Development of a chemically defined medium for optimal growth and recombinant protein production in HEK293 cells

The human cell line Human Embryonic Kidney (HEK) 293 is a popular workhorse for transient gene expression. Due to stringent regulatory requirement, better batch-to-batch consistency and simplified downstream processing, the demand for a chemically defined (CD) medium to support the growth of these cells is higher than ever. To address this need, Kerry has developed an animal component-free (ACF), chemically defined (CD) cell culture medium, to support the growth of HEK293 cells in suspension culture. AmpliHEK CD medium was designed from the knowledge gained in developing a medium for Chinese Hamster Ovary (CHO) cells and was refined by analysis of spent medium after growth of HEK293 cells in shake flasks to determine the nutrients needed for optimal growth. AmpliHEK is a versatile medium providing optimal cell culture conditions for the production of recombinant proteins, monoclonal antibodies and viral vectors. AmpliHEK medium formulation shows similar growth profile and viability compared to competitor CD media during and after sequential adaption. The medium also showed prolonged culture longevity and higher peak cell density in a batch culture (with glucose feeding), which is important for a higher production yield. The efficacy of this medium to support the expression of recombinant β-galactosidase in transient transfection was tested and found to be comparable with competitor CD media. Currently, the versatility of this medium using different HEK293 host cells is being tested. The effect of CD and complex feeds containing plant-based hydrolysates on transient gene expression was another scope of this study. In all, these results show that this medium could be used for industrial production with HEK293 cells

Implications of feeds and supplements on the productivity and quality of recombinant proteins produced in CHO cells

Chinese Hamster Ovary (CHO) cells have become the preferred host for recombinant protein production due to their ability to secrete desired proteins with post-translational modifications similar to those observed in humans. Biopharmaceutical companies routinely utilize supplements and feed systems for maximizing the yield of recombinant proteins in processes involving CHO cells. Although high productivity is desirable for monoclonal antibodies (mAbs) production, utilizing supplements and feeds might affect the quality of recombinant proteins. Clinical efficacy and safety of recombinant proteins is dependent on key quality attributes including glycosylation pattern, aggregates, charge variants and low molecular weight species. Our work outlines the comparison of productivity and quality of recombinant proteins produced in fed-batch CHO-cell based processes using commercially available chemically defined (CD) medium for CHO cells, CD feeds and a variety of complex supplements and feed systems. The CD medium, feeds and supplements were tested in three CHO cell lines, each expressing a different IgG molecule. Each cell line was sequentially adapted to all the different CD medium being evaluated. Multiple fed-batch experiments were performed in shake-flasks with combinations of different commercially available supplements and feeds. At regular intervals, samples were assessed for viable cell density, cell viability, nutrient metabolism and IgG titers for the different conditions. The key IgG quality attributes including glycosylation pattern, aggregates, charge variants, low molecular weight species were also evaluated for the samples obtained at the end of the fed-batch process. The medium/feed combinations which demonstrated high protein productivity and high cell viability at the end of the culture process were further tested in bioreactors to evaluate scalability of the medium/feed combinations in the different cell lines. The use of CD medium with either CD feed or complex feed supplements resulted in higher cell viability at the end of the fed-batch process in addition to higher IgG titers. After evaluation of the product quality, the desired glycosylation pattern was obtained in certain combinations of medium, supplements and feeds. Lower amount of IgG aggregation was also observed. Due to the unique nutritional requirements of each cell line, different combinations of medium/supplement/feed were needed for optimal cell growth and productivity without affecting the product quality