Investigating Cancer Stem Cells - Niche Interactions in Brain Tumor

描述(申请人提供)：胶质母细胞瘤(GBM)是人类最常见的恶性脑肿瘤，预后极差。迫切需要针对这种致命疾病的新的治疗策略。肿瘤干细胞(CSLC)是从基底膜中分离出来的，是肿瘤生长所必需的。然而，已有研究表明，GBM CSLC位于壁龛内(包括内皮细胞壁龛)，对化疗和放射治疗具有抵抗力。了解利基细胞和CSLC之间的分子通讯将有助于开发基于CSLC及其利基的GBM患者的新治疗策略。我们的长期目标是通过研究信号通路和肿瘤微环境调节GBM中CSLCs的分子机制，为GBM患者开发新的治疗策略。我们和其他人最近已经证明，用伽马分泌酶抑制剂(GSI)阻断Notch通路可以耗尽GBM CSLCs，抑制肿瘤生长，并延长携带颅内异种移植瘤的小鼠的生存时间。这项应用的总体目标是确定GBM CSLC中获得的Notch活性是否来自于内皮和分化的肿瘤细胞(NICE细胞)中表达的Notch配体，研究Notch信号在其缝隙内调节GBM CSLCs的分子机制，并在临床前模型中调查靶向GBM CSLCs和NICE细胞是否可以改善GBM的治疗。在特定的Aim1中，我们将研究表达在壁龛细胞(内皮细胞和分化的肿瘤细胞)中的Notch配体是否有助于GBM CSLCs中Notch的激活。我们将首先确定哪些Notch配体在原代GBM样本中的内皮细胞和分化的肿瘤细胞中表达。然后，我们将检查在体外和体内共同培养的GBM CSLCs中，敲除Notch配体是否会减少共同培养的GBM CSLCs的生长。在特定的AIM2中，我们将定义配体诱导的Notch激活调节GBM CSLC的分子机制。我们将利用Notch功能的获得和丢失研究来确定介导Notch调控的GBM CSLCs增殖、凋亡和分化的靶点。在特定的Aim3中，我们将研究同时针对CSLC及其利基细胞是否会在临床前小鼠模型中改善GBM的治疗。我们将研究GSI靶向GBM CSLCs和血管内皮生长因子抑制剂(阿瓦斯丁)靶向内皮细胞生态位的组合是否会提高携带人原代GBM来源的颅内异种移植的小鼠的存活率。目前的建议的成功不仅将加深我们对GBM CSLC与其利基之间的分子通讯的了解，还将有助于开发针对CSLC及其利基的GBM患者的新疗法。虽然目前的研究主要集中在基底膜，但这样的结果也将对多种形式的肿瘤具有普遍意义。

DESCRIPTION (provided by applicant): Glioblastoma (GBM) is the most common malignant brain tumor in human with extremely poor prognosis. Novel treatment strategies to this deadly disease are desperately needed. Cancer stem-like cells (CSLCs) have been prospectively isolated from GBM and shown required for tumor propagation. However, it has been shown that GBM CSLCs reside within niches (including endothelial niche) and are resistant to chemo- and radiation-therapy. Understanding the molecular communication between niche cells and CSLCs will help develop novel therapeutic strategies for GBM patients based on targeting both CSLCs and their niches. Our long-term goal is to develop novel therapeutic strategies for GBM patients through investigating the molecular mechanism by which signaling pathways and tumor microenvironment regulate CSLCs in GBM. We and others have demonstrated recently that Notch pathway blockade with a gamma-secretase inhibitor (GSI) depletes GBM CSLCs, inhibits tumor growth, and prolongs survival of mice bearing intracranial xenografts. The overall objective of this application is to define if acquired Notch activity in GBM CSLCs comes from Notch ligands expressed in endothelium and differentiated tumor cells (niche cells), to study the molecular mechanism by which Notch signaling regulates GBM CSLCs within their niches, and to investigate if targeting both GBM CSLCs and niche cells can improve the treatment of GBM in a pre-clinical model. In specific Aim1, we will examine if Notch ligands expressed in niche cells (endothelial cells and differentiated tumor cells) contribute to the activation of Notch in GBM CSLCs. We will first identify which Notch ligands are expressed in endothelial cells and differentiated tumor cells in primary GBM samples. Then, we will examine if knockdown of Notch ligands in niche cells will reduce growth of co-cultured GBM CSLCs in vitro and in vivo. In specific Aim2, we will define the molecular mechanism by which ligand-induced Notch activation regulates GBM CSLCs. We will use both gain and loss of Notch-function studies to identify the targets that mediate Notch-regulated proliferation, apoptosis, and differentiation in GBM CSLCs. In specific Aim3, we will examine if targeting both CSLCs and their niche will improve the treatment of GBM in a pre-clinical mouse model. We will examine if a combination targeting GBM CSLCs by GSI and targeting endothelial cell niche by VEGF inhibitor (Avastin) will improve survival of mice bearing intracranial xenografts derived from human primary GBMs. Success in the current proposal will not only enhance our understanding the molecular communication between GBM CSLCs and their niche, but will also help develop novel therapies for GBM patients based on targeting both CSLCs and their niches. Although the current research focuses on GBM, such results will also have general implications for multiple forms of neoplasia.