Tanaka H, Shimada Y, Imamura M, Shibagaki I, Ishizaki K
Laboratory of Experimental Radiology, Aichi Cancer Center Research Institute, Nagaya, Japan.
Int J Cancer. 1997 Feb 7;70(4):437-42. doi: 10.1002/(sici)1097-0215(19970207)70:4<437::aid-ijc11>3.0.co;2-c.
To determine the role and mode of inactivation of the p16 and p15 genes in human esophageal tumors, we examined alterations and expression of the alpha and beta forms of the p16 gene, 5' CpG island methylation of p16 exon 1 alpha, and alterations of the p15 gene in 30 esophageal squamous-cell-carcinoma cell lines. Of 30 such cell lines examined, 28 (93%) showed aberrations of the alpha form of the p16 gene: 18 homozygous deletions, 6 point mutations and 4 hypermethylation. Methylation was exclusively observed in cell lines with the wild-type alpha form. Of the 6 point mutations, one was observed in exon 1 alpha, one in the splice acceptor site of intron 1 and the remaining 4 were in exon 2. In the beta form, 18 homozygous deletions and 3 point mutations in exon 2 were detected, but no point mutation was found in exon 1 beta. All mutations in exon 2 gave rise to premature termination codons in the reading frame of the alpha transcript, while no non-sense mutations were observed in the reading frame of the beta transcript. Among 12 cell lines without homozygous deletions of the alpha and beta forms of the p16 gene, the expected wild-type beta transcript was observed in 8 cell lines, whereas only one cell line expressed the expected wild-type alpha transcript. Homozygous deletions of the p15 gene were observed in 16 cell lines (53%), and no point mutations were detected. Twelve cell lines had alterations only in the alpha form of the p16 gene, while none showed aberrations exclusively in the p15 gene. Taken together, these results indicate that inactivation of the beta form of the p16 gene and the p15 gene are not so frequent as that of the alpha form of the p16 gene in ESC cell lines, suggesting that aberration of the alpha form of p16 gene is the primary target of 9p loss in ESC.
为确定p16和p15基因在人食管肿瘤中的作用及失活模式，我们检测了30个人食管鳞状细胞癌细胞系中p16基因α和β形式的改变与表达、p16外显子1α的5' CpG岛甲基化以及p15基因的改变。在所检测的30个此类细胞系中，28个（93%）显示p16基因α形式存在畸变：18个纯合缺失、6个点突变和4个高甲基化。甲基化仅在具有野生型α形式的细胞系中观察到。在6个点突变中，1个在外显子1α中观察到，1个在内含子1的剪接受体位点，其余4个在外显子2中。在β形式中，检测到18个纯合缺失和外显子2中的3个点突变，但在外显子1β中未发现点突变。外显子2中的所有突变均导致α转录本阅读框中的过早终止密码子，而在β转录本阅读框中未观察到无义突变。在12个无p16基因α和β形式纯合缺失的细胞系中，8个细胞系观察到预期的野生型β转录本，而只有1个细胞系表达预期的野生型α转录本。在16个细胞系（53%）中观察到p15基因的纯合缺失，未检测到点突变。12个细胞系仅在p16基因的α形式上有改变，而没有一个细胞系仅在p15基因上显示畸变。综上所述，这些结果表明，在食管鳞状细胞癌细胞系中，p16基因β形式和p15基因的失活不如p16基因α形式频繁，提示p16基因α形式的畸变是食管鳞状细胞癌中9p缺失的主要靶点。
