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病毒，可导致人衰弱 人类脑炎疾病。大约三分之一的人类 EEEV 感染病例死亡，许多幸存者 长期的、使人衰弱的神经系统问题。该病毒在 Culiseta 之间维持地方性动物流行循环 黑色蚊子和鸟类宿主，但可以通过以下方式传播给人类和马： 一些伊蚊、 Coquillettidia 和库蚊属物种。这种感染在人类中并不常见，但频率正在增加 近年来，可能继发于气候变化、病媒扩张和其他不典型因素。 EEEV 由于通过气溶胶途径传播，也被视为潜在的生物恐怖主义威胁。尽管具有高致病性 由于该病毒的性质，目前尚无针对 EEEV 的具体治疗方法或疫苗。该项目的主要目标是 定义超强中和人类的分子、遗传、免疫学和结构特征 单克隆抗体在体内具有广泛的抗 EEEV 活性。其他目标包括定义以下机制的相关性： 这些超强中和单克隆抗体的保护，并确定优化功能和传递给 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如何在 细胞培养和体内。该方法将包括人类单克隆抗体的高效分离，以及 基于下一代测序的创新抗体基因库研究。将测试几个假设， 包括超强中和活性源自两种抗体特征的概念（选择 最佳 V-D-J 克隆型和关键体细胞突变的积累）和抗原（结合 多个相邻包膜蛋白上的四级表位以及对病毒至关重要的结构转变的阻断 进入或释放）。我们还将应用新技术进行受体介导的分子跨域转移 血脑屏障利用 Fc 区的工程序列变化。虽然我们的重点是了解 超强人类单克隆抗体如何以及为何抑制 EEEV，这些研究可能与以下一般原则相关： 抗体中和许多不同的脑炎病毒。除了定义分子和结构 EEEV 抗体中和的基础和部署向大脑输送生物制剂的新策略，这些 研究将产生一组完全人类单克隆抗体，可以预防和治疗 EEEV 感染，这可能是 在不久的将来被开发为一种可能的人类治疗方法。该项目的研究同时针对 EEEV，可能将为未来针对导致人类大脑的其他虫媒病毒的基于抗体和/或疫苗的努力提供信息 感染。我们组建了一个独特的研究小组，包括一名人类抗体专家、一名分子抗体专家、一名 具有抗体病毒相互作用经验的病毒学家、具有特定知识的动物模型和发病机制专家 包括 EEEV 在内的脑炎甲病毒方面的专业知识，以及从事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.