Circ_0030586 inhibits cell proliferation and stemness in bladder cancer by inactivating the ERK signaling via miR-665/NR4A3 axis

Increasing evidence reveals that circular RNAs (circRNAs) serve as oncogenes or tumor suppressors in the development of various tumors including bladder cancer (BCa). In this study, we explored the function and mechanism of circ_0030586 (also named circABCC4, ATP binding cassette subfamily C member 4) in BCa. The expression of circ_0030586 was significantly decreased in BCa tissues and cells, as suggested by RT-qPCR. The circular characteristics of circ_0030586 were verified by agarose gel electrophoresis and RNase R treatment. Colony formation, 5-Ethynyl-2'-deoxyuridine and sphere formation assays revealed that overexpression of circ_0030586 suppressed BCa cell proliferation and stemness in vitro. According to xenograft experiment, circ_0030586 overexpression suppressed tumor growth in vivo. Mechanistically, RNA pulldown and luciferase reporter assays were carried out to explore the interaction between genes. Circ_0030586 served as a competing endogenous RNA (ceRNA) for hsa-miR-665 to upregulate the expression of nuclear receptor subfamily 4 group A member 3 (NR4A3) which is a downstream target gene of miR-665 in BCa. MiR-665 exhibited high expression in BCa tissues and cells while NR4A3 expression was downregulated in BCa. MiR-665 overexpression or NR4A3 silencing reversed the suppressive effect of circ_0030586 overexpression on BCa cell proliferation and stemness. Moreover, western blot analysis revealed that circ_0030586 inactivated the extracellular signal-regulated kinase (ERK) pathway by upregulating NR4A3 expression. In conclusion, circ_0030586 inhibits BCa cell proliferation and stemness by serving as a ceRNA for miR-665 to upregulate NR4A3 expression and thus inactivate the ERK signaling.