Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Viral Transfer Technology
  • Published:

Functional long-term thymidine kinase suicide gene expression in human T cells using a herpesvirus saimiri vector

Gene Therapy volume 7, pages 664–674 (2000)Cite this article

Abstract

Herpesvirus saimiri transforms human T lymphocytes to stable growth and persists episomally without genomic integration and without virus production. The transformed T cells retain essential features of their parental cells including the MHC-restricted antigen specificity which may be useful for applications in adoptive immunotherapy. In order to improve the biological safety of such vectors, the prodrug activating gene thymidine kinase of herpes simplex virus was inserted into the genome of herpesvirus saimiri by homologous recombination. After infection with wild-type or cloned recombinant viruses, T cells from tamarin monkeys and from humans were transformed to stable growth. Thymidine kinase-expressing transformed T cells were efficiently eliminated in the presence of low concentrations of ganciclovir. This elimination mechanism remained fully functional over an observation period of 12 months. The potentially immunogenic neomycin resistance gene expression cassette was deleted from the genome of established mutant viruses by using the prokaryotic Cre/LoxP recombination system. At any time during the course of a therapeutic application, thymidine kinase-expressing transformed human T cells might be eliminated after administration of ganciclovir. In principle, this function could be useful for the T cell-dependent immunotherapy of resistant blood cancer while avoiding the risk of uncontrolled graft-versus-host disease.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

Similar content being viewed by others

References

  1. Riddell SR, Greenberg PD . Principles for adotive T cell therapy of human viral diseases Annu Rev Immunol 1995 13: 545–586

    Article  CAS  PubMed  Google Scholar 

  2. Kolb HJ, Holler E . Adoptive immunotherapy with donor lymphocyte transfusions Curr Opin Oncol 1997 9: 139–145

    Article  CAS  PubMed  Google Scholar 

  3. Riddell SR et al. Restoration of viral immunity in immunodeficient humans by the adoptive transfer of T cell clones Science 1992 257: 238–241

    Article  CAS  PubMed  Google Scholar 

  4. Walter EA et al. Reconstitution of cellular immunity against cytomegalovirus in recipients of allogeneic bone marrow by transfer of T-cell clones from the donor New Engl J Med 1995 333: 1038–1044

    Article  CAS  PubMed  Google Scholar 

  5. Bonini C et al. HSV-TK gene transfer into donor lymphocytes for control of allogeneic graft-versus-leukemia Science 1997 276: 1719–1724

    Article  CAS  PubMed  Google Scholar 

  6. Heslop HE et al. Long-term restoration of immunity against Epstein–Barr virus infection by adoptive transfer of gene-modified virus-specific T lymphocytes Nature Med 1996 2: 551–555

    Article  CAS  PubMed  Google Scholar 

  7. Papadopoulos EB et al. Infusions of donor leukocytes to treat Epstein-Barr virus-associated lymphoproliferative disorders after allogeneic bone marrow transplantation New Engl J Med 1994 330: 1185–1191

    Article  CAS  PubMed  Google Scholar 

  8. Rooney CM et al. Use of gene-modified virus-specific T lymphocytes to control Epstein–Barr-virus-related lymphoproliferation Lancet 1995 345: 9–14

    Article  CAS  PubMed  Google Scholar 

  9. Collins RH et al. Donor leukocyte infusions in 140 patients with relapsed malignancy after allogeneic bone marrow transplantation J Clin Oncol 1997 15: 433–444

    Article  PubMed  Google Scholar 

  10. Kolb HJ et al. Donor leukocyte transfusions for treatment of recurrent chronic myelogenous leukemia in marrow transplant patients Blood 1990 76: 2462–2465

    CAS  PubMed  Google Scholar 

  11. Kolb HJ et al. Graft-versus-leukemia effect of donor lymphocyte transfusions in marrow grafted patients Blood 1995 86: 2041–2050

    CAS  PubMed  Google Scholar 

  12. Or R et al. Allogeneic cell-mediated and cytokine-activated immunotherapy for malignant lymphoma at the stage of minimal residual disease after autologous stem cell transplantation J Immunother 1998 21: 447–453

    Article  CAS  PubMed  Google Scholar 

  13. Slavin S et al. Cellular mediated immunotherapy of leukemia in conjunction with autologous and allogeneic bone marrow transplantation in experimental animals and man Blood 1988 72 (Suppl. 1): 407

    Google Scholar 

  14. Slavin S et al. Allogeneic cell therapy for relapsed leukemia after bone marrow transplantation with donor peripheral blood lymphocytes Exp Hematol 1995 23: 1553–1562

    CAS  PubMed  Google Scholar 

  15. Slavin S et al. Allogeneic cell therapy with donor peripheral blood cells and recombinant human interleukin-2 to treat leukemia relapse after allogeneic bone marrow transplantation Blood 1996 87: 2195–2204

    CAS  PubMed  Google Scholar 

  16. Tiberghien P et al. Ganciclovir treatment of herpes simplex thymidine kinase-transduced primary T lymphocytes: an approach for specific in vivo donor T-cell depletion after bone marrow transplantation? Blood 1994 84: 1333–1341

    CAS  PubMed  Google Scholar 

  17. Contassot E et al. Retrovirus-mediated transfer of the herpes simplex type 1 thymidine kinase gene in alloreactive T lymphocytes Hum Gene Ther 1998 9: 73–80

    Article  CAS  PubMed  Google Scholar 

  18. Munshi NC et al. Thymidine kinase (TK) gene-transduced human lymphocytes can be highly purified, remain fully functional, and are killed efficiently with ganciclovir Blood 1997 89: 1334–1340

    CAS  PubMed  Google Scholar 

  19. Verzeletti S et al. Herpes simplex virus thymidine kinase gene transfer for controlled graft-versus-host disease and graft-versus-leukemia: clinical follow-up and improved new vectors Hum Gene Ther 1998 9: 2243–2251

    Article  CAS  PubMed  Google Scholar 

  20. Bordignon C et al. Transfer of the HSV-tk gene into donor peripheral blood lymphocytes for in vivo modulation of donor anti-tumor immunity after allogeneic bone marrow transplantation Hum Gene Ther 1995 6: 813–819

    Article  CAS  PubMed  Google Scholar 

  21. Tiberghien P et al. Use of donor T-lymphocytes expressingherpes-simplex thymidine kinase in allogeneic bone marrow transplantation: a phase I–II study Hum Gene Ther 1997 8: 615–624

    Article  CAS  PubMed  Google Scholar 

  22. Weijtens MEM, Willemsen RA, Hart EH, Bolhuis RLH . A retroviral vector system STITCH in combination with an optimized single chain antibody chimeric receptor gene structure allows efficient gene transduction and expression in human T lymphocytes Gene Therapy 1998 5: 1195–1203

    Article  CAS  PubMed  Google Scholar 

  23. Mavilio F et al. Peripheral blood lymphocytes as target cells of retroviral vector-mediated gene transfer Blood 1994 83: 1988–1997

    CAS  PubMed  Google Scholar 

  24. Melendez LV, Daniel MD, Hunt RD, Garcia RG . An apparently new herpesvirus from primary kidney cultures of the squirrel monkey (Saimiri sciureus) Lab Animal Care 1968 18: 374–381

    CAS  Google Scholar 

  25. Desrosiers RC et al. Synthesis of bovine growth hormone in primates by using a herpesvirus vector Mol Cell Biol 1985 5: 2796–2803

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Alt M, Fleckenstein B, Grassmann R . A pair of selectable herpesvirus vectors for simultaneous gene expression in human lymphoid cells Gene 1991 102: 265–269

    Article  CAS  PubMed  Google Scholar 

  27. Grassmann R, Fleckenstein B . Selectable recombinant herpesvirus saimiri is capable of persisting in a human T-cell line J Virol 1989 63: 1818–1821

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Grassmann R et al. Transformation to continuous growth of primary human T lymphocytes by human T-cell leukemia virus type I X-region genes transduced by a herpesvirus saimiri vector Proc Natl Acad Sci USA 1989 86: 3351–3355

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Grassmann R et al. Role of human T-cell leukemia virus type 1 X region proteins in immortalization of primary human lymphocytes in culture J Virol 1992 66: 4570–4575

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Rosin O et al. A human T-cell leukemia virus Tax variant incapable of activating NF-kB retains its immortalizing potential for primary T-lymphocytes J Biol Chem 1998 273: 6698–6703

    Article  CAS  PubMed  Google Scholar 

  31. Schmitt I et al. Stimulation of cyclin-dependent kinase activity and G1- to S-phase transition in human lymphocytes by the human T-cell leukemia/lymphotropic virus type 1 Tax protein J Virol 1998 72: 633–640

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Biesinger B et al. Stable growth transformation of human T lymphocytes by herpesvirus saimiri Proc Natl Acad Sci USA 1992 89: 3116–3119

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Fickenscher H et al. Regulation of the herpesvirus saimiri oncogene stpC, similar to that of T-cell activation genes, in growth-transformed human T lymphocytes J Virol 1996 70: 6012–6019

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Simmer B et al. Persistence of selectable herpesvirus saimiri in various human haematopoietic and epithelial cell lines J Gen Virol 1991 72: 1953–1958

    Article  PubMed  Google Scholar 

  35. Bröker BM et al. Immortalization of human T cell clones by herpesvirus saimiri. Signal transduction analysis reveals functional CD3, CD4, and IL-2 receptors J Immunol 1993 151: 1184–1192

    PubMed  Google Scholar 

  36. De Carli M et al. Immortalization with herpesvirus saimiri modulates the cytokine secretion profile of established Th1 and Th2 human T cell clones J Immunol 1993 151: 5022–5030

    CAS  PubMed  Google Scholar 

  37. Fickenscher H, Fleckenstein B . Growth-transformation of human T cells Meth Microbiol 1998 25: 573–603

    Article  Google Scholar 

  38. Mittrücker HW, Müller-Fleckenstein I, Fleckenstein B, Fleischer B . Herpesvirus saimiri-transformed human T lymphocytes: normal functional phenotype and preserved T cell receptor signalling Int Immunol 1993 5: 985–990

    Article  PubMed  Google Scholar 

  39. Weber F et al. Transformation of human T-cell clones by herpesvirus saimiri: intact antigen recognition by autonomously growing myelin basic protein-specific T cells Proc Natl Acad Sci USA 1993 90: 11049–11053

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Knappe A et al. The superantigen-homologous viral immediate–early gene ie14/vsag in herpesvirus saimiri-transformed human T cells J Virol 1997 71: 9124–9133

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Knappe A et al. Herpesvirus saimiri-transformed macaque T cells are tolerated and do not cause lymphoma after autologous reinfusion Blood 2000 (in press)

  42. Biesinger B et al. The product of the herpesvirus saimiri open reading frame 1 (tip) interacts with T cell-specific kinase p56lck in transformed cells J Biol Chem 1995 270: 4729–4734

    Article  CAS  PubMed  Google Scholar 

  43. Kraft MS et al. Herpesvirus saimiri transforms human T-cell clones to stable growth without inducing resistance to apoptosis J Virol 1998 72: 3138–3145

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Fickenscher H et al. Functional phenotype of transformed human alphabeta and gammadelta T cells determined by different subgroup C strains of herpesvirus saimiri J Virol 1997 71: 2252–2263

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Klein JL et al. Herpesvirus saimiri immortalized gamma delta T cell line activated by IL-12 J Immunol 1996 156: 2754–2760

    CAS  PubMed  Google Scholar 

  46. Pacheco-Castro et al. Herpesvirus saimiri immortalization of alpha/beta and gamma/delta human T-lineage cells derived from CD34+ intrathymic precursors in vitro Int Immunol 1996 8: 797–805

    Article  Google Scholar 

  47. Yasukawa M, Inoue Y, Kimura N, Fujita S . Immortalization of human T cells expressing T-cell receptor gamma delta by herpesvirus saimiri J Virol 1995 69: 8114–8117

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Fickenscher H et al. TcR expression of herpesvirus saimiri immortalized human T cells Immunologist 1996 4: 41–43

    CAS  Google Scholar 

  49. Jung JU, Desrosiers RC . Identification and characterization of the herpesvirus saimiri oncoprotein STP-C488 J Virol 1991 65: 6953–6960

    CAS  PubMed  PubMed Central  Google Scholar 

  50. Jung JU et al. Identification of transforming genes of subgroup A and C strains of herpesvirus saimiri Proc Natl Acad Sci USA 1991 88: 7051–7055

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Jung JU, Desrosiers RC . Association of the viral oncoprotein STP-C488 with cellular ras Mol Cell Biol 1995 15: 6506–6512

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Murphy C et al. Epithelial tumours induced by a herpesvirus oncogene in transgenic mice Oncogene 1994 9: 221–226

    CAS  PubMed  Google Scholar 

  53. Duboise SM et al. Stp and Tip are essential for herpesvirus saimiri oncogenicity J Virol 1998 72: 1308–1313

    CAS  PubMed  PubMed Central  Google Scholar 

  54. Albrecht JC et al. Primary structure of the herpesvirus saimiri genome J Virol 1992 66: 5047–5058

    CAS  PubMed  PubMed Central  Google Scholar 

  55. Cannon JS et al. Human herpesvirus 8-encoded thymidine kinase and phosphotransferase homologues confer sensitivity to ganciclovir J Virol 1999 73: 4786–4793

    CAS  PubMed  PubMed Central  Google Scholar 

  56. Cazaux C et al. Phosphorylation and cytotoxicity of therapeutic nucleoside analogues: a comparison of alpha and gamma herpesvirus thymidine kinase suicide genes Cancer Gene Ther 1998 5: 83–91

    CAS  PubMed  Google Scholar 

  57. Knappe A et al. T-cell lymphoma caused by herpesvirus saimiri C488 independently of ie14/vsag, a viral gene with superantigen homology J Virol 1998 72: 3469–3471

    CAS  PubMed  PubMed Central  Google Scholar 

  58. Knappe A et al. The interleukin-17 gene of herpesvirus saimiri J Virol 1998 72: 5797–5801

    CAS  PubMed  PubMed Central  Google Scholar 

  59. Bush TG et al. Fulminant jejuno-ileitis following ablation of enteric glia in adult transgenic mice Cell 1998 93: 189–201

    Article  CAS  PubMed  Google Scholar 

  60. Caruso M et al. Regression of established macroscopic liver metastases after in situ transduction of a suicide gene Proc Natl Acad Sci USA 1993 90: 7024–7028

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Cohen JL et al. Prevention of graft-versus-host disease in mice using a suicide gene expressed in T lymphocytes Blood 1997 89: 4636–4645

    CAS  PubMed  Google Scholar 

  62. Smith JG et al. Intracranial administration of adenovirus expressing HSV-TK in combination with ganciclovir produces a dose-dependent, self-limiting inflammatory response Hum Gene Ther 1997 8: 943–954

    Article  CAS  PubMed  Google Scholar 

  63. Haack K et al. Detection of cytosine deaminase in genetically modified tumor cells by specific antibodies Hum Gene Ther 1997 8: 1395–1401

    Article  CAS  PubMed  Google Scholar 

  64. Kenney S, Ge JQ, Westphal EM, Olsen J . Gene therapy strategies for treating Epstein–Barr virus-associated lymphomas: comparison of two different Epstein–Barr virus-based vectors Hum Gene Ther 1998 9: 1131–1141

    Article  CAS  PubMed  Google Scholar 

  65. Mullen CA, Kilstrup M, Blaese RM . Transfer of the bacterial gene for cytosine deaminase to mammalian cells confers lethal sensitivity to 5-fluorocytosine: a negative selection system Proc Natl Acad Sci USA 1992 89: 33–37

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Rogers RP et al. Killing Epstein–Barr virus-positive B lymphocytes by gene therapy: comparing the efficacy of cytosine deaminase and herpes simplex virus thymidine kinase Hum Gene Ther 1996 7: 2235–2245

    Article  CAS  PubMed  Google Scholar 

  67. Nestler U et al. Foamy virus vectors for suicide gene therapy Gene Therapy 1997 4: 1270–1277

    Article  CAS  PubMed  Google Scholar 

  68. Riddell SR et al. T-cell mediated rejection of gene-modified HIV-specific cytotoxic T lymphocytes in HIV-infected patients Nature Med 1996 2: 216–223

    Article  CAS  PubMed  Google Scholar 

  69. Sauer B, Henderson N . Site-specific DNA recombination in mammalian cells by the Cre recombinase of bacteriophage P1 Proc Natl Acad Sci USA 1988 85: 5166–5170

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  70. Duboise SM, Guo J, Desrosiers RC, Jung JU . Use of virion DNA as a cloning vector for the construction of mutant and recombinant herpesviruses Proc Natl Acad Sci USA 1996 93: 11389–11394

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. McBurney MW et al. The mouse Pgk-1 gene promoter contains an upstream activator sequence Nucleic Acids Res 1991 19: 5755–5761

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This project was funded in parts by grants to HF from the Bundesministerium für Bildung und Forschung (Bonn), the Bayerische Forschungsstiftung (München), and the Wilhelm Sander-Stiftung (Neustadt/Donau), as well as by a joint grant to SS and HF by the German Israeli Foundation (Jerusalem). We are grateful to Kenji Imai (Neuherberg) for the tk vector pGEM7TKSal, to Michael Blaese (Bethesda) for the cd plasmid pCD2, to Karin Haack (Heidelberg) for a monoclonal antibody against CD, and to David Gower (Glaxo Welcome, Stevenage) for a rabbit TK antiserum. The authors thank Ralph Grassmann (Erlangen) for providing a series of useful plasmids, Edgar Meinl (Munich) for experimental advice, and Bernhard Fleckenstein (Erlangen) for continuous support.

Author information

Authors and Affiliations

  1. Institut für Klinische und Molekulare Virologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany

    C Hiller, S Wittmann & H Fickenscher

  2. Department of Bone Marrow Transplantation, Hadassah University Hospital, Jerusalem, Israel

    S Slavin

Authors
  1. C Hiller
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. S Wittmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. S Slavin
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. H Fickenscher
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hiller, C., Wittmann, S., Slavin, S. et al. Functional long-term thymidine kinase suicide gene expression in human T cells using a herpesvirus saimiri vector. Gene Ther 7, 664–674 (2000). https://doi.org/10.1038/sj.gt.3301158

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.gt.3301158

Keywords

This article is cited by

Search

Advanced search

Quick links