Harnessing ZBP1-driven cell death to improve influenza vaccine efficacy

项目摘要 接种疫苗是预防甲型流感病毒(IAV)感染的最有效手段。当前 IAV疫苗的方法主要针对HA蛋白，但HA经历了显著的抗原性漂移，并 因此，对于持久的体液免疫来说，这是一个糟糕的目标。开发通用IAV疫苗的另一种方法 是针对内部病毒蛋白(如核蛋白)，这些蛋白在大多数IAV之间也是高度保守的 菌株。然而，这种方法的一个主要障碍是缺乏触发正确类型的免疫的佐剂。 对这些内部抗原的反应--I型反应(或Th1/CD8+T细胞驱动的)反应。在这份提案中，我们概述了 寻求利用免疫原性细胞死亡作为激活对IAV的I型免疫的一种新策略 疫苗。我们已经确定了一条宿主信号通路，它几乎解释了所有IAV激活的免疫原性 感染细胞死亡，包括原代呼吸道上皮细胞、巨噬细胞和树突状细胞。这条细胞死亡途径 当宿主传感器蛋白ZBP1检测到IAV基因组RNA并激活平行的 程序性坏死(坏死性下垂)和凋亡，以杀死感染的细胞并激发CD8+T细胞介导的 适应性免疫反应。除了ZBP1引发的细胞死亡作为疫苗方法的吸引力之外，还有 我们的发现，首先，IAV产生一种新形式的双链RNA-称为Z-RNA-激活ZBP1。 因此，Z-RNA是一种新的PAMP，可以很容易地被利用来触发免疫原性细胞死亡和佐剂IAV 疫苗。其次，ZBP1从细胞核引发坏死性下垂，导致核膜破裂和释放 具有高度免疫原性的核抑制物和IAV抗原。核性坏死性下垂的免疫原性比 传统的(细胞质引发的)坏死性下垂。AS ZBP1诱导的免疫原性细胞死亡(细胞凋亡和 核坏死性下垂)对于有效的抗IAV细胞毒性T细胞反应很重要，我们的发现使我们能够 提示Z-RNAs是一种有效的新型佐剂，可激活针对IAV的I型免疫应答。 制造Z-RNA的重组IAV代表了新的I型免疫激活疫苗。在这 提议，三个在IAV细胞死亡信号方面拥有丰富专业知识的实验室(Balachandran)、重组IAV 技术(朗格洛斯)和致病/疫苗策略(洛佩兹)将联合起来识别IAV产生的Z-RNA 激活ZBP1(目标1)；确定哪些基质和免疫细胞类型通过ZBP1介导的机制死亡 启动和调节有效的适应性免疫反应(目标2)；并测试是否触发ZBP1介导的细胞 死亡与合成和病毒产生的Z-RNA可以被利用来通知和供应下一代 能够对IAV提供有效交叉保护免疫的疫苗(目标3)。

Project Summary Vaccination represents the most effective means of preventing influenza A virus (IAV) infections. Current approaches to IAV vaccines primarily target the HA protein, but HA undergoes significant antigenic drift and is thus a poor target for lasting humoral immunity. An alternative approach to developing a universal IAV vaccine is to target internal viral proteins (such as the nucleoprotein), which are also highly conserved between most IAV strains. A major roadblock to this approach, however, is a lack of adjuvants that trigger the right kind of immune response - a ‘type I’ (or Th1/CD8+ T-cell driven) response - to these internal antigens. In this proposal, we outline a new strategy that seeks to harness immunogenic cell death as a means of activating type I immunity to IAV vaccines. We have identified a host signaling pathway which accounts for almost all IAV-activated immunogenic death in infected cells, including primary airway epithelial cells, macrophages and DCs. This cell death pathway is initiated when the host sensor protein ZBP1 detects IAV genomic RNA and activates parallel pathways of programmed necrosis (necroptosis) and apoptosis to kill the infected cell and instigate CD8+ T cell-mediated adaptive immune responses. Adding to the attractiveness of ZBP1-initiated cell death as a vaccine approach are our discoveries, first, that IAV produces a new form of double-stranded RNA - termed Z-RNA - to activate ZBP1. Z-RNA is thus a new PAMP, and can be readily harnessed to trigger immunogenic cell death and adjuvant IAV vaccines. Second, ZBP1 initiates necroptosis from the nucleus, resulting in nuclear envelope rupture and release of highly-immunogenic nuclear DAMPs and IAV antigens. Nuclear necroptosis is even more immunogenic than conventional (cytoplasm-initiated) necroptosis. As ZBP1-driven immunogenic cell death (both apoptosis and nuclear necroptosis) is important for effective anti-IAV cytotoxic T cell responses, our discoveries allow us to propose that Z-RNAs are potent new adjuvants for activating type I immune responses to IAV, and that recombinant IAVs engineered to produce Z-RNAs represent new type I immunity-activating vaccines. In this proposal, three labs with strong expertise in IAV cell death signaling (Balachandran), recombinant IAV technology (Langlois), and pathogenesis/vaccine strategies (Lopez) will unite to identify IAV-generated Z-RNAs that activate ZBP1 (Aim 1); determine which stromal and immune cell types die by ZBP1-mediated mechanisms to initiate and regulate effective adaptive immune responses (Aim 2); and test if triggering ZBP1-mediated cell death with synthetic and virus-generated Z-RNAs can be leveraged to inform and supply next-generation vaccines capable of providing effective cross-protective immunity to IAV (Aim 3).