A Novel First-in-class 3D Printing Technology for Advanced Manufacturing of Complex Vaccine Formulations against Influenza and Emerging Infectious Diseases

项目摘要：接种疫苗被认为是最有效的控制传播和 各种传染病的有害影响，包括最近出现的冠状病毒病2019年， 新冠肺炎。重组蛋白亚单位疫苗在免疫方面显示出良好的效果。 最近在对抗传染病方面。这些疫苗是通过重组DNA技术在 编码重组蛋白生产的基因片段被引入宿主细胞作为 一种表达系统。基因工程细胞可以增殖并产生大量的蛋白质。 可在后续步骤中分离和提纯的目标。遗传工具研究的最新进展 微生物操纵技术的发展及哺乳动物细胞系在生物制药中的应用 制造业预测，到今年年底，全球蛋白质市场的规模将达到2284亿美元。然而， 该行业的流程仍然过载，缺乏灵活性和流程控制或集成 连续或按需生产的生产能力。没有生物制造系统可以生产出 重组蛋白质通过一步连续制造过程。因此，由于需求旺盛， 在世界各地，对高效而廉价的技术有着巨大的需求。酵母菌 诸如巴斯德毕赤酵母的表达系统可用作表达宿主细胞 与传统系统相比有许多优点，包括高生长速度，容易的基因操作过程， 高效的蛋白质表达，执行真核翻译后修饰，适当的蛋白质折叠 和蛋白质在外部介质中的分泌，以及易于纯化的过程。在这个项目中，我们将使用一种新的 喷雾多吸附颗粒静置技术(智能3D打印技术，产生生物相容性 Pluronic(F127)-二氨基甲基丙烯酸酯(F127-BUM)聚合物微载体固定化巴斯德毕赤酵母 可用于大规模发酵生产重组蛋白。我们的智能技术 满足重组蛋白制造的要求，如易于放大，正确的蛋白质折叠， 和较短的后期制作过程。它还有可能提高敏捷性、灵活性、成本和健壮性 在复杂的基于蛋白质的生物制剂的制造过程中。此外，与其他颗粒相比 制造技术，SMART可以在单步微粒配方中加入活细胞 进程。这项技术可以很容易地支持进一步的辅助工艺，如超低温冷冻打印 床(-80oC或更低)，用于在线监控关键产品质量属性(CQA)的光纤探头，如 粘度、含量均匀度和稳定性，使其在短期内可用于工业，并具有强大的控制能力 策略。我们的SMART将在连续的装置中实施，以生产干粉生物工程P。 Pastoris包裹F127-BUM微载体生产用于感染疾病的重组蛋白 爱泼斯坦-巴尔病毒(EBV)疫苗和流感疫苗。

PROJECT SUMMARY: Vaccination is known to be the most effective strategy to manage the spread and deleterious impact of various infectious diseases including the most recent emerging, coronavirus disease 2019, COVID-19. Recombinant protein subunit vaccines have demonstrated promising results for immunization against infectious diseases recently. These vaccines are manufactured through recombinant DNA technology in which the gene fragment that encodes the production of the recombinant protein is introduced to a host cell as an expression system. The genetically engineered cells can proliferate and produce a high amount of the protein of the target which can be separated and purified in the succeeding steps. The recent progress in genetic tool development to manipulate the microorganisms and utilization of mammalian cell lines in biopharmaceutical manufacturing have projected the global protein markets to reach $228.4 billion by the end of this year. However, this industry is still overloaded with processes that lack flexibility and process controls or integration needed for continuous or on demand production capacity. There is no biomanufacturing system that can produce recombinant proteins through a single-step continuous manufacturing process. So, due to the high demand for vaccines all over the world, there’s an immense need for highly efficient yet inexpensive technologies. Yeast expression systems such as Pichia pastoris (P. pastoris) can be used as an expression host cell which offers numerous advantages over traditional systems including high growth rate, easy genetic manipulation process, high yield protein expression, performing eukaryotic post-translational modifications, appropriate protein folding and protein secretion in the external medium and easy purification process.In this project we will utilise a novel Sprayed Multi Adsorbed-particle Reposing Technology (SMART 3D printing technique to produce biocompatible Pluronic (F127)-bisurethane methacrylate (F127-BUM) polymers based microcarrier immobilised with P. pastoris which can be used in large-scale fermentations for production of recombinant proteins. Our SMART technology meets the requirements for recombinant proteins manufacturing such as ease of scale-up, correct protein folding, and short post-production processing. It also has the potential to improve agility, flexibility, cost, and robustness in the manufacturing processes for complex protein-based biologics.Additionally, in contrast to other particulate fabrication techniques, SMART can incorporate live cells during the single-step microparticle formulation process. This technology can easily host further ancillary processes such as ultra-low temperature freezing print bed (-80oC or lower), fibre optic probes for the inline monitoring of critical product quality attributes (CQAs) such as viscosity, content uniformity and stability, making it accessible to industry in the near term with a robust control strategy. Our SMART will be implemented in a continuous setup to manufacture dry powder bioengineered P. pastoris encapsulated F127-BUM microcarriers to produce recombinant proteins for infectious diseases such as vaccines against Epstein-Barr virus (EBV) and influenza vaccines.