OP092: Role of slug in head and neck squamous cell carcinoma (HNSCC)

Abstract

Objective hypothesis Hand neck malignancies account for 6% of all cancers diagnosed in the United States and result in ∼14,000 deaths annually. Our previous oligonucleotide microarray study (Ginos et al., 2004 Cancer Res.) identified Slug (SNAI2) over-expression in a signature associated with recurrent head and neck squamous cell carcinoma (HNSCC) following definitive treatment. Slug, a zinc finger transcription factor family member, is critical in mediating epithelial to mesenchymal transitions (EMT) in developing organisms and in modulating sensitivity to apoptosis. Since acquisition of an invasive phenotype and resistance to apoptosis are primary characteristics of human cancer we hypothesized that Slug over-expression would induce EMT, increase invasion and evade apoptosis in HNSCC cell lines. Methods Baseline levels of Slug and E-cadherin were determined using Real time PCR and Western blotting in a variety of HNSCC cell lines. A cell line with high E-cadherin expression and low Slug expression was selected for transfection studies. We overexpressed Slug in a HNSCC cell line (SCC-15) and probed for epithelial and mesenchymal markers (E-cadherin, Vimentin and α -smooth muscle actin) using Immunofluorescence and Western blotting. Apoptosis evasion was tested using serum starvation and clonogenic assays. Results Epithelial marker, E-cadherin, expression was decreased significantly in the clones compared to mock transfectants. Increased mesenchymal markers Vimentin and α -smooth muscle actin expression was observed using both Immunofluorescence and western blotting. Slug-transfected cells showed reduced cell death in a serum starvation assay. Preliminary data also suggested that Slug-transfected clones show increased resistance to radiation-induced apoptosis using clonogenic assay. Conclusion Slug induces EMT when overexpressed in SCC-15 cell line. EMT induction includes loss of E-cadherin and therefore loss of epithelial integrity. Epithelial integrity loss leads to an altered phenotype and increased expression of mesenchymal markers Vimentin and α -smooth actin. Preliminary data from serum starvation and clonogenic assays suggest that Slug has a protective effect on the Slug-transfected clones when compared to the mock. The precise mechanism of this has yet to be explored in HNSCC cell lines. The above results suggest that Slug plays a strong role in recurrent disease through the malignant phenotype and evasion of apoptosis.