硬脂醇半乳糖苷修饰的阿西替尼脂质体的制备及体外活性研究

Study of Preparation and in Vitro Activity of Stearyl Alcohol Galactosidase Modified Axitinib Liposomes

关键词:  阿西替尼  脂质体  硫酸铵梯度法  硬脂醇半乳糖苷  细胞毒性

Key words:  Axitinib  liposomes  transmembrane ammonium sulfate gradients  stearyl alcohol galactosidase  cytotoxicity

摘要:

目的 制备硬脂醇半乳糖苷修饰的阿西替尼脂质体，并进行处方筛选及体外活性研究。方法 采用硫酸铵梯度法制备硬脂醇半乳糖苷修饰的阿西替尼脂质体，以包封率及粒径为评价指标，采用Box-Behnken响应面设计法优化制备工艺，并研究硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株的增殖抑制及凋亡的诱导作用。采用CCK-8法检测硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株的生长抑制情况，采用了Annexin V/PI流式细胞分析法检测细胞凋亡。结果 最佳工艺：药与磷脂比为1∶14.95，胆固醇与磷脂之比为1∶4.45，水浴温度为61.93 ℃，硫酸铵溶液的体积为4.31 mL。硬脂醇半乳糖苷修饰的阿西替尼脂质体的粒径为(252±6.4)nm，包封率为(68.50±0.85)%。CCK-8细胞毒性试验结果显示，在药物浓度相同时，抑制率随时间的延长而增加；作用时间相同时，抑制率随药物浓度的增大而增加。Annexin V/PI流式试验结果显示，硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株的抑制率优于人肝癌A549细胞株。结论 硫酸铵梯度法制备硬脂醇半乳糖苷修饰的阿西替尼脂质体的处方合理，工艺可行，包封率高。硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株有更高的细胞毒性，诱导SMMC-7721细胞株凋亡能力比A549的强。初步判定硬脂醇半乳糖苷修饰的阿西替尼脂质体具有主动肝靶向性。

Abstract:

OBJECTIVE To study presciption screening and in vitro activity of stearyl alcohol galactosidase modified Axitinib liposomes. METHODS Stearyl alcohol galactosidase modified axitinib liposomes were prepared with transmembrane ammonium sulfate gradients. Encapsulation efficiency and particle size were used as the evaluation index. The role of stearyl alcohol galactosidase modified axitinib liposomes in inducing proliferation and apoptosis of liver cancer SMMC-7721 cells was studied. The inhibitory effect of stearyl alcohol galactosidase modified Axitinib liposomes on growth of SMMC-7721 was tested by CCK-8 assay. Apoptosis of SMMC-7721 cells was assayed by Annexin V/PI flow cytometry. RESULTS The best prescription were as follows: the ratio of axitinib to phospholipids was 1∶14.95, the cholesterol and phospholipids ratio was 1∶4.45, the optimum temperature was 61.93 ℃, volume of ammonium sulfate solution was 4.31 mL. The diameter and entrapment efficiency were (252±6.4)nm and (68.50±0.85)%. The result of CCK-8 assay showed that the inhibition rate increased with time going on at the same drug concentration, the inhibition rate increased with the drug concentration increasing at the same time. The result of Annexin V/PI flow cytometry assay showed that the inhibition effect of stearyl alcohol galactosidase modified axitinib liposomes on SMMC-7721 cells were better than A549 under the same conditions. CONCLUSION The selected formulation and preparation technique of stearyl alcohol galactosidase modified axitinib liposomes with transmembrane ammonium sulfate gradients is reasonable and practicable. The axitinib liposomes modified by stearyl alcohol galactosidase have higher cytotoxicity to SMMC-7721 cells, and stronger apoptosis-inducing of SMMC-7721 than A549. The preliminary determination is made that stearyl alcohol galactosidase modified axitinib liposomes are of specific active hepatic targeting.