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Efficient transduction and long-term retroviral expression of the melanoma-associated tumor antigen tyrosinase in CD34+ cord blood-derived dendritic cells

Gene Therapy volume 9, pages 1551–1560 (2002)Cite this article

Abstract

Differentiation of genetically modified CD34+ hematopoietic stem cells into dendritic cells (DCs) will contribute to the development of immunotherapeutic anticancer protocols. Retroviral vectors that have been used for the transduction of CD34+ cells face the problem of gene silencing when integrated into the genome of repopulating stem cells. We reasoned that a high copy number of retroviral DNA sequences might overcome silencing of transgene expression during expansion and differentiation of progenitor cells into functional DCs. To prove this, we utilized a retroviral vector with bicistronic expression of the melanoma-associated antigen tyrosinase and the enhanced green fluorescent protein (EGFP). Human cord blood CD34+ cells were transduced with vesicular stomatitis virus G-protein (VSV-G) pseudotyped Moloney murine leukemia virus (MoMuLV) particles using 100–150 multiplicity of infection. During expansion of transduced cells with immature phenotype, transgene expression was strongly silenced, but upon differentiation into mature DCs, residual transgene expression was retained. Intracellular processing of the provirally expressed tyrosinase was tested in a chromium release assay utilizing a cytotoxic T cell clone specific for a HLA-A*0201-restricted tyrosinase peptide. We suggest that retroviral transduction of tumor-associated antigens in hematopoietic progenitor cells and subsequent differentiation into DCs is a suitable basis for the development of potent anti-tumor vaccines.

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Acknowledgements

This work was supported by a grant of the Ministry of Environment and Agriculture (Az. 56-8811.61/71), the State of Saxony, Germany, to AT and EPR. We thank S Heinicke and B Utess for excellent technical assistance. We are grateful to Dr PI Schrier, Department of Clinical Oncology, Leiden, and to Prof Dr CJ Melief, Department of Immunohematology and Blood Transfusion, Tumor Immunology Laboratory, Leiden, The Netherlands, for providing the tyrosinase specific T cell clone TyrF8.

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  1. A Temme and A Morgenroth: The first two authors contributed equally to this work

Authors and Affiliations

  1. Institute of Immunology, Medical Faculty Carl Gustav Carus, Technical University Dresden, Dresden, Germany

    A Temme, A Morgenroth, M Schmitz, B Weigle, M Füssel & E P Rieber

  2. Department of Internal Medicine I, University Hospital Carl Gustav Carus, Dresden, Germany

    A Temme & G Ehninger

  3. Institute of Virology, Medical Faculty Carl Gustav Carus, Technical University Dresden, Dresden, Germany

    J Rohayem & D Lindemann

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  1. A Temme
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Temme, A., Morgenroth, A., Schmitz, M. et al. Efficient transduction and long-term retroviral expression of the melanoma-associated tumor antigen tyrosinase in CD34+ cord blood-derived dendritic cells. Gene Ther 9, 1551–1560 (2002). https://doi.org/10.1038/sj.gt.3301821

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