title: NDRG2gene copy number is not altered in colorectal carcinoma.
abstract: OBJECTIVE
  To investigate if the down-regulation ofN-myc Downstream Regulated Gene 2(NDRG2) expression in colorectal carcinoma (CRC) is due to loss of theNDRG2allele(s).
  
  
  METHODS
  The following were investigated in the human colorectal cancer cell lines DLD-1, LoVo and SW-480: NDRG2 mRNA expression levels using quantitative reverse transcription-polymerase chain reaction (qRT-PCR); interaction of the MYC gene-regulatory protein with theNDRG2promoter using chromatin immunoprecipitation; andNDRG2promoter methylation using bisulfite sequencing. Furthermore, we performed qPCR to analyse the copy numbers ofNDRG2andMYCgenes in the above three cell lines, 8 normal colorectal tissue samples and 40 CRC tissue samples.
  
  
  RESULTS
  As expected, NDRG2 mRNA levels were low in the three colorectal cancer cell lines, compared to normal colon. Endogenous MYC protein interacted with theNDRG2core promoter in all three cell lines. In addition, theNDRG2promoter was heavily methylated in these cell lines, suggesting an epigenetic regulatory mechanism. Unaltered gene copy numbers ofNDRG2were observed in the three cell lines. In the colorectal tissues, one normal and three CRC samples showed partial or complete loss of oneNDRG2allele. In contrast, theMYCgene was amplified in one cell line and in more than 40% of the CRC cases.
  
  
  CONCLUSIONS
  Our study suggests that the reduction inNDRG2expression observed in CRC is due to transcriptional repression by MYC and promoter methylation, and is not due to allelic loss.
authors.affiliation: Anders Lorentzen, Cathy Mitchelmore, Eucaryotic Cell Biology, Department of Science and Environment, Roskilde University, 4000 Roskilde, Denmark.
authors.forename: Anders
authors.initials: A
authors.lastname: Lorentzen
authors.affiliation: Anders Lorentzen, Cathy Mitchelmore, Eucaryotic Cell Biology, Department of Science and Environment, Roskilde University, 4000 Roskilde, Denmark.
authors.forename: Cathy
authors.initials: C
authors.lastname: Mitchelmore
keywords: Allelic loss; Colorectal carcinoma; Copy number; Gene amplification; MYC; N-myc downstream-regulated gene 2; Tumor suppressor