Inducible packaging cells for large-scale production of lentiviral vectors in serum-free suspension culture

Molecular Therapy : the Journal of the American Society of Gene Therapy
Sophie BroussauBernard Massie

Abstract

We have developed new packaging cell lines (293SF-PacLV) that can produce lentiviral vectors (LVs) in serum-free suspension cultures. A cell line derived from 293SF cells, expressing the repressor (CymR) of the cumate switch and the reverse transactivator (rtTA2(S)-M2) of the tetracycline (Tet) switch, was established first. We next generated clones stably expressing the Gag/Pol and Rev genes of human immunodeficiency virus-1, and the glycoprotein of vesicular stomatitis virus (VSV-G). Expression of Rev and VSV-G was tightly regulated by the cumate and Tet switches. Our best packaging cells produced up to 2.6 x 10(7) transducing units (TU)/ml after transfection with the transfer vector. Up to 3.4 x 10(7) TU/ml were obtained using stable producers generated by transducing the packaging cells with conditional-SIN-LV. The 293SF-PacLV was stable, as shown by the fact that some producers maintained high-level LV production for 18 weeks without selective pressure. The utility of the 293SF-PacLV for scaling up production in serum-free medium was demonstrated in suspension cultures and in a 3.5-L bioreactor. In shake flasks, the best packaging cells produced between 3.0 and 8.0 x 10(6) TU/ml/day for 3 days, and the best producer cells,...Continue Reading

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Citations

Sep 18, 2010·Bioprocess and Biosystems Engineering·Olivier HenryAmine Kamen
Jan 21, 2010·Molecular Therapy : the Journal of the American Society of Gene Therapy·Janka MátraiThierry VandenDriessche
Apr 8, 2010·Molecular Therapy : the Journal of the American Society of Gene Therapy·Yoon-Sang KimArthur W Nienhuis
Mar 2, 2010·Nature Reviews. Drug Discovery·UNKNOWN Committee for Advanced Therapies (CAT)Patrick Celis
Jun 19, 2013·Human Gene Therapy Methods·Anna StornaiuoloChiara Bovolenta
Sep 22, 2011·Human Gene Therapy·Scott R WittingMadhusudan V Peshwa
Nov 3, 2011·Human Gene Therapy·Sayandip Mukherjee, Adrian J Thrasher
Nov 16, 2010·Future Microbiology·Patrick MaierStephanie Laufs
Oct 26, 2013·Nanomedicine·Seog-Jin SeoHae-Won Kim
Jun 6, 2012·Journal of Visualized Experiments : JoVE·Mingjie LiB Joy Snider
Apr 18, 2013·Expert Opinion on Biological Therapy·Maria Mercedes SeguraAlain Garnier
Jul 18, 2012·Cytotherapy·David L Digiusto, Hans-Peter Kiem
Jun 1, 2011·Metabolic Engineering·Olivier Henry, Yves Durocher
Nov 16, 2010·Hematology/oncology Clinics of North America·Arthur Bank
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Feb 24, 2010·Biotechnology and Bioengineering·Bruno GailletBernard Massie
Nov 1, 2015·Human Molecular Genetics·Johannes C M van der Loo, J Fraser Wright
Apr 26, 2016·Molecular Therapy. Methods & Clinical Development·Otto-Wilhelm MertenChiara Bovolenta
Jan 3, 2012·Journal of Neuroscience Methods·David W ChittyMahmud Bani-Yaghoub
Apr 9, 2016·Molecular Therapy : the Journal of the American Society of Gene Therapy·Olivier HumbertHans-Peter Kiem
Mar 13, 2015·Scientific Reports·Khaled S SanberYasuhiro Takeuchi
May 19, 2010·Expert Review of Respiratory Medicine·Uta Griesenbach, Eric W F W Alton
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Sep 30, 2010·Experimental Biology and Medicine·Anthony J BellArthur Bank
Sep 2, 2017·Scientific Reports·Jean-François GélinasStephen C Hyde

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