RAB25 expression is epigenetically downregulated in oral and oropharyngeal squamous cell carcinoma with lymph node metastasis

Oral and oropharyngeal squamous cell carcinoma (OOSCC) have a low survival rate, mainly due to metastasis to the regional lymph nodes. For optimal treatment of these metastases, a neck dissection is required; however, inaccurate detection methods results in under- and over-treatment. New DNA prognostic methylation biomarkers might improve lymph node metastases detection. To identify epigenetically regulated genes associated with lymph node metastases, genome-wide methylation analysis was performed on 6 OOSCC with (pN+) and 6 OOSCC without (pN0) lymph node metastases and combined with a gene expression signature predictive for pN+ status in OOSCC. Selected genes were validated using an independent OOSCC cohort by immunohistochemistry and pyrosequencing, and on data retrieved from The Cancer Genome Atlas. A two-step statistical selection of differentially methylated sequences revealed 14 genes with increased methylation status and mRNA downregulation in pN+ OOSCC. <i>RAB25</i>, a known tumor suppressor gene, was the highest-ranking gene in the discovery set. In the validation sets, both <i>RAB25</i> mRNA (<i>P</i> = 0.015) and protein levels (<i>P</i> = 0.012) were lower in pN+ OOSCC. <i>RAB25</i> mRNA levels were negatively correlated with <i>RAB25</i> methylation levels (<i>P</i> &lt; 0.001) but RAB25 protein expression was not. Our data revealed that promoter methylation is a mechanism resulting in downregulation of RAB25 expression in pN+ OOSCC and decreased expression is associated with lymph node metastasis. Detection of <i>RAB25</i> methylation might contribute to lymph node metastasis diagnosis and serve as a potential new therapeutic target in OOSCC.