Modelling of Cell Metabolism to Reduce the Lactate Generation in CHO Cell Cultures

Chinese Hamster Ovary cells (CHO) display Warburg metabolism characterized by high glucose consumption and high lactate production under aerobic conditions. Lactate is a by-product widely reported to inhibit cell growth in culture, imposing an important burden to industrial processes. In order to reduce lactate secretion and thus its inhibitory effects, two different approaches have been applied: i) Bioprocess Engineering, controlling extracellular conditions in the bioreactor to trigger concomitant glucose/lactate consumption, and ii) Synthetic Biology, to generate non-lactate producer mutant cells knocking out lactate dehydrogenase and the regulators responsible for inhibiting pyruvate conversion to acetyl-CoA.

In the current study, the metabolic profile of CHO cells was investigated in batch bioreactor cultures performed under three conditions: normal WT, controlled WT, and zero-lactate CHO (CHO ZeLa). Exometabolomic data was integrated into a reduced genome-scale metabolic model using Flux Balance Analysis (FBA) during the mid-exponential phase, as well as Dynamic FBA (DFBA) to capture the dynamic changes occurring over time in CHO cell metabolism. Model reduction was performed through a novel semi-automated reduction protocol to generate fully functional metabolic models.

FBA showed that in wild-type cell line, lactate is produced to fulfill the NADH regeneration requirements in the cytoplasm and only a small amount of pyruvate is introduced into TCA through Acetyl-CoA. When concomitant glucose and lactate consumption was triggered, as well as in CHO ZeLa, glucose uptake was significantly reduced and a balance between glycolysis and TCA cycle fluxes was reached, yielding a more efficient substrate consumption. Moreover, DFBA illuminated the metabolic mechanisms by which wild-type CHO switches from a Warburg (glucose consumption/lactate production) phenotype to a glucose/lactate co-consumption phenotype.

Our approach enabled us to explore the mechanisms underlying dynamic metabolic response of wild-type CHO and CHO ZeLa with potential implications in the industry of bioproducts.