长春新碱抗CD20免疫脂质体的制备及其对人B细胞淋巴瘤的靶向特异性杀伤作用研究

The Preparation of Vincristine Encapsulated Anti-CD20 Immunoliposomes and the Preliminary Study of Its Target-Specificity and to Human B cell Lymphoma Cell

目的:利用单克隆抗体药物的靶向特异性,探索常规化疗药物长春新碱的免疫脂质体制剂制备方法.方法:通过逆向蒸发法制备空白脂质体,pH值梯度法包裹长春新碱,抗体修饰脂质体制备免疫脂质体制剂.采用流式细胞技术检测免疫脂质体对人B淋巴瘤细胞的靶向特异性.通过细胞增殖、细胞毒性实验来验证长春新碱免疫脂质体对人B淋巴瘤细胞的杀伤作用.结果:粒径分析表明,制备的空白脂质体平粒径在130 nm左右,包裹长春新碱后脂质体平均粒径在140-150 nm左右.流式细胞仪检测结果显示,抗CD20免疫脂质体对人B淋巴瘤细胞Romas和Raji细胞具有明显的靶向特异性.细胞增殖、细胞毒性实验表明,包裹长春新碱的抗CD20免疫脂质体较非免疫脂质体对人B淋巴瘤细胞杀伤效果更好.结论:获得了一种具有免疫靶向的长春新碱脂质体制剂的制备方法,初步的体外实验表明其在人B淋巴瘤治疗方向具有潜在的应用前景.

Objective:To explore a preparation method of vincristine encapsulated immunoliposomes dependent on the target specificity of monoclonal antibody drugs.Methods:Blank liposomes were prepared by reverse-phase evaporation and then the vincristine was encapsulated by pH gradient method.Immunoliposomes were prepared by the conjugation of thiol modified antibodies with maleimide modified PEGylated liposomes.Flow cytometry was used to test the specificity of immunoliposomes targeting human B lymphoma cells.Cell proliferation and cytotoxicity experiments were used to test cytotoxicity of vincristine immunoliposomes to human B lymphoma cells.Results:Size analysis showed that the average particle sizes of the blank liposomes,vincristine encapsulated liposomes and vincristine immunoliposomes were 130 nm,140 nm and 141 nm,respectively.Flow cytometry results showed that the anti-CD20 modified immunoliposomes specifically target the human B lymphoma cells.Cell proliferation and cytotoxicity experiments showed that the anti-CD20 modified vincristine immunoliposomes kill the human B lymphoma cells more effectively than vincristine non-immunoliposomes and free vincristine.Conclution:Obtain a preparation method of vincristine immunoliposomes formulation.Characterization and preliminary analysis of in vitro experiments showed that it has potential applications in the treatment of human B lymphoma.