Co-Presentation and Delivery of TLR Agonist Combinations with Subunit Antigens to Pathogen-Match the Immune Response

项目摘要/摘要 亚单位疫苗是预防传染病和相关死亡的关键战略。然而，使用 与完整病原体相比，亚单位抗原引起的免疫反应较弱，这与对 安全有效的佐剂，提高疫苗效力，增强免疫应答，缩小疫苗体积和 提供免疫所需的疫苗数量。然而，我们缺乏一张详细的图片，说明如何吸引多个 同一细胞或不同细胞上的先天免疫受体同时或顺序地改变结果 对免疫反应的影响。我们的研究团队已经设计出一种新的、灵活的、模仿病原体的 一种可共同呈递功能抗原的重组外膜泡囊平台 使用所需的佐剂组合。我们打算利用这项创新技术来研究如何 TLR和NOD激动剂的不同佐剂组合可激活先天性免疫系统，并帮助引导 对病毒和细菌病原体的免疫反应。我们的假设，将通过 特定的目的是rOMvs可以将抗原与特定的佐剂组合共同呈现以诱导 病原体匹配和保护性免疫反应(例如，抗病毒和抗细菌)。具体目标 目的：目标1：确定rOMV上单个佐剂或佐剂组合所引发的分子通路。 ROMV将被TLR和NOD激动剂装饰，以了解产生的先天免疫受体如何 激动剂利用已建立的报告细胞激活和影响细胞信号通路，并决定这些信号通路如何 TLR和NOD激动剂的共同呈递改变了通路。我们会将反应与可溶性进行比较 相同佐剂的组合对信号和树突状细胞的激活和成熟都有影响。目标2： 确定rOMV-佐剂组合如何引导适应性免疫反应到使用OVA的免疫 作为模型抗原。我们将使用OVA抗原来表征rOMV诱导的获得性免疫反应 表达单一或组合的佐剂，并将它们与单一或组合的可溶性佐剂进行比较。 我们将确定能促进强烈的指向性T细胞、B细胞和/或记忆反应的组合(S)。 目的3：确定量身定制的rOMV佐剂组合是否能指导特异性抗病毒和抗细菌 保护豁免权。我们将使用呈现OVA的rOMV，以及 ROMV，在两种病毒(流感)的挑战模型中装饰不同的单一或联合佐剂 以及一种细菌病原体(李斯特氏菌)，经修饰后可表达OVA以及本地病原体(即不表达OVA)。我们 将研究天然信号通路激活和免疫反应如何与保护性免疫相关。 拟议研究的预期结果是对佐剂组合如何进行机械性理解 在分子、细胞和有机体水平上工作，以防止病毒和细菌病原体。结果是 将有助于定义一套指导原则，我们和该领域的其他调查人员可以使用这些指导原则来更有力地 设计佐剂系统以最大限度地发挥亚单位疫苗的效力。

Project Summary/Abstract Subunit vaccines are a key strategy for preventing infectious disease and related deaths.. However, the use of subunit antigens, which elicit a weaker immune response than intact pathogen, has coincided with a need for safe and effective adjuvants to improve vaccine efficacy, increase immune response, and reduce the size and quantity of vaccine necessary to impart immunity. However, we lack a detailed picture of how engaging multiple innate immune receptors simultaneously or sequentially, on the same cell, or different cells, alters the outcome of the immune response. Our research team has engineered a new, flexible, pathogen-mimicking recombinant outer membrane vesicle (rOMV) platform capable of presenting functional antigen together with desired combinations of adjuvants. We intend to use this innovative technology to investigate how different adjuvant combinations of TLR and NOD agonists engage the innate immune system and help direct the immune response against viral and bacterial pathogens. Our hypothesis, to be evaluated through the Specific Aims, is that rOMVs can co-present antigen with specific combinations of adjuvants to induce pathogen-matched and protective immune responses (e.g., anti-viral and anti-bacterial). The specific aims are to: Aim 1: Identify the molecular pathways elicited by individual or combinations of adjuvants on rOMVs. rOMVs will be decorated with TLR and NOD agonists to understand how the resulting innate immune receptor agonists activate and influence cell signaling pathways using established reporter cells, and determine how those pathways are altered by co-presentation of TLR and NOD agonists. We will compare the response to soluble combinations of the same adjuvants on both signaling and dendritic cell activation and maturation. Aim 2: Determine how rOMV-adjuvant combinations direct the adaptive immune response to immunization using OVA as a model antigen. We will use OVA antigen to characterize the adaptive immune response induced by rOMV expressing single or combinations of adjuvants and compare them to single or combinations of soluble adjuvants. We will identify the combination(s) that promote strong and directive T cell, B cell and/or memory responses. Aim 3: Determine whether tailored rOMV adjuvant combinations can direct specific anti-viral and anti-bacterial protective immunity. We will use OVA-presenting rOMVs, as well as pathogen-specific antigens presented on rOMVs, decorated with different single or combined adjuvants in challenge models for both a viral (influenza) and a bacterial pathogen (Listeria) modified to express OVA as well as the native pathogens (i.e., no OVA). We will examine how innate signaling pathway activation and immune response correlate to protective immunity. The anticipated outcome of the proposed studies is a mechanistic understanding of how adjuvant combinations work at the molecular, cellular and organismal level to protect against viral and bacterial pathogens. The outcome will help to define a set of guiding principles that we and other investigators in the field can use to more robustly design adjuvant systems to maximize the efficacy of subunit vaccines.