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首页> 外文期刊>Bioprocess and Biosystems Engineering >Pre-stage perfusion and ultra-high seeding cell density in CHO fed-batch culture: a case study for process intensification guided by systems biotechnology
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Pre-stage perfusion and ultra-high seeding cell density in CHO fed-batch culture: a case study for process intensification guided by systems biotechnology

机译：CHO FED分批培养中的前阶段灌注和超高播种细胞密度：系统生物技术引导的过程强化案例研究

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Process intensification strategies are needed in the field of therapeutic protein production for higher productivities, lower cost of goods and improved facility utilization. This work describes an intensification approach, which connects a tangential-flow-filtration (TFF) based pre-stage perfusion process with a concentrated fed-batch production culture inoculated with an ultra-high seeding density (uHSD). This strategy shifted biomass production towards the pre-stage, reaching up to 45 x 10(6) cells/mL in perfusion mode. Subsequently, production in the intensified fed-batch started immediately and the product titer was almost doubled (1.9-fold) in an equivalent runtime and with comparable product quality compared to low-seeded cultures. Driven by mechanistic modelling and next-generation sequencing (NGS) the process had been optimized by selecting the media composition in a way that minimized cellular adaptation between perfusion and production culture. As a main feature, lactate feeding was applied in the intensified approach to promote cell culture performance and process scalability was proven via transfer to pilot-scale i.e., 20 L pre-stage perfusion and 80 L production reactor. Moreover, an earlier shift from a growth associated to a production stage associated gene expression pattern was identified for uHSD cultures compared to the reference. Overall, we showed that the described intensification strategy yielded in a higher volumetric productivity and is applicable for existing or already approved molecules in common, commercial fed-batch facilities. This work provides an in-depth molecular understanding of cellular processes that are detrimental during process intensification.
机译：治疗蛋白质生产领域需要加工强化策略，以获得更高的产品，更低的商品成本和改进的设施利用。这项工作描述了一种强化方法，其基于切向流过滤（TFF）的基级灌注过程，其具有浓缩的批量生产培养物，其接种具有超高播种密度（UHSD）。该策略将生物质生产转移到预阶段，在灌注模式下达到45×10（6）个细胞/ mL。随后，在加强的补料批料中产生立即开始，产品滴度几乎加倍（1.9倍），在等效的运行时，与低种子培养相比具有相当的产品质量。由机械建模和下一代测序（NGS）驱动，通过以最小化灌注和生产培养物之间的细胞适应的方式选择介质组成来优化该过程。作为主要特征，应用乳酸喂养以促进细胞培养性能，并通过转移到导频尺寸的过程可扩展性，即20L预阶段灌注和80L生产反应器。此外，与参考文献相比，针对UHSD培养物鉴定出与生产阶段相关基因表达模式相关的生长的早期转变。总体而言，我们表明，所描述的强化策略以较高的体积生产力产生，适用于现有或已经批准的普通商业美联储设施的分子。这项工作提供了对在过程强化期间有害的细胞过程的深入分子理解。

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