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Hendra病毒和Nipah病毒(HEV，NIV)的致死性和传播性使这些病原体令人严重关切。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。一种针对副粘病毒病毒融合蛋白(F)的疫苗可以对一组重要的人类病原体提供中和免疫。F的瞬时激活状态-只有在受体结合蛋白与受体结合时激活后才暴露，但在融合之前-是广泛中和抗体的最佳靶点。我们建议捕获和固定这种激活的融合中间体，暴露保守的F结构域，这是病毒进入必不可少的。然后，我们将使用这种捕获的活性状态分子-它通常高度保护免疫系统，不能在固定状态下正确呈现-来诱导广泛的中和抗体。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。人工原细胞将在仿生膜环境中呈现受体分子，以“触发”融合蛋白。我们实验室最近的三个进展使这一方法可行：(1)HEV/NIV在与受体部分相互作用时不可逆转地失活，这些受体部分导致F的构象变化，从而导致融合准备。(2)脂质包裹的硅胶原细胞可以在表面呈现受体分子，并通过过早地触发融合机制而不可逆转地灭活副粘病毒假型病毒。(3)与融合蛋白的七肽重复序列相对应的多肽仅在构象改变激活到融合就绪后但在融合进行之前才通过结合而抑制融合/进入。我们将使用以仿生方式呈现在原细胞表面的受体分子来激活F的构象变化，然后使用七肽重复序列来阻止F处于激活状态，从而将捕获的中间体与原细胞固定在一起。这个复合体随后将被用来诱导中和抗体。1.Nipah病毒融合(F)机制激活构象的捕获：携带受体加多肽的原细胞的发展，旨在将F固定在其过渡状态。捕获机制的验证。我们将验证这样的假设，即携带受体加肽的原始细胞可以有效地触发和固定F的中间构象状态。2.利用与F络合的原始细胞在其激活的构象中诱导免疫。免疫反应的评估。我们将测试这样一个假设，即暴露在仿生人工表面上的F的瞬时构象，可以诱导出强大的中和抗体，从而抑制尼帕病毒的感染。这些成果将为基于生物工程的疫苗接种建立一个新的平台。 与公共卫生的相关性：副粘病毒导致重要的人类疾病，这些疾病对全球疾病和死亡率有很大贡献。作为这项申请的主题的人畜共患副粘病毒，亨德拉病毒和尼帕病毒，由于其致命性和可传播性，是公共卫生的紧急关切。该项目的结果将导致建立一种新的基于生物工程的疫苗接种策略，将病毒学的新基本发现与新技术相结合，实现一种全新的方法，即诱导和捕获病毒融合蛋白的相关激活构象中间体。这种新的复合体将以一种通常只被免疫系统短暂看到的方式呈现关键的病毒抗原，密切模拟自然感染。鉴于副粘病毒对人类健康的重要性，以及针对这些和其他严重病原体的疫苗开发生物工程方法的潜在广泛适用性，这一结果将具有高度相关性。

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.