During the CHO cell culture process, dissolved carbon dioxide (pCO2) accumulation has been recognized as an important factor which needs to be well controlled for successful scale-up. Adverse effects on cell growth performance and protein production are observed either at low or high CO2 concentration in the medium. Strategies have been developed to alleviate the accumulation of pCO2, including replacement of pH buffers in the culture medium and using different ways to maintain culture pH other than CO2 and NaOH. However, the understanding of how CO2 affects cell growth, metabolism, productivity and product quality are still limited. Therefore, this work aims to reveal the influence of low and high pCO2 levels on cell metabolism and protein production to provide mechanistic information for further optimization. The pCO2 profiles of CHO cells in various culture scales with different production strategies were examined to find a wide range from 10mmHg to 300mmHg. To mimic the circumstance of large-scale reactors, low and high pCO2 levels were maintained in shake flasks to discover more details in intracellular process. Cell metabolism was investigated by transcriptome and metabolome analysis using RNA-sequencing and LC-MS, respectively. The results showed that elevated pCO2 decreased the metabolic activity of mitochondrial；However, low pCO2 took its effect mainly on pyruvate metabolism. The changes of key enzymes activities suggested that the elevated pCO2 inhibited cell growth by product inhibition, while low pCO2 reduced the metabolic activity by increasing the intracellular pH. Furthermore, TCA cycle intermediates or their precursors were added at low and high pCO2 levels to reveal and break through the bottlenecks of metabolic pathways. In short, this study not only provides insight into the metabolic responses of CHO cell lines to CO2 , but also solid background for further optimization of manufacturing process.

Elucidating the effects of CO2 on CHO cell metabolism in bioprocessing

253rd National Meeting (2017)

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