A mucosally targeted MERS-CoV vaccine produced in plants

在植物中生产的粘膜靶向 MERS-CoV 疫苗

• 批准号: 9141182
• 负责人: KEITH WYCOFF
• 金额: $ 23.65万
• 依托单位: PLANET BIOTECHNOLOGY, INC.
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-02-17 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9141182
• 关键词: Acute; Amino Acid Sequence; Amino Acids; Animals; Antibodies; Antigen Presentation; Antigen-Presenting Cells; Binding; Camels; Case Study; Cell Culture Techniques; Cells; Cellular Immunity; Cessation of life; Chimeric Proteins; Contracts; Coronavirus; Coronavirus spike protein; Country; Development; Dipeptidyl Peptidases; Epidemic; Fatality rate; Fc Receptor; Fc domain; Flow Cytometry; Human; IgG1; Immune response; Immunization; Immunize; In Vitro; Kilogram; Mammalian Cell; Measures; Mediating; Middle East; Middle East Respiratory Syndrome Coronavirus; Modification; Molecular Biology; Mucosal Immune Responses; Mus; Phase; Plants; Play; Process; Production; Proteins; Recombinant Fusion Proteins; Recombinant Proteins; Reporting; Research; Role; Saudi Arabia; Small Business Innovation Research Grant; Staining method; Stains; Surface; System; T cell response; Testing; Therapeutic; Time; Vaccine Design; Vaccines; Variant; Viral; Virus; Virus Receptors; World Health Organization; antigen binding; cost; cytokine; epidemiology study; immunogenicity; improved; in vivo; milligram; mortality; neonatal Fc receptor; neutralizing antibody; pandemic disease; public health relevance; receptor binding; residence; respiratory; trafficking; transmission process; vaccine candidate

 DESCRIPTION (provided by applicant): Middle East Respiratory Syndrome (MERS) coronavirus (MERS-CoV) causes severe acute respiratory illness with a high fatality rate among reported cases. MERS was first reported in 2012, but the virus has been circulating in camels, which are thought to be an animal reservoir, for at least 20 years. To date over 1000 MERS cases have been reported to the World Health Organization, including over 400 deaths. While most cases have been in Saudi Arabia and contact with camels or camel products has been implicated in some cases, human-to-human transmission has been documented as well and the virus is thought to have epidemic potential. There are currently no approved vaccines or therapeutics for MERS. The MERS-CoV spike (S) protein mediates viral entry into host cells expressing the viral receptor dipeptidyl peptidase 4 (DPP4). Within the S1 subunit, a Receptor Binding Domain (RBD) of about 200 amino acids has been identified. Immunization of mice with a recombinant fusion protein containing S377-588 fused to human IgG1 Fc (S377-588-Fc) induced high titers of neutralizing antibodies in immunized animals. Unfortunately, expression of S377-588-Fc (or RBD-Fc) in mammalian cell culture is low, which may be an impediment for further development of this promising candidate vaccine. We will circumvent the production problem by producing MERS-CoV RBD-Fc, and modifications thereof, using a well-established plant-expression system. Our scalable transient plant-expression system allows rapid production of recombinant protein (within a week); with protein production costs less than 10% that of mammalian cell systems. We will produce variants of MERS-CoV RBD-Fc with enhanced binding to the mouse neonatal Fc receptor (FcRn), in order to test the hypothesis that improved antigen presentation via FcRn will result in an enhanced immune response. We will evaluate the immunogenicity of mucosally-administered MERS-CoV RBD-Fc in mice, by analyzing both humoral and cell-mediated immunity. We will evaluate titers of RBD-specific antibodies, analyze T cell responses by intracellular cytokine staining followed by flow cytometry analysis, and will measure the ability of sera from immunized mice to neutralize live MERS-CoV in vitro.

 描述（由适用提供）：中东呼吸综合征（MERS）冠状病毒（MERS-COV）引起严重的急性呼吸道疾病，报告病例中死亡率很高。 MERS于2012年首次报道，但该病毒一直在骆驼中循环，至少20年。迄今为止，已向世界卫生组织报告了超过1000例MERS病例，其中包括400多人死亡。尽管大多数病例都在沙特阿拉伯，并且在某些情况下已经实施了与骆驼或骆驼产品的接触，但也已经记录了人与人之间的传播，并且该病毒被认为具有流行潜力。目前尚无批准的MERS疫苗或治疗。 MERS-COV尖峰（S）蛋白介导了表达病毒受体二肽基肽4（DPP4）的宿主细胞的病毒进入。在S1亚基中，已经鉴定出约200个氨基酸的受体结合结构域（RBD）。用含有S377-588的重组融合蛋白进行的小鼠免疫接种，该蛋白与人IgG1 Fc（S377-588-FC）融合在一起，诱导了免疫动物中和中和抗体的高滴度。不幸的是，哺乳动物细胞培养中S377-588-FC（或RBD-FC）的表达较低，这可能是进一步开发这种诺言候选疫苗的障碍。我们将使用建立良好的植物表达系统来避免生产问题及其修改。我们可扩展的瞬态植物表达系统允许快速生产重组蛋白（一周之内）；蛋白质生产的成本不到哺乳动物细胞系统的10％。我们将产生与小鼠新生儿FC受体（FCRN）结合增强的MERS-COV RBD-FC的变体，以检验以下假设：通过FCRN改善的抗原呈递将导致免疫响应增强。我们将通过分析体液和细胞介导的免疫史地，评估小鼠中粘液化的MERS-COV RBD-FC的免疫原性。我们将评估RBD特异性抗体的滴度，通过细胞内细胞因子染色，然后进行流式细胞仪分析来分析T细胞反应，并测量来自免疫小鼠的血清在体外中和实时MERS-COV的能力。