ALKBH5-regulated circ_0032649, acting as a sponge for miR-4755-3p to upregulate ORAI2, facilitates intrahepatic cholangiocarcinoma tumorigenicity.

OBJECTIVE: Circular RNAs (circRNAs) serve as critical biomarkers in intrahepatic cholangiocarcinoma (ICC). This study aimed to identify the key circRNA targeted by ALKBH5 in ICC and elucidate the underlying molecular mechanisms.

METHODS AND RESULTS: Sixteen clinical samples were collected to assess ALKBH5 expression pattern in ICC. CircRNA sequencing was performed to identify the key circRNA targeted by ALKBH5, followed by bioinformatic prediction of the associated microRNA and downstream mRNA. Following in vitro transfection, cell functions were evaluated using cell counting kit-8, colony formation, and flow cytometry. Molecular interactions were assessed by RNA immunoprecipitation and dual-luciferase reporter assays. A xenograft tumor model was established, followed by intratumoral injection of lentivirus, to assess the tumorigenic capacity of ALKBH5 and circ_0032649. The results indicated that ALKBH5 was significantly overexpressed in ICC tumors compared with paracancerous tissues. Circ_0032649 was identified as a key downstream circRNA of ALKBH5 to mediate the expression of the miR‑4755‑3p/ORAI2 axis. Depletion of ALKBH5 significantly suppressed ICC cell viability, colony formation, apoptosis resistance, and epithelial-mesenchymal transition, while inducing G1-phase cell cycle arrest. Mechanistically, ALKBH5 bound circ_0032649, which functioned as a sponge for miR-4755-3p to upregulate ORAI2 expression. In vivo, the ALKBH5/circ_0032649 axis similarly promoted ICC tumor progression and altered the expression of this competing endogenous RNA (ceRNA) regulatory axis.

CONCLUSION: ALKBH5-regulated circ_0032649 promotes ICC malignancy by a miR-4755-3p/ORAI2-dependent ceRNA network.