Cell-free synthesis of influenza virus-like particles (VLPs) as prototyping platform for vaccine development and variant characterization

流感病毒样颗粒 (VLP) 的无细胞合成作为疫苗开发和变体表征的原型平台

• 批准号: 10648472
• 负责人: Irina V Novikova
• 金额: $ 8.07万
• 依托单位: BATTELLE PACIFIC NORTHWEST LABORATORIES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10648472
• 关键词: Address; Antigens; Architecture; Biomanufacturing; Biomedical Engineering; Capsid; Cell Culture Techniques; Cell-Free System; Cells; Cryo-electron tomography; Cryoelectron Microscopy; Disease Outbreaks; Drug Delivery Systems; Electron Microscopy; Ensure; Future; Generations; Germ Cells; Glycoproteins; Goals; Hepatitis B Virus; Heterogeneity; Heterophile Antigens; Human; Individual; Influenza; Influenza A Virus, H5N1 Subtype; Influenza A Virus, H7N9 Subtype; Influenza A Virus, H9N2 Subtype; Influenza A virus; Learning; Mass Spectrum Analysis; Microsomes; Morphology; Mutation; Negative Staining; Pichia; Population; Production; Protein Biosynthesis; Proteins; Recombinants; Resolution; Safety; Sampling; Surface; System; Technology; Testing; Vaccine Research; Vaccines; Variant; Viral Genome; Virus; Virus-like particle; Wheat; cost; density; flexibility; future pandemic; glycosylation; immunogenic; influenza virus strain; influenza virus vaccine; influenzavirus; manufacture; pandemic disease; particle; prophylactic; protein expression; prototype; recombinant virus; research and development; response; self assembly; structural biology; three dimensional structure; tomography; vaccine development; vaccine platform; virology

SUMMARY The absence of prophylactic and low-cost influenza vaccines along with slow production rate makes us unprepared for the next pandemic outbreak worldwide. Recombinant virus-like particles (VLPs) vaccines provide us with unique opportunities to address these difficulties, but their recombinant synthesis strategies are limited to cell culture production, which has its challenges such as limited throughput and architectural heterogeneity of the obtained products. In this project, we aim to combine wheat germ cell-free protein expression platform with active microsomes of different origins (including human) to synthesize glycosylated influenza virus-like particles (VLPs). These particles will further undergo detailed morphological and compositional examination using electron microscopy and native mass spectrometry to understand the architecture of obtained products and understand the mechanisms of their assembly and composition variability. High- resolution tomography will be utilized to determine the best VLP candidates with maximal antigen occupancy and thus maximal vaccine efficiency. We hypothesize that this approach will be suitable for a stable and streamlined generation of Influenza VLPs and will offer much more flexibility and control over their architectural heterogeneity and eventually serve as a rapid prototyping platform for influenza vaccine development. The current project will also inform us about the utility of using such approaches for virology and for glycoprotein synthesis in general.

概括 缺乏预防性和低成本流感疫苗，且生产速度缓慢 使我们对下一次全球大流行爆发毫无准备。重组病毒样颗粒 （VLP）疫苗为我们提供了解决这些困难的独特机会，但它们的 重组合成策略仅限于细胞培养生产，这有其挑战 例如有限的吞吐量和所获得产品的架构异质性。在这个项目中， 我们的目标是将小麦胚芽无细胞蛋白表达平台与活性微粒体相结合 不同来源（包括人类）合成糖基化流感病毒样颗粒（VLP）。 这些颗粒将进一步接受详细的形态和成分检查 电子显微镜和天然质谱来了解所获得的结构 产品并了解其组装和成分变异的机制。高的- 将利用分辨率断层扫描来确定具有最大抗原的最佳 VLP 候选者 占用率，从而实现最大的疫苗效率。我们假设这种方法将 适用于稳定且精简的流感 VLP 生成，并将提供更多功能 灵活性和对其架构异构性的控制，并最终作为快速 流感疫苗开发的原型平台。目前的项目也会告诉我们 关于使用此类方法进行病毒学和糖蛋白合成的实用性。