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Retrovirus mediated gene transduction of human T-cell subsets

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Abstract

Purpose: Allogeneic bone marrow transplantation (AlloBMT) can be curative for patients with leukaemia. Graft versus host disease (GVHD) is a potentially life threatening complication of AlloBMT mediated by the T cells contained within the graft. In order to be able to control GVHD, the allogeneic T cells may be transduced with a suicide gene such as herpes simplex virus thymidine kinase (HSV-tk). For this strategy to be successful, all subsets of T cells should be transduced to a similar extent. Also, the transduction protocol should not induce expression of unwanted homing receptors, nor should it lead to unwanted skewing of the T-cell receptor repertoire. We have studied the transduction efficiency of naïve and memory subsets of CD4+ and CD8+ T cells, and examined the transduced T-cell subsets for possible changes in T-cell receptor (TCR) repertoire and homing receptor expression. Methods: The cells were transduced using a Moloney murine retroviral vector carrying a conjugate of the genes encoding the truncated form of the cell surface marker, low affinity nerve growth factor receptor (ΔLNGFR) and HSV-tk. Transduction efficiency and homing receptor expression were quantified by flow cytometry. TCR repertoire was determined by spectratyping. Results: We obtained a transduction efficiency of 30–50% of the cells, with no difference between the T-cell subsets. Cell surface receptors responsible for homing to skin, gastrointestinal tract or lymph nodes were practically absent at the end of 2 weeks in culture. The activation procedure seemed to favour the expansion of certain T-cell clones over polyclonal populations. However, there was no difference in the TCR repertoire between transduced and non-transduced cells. Conclusion: Changes in the composition of the T-cell subsets at the end of the cell culture were the results of the activation, and not the suicide gene transduction. The transduced T cells did not express unwanted homing receptors.

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Acknowledgements

This project was financed with the aid of EXTRA funds from the Norwegian Foundation for Health and Rehabilitation. Contribution from the following organizations is also acknowledged: The Norwegian Cancer Society, The Blix Family Foundation, Rakel and Otto Kr. Bruun’s Legacy, Astri and Birger Torsted’s Legacy, Asbjørn Sognnes’ Estate, and Medinnova. We thank Gøril Olsen and Marianne Enger for performing the cell sorting.

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Authors and Affiliations

  1. Institute of Immunology, Rikshospitalet University Hospital, Oslo, Norway

    Ida Aagård Hedfors, Karen Johanne Beckstrøm & Jan E. Brinchmann

  2. Molmed S.p.A., Milano, Italy

    Claudia Benati

  3. Cancer Immunotherapy and Gene Therapy Progam, Bone Marrow Transplantation Unit, Scientific Institute H. S. Raffaele, Milano, Italy

    Chiara Bonini

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  1. Ida Aagård Hedfors
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  2. Karen Johanne Beckstrøm
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  5. Jan E. Brinchmann
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Correspondence to Jan E. Brinchmann.

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Hedfors, I.A., Beckstrøm, K.J., Benati, C. et al. Retrovirus mediated gene transduction of human T-cell subsets. Cancer Immunol Immunother 54, 759–768 (2005). https://doi.org/10.1007/s00262-004-0647-0

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