Background and objective: In an attempt to discover a new biomarker for early diagnosis and prognosis of esophageal squamous cell carcinoma (ESCC), the regulation mechanism of large intergenic non-coding RNA-regulator of reprogramming (lincRNA-ROR) as a microRNA (miRNA) sponge was studied.
Patients and methods: ROR expression in 91 pairs of ESCC tissue samples and matched adjacent tissues was quantified with real-time fluorescent quantitative polymerase chain reaction (qRT-PCR). The ROR-miRNA-mRNA regulatory network was built with 161 esophageal cancer (EC) tissues and 11 adjacent tumor tissues from The Cancer Genome Atlas (TCGA) database. A total of 96 cases of ESCC from TCGA database were collected for analysis on survival rates. The regulatory relationship between ROR, miR-145 and FSCN1 was verified in ESCC cells via qRT-PCR, dual luciferase reporter (DLR) assay, RNA immunoprecipitation (RIP) and Western blotting. The transwell method was used to detect cell migration and invasion.
Results: ROR expression in ESCC tumor tissues was significantly higher than in the adjacent tissues, p<0.001. The survival rate of ESCC patients with high ROR expression levels was lower than that of patients with low ROR expression levels (p<0.001). ROR overexpression could downregulate miR-145 by up to 50% was proven by RIP, DLR assay, and qRT-PCR. Two effective binding sites of ROR to miR-145 were verified by DLR assay. One of the sites has never been cited in the literature. The Western blotting results showed that FSCN1 was a downstream target of ROR/miR-145 (p<0.05). Transwell assays were used to show that overexpression of ROR enhanced migration and invasion behavior of ESCC and miR-145 hindered these effects.
Conclusion: ROR acted as a competitive endogenous RNA (ceRNA) of miR-145 in ESCC. A novel, effective miR-145 binding site of ROR was discovered. The ROR/miR-145/FSCN1 pathway was shown to take part in the metastasis of ESCC. ROR is likely an oncogene biomarker for ESCC early diagnosis and prognosis.
Keywords: ESCC; FSCN1; ROR; ceRNA; lncRNA; miR-145.