ABSTRACT: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer in the world. Its mortality rate is high, and metastasis is one of the main reasons for the poor prognosis. Therefore, it is quite urgent to elucidate the mechanism of HNSCC metastasis and to discover valuable therapeutic targets. Circular RNAs (circRNAs) have been characterized as key regulators of gene expression in numerous malignances. However, the role of circRNAs in HNSCC metastasis remains largely unknown. In this study, circRFWD3, was defined. We demonstrated a novel circRNA, circRFWD3, was significantly upregulated in HNSCC tissues and cell lines by circRNA microarray analysis and qPCR. Notably, high expression of circRFWD3 is related to highly aggressive HNSCC cell lines and lymph node metastasis in HNSCC patients. After that, Sanger sequencing, RNase R and actinomycin D assay were performed to verify the ring structure of circRFWD3. Then functional experiments found it could promote the metastasis of HNSCC cells both in vitro and in vivo. Mechanistically, dual luciferase reporter assay, FISH, RIP, RNA pull-down, RNA-seq and western blot experiments were employed, and found that circRFWD3 served as a miRNAs sponge for miR-27a/27b, leading to upregulation of PPARγ, and then activate NF-κB/MMP13 signaling. Finally, ISH and IHC were carried out to determine the expression levels and clinical significances of circRFWD3 and PPARγ in clinical cohorts of HNSCC. According to the analysis results from two independent HNSCC cohorts, upregulated expression of circRFWD3 and PPARγ were positively associated with worse survival in patients with HNSCC. Overall, our results uncover that circRFWD3 acts a critical role in promoting the aggressiveness of HNSCC cells and is a prognostic marker for the disease, indicating that circRFWD3 may act as a potential therapeutic target in HNSCC. CircRNA microarray analysis of HNSCC
