Exosome-Mediated Transfer of circ_0000338 Enhances 5-Fluorouracil Resistance in Colorectal Cancer through Regulating MicroRNA 217 (miR-217) and miR-485-3p

Exosomes are microvesicles secreted by body cells for intercellular communication. The circular RNA circ_0000338 was found to be present in extracellular vesicles and improve the chemoresistance of colorectal cancer (CRC) cells. However, the role of exosomal circ_0000338 in 5-fluorouracil (5-FU) resistance in CRC is largely unknown. The levels of circ_0000338, microRNA 217 (miR-217), and miR-485-3p were detected using quantitative real-time PCR (qRT-PCR). The 50% inhibitory concentration (IC50) values of cells for 5-FU, cell proliferation, and apoptosis were evaluated using cell counting kit 8 (CCK-8), colony formation, flow cytometry, and Western blot assays. The interaction between miR-217 or miR-485-3p and circ_0000338 was confirmed by RNA immunoprecipitation (RIP), dual-luciferase reporter, and pulldown assays. Exosomes were isolated by ultracentrifugation and qualified by transmission electron microscopy (TEM), Nanosight tracking analysis (NTA), and Western blotting. Xenograft models were performed to analyze whether circ_0000338-loaded exosomes could increase resistance of CRC cells to 5-FU in vivo. The circ_0000338 level was elevated in 5-FU-resistant CRC tissues and cells, and circ_0000338 knockdown sensitized 5-FU-resistant CRC cells to 5-FU through enhancing apoptosis and decreasing proliferation in vitro. Mechanistically, circ_0000338 directly bound to miR-217 and miR-485-3p, and the inhibition of miR-217 or miR-485-3p reversed the effects of circ_0000338 knockdown on cell 5-FU resistance in CRC. Additionally, extracellular circ_0000338 could be incorporated into secreted exosomes and transmitted to 5-FU-sensitive cells. Treatment-sensitive cells with exosomes containing circ_0000338 reduced the 5-FU response in CRC both in vitro and in vivo. Besides that, the exosomal circ_0000338 concentration was higher in patients exhibiting resistance to 5-FU and showed good diagnostic efficiency in 5-FU -resistant CRC. The delivery of circ_0000338 via exosomes enhanced 5-FU resistance in CRC through negative regulation of miR-217 and miR-485-3p, indicating a promising diagnostic and therapeutic marker for 5-FU-based chemotherapy in CRC patients.