Methylation in head and neck squamous cell carcinoma

In order to improve treatment and facilitate timely detection in Head and Neck Squamous Cell Carcinoma (HNSCC), elucidation of early detection markers is crucial. DNA methylation markers are advantageous, because DNA methylation is an early event. Following an overview on methylation in HNSCC in Chapter 1, we describe a genome-wide screen using Restriction Landmark Genomic Scanning (RLGS) in Chapter 2. This analysis found a set of potential tumor suppressor genes that are commonly methylated and downregulated in HNSCC.; Not all relevant candidates are detected by RLGS because of limitations of the assay. In Chapter 3, a candidate gene approach identified C/CAAT enhancer binding protein alpha (C/EBPalpha), a gene previously shown to exhibit tumor suppressor activity in acute myeloid leukemia (AML) and lung cancer, as a gene of interest. C/EBPalpha tumor suppressor activity in lung cancer has previously been shown to be downregulated by epigenetic mechanisms. More recently, this gene has been found to be downregulated in HNSCC. This prompted investigation into the involvement of epigenetics in downregulating C/EBPalpha in HNSCC. It was revealed that C/EBPalpha is downregulated in HNSCC by loss of heterozygosity (LOH) and upstream DNA methylation. Also, C/EBPalpha overexpression in a HNSCC cell line (SCC22B) revealed its ability to provide tumor suppressor activity in HNSCC in vitro and in vivo.; Upstream methylation of C/EBPalpha correlates with decreased expression in HNSCC. In Chapter 4, investigation of previously unstudied AP2alpha binding sites within the upstream methylated region demonstrated that AP2alpha suppresses C/EBPalpha promoter activity and protein expression. Methylation analysis of the upstream C/EBPalpha sequence after AP2alpha downregulation revealed decreased methylation, suggesting that AP2alpha binding may preceed and facilitate methylation and stable silencing of the gene. Finally, in Chapter 5 we discuss the relevance of the findings in the preceeding chapters and the future direction of this body of work.