Zimmermann Sarah, Scheeder Christian, Zimmermann Philipp K, Bogsnes Are, Hansson Mattias, Staby Arne, Hubbuch Jürgen
Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.
Novo Nordisk A/S, Biopharm Purification Development & Virology, Gentofte, Denmark.
Biotechnol J. 2017 Feb;12(2). doi: 10.1002/biot.201600587. Epub 2016 Dec 21.
The availability of preparative-scale downstream processing strategies for cell-based products presents a critical juncture between fundamental research and clinical development. Aqueous two-phase systems (ATPS) present a gentle, scalable, label-free, and cost-effective method for cell purification, and are thus a promising tool for downstream processing of cell-based therapeutics. Here, the application of a previously developed robotic screening platform that enables high-throughput cell partitioning analysis in ATPS is reported. In the present case study a purification strategy for two model cell lines based on high-throughput screening (HTS)-data and countercurrent distribution (CCD)-modeling, and validated the CCD-model experimentally is designed. The obtained data are shown an excellent congruence between CCD-model and experimental data, indicating that CCD-models in combination with HTS-data are a powerful tool in downstream process development. Finally, the authors are shown that while cell cycle phase significantly influences cell partitioning, cell type specific differences in surface properties are the main driving force in charge-dependent separation of HL-60 and L929 cells. In order to design a highly robust purification process it is, however, advisable to maintain constant growth conditions.
用于基于细胞的产品的制备规模下游加工策略的可用性，是基础研究与临床开发之间的一个关键节点。双水相系统（ATPS）为细胞纯化提供了一种温和、可扩展、无标记且具有成本效益的方法，因此是基于细胞的治疗药物下游加工的一种有前景的工具。在此，报道了一种先前开发的机器人筛选平台的应用，该平台能够在ATPS中进行高通量细胞分配分析。在本案例研究中，基于高通量筛选（HTS）数据和逆流分配（CCD）建模设计了两种模型细胞系的纯化策略，并通过实验验证了CCD模型。获得的数据表明CCD模型与实验数据之间具有极好的一致性，这表明CCD模型与HTS数据相结合是下游工艺开发中的一个强大工具。最后，作者表明，虽然细胞周期阶段显著影响细胞分配，但表面性质的细胞类型特异性差异是HL-60和L929细胞电荷依赖性分离的主要驱动力。然而，为了设计一个高度稳健的纯化工艺，建议保持恒定的生长条件。
