Generation of a high-titer packaging cell line for the production of retroviral vectors in suspension and serum-free media

ABSTRACT Several patients with severe combined immunodeficiency-X1 disease and adenosine deaminase deficiency have been cured by retroviral-mediated gene therapy. Despite the earlier

success, the production of retroviral vectors for clinical gene therapy is cumbersome, costly and lacks safety features because of the adherent nature of packaging cells and the necessity to

supplement the culture media with bovine serum. The aim of this study was to generate a retrovirus packaging cell line that could be used for the production of large clinical batch vectors.

Bicistronic vectors containing an internal ribosomal entry site followed by a selection gene were used to express Moloney murine leukemia gag-pol and amphotropic envelope viral proteins in

HEK293 cells. The candidate clone (293GP-A2) that was selected as the packaging cell line could release recombinant green fluorescent protein retroviruses at 4 × 107 infectious viral

particles per ml. Similar titers were achieved after these cells were adapted to grow in suspension and serum-free media. Furthermore, using the same culture conditions viral titers proved

to be stable for a 3-month culture period. The 293GP-A2 packaging cell line has the potential to be cultured in bioreactors, opening the possibility for large-scale use of retroviral vectors

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FAQs * Contact customer support SIMILAR CONTENT BEING VIEWED BY OTHERS A GUIDE IN LENTIVIRAL VECTOR PRODUCTION FOR HARD-TO-TRANSFECT CELLS, USING CARDIAC-DERIVED C-KIT EXPRESSING CELLS AS A

MODEL SYSTEM Article Open access 28 September 2021 AAV PRODUCTION IN STABLE PACKAGING CELLS REQUIRES EXPRESSION OF ADENOVIRUS 22/33K PROTEIN TO ALLOW EPISOMAL AMPLIFICATION OF INTEGRATED

REP/CAP GENES Article Open access 07 December 2023 STABLE EXPRESSION OF LARGE TRANSGENES VIA THE KNOCK-IN OF AN INTEGRASE-DEFICIENT LENTIVIRUS Article 01 May 2023 REFERENCES *

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CAS  PubMed  Google Scholar  Download references ACKNOWLEDGEMENTS We are grateful to Pedro Otavio de Campos-Lima for critical reading of the manuscript and to Stephen Goff for the MLV

provirus used to generate the 3T3 chronically infected cell line. This study was initiated with a grant from the Canadian Institute of Health Research (CIHR) (IC074582). MC is a Senior

Research Scholar of the Fonds de la Recherche en Santé du Québec (FRSQ). AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Le Centre de Recherche en Cancérologie de l'Université Laval,

L'Hôtel Dieu de Québec, Centre Hospitalier Universitaire de Québec, Québec, Canada K Ghani, S Cottin & M Caruso * Biotechnology Research Institute, National Research Council Canada,

Montréal, Québec, Canada A Kamen Authors * K Ghani View author publications You can also search for this author inPubMed Google Scholar * S Cottin View author publications You can also

search for this author inPubMed Google Scholar * A Kamen View author publications You can also search for this author inPubMed Google Scholar * M Caruso View author publications You can also

search for this author inPubMed Google Scholar CORRESPONDING AUTHOR Correspondence to M Caruso. RIGHTS AND PERMISSIONS Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Ghani,

K., Cottin, S., Kamen, A. _et al._ Generation of a high-titer packaging cell line for the production of retroviral vectors in suspension and serum-free media. _Gene Ther_ 14, 1705–1711

(2007). https://doi.org/10.1038/sj.gt.3303039 Download citation * Received: 18 December 2006 * Revised: 23 July 2007 * Accepted: 23 July 2007 * Published: 11 October 2007 * Issue Date:

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shareable link is not currently available for this article. Copy to clipboard Provided by the Springer Nature SharedIt content-sharing initiative KEYWORDS * retroviral vectors * packaging

cells * suspension culture * serum-free media