title: Mechanisms of inactivation of p14ARF, p15INK4b, and p16INK4a genes in human esophageal squamous cell carcinoma.
abstract: The 9p21 gene cluster, harboring growth suppressive genes p14ARF, p15INK4b, and p16INK4a, is one of the major aberration hotspots in human cancers. It was shown that p14ARF and p16INK4a play active roles in the p53 and Rb tumor suppressive pathways, respectively, and p15INK4b is a mediator of the extracellular growth inhibition signals. To elucidate specific targets and aberrations affecting this subchromosomal region, we constructed a detailed alteration map of the 9p21 gene cluster by analyzing homozygous deletion, hypermethylation, and mutation of the p14ARF, p15INK4b, and p16INK4a genes individually in 40 esophageal squamous cell carcinomas (ESCCs) and compared the genetic alterations with mRNA expression in 18 of these samples. We detected aberrant promoter methylation of the p16INK4a gene in 16 (40%), of p14ARF in 6 (15%), and of p15INK4b in 5 (12.5%) tumor samples. Most p16INK4a methylations were exclusive, whereas all but one of the p14ARF/p15INK4b methylations were accompanied by concomitant p16INK4a methylation. We detected homozygous deletion of p16INK4a in 7 (17.5%), of p14ARF-E1beta in 13 (33%), and of p15INK4b in 16 (40%) tumor samples. Most deletions occurred exclusively on the E1beta-p15INK4b loci. Two samples contained p14ARF deletion but with p16INK4a and p15INK4b intact. No mutation was detected in the p14ARF and p16INK4a genes. Comparative RT-PCR showed good concordance between suppressed mRNA expression and genetic alteration for p15INK4b and p16INK4a genes in the 18 frozen samples, whereas 5 of the 13 cases with suppressed p14ARF mRNA expression contained no detectable E1beta alteration but aberrations in the p16INK4a locus. Our results show that in human ESCCs, p14ARF is a primary target of homozygous deletion along with p15INK4b, whereas p16INK4a is the hotspot of hypermethylation of the 9p21 gene cluster. The frequent inactivation of the p14ARF and p16INK4a genes may be an important mechanism for the dysfunction of both the Rb and p53 growth regulation pathways during ESCC development.
authors.affiliation: Laboratory for Cancer Research, College of Pharmacy, Rutgers, The State University of New Jersey, Piscataway 08854-8020, USA.
authors.forename: E P
authors.initials: EP
authors.lastname: Xing
authors.forename: Y
authors.initials: Y
authors.lastname: Nie
authors.forename: Y
authors.initials: Y
authors.lastname: Song
authors.forename: G Y
authors.initials: GY
authors.lastname: Yang
authors.forename: Y C
authors.initials: YC
authors.lastname: Cai
authors.forename: L D
authors.initials: LD
authors.lastname: Wang
authors.forename: C S
authors.initials: CS
authors.lastname: Yang
mesh_terms: D002294:Carcinoma, Squamous Cell / Q000235:genetics*; D018797:Cell Cycle Proteins; D002899:Chromosomes, Human, Pair 9; D018899:CpG Islands; D050762:Cyclin-Dependent Kinase Inhibitor p15; D019175:DNA Methylation; D004938:Esophageal Neoplasms / Q000235:genetics*; D017353:Gene Deletion; D016147:Genes, Tumor Suppressor; D019942:Genes, p16; D015801:Helminth Proteins / Q000235:genetics*; D006801:Humans; D005810:Multigene Family; D009124:Muscle Proteins / Q000235:genetics*; D009154:Mutation; D012333:RNA, Messenger / Q000032:analysis; D014157:Transcription Factors / Q000235:genetics*; D025521:Tumor Suppressor Proteins; D000096985:Calponins
chemical_list: D018797:Cell Cycle Proteins; D050762:Cyclin-Dependent Kinase Inhibitor p15; D015801:Helminth Proteins; D009124:Muscle Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C493966:CDKN2B protein, human; C118619:calponin homolog protein, Schistosoma japonicum; D000096985:Calponins