Retrovirus Associated With Chronic Neurologic Disease

逆转录病毒与慢性神经系统疾病相关

• 批准号: 7143860
• 负责人: Steven Jacobson
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7143860
• 关键词: autoimmunity; clinical research; epilepsy; glycoproteins; host organism interaction; human T cell lymphotropic virus type 1; human herpesvirus 6; human subject; microorganism immunology; multiple sclerosis; neuroimmunomodulation; protein structure function; spastic paralysis; spinal cord disorders; virus cytopathogenic effect

The human T lymphotropic virus type I (HTLV-I) is the etiologic agent of a chronic progressive myelopathy known as HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP), a disease clinically similar to the chronic progressive form of multiple sclerosis (MS). While many viruses have been associated with MS, particular attention has recently focused on the human herpes virus type 6 (HHV-6). An understanding of the pathogenesis of a neurologic disease with a known viral etiology (HTLV-I and HAM/TSP) will aid in defining similar mechanisms of pathogenesis in MS, a disease of unknown etiology in which viruses have been suspected to play a role. Areas of research addressing these neurovirological and neuroimmunological issues include: I) The host immune response to viruses from patients with HAM/TSP and MS with particular focus on cytotoxic virus-specific CD8+ and CD4+ CD25+ T regulatory cells; II) The role of HHV-6 in the pathogenesis of MS and other chronic neurological disorders of the central nervous system; III) The translation of an understanding of these basic virological and immunological mechanisms associated with viruses associated with neurologic disease to immunotherapeutic strategies for the treatment of these disorders. The major findings of these studies have been: 1) Increased HTLV-I specific CD8+ cells have been shown to be elevated in the peripheral blood and CSF of HAM/TSP patients and directly proportional to the amount of HTLV-I proviral DNA and RNA. These antigen-specific T cells are considered to be immunopathogenic and may be directly involved in virus-host interactions in the CNS. These observations have been applied to help in the diagnosis of patients with multiple sclerosis who are infected with HTLV-I and do not have increased HTLV-I viral loads in their CSF. 2) Recently, dysregulation of the newly described cytokine, interleukin-15 (IL-15), has been implicated in the pathogenesis of a number of immune mediated diseases including HAM/TSP. Collaborative research by the Viral Immunology Section, NINDS and the Metabolism Branch, NCI suggests that up-regulation of IL-15 may sustain the persistent expansion of virus-specific CD8+ T cells in HAM/TSP. A phase I clinical trial for the treatment of HAM/TSP has been approved that blocks the action of IL-15 using a novel humanized monoclonal antibody, Hu MiK-beta-1. We will assess the effects of intravenously administered Hu-MiK-beta-1 on the cellular immune response in patients with HAM/TSP, with particular focus on virus-specific memory CD8+ T cells. Secondary outcomes to be measured will be clinical responses, including toxicity, and the effect on CD4+CD25+ T regulatory cells. As IL-15 over expression has been demonstrated in patients in a wide variety of autoimmune disorders including, rheumatoid arthritis, inflammatory bowel disease, multiple sclerosis and psoriasis, the successful application of Hu MiK-beta-1 in HAM/TSP has significant and broad therapeutic implications. 3) An understanding of how HTLV-I infects cells is an important area of investigation that may also lead to therapeutic strategies to intervene in HTLV-I associated disease. Recently, the glucose transporter type 1 (GLUT1) was reported to be the main binding receptor for HTLV-I. However, we have demonstrated that while overexpression of GLUT1 increased cell-cell transmission of HTLV-1, it did not increase either the binding of soluble HTLV-1 surface glycoproteins or the entry of HTLV-1 virions. Rather, we provide evidence that the majority of binding to the involved interactions with heparin surface proteoglycans (HSPG). These results indicate that both GLUT1 and HSPGs play an important role in HTLV infection. 4) CD4+CD25+ regulatory T cells are important in the maintenance of immunological self-tolerance and in the prevention of autoimmune diseases. We have demonstrated that in HTLV-I infected CD4+CD25+ T cells of patients with HAM/TSP the expression of the forkhead transcription factor Foxp3, a specific marker of regulatory T cells, was lower than that of healthy individuals. Importantly, we have generated cell lines that stably overexpress Fox P3 and have tested panels of anti-FoxP3 antibodies. We can show reduced protein expression of Fox-P3 positive T regulatory cells in patients with HAM/TSP. We also have demonstrated that HTLV-I tax had a direct inhibitory effect for the Foxp3 expression and inhibited the regulatory function of these cells. We are defining transcriptional mechanisms that control human FoxP3 and how HTLV-I tax acts to down regulate this important molecule. These results suggest that direct human retroviral infection of CD4+CD25+ T cells may be associated with the pathogenesis of HTLV-I associated neurologic disease through the dysregulation of CD4+CD25+ regulatory T cells. These studies are now being extended to characterize T regulatory cells in patients with multiple sclerosis. 5) Using a proteomic approach to generate disease specific protein profiles (Surface Enhanced Laser Desorption Ionization Time-Of-Flight Mass Spectroscopy (SELDI)) we have defined phenotypic ?signatures? within HTLV-1-infected sera that can discriminate HTLV-I infected patients with neurologic disease from patients with HTLV-I associated malignancies. These protein ?fingerprint? changes can be used for diagnostics, prognostics and early detection of disease. From these profiles, we have identified potential disease-specific biomarker proteins in sera. The identification of these proteins will allow for future development of immunoassays as well as insight into disease development. These studies will provide a tool for diagnosis/prediction of virus associated neurologic and hematologic disease and will provide the groundwork for similar approaches in other chronic, neurologic disorders. Towards this goal, we have begun an extensive proteomic profile analysis of sera from patients with multiple sclerosis and controls including healthy individuals and patients with other inflammatory and neurologic disease. Discrete protein profiles have been obtained that distinguish these groups. We are presently expanding these observations to larger numbers of multiple sclerosis patients to determine if signatures can be defined that discriminate severity and type of disease. 6) We continue to extend our work on the detection of HHV-6 from brain resections of patients with mesial temporal lobe epilepsy and patients with neurologic complications following allogeneic bone marrow transplants. Our observations that HHV-6 variant B can be demonstrated in epilepsy CNS tissue continue to be supported with larger number of samples. We have isolated this virus from explanted primary astrocyte cell cultures and have shown a dysregulation of glutamate uptake in HHV-6 infected astrocyte cell lines. As there are a number of antiviral HHV-6 compounds currently available we have shown that these drugs have different sensitivities in HHV-6 infected glial cells compared to virus-infected lymphocytes. These data suggest that different treatment strategies should be considered when trying to clear this virus from the periphery or the CNS. 7) We have developed a novel virus chip containing the complete open reading frames of 8 divergent viruses. We have defined the specificity and sensitivity of this chip and successfully used this new methodology to detect the expression of viral gene products from the CNS of a patient who died from encephalitis of unknown cause. In addition, we have shown the utility of this virus chip to temporally map the virus gene expression profile of HHV-6 in vitro. We are considering expanding this chip to include other viruses that have been associated with infection of the CNS.

人类T淋巴病毒I型（HTLV-I）是一种慢性进行性骨髓病的病因，称为HTLV-I相关的骨髓病/热带痉挛性拼发（HAM/TSP），一种疾病，临床上与多发性孢子菌病的慢性渐进形式相似（MS）。尽管许多病毒与MS有关，但最近特别关注的是人类疱疹病毒6型（HHV-6）。对具有已知病毒病因（HTLV-I和HAM/TSP）神经系统疾病的发病机理的理解将有助于定义MS中类似的发病机理，MS是一种未知病因的疾病，其中怀疑病毒被怀疑起着作用。解决这些神经病毒学和神经免疫学问题的研究领域包括：i）HAM/TSP和MS患者对病毒的免疫反应，特别关注细胞毒性病毒特异性CD8+和CD4+ CD4+ CD25+ T调节细胞； ii）HHV-6在中枢神经系统的MS和其他慢性神经系统疾病的发病机理中的作用； iii）对与神经疾病有关的病毒相关的这些基本病毒学和免疫机制的理解转化为治疗这些疾病的免疫治疗策略。这些研究的主要发现是： 1）在HAM/TSP患者的外周血和CSF中，HTLV-I特异性CD8+细胞的增加升高，与HTLV-I前病毒DNA和RNA的量直接成正比。这些抗原特异性T细胞被认为是免疫发病的，可能直接参与中枢神经系统中的病毒宿主相互作用。这些观察结果已用于帮助诊断患有HTLV-I的多发性硬化症患者，并且在CSF中没有增加HTLV-I病毒载量。 2）最近，新描述的细胞因子白介素15（IL-15）的失调与包括HAM/TSP在内的多种免疫介导的疾病的发病机理有关。病毒免疫学部分，NINDS和代谢分支的合作研究NCI表明，IL-15的上调可以维持HAM/TSP中病毒特异性CD8+ T细胞的持续扩展。已经批准了一项用于治疗HAM/TSP的I期临床试验，该试验使用一种新型的人源化单克隆抗体Hu Mik-Beta-1阻断了IL-15的作用。我们将评估HAM/TSP患者的静脉内施用HU-MIK-BETA-1对细胞免疫反应的影响，特别关注病毒特异性记忆CD8+ T细胞。要测量的次要结果将是临床反应，包括毒性，以及对CD4+ CD25+ T调节细胞的影响。由于在多种自身免疫性疾病中的患者中已经证明了IL-15表达，包括类风湿关节炎，炎症性肠病，多发性硬化症和牛皮癣，因此HU Mik-Beta-1在HAM/TSP中成功地应用了显着且广泛的治疗含义。 3）了解HTLV-I如何感染细胞是一个重要的研究领域，也可能导致治疗策略干预HTLV-I相关疾病。最近，据报道，葡萄糖转运蛋白1型（GLUT1）是HTLV-I的主要结合受体。但是，我们已经证明，尽管GLUT1的过表达增加了HTLV-1的细胞细胞传递，但它并没有增加可溶性HTLV-1表面糖蛋白的结合或HTLV-1病毒体的进入。相反，我们提供的证据表明，大多数与肝素表面蛋白聚糖（HSPG）相互作用的结合。这些结果表明，GLUT1和HSPG在HTLV感染中都起着重要作用。 4）CD4+ CD25+调节性T细胞在维持免疫学自我耐受和预防自身免疫性疾病方面很重要。我们已经证明，在HTLV-I感染HAM/TSP患者的CD4+ CD25+ T细胞中，叉子转录因子Foxp3（调节性T细胞的特定标记）的表达低于健康个体的表达。重要的是，我们产生了稳定表达FOX P3的细胞系，并测试了抗Foxp3抗体的面板。我们可以显示HAM/TSP患者中FOX-P3阳性T调节细胞的蛋白质表达降低。我们还证明，HTLV-I税对FOXP3表达具有直接的抑制作用，并抑制了这些细胞的调节功能。我们正在定义控制人FOXP3的转录机制，以及HTLV-I税收税如何降低调节这一重要分子的作用。这些结果表明，通过CD4+ CD25+调节性T细胞的失调，CD4+ CD25+ T细胞的直接人逆转录病毒感染可能与HTLV-I相关神经疾病的发病机理有关。现在，这些研究正在扩展以表征多发性硬化症患者的T调节细胞。 5）使用蛋白质组学方法产生特异性蛋白质谱（表面增强的激光解吸电离电离飞行时间质谱（SELDI））我们已经定义了表型？在HTLV-1感染的血清中，可以区分HTLV-I与HTLV-I相关恶性肿瘤患者的HTLV-I感染患者。这些蛋白质？指纹？变化可用于诊断，预后和疾病的早期检测。从这些特征中，我们发现了血清中潜在的疾病特异性生物标志物蛋白。这些蛋白质的鉴定将允许未来的免疫测定以及对疾病发展的见解。这些研究将为诊断/预测与病毒相关的神经系统疾病和血液学疾病提供工具，并为其他慢性神经系统疾病的类似方法提供基础。为了实现这一目标，我们已经开始对多发性硬化症患者的血清进行广泛的蛋白质组学分析，包括健康的个体以及其他炎症性和神经系统疾病的患者。已经获得了区分这些组的离散蛋白质谱。目前，我们正在将这些观察结果扩展到大量的多发性硬化症患者，以确定是否可以定义签名，以区分严重性和类型的疾病。 6）我们继续扩展有关在同种异体骨骨髓移植后患有颞叶颞叶癫痫患者和神经系统并发症患者的大脑切除术中检测HHV-6的工作。我们认为，可以在癫痫中枢神经系统组织中证明HHV-6变体B继续受到更多样品的支持。我们已经从植入的原发性星形胶质细胞培养物中分离出该病毒，并显示出HHV-6感染的星形胶质细胞细胞系中谷氨酸摄取的失调。由于目前有许多抗病毒HHV-6化合物，因此与感染病毒感染的淋巴细胞相比，这些药物对HHV-6感染的神经胶质细胞具有不同的敏感性。这些数据表明，在试图从外围或中枢神经系统中清除该病毒时，应考虑不同的治疗策略。 7）我们已经开发了一种新型病毒芯片，其中包含8种发散病毒的完整开放式阅读框。我们已经定义了这种芯片的特异性和敏感性，并成功地使用了这种新方法来检测死于未知原因脑炎的患者中枢神经系统中的病毒基因产物的表达。此外，我们已经显示了该病毒芯片的实用性，可以在体外暂时绘制HHV-6的病毒基因表达谱。我们正在考虑将这种芯片扩展到包括与CNS感染有关的其他病毒。