Human Monoclonal Antibodies for Encephalitic Alphaviruses

摘要： 东部马脑炎病毒(EEEV)是一种重新出现的蚊媒甲型病毒，可导致衰弱 人类的脑炎疾病。大约三分之一的人类EEEV感染病例死亡，许多幸存者 长期的、令人衰弱的神经问题。病毒维持在库里塞塔之间的地方性循环中 黑斑蚊和禽类宿主，但可通过 一些伊蚊、Coquillettidia和库蚊物种。这种感染在人类中并不常见，但频率正在上升。 近年来，可能次要受到气候变化、病媒扩张等不明因素的影响。EEEV 也被认为是潜在的生物恐怖主义威胁，因为它通过气雾剂路线传播。尽管高致病性 由于病毒的性质，目前还没有针对EEEV的特定治疗方法或疫苗。这个项目的一个主要目标是 确定超强中和人的分子、遗传、免疫学和结构特征 具有广泛体内抗EEEV活性的单抗。其他目标包括定义 通过这些超强中和单抗提供保护，并确定优化功能和传递到 大脑。在这些研究中，我们将阐明具有特殊抑制活性的抗病毒抗体是如何在 细胞培养和体内实验。该方法将包括高效分离人mAbs，再加上 基于下一代测序的创新抗体基因谱系研究。我们将检验几个假设， 包括超强中和活性的概念是由两种抗体的特性(选择 最佳V-D-J克隆型和关键体细胞突变的积累)和抗原(与 多个相邻包膜蛋白上的四元表位与病毒关键结构转变的阻断 进入或释放)。我们还将应用新技术，通过受体介导的分子跨 利用基因工程技术改变Fc区的血脑屏障。尽管我们的重点是了解 超强抗人单抗是如何以及为什么抑制EEEV的，这些研究可能与以下一般原理有关 多种不同脑炎病毒的抗体中和。除了定义分子和结构 基于抗体中和EEEV和部署将生物制剂输送到大脑的新策略，这些 研究将产生一组完全人类的单抗，可以预防和治疗EEEV感染，这可能是 在不久的将来开发出来，可能是一种治疗人类的药物。本项目中的研究，虽然针对的是 EEEV，可能会为未来基于抗体和/或疫苗的努力提供信息，以对抗导致人类大脑的其他虫媒病毒 感染。我们已经组建了一个独特的研究小组，其中包括一名人类抗体专家，一名分子 在抗体-病毒相互作用方面有经验的病毒学家，动物模型和发病机制专家 包括EEEV在内的脑型甲型病毒方面的专业知识，以及进行这些研究的脑靶向科学家。

SUMMARY: Eastern equine encephalitis virus (EEEV) is a re-emerging mosquito-borne alphavirus that causes a debilitating encephalitic illness in humans. About a third of human cases of EEEV infection die and many survivors have long-term, debilitating neurologic problems. The virus is maintained in an enzootic cycle between Culiseta melanura mosquitoes and avian hosts but can be transmitted to humans and horses by some Aedes, Coquillettidia, and Culex species. The infection is unusual in humans but increasing in frequency in recent years, likely secondary to climate changes, vector expansion, and other uncharacterized factors. EEEV also is regarded as a potential bioterrorism threat due to spread via aerosol route. Despite the highly pathogenic nature of the virus, no specific treatment or vaccine for EEEV is available. A primary goal of this project is to define the molecular, genetic, immunologic, and structural characteristics of ultra-potent neutralizing human mAbs with broad activity in vivo against EEEV. Additional goals include defining the mechanistic correlates of protection by these ultra-potent neutralizing mAbs and determining ways to optimize function and deliver to the brain. In these studies, we will elucidate how antiviral Abs with exceptional inhibitory activity exert their action in cell culture and in vivo. The approach will include high efficiency isolation of human mAbs, coupled with innovative antibody gene repertoire studies based on next-gen sequencing. Several hypotheses will be tested, including the concept that ultra-potent neutralizing activity results from features of both the antibodies (selection of optimal V-D-J clonotypes and accumulation of critical somatic mutations) and the antigen (binding to quaternary epitopes on multiple adjacent envelope proteins and blockade of structural transitions critical for virus entry or release). We also will apply new technologies for receptor-mediated transfer of molecules across the blood-brain barrier using engineered sequence changes in the Fc region. Although our focus is to understand how and why ultra-potent human mAbs inhibit EEEV, the studies likely will be relevant to general principles of antibody neutralization of many different encephalitic viruses. In addition to defining the molecular and structural basis of Ab neutralization of EEEV and deploying new strategies for delivery of biologics to the brain, these studies will generate a group of fully human mAbs that can prevent and treat EEEV infection, which could be developed in the near future as a possible therapeutic for humans. Studies in this project, while targeted against EEEV, likely will inform future Ab-based and/or vaccine efforts against other arboviruses that cause human brain infections. We have assembled a unique group of investigators, including a human Ab expert, a molecular virologist with experience in Ab-virus interactions, an animal model and pathogenesis expert with specific expertise in encephalitic alphaviruses, including EEEV, and brain-targeting scientists to pursue these studies.