Promoter hypermethylation of the EDNRB gene in nasopharyngeal carcinoma.
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.
Lo
Kwok-Wai
KW
Department of Anatomical and Cellular Pathology, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong SAR, People's Republic of China. kwlo@cuhk.edu.hk
Tsang
Yuen-Shan
YS
Kwong
Joseph
J
To
Ka-Fai
KF
Teo
Peter M L
PM
Huang
Dolly P
DP
D001483:Base Sequence; D019175:DNA Methylation; D017931:DNA Primers / Q000737:chemistry; D004273:DNA, Neoplasm / Q000235:genetics*; D015870:Gene Expression; D020868:Gene Silencing; D006801:Humans; D008969:Molecular Sequence Data; D009092:Mucous Membrane / Q000502:physiology; D009303:Nasopharyngeal Neoplasms / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D004273:DNA, Neoplasm; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin