S53: Control of Lactate Consumption Metabolic State Through Manipulation of Glycolytic Activity

Cultured mammalian cells consume large amounts of glucose and divert most of it towards lactate whose accumulation is inhibitory to growth and product synthesis.   In fedbatch-cultures, fortuitous metabolic shifts to low lactate production or consumption leads to sustained viability and higher productivity of recombinant proteins.  This was evident in the strong correlation between superior productivity and the occurrence of metabolic shift in late stages in a survey of process data of over two hundred manufacturing runs.  The mechanism governing such metabolic shifts is elusive although a consistent occurrence of such shifts is highly desired.

Multivariate analysis of the process data revealed that, in late stages, runs consuming lactate had lower glucose uptake rates than those with no consumption.  Using a kinetic model for the central metabolic network, we identified low glucose flux and high lactate concentration as key factors for such a metabolic shift.  Simulations showed that the low glycolytic activity can be an outcome of low culture-glucose availability or inhibition of rate controlling glycolytic enzymes.  Experimentally, by controlling the culture-nutrient levels in late stages of the culture, we observed reproducible shift to lactate consumption state in fedbatch-cultures of both mouse myeloma and Chinese hamster ovary cell-lines.  Further, in the late stages of culture, inhibition of signaling pathways regulating glycolysis, such as AKT-mTOR, also led to lactate consumption.  As the metabolic state of a culture is also often related to product quality, this strategy of robust control of metabolic shift shall enhance process robustness and improve productivity and product quality.