Chinese hamster ovary cell growth and interferon production kinetics in stirred batch culture

Abstract

Recombinant human interferon-gamma production by Chinese hamster ovary cells was restricted to the growth phase of batch cultures in serum-free medium. The specific interferon production rate was highest during the initial period of exponential growth but declined subsequently in parallel with specific growth rate. This decline in specific growth rate and interferon productivity was associated with a decline in specific metabolic activity as determined by the rate of glucose uptake and the rates of lactate and ammonia production. The ammonia and lactate concentrations that had accumulated by the end of the batch culture were not inhibitory to growth. Glucose was exhausted by the end of the growth phase but increased glucose concentrations did not improve the cell yield or interferon production kinetics. Analysis of amino acid metabolism showed that glutamine and asparagine were exhausted by the end of the growth phase, but supplementation of these amino acids did not improve either cell or product yields. When glutamine was omitted from the growth medium there was no cell proliferation but interferon production occurred, suggesting that recombinant protein production can be uncoupled from cell proliferation.

Specific nutrient supplementation of defined serum-free medium for the improvement of CHO cells growth and t-PA production

“…Alanine, which usually accumulates during cultivation, competitively inhibits proline intake, therefore impairing CHO cell growth (Curriden and Englesberg, 1981). Alanine at 3 mM was reported inhibitory for growth of CHO-γ cells (Hayter et al 1991). This might have occurred in C4, where proline consumption rate decreased drastically over 2.4 mM alanine (data not shown).…”

Specific heat flow rate: An on-line monitor and potential control variable of specific metabolic rate in animal cell culture that combines microcalorimetry with dielectric spectroscopy

“…Amplification was made by selection with 0.1 M methotrexate. The cells were grown in an RPMI 1640 (Sigma-Aldrich, Poole, England; or Life Technologies, Paisley, Scotland) defined medium with the following supplements added: 2 mM Lglutamine, 24 mM sodium bicarbonate, 5 g dm −3 BSA free of fatty acids (PENTEX bovine albumin, Miles, Inc., Kankakee, IL), 5 mg dm −3 bovine insulin, 5 mg dm −3 hu-man transferrin, 1 mM sodium pyruvate, 0.1 mM L-alanine, 1 M putrescine, 3 M FeSO 4 , 3 M ZnSO 4 , 10 nM Na 2 SeO 4 , 10 nM CuSO 4 , and 50 mg dm −3 gentamycin (all from Sigma) (Hayter et al, 1991). Bovine serum albumin free of fatty acids was used to avoid the possible significant complications in enthalpy balance studies of fatty acid utilization (Kemp and Guan, 1998).…”

“…The experiments were performed using suspension-adapted CHO 320 cells genetically engineered to produce IFN-␥ (Hayter et al, 1991) after co-transfection of dhfr and human IFN-␥ genes in a plasmid derived from a pSV2-dhfr vector controlled by an SV-40 early promotor. Amplification was made by selection with 0.1 M methotrexate.…”

“…The generallyapplicable procedure is illustrated using batch cultures of recombinant Chinese hamster ovary (CHO) cells which grew in a stirred tank and produced the recombinant glycoprotein, human interferon-␥ (IFN-␥) (Hayter et al, 1991). Changes in specific heat flow rate are related to the specific rates of growth and of individual metabolites including the target protein, IFN-␥.…”

Plant protein hydrolysates support CHO-320 cells proliferation and recombinant IFN-γ production in suspension and inside microcarriers in protein-free media

“…This effect was related to an increase in the cell density, but mostly to a direct stimulation of the IFN-c specific production, which reached about 0.65 lg 10 À6 cells in PFS-WP to be compared to 0.5 lg 10 À6 cells in PFS. The IFN-c production by CHO-320 cells has already been described to be at least partly growth-associated (Hayter et al 1993;Leelavatcharamas et al 1999), even if this is sometimes unclear (Hayter et al 1991;Castro et al, 1992;Goldman et al 1998).…”