title: ADAMTS1, CRABP1, and NR3C1 identified as epigenetically deregulated genes in colorectal tumorigenesis.
abstract: BACKGROUND
  Gene silencing through CpG island hypermethylation is a major mechanism in cancer development. In the present study, we aimed to identify and validate novel target genes inactivated through promoter hypermethylation in colorectal tumor development.
  
  
  METHODS
  With the use of microarrays, the gene expression profiles of colon cancer cell lines before and after treatment with the demethylating agent 5-aza-2'-deoxycytidine were identified and compared. The expression of the responding genes was compared with microarray expression data of primary colorectal carcinomas. Four of these down-regulated genes were subjected to methylation-specific PCR, bisulphite sequencing, and quantitative gene expression analysis using tumors (n=198), normal tissues (n=44), and cell lines (n=30).
  
  
  RESULTS
  Twenty-one genes with a CpG island in their promoter responded to treatment in cell lines, and were simultaneously down-regulated in primary colorectal carcinomas. Among 20 colon cancer cell lines, hypermethylation was subsequently identified for three of four analyzed genes, ADAMTS1 (85%), CRABP1 (90%), and NR3C1 (35%). For the latter two genes, hypermethylation was significantly associated with absence or reduced gene expression. The methylation status of ADAMTS1, CRABP1, and NR3C1 was further investigated in 116 colorectal carcinomas and adenomas. Twenty-three of 63 (37%), 7/60 (12%), and 2/63 (3%) adenomas, as well as 37/52 (71%), 25/51 (49%), and 13/51 (25%) carcinomas were hypermethylated for the respective genes. These genes were unmethylated in tumors (n=82) from three other organs, prostate, testis, and kidney. Finally, analysis of normal colorectal mucosa demonstrated that the observed promoter hypermethylation was cancer-specific.
  
  
  CONCLUSIONS
  By using a refined microarray screening approach we present three genes with cancer-specific hypermethylation in colorectal tumors, ADAMTS1, CRABP1, and NR3C1.
authors.affiliation: Department of Cancer Prevention, Institute for Cancer Research, Rikshospitalet-Radiumhospitalet Medical Centre, Oslo, Norway.
authors.forename: Guro E
authors.initials: GE
authors.lastname: Lind
authors.forename: Kristine
authors.initials: K
authors.lastname: Kleivi
authors.forename: Gunn I
authors.initials: GI
authors.lastname: Meling
authors.forename: Manuel R
authors.initials: MR
authors.lastname: Teixeira
authors.forename: Espen
authors.initials: E
authors.lastname: Thiis-Evensen
authors.forename: Torleiv O
authors.initials: TO
authors.lastname: Rognum
authors.forename: Ragnhild A
authors.initials: RA
authors.lastname: Lothe
mesh_terms: D051722:ADAM Proteins / Q000235:genetics*; D000071097:ADAMTS1 Protein; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000235:genetics*; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D015972:Gene Expression Regulation, Neoplastic; D052138:Genes, Neoplasm; D006801:Humans; D046228:Microarray Analysis; D011401:Promoter Regions, Genetic / Q000235:genetics; D011965:Receptors, Glucocorticoid / Q000235:genetics*; D018168:Receptors, Retinoic Acid / Q000235:genetics*; D017422:Sequence Analysis, DNA; D014407:Tumor Cells, Cultured
chemical_list: C497713:NR3C1 protein, human; D011965:Receptors, Glucocorticoid; D018168:Receptors, Retinoic Acid; C082927:retinoic acid binding protein I, cellular; D051722:ADAM Proteins; D000071097:ADAMTS1 Protein; C103859:ADAMTS1 protein, human