title: Identification of known and novel immunogenic T-cell epitopes from tumor antigens recognized by peripheral blood T cells from patients responding to IL-2-based treatment.
abstract: In previous studies CD8+ T cells specific for melanocyte antigens have been frequently found in melanoma patients responding to interleukin-2 (IL-2)-based therapies. In our study we analyzed the suitability of using circulating T cells from melanoma patients with clinical response after IL-2-based therapy to identify novel T-cell epitopes from defined tumor antigens. Using unstimulated peripheral blood mononuclear cells and the interferon-gamma (IFN-gamma) ELISPOT assay, we studied CD8(+) T-cell responses against 5 peptides from the tumor antigen tyrosinase (Tyr) selected by epitope prediction using an HLA-A1-binding computer algorithm. T cells specifically secreting IFN-gamma in response to 3 of these 5 peptides, namely, Tyr (454-463), Tyr (146-156) and Tyr (243-251), could be detected in 4 of 4 HLA-A1-positive patients with clinical response. In contrast, no T-cell responses against these peptides were seen in 6 HLA-A1-positive melanoma patients with progressive disease and in 8 healthy subjects. We could generate specific cytotoxic T lymphocytes (CTL) against Tyr (454-463) using peptide-pulsed autologous dendritic cells as antigen-presenting cells. The induced CTLs efficiently killed melanoma cells that express HLA-A1 and tyrosinase. The peptides Tyr (146-156) and Tyr (243-251) had recently been identified as CTL epitopes by other groups. Further ex vivo characterization of the T cells reactive against the novel epitope Tyr (454-463) in 1 patient by multicolor flow cytometry showed specific CD3+/CD8+/IFN-gamma+ T cells with frequencies of up to 0.41% of the CD3+/CD8+ T-cell population. Most of this T-cell population also expressed granzyme B. Our data confirm that in patients with tumor regressions induced by immunotherapy or chemoimmunotherapy circulating T cells reactive with tyrosinase epitopes can frequently be detected. Peripheral blood T cells from such patients are a valuable source for screening peptides selected by epitope prediction This strategy facilitates the rapid identification of immunogenic T-cell epitopes that are probable targets of immune-mediated tumor rejection.
authors.affiliation: Medizinische Klinik III, Hematology, Oncology and Transfusion Medicine, Universitaetsklinikum Benjamin Franklin, Freie Universität Berlin, Berlin, Germany. scheibenbogen@ukfb.fu-berlin.de
authors.forename: Carmen
authors.initials: C
authors.lastname: Scheibenbogen
authors.forename: Yuansheng
authors.initials: Y
authors.lastname: Sun
authors.forename: Ulrich
authors.initials: U
authors.lastname: Keilholz
authors.forename: Mingxia
authors.initials: M
authors.lastname: Song
authors.forename: Stefan
authors.initials: S
authors.lastname: Stevanovic
authors.forename: Anne Marie
authors.initials: AM
authors.lastname: Asemissen
authors.forename: Dirk
authors.initials: D
authors.lastname: Nagorsen
authors.forename: Eckhard
authors.initials: E
authors.lastname: Thiel
authors.forename: Hans-Georg
authors.initials: HG
authors.lastname: Rammensee
authors.forename: Dirk
authors.initials: D
authors.lastname: Schadendorf
mesh_terms: D000951:Antigens, Neoplasm / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D016207:Cytokines / Q000378:metabolism; D003601:Cytotoxicity Tests, Immunologic; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D018984:Epitopes, T-Lymphocyte / Q000276:immunology*; D005434:Flow Cytometry; D015788:HLA-A1 Antigen / Q000276:immunology; D015789:HLA-A2 Antigen / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology; D007376:Interleukin-2 / Q000627:therapeutic use*; D008545:Melanoma / Q000097:blood / Q000628:therapy*; D012878:Skin Neoplasms / Q000097:blood / Q000628:therapy*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D014407:Tumor Cells, Cultured / Q000276:immunology / Q000378:metabolism
chemical_list: D000951:Antigens, Neoplasm; D016207:Cytokines; D018984:Epitopes, T-Lymphocyte; D015788:HLA-A1 Antigen; D015789:HLA-A2 Antigen; D007376:Interleukin-2; D007371:Interferon-gamma