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The role of TLR4 and RSV F protein in immunity to RSV

TLR4和RSV F蛋白在RSV免疫中的作用

基本信息

批准号：
8473654
负责人：
JORGE C BLANCO
金额：
$ 49.56万
依托单位：
SIGMOVIR BIOSYSTEMS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-01-01 至 2015-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8473654
关键词：
1 year old Adjuvant Agonist Alveolar Macrophages Alveolitis Animals Anti-Inflammatory Agents Anti-inflammatory Antibodies Antigens Binding Bronchiolitis Bronchopulmonary Dysplasia CD14 gene CXCL10 gene Case Series Cell surface Cells Cessation of life Child Clinical Trials Complex Computer Analysis Contracts Cotton Rats Coupled DNA DNA-Binding Proteins Data Detection Development Disease Elderly Employee Strikes Engineering Enzymes Exhibits Failure Family Formalin Frequencies Funding Gene Expression Profile Genes Genetic Polymorphism Genetic Transcription Genotype Goals Gram-Negative Bacteria Grant Histopathology Hospitalization IL8 gene Immune Immune response Immunity Immunologic Deficiency Syndromes In Vitro Infant Infection Inflammation Mediators Inflammatory Inflammatory Response Interferons Interleukin-12 Interleukin-6 Intervention Laboratories Lead Life Ligands Lipopolysaccharides Lung Lymphocyte Antigen 96 Mediating Membrane Modeling Molecular Molecular Conformation Molecular Genetics Monoclonal Antibodies Morbidity - disease rate Mus Outcome Palivizumab Pathologic Pathology Pathway interactions Pattern Pattern recognition receptor Phase Play Pneumonia Positioning Attribute Production Prostaglandins Protein Subunits Proteins Recombinants Recruitment Activity Reporting Resolution Respiratory Syncytial Virus Infections Respiratory Syncytial Virus Vaccines Respiratory System Respiratory syncytial virus Respiratory syncytial virus RSV F proteins Respiratory tract structure Risk Role STAT1 gene Sampling Sigmodon Signal Pathway Signal Transduction Signaling Molecule Single Nucleotide Polymorphism Structure Subunit Vaccines Surface TNF gene Testing Therapeutic Therapeutic Intervention Toll-like receptors Vaccinated Vaccination Vaccines Virus Diseases aged airway hyperresponsiveness analog autocrine base chemokine congenital heart disorder cyclooxygenase 2 cytokine high risk infant human IRF3 protein human NOS2A protein human TLR4 protein immunosuppressed in vivo innovation interferon regulatory factor-3 killings lung injury macrophage microbial monophosphoryl lipid A mortality novel novel therapeutics paracrine pathogen premature prophylactic protein activation prototype receptor expression response secondary infection therapeutic target toll-like receptor 4 transcription factor volunteer

项目摘要

The respiratory tract is a major portal for pathogens. The bronchoalveolar macrophage, positioned at the mucosal surface, recognizes "pathogen associated molecular patterns (PAMPs)" through "pattern recognition receptors (PRRs)." A family of mammalian PRRs, "Toll-like receptors" (TLRs), are transmembrane signaling molecules that respond to diverse PAMPs. Gram negative lipopolysaccharide (LPS) stimulates cells through TLR4 to elicit a strongly proinflammatory pattern of gene expression, resulting in a "Th1-type" cytokine milieu. Respiratory syncytial virus (RSV) is the leading cause of pneumonia and bronchiolitis in infants and young children worldwide, and has recently been attributed to increased morbidity and mortality in the elderly and immunosuppressed. The RSV fusion (F) protein is also a TLR4 agonist. Prophylactic administration of anti-F antibodies to high-risk infants is highly protective. In a failed clinical trial in the 1960's, a formalin-inactivated RSV (FI-RSV) vaccine led to exacerbated RSV disease, findings we have recapitulated in the cotton rat (S. hispidus), assessed by pulmonary histopathology and airway hyperreactivity. During the first cycle of this grant, we identified cyclooxygenase-2 and prostanoids as key therapeutic targets for RSV-induced lung pathology. We found that vaccination of cotton rats with the original FI-RSV used in the failed trials, newly formulated with a non-toxic adjuvant and TLR4 agonist, monophosphoryl lipid A (MPL), suppressed FI-RSV vaccine-enhanced disease by blunting the mixed Th1- and Th2-type "cytokine storm" that is elicited upon RSV infection of vaccinated subjects. In vitro, purified F protein activation of NF-¿B and IL-8 secretion in HEK293T cells is TLR4-, MD-2-, and CD14-dependent, and transfectants that express TLR4 proteins with one or both of two single nucleotide polymorphisms (SNPs), previously associated with LPS-hyporesponsiveness, were significantly less responsive to purified RSV F protein, under conditions of equal TLR expression. Importantly, we identified a highly significant overrepresentation of these TLR4 SNPs in DNA samples derived from a case series of high-risk infants and children with documented RSV infection. These data strongly support our overarching hypothesis that TLR4 plays a central role in the innate immune response to RSV and imply that initial engagement of TLR4 is required for development of a protective, adaptive immune response, rather than a pathological one. This proposal details innovative experimental approaches that will (i) lead to development a safe and effective RSV F protein subunit vaccine, (ii) lead to development of therapeutic intervention strategies based on a characterization of the interaction of F protein with the TLR4/MD-2/CD14 complex and examine the role(s) of TLR4 signaling in RSV infection/protection. It is expected that at the completion of this grant, we will have identified strategies that may lead to development of a RSV vaccine and new therapeutics for mitigating the pathologic host response to RSV.

呼吸道是病原体的主要门户。支气管肺泡巨噬细胞位于粘膜表面，通过“模式识别”病原体相关分子模式（PAMP）识别受体 (PRR)。”哺乳动物 PRR 家族“Toll 样受体”(TLR) 是响应不同 PAMP 的跨膜信号分子。革兰氏阴性脂多糖 (LPS) 通过 TLR4 刺激细胞，引发强烈的促炎基因表达模式，从而产生在“Th1型”细胞因子环境中。呼吸道合胞病毒（RSV）是引起肺炎和肺炎的主要原因全世界婴儿和幼儿中的细支气管炎，最近被归因于发病率增加以及老年人和免疫抑制者的死亡率。 RSV 融合 (F) 蛋白也是 TLR4 激动剂。对高危婴儿预防性施用抗 F 抗体具有高度保护作用。在一次失败的临床试验中在 20 世纪 60 年代，福尔马林灭活 RSV (FI-RSV) 疫苗导致 RSV 疾病加剧，我们发现通过肺组织病理学和气道评估，在棉鼠（S. hispidus）中进行了重述过度反应。在本次资助的第一个周期中，我们确定环氧合酶 2 和前列腺素类药物是关键 RSV 引起的肺部病理学的治疗靶点。我们发现用原疫苗接种棉鼠失败试验中使用的FI-RSV，采用无毒佐剂和TLR4激动剂新配制，单磷酰脂质 A (MPL)，通过削弱混合 Th1- 抑制 FI-RSV 疫苗增强的疾病接种疫苗的受试者感染 RSV 后引发的 Th2 型“细胞因子风暴”。体外纯化的 F HEK293T 细胞中 NF-¿B 和 IL-8 分泌的蛋白激活是 TLR4-、MD-2- 和 CD14 依赖性的，并且表达具有两个单核苷酸多态性（SNP）之一或两者的TLR4蛋白的转染子，以前与 LPS 反应性低下相关，对纯化的 RSV F 的反应明显较低蛋白质，在 TLR 表达相等的条件下。重要的是，我们发现了一个非常重要的来自高危婴儿病例系列的 DNA 样本中这些 TLR4 SNP 的比例过高有记录的 RSV 感染儿童。这些数据有力地支持了我们的总体假设： TLR4 在 RSV 的先天免疫反应中发挥着核心作用，这意味着 TLR4 的初始参与 TLR4 是保护性、适应性免疫反应（而非病理性免疫反应）发展所必需的。该提案详细介绍了创新的实验方法，这些方法将（i）导致开发安全且有效的 RSV F 蛋白亚单位疫苗，(ii) 导致基于的治疗干预策略的开发表征 F 蛋白与 TLR4/MD-2/CD14 复合物的相互作用，并检查 TLR4 信号传导在 RSV 感染/保护中的作用。预计在完成这笔赠款后，我们将确定可能导致 RSV 疫苗和新疗法开发的策略减轻宿主对 RSV 的病理反应。

项目成果

期刊论文数量（19）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Activation of interferon response through toll-like receptor 3 impacts viral pathogenesis and pulmonary toll-like receptor expression during respiratory syncytial virus and influenza infections in the cotton rat Sigmodon hispidus model.

DOI：
10.1089/jir.2009.0025
发表时间：
2010-04
期刊：
Journal of interferon & cytokine research : the official journal of the International Society for Interferon and Cytokine Research
影响因子：
0
作者：
Boukhvalova MS;Sotomayor TB;Point RC;Pletneva LM;Prince GA;Blanco JC
通讯作者：
Blanco JC

Role of the lipoxygenase pathway in RSV-induced alternatively activated macrophages leading to resolution of lung pathology.

DOI：
10.1038/mi.2013.71
发表时间：
2014-05
期刊：
Mucosal immunology
影响因子：
8
作者：
通讯作者：

Respiratory syncytial virus fusion protein-induced toll-like receptor 4 (TLR4) signaling is inhibited by the TLR4 antagonists Rhodobacter sphaeroides lipopolysaccharide and eritoran (E5564) and requires direct interaction with MD-2.

DOI：
10.1128/mbio.00218-12
发表时间：
2012
期刊：
mBio
影响因子：
6.4
作者：
Rallabhandi P;Phillips RL;Boukhvalova MS;Pletneva LM;Shirey KA;Gioannini TL;Weiss JP;Chow JC;Hawkins LD;Vogel SN;Blanco JC
通讯作者：
Blanco JC

New insights for development of a safe and protective RSV vaccine.