Downregulation of gene expression in laryngeal squamous cell carcinoma is an effect of DNA hypermethylation and correlates with disease progression.

We have turned our attention to gene, already described as deregulated in various types of cancer. By using the expression microarrays performed on the set of 16 laryngeal squamous cell carcinoma (LSCC) samples: 11 cell lines and 5 primary tumors we have shown downregulation of gene as compared to non cancer controls from head and neck region. gene downregulation, further confirmed by quantitative PCR on 25 LSCC cell lines, was observed in cell lines derived from recurrent tumors in comparison to controls. A significant gene downregulation was observed in cell lines derived from advanced, high grade tumors in comparison to controls. Intrigued by the recurrent transcriptional loss of we searched for the mechanism potentially responsible for its downregulation and hence we analyzed DNA copy number changes (a-CGH), promoter DNA methylation status and occurrence of gene mutations (). Neither the analysis of gene copy number, nor the mutation screen has shown recurrent deletions or mutations, that could contribute to the observed downregulation of the gene. However, by using bisulfite pyrosequencing, we have shown DNA hypermethylation (mean DNA methylation > 78%) of promoter region in 9/25 (36%) LSCC cell lines. Importantly, the 5-aza-2-deoxycytidine-induced inhibition of DNA methylation resulted in restoration of expression in the two LSCC cell lines on mRNA level. In summary, we have shown that recurrent downregulation of in LSCC is dependent on the gene's promoter DNA methylation and is observed predominantly in large, poorly differentiated tumors and recurrences.