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Annexin A1 (ANXA1) down-regulation is an early and frequent event in the development of head and neck squamous cell carcinomas (HNSCC). In an attempt to identify the underlying mechanisms of reduced ANXA1 protein expression, this study investigated ANXA1 mRNA expression in HNSCC specimens by both  in situ  hybridization and RT-qPCR. Results showed a perfect concordance between the pattern of ANXA1 mRNA and protein detected by immunofluorescence in tumors, precancerous lesions and normal epithelia, reflecting that ANXA1 down-regulation occurs at transcriptional level. We also found that both miR-196a and miR-196b levels inversely correlated with ANXA1 mRNA levels in paired HNSCC tissue samples and patient-matched normal mucosa. In addition, endogenous levels of ANXA1 mRNA and protein were consistently and significantly down-regulated upon miR-196a and miR-196b over-expression in various HNSCC-derived cell lines. The direct interaction of both mature miR-196a and miR-196b was further confirmed by transfection with  Anxa1  3′UTR constructs. Combined bioinformatics and functional analysis of  ANXA1  promoter activity contributed to identify key regions and potential mediators of  ANXA1  transcriptional control. This study unveils that, in addition to miR-196a, miR-196b also directly targets ANXA1 in HNSCC.

Annexin A1 down-regulation in head and neck squamous cell carcinoma is mediated via transcriptional control with direct involvement of miR-196a/b