Structure based design of dengue subunit vaccines for inducing protective but not disease enhancing antibodies

摘要 由于存在4种血清型，登革病毒(DENV)疫苗的开发一直具有挑战性 (DENV1-4)和疫苗的可能性增强了严重疾病。领先的减毒活四价DENV 疫苗一直受到疫苗成分复制不平衡导致效果不稳定的困扰 疫苗在一些儿童中引发了严重的登革热。这项提议的目标是开发小说 重组DENV包膜(E)蛋白和病毒样颗粒(VLP)疫苗克服面临的障碍 减毒四价活疫苗。我们发现DENV E蛋白是以一种分泌的方式产生的 蛋白质是一种很差的疫苗，因为它是一种单体，不显示主要的四元表位 人中和保护性抗体(Abs)。这项建议是基于我们小组的新发现。 关于如何使用基于结构的计算方法来生产高度稳定和适当折叠的产品 由哺乳动物细胞高效分泌的DENV E二聚体。我们将使用DENV的小鼠模型 疫苗接种和感染以测试人工稳定的DENV E二聚体是否刺激类似于 暴露于原发和继发野生环境的人群的血清型特异性和交叉保护性抗体反应 输入DENV感染。稳定的E二聚体将被进一步修饰，以测试E是否大规模重新表面 二聚体可用于将免疫反应集中在强大的中和抗体识别的表位上，而 消除对偏离目标的疾病增强表位的反应。最后，我们将设计锚定的膜 促进登革病毒样颗粒(VLP)候选疫苗形成的稳定E二聚体的变体 这更像是成熟的感染性病毒粒子，而不是用当前技术制造的VLP。我们的研究，这是 探索如何设计和传递重组E抗原，使宿主抗体反应集中在重要的 四级结构中和表位，将在黄病毒亚单位领域激发新的研究方向 疫苗。

ABSTRACT Dengue virus (DENV) vaccine development has been challenging because of the presence of 4 serotypes (DENV1-4) and the potential for vaccine enhanced severe disease. The leading live attenuated tetravalent DENV vaccines have been plagued by poorly balanced replication of vaccine components leading to variable efficacy and vaccine primed severe dengue disease in some children. The goal of this proposal is to develop novel recombinant DENV envelope (E) protein and virus like particle (VLP) vaccines that overcome barriers faced by live attenuated tetravalent vaccines. We have discovered that the DENV E protein produced as a secreted protein is a poor vaccine because it is a monomer that does not display major quaternary epitopes targeted by human neutralizing and protective antibodies (Abs). This proposal is based on new discoveries from our group about how structure based, computational approaches can be used to produce highly stable and properly folded DENV E dimers that are efficiently secreted from mammalian cells. We will use mouse models of DENV vaccination and infection to test if artificially stabilized DENV E dimers stimulate Ab responses that are similar to serotype-specific and serotype-cross-protective Ab responses in people exposed to primary and secondary wild type DENV infections. The stabilized E dimers will be further modified to test if large-scale resurfacing of the E dimer can be used to focus the immune response on epitopes recognized by potent neutralizing Abs, while eliminating responses to off target, disease enhancing epitopes. Finally, we will design membrane anchored variants of stabilized E dimers to promote the formation of dengue virus-like particle (VLP) vaccine candidates that better resemble mature infectious virions than VLPs made with current techniques. Our studies, which explore how to design and deliver recombinant E antigens to focus the host antibody response on important quaternary structure neutralizing epitopes, will stimulate new research directions in the field of flavivirus subunit vaccines.