Career Development Program
Project Four: Epigenetic Patterns in Head and Neck Squamous Cell Carcinoma
Laura Rozek, Ph.D.
The development of head and neck squamous cell carcinoma (HNSCC) is a multistep process requiring the accumulation of genetic and epigenetic alterations influenced by an individual's genetic predisposition and environmental risk factors. Despite advances in treatment, five-year survival remains at 50% (Surveillance, Epidemiology and End Results; www.seer.cancer.gov). Research studies indicate molecular analysis of somatic changes in HNSCC tumors have interest because they represent a potentially reversible target for prevention and treatment. This proposal outlines a case-only study of existing cases from the University of Michigan Head and Neck SPORE study to examine the prevalence of hypermethylated regions in cancer-related genes in tumors; utilize hierarchical clustering methods to determine if any methylation pattern corresponds to a distinct, clinical phenotype; and characterize hypermethylation as a possible independent prognostic factor for overall survival. A more thorough understanding of methylation patterns in HNSCC could identify a panel of markers for the prediction of outcome, including sensitivity to chemotherapy, and also serve as preliminary data for a R01 submission to further evaluate the relationship between somatic changes in HNSCC and prevention and prognosis of this disease.
Specific Aims are:
- Measure the prevalence of hypermethylated genes in a test set of 96 HNSCC using a commercially-available DNA Illumina methylation panel representing CpG islands in 807 cancer-related genes;
- Determine if hypermethylation corresponds to specific subset of HNSCC in these tumors using unsupervised hierarchical cluster analysis;
- Evaluate hypermethylation as a prognostic factor in HNSCC
- Measure methylation in genes identified in Aim 2 in a nonoverlapping validation set of 300 HNSCC tumors using Pyrosequencing;
- Evaluate the predictive value of these markers on survival in the presence of classically reported risk factors.
We have begun to define methylation profiles using the Illumina Goldengate Methylation chip on recurrent and nonrecuurent head and neck cancers. The first step of this project is to identify tumor tissue archived as a part of the SPORE project. We are currently processing these samples to extract both DNA and RNA (to be archived for future use) and bisulfite converting them for analysis using the Illumina chip in the University of Michigan Sequencing Core. We are concurrently processing samples for a non-overlapping validation set of cases using pyrosequencing in the Rozek laboratory. Novel planned analyses are also taking advantage of recently published associations between methylation of CpG sites in the HPV genome and cervical cancer prognosis. We have optimized methylation markers using pyrosequencing for these HPV CpG sites in head and neck cancer cell lines and are incorporating these into our repertoire of methylation markers. These analyses will not only identify novel prognostic markers for HN cancer, but also lead to a better understanding of the relationship between epidemiologic risk factors and the head and neck cancer epigenome.