Designed Amino Acid Feed in Improvement of Production and Quality Targets of a Therapeutic Monoclonal Antibody

Abstract

Cell culture feeds optimization is a critical step in process development of pharmaceutical recombinant protein production. Amino acids are the basic supplements of mammalian cell culture feeds with known effect on their growth promotion and productivity. In this study, we reported the implementation of the Plackett-Burman (PB) multifactorial design to screen the effects of amino acids on the growth promotion and productivity of a Chinese hamster ovary DG-44 (CHO-DG44) cell line producing bevacizumab. After this screening, the amino acid combinations were optimized by the response surface methodology (RSM) to determine the most effective concentration in feeds. Through this strategy, the final monoclonal antibody (mAb) titre was enhanced by 70%, compared to the control group. For this particular cell line, aspartic acid, glutamic acid, arginine and glycine had the highest positive effects on the final mAb titre. Simultaneously, the impact of the designed amino acid feed on some critical quality attributes of bevacizumab was examined in the group with highest productivity. The product was analysed for N-glycan profiles, charge variant distribution, and low molecular weight forms. The results showed that the target product quality has been improved using this feeding strategy. It was shown how this strategy could significantly diminish the time and number of experiments in identifying the most effective amino acids and related concentrations in target product enhancement. This model could be successfully applied to other components of culture media and feeds.

Cell culture media for recombinant protein expression in Chinese hamster ovary (CHO) cells: History, key components, and optimization strategies

“…In a more comprehensive study, Torkashvand et al developed a workflow utilizing several Plackett‐Burman experiments to optimize amino acid concentration in a cell culture medium by identifying key amino acids influencing cell growth and titers, and analyzing binary effects of those amino acids with response surface methodology. Using this approach, titer was increased by 70% without altering product quality . In addition to enhancing titer and peak cell density, selected amino acids at certain concentrations may have protective effects for cells growing in bioreactors.…”

“…Using this approach, titer was increased by 70% without altering product quality. 67 In addition to enhancing titer and peak cell density, selected amino acids at certain concentrations may have protective effects for cells growing in bioreactors. Certain amino acids have been shown to eliminate or alleviate some of the negative effects of ammonium and pCO 2 accumulation, as well as high osmolality.…”

Automatic real‐time calibration, assessment, and maintenance of generic Raman models for online monitoring of cell culture processes

“…Stable production of biotherapeutic proteins from modified cell lines requires real‐time monitoring and control of cell culture processes to ensure optimal cell growth profiles and consistent product quality (Fan et al, ; Kim, Kim, & Lee, ; Torkashvand et al, ). Barring few real‐time measurements, such as pH, dissolved oxygen (DO), and temperature, much of the cell culture parameters, such as metabolite and product concentrations, cell growth, and other product quality attributes are monitored using offline methods (Santos, Kessler, Salou, Menezes, & Peinado, ).…”

Real‐time quantification and supplementation of bioreactor amino acids to prolong culture time and maintain antibody product quality

“…Specifically, we are studying the possibility that replenishment of selected amino acids could help maintain the product quality of the bioreactor cell culture. Altered supplementation of amino acids in cell culture can modify the critical quality attributes of the antibody, including glycosylation . These changes are not limited to antibody products: an increase of recombinant human erythropoietin with decreased sialylation has been attributed to the addition of amino acids as feeds during cell culture .…”

“…Altered supplementation of amino acids in cell culture can modify the critical quality attributes of the antibody, including glycosylation. 15 These changes are not limited to antibody products: an increase of recombinant human erythropoietin with decreased sialylation has been attributed to the addition of amino acids as feeds during cell culture. 16 In a study of tissue plasminogen activator produced in ammonium-stressed CHO cells, the addition of amino acids leads to increased glycosylation site occupancy by mature glycoforms.…”