Modulation of nutrient precursors for controlling metabolic inhibitors by genome‐scale flux balance analysis

Therapeutic protein productivity and glycosylation pattern highly rely on cell metabolism. Cell culture medium composition and feeding strategy are critical to regulate cell metabolism. In this study, the relationship between toxic metabolic inhibitors and their nutrient precursors was explored to identify the critical medium components toward cell growth and generation of metabolic by‐products. Generic CHO metabolic model was tailored and integrated with CHO fed‐batch metabolomic data to obtain a cell line‐ and process‐specific model. Flux balance analysis study was conducted on toxic metabolites cytidine monophosphate, guanosine monophosphate and n‐acetylputrescine—all of which were previously reported to generate from endogenous cell metabolism—by mapping them to a compartmentalized carbon utilization network. Using this approach, the study projected high level of inhibitory metabolites accumulation when comparing three industrially relevant fed‐batch feeding conditions one against another, from which the results were validated via a dose‐dependent amino acids spiking study. In the end, a medium optimization design was employed to lower the amount of supplemented nutrients, of which improvements in critical process performance were realized at 40% increase in peak viable cell density (VCD), 15% increase in integral VCD, and 37% increase in growth rate. Tight control of toxic by‐products was also achieved, as the study measured decreased inhibitory metabolites accumulation across all conditions. Overall, the study successfully presented a digital twin approach to investigate the intertwined relationship between supplemented medium constituents and downstream toxic metabolites generated through host cell metabolism, further elucidating different control strategies capable of improving cellular phenotypes and regulating toxic inhibitors.