title: Frequency of circulating Tregs with demethylated FOXP3 intron 1 in melanoma patients receiving tumor vaccines and potentially Treg-depleting agents.
abstract: OBJECTIVE
  Regulatory T cells (Tregs) are thought to inhibit antitumor immune responses, and their depletion could therefore have a synergistic effect with therapeutic cancer vaccines. We investigated the impact of three medications on blood Treg frequency in vaccinated cancer patients.
  
  
  METHODS
  To date, the most specific marker for human Tregs is demethylation in the DNA that encodes the transcription factor FOXP3. Thus, we used a FOXP3 methylation-specific quantitative PCR assay (MS-qPCR) to measure Treg frequencies in the peripheral blood mononuclear cells (PBMCs) of melanoma patients. The patients participated in three clinical trials that combined tumor vaccines with potential Treg-depleting agents: low-dose cyclophosphamide, anti-CD25 monoclonal antibody daclizumab, and the IL-2/diphtheria toxin fusion protein denileukin diftitox.
  
  
  RESULTS
  In the nine control patients, blood Treg frequencies varied over time; there was a 46% reduction in one patient. In treated patients, a more than 2-fold decrease in Tregs was observed in one out of 11 patients receiving cyclophosphamide and in four out of 13 receiving daclizumab, but there was no such Treg decrease in any of the six patients who received denileukin diftitox. As a positive control, a more than 2-fold increase in blood Tregs was detected in four out of nine patients who were treated with interleukin-2.
  
  
  CONCLUSIONS
  We used a MS-qPCR method that detects Tregs but not other activated T lymphocytes; however, none of the Treg-depleting strategies that we tested led, in the majority of patients, to a conservative 50% reduction in blood Tregs.
authors.affiliation: Department of Medical Oncology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, The Netherlands.
authors.forename: I Jolanda M
authors.initials: IJ
authors.lastname: de Vries
authors.forename: Chiara
authors.initials: C
authors.lastname: Castelli
authors.forename: Caroline
authors.initials: C
authors.lastname: Huygens
authors.forename: Joannes F M
authors.initials: JF
authors.lastname: Jacobs
authors.forename: Julie
authors.initials: J
authors.lastname: Stockis
authors.forename: Beatrice
authors.initials: B
authors.lastname: Schuler-Thurner
authors.forename: Gosse J
authors.initials: GJ
authors.lastname: Adema
authors.forename: Cornelis J A
authors.initials: CJ
authors.lastname: Punt
authors.forename: Licia
authors.initials: L
authors.lastname: Rivoltini
authors.forename: Gerold
authors.initials: G
authors.lastname: Schuler
authors.forename: Pierre G
authors.initials: PG
authors.lastname: Coulie
authors.forename: Sophie
authors.initials: S
authors.lastname: Lucas
mesh_terms: D000911:Antibodies, Monoclonal / Q000627:therapeutic use; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents / Q000627:therapeutic use; D019496:Cancer Vaccines / Q000627:therapeutic use*; D002452:Cell Count; D003520:Cyclophosphamide / Q000627:therapeutic use; D019175:DNA Methylation; D000077561:Daclizumab; D004167:Diphtheria Toxin / Q000627:therapeutic use; D005260:Female; D051858:Forkhead Transcription Factors / Q000235:genetics*; D005819:Genetic Markers / Q000235:genetics; D006801:Humans; D007074:Immunoglobulin G / Q000627:therapeutic use; D007376:Interleukin-2 / Q000627:therapeutic use; D007438:Introns; D008212:Lymphocyte Depletion; D008297:Male; D008545:Melanoma / Q000276:immunology / Q000473:pathology* / Q000628:therapy; D011993:Recombinant Fusion Proteins / Q000627:therapeutic use; D012189:Retrospective Studies; D050378:T-Lymphocytes, Regulatory / Q000378:metabolism* / Q000473:pathology
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D019496:Cancer Vaccines; D004167:Diphtheria Toxin; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D005819:Genetic Markers; D007074:Immunoglobulin G; D007376:Interleukin-2; D011993:Recombinant Fusion Proteins; C078456:denileukin diftitox; D003520:Cyclophosphamide; D000077561:Daclizumab