Development of Nanotherapeutics for RSV-induced Lung Disease

RSV 引起的肺部疾病纳米疗法的开发

• 批准号: 8140565
• 负责人: Shyam S Mohapatra
• 金额: --
• 依托单位: JAMES A. HALEY VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-10-01 至 2015-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8140565
• 关键词: 3-Dimensional; A549; Address; Adult; Affect; Aging; Antiviral Agents; Apoptosis; Apoptotic; Area; Attenuated; Binding; Biology; Bronchiolitis; CCL4 gene; Caspase; Cell Culture Techniques; Cell Death; Cell Membrane Permeability; Cell Survival; Cells; Cessation of life; Chitosan; Chronic lung disease; Complex; Cyclophosphamide; Data; Development; Dissociation; Doctor of Philosophy; Effectiveness; Elderly; Encapsulated; Epithelial Cells; Fibroblasts; Fluorescein-5-isothiocyanate; Fluorescence Microscopy; Funding; Genes; Genetic; Goals; Grant Review; Host Defense; Hour; Human; Hybrids; Immune; Immune response; Immune system; Immunocompromised Host; In Vitro; Inbred BALB C Mice; Infant; Infection; Influenza; Intercellular adhesion molecule 1; Interferons; Knowledge; Label; Laboratories; Lead; Leadership; Link; Lipids; Lung; Lung Inflammation; Lung diseases; Membrane Potentials; Metabolic; Methods; Mitochondria; Modeling; Molecular Medicine; Motivation; Mucous Membrane; Mus; Natural Immunity; Nature; Nonstructural Protein; Palivizumab; Pathogenesis; Peptide Signal Sequences; Peptides; Physiological; Physiology; Plasmids; Play; Pneumonia; Postdoctoral Fellow; Principal Investigator; Process; Production; Prophylactic treatment; Proteins; Publications; RNA Helicase; RNA Interference; Recombinants; Regulation; Reporting; Research; Research Personnel; Research Project Grants; Respiratory Syncytial Virus Infections; Respiratory Syncytial Virus Vaccines; Respiratory syncytial virus; Role; Safety; Scientist; Signal Pathway; Signal Transduction; Signaling Protein; Small Interfering RNA; Structure-Activity Relationship; System; Technology; Testing; Therapeutic; Therapeutic Agents; Time; Tissue Model; Translations; Treatment Efficacy; Tretinoin; Veterans; Viral; Virus; Virus Diseases; Virus Replication; Work; accomplished suicide; attenuation; base; career; caspase-9; cytochrome c; effective therapy; efficacy testing; fetal; in vivo; mitochondrial membrane; mouse model; nanoparticle; nanotherapeutic; novel; novel therapeutics; pathogen; preclinical study; premature; prevent; professor; programs; promoter; protein structure function; receptor; red fluorescent protein; respiratory; response; skills; targeted delivery; vector control; viral RNA

DESCRIPTION (provided by applicant): The primary goal of this proposal is to investigate the mechanism whereby respiratory syncytial virus (RSV) nonstructural protein 1 (NS1) modulates innate immunity and cell survival through the mitochondrial system following RSV infection, and to combine this knowledge with nanoparticle technology to develop new ways to prevent and treat RSV infections. Veterans commonly have some form of chronic lung disease and are especially susceptible to respiratory viral infections like those caused by RSV and influenza. RSV is an opportunistic human respiratory pathogen that causes bronchiolitis in infants and pneumonia in immunocompromised adults and the elderly with an estimated 64 million infections and 166,000 deaths annually worldwide. RSV is becoming more and more of a serious threat to aging veterans. Despite progress, the precise nature of the RSV-induced innate immune response remains unclear. Our preliminary evidence shows that the NS1 protein associates with mitochondria and is found in complex with the mitochondrial antiviral signaling (MAVS) protein in RSV-infected A549 epithelial cells and inhibits the signaling of retinoic acid-inducible gene-I (RIG-I) and MAVS occurring via the caspase activation and recruitment domain (CARD). This interferes with IFN-2 production which inhibits the innate immune response. NS1 also blocks programmed cell death (apoptosis) of infected cells and prevents loss of mitochondrial membrane potential, thus promoting cell (and viral) survival. These data have led to the hypothesis that RSV utilizes NS1 to attenuate RIG-I- MAVS-induced antiviral IFN-2 production and to modulate mitochondrial function for increasing cell survival. The study of the precise mechanisms underlying these processes is expected to lead to the discovery of new targets for preventing or limiting RSV infection. To test these hypotheses, the following specific aims are proposed. In the first aim, it is planned to examine the role of NS1 in attenuating RIG-I/MAVS signaling during RSV infection. This includes analysis of the role of NS1 in regulating MAVS, LGP2 and RIG-1 expression and in attenuation of IFN-2 response during RSV infection, mitochondrial localization of NS1 protein and structure-function relationships of NS1- MAVS interaction. The second aim will focus on determining whether the NS1- MAVS interaction is required to prevent premature apoptosis and intrinsic cell death. This also includes addressing whether the NS1-MAVS complex is associated with anti- or pro-apoptotic factors and modulates their expression on the mitochondrial membrane. In the third aim, it is planned to develop and test targeted chlipid (chitosan-lipid) nanoparticles as a therapeutic agent for RSV-induced lung disease. These chlipids encapsulate a plasmid that encodes siRNAs for LGP2 and NS1 and a peptide for MAVS (referred to as pLMNS1), each of which individually has been shown to significantly down- regulate RSV replication in human cells. Lastly, in the fourth aim it is proposed to evaluate targeted nanoparticle-encapsulated pLMNS1 for treating RSV disease in a mouse model. The proposed research will be conducted by an excellent group of investigators with a proven track record in RSV disease, apoptosis and nanoparticle technology. The results are expected to lead to the discovery of novel targets and to the initiation of preclinical studies of these targets against RSV lung disease.

描述（由申请人提供）： 该提案的主要目的是研究该机制，该机制通过RSV感染后通过线粒体系统调节了先天性免疫和细胞存活的机制，在RSV感染后调节先天免疫和细胞存活，并将这种知识与纳米技术结合起来，以开发新的方法来预防和治疗RSV Infections。退伍军人通常具有某种形式的慢性肺部疾病，并且特别容易受到RSV和流感引起的呼吸道病毒感染的影响。 RSV是一种机会主义的人类呼吸道病原体，可在免疫功能低下的成年人和老年人中引起婴儿和肺炎的细支气管炎，估计有6400万感染和每年166,000人死亡。 RSV正越来越成为对老年退伍军人的严重威胁。尽管取得了进展，但RSV诱导的先天免疫反应的确切性质尚不清楚。我们的初步证据表明，在RSV感染的A549上皮细胞中，NS1蛋白与线粒体相关，并与线粒体抗病毒信号传导（MAVS）蛋白相关，并抑制视黄酸诱导的基因-I（RIG-I）的信号，并通过Casspase（Caspase）抑制了信号。这会干扰IFN-2产生，从而抑制先天免疫反应。 NS1还阻止了被感染细胞的编程细胞死亡（凋亡），并防止线粒体膜电位的损失，从而促进细胞（和病毒）生存。这些数据导致了这样的假设：RSV利用NS1减轻了RIG-MAVS诱导的抗病毒IFN-2产生，并调节线粒体功能以增加细胞存活。对这些过程基础的确切机制的研究有望导致发现预防或限制RSV感染的新靶标。为了检验这些假设，提出了以下特定目标。在第一个目标中，计划检查NS1在RSV感染过程中衰减RIG-I/MAV信号传导中的作用。这包括分析NS1在调节MAV，LGP2和RIG-1表达以及在RSV感染期间IFN-2反应的衰减中的作用，NS1蛋白的线粒体定位以及NS1- MAVS相互作用的结构功能关系。第二个目标将重点侧重于确定是否需要NS1-MAVS相互作用以防止过早细胞凋亡和内在细胞死亡。这还包括解决NS1-MAVS复合物是否与抗凋亡因素有关，并调节其在线粒体膜上的表达。在第三个目的中，计划开发和测试目标chlipid（壳聚糖 - 脂质）纳米颗粒作为RSV诱导的肺部疾病的治疗剂。这些Chlipids封装了一个编码LGP2和NS1的siRNA的质粒，以及MAVS的肽（称为PLMNS1），每个siRNAS已被证明在人类细胞中显着下调了RSV复制。最后，在第四个目标中，提议评估靶向的纳米颗粒封装的PLMNS1，用于在小鼠模型中处理RSV疾病。拟议的研究将由一群出色的研究人员进行，在RSV疾病，凋亡和纳米颗粒技术方面具有良好的记录。预计结果将导致发现新靶标，并启动针对RSV肺部疾病的临床前研究。