Elicitation of antibodies broadly neutralizing the porcine reproductive and respiratory syndrome virus (PRRSV) using reverse vaccinology

使用反向疫苗学诱导广泛中和猪繁殖与呼吸综合征病毒 (PRRSV) 的抗体

• 批准号: 511670106
• 负责人: Professor Dr. Thomas Krey
• 金额: --
• 依托单位: Institut für Virologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/511670106?language=en
• 关键词: Elicitation; antibodies; broadly; neutralizing; porcine

Porcine reproductive and respiratory syndrome virus (PRRSV) is an economically important pathogen of swine. Unfortunately, the available live attenuated and inactivated vaccines provide only limited protection against symptomatic disease and cannot effectively prevent the spread of virulent field viruses in swine herds. Although a strong humoral immune response against homologous viral strains is elicited after vaccination, the achieved protection against genetically divergent viral strains is insufficient. Traditional vaccine approaches apparently cannot cover the large genetic variability of PRRSV comprising two separate species (PRRSV-1 and -2). The structure of PRRSV glycoproteins remains elusive to date, complicating the use of structure-based vaccine design to improve vaccine efficacy. In this project, we will apply reverse vaccinology approaches to fill these gaps. Using multi-color FACS with fluorescently labeled PRRSV glycoproteins and virions followed by state-of-the-art single B cell sequencing we will analyze the PRRSV-specific porcine antibody repertoire. Individual antibodies will be expressed recombinantly and the recombinant antibodies characterized with respect to their neutralization profile and potency as well as their cognate antigen. In parallel, we will build on our successful pipeline to identify PRRSV neutralization epitopes to focus on highly conserved epitopes targeted by broadly neutralizing antibodies that are of particular interest for vaccine design. For this purpose, we will immunize mice sequentially with PRRSV-1 and PRRSV-2 virions followed by isolation of memory B cells reacting with glycoproteins of both species and characterization of bnAbs as described above. We will also investigate interesting neutralizing antibodies generated in pigs or mice in this study with respect to their neutralization mechanism and whether the virus can escape these antibodies by mutation in a so-called "immune escape". Finally, we will structurally characterize the neutralizing antibody mAb18, which we have already identified in a proof-of-concept study to recognize PRRSV gp2, in complex with an epitope peptide and use the resulting structure to develop an epitope-focused vaccine. To this end, we will transfer this epitope to suitable "scaffold" proteins, produce a scaffold-based vaccine candidate in the form of nanoparticles and validate this candidate in a challenge trial in pigs. Taken together, this project will provide important insights into the immune response to PRRSV glycoproteins to facilitate informed vaccine design.

猪繁殖与呼吸综合征病毒（Porcine reproductive and respiratory syndrome virus，PRRSV）是一种重要的经济病原。不幸的是，现有的减毒活疫苗和灭活疫苗仅提供有限的针对症状性疾病的保护，并且不能有效地防止猪群中强毒田间病毒的传播。尽管疫苗接种后引发了针对同源病毒株的强烈体液免疫应答，但针对遗传上不同的病毒株所实现的保护是不够的。传统的疫苗方法显然无法覆盖由两个不同物种（PRRSV-1和-2）组成的PRRSV的巨大遗传变异性。迄今为止，PRRSV糖蛋白的结构仍然难以捉摸，使得使用基于结构的疫苗设计来提高疫苗效力变得复杂。在这个项目中，我们将应用反向疫苗学方法来填补这些空白。我们将使用荧光标记的PRRSV糖蛋白和病毒体的多色FACS，然后使用最先进的单B细胞测序，分析PRRSV特异性猪抗体库。将重组表达单个抗体，并就其中和谱和效价以及其同源抗原对重组抗体进行表征。与此同时，我们将建立在我们成功的管道，以确定PRRSV中和表位，重点是高度保守的表位靶向广泛中和抗体是特别感兴趣的疫苗设计。为此目的，我们将用PRRSV-1和PRRSV-2病毒体依次免疫小鼠，然后分离与两个物种的糖蛋白反应的记忆B细胞，并如上所述表征bnAb。我们还将研究在本研究中在猪或小鼠中产生的有趣的中和抗体的中和机制，以及病毒是否可以通过所谓的“免疫逃逸”中的突变来逃避这些抗体。最后，我们将在结构上表征中和抗体mAb 18，我们已经在概念验证研究中鉴定了该抗体，以识别与表位肽复合的PRRSV gp 2，并使用所得结构开发表位聚焦疫苗。为此，我们将把这种表位转移到合适的“支架”蛋白上，以纳米颗粒的形式生产基于支架的候选疫苗，并在猪的挑战试验中验证这种候选疫苗。总之，该项目将为PRRSV糖蛋白的免疫反应提供重要的见解，以促进明智的疫苗设计。