Epigenetic Profiling of Oral Cancer Cells

The proposed work will evaluate an innovative technology for profiling cancer initiator cells to identify markers that can be used in diagnostics and molecular targets that can guide drug development programs. Oral squamous cell carcinomas afflict nearly 25,000 patients in the US, with more than ten-fold that number worldwide. Treatment of oral cancers still relies on surgical intervention and cytotoxic drug cocktails and survival rates have remained unchanged over the past thirty years. Hence, there remains a pressing need for strategies that can give earlier detection and new drug therapies that have higher efficacy in treatment. It has become clear that the invasiveness and progression of oral cancers (among others) relies on a small population of ‘cancer initiator cells’ (CICs), which share an invasive phenotype that facilitates metastasis and also a stem cell phenotype that allows these cells to proliferate to give macroscopic metastases. It is further clear that epigenetic factors are involved in the generation and maintenance of these cells. These recent insights have motivated efforts to understand the molecular pathways that distinguish the CICs from the normal epithelial cells. The proposed work will apply a novel peptide array technology to profile lysates of normal epithelial cells and of CICs to identify patterns of deacetylase activity that may underlie the cancer cell phenotype. The technology is based on the SAMDI mass spectrometry technique developed by the PI and which uses matrix-assisted laser desorption-ionization mass spectrometry to analyze self-assembled monolayers. Peptide substrates for an enzyme of interest can be immobilized to the monolayer. SAMDI mass spectrometry detects the mass of the peptide-alkanethiolate conjugate. In this way, when the monolayer is treated with an enzyme that modifies the peptide, the resulting mass change of the peptide can be detected with mass spectrometry. The applicants have demonstrated that this assay is applicable to a broad range of post-translational activities, can be performed in high throughput using plates having 1,536 distinct reaction zones—and offering a throughput of 50,000 assays per day—and is quantitative, with Z-factors greater than 0.8. The proposed work will apply this method to profile deacetylase activities in cell cultures and identify differences in the activity profiles between normal and cancer cells as described in two specific aims.