Downregulation of long non-coding RNA MEG3 in nasopharyngeal carcinoma.

In our previous whole-transcriptome sequencing analysis, downregulation of a long non-coding RNA, maternally expressed gene 3 (MEG3), was identified in NPC samples. This finding suggests the possible role of MEG3 as a tumor suppressor in this distinctive disease. In the present study, two MEG3 variants, AF119863 (MEG3-AF) and BX247998 (MEG3-BX), were found abundantly expressed in a normal nasopharyngeal epithelial cell line, NP69. Significant downregulation of MEG3-AF was further verified in a panel of NPC samples including xenografts and primary biopsies. MEG3 is an imprinted gene located within chromosome 14q32, a common deleted region in NPC. Both DNA copy number loss and aberrant promoter methylation contributed to MEG3 inactivation. Interestingly, MEG3 expression could successfully be rescued by the treatment of a demethylation agent. Besides, ectopic expression of MEG3 in NPC cell lines resulted in considerable repression of in vitro anchorage-independent growth and in vivo tumorigenicity, in addition to significant inhibition in cell proliferation, colony formation, and induction of cell cycle arrest. Finally, we revealed the association between MEG3 activity and the p53 signaling cascade. Our findings characterize MEG3 as a tumor suppressive long non-coding RNA in NPC and encourage the development of precise long non-coding RNA-targeted epigenetic therapy against this malignancy. © 2016 Wiley Periodicals, Inc.

14-3-3sigma expression as a prognostic marker in undifferentiated nasopharyngeal carcinoma.

In this study, we investigated the expression of 14-3-3sigma tumor suppressor gene in a panel of NPC cell lines, xenografts and primary tumors. Our objective was to determine the correlation between 14-3-3sigma expression and clinical outcome in NPC. We detected reduced 14-3-3sigma expression in 5/6 NPC tumor lines by quantitative RT-PCR and Western blotting. By immunohistochemical staining, significant down-regulation of 14-3-3sigma was also found in 26/72 (36.1%) primary tumors of NPC patients, who were treated with curative radiotherapy. Promoter methylation was confirmed in a subset of primary tumors by methylation-specific PCR analysis. Importantly, we demonstrated that 14-3-3sigma expression is significantly associated with both overall survival (OS) and cancer-specific survival (CSS), but not with the clinical staging of NPC patients. The low 14-3-3sigma expression was associated with improved overall (p=0.029) and cancer-specific survival (p=0.042) on univariate analysis. 14-3-3sigma expression and staging were also independent variables to all the prognostic factors under multivariate analysis. In conclusion, low expression of 14-3-3sigma appears to be a valuable marker for better survival in patient with NPC. These results provide the evidence that 14-3-3sigma expression is a significant prognostic factor for NPC patients.

Epigenetic inactivation of TSLC1 gene in nasopharyngeal carcinoma.

Deletion of 11q23 is a common genetic aberration in nasopharyngeal carcinoma (NPC). Multiple candidate tumor suppressor genes (TSG) were mapped to this region but few of them were investigated in NPC. TSLC1 (tumor suppressor in lung cancer) is recently reported to be a putative TSG on 11q23. This gene was found to be inactivated by promoter hypermethylation in non-small cell lung carcinoma (NSCLC), liver cancer, and breast cancer. To study the role of TSLC1 gene in NPC tumorigenesis, we screened for mutations and aberrant methylation of TSLC1 gene in 5 NPC cell lines, 3 NPC xenografts, and 38 primary NPC cases. No somatic mutations of TSLC1 were detected in the NPC samples, but a 9-bp (CCACCACCA) deletion in exon 8 was found in a primary NPC and its corresponding blood sample. Bisulfite sequencing revealed aberrant methylation of TSLC1 promoter in four NPC cell lines. Loss of TSLC1 gene expression was found in two cell lines (HK-1 and CNE-2) with dense methylation. Expression of this gene was restored in these cell lines after treatment with demethylating agent 5-aza-2'-deoxycytidine. Our results showed that silencing of TSLC1 gene expression in NPC was associated with promoter hypermethylation. Promoter hypermethylation of TSLC1 gene was further illustrated in 34.2% (13/38) of primary NPCs. No aberrant promoter methylation was found in any of the four investigated normal nasopharyngeal epithelia. Frequent epigenetic inactivation of TSLC1 gene in NPC suggested that this gene is one of the target tumor suppressor genes of this endemic cancer.