Tumor suppressor genes (TSGs) are commonly inactivated by somatic mutation and/or promoter methylation; yet, recent high-throughput genomic studies have not identified key TSGs inactivated by both mechanisms. We pursued an integrated molecular analysis based on methylation binding domain sequencing (MBD-seq), 450K Methylation arrays, whole exome sequencing, and whole genome gene expression arrays in primary head and neck squamous cell carcinoma (HNSCC) tumors and matched uvulopalatopharyngoplasty tissue samples (UPPPs). We uncovered 186 downregulated genes harboring cancer specific promoter methylation including PAX1 and PAX5 and we identified 10 key tumor suppressor genes (GABRB3, HOXC12, PARP15, SLCO4C1, CDKN2A, PAX1, PIK3AP1, HOXC6, PLCB1, and ZIC4) inactivated by both promoter methylation and/or somatic mutation. Among the novel tumor suppressor genes discovered with dual mechanisms of inactivation, we found a high frequency of genomic and epigenomic alterations in the PAX gene family of transcription factors, which selectively impact canonical NOTCH and TP53 pathways to determine cell fate, cell survival, and genome maintenance. Our results highlight the importance of assessing TSGs at the genomic and epigenomic level to identify key pathways in HNSCC, deregulated by simultaneous promoter methylation and somatic mutations.

Department of Head and Neck Surgery; University of Texas M D Anderson Cancer Center; Houston TX USA

Department of Oncology Biostatistics; Johns Hopkins University; School of Medicine; Baltimore MD USA

Department of Otolaryngology and Head and Neck Surgery; Johns Hopkins University; School of Medicine; Baltimore MD USA

Department of Otolaryngology and Head and Neck Surgery; Johns Hopkins University; School of Medicine; Baltimore MD USA; Department of Obstetrics and Gynecology; University of Puerto Rico School of Medicine; Río Piedras Puerto Rico

Department of Otolaryngology and Head and Neck Surgery; Johns Hopkins University; School of Medicine; Baltimore MD USA; Department of Otorhinolaryngology Head and Neck Surgery; University of Groningen; University Medical Center; Groningen The Netherlands

Department of Otolaryngology and Head and Neck Surgery; Johns Hopkins University; School of Medicine; Baltimore MD USA; Laboratory of Molecular Pathology and Institute of Molecular and Translational Medicine; Faculty of Medicine and Dentistry; Palacky University; Olomouc Czech Republic

Department of Otolaryngology and Head and Neck Surgery; Johns Hopkins University; School of Medicine; Baltimore MD USA; Milton J Dance Head and Neck Center; Greater Baltimore Medical Center; Baltimore MD USA

Department of Otolaryngology and Head and Neck Surgery; Johns Hopkins University; School of Medicine; Baltimore MD USA; Sidney Kimmel Comprehensive Cancer Center; Johns Hopkins University; Baltimore MD USA

Division of Biostatistics and Bioinformatics; Department of Oncology; Sidney Kimmel Comprehensive Cancer Center; Baltimore MD USA

Sidney Kimmel Comprehensive Cancer Center; Johns Hopkins University; Baltimore MD USA

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