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


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2001-05

AUTHORS

M Pizzato, O W Merten, E D Blair, Y Takeuchi

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 10(7) 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. More... »

PAGES

3301457

References to SciGraph publications

Identifiers

URI

http://scigraph.springernature.com/pub.10.1038/sj.gt.3301457

DOI

http://dx.doi.org/10.1038/sj.gt.3301457

DIMENSIONS

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PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/11420637


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56 schema:description 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 10(7) 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.
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