Inactivation of RASSF1A, RARβ2 and DAP-kinase by promoter methylation correlates with lymph node metastasis in nasopharyngeal carcinoma
Inactivation of RASSF1A, RARβ2 and DAP-kinase by promoter methylation correlates with lymph node metastasis in nasopharyngeal carcinoma
复制标题
DOI：
10.4161/cbt.8.5.7686
复制
发表时间：
2009-03-01
影响因子：
3.6
通讯作者：
Mokdad-Gargouri, Raja
中科院分区：
文献类型：
--
作者：
Fendri, Ali;Masmoudi, Asma;Mokdad-Gargouri, Raja
表观遗传修饰是导致肿瘤细胞基因沉默的机制之一。采用甲基化特异性PCR技术，检测了两例鼻咽癌移植瘤（C15和C17）、68例原发性鼻咽癌和9例正常鼻咽上皮组织中Ras相关结构域家族1A（RASSF 1A）、死亡相关蛋白激酶（DAP-kinase）和视黄酸受体β 2（RAR β 2）基因启动子区的甲基化。我们发现C15和C17显示RASSF 1A、RAR β 2和DAP激酶基因的完全启动子甲基化。在原发性NPC肿瘤中，所有三种测试基因的启动子甲基化发生率非常高：RASSF 1A为91%，RAR β 2和DAP激酶均为88%，而所有正常鼻咽上皮均未甲基化。有趣的是，我们的研究揭示了这三个基因的异常启动子甲基化与淋巴结受累有统计学相关性（p < 0.0001）。此外，RASSF 1A的高甲基化与诊断时的年龄（p = 0.047）和T分期（p = 0.037）相关，而RAR β 2的高甲基化与组织学类型相关（p = 0.011）。综上所述，我们的研究结果表明，RASSF 1A和RAR β 2表达的沉默与高分化肿瘤，晚期肿瘤阶段和淋巴结转移的存在有关。采用逆转录聚合酶链反应（RT-PCR）和免疫组化（IHC）方法检测两个下游靶基因考克斯-2和p53的表达。我们揭示了鼻咽癌活检组织中考克斯-2的表达与RAR β 2异常甲基化丢失之间的显著相关性（p = 0.003），我们的结论是，RASSF 1A、RAR β 2和DAP激酶的超甲基化失活是鼻咽癌发生和发展的关键步骤。
Epigenetic modification is one of the mechanisms leading to gene silencing in neoplastic cells. By methylation-specific PCR, we analyzed the promoter methylation of three cancer-related genes: Ras Association domain Family 1A (RASSF1A), Death Associated Protein kinase (DAP-kinase) and Retinoic Acid Receptor beta 2 (RAR beta 2) in two NPC xenografts (C15 and C17), 68 primary NPC tumors, and 9 normal nasopharyngeal epithelia. We showed that C15 and C17 displayed a complete promoter methylation of RASSF1A, RAR beta 2 and DAP-kinase genes. In primary NPC tumors, the incidence of promoter methylation was very high for all three tested genes: 91% for RASSF1A, 88% for both RAR beta 2 and DAP-kinase whereas all normal nasopharyngeal epithelia were unmethylated. Interestingly, our study revealed that aberrant promoter methylation of the three genes were statistically associated with the lymph node involvement (p < 0.0001). In addition, hypermethylation of RASSF1A was correlated with age at diagnosis (p = 0.047) and T stage (p = 0.037) while the RAR beta 2 hypermethylation was associated with histological type (p = 0.011). Taken together, our results demonstrate that silencing of RASSF1A and RAR beta 2 expression by promoter hypermethylation is associated with highly differentiated tumors, advanced tumor stage and the presence of lymph node metastasis.To assess the functional significance of the epigenetic silencing of RAR beta 2 and DAP-kinase in NPC, we analysed the expression of two downstream target genes COX-2 and p53 by reverse PCR (RT-PCR) and immunohistochemistry (IHC). We revealed a significant association between expression of COX-2 and loss of RAR beta 2 through aberrant methylation (p = 0.003) in NPC biopsies.We concluded that the inactivation of RASSF1A, RAR beta 2 and DAP-Kinase by hypermethylation is a key step in NPC tumorigenesis and progression.