Promoter hypermethylation of the EDNRB gene in nasopharyngeal carcinoma
Promoter hypermethylation of the EDNRB gene in nasopharyngeal carcinoma
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DOI：
10.1002/ijc.10271
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发表时间：
2002-04-10
影响因子：
6.4
通讯作者：
Huang, DP
中科院分区：
文献类型：
--
作者：
Lo, KW;Tsang, YS;Huang, DP
为了确定鼻咽癌（NPC）的表观遗传变化，我们对鼻咽癌细胞系和异种移植物进行了甲基化敏感限制性指纹图谱（MSRF）分析。在鼻咽癌肿瘤中分离到一个190 bp的甲基化序列，该序列与内皮素受体B （EDNRB）基因的5' CpG岛高度同源。由于EDNRB基因在前列腺癌和膀胱癌中通常失活，因此它可能是参与鼻咽癌肿瘤发生的候选靶基因。通过亚硫酸氢盐测序，我们证实了EDNRB的5' CpG岛的超甲基化发生在异种移植物和所有4种细胞系中，但在2种正常的鼻咽生长中没有发生。RT-PCR结果显示，正常鼻咽上皮细胞中只表达EDNRB原始转录本，而不表达剪接转录本。原始EDNRB表达的缺失在2个异种移植物和3个具有密集甲基化模式的细胞系中一致发现。用5'-aza-2'-脱氧胞苷处理这3种细胞系导致EDNRB转录物的重新表达和其启动子区域的去甲基化。我们的研究结果表明，鼻咽癌中EDNRB基因表达的沉默与启动子超甲基化有关。通过甲基化特异性PCR，我们还在19/21（90.5%）原发性肿瘤中检测到EDNRB的5' CpG岛甲基化，而在所有6个正常鼻咽上皮中均未发现甲基化。高频率的启动子超甲基化表明EDNRB基因的抑制可能在鼻咽癌的发展中起作用。(C) 2002 Wiley-Liss, Inc。
To identify the epigenetic changes in nasopharyngeal carcinoma (NPC), we performed methylation-sensitive restriction fingerprinting (MSRF) analysis on NPC cell lines and xenografts. A 190 bp sequence methylated in NPC tumors was isolated and showed high homology to the 5' CpG island of the endothelin receptor B (EDNRB) gene. Since the EDNRB gene is commonly inactivated in prostate and bladder cancers, it may be a candidate target gene involved in NPC tumorigenesis. By bisulfite sequencing, we have confirmed that hypermethylation of the 5' CpG island of EDNRB occurred in both xenografts and all 4 cell lines but not in 2 normal nasopharyngeal outgrowths. RT-PCR demonstrated that only original EDNRB transcripts, but not the splicing transcripts, were expressed in normal nasopharyngeal epithelial cells. Loss of the original EDNRB expression was consistently found in 2 xenografts and 3 cell lines with dense methylation patterns. Treatment of these 3 cell lines with 5'-aza-2'-deoxycytidine led to re-expression of the EDNRB transcript and demethylation of its promoter regions. Our results demonstrate that silencing of EDNRB gene expression in NPC is associated with promoter hypermethylation. Using methylation-specific PCR, we also detected methylation of the 5' CpG island of EDNRB in 19/21 (90.5%) primary tumors, while no methylation was found in all 6 normal nasopharyngeal epithelia. The high frequencies of promoter hypermethylation suggest that repression of the EDNRB gene may play a role in the development of NPC. (C) 2002 Wiley-Liss, Inc.