长效重组人白介素2药物的生物学活性和抑瘤作用  

Biological activity and antitumor effect of long-acting recombinant human interleukin-2 drug

作　　者：梁学军 张凤霞 靳婷 祝静静 LIANG Xuejun;ZHANG Fengxia;JIN Ting;ZHU Jingjing(Novocodex Biopharmaceuticals Company Limited,Shaoxing 312000,Zhejiang,China)

机构地区：[1]浙江新码生物医药有限公司,浙江绍兴312000

出　　处：《北京大学学报(医学版)》2025年第2期253-261,共9页Journal of Peking University:Health Sciences

摘　　要：目的:考察运用非天然氨基酸定点偶联聚乙二醇(polyethylene glycol,PEG)获得的PEG化重组人白介素2(pegylated recombinant human interleukin 2,PEG-rhIL-2)的生物学活性和抑瘤作用,并探索其抑瘤机制。方法:用表面等离子共振(surface plasmon resonance,SPR)技术检测T41、Y45和V91三个不同位点PEG-rhIL-2与人IL-2受体α(interleukin 2 receptor α,IL-2R_(α))和IL-2受体β(interleukin 2 receptor β,IL-2R_(β))的平衡解离常数(equilibrium dissociation constant,KD)。采用Western blot检测不同剂量rhIL-2和PEG-rhIL-2激活CTTL-2和YT细胞中Janus激酶-信号转导和转录激活因子5(Janus kinase-signal transducer and activator of transcription 5,JAK-STAT5)信号通路的水平。将小鼠单次给药后采血,检测不同时间点药物的浓度,评估Y45-PEG-rhIL-2的药代动力学参数。选择小鼠肝癌细胞系Hepa1-6、胰腺癌细胞系Pan-02和结肠癌细胞系MC-38,用C57BL/6品系小鼠构建肿瘤模型,分别注射不同剂量的Y45-PEG-rhIL-2及赋形剂对照,评估药物的抑瘤作用,并在MC-38模型中评估Y45-PEG-rhIL-2与抗小鼠程序性死亡受体-1(programmed death-1,PD-1)单抗联用的抑瘤作用。构建Hepa1-6小鼠肿瘤模型,分别注射rhIL-2、Y45-rhIL-2和Y45-PEG-rhIL-2,用流式细胞仪分析肿瘤浸润淋巴细胞的比例。结果:SPR检测结果显示PEG-rhIL-2与IL-2R_(α)/IL-2R_(β)亲和力均降低,Y45-PEG-rhIL-2与IL-2R_(α)亲和力降低至约1/250,与IL-2R_(β)亲和力降低至1/3。Western blot结果显示Y45-PEG-rhIL-2激活表达IL-2受体异源三聚体αβγ(heterotrimeric IL-2 receptor complex αβγ,IL-2R_(αβγ))的CTLL-2细胞JAK-STAT5信号的活性降低至约1/300,激活表达IL-2受体异源二聚体βγ(heterodimeric IL-2 receptor complex βγ,IL-2R_(βγ))的YT细胞JAK-STAT5信号的活性降低至1/3。小鼠单次给药后药代动力学评估结果显示Y45-PEG-rhIL-2消除半衰期为17.7 h,具有优于rhIL-2的药代动力学特征。在小鼠肿瘤模型中,Y45-PEG-rhIL-2显Objective:To investigate the biological activity and antitumor effect of pegylated recombinant human interleukin 2(PEG-rhIL-2)obtained by site-specific conjugation of polyethylene glycol(PEG)with non-natural amino acids,and to explore its antitumor mechanism.Methods:The binding activities of PEG-rhIL-2 at three different sites(T41,Y45,and V91)to human interleukin 2 receptors α(IL-2R_(α))and β(IL-2R_(β))and were detected by surface plasmon resonance(SPR)technology.Western blot was used to detect the levels of the Janus kinase-signal transducer and activator of transcription 5(JAK-STAT5)signaling pathway activated by different doses of rhIL-2 and PEG-rhIL-2 in CTTL-2 and YT cells.Blood was collected after a single administration in mice to detect the drug concentration at different time points and evaluate the pharmacokinetic parameters of Y45-PEG-rhIL-2.Mouse hepatoma cell line Hepa1-6,pancreatic cancer cell line Pan-02,and colon cancer cell line MC-38 were selected.Tumor models were constructed in C57BL/6 mice.Different doses of Y45-PEG-rhIL-2 and excipient control were administrated respectively to evaluate the tumor suppression effect of the drug.In the MC-38 colon cancer model,the tumor suppression effect of Y45-PEG-rhIL-2 combined with anti-programmed death-1(PD-1)monoclonal antibody was evaluated.Hepa1-6 mouse tumor models were constructed and rhIL-2,Y45-rhIL-2 and Y45-PEG-rhIL-2 were administrated respectively.The proportion of tumor-infiltrating lymphocytes was analyzed by flow cytometry.Results:The SPR detection results showed that the binding activities of PEG-rhIL-2 to IL-2R_(α)/IL-2R_(β)were both reduced.The affinity of Y45-PEG-rhIL-2 to IL-2R_(α)was reduced to approximately 1/250,and its affinity to IL-2R_(β)was reduced to 1/3.Western blot results showed that the activity of Y45-PEG-rhIL-2 in stimulating JAK-STAT5 signaling in CTLL-2 cells expressing heterotrimeric IL-2 receptor complex IL-2R_(αβγ)was reduced to approximately 1/300,while its activity in YT cells expressing heterodimeric IL-

关 键 词：重组人白介素2 非天然氨基酸 免疫治疗 

分 类 号：R966[医药卫生—药理学]