Enhanced antibody production following intermediate addition based on flux analysis in mammalian cell continuous culture

Abstract

Generally, mammalian cells utilize glucose and glutamine as primary energy sources. To investigate the effect of energy sources on metabolic fluxes and antibody production, glucose- or glutamine-limited serum-free continuous culture of hybridoma 3A21 cells, which produce anti-ribonuclease A antibody, was carried out. The cell volume and dry cell weight were evaluated under various steady-state conditions. The specific consumption and production rates were evaluated on the basis of dry cell weight. On the basis of these results, the fluxes of the metabolic pathway were calculated. It was found that increasing the specific growth rate causes the specific ATP and antibody production rates to decrease. The fluxes between malate and pyruvate also decreased with the increase in specific growth rate. To increase the ATP production rate under steady-state conditions by the enhancement of fluxes between malate and pyruvate, the reduced metabolic fluxes were increased by an intermediate (pyruvate, malate, and citrate) addition. As a result, higher specific ATP and antibody production rates were achieved following the intermediate addition at a constant dilution rate.

Quantitative characterization of metabolism and metabolic shifts during growth of the new human cell line AGE1.HN using time resolved metabolic flux analysis

“…Pyruvate represents an extracellular metabolite that was often neglected in other studies [26, 28]. However, as shown in this study and in MDCK cells [23, 49] or CHO cells [50], it is a very interesting metabolite that can induce huge changes in the metabolism. In AGE1.HN cells pyruvate uptake rate was by far lower than in MDCK cells that were grown in high pyruvate medium (Table 2, M2).…”

Enhancement of antibody production by the addition of Coenzyme-Q10

“…To illustrate the potential effectiveness of such enhancement, a 20% increase in SPR could reduce the annual cost ($65 million) of obtaining 250 kg of MAbs with a yield of 1,000 mg/ L in a 10,000 L plant (Werner 2004) by $6 million. In fact, increases of 10%-20% in SPR of various MAbs have already been reported (Arden et al 2007;Kim et al 2005;Omasa et al 2010). …”

Enhancement of sialylation on humanized IgG-like bispecific antibody by overexpression of α2,6-sialyltransferase derived from Chinese hamster ovary cells

“…The cell line was maintained in RDF media [a mixture of RPMI1640, Dulbecco's Modified Eagle Medium, and HamF12 media (Nissui, Tokyo, Japan) containing 10% dialyzed fetal bovine serum (FBS) (JRH Biosciences, Kansas, USA); Omasa et al 2010b]. The media was supplemented with 1,000 nM methotrexate (MTX) and 1 mM G418.…”