Molecular diagnosis of oral cancer based on aberrant expression and structural alterations of the p53 tumor suppressor gene

本研究旨在建立基于p53抑癌基因增强表达和改变的鳞状细胞癌的分子诊断方法，获得了以下结果：(1)应用PCR-SSCP和直接测序的方法，发现93%的口腔鳞状细胞癌细胞系和63%的肿瘤组织中存在p53基因突变，表明存在p53突变是一种很好的鳞癌分子诊断标志。(2)对鳞状细胞癌细胞系进行P53过表达的免疫组织化学分析，发现错义突变的细胞表达增强，而产生C端截短P53的细胞表达阴性。突变特异性抗体PAb240可识别部分错义突变的P53蛋白。因此，虽然免疫组织化学方法简单、快速，但并不能检测到所有的P53突变。(3)15例口腔白斑中有6例检测到P53过表达，1例口腔鳞癌患者检测到P53过表达。有趣的是，该患者的LK活检组织具有与SCC相同的突变，表明LK转化为SCC。(4)从REF与激活的ras的转化活性判断，预后因素与突变的p53基因的生物学活性无显著相关性。(5)SSCP分析采用快速简便的P53基因突变检测方法。由于不使用放射性物质，这种方法可以作为医院实验室的诊断方法之一。

Research to develop molecular diagnostic method of squamous cell carcinoma(SCC) based on the enhanced expression and alteration of the p53 tumor suppressor gene was conducted and the following results were obtained.(1) Using PCR-SSCP and direct sequencing methods, 93% of oral SCC cell lines and 63% of tumor tissues were found to have p53 gene mutations, thus indicating that presence of p53 mutations is a good molecular diagnostic marker of SCC.(2) Immunohistochemical analysis of SCC cell lines for overexpression of p53 showed that enhanced expression was detected in cells with missense mutations while the expression was negative in cells which produce p53 with C-terminal truncation. A part of missense-mutated p53 proteins were recognized by a mutant-specific antibody PAb240. Thus, not all p53 mutations were detected by immunohistochemical method, although the method is easy and quick to analyze mutation.(3) p53 overexpression was detected in 6 out of 15 leukoplakia(LK), and LK of a patient who had oral SCC at the same region. Interestingly, LK biopsy materials of this patient had the same mutation as of SCC, indicating that LK converts to SCC.(4) No significant correlation of prognostic factors was found with biological activities of mutated p53 genes as judged from transforming activities of REF with activated ras.(5) The quick and easy method for detection of p53 gene mutations was applied for SSCP analyses. This method can be incorporated as one of diagnoses in laboratories of hospitals, since no radioactive material is used.