背景:环状RNARNA结合基序蛋白23(circ_RBM23;ID:hsa_circ_0000524)是肝细胞癌(HCC)的新型调节因子。在这里,我们计划研究其在肝癌索拉非尼耐药中的作用.
方法:circ_RBM23,microRNA(miR)-338-3p,Ras相关GTP酶运输蛋白(RAB1B),通过实时定量PCR和蛋白质印迹法检测蜗牛和E-cadherin。索拉非尼抗性(SR)HCC细胞(Huh7/SR和SK-HEP-1/SR)通过获得索拉非尼抗性来建立,用MTT法测定细胞功能,Edu测定,集落形成试验,凋亡测定,transwell分析,和体内异种移植物形成试验。miR-338-3p与circ_RBM23或RAB1B之间的交联通过生物信息学分析和双荧光素酶报告基因测定得到证实。
结果:在SR患者和SR细胞的组织中发现Circ_RBM23上调,伴随着miR-338-3p下调和RAB1B上调。通过干扰circ_RBM23或增强miR-338-3p,索拉非尼在SR细胞中的50%抑制浓度(IC50)受到极大抑制,与此相关的是抑制EdU阳性细胞率,索拉非尼治疗下的集落形成和迁移/侵袭能力,以及细胞凋亡率的提高。此外,circ_RBM23抑制在体内sorfanib治疗下延迟Huh7/SR细胞的肿瘤生长。
结论:Circ_RBM23促进化学抗性,恶性增殖,通过调节miR-338-3p/RAB1B轴对SRHCC细胞的迁移和侵袭。
BACKGROUND: Circular RNA RNA-binding motif protein 23 (circ_RBM23; ID: hsa_circ_0000524) is a novel regulator in hepatocellular carcinoma (HCC). Herein, we planned to investigate its role in sorafenib resistance in HCC.
METHODS: Levels of circ_RBM23, microRNA (miR)-338-3p, Ras-related GTPase-trafficking protein (RAB1B), Snail and E-cadherin were detected by real-time quantitative PCR and western blotting. Sorafenib resistant (SR) HCC cells (Huh7/SR and SK-HEP-1/SR) were established by acquisition of sorafenib resistance, and cell functions were measured by MTT assay, Edu assay, colony formation assay, apoptosis assay, transwell assay, and in vivo xenograft formation assay. Crosslink between miR-338-3p and circ_RBM23 or RAB1B was confirmed by bioinformatics analysis and dual-luciferase reporter assay.
RESULTS: Circ_RBM23 upregulation was discovered in the tissues of SR patients and SR cells, which was accompanied with miR-338-3p downregulation and RAB1B upregulation. The 50% inhibitory concentration (IC50) of sorafenib in SR cells was greatly suppressed by interfering circ_RBM23 or reinforcing miR-338-3p, allied with this was the inhibition of EdU-positive cell rate, colony formation and migration/invasion abilities under sorafenib treatment, as well as the enhancement of apoptotic rate. Moreover, circ_RBM23 inhibition delayed tumor growth of Huh7/SR cells under sorfanib treatment in vivo.
CONCLUSIONS: Circ_RBM23 promoted chemoresistance, malignant proliferation, migration and invasion of SR HCC cells by modulating miR-338-3p/RAB1B axis.