title: Quantitative global and gene-specific promoter methylation in relation to biological properties of neuroblastomas.
abstract: BACKGROUND
  In this study we aimed to quantify tumor suppressor gene (TSG) promoter methylation densities levels in primary neuroblastoma tumors and cell lines. A subset of these TSGs is associated with a CpG island methylator phenotype (CIMP) in other tumor types.
  
  
  METHODS
  The study panel consisted of 38 primary tumors, 7 established cell lines and 4 healthy references. Promoter methylation was determined by bisulphate Pyrosequencing for 14 TSGs; and LINE-1 repeat element methylation was used as an indicator of global methylation levels.
  
  
  RESULTS
  Overall mean TSG Z-scores were significantly increased in cases with adverse outcome, but were unrelated to global LINE-1 methylation. CIMP with hypermethylation of three or more gene promoters was observed in 6/38 tumors and 7/7 cell lines. Hypermethylation of one or more TSG (comprising TSGs BLU, CASP8, DCR2, CDH1, RASSF1A and RASSF2) was evident in 30/38 tumors. By contrast only very low levels of promoter methylation were recorded for APC, DAPK1, NORE1A, P14, P16, TP73, PTEN and RARB. Similar involvements of methylation instability were revealed between cell line models and neuroblastoma tumors. Separate analysis of two proposed CASP8 regulatory regions revealed frequent and significant involvement of CpG sites between exon 4 and 5, but modest involvement of the exon 1 region.
  
  
  CONCLUSIONS
  The results highlight the involvement of TSG methylation instability in neuroblastoma tumors and cell lines using quantitative methods, support the use of DNA methylation analyses as a prognostic tool for this tumor type, and underscore the relevance of developing demethylating therapies for its treatment.
authors.affiliation: Departments of Molecular Medicine and Surgery, Stockholm, Sweden.
authors.forename: Nimrod B
authors.initials: NB
authors.lastname: Kiss
authors.forename: Per
authors.initials: P
authors.lastname: Kogner
authors.forename: John Inge
authors.initials: JI
authors.lastname: Johnsen
authors.forename: Tommy
authors.initials: T
authors.lastname: Martinsson
authors.forename: Catharina
authors.initials: C
authors.lastname: Larsson
authors.forename: Janos
authors.initials: J
authors.lastname: Geli
mesh_terms: D001483:Base Sequence; D053181:Caspase 8 / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D018899:CpG Islands; D019175:DNA Methylation; D005091:Exons; D006801:Humans; D053208:Kaplan-Meier Estimate; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000235:genetics; D009447:Neuroblastoma / Q000235:genetics* / Q000401:mortality; D010641:Phenotype; D011401:Promoter Regions, Genetic; D025521:Tumor Suppressor Proteins / Q000235:genetics
chemical_list: D025521:Tumor Suppressor Proteins; D053181:Caspase 8