Epigenetic silencing of MAL, a putative tumor suppressor gene, can contribute to human epithelium cell carcinoma

Background: To identify new and useful candidate biomarkers in head and neck squamous cell carcinoma (HNSCC), we performed a genome-wide survey and found that Myelin and lymphocyte-associated protein (MAL) was a gene that was markedly down-regulated in HNSCC. Hence, we investigated the mechanism of MAL silencing and the effects of MAL on the proliferation, invasion, and apoptotic potential in HNSCC. Results: MAL was significantly down-regulated in 91.7% of HNSCC specimens at the mRNA level as compared with adjacent normal tissues (P = 0.0004). Moreover, the relative transcript levels of the MAL gene were remarkably decreased by five-fold in nine HNSCC cell lines as compared with normal head and neck epithelium cells. MAL gene expression was restored in 44%, 67%, and 89% in HNSCC cell lines treated with TSA, 5-Aza-dC, and TSA plus 5-Aza-dC, respectively. Furthermore, bisulfate-treated DNA sequencing demonstrated that the two CpG islands (that is, M-1 and M-2) located in MAL promoter region were completely methylated in the HNSCC cell lines (CpG methylated ratio was more than 90%), and only one CpG island (that is, M1) was partially methylated in HNSCC tissues (CpG methylated ratio between 20% and 90%). A significant reduction in cell proliferation and a change in the cell cycle profile were also observed in MAL transfectants. Matrigel assay demonstrated that the invasiveness of HNSCC cells significantly decreased. A significant increase in the population of apoptotic cells was observed in MAL transfected cells. The exogenous expression of the MAL gene suppressed malignant phenotypes, while the cell death induced by MAL gene transfer was a result of apoptosis as demonstrated by the induction of cleavage of the poly (that is, ADP-ribose) polymerase. Additionally, tumor growth was suppressed in cells expressing MAL as compared with cells not expressing MAL. Conclusion: Our data suggest that the epigenetic inactivation of MAL, as a candidate tumor suppressor gene, can contribute to human epithelial cell carcinoma and may be served as a biomarker in HNSCC.