3D bioprinted microenvironments shape growth and IgG1 monoclonal antibody production of CHO DG44 cells

Abstract

Three-dimensional (3D) bioprinting offers new possibilities to recreate in vivo-like microenvironments for mammalian cells, with potential applications in therapeutic bioproduction. In this study, we evaluate the growth, central carbon metabolism, and IgG1 monoclonal antibody (MAb) bioproduction of CHO DG44 cells embedded in 3D bioprinted hydrogel constructs under different cultivation strategies: batch, semi-continuous, and both modes preceded by a 3D amplification step in amplification medium. Across all 3D conditions, CHO DG44 cells exhibited markedly reduced and limited growth over 14 days, most likely due to restricted nutrient diffusion and limited available space imposed by the intrinsic porosity of the bioink. Glucose was not depleted in any condition. In semi-continuous cultures, glucose consumption and lactate production remained approximately constant over the full culture duration, whereas both processes stopped mid-run in batch cultures. The relatively low overall glucose consumption indicates that higher cell-to-medium ratios could be implemented to improve nutrient utilization. MAb production closely followed lactate metabolism, with sustained production in semi-continuous cultures and an early plateau in batch mode, resulting in 2-3-fold lower final mAb amounts in batch compared with semi-continuous cultures. Introducing a 3D amplification phase more than doubled cumulative mAb production after 14 days. Overall, low growth, low titers, low space–time yields, and residual nutrients in the supernatant show that the current 3D bioprinting strategy is not yet optimal for mAb manufacturing. However, despite this apparent weakness of the 3D bioprinted environment for production, 3D specific productivity of 3D Amplified Semi-continuous conditions was found to be higher than the global suspension specific productivity. Consequently, our data suggest that increasing the cell-to-medium ratio and reducing construct size to mitigate diffusion limitations could enhance cell-specific productivity and process efficiency in 3D bioprinted CHO-based bioproduction systems.