羟基喜树碱铁蛋白包合物的制备及其抗结肠癌活性的研究

Study on the preparation and anti-colon cancer activity of hydroxycamptothecin loaded ferritin inclusion complex

目的用铁蛋白(ferritin,Fn)包合羟基喜树碱(hydroxycamptothecin,HCPT)制备羟基喜树碱铁蛋白包合物(hydroxycamptothecin@ferritin,HCPT@Fn),增加HCPT的溶解性,提高治疗结肠癌的效果。方法用透析法制备HCPT@Fn;用激光粒度测定仪检测HCPT@Fn的粒径及Zeta电位;用超高效液相色谱法(ultra-high performance liquid chromatography,UPLC)检测HCPT@Fn中HCPT的载药量;用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide,MTT]实验、活死细胞染色实验及克隆形成实验考察HCPT@Fn对人结肠癌细胞(human colon cancer cells,HT-29)的毒性;用荧光显微镜观察HCPT@Fn在HT-29细胞内的蓄积;用溶血实验及HE染色实验初步考察HCPT@Fn的安全性。结果用透析法制备的HCPT@Fn的粒径为148.4 nm,Zeta电位为−34.4 mV,载药量为37.46%;HCPT@Fn能大量蓄积于HT-29细胞中,可显著抑制HT-29细胞的增殖及克隆形成;在设定的质量浓度范围内,HCPT@Fn均未造成明显的溶血,HCPT@Fn对小鼠正常组织无明显破坏性。结论HCPT@Fn能够显著增加HCPT的溶解性,提高抗结肠癌活性并降低毒性和不良反应。

Objective In order to increase the solubility and improve the therapeutic effect of colon cancer,a hydroxycamptothecin(HCPT)loaded ferritin(Fn)inclusion complex(hydroxycamptothecin@ferritin,HCPT@Fn)was prepared.Methods Dialysis method was used to prepare HCPT@Fn.Then,the particle size and Zeta potential of HCPT@Fn were evaluated by laser particle size analyzer.The drug loading of HCPT@Fn was determined by ultra-high performance liquid chromatography(UPLC).The cytotoxicity of HCPT@Fn on human colon cancer cells(HT-29)cells was investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assay,alive/dead cell staining test and clone formation assay.The uptake of HCPT@Fn in HT-29 cells was detected by fluorescence microscope.The safety of HCPT@Fn was investigated by hemolysis test and HE staining preliminarily.Results The particle size and Zeta potential of HCPT@Fn were 148.4 nm and−34.4 mV,respectively.The drug loading of HCPT in HCPT@Fn was 37.46%.HCPT@Fn can accumulate in large amounts in HT-29 cells,inhibiting the proliferation and clone formation of HT-29 cells significantly.In the set concentration range,HCPT@Fn did not cause significant hemolysis,and did not cause significant damage to normal tissue in mice.Conclusion HCPT@Fn could increase the solubility of HCPT,enhance the anti-colon cancer activity of HCPT and reduce toxic side effects.