HOXA-AS2 Promotes Proliferation and Induces Epithelial-Mesenchymal Transition via the miR-520c-3p/GPC3 Axis in Hepatocellular Carcinoma

Abstract

Background/aims: Previous studies have demonstrated that long non-coding RNAs (lncRNAs) may play critical roles in cancer biology, including Hepatocellular carcinoma (HCC). The HOXA cluster antisense RNA2 (HOXA-AS2) lncRNA plays an important role in carcinogenesis, however, the underlying role of HOXA-AS2 in HCC remains unknown. The present study examined the effects of HOXA-AS2 on the progression of HCC, and explored the underlying molecular mechanisms.

Methods: Quantitative real-time PCR was used to detect HOXA-AS2 expression in HCC tissues and cell lines. Furthermore, the effects of HOXA-AS2 silencing and overexpression on cell proliferation, cell cycle, apoptosis, migration, and invasion were assessed in HCC in vitro and in vivo. Furthermore, bioinformatics online programs predicted and luciferase reporter assay were used to validate the association of HOXA-AS2 and miR-520c-3p in HCC cells.

Results: We observed that HOXA-AS2 was up-regulated in HCC tissues and cell lines. In vitro experiments revealed that HOXA-AS2 knockdown significantly inhibited HCC cells proliferation by causing G1 arrest and promoting apoptosis, whereas HOXA-AS2 overexpression promoted cell growth. Further functional assays indicated that HOXA-AS2 significantly promoted HCC cell migration and invasion by promoting EMT. Bioinformatics online programs predicted that HOXA-AS2 sponge miR-520c-3p at 3'-UTR with complementary binding sites, which was validated using luciferase reporter assay. HOXA-AS2 could negatively regulate the expression of miR-520c-3p in HCC cells. MiR-520c-3p was down-regulated and inversely correlated with HOXA-AS2 expression in HCC tissues. miR-520c-3p suppressed cell proliferation, invasion and migration in HCC cells, and enforced expression of miR-520c-3p attenuated the oncogenic effects of HOXA-AS2 in HCC cells. By bioinformatic analysis and dual-luciferase reporter assay, we found that miR-223-3p directly targeted the 3'-untranslated region (UTR) of Glypican-3 (GPC3), one of the key players in HCC. GPC3 was up-regulated in HCC tissues, and was negatively correlated with miR-520c-3p expression and positively correlated with HOXA-AS2 expression.

Conclusion: In summary, our results suggested that the HOXA-AS2/miR-520c-3p/GPC3 axis may play an important role in the regulation of PTC progression, which could serve as a biomarker and therapeutic target for HCC.

Keywords: EMT; HCC; Hoxa-AS2; LncRNA; Therapeutic target; miR-520c-3p.