Ex Vivo Staining of Metastatic Lymph Nodes by Class I Major Histocompatibility Complex Tetramers Reveals High Numbers of Antigen-experienced Tumor-specific Cytolytic T Lymphocytes

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  • Pedro Romero
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • P. Rod Dunbar
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Danila Valmori
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Mikaël Pittet
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Graham S. Ogg
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Donata Rimoldi
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Ji-Li Chen
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Danielle Liénard
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Jean-Charles Cerottini
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom
  • Vincenzo Cerundolo
    From the *Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, and the ‡Multidisciplinary Oncology Center, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland; and the §Institute of Molecular Medicine, Nuffield Department of Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom

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<jats:p>Characterization of cytolytic T lymphocyte (CTL) responses to tumor antigens has been impeded by a lack of direct assays of CTL activity. We have synthesized reagents (“tetramers”) that specifically stain CTLs recognizing melanoma antigens. Tetramer staining of tumor-infiltrated lymph nodes ex vivo revealed high frequencies of tumor-specific CTLs which were antigen-experienced by surface phenotype. In vitro culture of lymph node cells with cytokines resulted in very large expansions of tumor-specific CTLs that were dependent on the presence of tumor cells in the lymph nodes. Tetramer-guided sorting by flow cytometer allowed isolation of melanoma-specific CTLs and confirmation of their specificity and their ability to lyse autologous tumor cells. Our results demonstrate the value of these novel reagents for monitoring tumor-specific CTL responses and for generating CTLs for adoptive immunotherapy. These data also indicate that strong CTL responses to melanoma often occur in vivo, and that the reactive CTLs have substantial proliferative and tumoricidal potential.</jats:p>

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