Review Article | Published:

Production of recombinant protein therapeutics in cultivated mammalian cells

Nature Biotechnology volume 22, pages 13931398 (2004) | Download Citation

Subjects

Abstract

Cultivated mammalian cells have become the dominant system for the production of recombinant proteins for clinical applications because of their capacity for proper protein folding, assembly and post-translational modification. Thus, the quality and efficacy of a protein can be superior when expressed in mammalian cells versus other hosts such as bacteria, plants and yeast. Recently, the productivity of mammalian cells cultivated in bioreactors has reached the gram per liter range in a number of cases, a more than 100-fold yield improvement over titers seen for similar processes in the mid-1980s. This increase in volumetric productivity has resulted mainly from improvements in media composition and process control. Opportunities still exist for improving mammalian cell systems through further advancements in production systems as well as through vector and host cell engineering.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    & Kinetics of recombinant immunoglobulin production by mammalian cells in continuous culture. Biotechnol. Bioeng. 38, 972–976 (1991).

  2. 2.

    , & Co-expression and amplification of dihydrofolate reductase cDNA and the Escherichia coli XGPRT gene in Chinese hamster ovary cells. J. Mol. Appl. Genet. 1, 165–175 (1981).

  3. 3.

    et al. Use of the human EF-1alpha promoter for expression can significantly increase success in establishing stable cell lines with consistent expression: a study using the tetracycline-inducible system in human cancer cells. Nucleic Acids Res. 27, 4775–4782 (1999).

  4. 4.

    , , How introns influence and enhance eukaryotic gene expression. Trends in Biochem. Sci. 215–220 (2004).

  5. 5.

    Components of vectors for gene transfer and expression in mammalian cells. Protein Expr. Purif. 17, 183–202 (1999).

  6. 6.

    & A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology 52, 456–467 (1973).

  7. 7.

    & Methods for DNA introduction into mammalian cells. in Gene Transfer and Expression in Mammalian Cells. (ed. Makrides, S.C.) 265–277 (Elsevier Science BV, Amsterdam, 2003).

  8. 8.

    et al. Characterisation of two differently processed forms of human recombinant factor IX synthesised in CHO cells transformed with a polycistronic vector. Eur. J. Biochem. 172, 565–572 (1988).

  9. 9.

    , , & Homologous plasmid recombination is elevated in immortally transformed cells. Mol. Cell. Biol. 9, 4009–4017 (1989).

  10. 10.

    , & Genetic and physical linkage of exogenous sequences in transformed cells. Cell 22, 309–317 (1980).

  11. 11.

    & Epigenetic codes for heterochromatin formation and silencing: rounding up of the usual suspects. Cell 108, 489–500 (2002).

  12. 12.

    & Silencing of transgene transcription precedes methylation of promoter DNA and histone H3 lysine. EMBO J. 23, 138–149 (2004).

  13. 13.

    & Use of scaffold/matrix-attachment regions for protein production. in Gene Transfer and Expression in Mammalian Cells. (ed. Makrides, S.C.) 359–379 (Elsevier Science BV, Amsterdam, 2003).

  14. 14.

    et al. Transgenese encompassing dual-promoter CpG islands from the human TBP and H NRPA2B1 loci are resistant to heterochromatin-mediated silencing. Genomics 82, 269–279 (2003).

  15. 15.

    et al. Identification of anti-repressor elements that confer high and stable protein production in mammalian cells. Nat. Biotechnol. 21, 553–558 (2003).

  16. 16.

    , & Matrix attachment region-dependent function of the immunoglobulin mu enhancer involves histone acetylation at a distance without changes in enhancer occupancy. Mol. Cell. Biol. 21, 196–208 (2001).

  17. 17.

    et al. Development of stable cell lines for production or regulated expression using matrix attachment regions. J. Biotechnol. 87, 29–42 (2001).

  18. 18.

    , & Expression of recombinant plasmids in mammalian cells is enhanced by sodium butyrate. Nucleic Acids Res. 11, 7631–7648 (1983).

  19. 19.

    , & Protein phosphatase is involved in the inhibition of histone diacetylation by sodium butyrate. Biochem. Biophys. Res. Comm. 246, 760–764 (1998).

  20. 20.

    & Gene targeting in normal and amplified cell lines. Nature 344, 170–173 (1990).

  21. 21.

    , , , & Host cell derived retroviral sequences enhance transfection and expression efficiency in CHO cells. in Animal Cell Technology: Developments, Processes and Products (eds. R.E. Spier, J.B. Griffiths, C. MacDonnald) 35–41 (Butterworth-Heinemann, Oxford, UK, 1992).

  22. 22.

    et al. Architecture and utilization of highly expressed genomic sites. in Gene Transfer and Expression in Mammalian Cells. (ed. Makrides, S.C.) 551–571 (Elsevier Science, BV, Amsterdam, 2002).

  23. 23.

    & The LoxP/CRE system and genome modification. Methods Mol. Biol. 158, 83–94 (2001).

  24. 24.

    , & Cell Engineering blocks stress and improves biotherapeutic production. Bioprocessing 3, 23–28 (2004).

  25. 25.

    , , , & In hybridoma cultures, deprivation of any single amino acid leads to apoptotic death, which is suppressed by expression of the bcl-2 gene. Biotechnol. Bioeng. 59, 90–98 (1999).

  26. 26.

    , , , & Engineered glycoforms of an antineuroblastoma IgG1 with optimized antibody-dependent cellular cytotoxic activity. Nat. Biotechnol. 17, 176–180 (1999).

  27. 27.

    et al. Effects of methotrexate on transfected DNA stability in mammalian cells. Mol. Cell. Biol. 10, 401–404 (1990).

  28. 28.

    , , & Amplification and expression of recombinant genes in serum-independent Chinese hamster ovary cells. FEBS Lett. 377, 290–294 (1995).

  29. 29.

    , & Inducible overexpression of the mouse c-myc protein in mammalian cells. Proc. Natl. Acad. Sci. USA 83, 5414–5418 (1986).

  30. 30.

    , , & Isolation of overproducing recombinant mammalian cells by a fast and simple selection procedure. Gene 73, 419–426 (1988).

  31. 31.

    , et al. High level expression of a recombinant antibody from myeloma cells using a glutamine synthetase gene as an amplifiable selectable marker. Bio/Technology 10, 169–175 (1992).

  32. 32.

    , & Advances in animal cell recombinant protein production: GS-NS0 expression system. Cytotechnology 32, 109–123 (2000).

  33. 33.

    , & Molecular definition of predictive indicators of stable protein expression in recombinant NS0 myeloma cells. Biotechnol. Bioeng. 85, 115–121 (2004).

  34. 34.

    et al. High level expression of recombinant IgG in the human cell line PER.C6. Biotechnol. Prog. 19, 163–168 (2003).

  35. 35.

    , , & Higher productivity of growth-arrested Chinese hamster ovary cells expressing the cyclin-dependent kinase inhibitor p27. Biotechnol. Prog. 14, 705–713 (1998).

  36. 36.

    , & Expression of recombinant plasmids in mammalian cells is enhanced by sodium butyrate. Nucleic Acids Res. 11, 7631–7648 (1983).

  37. 37.

    Chromosomal evolution in cell populations. Int. Rev. Cytol. 12, 69–121 (1961).

  38. 38.

    , & Characterization of the stability of recombinant protein production in the GS-NS0 expression system. Biotechnol. Bioeng. 73, 261–270 (2001).

  39. 39.

    , & Integration and stability of CHO amplicons containing plasmid sequences. Dev. Biol. Stand. 76, 69–82 (1992).

  40. 40.

    & 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 (1999).

  41. 41.

    et al. Recombinant follicle stimulating hormone: development of the first biotechnology product for the treatment of infertility. Hum. Reprod. Update 4, 862–881 (1998).

  42. 42.

    & The evolution of processes for the commercial exploitation of anchorage-dependent animal cells. Enzyme Microb. Technol. 21, 2–8 (1997).

  43. 43.

    , , , & Production of recombinant Factor VIII from perfusion cultures: I. Large Scale Fermentation. in Animal Cell Technology, Products of Today, Prospects for Tomorrow. (eds. R.E. Spier, J.B. Griffiths, and W. Berthold) 580–590 (Butterworth-Heinemann, Oxford, UK, 1994).

  44. 44.

    & Large scale transient expression in mammalian cells for recombinant protein production. Curr. Opin. Biotechnol. 10, 156–159 (1999).

  45. 45.

    , & Adaptation of mammalian cells to growth in serum-free media. Mol. Biotechnol. 15, 249–257 (2000).

  46. 46.

    Laboratory scale up of cell cultures (0.5–50 liters). Methods Cell Biol. 57, 219–227 (1998).

  47. 47.

    , et al. TubeSpin satellites: a fast track approach for process development with animal cells using shaking technology. Biochem. Eng. J. 17, 217–223 (2004).

  48. 48.

    et al. Gas-inducible transgene expression in mammalian cells, mice and bioreactor. Nat. Biotechnol. 22, 1440–1444 (2004).

  49. 49.

    & Metabolic engineering of mammalian cells for higher protein yield. in Gene Transfer and Expression in Mammalian Cells (ed. Makrides, S.C.) 457–569 (Elsevier Science, BV, Amsterdam, 2003).

Download references

Acknowledgements

I am very grateful to my colleagues David Hacker and Martin Jordan for discussions and extensive editing in the context of the writing of this review.

Author information

Affiliations

  1. Laboratory of Cellular Biotechnology, Institute of Biological Engineering and Biotechnology, Faculty of Life Sciences, Swiss Federal Institute of Technology, Lausanne (EPFL), 1015 Lausanne, Switzerland.

    • Florian M Wurm

Authors

  1. Search for Florian M Wurm in:

Corresponding author

Correspondence to Florian M Wurm.

About this article

Publication history

Published

DOI

https://doi.org/10.1038/nbt1026

Further reading