Human epididymis protein 4 promotes P-glycoprotein-mediated chemoresistance in ovarian cancer cells through interactions with Annexin II

The aim of the present study was to investigate the effects of human epididymis protein 4 (HE4) on drug resistance and its underlying mechanisms. The associations among proteins were detected by immunoprecipitation and immunofluorescence assays. Then, stably transfected cell lines CAOV3-HE4-L and CAOV3-A2-L expressing HE4 short hairpin (sh)RNAs and ANXA2 shRNAs, respectively, were constructed. MTT assay, immunocytochemistry, western blotting, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and flow cytometry were employed to examine drug sensitivity, as well as the expression and activity of P-glycoprotein (P-gp). HE4 and P-gp in epithelial ovarian cancer tissues were assessed via immunohistochemistry. MicroRNAs that targeted the P-gp gene, ABCB1, were predicted using bioinformatics methods, and their expression was evaluated by RT-qPCR. The common signaling pathways shared by HE4, ANXA2 and P-gp were selected by Gene Set Enrichment Analysis (GSEA). The interaction of HE4, ANXA2 and P-gp were confirmed. P-gp expression was positively associated with HE4 and ANXA2 expression, respectively. Moreover, it was observed that there was no significant rescue of P-gp expression in CAOV3-A2-L cells following the administration of active HE4 protein. In addition, the expression of HE4 and P-gp in ovarian cancer tissues of drug-resistant patients were higher compared with that of the drug-sensitive group (P<0.05). Furthermore, the results revealed that hsa-miR-129-5p was significantly increased accompanied by decreased HE4 or ANXA2 expression and P-gp expression in CAOV3-HE4-L and CAOV3-A2-L cells. GSEA analyses disclosed that HE4, ANXA2 and P-gp genes were commonly enriched in the signaling pathway involved in regulating the actin cytoskeleton. These results indicated that HE4 promotes P-gp-mediated drug resistance in ovarian cancer cells through the interactions with ANXA2, and the underlying mechanism may be associated with decreased expression of hsa-miR-129-5p and dysregulation of the actin cytoskeleton signaling pathway.