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Abstract：
Background: Cisplatin (DDP) based combination chemotherapy is a vital method for the treatment of bladder cancer (BLca). Chemoresistance easily occurs in the course of cisplatin chemotherapy, which is one of the important reasons for the unfavorable prognosis of BLca patients. Circular RNAs (circRNAs) are widely recognized for their role in the development and advancement of BLca. Nevertheless, the precise role of circRNAs in DDP resistance for BLca remains unclear. Methods: To study the properties of circATIC, sanger sequencing, agarose gel electrophoresis and treatment with RNase R/Actinomycin D were utilized. RT-qPCR assay was utilized to assess the expression levels of circRNA, miRNA and mRNA in BLca tissues and cells. Functional experiments were conducted to assess the function of circATIC in BLca progression and chemosensitivity in vitro. Various techniques such as FISH, Dual-luciferase reporter assay, TRAP, RNA digestion assay, RIP and ChIRP assay were used to investigate the relationships between PTBP1, circATIC, miR-1247-5p and RCC2. Orthotopic bladder cancer model, xenograft subcutaneous tumor model and xenograft lung metastasis tumor model were performed to indicate the function and mechanism of circATIC in BLca progression and chemosensitivity in vivo. Results: In our study, we observed that circATIC expression was significantly enhanced in BLca tissues and cells and DDP resistant cells. Patients with higher circATIC expression have larger tumor diameter, higher incidence of postoperative metastasis and lower overall survival rate. Further experiments showed that circATIC accelerated BLca cell growth and metastasis and induced DDP resistance. Mechanistically, alternative splicing enzyme PTBP1 mediated the synthesis of circATIC. circATIC could enhance RCC2 mRNA stability via sponging miR-1247-5p or constructing a circATIC/LIN28A/RCC2 RNA-protein ternary complex. Finally, circATIC promotes RCC2 expression to enhance Epithelial-Mesenchymal Transition (EMT) progression and activate JNK signal pathway, thus strengthening DDP resistance in BLca cells. Conclusion: Our study demonstrated that circATIC promoted BLca progression and DDP resistance, and could serve as a potential target for BLca treatment.
摘要：
背景：基于顺铂（DDP）的联合化疗是治疗膀胱癌（BLca）的重要方法。顺铂化疗过程中容易发生化疗耐药，是BLca患者预后不良的重要原因之一。环状RNA(circularRNAs,circRNAs)在BLca的发育和发展中的作用被广泛认可。然而，circRNAs在BLca的DDP抗性中的确切作用尚不清楚。方法：研究circATIC的性质，Sanger测序,利用琼脂糖凝胶电泳和RNA酶R/放线菌素D处理。RT-qPCR测定用于评估circRNA的表达水平，BLca组织和细胞中的miRNA和mRNA。进行功能实验以评估circATIC在BLca进展和体外化学敏感性中的功能。各种技术，如FISH，双荧光素酶报告基因测定,陷阱,RNA消化试验,RIP和ChIRP测定用于研究PTBP1，circATIC，miR-1247-5p和RCC2。原位膀胱癌模型,进行了异种皮下移植瘤模型和异种肺转移瘤模型，以表明circATIC在BLca进展和体内化学敏感性中的功能和机制。结果：在我们的研究中，我们观察到在BLca组织和细胞以及DDP抗性细胞中circATIC表达显着增强。circATIC表达较高的患者肿瘤直径较大，术后转移发生率较高，总生存率较低。进一步的实验表明circATIC加速BLca细胞的生长和转移并诱导DDP抗性。机械上,选择性剪接酶PTBP1介导circATIC的合成。circATIC可以通过形成miR-1247-5p或构建circATIC/LIN28A/RCC2RNA-蛋白三元复合物来增强RCC2mRNA的稳定性。最后,circATIC促进RCC2表达以增强上皮-间质转化（EMT）进程并激活JNK信号通路，从而增强BLca细胞的DDP抗性。结论：我们的研究表明circATIC促进BLca进展和DDP抵抗，并可作为BLca治疗的潜在靶标。
