Department of Chemical, Biological and Environmental Engineering, Universitat Autònoma de Barcelona, 08193, Cerdanyola del Vallès, Spain.
IQS School of Engineering, Universitat Ramon Llull, 08017, Barcelona, Spain.
Appl Microbiol Biotechnol. 2018 Dec;102(24):10469-10483. doi: 10.1007/s00253-018-9388-4. Epub 2018 Oct 5.
The increasing demand for biopharmaceuticals produced in mammalian cells has driven the industry to enhance productivity of bioprocesses through different strategies. This is why fed-batch and perfusion cultures are considered more attractive choices than batch processes. In this context, the availability of reliable online measuring systems for cell density and metabolic activity estimation will help the application of these processes. The present work focuses on the comparison of two different monitoring tools for indirect estimation of biomass concentration in a HEK293 cell cultures producing IFN-γ: on one side, the oxygen uptake rate (O.U.R.) determination, by means of application of the dynamic method measurement which is already a widely used tool and, on the other side, a new robust online monitoring tool based on the alkali buffer addition used to maintain the pH set point. Both strategies allow a proper monitoring of cell growth and metabolic activity, with precise identification of the balanced cell growth and the most important action in the process, as is the media feeding. The application of these monitoring systems in fed-batch processes allows extending the growth of HEK293 cells, which in turn results in higher final cell concentrations compared with Batch strategy (7 · 10 cells mL), achieving 14 · 10 cells mL for the fed-batch based on O.U.R. and 19 · 10 cells mL for the fed-batch based on the alkali addition. Product titter is also increased in respect of the batch strategy (3.70 mg L), resulting in 8.27 mg L when fed-batch was based on O.U.R. and 11.49 mg L when it was based on the alkali buffer strategy. Results prove that fed-batch strategy based on the alkali buffer addition is a robust online monitoring method that has shown its great potential to optimize the feeding strategy in HEK293 fed-batch cultures.
由于对哺乳动物细胞生产的生物制药的需求不断增加，业界通过不同的策略来提高生物工艺的生产力。这就是为什么与分批过程相比，补料分批和灌注培养被认为是更有吸引力的选择。在这种情况下，可靠的在线测量系统可用于细胞密度和代谢活性估计，这将有助于这些过程的应用。本工作重点比较了两种不同的监测工具，用于间接估计生产 IFN-γ的 HEK293 细胞培养物中的生物量浓度：一方面，通过应用已经广泛使用的动态方法测量来确定耗氧率 (O.U.R.)，另一方面，使用添加碱性缓冲液来维持设定 pH 值的新的稳健在线监测工具。这两种策略都可以对细胞生长和代谢活性进行适当的监测，并精确识别细胞生长和过程中的重要操作，如补料。这些监测系统在补料分批过程中的应用可以延长 HEK293 细胞的生长，从而与分批策略相比，最终细胞浓度更高（7×10^6 细胞/mL），基于 O.U.R 的补料分批达到 14×10^6 细胞/mL，基于碱添加的补料分批达到 19×10^6 细胞/mL。与分批策略相比，产物滴度也有所提高（3.70mg/L），基于 O.U.R 的补料分批达到 8.27mg/L，基于碱缓冲策略的补料分批达到 11.49mg/L。结果表明，基于碱性缓冲液添加的补料分批策略是一种稳健的在线监测方法，已显示出在优化 HEK293 补料分批培养物的补料策略方面的巨大潜力。
