曲妥珠单抗-纳米金探针的制备及对乳腺癌细胞的抑制作用  

Preparation of herceptin-gold nanoprobe and its inhibitory effect on breast cancer cells

作　　者：苗凤真 刘琳[1] 宋丽娜[2] 周彦生[2] 张宇[2] 李苏宜[1,3] 

机构地区：[1]东南大学附属中大医院肿瘤科,江苏南京210009 [2]东南大学生物科学与医学工程学院江苏省生物材料与器件重点实验室,江苏南京210009 [3]安徽省肿瘤医院肿瘤内三科,安徽合肥230031

出　　处：《东南大学学报（医学版）》2015年第4期507-513,共7页Journal of Southeast University(Medical Science Edition)

基　　金：江苏省基础研究计划(自然科学基金-重点研究专项)项目(BK2011036)

摘　　要：目的:构建新型曲妥珠单抗-纳米金生物探针(herceptin-GNPs),探寻纳米靶向治疗乳腺癌新方法。方法:采用柠檬酸三钠还原法制备金纳米粒子(GNPs),通过静电吸附作用偶联曲妥珠单抗(herceptin)合成herceptin-GNPs;MTT法检测GNPs、herceptin、herceptin-GNPs对HER2阳性乳腺癌BT474细胞体外增殖抑制作用;流式细胞术检测BT474细胞凋亡率和周期变化;蛋白质印迹技术检测p-AKT和Bcl-2蛋白表达。结果:GNPs水溶液澄清透亮,静电吸附成功制备herceptin-GNPs探针。GNPs浓度在100μg·ml-1及以下未显示明显抑制BT474细胞增殖作用。不同浓度herceptin和herceptin-GNPs对BT474细胞生长具有剂量依赖的抑制作用;与herceptin相比,herceptin-GNPs抑制作用更强。流式细胞术检测显示herceptin、herceptinGNPs均诱导BT474细胞凋亡,阻滞细胞于G0/G1期,且herceptin-GNPs作用强于herceptin。herceptin、herceptin-GNPs均可下调抑凋亡蛋白Bcl-2表达,降低AKT的磷酸化水平,且herceptin-GNPs作用强于herceptin。结论:本研究合成的herceptin-GNPs具有良好的生物相容性,较herceptin抑制乳腺癌BT474细胞生长作用更强,可减少herceptin给药剂量。herceptin-GNPs可能为HER2过表达乳腺癌的靶向治疗提供新方法。Objective： To construct an innovative biocompatible nanoprobe-herceptin-gold nanoparticles（ GNPs）,and to explore new targeted therapies in breast cancer. Methods： GNPs were synthesized by the classic citrate reduction method and the herceptin conjugated to GNPs by electrostatic adsorption. Setting 5 concentration gradients of GNPs from 6. 25 μg·ml^-1to 100 μg·ml^-1,herceptin and herceptin-GNPs from 0. 625 μg·ml^-1to10 μg· ml^-1along with their control group to detect the cytotoxicity by MTT assay. The apoptosis,cell cycle changes were detected by flow cytometry and the expressions of p-AKT and Bcl-2 protein were detected by Western Blot,respectively. Results： The GNPs aqueous solution obtained were fuchsia and clear. The GNPs showed no cytotoxicity under the designed concentration gradients,which displayed good biocompatibility. Herceptin and herceptin-GNPs both could suppress cell growth,induce apoptosis and block cell proliferation in G0/ G1phase; the latter had better effect. Moreover,both herceptin and herceptin-GNPs could inhibit the expressions of p-AKT and Bcl-2 protein,and the latter had better inhibitory effect. Conclusion： The herceptin-GNPs are biocompatible.Compared with herceptin,the herceptin-GNPs have better inhibitory effect on HER2 overexpressing breast cancer cell proliferation,which can reduce the dosage of herceptin. It would provide a nanoparticle-based targeted delivery system for the treatment of HER2 overexpressing breast cancer.

关 键 词：乳腺癌细胞 人表皮生长因子受体-2 曲妥珠单抗 纳米探针 金纳米粒子 细胞凋亡 

分 类 号：R737.9[医药卫生—肿瘤] R318[医药卫生—临床医学]