基于纳米胶束的肿瘤治疗和成像一体化的多功能药物传递系统的构建与评价

Multidrug resistance and bioimaging are important problems to be sloved in cancer treatment. Our research team found that miR-302S can directly regulate BCRP expression and reverse chemotherapeutic drug resistance in breast cancer. Given the growing needs for theranostic micelle systems, on the basis of polymer synthesis, we construct a multi-functional drug delivery system for co-delivery of miRNA and chemotherapeutic drugs. With the help of aptamer, pH-responsive hydrazone bond and biodegradable photo-luminescent polymers, this delivery system exhibits tumor-targeting, pH-responsive controlled drug release and imaging abilities. This drug delivery system with high miRNA and chemotherapeutic drugs loading is prepared to improve the stability of miRNA, reduce the toxicity of chemotherapeutic drugs, reduce chemotherapy drug efflux and furthermore enhance the antitumor effect of chemotherapeutic agents. Cytotoxicity assay, flow cytometry, confocal microscopy, in vivo imaging instrument, real-time PCR and western blot are employed to verify the abilities of this delivery to be functioning as an imaging probe, improve intracellular chemotherapeutic drug concentration and then suppress tumor growth. They are also used to clarify the mechanisms of reversing multidrug resistance in vitro and in vivo. The development of this novel multifunctional drug delivery system not only lays a foundation for the universal drug co-delivery system for nucleic acid and chemotherapy drug, but also provides a new approach for targeted cancer theranostics.

肿瘤治疗与诊断一体化是临床中提高肿瘤治愈率急需解决的问题。课题组研究发现miR-302S可以调节乳腺癌耐药蛋白表达，基于聚合物合成的工作基础，为解决肿瘤成像诊断、多药耐药及miRNA体内递送的问题，本研究拟构建共传递miRNA和化疗药物的多功能递送系统。借助Aptamer使系统靶向肿瘤部位，通过pH敏感的腙键控制药物释放，借助生物可降解荧光聚合物使胶束可作为荧光探针。通过制备该传递系统，提高基因药物稳定性，降低化疗药毒性，提高化疗药的抗肿瘤效果并对肿瘤实时诊断成像。通过流式细胞仪、共聚焦显微镜、活体成像仪和western blot等表征手段在体内外验证该传递系统的靶向性、荧光成像诊断、减少化疗药物外排和抑制肿瘤生长的能力，阐明其逆转多药耐药的机制。该多功能传递系统的构建，不仅为建立一种通用性较强的核酸类药物和化疗药物共传递系统奠定了基础，也为基于临床诊断治疗学进行载体设计提供了一个新思路。

为逆转肿瘤多药耐药及提高肿瘤治愈率，构建了高分子载体COOH-PEG-PAHy-GTA-OCC-DOX，制备了以AS1411为靶向基团的同时包载DOX和miR-302S的pH敏感聚合物胶束。由于载体中具有腙键，该胶束在pH5.0的酸性介质中36h时可以释放药物约42%，释放速率大于pH6.8和7.4，腙键的断裂造成胶束的解聚，使载体骨架上连接的DOX和内部包载的DOX释放较快。在体外实验中，该胶束对基因药物miR-203S具有一定的保护作用。通过细胞内药物蓄积实验、细胞毒实验及western blot实验表明，在耐药细胞中，该胶束使细胞内化疗药物浓度及细胞毒性均高于DOX组，一是由于胶束的靶向作用和内吞途径，二是因为miR-302S下调耐药蛋白的表达，减少了细胞内化疗药物的外排，逆转了肿瘤多药耐药，抑制肿瘤细胞的生长。在体内实验中，该胶束具有抑制肿瘤生长的作用，使基因药物和化疗药物协同发挥作用，抑制肿瘤生长的效果优于单一使用化疗药物。本研究制备的聚合物胶束具有pH敏感性及靶向性，实现了化疗药物和基因药物的协同发挥作用，逆转了肿瘤多药耐药，对肿瘤的治疗具有一定的研究意义。