Process for the continuous culture of cells

Details Process for the continuous culture of cells Process for the continuous culture of cells

1998-07-10

2000-12-05

Lankford, Jr., Leon B.

Chemistry: molecular biology and microbiology

Animal cell, per se ; composition thereof; process of...

Method of regulating cell metabolism or physiology

435 711, 435139, 4352891, 435325, 435383, C12N 500

Patent

active

061565709

DESCRIPTION:

BRIEF SUMMARY
BACKGROUND OF THE INVENTION

The past fifteen years witnessed the transformation of animal cell culture from an exploratory protein production method to a mature manufacturing process. This technological transformation was aided by significant advances in bioprocessing research and development. As the technology becomes more mature the pressure on driving the production cost down is mounting.
Mammalian cells are capable of utilizing different proportions of glucose and glutamine and metabolize them differently under different culture conditions. Early work by Zielke et. al. (J. Cell. Physiol., 95, 41-48 (1978) showed that glutamine becomes a predominant source of energy at low glucose concentrations. The metabolism of glucose is also greatly affected by glucose concentration in the medium (W.-S. Hu et al., Dev. Biol. Standard., 66, 279-290 (1987); W.-S. Hu et al. in: Large Scale Mammalian Cell Culture Technology, Ed. A. S. Lubineicky, pp. 451-481, Marcel Dekker, Inc., New York, N.Y. (1990)). At high glucose levels, the consumption rate of glucose is higher, although most of the glucose consumed (80-90%) is converted to lactate; at low glucose concentrations, the consumption rate for glucose is lower and a larger proportion is completely oxidized to CO.sub.2.
This tendency of mammalian cells to metabolize glucose to lactate greatly restricts the cell concentration achievable in the bioreactor. Accumulation of lactate in the medium is detrimental to cell growth and is one of the factors that limits the maximum cell concentration that can be achieved in batch culture. In a typical batch cell culture, growth is inhibited after lactate concentration in the culture reaches approximately 30-50 mM and/or ammonia concentration reaches 3-5 mM. As a result, cell concentration in a batch culture is typically in the range of 1 to 3.times.10.sup.6 cells/ml.
Accumulation of lactate can be reduced by changing from a batch mode into a simple continuous operation, thereby continuously removing the spent culture medium containing lactate and cells. Eventually the concentrations of cells, medium components, and metabolites all stabilize, reaching a steady state. However, using the same medium and resulting steady state cell concentration is, at most, about the same level as that reached in batch mode. Cell concentrations usually cannot be much improved by merely increasing the nutrient concentration in the feed, because an increase in nutrient levels in the feed is generally followed by increased lactate and ammonia production and accumulation, thereby preventing the desired increase in cell concentration.
Perfusion cultures have also been used in attempts to achieve high cell concentrations in a bioreactor, and productivity is significantly higher than in conventional batch cultures. However, the large through-put requirement makes medium preparation for perfusion cultures a daunting task. In a perfusion culture, medium is perfused through the reactor at a high rate while cells are retained or recycled back into the reactor by sedimentation, centrifugation or filtration. Up to ten reactor volumes of medium is perfused through the bioreactor in a day. The major function of perfusing such a large volume of medium is primarily to remove the metabolites, mainly lactate, from the culture fluid. The large demand for medium, the high flow rates, and the multitudinous technical problems associated with cell retention devices make the perfusion cultures expensive, labor-intensive, and prone to technical difficulties.
Attempts have been made to increase cell concentration in fed-batch cultures by controlling glucose and/or glutamine concentrations in an effort to reduce the accumulation of inhibitory metabolites. See, e.g., Kurokowa et al., Biotechnol. Bioeng., 44, 95-103(1994); G.-S. Oh et al., J. Fermentation Bioeng., 81, 319-336 (1996); A. Sanfeliu et al., Biotechnol. Prog., 12, 209-216 (1996); W. Zhou et al., Biotechnol. Bioeng., 46, 579-587 (1995). However, in most cases, growth rates and maximum cell concentration are still limi

REFERENCES:
patent: 5443968 (1995-08-01), Takazawa et al.
Bibila et al., "Monoclonal Antibody Process Development Using Medium Concentrates," Biotechnol. Prog., 10, 87-96 (1994).
Carlsen et al., "Growth and .alpha.-amylase production by Aspergillus oryzae during continuous cultivations," Journal of Biotechnology, 45, 81-93 (1996).
Europa et al., "Multiple Steady States in Continuous Cultivation of Animal Cells,"Fourth Asia-Pacific Biochemical Engineering Conference, Beijing, China, Oct. 20-23 (1997) (Abstract and Poster (10 pp.)).
Europa et al., "High Density Continuous Culture of Hybridoma Cells," 1997 National AIChE Meeting, Los Angeles, CA, Nov. 16-21 (1997) (Poster only (13 pp.)).
Gambhir et al., "A Strategy for High Cell Density in Continuous Culture of Hybridoma Cells," Abstracts of Papers, American Chemical Society, 213.sup.th ACS National Meeting, San Francisco, CA, Apr. 13-17 (1997) (Abstract No. 130).
Haggstrom et al., "Metabolic Engineering of Animal Cells," Annals of the New York Academy of Sciences, 782, 40-52 (1996).
Hosobuchi et al., "Dynamic Nutrient Feeding in Hybridoma Cell Culture by using a Statistical Analysis," Abstracts of Papers, American Chemical Society, 213.sup.th ACS National Mtg., San Francisco, CA, Apr. 13-17 (1997) (Abstract No. 103).
Hu et al., "Controlling Mammalian Cell Metabolism in Bioreactors," J. Microbiol. Biotechnol., 8, 8-13 (1998).
Hu et al., "Dynamic Medium Feeding to Manipulate Cell Metabolism in High Density Fed-batch and Continuous Cultures of Mammalian Cells (conference abstract)," In Vitro, 33, V-15 (1997).
Hu et al., "Effect of Glucose on the Cultivation of Mammalian Cells," Dev. Biol. Standard., 66, 279-290 (1987).
Hu et al., "Monitoring and Control of Animal Cell Bioreactors: Biochemical Engineering Considerations," Large Scale Mammalian Cell Culture Technology, Ed. A.S. Lubineicki, 451-459, Marcel Dekker, Inc., New York, NY (1990).
Hu, "Needs and Challenges in the Quatitative Descriptions of Animal Cell Culture Process," Abstracts of Papers, American Chemical Society, 213.sup.th ACS National Meeting, San Francisco, CA, Apr. 13-17 (1997) (Abstract No. 060).
Kurokowa et al., "Growth Characteristics in Fed-Batch Culture of Hybridoma Cells with Control of Glucose and Glutamine Concentrations," Biotechnol. Bioeng., 44, 95-103 (1994).
Kurokawa et al., "Kinetic Study of Hybridoma Metabolism and Antibody Production in Continuous Culture Using Serum-Free Medium," J. Fermentation and Bioeng., 76, 128-133 (1993).
Kyung et al., "Enhanced productivity of Protein C by recombinant human cells in automated fed-batch cultures," Cytotechnology, 17, 109-115 (1995).
Linz et al., "Stoichiometry, Kinetics, and Regulation of Glucose and Amino Acid Metabolism of a Recombinant BHK Cell Line in Batch and Continuous Cultures," Biotechnol. Prog., 13, 453-463 (1997).
Ljunggren et al., "Catabolic Control of Hybridoma Cells by Glucose and Glutamine Limited Fed Batch Cultures," Biotechnol. Bioeng., 44, 808-818 (1994).
Oh et al., "Interactive Dual Control of Glucose and Glutamine Feeding in Hybridoma Cultivation," J. Fermentation Bioeng., 81, 329-336 (1996).
Portner et al., "High density fed-batch cultures for hybridoma cells performed with the aid of a kinetic model," Bioprocess Engineering, 15, 117-124 (1996).
Sanfeliu, "Production of Monoclonal Antibodies by In Vitro Hybridoma Cultures in Bioreactors: Analysis of Physiology and Cellular Metabolism (Igg, Fetal Calf Serum)," Dissertation Abstracts International, 58/02, 457 (1995) (Abstract only).
Sanfeliu et al., "Analysis of Nutritional Factors and Physical Conditions Affecting Growth and Monoclonal Antibody Production of the Hybridoma KB-26.5 Cell Line," Biotechnol. Prog., 12, 209-216 (1996).
Snay et al., "Effects of Growth Conditions on Carbon Utilization and Organic By-Product Formation in B. subtilis", Biotechnology Progress, 5, 63-69 (1989).
Wang et al., Fermentation and Enzyme Technology, p. 79, John Wiley & Sons (1979).
Xie et al., "Fed-Batch Cultivation of Animal Cells Using Di

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