Fully continuous manufacturing of therapeutic proteins is an emerging trend in the biopharmaceutical industry. Integration of the upstream and downstream processes and conversion to continuous manufacturing leads to increased productivities, decreased equipment size, improved product quality, and overall reduced production costs. An integrated continuous bioprocess (ICB) from the early stages of process development can accelerate the transfer of a new drug candidate to commercial manufacturing. The process presented in this work is a proof-of-concept of an end-to-end monoclonal antibody (mAb) production platform at small scale. It was implemented by a 200 mL ATF perfusion bioreactor, integrated with a single lab-scale chromatography system, performing all purification steps for the mAb from the cell culture harvest in a continuous way. The downstream process consisted of a periodic twin-column capture with protein A resin, followed by a virus inactivation step, a cation exchange step in bind-elute mode, and an anion exchange step in flow-through mode. MAbs were produced for 17 days in a high cell density perfusion culture of CHO cells and purified continuously with a recovery yield of up to 60 % by the following downstream train. A 5-log reduction of host cell protein levels revealed that impurities were sufficiently removed. A consistent glycosylation pattern of the purified product was ensured by the steady-state operation of the process. With this proof-of-concept, we demonstrated the technical feasibility of a fully continuous end-to-end process with a compact design, integrating several unit operations in a single chromatography station and using small-size equipment for the upstream and downstream operations. The work presented here can become a useful development tool for the future of continuous bioprocesses.
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机译：完全连续生产治疗性蛋白质是生物制药行业的一种新兴趋势。上游和下游工艺的集成以及向连续制造的转换可提高生产率，减小设备尺寸，提高产品质量并总体上降低生产成本。从过程开发的早期阶段开始的集成连续生物过程（ICB）可以加速新药候选物向商业生产的转移。这项工作中提出的过程是小规模端对端单克隆抗体（mAb）生产平台的概念验证。它是由200 mL ATF灌注生物反应器实现的，该反应器与单个实验室规模的色谱系统集成在一起，以连续方式执行来自细胞培养物收获物中mAb的所有纯化步骤。下游过程包括用蛋白A树脂定期进行双柱捕获，然后进行病毒灭活步骤，结合-洗脱模式下的阳离子交换步骤和流通模式下的阴离子交换步骤。在CHO细胞的高细胞密度灌注培养中产生单克隆抗体17天，并通过随后的下游培养连续纯化，回收率高达60％。宿主细胞蛋白质水平降低了5个对数，表明杂质已被充分去除。该方法的稳态操作确保了纯化产物的一致的糖基化模式。通过这一概念验证，我们以紧凑的设计展示了完全连续的端到端过程的技术可行性，该过程紧凑，将多个单元操作整合到一个色谱站中，并使用小型设备进行上游和下游操作。这里介绍的工作可以成为未来连续生物过程的有用的开发工具。
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