Patel Nisarg, Davis Zach, Hofmann Carl, Vlasak Josef, Loughney John W, DePhillips Pete, Mukherjee Malini
Analytical Research & Development, Merck & Co., Inc., 770 Sumneytown Pike, West Point, PA 19486, USA.
Vaccines (Basel). 2023 Jul 10;11(7):1224. doi: 10.3390/vaccines11071224.
Messenger RNA (mRNA) vaccines have emerged as a flexible platform for vaccine development. The evolution of lipid nanoparticles as effective delivery vehicles for modified mRNA encoding vaccine antigens was demonstrated by the response to the COVID-19 pandemic. The ability to rapidly develop effective SARS-CoV-2 vaccines from the spike protein genome, and to then manufacture multibillions of doses per year was an extraordinary achievement and a vaccine milestone. Further development and application of this platform for additional pathogens is clearly of interest. This comes with the associated need for new analytical tools that can accurately predict the performance of these mRNA vaccine candidates and tie them to an immune response expected in humans. Described here is the development and characterization of an imaging based in vitro assay able to quantitate transgene protein expression efficiency, with utility to measure lipid nanoparticles (LNP)-encapsulated mRNA vaccine potency, efficacy, and stability. Multiple biologically relevant adherent cell lines were screened to identify a suitable cell substrate capable of providing a wide dose-response curve and dynamic range. Biologically relevant assay attributes were examined and optimized, including cell monolayer morphology, antigen expression kinetics, and assay sensitivity to LNP properties, such as polyethylene glycol-lipid (or PEG-lipid) composition, mRNA mass, and LNP size. Collectively, this study presents a strategy to quickly optimize and develop a robust cell-based potency assay for the development of future mRNA-based vaccines.
信使核糖核酸（mRNA）疫苗已成为一种灵活的疫苗开发平台。脂质纳米颗粒作为编码疫苗抗原的修饰mRNA的有效递送载体的发展，在对新冠疫情的应对中得到了体现。从刺突蛋白基因组快速开发出有效的严重急性呼吸综合征冠状病毒2（SARS-CoV-2）疫苗，然后每年生产数十亿剂，这是一项非凡的成就，也是疫苗领域的一个里程碑。显然，将该平台进一步开发并应用于其他病原体是很有意义的。这伴随着对新分析工具的相关需求，这些工具能够准确预测这些mRNA候选疫苗的性能，并将其与人类预期的免疫反应联系起来。本文描述了一种基于成像的体外分析方法的开发和表征，该方法能够定量转基因蛋白表达效率，可用于测量脂质纳米颗粒（LNP）包裹的mRNA疫苗的效力、功效和稳定性。对多种具有生物学相关性的贴壁细胞系进行了筛选，以确定一种能够提供宽剂量反应曲线和动态范围的合适细胞底物。研究并优化了具有生物学相关性的分析属性，包括细胞单层形态、抗原表达动力学以及分析方法对LNP特性（如聚乙二醇脂质（或PEG脂质）组成、mRNA质量和LNP大小）的敏感性。总体而言，本研究提出了一种策略，可快速优化并开发一种强大的基于细胞的效力分析方法，用于未来基于mRNA的疫苗的开发。
