The role of p21 protein and HPV infection on Rb protein inactivation in Taiwanese lung tumorigenesis

• Main Authors: Pei-Liang Chen, 陳培亮
• Other Authors: Huei Lee
• Format: Others
• Language: en_US
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/16687235466284578577

碩士 === 中山醫學院 === 毒理學研究所 === 87 === In this study, we propose that Rb protein inactivation may play a role on Taiwanese lung tumorigenesis, because relatively lower frequency of p53 gene mutation (18%) was found compared with previous data reported in other countries (50-80%). To test the hypothesis, immunohistochemical analysis was used to evaluate the expression of Rb protein in 247 primary non-small-cell lung cancer (NSCLCs) specimens. High frequency of Rb protein was undetectable by immunostaining (169/247, 68.4%). We suspect that the high frequency of Rb protein inactivation may be caused by (1) alteration of genes involved in Rb protein phosphorylation, such as p21, cyclins and CDKs, (2)inactivation by other oncoproteins from HPV infection, and (3) alteration of Rb gene transcription. Thus, we provide the following evidence to demonstrate which mechanism was important in Rb protein inactivation. Therefore, we first analyzed the protein expressions of p21 and cyclin D1 using immunohistochemistry. We found that high frequency of p21 negative immunostaining (113/247, 45.7%) and its expression was significantly associated with Rb protein expression (P = 0.008). Cyclin D1 protein expression was also correlated with Rb protein expression (P < 0.001). These results suggest that Rb protein inactivation may be caused by the increased its phosphorylation. Secondly, genomic nested PCR was used to detect whether HPV DNA was existed in lung tumor. Surprisingly, 55.2% (32/58) of HPV-18 DNA was detected in lung tumors. The HPV infection frequency in lung cancer found in this study was significantly higher than previous reports (0-36%). We thus strongly suggest that high HPV infection may partly result in the absence of p21 and Rb proteins through the induction of ubiquitin-proteosome pathway. Finally, the defect of Rb gene transcription was examined by reverse transcription PCR (RT-PCR) with the primer which was used to hybridize exon 2 of Rb gene and we used genomic PCR to examine whether promoter site of Rb gene was defected to cause down-regulated transcription. Our results showed that Rb protein inactivation were only 16-19% of cases to be explained by the defect of Rb gene transcription. In clinical outcome, the survival rate of patients with the p21 negative/Rb negative immunostaining was significantly shorter than patients with p21 negative/Rb positive (P = 0.0078). This finding further support that p21 negative immunostaining plays an important role on Rb protein inactivation. From the above findings, we strongly suggest that high frequency of Rb protein inactivation caused by p21 gene alterations and HPV infection may play an important role on Taiwanese lung tumorigenesis.