Development of a Flavivirus E Protein Fusion Loop Vaccine

描述(申请人提供)：该提案的目标是为基于合成肽的多黄病毒疫苗的全面开发工作奠定基础。在我们以前研究黄病毒的工作中，我们团队使用噬菌体展示人库屏幕制备了针对西尼罗河病毒(WNV)包膜(E)蛋白的重组单抗。出乎意料的是，抗体识别了融合环，这是一种黄病毒E蛋白表位，通过膜融合介导病毒进入，在感染WN病毒或E蛋白免疫后通常不会引起抗体反应。该单抗对小鼠的WN病毒感染具有保护作用，并能清除正在进行的WN感染。值得注意的是，mAb还与其他几种黄病毒，如登革热病毒反应，这表明它可能具有广泛的交叉保护作用(1)。在这里，我们建议以单抗识别的表位为靶点的重组融合环肽疫苗可以预防和治疗广泛的黄病毒感染，以WN病毒作为我们的初始模型感染。不幸的是，代表这个表位的多肽在小鼠身上没有很强的免疫原性。因此，我们建议合成一组改进的融合环肽，对其进行修饰以提高天然多肽的免疫原性。包括多种抗原肽修饰在内的一组多肽将用于使用已建立的佐剂和递送技术来测试该方法。然后，我们将挑战小鼠，并评估体液和细胞反应。然后将评估引起反应的候选配方对西尼罗河病毒的中和能力。这项建议的主要目标是评估这些候选制剂对广泛的黄病毒，如WN病毒、昆津病毒、日本脑炎病毒、墨累谷脑炎、圣路易斯脑炎病毒(SLEV)、登革病毒(血清1至4)和黄热病毒的抗药性。最后，任何确实显示出针对西尼罗河病毒和更广泛的黄病毒集的特定活性的配方都将在体内与我们建立的神经侵袭性西尼罗河病毒的C3H/HEN小鼠模型进行测试。这种基础广泛的多肽疫苗战略的试点方法可能对多种黄病毒病原体产生深远影响，包括目前尚无疫苗可用的黄病毒。 公共卫生相关性：黄病毒属中的人类病原体在世界范围内导致显著的发病率和死亡率。该项目旨在开发一种新的候选疫苗，可能会保护人类免受各种黄病毒感染。

DESCRIPTION (provided by applicant): The goal of this proposal is to lay the foundation for a full development effort of a synthetic peptide-based multi-flavivirus vaccine. In our previous work on flaviviruses, our group produced a recombinant monoclonal antibody against West Nile virus (WNV) envelope (E) protein using a phage display human library screen. Unexpectedly, the antibody recognized the fusion loop, a flavivirus E protein epitope that mediates viral entry by membrane fusion and that does not normally elicit an antibody response following infection with WN virus or immunization with E protein. The mAb was protective against WN virus infection in mice and could also clear an ongoing WN infection. Remarkably, the mAb also reacted with several other flaviviruses, such as dengue virus, suggesting it may be broadly cross-protective (1). Here we propose that a recombinant fusion loop peptide vaccine that targets the epitope recognized by the mAb can prevent and treat a broad spectrum of flavivirus infections, with WN virus serving as our initial model infection. Unfortunately, Peptides representing this epitope are not highly immunogenic in mice. We therefore propose to synthesize a set of improved fusion loop peptides, modified to increase the immunogenicity of the native peptides. A set of peptides including, multiple antigenic peptide modifications will be used to test this approach using established adjuvant and delivery techniques. We will then challenge mice and evaluate humoral and cellular responses. Candidate formulations that elicit a response will then be evaluated for their neutralizing capacity of WNV. The main goal of this proposal is then to evaluate these candidate formulations against a broad array of Flaviviruses such as WN virus, Kunjin, Japanese encephalitis virus, Murray valley encephalitis, St. Louis encephalitis virus (SLEV), Dengue virus (serotypes 1 to 4), and Yellow fever virus. Finally, any formulations that do show both specific activity against WNV and against a broader set of flaviviruses will be tested in vivo against our well established C3H/HeN mouse Model of neuroinvasive WNV. This pilot approach to a broad based peptide vaccine strategy could have profound impact against multiple flavivirus pathogens including flaviviruses for which no vaccines are currently available. PUBLIC HEALTH RELEVANCE: Human pathogens in the Flavivirus genus cause significant morbidity and mortality worldwide. This project is designed to develop a new vaccine candidate that might protect humans against a variety of Flavivirus infections.