The Apparent Silencing Mechanism And Tumor Suppressor Function Of R-cadherin Gene In Nasopharyngeal Carcinoma

Nasopharyngeal carcinoma (NPC) is one of the most common cancers in some specific regions, especially southern China. Besides its significant association with EBV infection, both genetic and epigenetic events play indispensable roles in this complicated multi-step process of NPC carcinogenesis.The cadherins are a superfamily of cell surface glycoproteins, consisting of an extracellular domain, which can specially interact with other cell adhesion molecules on the surface of neighboring cells. Recently, much attention has been paid to the epigenetic alternations of cadherin adhesion molecules, since the loss of intercellular adhesion could release individual tumor cell from the tumor mass, which is the first step of the initiation of metastases.In the present study, we focused on the epigenetic regulation of R-cadherin gene, a cell adhesion molecule, in NPC. We evaluated the transcriptional levels and promoter methylation status of R-cadherin gene in NPC cell lines, NPC tumor biopsies, NPC xenografts and normal nasopharyngeal epithelial cell line NP69 and normal nasopharyngeal epithelia by RT-PCR and methylation specific PCR. We found that R-cadherin gene was completely silenced in 4 out of 5 NPC cell lines, and in all the 2 NPC xenografts. Decreased R-cadherin gene transcriptional expression was also observed in NPC primary tumors compared with normal nasopharyngeal epithelia. Promoter methylation of R-cadherin gene could be detected in all the NPC cell lines (5/5) and 94.3% (50/53) of primary tumors,but not in any of the 10 normal epithelia. Loss of R-cadherin gene expression can be greatly restored by the methyltransferase inhibitor 5-aza-dC in 2 NPC cell lines.To address the role of R-cadherin in NPC carcinogenesis, we clone the complete coding region of R-cadherin gene from a human brain cDNA,subcolone to an expression vector p-3×FLAG. Stable transfectance of R-cadherin was established. By cell proliferation assay, cloning formation assay,wound healing assay and scrape loading and dye transfer assay, we demonstrated that ectopic expression of R-cadherin in NPC cells could inhibit tumor cells proliferation, colony formation and cell migration and increase gap junction. Our results support R-cadherin gene as a tumor suppressor gene in NPC.