Skip to main content

Industrial Production of Therapeutic Proteins: Cell Lines, Cell Culture, and Purification

  • Chapter
Kent and Riegel’s Handbook of Industrial Chemistry and Biotechnology

Abstract

The biotechnology and pharmaceutical industries have seen a recent surge in the development of biological drug products manufactured from engineered mammalian cell lines. Since the hugely successful launch of human tissue plasminogen activator in 1987 and erythropoietin in 1988, the biopharmaceutical market has grown immensely. Global sales in 2003 exceeded US $30 billion.1 Currently, a total of 108 biotherapeutics are approved and available to patients (Table 32.1). In addition, 324 medically related, biotechnology-derived medicines for nearly 150 diseases are in clinical trials or under review by the U.S. Food and Drug Administration.2 These biopharmaceutical candidates promise to bring more and better treatments to patients. Compared to small molecule drugs, biotherapeutics show exquisite specificity with fewer off-target interactions and improved safety profiles.

Approved Biotechnology Products

Michael Mollet is working at Medimmune now

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 249.00
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Walsh, G. 2003. “Biopharmaceutical benchmarks—2003.” Nat Biotechnol 21:865–870.

    Article  CAS  Google Scholar 

  2. PhRMA. 2004. 324 biotechnology medicines in testing promise to bolster the arsenal against disease. New medicines in development. Oct 25, 2004. Available at: http://www.phrma.org/newmedicines/biotech/.

    Google Scholar 

  3. Graddis, T.J., R.L. Remmele, Jr., and J.T. McGrew. 2002. “Designing proteins that work using recombinant technologies.” Curr Pharm Biotechnol 3:285–297.

    Article  CAS  Google Scholar 

  4. Brekke, O.H., and G.A. Loset. 2003. “New technologies in therapeutic antibody development.” Curr Opin Pharmacol 3:544–550.

    Article  CAS  Google Scholar 

  5. Lazar, G.A., S.A. Marshall, J.J. Plecs, S.L. Mayo, and J.R. Desjarlais. 2003. “Designing proteins for therapeutic applications.” Curr Opin Struct Biol 13:513–518.

    Article  CAS  Google Scholar 

  6. Marshall, S.A., G.A. Lazar, A.J. Chirino, and J.R. Desjarlais. 2003. “Rational design and engineering of therapeutic proteins.” Drug Discov Today 8:212–221.

    Article  CAS  Google Scholar 

  7. Vasserot, A.P., C.D. Dickinson, Y. Tang, W.D. Huse, K.S. Manchester, and J.D. Watkins. 2003. “Optimization of protein therapeutics by directed evolution.” Drug Discov Today 8:118–126.

    Article  CAS  Google Scholar 

  8. Wurm, F.M. 2004. “Production of recombinant protein therapeutics in cultivated mammalian cells.” Nat Biotechnol 22:1393–1398.

    Article  CAS  Google Scholar 

  9. Arden, N., T. Nivtchanyong, and M.J. Betenbaugh. 2004. “Cell engineering blocks stress and improves biotherapeutic production.” Bioprocessing 3:23–28.

    Google Scholar 

  10. Running Deer, J., and D.S. Allison. 2004. “High-level expression of proteins in mammalian cells using transcription regulatory sequences from the Chinese hamster EF-1 alpha gene.” Biotechnol Prog 20:880–889.

    Article  Google Scholar 

  11. Brinster, R.L., J.M. Allen, R.R. Behringer, R.E. Gelinas, and R.D. Palmiter. 1988. “Introns increase transcriptional efficiency in transgenic mice.” Proc Natl Acad Sci U S A 85:836–840.

    Article  CAS  Google Scholar 

  12. Palmiter, R.D., E.P. Sandgren, M.R. Avarbock, D.D. Allen, and R.L. Brinster. 1991. “Heterologous introns can enhance expression of transgenes in mice.” Proc Natl Acad Sci U S A 88:478–182.

    Article  CAS  Google Scholar 

  13. Pestova, T.V., I.B. Lomakin, and C.U. Hellen. 2004. “Position of the CrPV IRES on the 40S subunit and factor dependence of IRES/80S ribosome assembly.” EMBO Rep 5:906–913.

    Article  CAS  Google Scholar 

  14. Zahn-Zabal, M., M. Kobr, P.A. Girod, M. Imhof, P. Chatellard, M. de Jesus, F. Wurm, and N. Mermod. 2001. “Development of stable cell lines for production or regulated expression using matrix attachment regions.” J Biotechnol 87:29–42.

    Article  CAS  Google Scholar 

  15. Kim, J.M., J.S. Kim, D.H. Park, H.S. Kang, J. Yoon, K. Baek, and Y. Yoon. 2004. “Improved recombinant gene expression in CHO cells using matrix attachment regions.” J Biotechnol 107:95–105.

    Article  CAS  Google Scholar 

  16. Harland, L., R. Crombie, S. Anson, J. deBoer, P.A. Ioannou, and M. Antoniou. 2002. “Transcriptional regulation of the human TATA binding protein gene.” Genomics 79:479–482.

    Article  CAS  Google Scholar 

  17. Wong, T.K., and E. Newmann. 1982. “Electric-field mediated gene-transfer.” Biochem Biophys Res Commun 107:584–587.

    Article  CAS  Google Scholar 

  18. Kichler, A. 2004. “Gene transfer with modified polyethylenimines.” J Gene Med 6Suppl 1:S3–10.

    Article  CAS  Google Scholar 

  19. Cockett, M.I., C.R. Bebbington, and G.T. Yarranton. 1990. “High level expression of tissue inhibitor of metalloproteinases in Chinese hamster ovary cells using glutamine synthetase gene amplification.” Biotechnol (NY) 8:662–667.

    Article  CAS  Google Scholar 

  20. Lucas, B.K., L.M. Giere, R.A. DeMarco, A. Shen, V. Chisholm, and C.W. Crowley. 1996. “High-level production of recombinant proteins in CHO cells using a dicistronic DHFR intron expression vector.” Nucleic Acids Res 24:1774–1779.

    Article  CAS  Google Scholar 

  21. Kim, S.J., and G.M. Lee. 1999. “Cytogenetic analysis of chimeric antibody-producing CHO cells in the course of dihydrofolate reductase-mediated gene amplification and their stability in the absence of selective pressure.” Biotechnol Bioeng 64:741–749.

    Article  CAS  Google Scholar 

  22. Fann, C.H., F. Guirgis, G. Chen, M.S. Lao, and J.M. Piret. 2000. “Limitations to the amplification and stability of human tissue-type plasminogen activator expression by Chinese hamster ovary cells.” Biotechnol Bioeng 69:204–212.

    Article  CAS  Google Scholar 

  23. Yoshikawa, T., F. Nakanishi, Y. Ogura, D. Oi, T. Omasa, Y. Katakura, M. Kishimoto, and K. Suga. 2000. “Amplified gene location in chromosomal DNA affected recombinant protein production and stability of amplified genes.” Biotechnol Prog 16:710–715.

    Article  CAS  Google Scholar 

  24. Brezinsky, S.C., G.G. Chiang, A. Szilvasi, S. Mohan, R.I. Shapiro, A. MacLean, W. Sisk, and G. Thill. 2003. “A simple method for enriching populations of transfected CHO cells for cells of higher specific productivity.” J Immunol Methods 277:141–155.

    Article  CAS  Google Scholar 

  25. Ham, R.G. 1981. “Tissue growth factors.” In Handbook of Experimental Pharmacology. R. Baserga, ed., Springer Verlag, New York, p. 13.

    Google Scholar 

  26. Sato, G.H., A. Pardee, and D.A. Sirbasku, editors. 1982. Growth of Cells in Hormonally Defined Media, Cold Spring Harbor Conference on Cell Proliferation. Vol. 9, Cold Spring Harbor Press, Cold Spring Harbor, NY.

    Google Scholar 

  27. Fletcher, T. 2005. “Designing culture media for recombinant protein production: A rational approach.” BioProcess Int 3:30–36.

    CAS  Google Scholar 

  28. Lee, G.M., E.J. Kim, N.S. Kim, S.K. Yoon, Y.H. Ahn, and J.Y. Song. 1999. “Development of a serum-free medium for the production of erythropoietin by suspension culture of recombinant Chinese hamster ovary cells using a statistical design.” J Biotechnol 69:85–93.

    Article  CAS  Google Scholar 

  29. Liu, C., I. Chu, and S. Hwang. 2001. “Factorial designs combined with the steepest ascent method to optimize serum-free media for CHO cells.” Enzyme Microb Technol 28:314–321.

    Article  CAS  Google Scholar 

  30. Chun, C., K. Heineken, D. Szeto, T. Ryll, S. Chamow, and J.D. Chung. 2003. “Application of factorial design to accelerate identification of CHO growth factor requirements.” Biotechnol Prog 19:52–57.

    Article  CAS  Google Scholar 

  31. Allison, D.W., K.A. Aboytes, D.K. Fong, S.L. Leugers, T.K. Johnson, H.N. Loke, and L.M. Donahue. 2005. “Development and optimization of cell culture media: Genomic and proteomic approaches.” BioProcess Int 31:38–15.

    Google Scholar 

  32. Zhu, M.M., E.S. Lee, W.R. Hermans, and D.J. Wasilko. 2001. “Overview and serum-free medium development for mammalian cell culture.” Fourth Conference on Recent Advances in Fermentation Technology (RAFTIV), Nov 11–13, Long Beach, CA.

    Google Scholar 

  33. Varley, J., and J. Birch. 1999. “Reactor design for large scale suspension animal cell culture.” Cytotechnol 29:177–205.

    Article  CAS  Google Scholar 

  34. Ozturk, S.S. 1996. “Engineering challenges in high density cell culture systems.” Cytotechnol 22:3–16.

    Article  Google Scholar 

  35. Myers, K.J., M.F. Reeder, and J.B. Fasano. 2002. “Optimize mixing by using the proper baffles.” Chem Eng Progress 98:42–47.

    CAS  Google Scholar 

  36. Chisti, Y. 1993. “Animal cell culture in stirred bioreactors: Observations on scale-up.” Bioprocess Eng 9:191–196.

    Article  CAS  Google Scholar 

  37. Pattison, R.N., J. Swamy, B. Mendenhall, C. Hwang, and B.T. Frohlich. 2000. “Measurement and control of dissolved carbon dioxide in mammalian cell culture processes using an in situ fiber optic chemical sensor.” Biotechnol Prog 16:769–774.

    Article  CAS  Google Scholar 

  38. Dowd, J.E., A. Jubb, K.E. Kwok, and J.M. Piret. 2003. “Optimization and control of perfusion cultures using a viable cell probe and cell specific perfusion rates.” Cytotechnol 42:35–45.

    Article  CAS  Google Scholar 

  39. Noll, T., and M. Biselli. 1998. “Dielectric spectroscopy in the cultivation of suspended and immobilized hybridoma cells.” J Biotechnol 63:187–198.

    Article  CAS  Google Scholar 

  40. Gupta, A., and G. Rao. 2003. “A study of oxygen transfer in shake flasks using a non-invasive oxygen sensor.” Biotechnol Bioeng 84:351–358.

    Article  CAS  Google Scholar 

  41. Ge, X., Y. Kostov, and G. Rao. 2005. “Low-cost noninvasive optical CO2 sensing system for fermentation and cell culture.” Biotechnol Bioeng 89:329–334.

    Article  CAS  Google Scholar 

  42. Singh, V. 1999. “Disposable bioreactor for cell culture using wave-induced agitation.” Cytotechnol 30:149–158.

    Article  CAS  Google Scholar 

  43. Ozturk, S.S. 2005. “Batch versus perfusion: A real case comparison of highly developed cell culture processes for the production of monoclonal antibodies.” 229th National Meeting American Chemical Society, Mar 13–17, San Diego, CA.

    Google Scholar 

  44. Voisard, D., F. Meuwly, P.A. Ruffieux, G. Baer, and A. Kadouri. 2003. “Potential of cell retention techniques for large-scale high-density perfusion culture of suspended mammalian cells.” Biotechnol Bioeng 82:751–765.

    Article  CAS  Google Scholar 

  45. Alrubeai, M., R.P Singh, A.N. Emery, and Z. Zhang. 1995. “Cell cycle and cell size dependence of susceptibility to hydrodynamic forces.” Biotechnol Bioeng 46:88–92.

    Article  CAS  Google Scholar 

  46. Alrubeai, M., R.P. Singh, M.H. Goldman, and A.N. Emery. 1995. “Death mechanisms of animal cells in conditions of intensive agitation.” Biotechnol Bioeng 45:463–472.

    Article  CAS  Google Scholar 

  47. Gregoriades, N., J. Clay, N. Ma, K. Koelling, and J.J. Chalmers. 2000. “Cell damage of microcarrier cultures as a function of local energy dissipation created by a rapid extensional flow“ Biotechnol Bioeng 69:171–182.

    Article  CAS  Google Scholar 

  48. Ma, N., K.W. Koelling, and J.J. Chalmers. 2002. “Fabrication and use of a transient contractional flow device to quantify the sensitivity of mammalian and insect cells to hydrodynamic forces.” Biotechnol Bioeng 80:428–437.

    Article  CAS  Google Scholar 

  49. Kunas, K.T., and E.T. Papoutsakis. 1990. “Damage mechanisms of suspended animal cells in agitated bioreactors with and without bubble entrainment.” Biotechnol Bioeng 36:476–483.

    Article  CAS  Google Scholar 

  50. Chattopadhyay, D., J.F. Rathman, and J.J. Chalmers. 1995. “Thermodynamic approach to explain cell adhesion to air-medium interfaces.” Biotechnol Bioeng 48:649–658.

    Article  CAS  Google Scholar 

  51. Ma, N., J.J. Chalmers, J.G. Aunins, W. Zhou, and L. Xie. 2004. “Quantitative studies of cell-bubble interactions and cell damage at different Pluronic F-68 and cell concentrations.” Biotechnol Prog 20:1183–1191.

    Article  CAS  Google Scholar 

  52. Osman, J.J., J. Birch, and J. Varley. 2001. “The response of GS-NSO myeloma cells to pH shifts and pH perturbations.” Biotechnol Bioeng 75:63–73.

    Article  CAS  Google Scholar 

  53. Sauer, P.W., J.E. Burky, M.C. Wesson, H.D. Sternard, and L. Qu. 2000. “A high-yielding, generic fed-batch cell culture process for production of recombinant antibodies.” Biotechnol Bioeng 67:585–597.

    Article  CAS  Google Scholar 

  54. Lao, M.S., and D. Toth. 1997. “Effects of ammonium and lactate on growth and metabolism of a recombinant Chinese hamster ovary cell culture.” Biotechnol Prog 13:688–691.

    Article  CAS  Google Scholar 

  55. Ozturk, S.S., J.C. Thrift, J.D. Blackie, and D. Naveh. 1997. “Real-time monitoring and control of glucose and lactate concentrations in a mammalian cell perfusion reactor.” Biotechnol Bioeng 53:372–378.

    Article  CAS  Google Scholar 

  56. Martinelle, K., A. Westlund, and L. Haggstrom. 1996. “Ammonium ion transport: A cause of cell death.” Cytotechnol 22:251–254.

    Article  CAS  Google Scholar 

  57. Genzel, Y., J.B. Ritter, S. Konig, R. Alt, and U. Reichl. 2005. “Substitution of glutamine by pyruvate to reduce ammonia formation and growth inhibition of mammalian cells.” Biotechnol Prog 21:58–69.

    Article  CAS  Google Scholar 

  58. Yang, M., and M. Butler. 2000. “Effect of ammonia on the glycosylation of human recombinant erythropoietin in culture.” Biotechnol Prog 16:751–759.

    Article  CAS  Google Scholar 

  59. Gray, D.R., S. Chen, W. Howarth, D. Inlow, and B.L. Maiorella. 1996. “CO2 in large-scale and high-density CHO cell perfusion culture.” Cytotechnol 22:65–78.

    Article  CAS  Google Scholar 

  60. deZengotita, V.M., A.E. Schmelzer, and W.M. Miller. 2002. “Characterization of hybridoma cell responses to elevated pCO2 and osmolality: Intracellular pH, cell size, apoptosis, and metabolism.” Biotechnol Bioeng 77:369–380.

    Article  CAS  Google Scholar 

  61. Zhu, M.M., A. Goyal, D.L. Rank, S.K. Gupta, T. Vanden Boom, and S.S. Lee. 2005. “Effects of elevated pCO2 and osmolality on growth of CHO cells and production of antibody-fusion protein Bl: A case study.” Biotechnol Prog 21:70–77.

    Article  Google Scholar 

  62. Mostafa, S.S., and X. Gu. 2003. “Strategies for improved dCO2 removal in large-scale fed-batch cultures.” Biotechnol Prog 19:45–51.

    Article  CAS  Google Scholar 

  63. Chen, Z.L. 2004. “Temperature shift as a process optimization step for the production of pro-urokinase by a recombinant Chinese hamster ovary cell line in high-density perfusion culture.” J Biosci Bioeng 97:239–243.

    CAS  Google Scholar 

  64. Clark, K.J., F.W. Chaplin, and S.W. Harcum. 2004. “Temperature effects on product-quality-related enzymes in batch CHO cell cultures producing recombinant tPA.” Biotechnol Prog 20:1888–1892.

    Article  CAS  Google Scholar 

  65. Fox, S.R., U.A. Patel, M.G. Yap, and D.I. Wang. 2004. “Maximizing interferon-gamma production by Chinese hamster ovary cells through temperature shift optimization: Experimental and modeling.” Biotechnol Bioeng 85:177–184.

    Article  CAS  Google Scholar 

  66. Bollati-Fogolin, M., G. Forno, M. Nimtz, H.S. Conradt, M. Etcheverrigaray, and R. Kratje. 2005. “Temperature reduction in cultures of hGM-CSF-expressing CHO cells: Effect on productivity and product quality.” Biotechnol Prog 21:17–21.

    Article  CAS  Google Scholar 

  67. Dempsey, J., S. Ruddock, M. Osborne, A. Ridley, S. Sturt, and R. Field. 2003. “Improved fermentation processes for NSO cell lines expressing human antibodies and glutamine synthetase.” Biotechnol Prog 19:175–178.

    Article  CAS  Google Scholar 

  68. Wong, D.C.F., K.T.K. Wong, L.T. Goh, C.K. Heng, and M.G.S. Yap. 2005. “Impact of dynamic online fed-batch strategies on metabolism, productivity and N-glycosylation quality in CHO cell cultures.” Biotechnol Bioeng 89:164–177.

    Article  CAS  Google Scholar 

  69. Dowd, J.E., K.E. Kwok, and J.M. Piret. 2000. “Increased t-PA yields using ultrafiltration of an inhibitory product from CHO fed-batch culture.” Biotechnol Prog 16:786–794.

    Article  CAS  Google Scholar 

  70. Dowd, J.E., K.E. Kwok, and J.M. Piret. 2001. “Predictive modeling and loose-loop control for perfusion bioreactors.” Biochem Eng J 9:1–9.

    Article  CAS  Google Scholar 

  71. Nienow, A.W., C. Langheinrich, N.C. Stevenson, A.N. Emery, T.M. Clayton, and N.K.H. Slater. 1996. “Homogenisation and oxygen transfer rates in large agitated and sparged animal cell bioreactors: Some implications for growth and production.” Cytotechnol 22:87–94.

    Article  CAS  Google Scholar 

  72. Gardner, A.R., and T.M. Smith. 2000. “Identification and establishment of operating ranges of critical process variables.” In Biopharmaceutical Process Validation. G. Sofer and D.W. Zabriskie, Ed., Marcel Dekker, New York, pp. 61–76.

    Google Scholar 

  73. Moran, E.B., S.T. McGowan, J.M. McGuire, J.E. Frankland, I.A. Oyebade, W. Waller, L.C. Archer, L.O. Morris, J. Pandya, S.R. Nathan, L. Smith, M.L. Cadette, and J.T. Michalowski. 2000. “A systematic approach to the validation of process control parameters for monoclonal antibody production in fed-batch culture of a murine myeloma.” Biotechnol Bioeng 69:242–255.

    Article  CAS  Google Scholar 

  74. Lyddiatt, A. 1981. “Downstream processing: Protein recovery.” In Mammalian Cell Biotechnology: A Practical Approach. M. Butler, Ed., IRL Press at Oxford University Press, New York, pp. 187–206.

    Google Scholar 

  75. Bennan, J., F. Bing, H. Boone, J. Fernandez, B. Seely, H. van Deinse, and D. Miller. 2002. “Evaluation of extractables from product-contact surfaces.” Biopharm Int 15:22–34.

    Google Scholar 

  76. Kemp, G., and P. O’Neil. 2004. “Large-scale production of therapeutic antibodies: Considerations for optimizing product capture and purification.” In Antibodies, Volume 1: Production and Purification. G. Subramanian, Ed., Kluwer Academic, Boston, pp. 75–100.

    Google Scholar 

  77. Burton, S. 2002. “A generic approach to the purification of monoclonal antibodies: An alternative to protein A.” IBC Conference: Antibody Production & Downstream Processing, Feb 13–15, San Diego, CA.

    Google Scholar 

  78. Jacob, L.R., and M. Frech. 2004. “Scale-up of antibody purification.” In Antibodies, Volume 1: Production and Purification. G. Subramanian, Ed., Kluwer Academic, Boston, pp. 101–131.

    Google Scholar 

  79. Gattschalk, U. 2005. “Large scale manufacturing of mAbs and the backlog in bioseparation technologies.” IBC Conference: Antibody Production & Downstream Processing, Mar 8–11, San Diego, CA.

    Google Scholar 

  80. Hubbard, B. 2005. “Platform approaches to monoclonal antibody purification.” IBC Conference: Antibody Production & Downstream Processing, Mar 8–11, San Diego, CA.

    Google Scholar 

  81. Fish, B. 2002. “Taking a monoclonal antibody from mg to kg scale: Production strategies, issues and successes.” IBC Conference: Scaling-up From Bench to Clinic and Beyond, Aug 14–16, San Diego, CA.

    Google Scholar 

  82. Aranha-Creado, H. 1998. “Clearance of murine leukaemia virus from monoclonal antibody solutions by a hydrophilic PVDF microporous membrane filter.” Biologicals 26:167–172.

    Article  CAS  Google Scholar 

  83. Rathore, A., and A. Velaydhan. 2003. “Guidelines for optimization and scale-up in preparative chromatography.” Biopharm Int 16:34–42.

    CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

James A. Kent Ph.D. (Professor of Chemical Engineering and Dean of Engineering)

Rights and permissions

Reprints and permissions

Copyright information

© 2007 Springer Science+Business Media, LLC

About this chapter

Cite this chapter

Zhu, M.M., Mollet, M., Hubert, R.S. (2007). Industrial Production of Therapeutic Proteins: Cell Lines, Cell Culture, and Purification. In: Kent, J.A. (eds) Kent and Riegel’s Handbook of Industrial Chemistry and Biotechnology. Springer, Boston, MA. https://doi.org/10.1007/978-0-387-27843-8_32

Download citation

Publish with us

Policies and ethics