APE1线粒体调控ROS介导骨肉瘤化疗耐药的分子机制

The inhibition effect of intracellular reactive oxygen species (ROS) generation is an important mechanism of cancer chemoresistance. Apurinic/ apyrimidinic endonuclease 1 (APE1) is a nuclear protein with DNA damage repair activity which regulates nuclear ROS. The recent research indicated that APE1 could also be localized in mitochondria; however, it was unknown whether mitochondrial APE1 can regulate ROS. In previous studies, we found that the cytoplasmic APE1 expression increased with the resistance to chemotherapy in osteosarcoma; after the oxidative stress, the translocation of APE1 to mitochondria could inhibit cell apoptosis; APE1 in mitochondria could regulate mitochondrial electron transport chain (ETC) and further suppress the apoptosis; furthermore, ETC was the main source of ROS. After the chemotherapy, the high cytoplasmic APE1 expression in osteosarcoma and regulatory effects of ETC suggested that cytoplasmic APE1 and osteosarcoma chemotherapy resistance were closely related. Thus, the regulation of mitochondrial ROS may play an essential role. Therefore, we raised a hypothesis that the high expression of mitochondrial APE1 downregulates the ROS level further promotes drug resistance in osteosarcoma chemotherapy. Through the establishment of xenograft model of osteosarcoma multidrug resistance, this project will detect the resistant and parental cell phenotype and the expression of mitochondrial APE1, combined with research methods, including RNA inference, Western blot,ROS detection etc, and will carry out studies in the regulation of mitochondrial APE1 on ROS in vivo and in vitro and the molecular mechanism, in order to provide new strategy to overcome the clinical drug resistance in osteosarcoma chemotherapy.

抑制胞内产生的ROS作用是肿瘤化疗耐药的重要机制。APE1是具有DNA损伤修复功能的核功能蛋白，可核内调控ROS。新近研究发现APE1亦可定位于线粒体，然线粒体APE1是否调节ROS鲜有报道。前期我们发现化疗耐药骨肉瘤中胞浆APE1表达增高；氧化应激后APE1转位至线粒体可抑制细胞凋亡；APE1在线粒体中可调控线粒体电子转运链（ETC）基因抑制凋亡，且ETC是ROS的主要来源。化疗后骨肉瘤中胞浆APE1高表达和ETC调控作用提示核外APE1与骨肉瘤化疗耐药密切相关，而线粒体调控ROS可能是重要作用点。由此我们提出假说：线粒体APE1高表达可通过下调ROS水平介导骨肉瘤化疗耐药。本项目通过建立骨肉瘤耐药细胞株和裸鼠移植瘤模型，检测耐药和亲本细胞表型及线粒体APE1表达，结合干扰表达、WB、ROS检测等方法，研究体内外线粒体APE1对ROS调控及其机制，为临床解决骨肉瘤化疗耐药问题提供新思路。