细胞周期调节基因甲基化在甲状腺癌发生发展中的作用及其与BRAF突变的关系

项目名称： 细胞周期调节基因甲基化在甲状腺癌发生发展中的作用及其与BRAF突变的关系

项目编号： No.30801120

项目类型： 青年科学基金项目

立项/批准年度： 2009

项目学科： 医药、卫生

项目作者： 关海霞

作者单位： 中国医科大学

项目金额： 18万元

中文摘要： 本研究从细胞周期调节基因甲基化角度，探讨甲状腺癌发生及BRAF突变所致肿瘤发生发展的分子机制。研究显示：1）12个候选细胞周期调节基因中，分别有7个、5个和5个基因在PTC、FTC和ATC中存在甲基化，证实了细胞周期调节基因甲基化是甲状腺癌中的常见事件。2）应用免疫组织化学染色（甲状腺组织）和Western杂交（细胞系）对异常甲基化的基因进行蛋白表达的检测，证实了甲基化的发生可使对应蛋白的表达"沉默"。3）结合临床病理资料分析得出，细胞周期调节基因甲基化与PTC的淋巴结转移、肿瘤分期为III/IV期、以及BRAF基因突变显著相关，提示它们可能成为新的肿瘤预后评估标记物。4）应用去甲基化试剂处理甲状腺癌细胞后，细胞中发生甲基化的基因的表达得以恢复，细胞生长减慢，提示发生甲基化的细胞周期调节基因可能成为甲状腺癌的治疗靶点之一。5）应用siRNA和通路抑制剂急性抑制甲状腺癌细胞系的MAP激酶通路，结果未能改变甲状腺癌细胞中细胞周期调节基因的甲基化和表达情况，提示尽管甲基化与BRAF基因突变存在联系，但似乎并非BRAF突变直接引起。长时间抑制MAP激酶通路对甲基化的影响还有待进一步研究。

中文关键词： 甲状腺癌；基因甲基化；细胞周期；BRAF突变

英文摘要： The present study was designed to explore the mechanism behind thyroid tumorgenesis and BRAF mutaion-promoted tumor aggressiveness in terms of aberrant methylation of 12 CKIs and checkpoints genes, which are involved in cell cycle regulation. It was indicated that: 1) p16Ink4a, p15Ink4b, p21Cip1, p27Kip1, 14-3-3 sigma, Gadd45gamma and RB1 genes were found aberrantly hypermethylated in papillary thyroid cancer; p16Ink4a, p27Kip1, 14-3-3 sigma, Gadd45gamma and RB1 genes were found hypermethylated in follicular thyroid cancer; as well as p16Ink4a, p15Ink4b, p27Kip1, 14-3-3 sigma and Gadd45gamma genes were found hypermethylated in anaplastic thyroid cancer. Hypermethylation of genes involved in cell cycle regualtion seems to be a common phenomenon in thyroid cancer. 2) Proteins expression were "silenced" upon aberrant hypermethylation of the corresponding genes, which was tested by immunohistochemisty staining and Western blotting. 3) Hypermethylation of CKIs and checkpoints genes has a close association with the BRAF mutation, lymph nodes metastasis and advanced cancer stages of thyroid cancers, which means this molecular event may become a new biomarker for prognostic prediction. 4) Thyroid cancer cells treated with 5-aza restored the expression of the hypermethylated genes, subsequently led to slowing down of the cell proliferation. This provide a theoretical basis for the demethylation therapy of thyroid cancer. 5) Acute suppression of the MAP kinase signaling activity by siRNA knock-down or specific inhibitor did not have any effect on the methylation status and expression of the CKIs and checkpoints genes in thyroid cancer. This result did not support the hypothesis that the BRAF mutation may cause aberrant hypermethylation of genes directly. However, chronic effects of long-term inhibition of MAP kinase signaling activity on gene methylation needs further studies.

英文关键词： thyroid cancer; gene methylation; cell cycle; BRAF mutation