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Suspension packaging cell lines for the simplified generation of T-cell receptor encoding retrovirus vector particles

Gene Therapy volume 14, pages 595–603 (2007)Cite this article

Abstract

The transfer of T-cell receptor (TCR) genes into primary human T-cells to endow their specificity toward virus-infected and tumor cells is becoming an interesting tool for immunotherapy. TCR-modified T cells are mainly generated by retrovirus-mediated gene transfer. To produce TCR-retrovirus particles, fibroblast packaging cell lines are the most common tool. We constructed two packaging cell lines based on the human suspension T-cell lymphoma line Δβ-Jurkat, which lacks endogenous TCRβ-chains and is therefore unable to express CD3 complexes on the cell surface. After supply of gag-pol (murine leukemia virus (Mo-MLV)) and env (GALV or MLV-10A1) genes, a green fluorescent protein (GFP)-encoding retrovirus vector was transduced into both packaging cell clones, which then stably produced GFP-retroviruses with titers of up to 4 × 105 infectious particles (IP)/ml. After transfer of a TCRα/β-encoding retrovirus vector, Δβ-Jurkat/GALV and Δβ-Jurkat/10A1 cells expressed CD3 molecules on the cell surface. CD3-high expressing packaging cells were enriched by fluorescence-activated cell sorter sorting. In these cells, the CD3 expression level directly correlated with the titer of vector particles. TCR-retroviruses efficiently transduced human T-cell lines and primary T cells. In conclusion, the method allowed the fast and easy generation of high virus titer supernatants for TCR gene transfer.

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Acknowledgements

We thank U Fischer and I Küttner for excellent technical assistance and T Blankenstein and J Charo for helpful discussions and critical reading of the manuscript. This work was supported by grants from the Deutsche Forschungsgemeinschaft (Sonderforschungsprogramm 1230, Transregio-Sonderforschungsbereich 36) and the Wilhelm Sander-Stiftung.

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Authors and Affiliations

  1. Max-Delbrück-Center for Molecular Medicine, Berlin, Germany

    S Reuß & W Uckert

  2. Institute of Biology, Humboldt-University of Berlin, Berlin, Germany

    P Biese & W Uckert

  3. INSERM, U758, Ecole Normale Supérieure of Lyon, Lyon, France

    F-L Cosset

  4. Wohl Virion Center, Windeyer Institute of Medical Sciences, University College London, London, UK

    Y Takeuchi

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  1. S Reuß
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  2. P Biese
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  3. F-L Cosset
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  4. Y Takeuchi
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  5. W Uckert
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Correspondence to W Uckert.

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Reuß, S., Biese, P., Cosset, FL. et al. Suspension packaging cell lines for the simplified generation of T-cell receptor encoding retrovirus vector particles. Gene Ther 14, 595–603 (2007). https://doi.org/10.1038/sj.gt.3302906

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