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.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Eshhar Z . Tumor-specific T-bodies: towards clinical application. Cancer Immunol Immunother 1997; 45: 131–136.
Hombach A, Heuser C, Abken H . The recombinant T cell receptor strategy: insights into structure and function of recombinant immunoreceptors on the way towards an optimal receptor design for cellular immunotherapy. Curr Gene Ther 2002; 2: 211–226.
Schumacher TN . T-cell-receptor gene therapy. Nat Rev Immunol 2002; 2: 512–519.
Miller A . Retrovirus packaging cells. Hum Gene Ther 1990; 1: 5–14.
Pear WS, Nolan GP, Scott ML, Baltimore D . Production of high-titer helper-free retroviruses by transient transfection. Proc Natl Acad Sci USA 1993; 90: 8392–8396.
Cornetta K, Matheson L, Ballas C . Retroviral vector production in the National Gene Vector Laboratory at Indiana University. Gene Therapy 2005; 12: S28–S35.
Farson D, McGuinness R, Dull T, Limoli K, Lazar R, Jalali S et al. Large-scale manufacturing of safe and efficient retrovirus packaging lines for use in immunotherapy protocols. J Gene Med 1999; 1: 195–209.
Sheridan PL, Bodner M, Lynn A, Phuong TK, DePolo NJ, de la Vega J et al. Generation of retroviral packaging and producer cell lines for large-scale vector production and clinical application: improved safety and high titer. Mol Ther 2000; 2: 262–275.
Jung D, Jaeger E, Cayeux S, Blankenstein T, Hilmes C, Karbach J et al. Strong immunogenic potential of a B7 retroviral expression vector: generation of HLA-B7-restricted CTL response against selectable marker genes. Hum Gene Ther 1998; 9: 53–62.
Emerman M, Temin HM . Genes with promoters in retroviral vectors can be independently suppressed by an epigenetic mechanism. Cell 1984; 39: 459–467.
Chan LM, Coutelle C, Themis M . A novel human suspension culture packaging cell line for production of high-titre retroviral vectors. Gene Therapy 2001; 8: 697–703.
Pizzato M, Merten OW, Blair ED, Takeuchi Y . Development of a suspension packaging cell line for production of high titre, serum-resistant murine leukemia virus vectors. Gene Therapy 2001; 8: 737–745.
Bunnell BA, Muul LM, Donahue RE, Blaese RM, Morgan RA . High-efficiency retroviral-mediated gene transfer into human and nonhuman primate peripheral blood lymphocytes. Proc Natl Acad Sci USA 1995; 92: 7739–7743.
Lam JS, Reeves ME, Cowherd R, Rosenberg SA, Hwu P . Improved gene transfer into human lymphocytes using retroviruses with the gibbon ape leukemia virus envelope. Hum Gene Ther 1996; 7: 1415–1422.
Uckert W, Becker C, Gladow M, Klein D, Kammertoens T, Pedersen L et al. Efficient gene transfer into primary human CD8+ T lymphocytes by MuLV-10A1 retrovirus pseudotype. Hum Gene Ther 2000; 11: 1005–1014.
Gladow M, Becker C, Blankenstein T, Uckert W . MLV-10A1 retrovirus pseudotype efficiently transduces primary human CD4+ T lymphocytes. J Gene Med 2000; 2: 409–415.
Kaptein LC, Greijer AE, Valerio D, van Beusechem VW . Optimized conditions for the production of recombinant amphotropic retroviral vector preparations. Gene Therapy 1997; 4: 172–176.
Merten OW . State-of-the-art of the production of retroviral vectors. J Gene Med 2004; 6 Suppl 1: S105–S124.
Kotani H, Newton III PB, Zhang S, Chiang YL, Otto E, Weaver L et al. Improved methods of retroviral vector transduction and production for gene therapy. Hum Gene Ther 1994; 5: 19–28.
Cosset FL, Takeuchi Y, Battini JL, Weiss RA, Collins MK . High-titer packaging cells producing recombinant retroviruses resistant to human serum. J Virol 1995; 69: 7430–7436.
Stitz J, Buchholz CJ, Engelstadter M, Uckert W, Bloemer U, Schmitt I et al. Lentiviral vectors pseudotyped with envelope glycoproteins derived from Gibbon Ape Leukemia Virus and Murine Leukemia Virus 10A1. Virology 2000; 273: 16–20.
Engels B, Cam H, Schuler T, Indraccolo S, Gladow M, Baum C et al. Retroviral vectors for high-level transgene expression in T lymphocytes. Hum Gene Ther 2003; 14: 1155–1168.
Engels B, Noessner E, Frankenberger B, Blankenstein T, Schendel DJ, Uckert W . Redirecting human T lymphocytes toward renal cell carcinoma specificity by retroviral transfer of T cell receptor genes. Hum Gene Ther 2005; 16: 799–810.
Jantzer P, Schendel DJ . Human renal cell carcinoma antigen-specific CTLs: antigen-driven selection and long-term persistence in vivo. Cancer Res 1998; 58: 3078–3086.
Horn PA, Topp MS, Morris JC, Riddell SR, Kiem HP . Highly efficient gene transfer into baboon marrow repopulating cells using GALV-pseudotype oncoretroviral vectors produced by human packaging cells. Blood 2002; 100: 3960–3967.
Uckert W, Kammertons T, Haack K, Qin Z, Gebert J, Schendel DJ et al. Double suicide gene (cytosine deaminase and herpes simplex virus thymidine kinase) but not single gene transfer allows reliable elimination of tumor cells in vivo. Hum Gene Ther 1998; 9: 855–865.
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.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
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
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/sj.gt.3302906
Keywords
This article is cited by
-
Enhanced functionality of T cell receptor-redirected T cells is defined by the transgene cassette
Journal of Molecular Medicine (2008)