Bioengineering protocells for a new class of viral vaccines

描述（由申请人提供）：应对该领域挑战的新战略。副粘病毒亨德拉病毒和尼帕病毒（HeV, NiV）的致死性和传染性使这些病原体受到严重关注。这些也是在生物工程和病毒学之间开发创新的新疫苗平台的理想模式。一种以副粘病毒融合蛋白(F)为靶点的疫苗可以对一组重要的人类病原体提供中和性免疫。F的瞬时激活状态-仅在受体结合蛋白与受体结合后激活，但在融合之前暴露-是广泛中和抗体的最佳靶点。我们建议捕获和固定这种活化的融合中间体，暴露对病毒进入至关重要的保守F结构域。然后，我们将使用这种捕获的活性状态分子（通常对免疫系统高度保护，不能在固定状态下适当呈现）来引发广泛中和的抗体。虽然这种激活的构象通常只存在于活细胞的表面，但我们建议使用工程原细胞来捕获和固定这种过渡状态。人造原细胞将在仿生膜设置中呈现受体分子以“触发”融合蛋白。我们实验室最近的三个进展使这种方法可行：(1)当HeV/NiV与受体相互作用时，它们会不可逆地失活，从而诱导F的构象变化，从而导致融合准备。(2)脂质包被的硅基原细胞可以在表面呈递受体分子，通过过早触发融合机制，不可逆地灭活副粘病毒假型病毒。(3)与融合蛋白七肽重复序列相对应的肽仅在激活融合准备构象变化后，而在融合进展之前通过结合来抑制融合/进入。我们将使用受体分子，以仿生的方式呈现在原始细胞表面，激活F的构象变化，然后使用七重重复肽来阻止处于激活状态的F，从而将捕获的中间体固定在原始细胞上。这种复合物将被用来诱导中和抗体。1. 捕获尼帕病毒融合(F)机制的激活构象：携带受体加肽的原细胞的开发，旨在将F固定在其过渡状态。捕获机制的验证。我们将验证携带受体加肽的原细胞可以有效地触发和固定F. 2的中间构象状态的假设。使用与活化构象F复合物的原始细胞来引起免疫。免疫反应评估。我们将检验这一假设，即在仿生人工表面上呈现和捕获的暴露的F的短暂构象，引发抑制尼帕病毒感染的强大中和抗体。这些结果将为基于生物工程的疫苗接种建立一个新的平台。

DESCRIPTION (provided by applicant): A new strategy that addresses challenges in the field. The lethal and transmissible nature of the paramyxoviruses Hendra virus and Nipah virus (HeV, NiV) makes these pathogens of serious concern. These are also ideal models for developing an innovative new vaccine platform at the interface between bioengineering and virology. A vaccine that targets the paramyxovirus viral fusion protein (F) could provide neutralizing immunity against an important group of human pathogens. The transient activated state of F - exposed only after activation when the receptor binding protein binds receptor, but before fusion - is an optimal target for broadly neutralizing antibodies. We propose to capture and immobilize this activated fusion intermediate, exposing the conserved F domains that are essential for viral entry. We will then use this captured active-state molecule - which is normally highly sheltered from the immune system and could not be properly presented in an immobilized state - to elicit broadly neutralizing antibodies. While this activated conformation is normally present only at the surface of live cells, we propose to use engineered protocells to capture and immobilize this transitional state. Artificial protocells will present the receptor molecules in a biomimetic membrane setting to "trigger" the fusion protein. Three recent advances in our lab make this approach feasible: (1) HeV/NiV are irreversibly inactivated when they interact with receptor moieties that induce the conformational changes in F that lead to fusion-readiness. (2) Lipid-coated silica-based protocells can present receptor molecules on the surface, and irreversibly inactivate paramyxovirus pseudotyped viruses by prematurely triggering the fusion mechanism. (3) Peptides corresponding to the heptad repeats of the fusion protein inhibit fusion/entry by binding only after activation of the conformational change to fusion-readiness, but before progression of fusion. We will use receptor molecules, presented in a biomimetic fashion on the surface of protocells, to activate the conformational change in F, and then use heptad-repeat peptides to arrest F in its activated state and thus immobilize the captured intermediate with the protocell. This complex will then be used to induce neutralizing antibodies. 1. Capture of the activated conformation of Nipah virus fusion (F) machinery: Development of protocells bearing receptor plus peptide, designed to immobilize F in its transition state. Validation of capture mechanism. We will test the hypothesis that protocells bearing receptor plus peptide can effectively trigger and immobilize the intermediate conformational state of F. 2. Use of protocells complexed with F in its activated conformation to elicit immunity. Assessment of immune response. We will test the hypothesis that the exposed transient conformation of F, presented and captured on a biomimetic artificial surface, elicits powerful neutralizing antibodies that inhibit Nipah virus infection. These results will establish a new platform for bioengineering-based vaccination. PUBLIC HEALTH RELEVANCE: Paramyxoviruses cause important human illnesses that contribute significantly to global disease and mortality. The zoonotic paramyxoviruses that are the subject of this application, Hendra virus and Nipah virus, are an urgent concern for public health due to their lethal and transmissible nature. The results of this project will lead to setting up a new bioengineering-based strategy for vaccination that combines new basic findings from virology with new technology to achieve an entirely new approach that of inducing and capturing the relevant activated conformational intermediate of a viral fusion protein. The novel complex will present the key viral antigen in a way normally only seen transiently by the immune system, closely mimicking natural infection. The results will be highly relevant in light of the importance of paramyxoviruses to human health and the potential broad applicability of the bioengineering approach to vaccine development for these and other serious pathogens.