title: Methylation of a novel panel of tumor suppressor genes in urine moves forward noninvasive diagnosis and prognosis of bladder cancer: a 2-center prospective study.
abstract: OBJECTIVE
  Changes in DNA methylation of tumor suppressor genes early in carcinogenesis represent potential indicators of cancer detection and disease evolution. We examined the diagnostic, stratification and prognostic biomarker roles in urine of the methylation of a novel panel of tumor suppressor genes in bladder cancer.
  
  
  METHODS
  We evaluated the methylation of 18 tumor suppressor genes in 2 prospective, independent sets of urine samples (training set of 120 preparations and validation set of 128) from patients with bladder cancer (170) and controls (78) using methylation specific multiplex ligation-dependent probe amplification. Diagnostic performance was evaluated with ROC curves. Recurrence, progression and disease specific survival were analyzed using univariate and multivariate Cox models.
  
  
  RESULTS
  PRDM2, HLTF, ID4, DLC1, BNIP3, H2AFX, CACNA1G, TGIF and CACNA1A were methylated in bladder cancer. CCND2, SCGB3A1, BNIP3, ID4 and RUNX3 were the most frequently methylated tumor suppressor genes in each urine set. Methylation of several tumor suppressor genes correlated with clinicopathological variables, such as stage, tumor grade, focality or age. ROC analysis revealed significant diagnostic accuracy for RUNX3 and CACNA1A in the training set, and for RUNX3 and ID4 in the validation set. On univariate and multivariate analysis CACNA1A methylation correlated with recurrence in the training set, while in the validation set PRDM2 and BNIP3 were significantly associated with recurrence and disease specific survival, respectively.
  
  
  CONCLUSIONS
  Tumor suppressor gene methylation allowed for histopathological and clinical stratification. Urine methylation has noninvasive usefulness not only for diagnostic assessment but also as independent bladder cancer prognosticators.
authors.affiliation: Tumor Markers Group, Molecular Pathology Program, Spanish National Cancer Center, Madrid, Spain; Urology Department, Hospital Puerta del Mar, Cadiz, Spain.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: García-Baquero
authors.forename: Patricia
authors.initials: P
authors.lastname: Puerta
authors.forename: Manuel
authors.initials: M
authors.lastname: Beltran
authors.forename: Miguel
authors.initials: M
authors.lastname: Alvarez
authors.forename: Raquel
authors.initials: R
authors.lastname: Sacristan
authors.forename: Jose Luis
authors.initials: JL
authors.lastname: Alvarez-Ossorio
authors.forename: Marta
authors.initials: M
authors.lastname: Sánchez-Carbayo
mesh_terms: D019175:DNA Methylation; D018450:Disease Progression; D016147:Genes, Tumor Suppressor; D006801:Humans; D009361:Neoplasm Invasiveness; D016133:Polymerase Chain Reaction; D011379:Prognosis; D016016:Proportional Hazards Models; D011446:Prospective Studies; D012372:ROC Curve; D015203:Reproducibility of Results; D015996:Survival Rate; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000473:pathology / Q000652:urine*
keywords: CIS; HG; LG; M-ratio; MS-MLPA; TSG; carcinoma in situ; genes, tumor suppressor; high grade; low grade; methylation; methylation ratio; methylation-specific multiplex ligation-dependent probe amplification; multiplex polymerase chain reaction; tumor suppressor gene; urinary bladder; urinary bladder neoplasms