AKR1C1促CSC样细胞形成介导膀胱癌奥沙利铂耐药的分子机制

AKR1C1, a CSC-associated marker, is one of the important factors mediating resistance of bladder cancer to platinum drugs. However, the explicit mechanism of AKR1C1 on chemoresistance is still unclear, and the signaling pathways involved remain to be discussed. Previous studies from our lab showed the acquiring of the resistance of bladder cancer to oxaliplatin resulting in many cellular alterations, such as less drug accumulation, lower Pt-DNA adduct formation, higher DNA repair capacity, lower rate of apoptosis, and multidrug resistance. Comprehensive data reflected the similarity to the resistance of CSC, accompanied with the super higher expression of AKR1C1 synchronously. Based on the information, we hypothesized that AKR1C1 may mediate chemoresistance of bladder cancer to oxaliplatin via promoting the CSC-like cell formation. In order to confirm this hypothesis, this project will establish different chemoresistant or chemosensitive models based on the various of genotype and phenotype of AKR1C1 by using the pharmacological and molecular biology methods. The aim to demonstrate the following three parts: first, the increase of AKR1C1 induced by oxaliplatin through KEAP1/NRF2 signaling pathway; second, the correlation between AKR1C1 expression and the phenotype and function of CSC-like cell; final, the CSC-like cell formation mediated by AKR1C1 via Wnt/β-catenin mechanism. We plan to uncover the potential mechanism of AKR1C1 in the process of chemoresistance of bladder cancer to oxaliplatin. The elucidation of the mechanisms by which bladder cancer become refractory to oxaliplatin mediated by AKR1C1 will lead not only to optimal chemosensitization strategies, but also to the discovery of new prognostic and predictive biomarkers.

CSC相关标志物AKR1C1是介导膀胱癌铂药耐药的重要因素，但AKR1C1发挥肿瘤耐药作用的明确机制尚不清楚，其信号途径有待深入探讨。本课题组前期结果表明，奥沙利铂诱导膀胱癌获得性耐药表型表现为药物蓄积少、DNA损伤轻、DNA修复强、细胞凋亡少、多药耐药等多方改变，综合体现了CSC具备的耐药性；且平行伴有CSC相关基因AKR1C1超高表达。综上提出AKR1C1参与膀胱癌CSC样细胞形成，介导奥沙利铂耐药这一假说。本项目拟采用药理学及分子生物学方法，建立多种AKR1C1基因与表型相关耐药或敏感模型，分别从奥沙利铂激活AKR1C1的信号通路、AKR1C1与CSC样细胞表型与功能的相关性、AKR1C1介导CSC样细胞形成途径三方面研究，揭示AKR1C1在膀胱癌对奥沙利铂耐药进程中的调节机制，明确抗癌治疗靶点，以寻求新的具高度针对性的分子靶向药物，为临床抗膀胱癌的治疗提供新思路。

以奥沙利铂为基础的联合化疗可作为对顺铂毒副作用无法耐受者的一线治疗方案，而化学耐药常常是铂药治疗失败的主要原因。AKR1C1是介导实体肿瘤铂药耐药的重要因素，AKR1C1分子靶向治疗有望延缓或抑制实体肿瘤的进程与发展，但AKR1C1发挥肿瘤耐药作用的明确机制尚不清楚，其信号途径亦有待深入探讨。奥沙利铂诱导膀胱癌获得性耐药表型表现为药物蓄积少、DNA损伤轻、DNA修复强、细胞凋亡少、多药耐药等多种改变，综合体现CSC具备的耐药性；且平行伴有CSC相关基因AKR1C1超高表达。综上提出AKR1C1参与膀胱癌CSC样细胞形成，介导奥沙利铂耐药这一假说。基于课题组前期结果，本研究分别从奥沙利铂激活AKR1C1的信号通路、AKR1C1与CSC样细胞表型与功能的相关性、AKR1C1介导CSC样细胞形成途径及AKR1C1与膀胱癌病理参数相关性四方面研究，以揭示AKR1C1在膀胱癌对奥沙利铂耐药进程中的调节机制，明确抗癌治疗靶点。本研究结果显示，伴随AKR1C1表达增高，膀胱癌细胞对奥沙利铂耐药性逐渐增强，反之，敲除AKR1C1可明显增强膀胱癌细胞对奥沙利铂敏感性。AKR1C1的活性及表达受转录因子NRF2调控，奥沙利铂刺激膀胱癌细胞耐药可诱导NRF2从胞浆向胞核转位，进而促进AKR1C1表达。本研究亦显示，培养膀胱癌CSC细胞所获得的成球细胞，其AKR1C1表达明显高于亲代细胞；抑制AKR1C1活性亦明显增强CSC细胞对奥沙利铂敏感性；同时，过表达AKR1C1膀胱癌细胞的促EMT相关蛋白Vimentin与Slug表达增高，促CSC形成蛋白β-Catenin及磷酸化β-Catenin表达亦升高，且抑制β-Catenin可减少AKR1C1表达并逆转AKR1C1介导的膀胱癌对奥沙利铂耐药。临床样本AKR1C1表达与膀胱癌肿瘤体积呈正相关，且具有降低生存率的趋势。本项研究证明，奥沙利铂可通过NRF2信号途径诱导AKR1C1的表达及活性升高，Wnt信号通路激活NRF2-AKR1C1信号，诱导CSC样细胞生成，最终导致膀胱癌对奥沙利铂耐药。因此，靶向抑制AKR1C1的表达及活性，对协同或优化奥沙利铂的抗肿瘤治疗方案具有重要意义，可为抗肿瘤靶向药物的设计提供新的理论依据，并有望改善膀胱癌患者的治疗反应性，提高膀胱癌患者的生存率及生存质量。