title: Promoter methylation correlates with reduced NDRG2 expression in advanced colon tumour.
abstract: BACKGROUND
  Aberrant DNA methylation of CpG islands of cancer-related genes is among the earliest and most frequent alterations in cancerogenesis and might be of value for either diagnosing cancer or evaluating recurrent disease. This mechanism usually leads to inactivation of tumour-suppressor genes. We have designed the current study to validate our previous microarray data and to identify novel hypermethylated gene promoters.
  
  
  METHODS
  The validation assay was performed in a different set of 8 patients with colorectal cancer (CRC) by means quantitative reverse-transcriptase polymerase chain reaction analysis. The differential RNA expression profiles of three CRC cell lines before and after 5-aza-2'-deoxycytidine treatment were compared to identify the hypermethylated genes. The DNA methylation status of these genes was evaluated by means of bisulphite genomic sequencing and methylation-specific polymerase chain reaction (MSP) in the 3 cell lines and in tumour tissues from 30 patients with CRC.
  
  
  RESULTS
  Data from our previous genome search have received confirmation in the new set of 8 patients with CRC. In this validation set six genes showed a high induction after drug treatment in at least two of three CRC cell lines. Among them, the N-myc downstream-regulated gene 2 (NDRG2) promoter was found methylated in all CRC cell lines. NDRG2 hypermethylation was also detected in 8 out of 30 (27%) primary CRC tissues and was significantly associated with advanced AJCC stage IV. Normal colon tissues were not methylated.
  
  
  CONCLUSIONS
  The findings highlight the usefulness of combining gene expression patterns and epigenetic data to identify tumour biomarkers, and suggest that NDRG2 silencing might bear influence on tumour invasiveness, being associated with a more advanced stage.
authors.affiliation: Gastroenterology Unit and Research Laboratory, Casa Sollievo della Sofferenza, Hospital, IRCCS, San Giovanni Rotondo, Italy. a.piepoli@operapadrepio.it
authors.forename: Ada
authors.initials: A
authors.lastname: Piepoli
authors.forename: Rosa
authors.initials: R
authors.lastname: Cotugno
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Merla
authors.forename: Annamaria
authors.initials: A
authors.lastname: Gentile
authors.forename: Bartolomeo
authors.initials: B
authors.lastname: Augello
authors.forename: Michele
authors.initials: M
authors.lastname: Quitadamo
authors.forename: Antonio
authors.initials: A
authors.lastname: Merla
authors.forename: Anna
authors.initials: A
authors.lastname: Panza
authors.forename: Massimo
authors.initials: M
authors.lastname: Carella
authors.forename: Rosalia
authors.initials: R
authors.lastname: Maglietta
authors.forename: Annarita
authors.initials: A
authors.lastname: D'Addabbo
authors.forename: Nicola
authors.initials: N
authors.lastname: Ancona
authors.forename: Saverio
authors.initials: S
authors.lastname: Fusilli
authors.forename: Francesco
authors.initials: F
authors.lastname: Perri
authors.forename: Angelo
authors.initials: A
authors.lastname: Andriulli