NDRG2调控IRE1α-XBP1介导内质网应激逆转ER+乳腺癌他莫昔芬耐药  

NDRG2 Activates Endoplasmic Reticulum Stress via IRE1α-XBP1 to Reverse Tamoxifen Resistance in ER+Breast Cancer

作　　者：王守莹 杜彦艳[2] 曹鹏 刘文宇 齐俊愉 石炜业 张春晓[1] 周晓雷[1] WANG Shou-Ying;DU Yan-Yan;CAO Peng;LIU Wen-Yu;QI Jun-Yu;SHI Wei-Ye;ZHANG Chun-Xiao;ZHOU Xiao-Lei(College of Food Science and Biology,Hebei University of Science and Technology,Shijiazhuang 050091,China;Department of Clinical Laboratory,The Fourth Hospital of Hebei Medical University,Shijiazhuang 050011,China)

机构地区：[1]河北科技大学食品与生物学院,石家庄050091 [2]河北医科大学第四医院临床检验科,石家庄050011

出　　处：《中国生物化学与分子生物学报》2024年第10期1409-1416,共8页Chinese Journal of Biochemistry and Molecular Biology

基　　金：河北省自然科学基金项目(No.H2024208001);国家自然科学基金项目(No.82003601)资助。

摘　　要：他莫昔芬(tamoxifen,TAM)作为雌激素受体阳性(estrogen receptor,ER+)乳腺癌的一线化疗药物使大多数患者受益,但原发性和继发性耐药问题严重影响临床治疗效果。深入研究ER+乳腺癌TAM耐药机制,改善治疗效果是当前亟待解决的问题。抑癌因子NDRG2(N-myc downstream regulated gene 2,NDRG2)在肿瘤发生发展中发挥重要作用,但是否参与ER+乳腺癌TAM耐药尚不清楚。本研究旨在探明NDRG2在ER+乳腺癌TAM耐药中发挥的作用和机制。通过RT-PCR与免疫印迹分析对比TAM敏感型和耐药型ER+乳腺癌细胞发现,NDRG 2的mRNA转录水平和蛋白质翻译水平在TAM耐药细胞中表达显著下调,且与耐药能力负相关(P<0.001);CCK-8细胞毒性实验和软琼脂克隆形成实验证实,在耐药细胞中过表达NDRG2可显著降低TAM药物半抑制浓度IC 50和软琼脂克隆形成率(P<0.001),逆转耐药表型。分子机制上,X-box结合蛋白1(X-box binding protein 1,XBP1)mRNA剪切实验与内质网相关降解(endoplasmic-reticulum associated degradation,ERAD)报告蛋白的结果显示,过表达NDRG2可增强耐药细胞中剪切型XBP1s mRNA转录与ERAD报告蛋白CD3ε-YFP表达(P<0.001),引发耐药细胞内质网强应激反应;免疫印迹检测结果显示,过表达NDRG2可显著提高耐药细胞中内质网应激感受器肌醇需要激酶1α(inositol requiring enzyme 1,IRE1α)的磷酸化水平及其下游因子,例如内质网EIP辅助因子(endoplasmic reticulum-localized DnaJ 4,ERdj4)、PKR蛋白激酶的细胞抑制剂(cellular Inhibitor of the PKR protein kinase,P58 IPK)、α甘露糖苷酶样应激蛋白(er degradation enhancingαmannosidase likeprotein,EDEM)和蛋白质二硫键异构酶家族A成员5(protein disulfide isomerase family a member 5,PDIA5)的表达水平(P<0.001)。小鼠异种移植瘤研究进一步证实,在耐药细胞中过表达NDRG2可增强TAM治疗效果,显著抑制耐药移植瘤生长(P<0.001)。以上研究结果表明,通过提高耐药细胞中NDRG2表达,Tamoxifen(TAM)has been widely used for the treatment of ER+breast cancer.However,the inevitable emergence of resistance to tamoxifen obstructs the successful treatment of this cancer.The tumor suppressor gene N-myc downstream-regulated gene 2(NDRG2)plays a significant role in the development of ER+breast cancer.However,it is unclear whether NDRG2 participates in mediating TAM resistance in ER+breast cancer.Here,we investigate the expression of NDRG2 mRNA and protein in TAM-sensitive and TAM-resistant ER+breast cancer cells.The results of immunoblotting experiments revealed a negative correlation between NDRG2 expression and TAM resistance ability in ER+breast cancer cells(P<0.001).CCK-8 cell viability assays and soft agar colony formation assays showed that NDRG2 overexpression in TAM resistant cells significantly reduced the TAM IC_(50)value and the soft agar colony formation rate(P<0.001).For the mechanism,the ERAD reporter protein assays showed that NDRG2 overexpression upregulated the expression of the ERAD reporter protein CD3ε-YFP and increased the levels of spliced XBP1s mRNA,leading to severe endoplasmic reticulum stress in TAM resistant cells(P<0.001).Immunoblot analysis confirmed that overexpression of NDRG2 significantly increased the level of phosphorylation of the endoplasmic reticulum stress sensor IRE1αand the expression levels of its downstream protein factors,including ERdj4,P58IPK,EDEM and PDIA5(P<0.001).The in vivo xenograft tumor experiments in mice further verified that NDRG2 overexpression significantly inhibited the growth of resistant tumors,which enhanced the therapeutic effect of TAM(P<0.001).These findings indicate that increasing NDRG2 expression and triggering severe endoplasmic reticulum stress upon TAM treatment can reverse the resistance of ER+breast cancer cells to TAM and inhibits the growth of ER+breast cancer tumors.Our results provide valuable new insights and potential targets for improving the clinical management of TAM-resistance and prognosis in ER+breast cancer.

关 键 词：雌激素受体阳性乳腺癌 N-myc下游调节基因2 他莫昔芬 耐药 内质网应激 

分 类 号：Q7[生物学—分子生物学]