Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

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Lemoine, F.M. et al.

Lemoine, Mesel-Lemoine, Cherai, Gallot, Vié, Leclercq, Trébèden-Negre, Mammès, Boyer, Noguiez-Hellin, Klatzmann,

Efficient transduction and selection of human T-lymphocytes with bicistronic Thy1/HSV1-TK retroviral vector produced by a human packaging cell line.

BACKGROUND: T-cells expressing the HSV1-TK suicide gene can be used for the control of graft-versus-host disease following allogeneic stem cell transplantation. To develop clinical trials based on such a strategy, we have generated under good manufacturing procedures a novel 'split genome' human packaging cell line (1704 cells). METHODS: To minimize the risk of generating replication-competent retroviruses, pol was truncated to remove sequences overlapping with env. To improve retroviral infection and selection of transduced T-cells, high titers of GALV-pseudotyped retroviral particles harboring a bicistronic Thy1-IRES-TK vector coding for the CD90 GPI-anchored membrane molecule were produced by 1704 cells. RESULTS: Using 1704 cell supernatant and an optimized transduction protocol, approximately 50% of primary T-cells were transduced and could then be purified (approximately 95%) using clinical-grade immunomagnetic beads directed against CD90. Over 96% of these OKT3/IL-2-activated CD90(+)-selected T-cells were killed by ganciclovir. Cell proliferation and cytokine production of transduced T-cells and HLA-restricted cytotoxicity of transduced T-cell clones were identical to those of their non-transduced counterparts cultured under the same conditions. CONCLUSIONS: GALV-pseudotyped retroviral particles harboring a bicistronic Thy1-IRES-TK vector allow efficient transduction and rapid selection of human T-cells under conditions applicable for clinical trials using the new human 1704 packaging cell line.[1]

References

Efficient transduction and selection of human T-lymphocytes with bicistronic Thy1/HSV1-TK retroviral vector produced by a human packaging cell line. Lemoine, F.M., Mesel-Lemoine, M., Cherai, M., Gallot, G., Vié, H., Leclercq, V., Trébèden-Negre, H., Mammès, O., Boyer, O., Noguiez-Hellin, P., Klatzmann, D. The journal of gene medicine. (2004) [Pubmed]

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