Modulation of Glioma Growth and Invasion by HCMV Glycoprotein B

描述(申请人提供)：胶质母细胞瘤(GBM)是成人最常见的原发脑肿瘤，通常是致命的。本实验室首次证实人类巨细胞病毒(HCMV)核酸和蛋白质存在于90%以上的人类恶性胶质瘤中，这一发现现已被三个独立的研究小组证实。最近的研究表明，HCMV感染水平与患者生存期呈负相关，提示HCMV可能诱导向更具侵袭性和侵袭性的胶质瘤表型转变。我们最近发表了两篇报道，证明在GBM细胞中，HCMV感染和基因表达可以诱导肿瘤生长关键的细胞酪氨酸激酶信号通路(Akt、PLC3、FAK)，抑制P53和Rb肿瘤抑制活性，改变细胞周期检查点控制，促进细胞增殖。我们还发现，HCMV激活了GBM细胞中强大的PDGFR1信号，而PDGFR1的激活实际上是HCMV感染所必需的。这一发现与胶质瘤生物学特别相关，因为PDGFR1在高比例的GBM细胞中被扩增/过表达，而在正常的NPC中，PDGFR1的激活与胶质瘤形成的早期阶段有关。我们已发表的和初步的数据表明，HCMV糖蛋白B(GB，最丰富的病毒包膜蛋白)是胶质瘤细胞中结合和酪氨酸磷酸化PDGFR1并激活下游PI3K-Akt信号的病毒部分。最近，我们有证据表明，GB诱导胶质瘤细胞向肝细胞迁移，并且HCMV诱导的胶质瘤跨壁迁移被PDGFR1阻断抗体或GB中和抗体抑制。综上所述，我们的数据提示，表达在胶质瘤细胞表面的HCMV-GB和PDGFR1之间的相互作用可能在调节胶质瘤的生长和侵袭中发挥关键作用。为了研究这些假说，我们建议使用体外、体外和体内试验，以测试HCMV感染和表达GB的人脑胶质瘤细胞是否表现出增强的生长和侵袭力。我们的实验室有独特的途径获得人类原代胶质母细胞瘤来源的培养物，这些培养物内源性表达几种HCMV基因产物，包括GB。我们计划使用这些原代胶质瘤培养物，超级感染临床HCMV分离株，并设计成异位表达GB，用于跨井迁移、脑片侵袭和体内肿瘤生长分析。如果我们发现HCMV和GB确实促进了胶质瘤的生长和侵袭性，我们计划研究这种调节的特定机制，包括细胞增殖、细胞侵袭和肿瘤血管生成。这些实验的结果将是理解巨细胞病毒在人类胶质瘤发病机制中所起作用的关键，并可能揭示基于抗病毒策略的针对这种无法治愈的人类癌症的新的治疗方法。 公共卫生相关性：我们的实验室发现一种常见的人类病毒，巨细胞病毒(HCMV)存在于90%以上的恶性胶质瘤中，这是一种普遍致命的高度侵袭性癌症，目前尚不清楚病因或治疗方法。本研究将探讨人巨细胞病毒包膜糖蛋白B在促进胶质瘤生长和侵袭中的作用。本文提出的实验结果可能导致基于抗病毒治疗的胶质瘤患者的新的治疗方法。

DESCRIPTION (provided by applicant): Glioblastoma (GBM) is the most common primary brain tumor in adults and is uniformly fatal. Our laboratory was the first to show that human cytomegalovirus (HCMV) nucleic acids and proteins are present in over 90% of human malignant gliomas, and these findings have now been confirmed by three independent groups. Recent studies have shown that HCMV infection levels negatively correlate with patient survival, suggesting that HCMV may induce a shift towards a more aggressive and invasive glioma phenotype. We recently published two reports demonstrating that HCMV infection and gene expression in GBM cells can induce cellular tyrosine kinase signaling pathways critical for tumor growth (Akt, PLC3, FAK), inhibit p53 and Rb tumor suppressor activity, alter cell cycle checkpoint controls, and promote cell proliferation. We have also discovered that HCMV activates robust PDGFR1 signaling in GBM cells, and that PDGFR1 activation is actually required for HCMV infection. This finding is particularly relevant to glioma biology since PDGFR1 is amplified/overexpressed in a high percentage of GBM cells and activation of PDGFR1 in normal NPCs is implicated in the earliest stages of gliomagenesis. Our published and preliminary data indicate that HCMV glycoprotein B (gB, the most abundant viral envelope protein) is the viral moiety that binds and Tyrosine phosphorylates PDGFR1 and activates downstream PI3K-Akt signaling in glioma cells. Most recently, we have evidence that gB induces haptotactic glioma cell migration and that HCMV-induced glioma transwell migration is inhibited both by PDGFR1 blocking antibodies or gB neutralizing antibodies. Taken together, our data suggest that the interaction between HCMV gB and PDGFR1 expressed on the surface of glioma cells may play a critical role in modulating glioma growth and invasion. To investigate these hypotheses, we propose to use in vitro, ex vivo, and in vivo assays which will test whether HCMV-infected and gB -expressing human glioma cells exhibit enhanced growth and invasiveness. Our laboratory has unique access to human primary glioblastoma-derived cultures which endogenously express several HCMV gene products, including gB. We plan to use these primary glioma cultures, super-infected with a clinical HCMV isolate and engineered to ectopically express gB for transwell migration, brain slice invasion, and in vivo tumor growth assays. If we find that HCMV and gB do promote glioma growth and invasiveness, we plan to investigate specific mechanisms underlying this modulation, including cell proliferation, cell invasion, and tumor angiogenesis. Results from these experiments will be critical in understanding the role that HCMV plays in the pathogenesis of human gliomas and may uncover novel therapeutic approaches against this incurable human cancer, based on antiviral strategies. PUBLIC HEALTH RELEVANCE: Our laboratory has found that a common human virus, cytomegalovirus (HCMV) is present in over 90% of malignant gliomas, a uniformly fatal, highly aggressive cancer, with no known etiology or treatment. This proposal will investigate the role of HCMV envelope glycoprotein B in promoting glioma growth and invasion. Results from experiments proposed herein may lead to novel therapeutic approaches based on antiviral treatments for glioma patients.