CDKN2 gene silencing in lung cancer by DNA hypermethylation and kinetics of p16INK4 protein induction by 5-aza 2'deoxycytidine

Oncogene. 1995 Sep 21;11(6):1211-6.


Absent expression of the cyclin dependent kinase-inhibitor, p16INK4, is observed in a wide range of primary human cancers. Although homozygous deletions and point mutations have been reported in a subset of these tumors, the molecular basis for absent p16INK4 in other samples is unknown. We have examined 33 tumor cell lines and have shown that hypermethylation of a G:C-rich region within exon 1 of the CDKN2 gene was present in 100% of samples with wildtype RB expression and no detectable CDKN2 mutations. Treatment for at least 4 hours with the demethylating agent 5-aza 2'deoxycytidine, but not 5-azacytidine or 6-azacytidine, induces the prolonged expression of p16INK4 protein in each of these samples following a discrete 24-48 hour lag period. Consistent with the hypothesis that hypermethylation of the CDKN2 gene is a tumor-specific mechanism for gene inactivation, we observed hypomethylation at the exon 1 site exclusively in tumor lines that expressed p16INK4 or that had sustained inactivating point mutations within the CDKN2 open reading frame. These findings demonstrate a link between DNA methylation and the p16INK4:RB tumor suppressor pathway.

MeSH terms

  • Azacitidine / analogs & derivatives*
  • Azacitidine / pharmacology
  • Azacitidine / therapeutic use
  • Base Sequence
  • Carrier Proteins / biosynthesis*
  • Carrier Proteins / genetics*
  • Cyclin-Dependent Kinase Inhibitor p16
  • DNA / metabolism
  • DNA Modification Methylases / antagonists & inhibitors*
  • Decitabine
  • Enzyme Inhibitors / pharmacology*
  • Gene Expression Regulation / drug effects
  • Humans
  • Lung Neoplasms / drug therapy
  • Lung Neoplasms / genetics*
  • Lung Neoplasms / metabolism
  • Methylation
  • Molecular Sequence Data
  • Protein Kinase Inhibitors*
  • Tumor Cells, Cultured


  • Carrier Proteins
  • Cyclin-Dependent Kinase Inhibitor p16
  • Enzyme Inhibitors
  • Protein Kinase Inhibitors
  • Decitabine
  • DNA
  • DNA Modification Methylases
  • Azacitidine