Abstract 3557: Epigenetic modulation of microRNA-363 in human papillomavirus-positive head and neck squamous cell carcinoma

Abstract

Abstract The incidence of human papillomavirus (HPV)-associated head and neck squamous cell carcinoma (HNSCC) of the oropharynx has significantly increased in recent decades, despite decreasing rates of tobacco and alcohol-related HNSCC. HPV-positive HNSCC is often characterized as a distinct malignancy due to unique epidemiological features and improved prognoses compared with HPV-negative HNSCC. MicroRNAs (miRNAs) are small non-coding RNAs that regulate mRNA translation and decay. Altered miRNA signatures have been identified in numerous types of solid tumors, including HNSCC, but their functional consequences are largely unknown. Our studies show that miR-363 is significantly overexpressed in HPV-positive compared to HPV-negative HNSCC patient tumors and cell lines and that the HPV-16 E6 oncoprotein positively modulates miR-363 expression. We hypothesize that the HPV-16 E6 oncoprotein upregulates miR-363 at the epigenetic level by decreasing miR-363 promoter methylation. The Illumina Genome Analyzer miRNA Sequencing and Illumina HiSeq 2000 miRNA Sequencing platforms were utilized to assess miRNA levels in HPV-positive and HPV-negative tumors from The Cancer Genome Atlas (TCGA) database. miR-363 levels in HPV-positive and HPV-negative tumors were compared to their respective paired normal tissue samples. The Applied Biosystems Cells-to-CpG Bisulfite Conversion Kit was employed to investigate the methylation status of the miR-363 promoter in HNSCC cell lines. Our analysis of 280 (35 HPV-positive, 245 HPV-negative) TCGA HNSCC patient tumors identified a 3.36-fold increase in miR-363 in HPV-positive compared to HPV-negative tumors (p<0.0001). While matched HPV-positive tumor and paired normal samples indicate miR-363 overexpression in tumor tissues, additional samples are necessary to increase the power and statistical significance of the study. Bisulfite conversion methylation-specific PCR demonstrates that HPV-positive cell lines are unmethylated at the miR-363 promoter whereas HPV-negative cell lines generally possess methylated profiles. Cell proliferation, soft agar colony formation, migration, and invasion functional assays following transient and stable transfection with premiR-363 or microRNA-363 inhibitor are in progress to assess HPV-positive and HPV-negative HNSCC phenotypes. Our overall goal is to elucidate the molecular mechanisms underlying HPV-associated cancers, to identify specific pathways perturbed by HPV, and to develop improved prognostic biomarkers and targeted therapies for HNSCC. Citation Format: Bhavana S. Vangara, Parvez Akhtar, Jennifer R. Grandis, Saleem A. Khan. Epigenetic modulation of microRNA-363 in human papillomavirus-positive head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3557. doi:10.1158/1538-7445.AM2014-3557