Da Silva Damas Pinto, Ana Catarina; (2016) Evaluating the response of mammalian cells to pH perturbations using a parallel microwell approach. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Currently, the production of mAbs in mammalian cells plays a significant part in the biopharmaceutical industry. Consequently, the increased market has led to a need for increased production volumes and a drive to improve process efficiency. One of the key parameters that has a significant impact on the cell growth and metabolism, mAb productivity and quality is the pH. The understanding of the response of mammalian cells to pH perturbations is critical. Therefore, a scale-down model of the pH effects on an industrial GSCHO cell line (CHO-CY01) growth and mAb (IgG4) production was developed. A scale translation criteria of matching mixing times (< 5 ) and energy dissipation rates (10 −3 ) was found to be more reliable to set the microwell plates (24-SRW MTPs) and shake flasks as a scale-down models of a 5L-STR bioreactor CHO-CY01 cell line bath and fed-batch cultures. A high-throughput online multi-parametric analysis method of mammalian cell cellular activity at 24-SRW MTP was developed, where the Presens 24-SRW MTPs were exclusively used for the online monitoring of the process parameter. Therefore, it was possible to evaluate the effects of different 2 and pH profiles created with 2/pH shifts on the CHO-CY01 cell line kinetics. A culture pH higher than 7.50 caused an excessive lactate production. While, culture pHs between pH 6.70 - 6.80 inhibited the cell growth and lactate production, pH 6.60 – 6.70 caused an increase in ammonium production and pH < 6.60 triggered cell apoptosis. Therefore, it became important to identify the lactate and ammonium inhibition kinetics. It was found that an initial lactate concentration of 4.08 −1 and an initial ammonium concentration of 18.1 −1 reduced to half the specific growth rate. Ultimately, the optimal pH for IgG4 production was at pH 6.85 ± 0.02, where the maximum specific antibody production rate (10.50 10−9 −1 −1 ) with the minimal lactate production was reached.

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