Analysis of oncogenes and tumor-suppressor genes in oral cancers and the application to diagnosis

人类肿瘤是在正常细胞中原癌基因和抑癌基因发生多种遗传变化后，经过多步骤发生的。本研究以口腔鳞状细胞癌（SCC）为研究对象，检测了ras和erbB1 （EGFR）原癌基因以及p53抑癌基因的异常。ras突变：对29例口腔鳞状细胞癌的肿瘤病变进行N-、H- K-ras基因的密码子12、13和61的激活突变检测。通过PCR（聚合酶链反应）扩增的DNA与等位基因特异性寡核苷酸探针杂交或PCR- sscp（单链构象多态性）分析检测突变。结果表明，除H-ras密码子12 （HOC313）和13 （ZA）突变的两株细胞系外，未检测到突变。石蜡包埋切片分析显示HOC313肿瘤样本在初诊时存在突变。虽然在用于细胞构建的肿瘤组织中检测到ZA突变，但正常上皮和白斑病变均为阴性。从这些结果来看，与腺瘤相比，口腔鳞状细胞癌中ras突变的频率相当低，因此提示ras突变在口腔鳞状细胞癌的肿瘤发生中可能并不重要。p53基因突变：通过dna - sscp和外显子- sscp分析检测15株口腔SCC细胞株是否存在突变，然后直接测序。结果，在14个细胞系中发现p53基因突变，定位在cadon 126到cadon 305之间。在密码子248（3例）和外显子6剪接供体序列（2例）发现两个突变热点。14个突变中有13个为点突变，包括10个错义突变。EGFR分析：4例口腔鳞状细胞癌患者均观察到EGF结合能力增加。根据这些结果，我们预计p53突变和EGF结合能力的增加在口腔SCC的发生中起重要作用，因此表明这两者都将是SCC的良好诊断指标。少

Human cancers emerge from a normal cell after multiple genetic changes of protooncogenes and tumor suppressor genes by multistep process. In the present research we focused on squamous cell carcinomas (SCC) in the oral cavity and examined abnormalities of ras and erbB1 (EGFR) protooncogenes, and p53 tumor suppressor gene.ras mutation : 29 tumor lesions of oral SCC were examined for the activational mutations of codons 12, 13, and 61 of N-, H- K-ras genes. Mutations were detected by hybridization of PCR (polymerase chain reaction)-amplified DNA with allele specific oligonucleotide probes or by PCR-SSCP (single strand conformation polymorphysm) analysis. As a result we could not detect mutation except for two cell lines which had mutations of codons 12 (HOC313) and 13 (ZA) of H-ras. Analysis of parafin-embedded sections showed that the tumor sample of HOC313 had the mutation at the time of initial diagnosis. Although the mutation of ZA was detected in the tumor tissue used for the cell l … More ine establishment, the normal epithelium and leukoplakia lesion were negative. From these results frequency of ras mutation in oral SCC is considerablly low compared with those of adenomas, thus suggesting that ras mutation may not important in tumorigenesis of oral SCC.p53 gene mutation : 15 oral SCC cell lines were examined for the presence of the mutation by cDNA-SSCP and exon-SSCP anlaysis, followed by direct sequencing. As a result, p53 gene mutations were found in 14 cell lines and were localized from cadon 126 to cadon 305. Two mutational hot-spots were found in codon 248 (3 cases) and the spliincg donor sequence of exon 6 (2 cases). Thirteen out of 14 mutations were point mutations including 10 missense mutations.Analysis of EGFR : increased capacity for EGF binding was observed in all the examined 4 cases of oral SCC.From these results it is anticipated that p53 gone mutations and increased capacity for EGF binding play important roles in the genesis of oral SCC, thus suggesting that both will be good diagnostic markers for SCC. Less