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PubMed · 12679787

Continuous high-titer HIV-1 vector production.

Yasuhiro Ikeda·Yasuhiro Takeuchi·Francisco Martin·Francois-Loic Cosset·Kyriacos Mitrophanous·Mary Collins
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Abstract

Human immunodeficiency virus type 1 (HIV-1)-based vectors are currently made by transient transfection, or using packaging cell lines in which expression of HIV-1 Gag and Pol proteins is induced. Continuous vector production by cells in which HIV-1 Gag-Pol is stably expressed would allow rapid and reproducible generation of large vector batches. However, attempts to make stable HIV-1 packaging cells by transfection of plasmids encoding HIV-1 Gag-Pol have resulted in cells which secrete only low levels of p24 antigen (20-80 ng/ml), possibly because of the cytotoxicity of HIV-1 protease. Infection of cells with HIV-1 can result in stable virus production; cell clones that produce up to 1,000 ng/ml secreted p24 antigen have been described. Here we report that expression of HIV-1 Gag-Pol by a murine leukemia virus (MLV) vector allows constitutive, long-term, high-level (up to 850 ng/ml p24) expression of HIV-1 Gag. Stable packaging cells were constructed using codon-optimized HIV-1 Gag-Pol and envelope proteins of gammaretroviruses; these producer cells could make up to 10(7) 293T infectious units (i.u.)/ml (20 293T i.u./cell/day) for at least three months in culture.

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Gene Expression Regulation, ViralGenetic VectorsHIV-1HeLa CellsHumansQuality ControlTransfectionEvaluation StudyResearch Support, Non-U.S. Gov'tTechnical ReportJournal Article

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Yasuhiro Ikeda, Yasuhiro Takeuchi, Francisco Martin, Francois-Loic Cosset, Kyriacos Mitrophanous, Mary Collins. 2003-04-07. Continuous high-titer HIV-1 vector production.. https://doi.org/10.1038/nbt815

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