Cancer Immunology, Immunotherapy

, Volume 39, Issue 2, pp 93–99 | Cite as

Frequency analysis of tumor-reactive cytotoxic T lymphocytes in peripheral blood of a melanoma patient vaccinated with autologous tumor cells

  • Wolfgang Herr
  • Thomas Wölfel
  • Michael Heike
  • Karl-Hermann Meyer zum Büschenfelde
  • Alexander Knuth
Original Articles Human Melanoma, Limiting-Dilution Assay, Tumor-Reactive CTL, Tumor Cell Vaccination


A limiting-dilution assay was developed and used to determine the frequency of autologous tumor-reactive cytotoxic T lymphocytes (CTL) in peripheral blood of a melanoma patient MZ2, who has been free of detectable disease since several years. In this patient, the frequencies of tumor-reactive CTL spontaneously varied only by a factor of 1.5. After vaccinations with autologous mutagenized and lethally irradiated tumor cells a two- to tenfold increase in frequencies of tumor-reactive CTL was found within the first 2 weeks. Thereafter, CTL frequencies returned to values measured prior to vaccinations. We conclude, that the limiting-dilution assay applied in this study can detect changes in the T cell response to autologous tumor cells. The frequency of tumor-reactive CTL determined with this approach can serve as an immunological parameter for monitoring the T cell response to autologous tumor cells in individual cancer patients receiving tumor cell vaccinations.

Key words

Human melanoma Limiting-dilution assay Frequency of tumor-reactive CTL Tumor cell vaccination 


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  1. 1.
    Boon T, Kellermann O (1977) Rejection by syngeneic mice of cell variants obtained by mutagenesis of a malignant teratocarcinoma cell line. Proc Natl Acad Sci USA 74: 272PubMedGoogle Scholar
  2. 2.
    Borysiewicz LK, Graham S, Hickling JK, Mason PD, Sissons JGP (1988) Human cytomegalovirus-specific cytotoxic T cells: their precursor frequency and stage specificity. Eur J Immunol 18: 269PubMedGoogle Scholar
  3. 3.
    Brasseur F, Marchand M, Vanwijck R, Herin M, Chomez P, Boon T (1992) Human gene MAGE-1, which codes for a tumor-rejection antigen, is expressed by some breast tumors. Int J Cancer 52: 839PubMedGoogle Scholar
  4. 4.
    Brunner KT, Engers HD, Cerottini J-C (1976) In vitro methods in cell-mediated and tumor immunity. In: Bloom BR, David JR (eds) In vitro methods in cell-mediated and tumor immunity. Academic Press, New York, p 423Google Scholar
  5. 5.
    Coulic PG, Van Pel A (1993) Defined antigens recognized by T lymphocytes on human tumors. Curr Opin Oncol 5: 1043PubMedGoogle Scholar
  6. 6.
    Coulie PG, Somville M, Lehmann F, Hainaut P, Brasseur RD, Devos R, Boon T (1992) Precursor frequency analysis of human cytolytic T lymphocytes directed against autologous melanoma cells. Int J Cancer 50: 289PubMedGoogle Scholar
  7. 7.
    Degiovanni G, Lahaye T, Herin M, Hainaut P, Boon T (1988) Antigenic heterogeneity of human melanoma tumor detected by autologous CTL clones. Eur J Immunol 18: 671PubMedGoogle Scholar
  8. 8.
    Herin M, Lemoine C, Weynants P, Vessiere F, Van Pel A, Knuth A, Devos R, Boon T (1987) Production of stable cytolytic T cell clones directed against autologous human melanoma. Int J Cancer 39: 390PubMedGoogle Scholar
  9. 9.
    Herzog W-R, Zanker B, Irschick E, Huber C, Franz H, Wagner H, Kabelitz D (1987) Selective reduction of donor-specific cytotoxic T lymphocyte precursors in patients with a well-functional kidney allograft. Transplantation 43: 384PubMedGoogle Scholar
  10. 10.
    Hoffenbach A, Langlase-Demoyen P, Dadaglio G, Vilmer E, Michel F, Mayand C (1989) Unusually high frequencies of HIV-specific cytotoxic T lymphocytes in humans. J Immunol 142: 452PubMedGoogle Scholar
  11. 11.
    Ioannides CG, Freedman RS, Platsoucas CD, Rashed S, Kim YP (1991) Cytotoxic T cell clones isolated from ovarian tumor-infiltrating lymphocytes recognize multiple antigeneic epitopes on autologous tumor cells. J Immunol 146: 1700PubMedGoogle Scholar
  12. 12.
    Kabelitz D, Herzog W-R, Zanker B, Wagner H (1985) Human cytotoxic T lymphocytes. I. Limiting dilution analysis of alloreactive cytotoxic T-lymphocyte precursor frequencies. Scand J Immunol 22: 329PubMedGoogle Scholar
  13. 13.
    Knuth A, Wölfel T, Klehmann E, Boon T, Meyer zum Büschenfelde K-H (1989) Cytolytic T-cell clones against an autologous malignant human melanoma: specificity study and definition of three antigens by immunoselection. Proc Natl Acad Sci USA 86: 2804PubMedGoogle Scholar
  14. 14.
    Knuth A, Wölfel T, Meyer zum Büschenfelde K-H (1991) Cellular and humoral responses against cancer: implications for cancer vaccines. Curr Opin Immunol 3: 659PubMedGoogle Scholar
  15. 15.
    Knuth A, Wölfel T, Meyer zum Büschenfelde K-H (1992) T cell responses to human malignant tumors. Cancer Surv 13: 39PubMedGoogle Scholar
  16. 16.
    Moretta A, Pantaleo G, Moretta L, Mingari M, Cerottini J-C (1983) Quantitative assessment of the pool size and subset distribution of cytolytic T lymphocytes within human resting or alloactivated peripheral blood T cell populations. J Exp Med 158: 571PubMedGoogle Scholar
  17. 17.
    Parmiani G, Anichini A, Fossati G (1990) Cellular immune response against autologous human malignant melanoma: are in vitro studies providing a framework for a more effective immunotherapy? J Natl Cancer Inst 82: 361PubMedGoogle Scholar
  18. 18.
    Sharrock CEM, Man S, Wanachiwanawin W, Batchelor JR (1987) Analysis of the alloreactive T cell repertoire in man. Transplantation 43: 699PubMedGoogle Scholar
  19. 19.
    Sharrock CEM, Kaminsky E, Man S (1990) Limiting dilution analysis of human T cells: a useful clinical tool. Immunol Today 11: 281PubMedGoogle Scholar
  20. 20.
    Taswell C (1981) Limiting dilution assays for the determination of immunocompetent cell frequencies. I. Data Analysis. J Immunol 126: 1614PubMedGoogle Scholar
  21. 21.
    Taswell C, MacDonald R, Cerottini J-C (1979) Limiting dilution analysis of alloantigen-reactive T lymphocytes. II. Effect of cortisone and cyclophosphamide on cytolytic T lymphocyte precursor frequencies in the thymus. Thymus 1: 119PubMedGoogle Scholar
  22. 22.
    Van den Eynde B, Hainaut P, Herin M, Knuth A, Lemoine C, Weynants P, Van der Bruggen P, Fauchet R, Boon T (1989) Presence on a human melanoma of multiple antigens recognized by autologous CTL. Int J Cancer 44: 634PubMedGoogle Scholar
  23. 23.
    Van der Bruggen P, Traversari C, Chomez P, Lurquin C, De Plaen E, Van den Eynde B, Knuth A, Boon T (1991) A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science 254: 1643PubMedGoogle Scholar
  24. 24.
    Wölfel T, Klehmann E, Müller C, Schütt K-H, Meyer zum Büschenfelde K-H, Knuth A (1989) Lysis of human melanoma cells by autologous cytolytic T cell clones. Identification of human histocompatibility leukocyte antigen A2 as a restriction element for three different antigens. J Exp Med 170: 797PubMedGoogle Scholar
  25. 25.
    Zhang L, Guang Li S, Vandekereckhove B, Termijtelen A, Van Rood JJ, Class FHJ (1989) Analyses of cytotoxic T cell precursor frequencies directed against individual HLA-A and-B allo-antigens. J Immunol Methods 121: 39PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1994

Authors and Affiliations

  • Wolfgang Herr
    • 1
  • Thomas Wölfel
    • 1
  • Michael Heike
    • 1
  • Karl-Hermann Meyer zum Büschenfelde
    • 1
  • Alexander Knuth
    • 2
  1. 1.I. Medizinische Klinik und Poliklinik der Johannes Gutenberg-Universität MainzMainzGermany
  2. 2.II. Medizinische Klinik, Krankenhaus NordwestFrankfurt am MainGermany

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