Neutralization of Eastern equine encephalitis virus by human monoclonal antibodies

人单克隆抗体中和东方马脑炎病毒

• 批准号: 9757522
• 负责人: Lauren Williamson
• 金额: $ 2.97万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9757522
• 关键词: Address; Alanine; Alphavirus; American; Antibodies; Antibody Repertoire; Antibody Response; Antibody Specificity; Antigens; Antiviral Agents; Binding; Biological Assay; Bioterrorism; Capsid Proteins; Category B pathogen; Cells; Centers for Disease Control and Prevention (U.S.); Complex; Culicidae; Detection; Deterioration; Disease Outbreaks; Eastern Equine Encephalitis Virus; Electron Microscopy; Epitope Mapping; Epitopes; Equus caballus; Fellowship; Glycoproteins; Goals; Heterodimerization; Human; Human papillomavirus 16 E1 protein; Hybridomas; Immune; Immune response; Immune system; Immunity; Immunology; In Vitro; Infection; Knowledge; Laboratories; Libraries; Membrane; Molecular Conformation; Molecular Structure; Monoclonal Antibodies; Mus; Mutagenesis; Neurologic Signs; Neutralization Tests; Plaque Assay; Play; Prevention; Principal Investigator; Quantitative Reverse Transcriptase PCR; Recombinants; Recovery; Reporting; Research; Resolution; Role; Scanning; Scientist; Structural Protein; Structure; Surface; Survivors; Techniques; Technology; Testing; Therapeutic; Training; Transmembrane Domain; United States National Institutes of Health; Vaccines; Viral; Virion; Virulent; Virus; Virus Diseases; Virus Replication; X-Ray Crystallography; career; clinically relevant; clinically significant; crosslink; design; experience; feeding; human monoclonal antibodies; human mortality; insight; mortality; murine antibody; neutralizing antibody; neutralizing monoclonal antibodies; particle; prevent; programs; protein structure; prototype; receptor; stoichiometry; structural biology; virology

PROJECT SUMMARY/ABSTRACT The overall goal of this project addresses the molecular and structural basis of neutralization utilized by human monoclonal antibodies (mAbs) against Eastern equine encephalitis virus (EEEV). The North American lineage of EEEV (NA-EEEV) is the most virulent of the encephalitic alphaviruses with up to 70% human and 90% equine mortality rate. In addition, of those that survive infection, up to 80% show signs of neurological deterioration. On average, eight human cases a year in the US are reported. However, the recent rise in detection of NA-EEEV in human-feeding mosquito species raises concern for a large outbreak in the eastern US. Moreover, NA-EEEV is considered a NIH Category B priority pathogen and USDA/CDC Select Agent due to its potential threat as a bioterrorism agent. There are no approved antiviral drugs or licensed human vaccines available for NA-EEEV. The antibody response to alphaviruses has been shown to be an important part of the immune response in conferring protective immunity and aiding in the clearance and recovery from infection. However, the fundamental molecular and structural mechanisms of action of antibodies in humans to NA-EEEV remain poorly defined. To fill this gap in knowledge, the objective of this study is to determine the mechanisms by which human mAbs neutralize NA-EEEV. The overarching hypothesis of this study is that the principle mode of action of human neutralizing mAbs is to stabilize the virus particle and inhibit viral fusion to host cells. To test this hypothesis, I will isolate human mAbs from naturally infected survivors of NA-EEEV to the BSL-2 chimeric virus, Sindbis/EEEV (SINV/NA-EEEV) and recombinant NA-EEEV structural proteins. I will characterize the panel of human mAbs isolated for specific reactivity and neutralization potency against SINV/NA-EEEV. Of the human mAbs that neutralize NA-EEEV, I then will determine the step(s) in the replication cycle the mAbs neutralize the virus through in vitro mechanistic assays. To identify neutralizing antigenic determinants, I will use epitope mapping, alanine scanning mutagenesis, and structural biology techniques. Determination of the neutralizing antigenic determinants recognized by human NA-EEEV mAbs will inform how mAbs interact with NA-EEEV. The results obtained from this project will fill major gaps in knowledge about the human antibody response to NA-EEEV, will identify potential correlates of protection, and may facilitate the design of efficient therapeutics and vaccines against this clinically relevant alphavirus. The proposal outlines a comprehensive fellowship training plan that will prepare me for a high-impact career in the field of viral immunology. To accomplish this program and research, we have assembled a collaborative team consisting of principal investigators and staff scientists with exceptional expertise in the immunology, virology, and structural biology fields to provide support and guidance for this study.

项目概要/摘要 该项目的总体目标是解决人类利用的中和的分子和结构基础 针对东部马脑炎病毒 (EEEV) 的单克隆抗体 (mAb)。北美血统 EEEV (NA-EEEV) 是脑炎甲病毒中毒性最强的，其中高达 70% 为人类脑炎病毒，90% 为脑炎甲病毒。 马死亡率。此外，在感染后存活下来的人中，高达 80% 的人表现出神经系统症状 恶化。美国平均每年报告八起人类病例。然而，近期的上涨 在喂食人类的蚊子中检测到 NA-EEEV 引起人们对东部地区大规模疫情爆发的担忧 我们。此外，NA-EEEV 被认为是 NIH B 类优先病原体和 USDA/CDC 选择剂，因为 其作为生物恐怖分子的潜在威胁。没有批准的抗病毒药物或许可的人类药物 可用于 NA-EEEV 的疫苗。对甲病毒的抗体反应已被证明是一个重要的 免疫反应的一部分，赋予保护性免疫力并帮助清除和恢复 感染。然而，抗体在人体中作用的基本分子和结构机制 NA-EEEV 的定义仍然不明确。为了填补这一知识空白，本研究的目的是确定 人类单克隆抗体中和 NA-EEEV 的机制。这项研究的总体假设是 人中和单克隆抗体的主要作用方式是稳定病毒颗粒并抑制病毒融合 宿主细胞。为了检验这个假设，我将从自然感染的 NA-EEEV 幸存者中分离出人类单克隆抗体，以 BSL-2 嵌合病毒、Sindbis/EEEV (SINV/NA-EEEV) 和重组 NA-EEEV 结构蛋白。我会 表征分离的人类单克隆抗体组的特定反应性和中和效力 SINV/NA-EEEV。对于中和 NA-EEEV 的人类单克隆抗体，我将确定以下步骤： 在复制周期中，单克隆抗体通过体外机制测定来中和病毒。识别中和 抗原决定簇，我将使用表位作图、丙氨酸扫描诱变和结构生物学 技术。人 NA-EEEV mAb 识别的中和抗原决定簇的测定 将告知 mAb 如何与 NA-EEEV 相互作用。该项目取得的成果将填补国内重大空白 关于人类抗体对 NA-EEEV 反应的知识，将确定保护的潜在相关性，以及 可能有助于设计针对这种临床相关甲病毒的有效疗法和疫苗。这 提案概述了一项全面的奖学金培训计划，该计划将使我为在该领域的高影响力职业生涯做好准备 病毒免疫学领域。为了完成这个项目和研究，我们组建了一个协作团队 由在免疫学、病毒学、 和结构生物学领域为本研究提供支持和指导。