title: Detection and functional analysis of CD8+ T cells specific for PRAME: a target for T-cell therapy.
abstract: OBJECTIVE
  Preferentially expressed antigen on melanomas (PRAME) is an interesting antigen for T-cell therapy because it is frequently expressed in melanomas (95%) and other tumor types. Moreover, due to its role in oncogenic transformation, PRAME-negative tumor cells are not expected to easily arise and escape from T-cell immunity. The purpose of this study is to investigate the usefulness of PRAME as target for anticancer T-cell therapies.
  
  
  METHODS
  HLA-A*0201-subtyped healthy individuals and advanced melanoma patients were screened for CD8+ T cells directed against previously identified HLA-A*0201-binding PRAME peptides by IFN-gamma enzyme-linked immunosorbent spot assays and tetramer staining. PRAME-specific T-cell clones were isolated and tested for recognition of melanoma and acute lymphoid leukemia (ALL) cell lines. PRAME mRNA expression was determined by quantitative real-time reverse transcription-PCR.
  
  
  RESULTS
  In 30% to 40% of healthy individuals and patients, PRA(100-108)-specific CD8+ T cells were detected both after in vitro stimulation and directly ex vivo after isolation by magnetic microbeads. Although CD45RA- memory PRA(100-108)-specific T cells were found in some individuals, the majority of PRA(100-108)-tetramer+ T cells expressed CD45RA, suggesting a naive phenotype. PRA(100-108)-tetramer+ T-cell clones were shown to recognize and lyse HLA-A*0201+ and PRAME+ melanoma but not ALL cell lines. Quantitative real-time reverse transcription-PCR showed significantly lower PRAME mRNA levels in ALL than in melanoma cell lines, suggesting that PRAME expression in ALL is below the recognition threshold of our PRA(100-108)-tetramer+ T cells.
  
  
  CONCLUSIONS
  These data support the usefulness of PRAME and in particular the PRA(100-108) epitope as target for T-cell therapy of PRAME-overexpressing cancers.
authors.affiliation: Department of Clinical Oncology, Leiden University Medical Center, Leiden, Netherlands. M.Griffioen@lumc.nl
authors.forename: Marieke
authors.initials: M
authors.lastname: Griffioen
authors.forename: Jan H
authors.initials: JH
authors.lastname: Kessler
authors.forename: Martina
authors.initials: M
authors.lastname: Borghi
authors.forename: Ronald A
authors.initials: RA
authors.lastname: van Soest
authors.forename: Caroline E
authors.initials: CE
authors.lastname: van der Minne
authors.forename: Jan
authors.initials: J
authors.lastname: Nouta
authors.forename: Sjoerd H
authors.initials: SH
authors.lastname: van der Burg
authors.forename: Jan Paul
authors.initials: JP
authors.lastname: Medema
authors.forename: Peter I
authors.initials: PI
authors.lastname: Schrier
authors.forename: J H Frederik
authors.initials: JH
authors.lastname: Falkenburg
authors.forename: Susanne
authors.initials: S
authors.lastname: Osanto
authors.forename: Cornelis J M
authors.initials: CJ
authors.lastname: Melief
mesh_terms: D000951:Antigens, Neoplasm / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D015234:HLA-A Antigens / Q000276:immunology; D015789:HLA-A2 Antigen; D006801:Humans; D007167:Immunotherapy / Q000379:methods; D008545:Melanoma / Q000276:immunology* / Q000628:therapy; D012878:Skin Neoplasms / Q000276:immunology* / Q000628:therapy; D014407:Tumor Cells, Cultured
chemical_list: D000951:Antigens, Neoplasm; D000939:Epitopes; D015234:HLA-A Antigens; C435939:HLA-A*02:01 antigen; D015789:HLA-A2 Antigen; C404944:PRAME protein, human