Laboratory of Malaria Immunology and Vaccinology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 5640 Fishers Lane, Rockville, MD 20852, United States.
Research Technology Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 5640 Fishers Lane, Rockville, MD 20852, United States.
Vaccine. 2019 Sep 10;37(38):5762-5769. doi: 10.1016/j.vaccine.2018.09.037. Epub 2018 Sep 24.
Efforts to develop a vaccine for the elimination of malaria include the use of carrier proteins to assemble monomeric antigens into nanoparticles to maximize immunogenicity. Recombinant ExoProtein A (EPA) is a detoxified form of Pseudomonas aeruginosa Exotoxin A which has been used as a carrier in the conjugate vaccine field. A pilot-scale process developed for purification of EPA yielded product that consistently approached a preset upper limit for host cell protein (HCP) content per human dose. To minimize the risk of bulk material exceeding the specification, the purification process was redeveloped using mixed-mode chromatography resins. Purified EPA derived from the primary and redeveloped processes were comparable following full biochemical and biophysical characterization. However, using a process specific immunoassay, the HCP content was shown to decrease from a range of 0.14-0.24% w/w of total protein to below the level of detection with the revised process. The improved process reproducibly yields EPA with highly similar quality characteristics as the original process but with an improved profile for the HCP content.
为了消除疟疾而开发疫苗的努力包括使用载体蛋白将单体抗原组装成纳米颗粒，以最大限度地提高免疫原性。重组 ExoProtein A（EPA）是铜绿假单胞菌外毒素 A 的一种解毒形式，已被用作结合疫苗领域的载体。为 EPA 的纯化开发的中试规模工艺产生的产品，其宿主细胞蛋白（HCP）含量始终接近每人类剂量的预设上限。为了最大限度地降低散装物料超过规格的风险，使用混合模式色谱树脂重新开发了纯化工艺。经过全面的生化和生物物理特性分析后，发现源自原始工艺和重新开发工艺的纯化 EPA 是可比的。然而，使用特定于工艺的免疫测定法，显示 HCP 含量从总蛋白的 0.14-0.24%w/w 范围下降到用修订后的工艺检测不到的水平。改进后的工艺可重复性地产生具有与原始工艺高度相似质量特征的 EPA，但 HCP 含量的情况有所改善。
