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Development of a suspension packaging cell line for production of high titre, serum-resistant murine leukemia virus vectors

Gene Therapy volume 8, pages 737–745 (2001)Cite this article

An Erratum to this article was published on 11 October 2001

Abstract

To date, only adherent cell lines have been used for the generation of packaging cells for the production of type C retrovirus vectors. The large-scale production of high titre retrovirus vectors could benefit from the development of packaging cells growing in suspension. Here, we describe the ability of two different lymphoid cell lines, one B- and one T-lymphoblastoid cell line (Namalwa and CEM, respectively), to produce MLV-based vectors. Upon transfection with a third generation packaging construct, the virus particle production by Namalwa cells was characterised by low RT-activity, and by CEM cells as high RT activity as previously established adherent packaging cells. An amphotropic packaging cell line (CEMFLYA) was therefore established from CEM cells. Upon introduction of a lacZ vector genome, the novel packaging cell line produced vector particles routinely in the region of 107 infectious units/ml. The vectors were helper-free and highly stable in fresh human serum. The potential for scaled up vector production was demonstrated by continuous culture of the new packaging cells for 14 days in a 250 ml spinner flask. These suspension packaging cells should be applicable to large bioreactor systems to bulk produce high titre, complement-resistant retrovirus vectors for gene therapy. Gene Therapy (2001) 8, 737–745.

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Acknowledgements

This work is supported by GlaxoWellcome and the Medical Research Council. MP was funded by a EU TMR grant (contr. number FMBICT961804). We thank Brendan Hughes at GlaxoWellcome for seeding the idea of suspension packaging cells, Robin Weiss at Wohl Virion Centre for helpful advice, and Sabine Low-Hong and Olivier Danos at Genethon for excellent experimental work and kindly support, respectively. We also thank Michael Themis at Imperial College London, who has developed suspension packaging cell lines producing ecotropic retrovirus vectors, for helpful discussion and sharing data.

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Author notes

  1. M Pizzato

    Present address: Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02115, USA

Authors and Affiliations

  1. Wohl Virion Centre, Windeyer Institute of Medical Sciences, University College London, UK

    M Pizzato & Y Takeuchi

  2. Virology Unit, GlaxoWellcome, Medicines Research Centre, Stevenage, UK

    M Pizzato & E D Blair

  3. Généthon III, Evry, France

    O W Merten

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  4. Y Takeuchi
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Pizzato, M., Merten, O., Blair, E. et al. Development of a suspension packaging cell line for production of high titre, serum-resistant murine leukemia virus vectors. Gene Ther 8, 737–745 (2001). https://doi.org/10.1038/sj.gt.3301457

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