分子伴侣AGR2调控26S蛋白酶体活性及介导蛋白酶体抑制剂耐药的分子机制研究

蛋白酶体是维持细胞稳态的重要分子机器，亦是良好的抗肿瘤药物靶点。硼替佐米是首个应用于临床的蛋白酶体抑制剂，但其在实体瘤治疗中效果不佳且耐药机制不明确。本课题前期发现二硫键异构酶AGR2高表达可介导肿瘤细胞耐受硼替佐米。进一步实验表明，AGR2通过二硫键异构酶活性位点与蛋白酶体调节颗粒蛋白PSMD2相互作用，阻碍调节颗粒与核心颗粒的结合，从而抑制26S蛋白酶体活性。因此，我们提出二硫键异构酶AGR2可能通过抑制蛋白酶体活性介导肿瘤细胞对硼替佐米耐受。本课题在前期工作的基础上，拟通过构建高表达与低表达AGR2的肿瘤细胞系，利用体内外实验确定AGR2表达对硼替佐米药效的影响。采用生物化学与分子互作等技术手段，鉴定AGR2与蛋白酶体亚基的互作网络，研究分子伴侣AGR2对蛋白酶体的组装，解聚及蛋白酶活性的调控作用。本项目将进一步揭示蛋白酶体活性精细调节过程，并为硼替佐米合理用药奠定良好的研究基础。

Proteasome is an important molecular machine that maintains homeostasis of cells, and is also a good target for antitumor drugs. Bortezomib is the first proteasome inhibitor to be used clinically, but its efficacy in solid tumors is poor and the drug resistance mechanism is unclear. Earlier in this project, we found that the high expression of disulfide isomerase AGR2 can mediate the tolerance of bortezomib to tumor cells. Further experiments showed that AGR2 interacted with the proteasome regulatory granule protein PSMD2 through the disulfide isomerase active site, hindering the binding of the regulatory granule to the core granule, thereby inhibiting 26S proteasome activity. Therefore, we propose that the disulfide isomerase AGR2 may mediate tumor cell tolerance to bortezomib by inhibiting proteasome activity. Based on the previous work, this project intends to determine the effect of AGR2 expression on the efficacy of bortezomib by constructing tumor cell lines with high and low expression of AGR2 and using in vivo and in vitro experiments. Biochemical and molecular interaction techniques were used to identify the interaction network of AGR2 and proteasome subunits, and to study the regulation of the proteasome assembly, depolymerization, and protease activity by the molecular chaperone AGR2. This project will further reveal the fine-tuning process of proteasome activity and lay a good research foundation for the rational use of bortezomib.