title: CD4(+)and CD8(+)T-cell reactions against leukemia-associated- or minor-histocompatibility-antigens in AML-patients after allogeneic SCT.
abstract: T-cells play an important role in the remission-maintenance in AML-patients (pts) after SCT, however the role of LAA- (WT1, PR1, PRAME) or minor-histocompatibility (mHag, HA1) antigen-specific CD4(+) and CD8(+)T-cells is not defined. A LAA/HA1-peptide/protein stimulation, cloning and monitoring strategy for specific CD8(+)/CD4(+)T-cells in AML-pts after SCT is given. Our results show that (1) LAA-peptide-specific CD8+T-cells are detectable in every AML-pt after SCT. CD8(+)T-cells, recognizing two different antigens detectable in 5 of 7 cases correlate with long-lasting remissions. Clonal TCR-Vβ-restriction exemplarily proven by spectratyping in PRAME-specific CD8(+)T-cells; high PRAME-peptide-reactivity was CD4(+)-associated, as shown by IFN-γ-release. (2) Two types of antigen-presenting cells (APCs) were tested for presentation of LAA/HA1-proteins to CD4(+)T-cells: miniEBV-transduced lymphoblastoid cells (B-cell-source) and CD4-depleted MNC (source for B-cell/monocyte/DC). We provide a refined cloning-system for proliferating, CD40L(+)CD4(+)T-cells after LAA/HA1-stimulation. CD4(+)T-cells produced cytokines (GM-CSF, IFN-γ) upon exposure to LAA/HA1-stimulation until after at least 7 restimulations and demonstrated cytotoxic activity against naive blasts, but not fibroblasts. Antileukemic activity of unstimulated, stimulated or cloned CD4(+)T-cells correlated with defined T-cell-subtypes and the clinical course of the disease. In conclusion we provide immunological tools to enrich and monitor LAA/HA1-CD4(+)- and CD8(+)T-cells in AML-pts after SCT and generate data with relevant prognostic value. We were able to demonstrate the presence of LAA-peptide-specific CD8(+)T-cell clones in AML-pts after SCT. In addition, we were also able to enrich specific antileukemic reactive CD4(+)T-cells without GvH-reactivity upon repeated LAA/HA1-protein stimulation and limiting dilution cloning.
authors: Helmholtz Center Munich (German Research Center for Environmental Health and Clinical Cooperative Group Hematopoetic Cell-Transplantation), 81377 Munich, Germany; University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Brigitte
authors: B
authors: Steger
authors: Helmholtz Center Munich (German Research Center for Environmental Health and Clinical Cooperative Group Hematopoetic Cell-Transplantation), 81377 Munich, Germany.
authors: Slavoljub
authors: S
authors: Milosevic
authors: Institute for Medical Microbiology, Immunology and Hygiene, and Focus Group'Clinical Cell Processing and Purification', Institute for Advanced Study, Technical University Munich, 81675 Munich, Germany.
authors: Georg
authors: G
authors: Doessinger
authors: Department of Pediatric Oncology, Hematology and Immunology, University Hospital Duesseldorf, 40225 Duesseldorf, Germany.
authors: Susanne
authors: S
authors: Reuther
authors: University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Anja
authors: A
authors: Liepert
authors: Helmholtz Center Munich (German Research Center for Environmental Health and Clinical Cooperative Group Hematopoetic Cell-Transplantation), 81377 Munich, Germany.
authors: Marion
authors: M
authors: Braeu
authors: University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Julia
authors: J
authors: Schick
authors: University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Valentin
authors: V
authors: Vogt
authors: Department of Pediatric Oncology, Hematology and Immunology, University Hospital Duesseldorf, 40225 Duesseldorf, Germany.
authors: Friedhelm
authors: F
authors: Schuster
authors: University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Tanja
authors: T
authors: Kroell
authors: Institute for Medical Microbiology, Immunology and Hygiene, and Focus Group'Clinical Cell Processing and Purification', Institute for Advanced Study, Technical University Munich, 81675 Munich, Germany; Clinical Cooperation Groups "Antigen-specific Immunotherapy" and "Immune Monitoring", Helmholtz Center Munich and Technical University Munich, 81675 Munich, Germany.
authors: Dirk H
authors: DH
authors: Busch
authors: Department of Pediatric Oncology, Hematology and Immunology, University Hospital Duesseldorf, 40225 Duesseldorf, Germany.
authors: Arndt
authors: A
authors: Borkhardt
authors: Helmholtz Center Munich (German Research Center for Environmental Health and Clinical Cooperative Group Hematopoetic Cell-Transplantation), 81377 Munich, Germany; University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Hans-Jochem
authors: HJ
authors: Kolb
authors: University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Johanna
authors: J
authors: Tischer
authors: Helmholtz Center Munich (German Research Center for Environmental Health and Clinical Cooperative Group Hematopoetic Cell-Transplantation), 81377 Munich, Germany; University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany.
authors: Raymund
authors: R
authors: Buhmann
authors: Helmholtz Center Munich (German Research Center for Environmental Health and Clinical Cooperative Group Hematopoetic Cell-Transplantation), 81377 Munich, Germany; University Hospital of Munich, Department for Hematopoetic Cell Transplantation, Med. Dept. 3, 81377 Munich, Germany. Electronic address: Helga.Schmetzer@med.uni-muenchen.de.
authors: Helga
authors: H
authors: Schmetzer
mesh_terms: D000951:Antigens, Neoplasm / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D023201:CD40 Ligand / Q000378:metabolism; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D018929:Cell Culture Techniques; D002999:Clone Cells; D003602:Cytotoxicity, Immunologic; D005260:Female; D020544:Graft vs Leukemia Effect / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D015470:Leukemia, Myeloid, Acute / Q000276:immunology* / Q000628:therapy*; D008213:Lymphocyte Activation; D008297:Male; D015778:Minor Histocompatibility Antigens / Q000276:immunology; D009367:Neoplasm Staging; D010446:Peptide Fragments / Q000276:immunology; D033581:Stem Cell Transplantation; D014184:Transplantation, Homologous; D025721:WT1 Proteins / Q000276:immunology
chemical_list: D000951:Antigens, Neoplasm; D015778:Minor Histocompatibility Antigens; C404944:PRAME protein, human; D010446:Peptide Fragments; D025721:WT1 Proteins; C583751:WT1 protein, human; D023201:CD40 Ligand; D007371:Interferon-gamma
keywords: Acute myeloid leukemia; CD4/CD8 T-cells; Dendritic cells; Immunophenotyping; Minor antigens; Multimer; TAA