Co-targeting of multiple microRNAs on factor-Inhibiting hypoxia-Inducible factor gene for the pathogenesis of head and neck carcinomas.

BACKGROUND MicroRNAs (miRNAs) are short, noncoding RNAs that inhibit the expression of target genes that play roles in tumorigenesis. MiR-21, miR-31, and miR-184 are oncogenic miRNAs for head and neck squamous cell carcinoma (HNSCC). Factor-inhibiting hypoxia (FIH)-inducible factor is known to inactivate hypoxia-induced downstream effectors and is involved in HNSCC suppression. This study investigates whether miR-21, miR-31, and miR-184 target FIH in HNSCC. METHODS Reporter assays, Western blot analysis, quantitative reverse transcriptase-polymerase chain reaction (PCR) analysis, and phenotypic assays were used to prove that miR-21, miR-31, and miR-184 directly target FIH. Clinicopathological implications of the gene expression were also analyzed. RESULTS MiR-21, miR-31, and miR-184 directly bind to various sites in the 3' untranslated region (UTR) of FIH transcript, and this binding is associated with decreased FIH protein expression in HNSCC cells. Treatment with the precursors of these miRNAs increases the proliferation and migration of HNSCC cells. Concomitant treatment with precursors repressed FIH and enhanced oncogenicity most profoundly. Upregulation of miR-21, miR-31, and miR-184 expression is found in more than 80% of HNSCC tumors and 72% of tumors have concordant upregulation of these 3 oncogenic miRNAs. The highest expression of these miRNAs is present in T4b and stage IVB tumors. Downregulation of FIH mRNA expression is noted in 69% of HNSCC tumors, and in tumors exhibiting high expression of these miRNAs, the FIH mRNA expression is consistently downregulated. CONCLUSION This study provides novel clues indicating that miR-21, miR-31, and miR-184 co-target FIH tumor suppressor during pathogenesis in the vast majority of HNSCC.