title: Activation of tumor-specific T cells by dendritic cells expressing the NY-ESO-1 antigen after transfection with the cationic lipophosphoramide KLN5.
abstract: BACKGROUND
  Genetic modification of human monocyte-derived dendritic cells (DC) with cDNA sequences encoding tumor-associated antigens (TAA) is a promising strategy for cancer immunotherapy. The present study aimed to develop a nonviral gene transfer method based on the use of the cationic lipophosphoramide reagent, KLN-5, as an alternative to the commonly used viral vectors.
  
  
  METHODS
  First, the efficiency of KLN5 for gene transfection into DC was investigated using the green fluorescent protein (GFP) reporter gene. The highest transfection efficiency/cell viability ratio was determined by flow cytometry. Next, DC were transfected with a plasmid encoding NY-ESO-1, a TAA expressed in numerous cancers, according to the transfection protocol previously established with the GFP reporter. Transfected DC were then co-cultured with a CD8+ NY-ESO-1 specific HLA-A*02.01 T cell clone to control their ability to correctly process and present the corresponding epitope in the HLA-A*02.01 context. Finally, T cell activation was assessed via flow cytometry-based detection of interferon-gamma production.
  
  
  RESULTS
  An optimal KLN5/plasmid DNA ratio allowing both significant transgene expression and high viability of DC could be determined. Under the established experimental conditions, antigen processing and presentation of the immunodominant (SLLMWITQC(157-165)) epitope in the HLA-A*0201 context was demonstrated by activation of the NY-ESO-1-specific CD8+ T cell clone.
  
  
  CONCLUSIONS
  KLN5-based gene transfection into DC allows the efficient induction of TAA presentation and may thus represent a novel attractive nonviral approach for cancer vaccination.
authors.affiliation: Université de Rennes 1, Faculté de Médecine, UPRES 3891, Rennes, France.
authors.forename: Matthieu
authors.initials: M
authors.lastname: Le Gallo
authors.forename: Olivier
authors.initials: O
authors.lastname: Toutirais
authors.forename: Tristan
authors.initials: T
authors.lastname: Montier
authors.forename: Florian
authors.initials: F
authors.lastname: Cabillic
authors.forename: Françoise
authors.initials: F
authors.lastname: Bouet
authors.forename: Pascal
authors.initials: P
authors.lastname: Delépine
authors.forename: Pierre
authors.initials: P
authors.lastname: Lehn
authors.forename: Francine
authors.initials: F
authors.lastname: Jotereau
authors.forename: Véronique
authors.initials: V
authors.lastname: Catros
mesh_terms: D000951:Antigens, Neoplasm / Q000378:metabolism*; D003713:Dendritic Cells / Q000276:immunology*; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins; D006801:Humans; D007167:Immunotherapy / Q000379:methods*; D007371:Interferon-gamma / Q000378:metabolism; D008213:Lymphocyte Activation / Q000276:immunology*; D008565:Membrane Proteins / Q000378:metabolism*; D009369:Neoplasms / Q000628:therapy*; D009943:Organophosphorus Compounds / Q000627:therapeutic use*; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: D000951:Antigens, Neoplasm; C104948:CTAG1B protein, human; C493328:KLN-5 compound; D008565:Membrane Proteins; D009943:Organophosphorus Compounds; D049452:Green Fluorescent Proteins; D007371:Interferon-gamma