包载达沙替尼的仿生外泌体治疗肺癌的研究  

Study on tumor cell-derived biomimetic exosomes encapsulated dasatinib against lung cancer

作　　者：史雨宸 张立山[1] 刘晓红 张丽 袁陈雨 赵业霖 罗廖欣 秦烨 何钟 姚红娟 李亮[2] SHI Yuchen;ZHANG Lishan;LIU Xiaohong;ZHANG Li;YUAN Chenyu;ZHAO Yelin;LUO Liaoxin;QIN Ye;HE Zhong;YAO Hongjuan;LI Liang(Dongzhimen Hospital,Beijing University of Chinese Medicine,Beijing 100010,China;State Key Laboratory of Respiratory Health and Multimorbidity,the Institute of Medicinal Biotechnology,Chinese Academy of Medical Sciences&Peking Union Medical College,Beijing 100050,China;Shanghai Institute of Materia Medica,Chinese Academy of Sciences,Shanghai 201203,China)

机构地区：[1]北京中医药大学东直门医院,100010 [2]中国医学科学院北京协和医学院医药生物技术研究所肿瘤室/呼吸和共病全国重点实验室,北京100050 [3]中国科学院上海药物研究所分子影像中心,上海201203

出　　处：《中国医药生物技术》2025年第2期133-145,共13页Chinese Medicinal Biotechnology

基　　金：中国医学科学院医学与健康科技创新工程(CIFMS-2021-I2M-026);科技部“一带一路”创新人才交流外国专家项目(DL2022194002L);国家中医药管理局中医药创新团队及人才支持计划(ZYYCXTD-C-202205);中央高水平中医医院临床科研业务费(HLCMHPP2023130);全国重点实验室专项经费(2060204);卫健委基于大人群队列探索呼吸共病的共性机制和干预手段(2023-I2M-2-001)

摘　　要：目的制备包载达沙替尼的仿生外泌体,并对其进行表征和抗肺癌活性考察。方法利用人肺癌H460细胞外泌体与载药脂质体的各自特点,通过膜融合后自组装,制备包载达沙替尼的仿生外泌体。采用动态光散射法测定其粒径与zeta电位。通过蛋白免疫印迹和荧光共振能量转移(FRET)法验证外泌体-脂质体融合效果。利用荧光探针成像技术定量分析肿瘤细胞对仿生外泌体的摄取能力。MTS法检测体外细胞杀伤活性,并通过皮下荷瘤小鼠模型的活体荧光成像和药效学实验评价其体内肿瘤靶向性、抑瘤效果及安全性。结果达沙替尼仿生外泌体的粒径为(111.3±0.1)nm,电位(-30.1±0.3)mV。蛋白免疫印迹实验证实了仿生外泌体上外泌体特异性标志蛋白CD9、CD81、OXER1的表达。荧光共振能量转移法检测到外泌体与脂质体的膜融合指数为2.171±0.044,提示外泌体与载药脂质体的融合较高。荧光成像显示,同源肺癌细胞对仿生外泌体的摄取显著高于脂质体(P<0.05),而异源胰腺癌细胞对两者的摄取无差异。与游离达沙替尼及脂质体制剂相比,仿生外泌体的体外细胞毒性显著增强。体内动物模型评价结果显示其在荷瘤小鼠体内的肿瘤组织中更易富集,具有较好的归巢能力,显著的体内抑瘤作用(抑瘤率达75.0%),且给药期间未观察到小鼠体重的显著下降。结论达沙替尼仿生外泌体纳米制剂具有显著的抗肺癌活性和良好的安全性,为后续非小细胞肺癌的治疗提供了科学依据和前期基础。Objective The goal of this study was to create dasatinib-loaded biomimetic exosomes and assess their properties and effectiveness in treating lung cancer.Methods Dasatinib-loaded biomimetic exosomes were prepared by merging exosomes derived from human lung cancer H460 cells with drug-loaded liposomes through membrane fusion and self-assembly.The particle size and zeta potential were measured using dynamic light scattering(DLS).The successful fusion of exosomes and liposomes was verified via Western blot and fluorescence resonance energy transfer(FRET)assay.Cellular uptake was quantitatively analyzed using fluorescent probe-based imaging.The in vitro cytotoxicity was assessed by MTS assay,while in vivo tumor targeting,anti-tumor efficacy,and safety were evaluated in subcutaneous tumor-bearing mice through fluorescent imaging and pharmacological studies.Results The dasatinib-loaded biomimetic exosomes exhibited a particle size of(111.3±0.1)nm and a zeta potential of(-30.1±0.3)mV.Western blot confirmed the expression of exosomal markers(CD9,CD81,OXER1)on the biomimetic exosomes.The FRET assay revealed a membrane fusion index of 2.171±0.044,indicating efficient fusion between exosomes and drug-loaded liposomes.Fluorescent imaging demonstrated significantly higher uptake of biomimetic exosomes by homologous lung cancer cells compared to liposomes(P<0.05),with no significant difference observed in heterologous pancreatic cancer cells.The biomimetic exosomes exhibited markedly enhanced cytotoxicity in vitro compared to free dasatinib and liposomal formulations.In vivo studies demonstrated superior tumor accumulation(homing ability),a tumor inhibition rate of 75.0%,and no significant weight loss in mice during treatment.Conclusion Dasatinib-loaded biomimetic exosomes display potent anti-lung cancer activity with favorable safety profiles,suggesting their potential as a therapeutic option for non-small cell lung cancer.

关 键 词：达沙替尼 仿生外泌体 脂质体 非小细胞肺癌 

分 类 号：R734.2[医药卫生—肿瘤]