Role of PTEN and PI-3 Kinase in Medulloblastomagenesis

髓母细胞瘤(MB)是一种发生在小脑的癌症，是最常见的恶性脑肿瘤。 儿童肿瘤。目前依靠放疗和化疗的治疗只治愈了60%的患者， 并在幸存者中有严重的长期神经认知后遗症，因此未得到满足的医学需求需要发展 治疗这种疾病的更有效、毒性更低的疗法。包括声波刺猬在内的恶性脑瘤 (Shh)MB亚群由表达该细胞的肿瘤增殖细胞(肿瘤干细胞；CSCs)驱动 表面糖蛋白CD15。存在知识鸿沟，因为控制CSC和Non的分子机制 CSC表型描述较差。这一提议的中心假设是PI3K/PTEN 信号轴控制SHH MB的CSC表型，因此是治疗的靶点 使用PI3K抑制剂进行开发。重要的是，我们实验室的研究表明，PTEN 在Smo转基因(SHH途径)小鼠MB模型的CSC隔室中，表达被抑制。 我们的发现突显了PTEN缺陷的Smo转基因小鼠模型的临床相关性 PTEN在人MB中的表达缺失，并且观察到PTEN缺失与MB的低存活率相关 病人。我们的资助提案，根据我们的初步数据，提出了3个具体目标：1)确定基因 PTEN或PI3K特定亚型的改变对于SHH驱动的CSC表型是必要的和/或充分的 和肿瘤发生2)利用高通量的基因文库筛选和小鼠和 人类MB CSCs和非CSCs用于识别分子签名和合成致死效应 预测对PI-3K抑制剂的敏感性或耐药性。3)阐明PTEN的表观遗传学机制。 在人SHH PDX模型和SmoTg GEMM模型中调节CSC间隔；重点是 PTEN特异的miRNA调控网络。我们将使用Math1creER重组酶来生成 正常小脑颗粒神经元前体细胞中PTEN、p110和/或p110的条件性缺失 (GNP)或在CSC隔室，并确定对MB和CSC表型发育的影响。 在目标2中，我们将扩展我们最近的报告，展示了PI3K抑制剂BKM120的活性 针对CSCS，将合成致命性筛选与其他靶向制剂和常规 化疗药物。我们应用程序的一个创新部分是最近发现的一种新的miRNA 在与CSC相关的CSC间隔中，表观遗传调节PTEN的网络 表型。这一观察结果将在我们的GEMM、MB PDX模型和人类MB肿瘤中进行探索 以确定这些miRNAs在调节PTEN、PI3K和CSC表型中的作用 包括肿瘤的启动活性。拟议的工作具有重要意义，因为它将：1)建立PTEN的作用， 脑脊液干细胞生物学中的p110和p1102)结合抑制PI3K鉴定新的合成致死性 在MB CSCs中，3)阐明miRNA网络在控制PTEN和CSC表型中的作用。

Medulloblastoma (MB) is a cancer arising within the cerebellum, and is the most common malignant brain tumor in children. Current care which relies on radiotherapy and chemotherapy, only cures 60% of patients, and has serious long-term neurocognitive sequelae in survivors, hence the unmet medical need to develop more efficacious, less toxic therapies for this disease. Malignant brain tumors including the sonic hedgehog (SHH) subgroup of MB are driven by tumor propagating cells (cancer stem cells; CSCs) expressing the cell surface glycoprotein CD15. A knowledge gap exist in that the molecular mechanisms controlling CSC and non CSC phenotypes are poorly delineated. The central hypothesis for this proposal is that the PI3K/PTEN signaling axis exerts control over the CSC phenotype in SHH MB and hence represents a target for therapeutic exploitation with PI3K inhibitors. Importantly, work from our laboratory has demonstrated that PTEN expression is suppressed in the CSC compartment of the Smo transgenic (SHH pathway) mouse model of MB. The clinical relevance of the PTEN deficient Smo transgene mouse model is highlighted by our finding that PTEN expression is lost in human MB, and the observation that PTEN loss correlates with poor survival in MB patients. Our grant proposal, guided by our preliminary data proposes 3 specific aims: 1) Determine if genetic alterations in PTEN or specific isoforms of PI3K are necessary and/or sufficient for SHH driven CSC phenotype and tumorigenesis 2) Utilize high throughput kinome library screens and genomic profiling of mouse and human MB CSCs and non CSCs to identify a molecular signature and synthetic lethality effects which will predict sensitivity or resistance to a PI-3K inhibitor. 3) Elucidate the epigenetic mechanism by which PTEN is regulated in the CSC compartment in human SHH PDX models and the SmoTg GEMM model; Focused on PTEN specific miRNA regulatory networks. We will use the Math1creER recombinase to generate the conditional deletion of PTEN, p110 and/or p110 in normal cerebellar granule neuronal precursor cells (GNPs) or in the CSC compartment and determine the effects on the development of MB and CSC phenotype. In Aim 2, we will expand upon our recent report demonstrating the activity of the PI3K inhibitor, BKM120 against CSCs to include synthetic lethality screens with other targeted agents and conventional chemotherapeutics. An innovative component of our application is the recent discovery of a novel miRNA network which epigenetically regulates PTEN in the CSC compartment which is associated with the CSC phenotype. This observation will be explored in our GEMMs , MB PDX models and in human MB tumor samples to determine the role of these miRNAs in the regulation of PTEN, PI3K and the CSC phenotypes including tumor initiating activity. The proposed work is significant in that it will: 1) establish a role for PTEN, p110 and p110 in CSC biology in MB 2) identify novel synthetic lethalities in combination with PI3K inhibition in MB CSCs and 3) elucidate a role for miRNA networks in the control of PTEN and CSC phenotypes.