞样细胞的化学耐药机制。我们将使用细胞培养、体内实验和一种新的基于微流体的免疫细胞化学分析（MIC）系统来确定雷帕霉素是否选择具有增强致瘤性和途径激活的干细胞样细胞。我们还将确定是否可以通过抑制过度激活的途径来克服对雷帕霉素治疗的耐药性。然后，我们将基于完整的磷酸化蛋白质组学筛选来确定新的耐药途径，以发现在雷帕霉素耐药发展过程中磷酸化或去磷酸化的蛋白质。我们将确定通过该筛选确定的蛋白质在耐药性发展中的潜在作用。这些合作研究将为更深入地了解GBM生物学铺平道路，并为GBM机制和治疗的未来临床、转化和临床研究提供信息。

DESCRIPTION (provided by applicant): Glioblastoma multiforme (GBM) is almost universally fatal. The discovery of tumor-initiating cells with the capacity to self-renew, sometimes termed "cancer stem cells", has created tremendous enthusiasm for the development of new avenues of therapy. These cells utilize familiar pathways for their proliferation, such as the PI3 Kinase pathway. Despite the hope raised by the discovery of brain tumor stem cell-like cells, numerous obstacles lie in the path of therapeutic development. One complication is that these cells have significant resistance to conventional therapies and to inhibition of pathways. Another is that there are differences amongst brain tumor stem-like cells that are present in the tumors of different patients. The goals of this study are to critically examine brain tumor stem cell-like cell biology in order to develop the means to attack them and to overcome their mechanisms of resistance. First, we will examine the heterogeneity of GBM stem cell-like cells through the use of recent advances by the The Cancer Genome Atlas (TCGA). We will obtain samples from patients and group them according to molecular subclasses defined through the analysis of gene expression. We will evaluate the ability of these cells to give rise to neurospheres in vitro as well as to form tumors in xenografts. We will then use a pharmacologic and gene manipulation strategy to determine the dependence of GBM stem cell-like cells on different nodes of the PI3K pathway. We will determine whether the four subgroups defined by the TCGA--Neural, Proneural, Mesenchymal and Classical--confer different levels of dependency on these nodes for proliferation and tumorigenesis. We will next assess the role of the PI3K pathway in mediating the enhanced resistance to radiation observed in brain tumor stem cell-like cells. We will test the hypothesis that activation of the pathway results in enhanced resistance to radiation in vitro and determine whether we can reverse this resistance through inhibition of specific pathway components. Then we will test the hypothesis that one of the mechanisms by which pathway activation promotes radiation resistance is through the activation of the Nrf2 oxidative stress-response mechanism. We will then explore mechanisms of chemoresistance in GBM stem cell-like cells. We will use cell culture, in vivo assays and a new microfluidicsbased immunocytochemical analysis (MIC) system to determine whether rapamycin selects for stem cell-like cells with enhanced tumorigenicity and pathway activation. We will also determine whether resistance to rapamycin treatment can be overcome through inhibition of hyperactivated pathways. Then, we will identify novel pathways of resistance based on a completed phosphoproteomic screen to discover proteins that are phosphorylated or dephosphorylated during the development of rapamycin resistance. We will determine the potential role of the proteins identified by this screen in the development of resistance. These collaborative studies will pave the way for a deeper understanding of GBM biology and inform future clinical and translational and clinical research into the mechanisms and treatment of GBM. PUBLIC HEALTH RELEVANCE: Brain tumors, especially glioblastoma multiforme (GBM) are highly lethal. This project seeks to understand cells that are found within GBM, sometimes called cancer stem cells, and how these cells differ from patient to patient and how they resist conventional therapies, such as radiation and chemotherapy. In performing these studies, we hope to develop better treatment strategies for GBM.