Effect of different cell culture media on the production and glycosylation of a monoclonal antibody from a CHO cell line

Abstract

Recombinant monoclonal antibodies (mAbs) are commonly produced using Chinese hamster ovary (CHO) cells and the cell culture medium used in bioreactors influences the yield and quality attributes of the protein drug products. The COVID 19 pandemic revealed a vulnerability in the supply chain for necessary reagents (such as culture medium and raw material) for maintaining un-interrupted production of protein drugs with consistent quality. The supply interruption for the cell culture medium ActiPro™ optimized for producing VRC01, an IgG1-κ mAb, from a CHO-K1 cell line, necessitated the search for alternate media. VRC01 mAb is highly glycosylated and can broadly neutralize several strains of Human Immunodeficiency Virus (HIV). We investigated to see if an alternate medium can be used in the production without impacting quality attributes like glycosylation. In our strategy, we used 3 different commercially available media, performed two sets of experiments-with and without media supplements, Cell boost 7a and Cell boost 7b. Cell growth, volumetric production of the mAb protein and glycosylation pattern were compared to identify an alternative medium. Among the tested media based on cell growth, mAb production potential and glycosylation analysis, ActiCHO™ P was found to be a better alternate medium to ActiPro™ medium than EX-CELL® 325 PF CHO medium to produce VRC01 mAb. Overall, the approach used here to establish the impact of variation in medium on protein therapeutic attributes may be used during product development to build in supply chain resilience in drug manufacturing.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00733-7.

Keywords: Chemically defined medium; Glycosylation; Monoclonal antibody (mAb); N-glycans; Titer.

Fig. 1

VRC01 mAb CHO-K1 cell growth and viability profiles measured during the experiments. Viable cell density for each medium is given without the additives (a) and with the additives (b). Corresponding viability (%) profiles are given in panel (c) for without additives and panel (d) for with additives

Fig. 2

Representative profiles of metabolites in the three cell culture media. Data show the representative cultures from group 1 (absence of Cell boost additives, − CB) and group 2 (presence of Cell boost additives, + CB) for the duration of VRC01 CHO-K1 cell culture. Cell boost 7a and Cell boost 7b were used as additives. Metabolite concentrations were determined using Nova BioProfile Flex 2 instrument. Legend shown in A & D is common to all graphs

Fig. 3

VRC01 CHO-K1 cell growth and mAb production in three media. Profiles of production and viable cell density are given for EX-CELL® 325 PF CHO (a and b), ActiCHO™ P (c and d) and ActiPro™ media (e and f). The legend shown in (a) is common to all other panels. In panel c the last three datapoints of + CB group 2 (open triangles) were outside the standard curve range and re-measured by diluting the samples. This might have contributed to a little over estimation

Fig. 4

Relative abundance (% of total) of major glycan types quantified from the VRC01 mAb by HPLC analysis. The effect of Cell boost 7a and Cell boost 7b supplementation (+ CB; group 2) of the VRC01 CHO cells on various glycan type and content of VRC01 mAb was statistically compared to the un-supplemented (− CB; group 1). The significance was determined (p < 0.05) by a Dunnett’s test and only significant differences are indicated on the bars (n = 6). JMP 17.0.0 was used for statistical analysis