HIC1基因變異參與臺灣肺癌形成及其受p53/SIRT1蛋白調控之機制研究

HIC1 alteration and its association with deregulation of p53/SIRT1-mediated control in lung cancer

李金祝(Chin Chu Lee)

指導教授 ： 王憶卿

國立臺灣師範大學/理學院/生命科學研究所/碩士(2005年)

並列摘要

Purpose: We have previously reported that the chromosomal regions at 17p13.3 showed a high frequency of loss of heterozygosity in tumors from non-small cell lung cancer (NSCLC) patients. HIC1 (hypermethylated in cancer 1) is a candidate tumor suppressor gene at 17p13.3 and is epigenetically inactivated in many human cancers. A circular regulation of HIC1, SIRT1 deacetylase, and p53 is proposed for modulation of cellular responses to DNA damage in cell and animal studies. However, the etiological role of HIC1 alteration and its correlation with p53 and SIRT1 deregulation have never been examined in the same series of human cancer patients. Therefore, we investigated the alterations of HIC1 at the DNA, RNA, and protein levels, and their correlation with deregulation of p53/SIRT-mediated control in lung cancer. Materials and Methods: The alterations of HIC1 including loss of protein/mRNA expression and promoter hypermethylation as well as their clinical correlations were examined in 108 NSCLC patients. In addition, the expression of acetylated p53 and SIRT1 deacetylase was examined by Western blot. We also treated A549 lung cancer cell line with demethylation reagent 5’aza 2’deoxycytidine (5’-Aza-2’-dC) and SIRT1 inhibitor nicotinamide (vitamin B3) to investigate the effects in the regulation of HIC1, p53 and SIRT1. Results: Overall, 65.5%, 75.9%, and 40% of NSCLC patients showed low protein, low mRNA, and promoter hypermethylation for HIC1 gene, respectively. A high concordance was observed between low mRNA expression and promoter hypermethylation for the HIC1 gene (P<0.05). Low mRNA expression of HIC1 was significantly associated with low acetylated p53, supporting that HIC1 is a transactivating target of p53 (P=0.016). In addition, low acetylated p53 is significantly associated with high SIRT1 protein (P=0.002), and high SIRT1 protein is significantly associated with low HIC1 protein (P=0.004). In addition, in the A549 lung cancer cells, which showed low levels of HIC1 mRNA and promoter hypermethylation, treated with demethylation reagent 5’-Aza-2’-dC, the HIC1 mRNA and protein expressions were restored along with the demethylation of HIC1 promoter. In the A549 cells treated with SIRT1 inhibitor nicotinamide, an increase of p53 acetylation and HIC1 re-expression were seen. In addition, promoter hypermethylation of HIC1 gene was tended to be associated with adenocarcinomas than with squamous carcinomas (P=0.059), whereas deacetylation of p53 was tended to be associated with squamous carcinomas than with adenocarcinomas (P=0.046). Conclusion: HIC1 alteration plays an important role in lung tumorigenesis and the predominant mechanisms of HIC inactivation were HIC1 promoter hypermethylation and p53 deacetylation. The present study shows the first clinical evidence that alteration of HIC1/SIR1/p53 pathway is involved in tumorigenesis.

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