Purpose: The purpose of this study is to investigate the expression and functional role of Annexin (ANXA1) in lymph node (LN) metastasis of hypopharyngeal carcinoma (HSCC).
Methods: Differentially expressed genes in tissue from HSCC with or without LN metastasis were obtained from a previous RNA sequencing experiment. The presence of LN metastasis is determined by pathological diagnosis after neck dissection. ANXA1 expression was detected by qRT-PCR and Western blotting. Immunohistochemistry was used to detect the expression of ANXA1 in 74 cases of HSCC and normal control tissues. We also evaluated the clinical significance of ANXA1 in HSCC. Differentially expressed genes related to ANXA1 were analyzed using bioinformatic tools, and potential mechanisms of action of ANXA1 were assessed using in vitro experiments. In these in vitro experiments, cell proliferation was detected by CCK8 staining, and colony formation, migration and invasion were assessed using Transwell assays, and apoptosis as well as cell cycle status were quantified by flow cytometry.
Results: ANXA1 was significantly downregulated in HSCC with LN metastasis. The survival rate of patients with low ANXA1 expression was significantly worse than that of patients with high ANXA1 expression (p< 0.05). Silencing ANXA1 in cell culture experiments promoted the proliferation, migration and invasion of FaDu cells, inhibited apoptosis, and increased the proportion of cells in S phase. We furthermore found that the mRNA expression of ANXA1 was positively correlated with Yap1 expression (p< 0.0001). Our in vitro experiments showed that ANXA1 regulates the expression of Yap1, and over-expression of Yap1 could reverse the effect of ANXA1 silencing on cancer cell progression.
Conclusion: Our findings suggest that ANXA1 is a putative LN metastasis suppressor gene in tumor, which may suppress the LN metastasis of HSCC by regulating the expression of Yap1.
Keywords: HSCC, ANXA1, Yap1, LN metastasis