Abstract
Problem: Hypermethylation of tumor suppressor gene (TSG) promoter regions has been described for a limited number of genes in head and neck squamous cell carcinoma (HNSC). We describe a genome-wide strategy to identify putative tumor suppressor genes in HNSC that are inactivated by promoter methylation.
Methods: HNSC cell lines were treated with a demethylating agent to disinhibit expression of putative TSGs. Identification of candidates was based upon significant expression increases compared to untreated controls on cDNA microarrays. Genes downregulated in primary HNSC were identified by comparison of normal tissue and tumors using similar arrays. Downregulated genes common to these groups were verified by RT-PCR and subjected to PCR-based methylation analysis in primary tumors.
Results: Demethylating cell lines exhibited expression increases in 364 genes, and primary tumors showed 1106 genes with expression decreases, with 52 genes common to both groups. RT-PCR confirmed increased expression in cell lines for approximately 30% of these genes.
Conclusion: Genome-wide screening for promoter hyper-methylation-inactivated genes in primary HNSC and cell lines facilitates identification of putative tumor suppressor genes that may be important in HNSC development.
Significance: This study demonstrates a powerful method of identifying and testing putative HNSCC tumor suppressor genes. Identification of tumor suppressors may eventually lead to discovery of novel therapeutic targets and cancer screening markers and a better understanding of this deadly disease.
Support: Supported in part by an NIH T32 grant.