Structure-based design of broad flavivirus immunogens

广泛黄病毒免疫原的基于结构的设计

基本信息

批准号：
10685350
负责人：
Jonathan R. Lai
金额：
$ 73.88万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-24 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10685350
关键词：
Acute Address Antibodies Antibody Response Antibody titer measurement Antibody-Dependent Enhancement Antigens Attenuated Attenuated Vaccines Binding Binding Sites Cells Cessation of life Characteristics Complex Country Culicidae Dengue Dengue Fever Dengue Vaccine Dengue Virus Dengvaxia Development Disease Emerging Communicable Diseases Engineering Epitopes Europe European Union Evaluation Exhibits Fever Flavivirus Geometry Glycoproteins Goals Head Hospitalization Human IgG Receptors Immune Immune response Immunity Immunization Immunize Individual Infection Lateral Licensing Licensure Masks Membrane Membrane Fusion Membrane Proteins Methods Molecular Monoclonal Antibodies Motion Mus Mutation Patients Phage Display Polysaccharides Primary Infection Protein Engineering Proteins Rapid screening Reporting Risk Safety Serotyping Shock Site Structure Subunit Vaccines Surface Syndrome System Tail Testing Vaccine Design Vaccines Variant Viral Virus Work Yellow fever virus ZIKA Zika Virus cross reactivity design dimer flavivirus glycoprotein E glycosylation innovation manufacture nanoparticle neutralizing antibody neutralizing monoclonal antibodies next generation novel particle phase III trial premature receptor receptor binding response secondary infection severe dengue transmission process vaccine candidate vaccine development vaccine strategy vector vaccine

项目摘要

SUMMARY Dengue virus is a mosquito-transmitted flavivirus that causes an estimated 390 million human infections each year. There are four serotypes of Dengue (DENV1-4) that co-circulate in hyperendemic regions. Primary infection by a single DENV serotype results in febrile illness and subsequent durable immunity to that serotype. Secondary infections by heterotypic serotypes can lead to severe shock syndrome and death. Severe Dengue disease is caused in part by cross-reactive antibodies elicited during primary infection that can bind heterologous DENV serotypes but cannot neutralize them. Instead, these non-neutralizing antibodies facilitate entry and infection in Fcγ receptor-positive cells, thus causing "antibody-dependent enhancement" (ADE) of infection. While a live-attenuated four-component chimeric vaccine was recently deployed in 19 countries and Europe, this vaccine does not protect naïve individuals against symptomatic or severe infection, and may even exacerbate disease in some cases. Furthermore, the global emergence of Zika virus (ZIKV), and the potential for ADE between DENV and ZIKV, raises concerns for vaccine strategies containing most or all epitopes in the E glycoprotein. Nonetheless, the isolation and characterization of protective and, in some cases, broadly-neutralizing antibodies indicates that certain epitopes within the E glycoprotein may have the capacity to elicit broadly protective responses. Here, we utilize innovative protein engineering approaches to develop “immune-focused” antigens as potential vaccine candidates, in which epitopes that induce non-neutralizing antibodies are masked by engineered mutations or glycosylation. Our hypothesis is that masking of these unfavorable epitopes will skew the immune response toward a stronger neutralizing, protective, and broad response. Aims 1 and 2 focus on critical epitopes in DENV and ZIKV E domain III (EDIII), and Aim 3 explores glycan masking of the ZIKV E prefusion dimer to immune focus on the E-dimer epitope (EDE). EDIII is attractive for subunit vaccine design because it is the target of potently neutralizing and protective antibodies for both DENV and ZIKV. However, immunization with wild-type EDIII protein results in induction of both neutralizing and non-neutralizing antibodies that engage a variety of epitopes. We have used phage display to mask unproductive epitopes of DENV and ZIKV EDIIIs by mutation, while maintaining neutralizing epitopes. These “resurfaced EDIIIs” (rsDIIIs) will be conjugated to protein nanoparticles and their capacity to induce neutralizing and protective antibody response in mice evaluated. To immune focus the prefusion E dimer on the EDE, we have developed a mammalian display system that allows for rapid evaluation of E dimer constructs for binding to EDE mAbs. We will utilize this system to screen variants with multiple engineered glycosylation sites that mask the surface outside of the EDE. The most promising candidates will be tested for their capacity to induce EDE-like mAbs in mice. This work will provide a proof-of-concept for novel subunit vaccine candidates against DENV, ZIKV, and possibly other flaviviruses of global concern.

概括登革热病毒是一种蚊子传播的黄病毒，估计会引起大约3.9亿人的感染年。高流血区域中有四种登革热（DENV1-4）的血清型。基本的单个DENV血清型感染导致发热疾病，随后耐用的免疫力血清型。异型血清型的继发感染会导致严重的休克综合征和死亡。严重的登革热疾病部分是由于原发性感染过程中引起的交叉反应性抗体引起的可以结合异源DENV血清型，但不能中和它们。相反，这些不中和抗体有助于进入和感染Fcγ受体阳性细胞，从而导致“抗体依赖性感染的增强（ADE）。该疫苗在19个国家和欧洲部署，不能保护个人免受症状或严重的感染，甚至可能在某些情况下加剧疾病。此外，全球出现 Zika病毒（ZIKV），以及DENV和ZIKV之间ADE的潜力，引起了人们对疫苗策略的关注在糖蛋白中包含大多数或所有表位。尽管如此，保护性，在某些情况下，广泛中和化的抗体表明e内某些表位糖蛋白可能具有引起广泛保护的反应的能力。在这里，我们利用创新蛋白工程方法将“以免疫为中心”的抗原作为潜在疫苗候选者影响非中和抗体的表位被工程突变或糖基化掩盖。我们的假设是，掩盖这些不利的表位会使免疫反应偏向更强中和，保护和广泛的反应。目标1和2关注DENV和ZIKVE的关键表位域III（EDII），AIM 3探索了zikv e的聚糖掩蔽，以免疫聚焦于对该二聚体的聚焦电子二聚体（EDE）。 EDIII对亚基疫苗设计具有吸引力，因为它是潜在的目标中和和保护DENV和ZIKV的抗体。但是，用野生型EDIII免疫蛋白质会导致诱导中和和非中和抗体的诱导表位。我们已经使用噬菌体显示来掩盖DENV和ZIKV EDIIIS的非生产性表位，同时保持中和表位。这些“重新浮出水面的Ediiis”（RSDIIIS）将与蛋白质结合纳米颗粒及其在评估的小鼠中诱导中和和保护抗体反应的能力。为了免疫聚焦在EDE上的浓度二聚体，我们开发了一个哺乳动物显示系统允许快速评估E二聚体构建体，以结合EDE MAB。我们将利用此系统具有多个工程糖基化位点的屏幕变体，它们掩盖了EDE外面的表面。这最有前途的候选人将测试其在小鼠中诱导类似Ede的单盘的能力。这项工作将为针对DENV，ZIKV和其他可能的新型亚基疫苗提供概念证明全球关注的黄病毒。