title: Humoral anti-KLH responses in cancer patients treated with dendritic cell-based immunotherapy are dictated by different vaccination parameters.
abstract: OBJECTIVE
  Keyhole limpet hemocyanin (KLH) attracts biomedical interest because of its remarkable immunostimulatory properties. Currently, KLH is used as vaccine adjuvant, carrier protein for haptens and as local treatment for bladder cancer. Since a quantitative human anti-KLH assay is lacking, it has not been possible to monitor the dynamics of KLH-specific antibody (Ab) responses after in vivo KLH exposure. We designed a quantitative assay to measure KLH-specific Abs in humans and retrospectively studied the relation between vaccination parameters and the vaccine-induced anti-KLH Ab responses.
  
  
  METHODS
  Anti-KLH Abs were purified from pooled serum of melanoma patients who have responded to KLH as a vaccine adjuvant. Standard isotype-specific calibration curves were generated to measure KLH-specific Ab responses in individual serum samples using ELISA.
  
  
  RESULTS
  KLH-specific IgM, IgA, IgG and all IgG-subclasses were accurately measured at concentrations as low as 20 μg/ml. The intra- and inter-assay coefficients of variation of this ELISA were below 6.7 and 9.9 %, respectively. Analyses of 128 patients demonstrated that mature DC induced higher levels of KLH-specific IgG compared to immature DC, prior infusion with anti-CD25 abolished IgG and IgM production and patients with locoregional disease developed more robust IgG responses than advanced metastatic melanoma patients.
  
  
  CONCLUSIONS
  We present the first quantitative assay to measure KLH-specific Abs in human serum, which now enables monitoring both the dynamics and absolute concentrations of humoral immune responses in individuals exposed to KLH. This assay may provide a valuable biomarker for the immunogenicity and clinical effectiveness of KLH-containing vaccines and therapies.
authors.affiliation: Department of Medical Oncology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.
authors.forename: Erik H J G
authors.initials: EH
authors.lastname: Aarntzen
authors.forename: I Jolanda M
authors.initials: IJ
authors.lastname: de Vries
authors.forename: Joop H
authors.initials: JH
authors.lastname: Göertz
authors.forename: Marjo
authors.initials: M
authors.lastname: Beldhuis-Valkis
authors.forename: Huberdina M L M
authors.initials: HM
authors.lastname: Brouwers
authors.forename: Mandy W M M
authors.initials: MW
authors.lastname: van de Rakt
authors.forename: Renate G
authors.initials: RG
authors.lastname: van der Molen
authors.forename: Cornelis J A
authors.initials: CJ
authors.lastname: Punt
authors.forename: Gosse J
authors.initials: GJ
authors.lastname: Adema
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Tacken
authors.forename: Irma
authors.initials: I
authors.lastname: Joosten
authors.forename: Joannes F M
authors.initials: JF
authors.lastname: Jacobs
mesh_terms: D000276:Adjuvants, Immunologic / Q000627:therapeutic use; D000906:Antibodies / Q000097:blood*; D019496:Cancer Vaccines / Q000276:immunology* / Q000627:therapeutic use; D003713:Dendritic Cells / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006433:Hemocyanins / Q000276:immunology*; D006801:Humans; D056724:Immunity, Humoral; D007167:Immunotherapy; D008297:Male; D008545:Melanoma / Q000276:immunology / Q000628:therapy*; D015203:Reproducibility of Results; D012189:Retrospective Studies
chemical_list: D000276:Adjuvants, Immunologic; D000906:Antibodies; D019496:Cancer Vaccines; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin