Detection of hypermethylated RIZ1 gene in primary tumor, mouth, and throat rinsing fluid, nasopharyngeal swab, and peripheral blood of nasopharyngeal carcinoma patient.
OBJECTIVE
The aims of this study are to examine the methylation status of the Fn2 RIZ1 gene in nasopharyngeal (NP) tumors and nasopharyngeal carcinoma (NPC) cell lines and to evaluate the clinical value of methylated RIZ1 gene in body fluid samples of NPC patients.


METHODS
Methylation status of RIZ1 was evaluated by MSP on CNE-2 and M1 cell lines, 30 tumor biopsies, and their matched body fluid samples, including mouth and throat (M & T) rinsing fluid, NP swabs, plasma, and buffy coat. Normal controls included 8 normal NP biopsies and body fluid samples from 29 healthy volunteers. Sequencing was performed on MSP products from one NP tumor and one M & T rinsing fluid. Transcription of the RIZ1 gene before and after 5-aza-2'-deoxycytidine treatment was examined on CNE-2.


RESULTS
The methylated RIZ1 gene was detected in both CNE-2 and M1, 18 (60%) NP tumors, but not in any of the normal controls. Of 30 matched body fluid samples, methylated RIZ1 DNA was found in 11 (37%) NP swabs, 9 (30%) M & T rinsing fluid, 7 (23%) plasma, and 3 (10%) buffy coat samples. Sequencing analysis confirmed all cytosines to uracils conversion, excluding cytosines in CpG dinucleotides in methylated PCR products. Promoter methylation correlated with loss of RIZ1 mRNA expression, and 5-aza-2'-deoxycytidine treatment restored its expression in CNE-2.


CONCLUSIONS
Our results suggest that promoter hypermethylation of the RIZ1 gene is commonly found in NPC. Its detection in body fluid samples of NPC patients but not in normal controls indicates that it is worth to further evaluate its clinical application in assisting screening of NPC and monitoring recurrence after treatment.
Chang
Hsiao Wen
HW
Department of Surgery, The University of Hong Kong, Hong Kong, People's Republic of China.
Chan
Amy
A
Kwong
Dora Lai Wan
DL
Wei
William Ignace
WI
Sham
Jonathan Shun Tong
JS
Yuen
Anthony Po Wing
AP
D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002277:Carcinoma / Q000097:blood / Q000235:genetics* / Q000378:metabolism*; D016022:Case-Control Studies; D004247:DNA / Q000378:metabolism; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000235:genetics*; D000077209:Decitabine; D011495:Histone-Lysine N-Methyltransferase; D006801:Humans; D009055:Mouth / Q000378:metabolism*; D009303:Nasopharyngeal Neoplasms / Q000097:blood / Q000235:genetics* / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics*; D010614:Pharynx / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013447:Sulfites / Q000494:pharmacology; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D013447:Sulfites; D014157:Transcription Factors; D000077209:Decitabine; D004247:DNA; D011495:Histone-Lysine N-Methyltransferase; C093563:PRDM2 protein, human; D001374:Azacitidine