RGD类似物修饰的阿霉素隐形脂质体的制备及体外细胞结合试验

目的研究用精氨酸-甘氨酸-天冬氨酸(RGD)类似物(RGDm)修饰隐形脂质体(SL)，以增加抗癌药物在肿瘤部位积蓄的同时，增加抗癌药物向肿瘤细胞内的传递。方法合成RGDm，将其通过PEG链与二硬脂酰磷脂酰乙醇胺(DSPE)连接形成导向化合物DSPE-PEG-RGDm，在此基础上制备RGDm修饰的隐形脂质体(RGDm-SL)，阿霉素(DOX)作为模型药物通过硫酸铵梯度法装载。体外实验中，用pH探针(BCECF-AM)标记黑色素瘤细胞，通过细胞黏附试验考察导向化合物与肿瘤细胞的黏附情况；通过流式细胞实验和激光共聚焦显微实验考察肿瘤细胞对SL包封的阿霉素(SL-DOX)及RGDm-SL包封的阿霉素(RGDm-SL-DOX)的结合或摄取情况。结果与DSPE-PEG相比，黑色素瘤细胞与导向化合物DSPE-PEG-RGDm的黏附显著增加，过量游离RGDm的加入能抑制其黏附；与SL-DOX相比，RGDm-SL-DOX与黑色素瘤细胞共同孵育后，细胞对阿霉素的结合及摄取均显著增加。结论RGDm修饰的隐形脂质体可作为肿瘤靶向的载体通过受体介导的方式促进抗肿瘤药物向肿瘤细胞内的传递。

Preparation of doxorubicin-loaded stealth liposomes modified with RGD mimetic and cellular association in vitro

Aim To investigate the possibility of using stealth liposomes modified with arginine-glycine-aspartic acid(RGD) mimetic as the targeted carriers to achieve increased accumulation in tumor and enhanced intracellular delivery for the encapsulated anticancer drugs.Methods RGD mimetic(RGDm) as a ligand for integrins was synthesized and covalently conjugated to the active PEGylated phospholipids(DSPE-PEG-BTC) to form RGDm conjugate(DSPE-PEG-RGDm).Then RGDm-modified SL(RGDm-SL) containing DOX(RGDm-SL-DOX) and SL containing DOX(SL-DOX) were prepared by film dispersion followed by ammonium sulfate gradient method.The pH-sensitive probe,BCECF-AM,was used to study the binding of melanoma cells to DSPE-PEG-RGDm.Flow cytometry and confocal microscopy were performed to evaluate the cellular association or DOX uptake for RGDm-SL-DOX or SL-DOX in vitro.Results The melanoma cells A375 and B16 showed enhanced binding to the immobilized DSPE-PEG-RGDm.The cells treated with RGDm-SL-DOX showed remarkable increase in cellular association or DOX uptake compared with SL-DOX.Conclusion The RGDm-modified SL could be as the targeted carriers to facilitate the delivery of the encapsulated anti-cancer drugs into tumor cells by receptor-mediated way.