HAW-Hamburg University of Applied Sciences, D-21033 Hamburg, Germany.
Biotechnol J. 2013 Jun;8(6):738-47. doi: 10.1002/biot.201200190.
The identification of optimal expression conditions for state-of-the-art production of pharmaceutical proteins is a very time-consuming and expensive process. In this report a method for rapid and reproducible optimization of protein expression in an in-house designed small-scale BIOSTAT® multi-bioreactor plant is described. A newly developed BioPAT® MFCS/win Design of Experiments (DoE) module (Sartorius Stedim Systems, Germany) connects the process control system MFCS/win and the DoE software MODDE® (Umetrics AB, Sweden) and enables therefore the implementation of fully automated optimization procedures. As a proof of concept, a commercial Pichia pastoris strain KM71H has been transformed for the expression of potential malaria vaccines. This approach has allowed a doubling of intact protein secretion productivity due to the DoE optimization procedure compared to initial cultivation results. In a next step, robustness regarding the sensitivity to process parameter variability has been proven around the determined optimum. Thereby, a pharmaceutical production process that is significantly improved within seven 24-hour cultivation cycles was established. Specifically, regarding the regulatory demands pointed out in the process analytical technology (PAT) initiative of the United States Food and Drug Administration (FDA), the combination of a highly instrumented, fully automated multi-bioreactor platform with proper cultivation strategies and extended DoE software solutions opens up promising benefits and opportunities for pharmaceutical protein production.
确定最先进的药物蛋白生产的最佳表达条件是一个非常耗时和昂贵的过程。在本报告中，描述了一种在内部设计的小型 BIOSTAT®多生物反应器工厂中快速且可重复地优化蛋白表达的方法。一种新开发的 BioPAT®MFCS/win 实验设计（DoE）模块（德国 Sartorius Stedim Systems）将过程控制系统 MFCS/win 和 DoE 软件 MODDE®（瑞典 Umetrics AB）连接起来，从而实现了全自动优化程序的实施。作为概念验证，已对商业毕赤酵母 KM71H 株进行了转化，以表达潜在的疟疾疫苗。与初始培养结果相比，通过 DoE 优化程序，这种方法使完整蛋白分泌生产力提高了一倍。在下一步中，已证明在确定的最佳条件下，该过程对工艺参数变化的敏感性具有稳健性。通过七个 24 小时培养周期，建立了一种显著提高的药物生产工艺。具体而言，针对美国食品和药物管理局（FDA）过程分析技术（PAT）倡议中指出的监管要求，将高度仪器化的、全自动的多生物反应器平台与适当的培养策略和扩展的 DoE 软件解决方案相结合，为药物蛋白生产带来了有希望的益处和机遇。
