The impact of pH inhomogeneities on CHO cell physiology and fed‐batch process performance – two‐compartment scale‐down modelling and intracellular pH excursion

Abstract

Due to high mixing times and base addition from top of the vessel, pH inhomogeneities are most likely to occur during large-scale mammalian processes. The goal of this study was to set-up a scale-down model of a 10-12 m stirred tank bioreactor and to investigate the effect of pH perturbations on CHO cell physiology and process performance. Short-term changes in extracellular pH are hypothesized to affect intracellular pH and thus cell physiology. Therefore, batch fermentations, including pH shifts to 9.0 and 7.8, in regular one-compartment systems are conducted. The short-term adaption of the cells intracellular pH are showed an immediate increase due to elevated extracellular pH. With this basis of fundamental knowledge, a two-compartment system is established which is capable of simulating defined pH inhomogeneities. In contrast to state-of-the-art literature, the scale-down model is included parameters (e.g. volume of the inhomogeneous zone) as they might occur during large-scale processes. pH inhomogeneity studies in the two-compartment system are performed with simulation of temporary pH zones of pH 9.0. The specific growth rate especially during the exponential growth phase is strongly affected resulting in a decreased maximum viable cell density and final product titer. The gathered results indicate that even short-term exposure of cells to elevated pH values during large-scale processes can affect cell physiology and overall process performance. In particular, it could be shown for the first time that pH perturbations, which might occur during the early process phase, have to be considered in scale-down models of mammalian processes.

Analysis of Golgi pH in Chinese hamster ovary cells using ratiometric pH‐sensitive fluorescent proteins

“…As observed previously (Brunner et al, ; Kim et al, ), cytoplasm pH, unlike the Golgi pH, was affected by the pH of the culture medium. The cytoplasm pH remained almost constant during bioreactor cultures with pH maintained at 7.2.…”

Workflow for Target‐Oriented Parametrization of an Enhanced Mechanistic Cell Culture Model

“…Therefore several models describing mammalian cell cultures exist, for extensive reviews and collections see e.g . Additionally, macroscopic models typically ignore factors like pH, pCO 2 , or osmolality despite their effect, e.g., on the growth of CHO cells . Also amino acid concentrations or other unknown factors can differ with possibly high effects .…”

Scale-down of CHO cell fed-batch cultures

“…The study showed that the temporary pH zone has a strong impact on maximum viable cell density and final product titer. Unlike the previous study done by Nienow et al 171 , in the study of Brunner, M. et al 213 , the use of peristaltic pumps did not show an effect on performance of the applied cell line. In conclusion, only a few experiments have been performed to understand the effects of gradient and poor mixing conditions in mammalian cell culture bioreactors.…”

“…Furthermore, the oscillating DO resulted in a higher amount of triantennary and sialylated glycans, which may affect the quality of the mAb. Recently, Brunner, M. et al 213 studied the impact of pH inhomogeneity on CHO cell fed-batch process performance using a two-compartment (a 3 L STR and a 0.7 L STR) scale-down modelling. Their study simulated a well-mixed zone of pH 7.0 in the 3 L STR and a temporary pH zone of pH 9.0 in the 0.7 L STR, connected by recirculation peristaltic pumps.…”