抑癌基因p16和Rb与肺癌早期诊断的临床实验研究

目的　研究肺癌组织中p16和Rb基因失活的比率和失活的机制 ,探讨其与肺癌生物学特性及临床、病理和基因分型诊断的关系。　方法　采用免疫组化、双重原位杂交、PCR、PCR SSCP以及序列分析等方法 ,检测 10 6例肺癌及 2 3例肺良性疾病组织标本抑癌基因p16和Rb的改变。　结果　肺癌细胞p16、Rb在蛋白和mRNA水平的表达明显低于正常肺组织和良性疾病肺组织 ,且与肺癌组织学类型、有无淋巴结转移以及临床病理分期密切相关。较早期肺癌 (I、II期 )病例即有较为显著的抑癌基因p16和Rb的失活 (32 6 %和 2 8 3% ) ;非小细胞肺癌以p16基因失活为主 (5 0 1% ) ,小细胞肺癌以Rb基因失活为主 (88 2 % )。p16基因失活的主要机制有纯合缺失、甲基化和点突变。　结论　抑癌基因p16和Rb在肺癌发生发展中起重要作用 ;p16和Rb基因的失活 ,有可能是肺癌发生的早期始动环节 ,对肺癌的早期诊断有重要意义 ;本研究结果提示 ,有可能建立肺癌基因分型诊断新模式。

Relationship between tumor suppressor gene p16 and Rb and early diagnosis of lung cancers

OBJECTIVES: To study the incidences and mechanisms of tumor suppressor gene p16 and Rb inactivation in lung cancers, and to investigate their relation to biological characters, clinical pathological diagnosis and gene classification diagnosis of lung cancers. METHODS: Immunohistochemistry, double in situ hybridization, PCR, PCR-SSCP and sequencing were applied to study the changes of p16 and Rb genes in 106 lung cancers and 23 lung tissues of benign diseases. RESULTS: Total expression rates of p16 and Rb proteins and mRNAs in lung cancers were lower than those in normal lung tissues and benign lung diseases, and they were specially related to histological type, metastasis and clinical stage of lung cancers. In stage I and II lung cancers, the obvious inactivation of tumor suppressor gene p16 or Rb was examined (32.6% or 28.3%); p16 inactivation was detected mainly in non-small cell lung cancers, and Rb inactivation mainly in small cell lung cancers. There were three mechanisms of homozygous deletions, methylations and mutations for p16 gene inactivation. The deletion rate of p16 exon1 and/or exon2 was 25.8%, mainly took place in p16 protein negative cases of non-small cell lung cancers. 15 cases (16.9%) took part in methylations on SmaI sites of CpG island of p16 gene. PCR-SSCP and sequencing showed that 9 cases had p16 gene mutations. CONCLUSIONS: p16 and Rb genes may play important roles in genesis and progression of lung cancers. Inactivation of p16 or Rb gene may be is an early link of lung canceration, that is very important for early diagnosis of lung cancers. A new gene classification model for lung cancer diagnosis would be set up based on the research of p16 and Rb genes.