自载体的pH响应核-壳型紫杉醇-阿霉素纳米棒协同靶向治疗非小细胞肺癌

Application of self-carrier pH-responsive core-shell paclitaxel-doxorubicin nanorods in the synergistic targeted treatment of non-small cell lung cancer

目的 制备自载体的pH响应核-壳型紫杉醇-阿霉素(PTX-DOX)双药纳米棒,用于非小细胞肺癌的协同性靶向治疗,评价其临床应用效果。方法 采用溶剂交换法制备PTX纳米棒,化学偶联法制备同时包含亲水基团PEG和疏水基团PMHC18的DOX前药DOX-PEG-PMHC18,按照5 ∶ 1的体积比均匀混合两种药物,依次经超声和搅拌处理后组装并制备出核-壳型双药联合纳米棒复合体系PTX-DOX,用扫描电镜(SEM)、透射电镜(TEM)表征纳米棒的形貌和尺寸,紫外-可见分光光度计表征两种药物的复合情况,在不同pH值(pH=7.4,5.0)的生理缓冲液中测试药物的释放曲线,用共聚焦激光扫描显微镜(CLSM)实时表征纳米药物的细胞内吞过程,MTT法测试纳米药物对肿瘤细胞的协同抑制效果,并评价功能化聚合物的生物相容性。结果 经过严格控制实验条件,成功地大量制备形貌尺寸均匀的核-壳型PTX-DOX双药纳米棒(长度500 nm,直径40 nm),505 nm处的特征吸收峰验证了DOX的成功组装,纳米双药可在PBS缓冲液中稳定分散6个月以上而不出现团聚体,当体外释放250 h时,药物在pH=7.4的缓冲液中的释放率为30%,而在pH=5.0的酸性缓冲液中的释放率大于75%,该纳米双药可被肿瘤细胞快速内吞,当细胞与药物纳米棒共孵育2 h时达到内吞峰值且稳定,另外,纳米双药表现出协同增强的抗肿瘤活性,抗癌效果明显优于单药及双药混合物。结论 新型的自载体pH响应核-壳型PTX-DOX双药纳米棒分散性好、性能稳定,可被肿瘤细胞快速内吞并表现出酸性响应的药物释放和协同增强的抗肿瘤活性,为恶性肿瘤靶向治疗提供新的药物设计思路和用药指导。

Objective To prepare the pH-responsive core-shell paclitaxel-doxorubicin(PTX-DOX) dual-agent nanorods, and evalua-te their synergistic targeted efficacy for non-small cell lung cancer. Methods PTX nanorods were prepared by solvent exchange method. Amphiphilic DOX prodrug, DOX-PEG-PMHC18, containing hydrophilic PEG and hydrophobic PMHC18 group, was prepared by chemical coupling method. PTX-DOX core-shell composite nanorods were produced by mixing the two drugs in a volume ratio of 5 ∶ 1 and successively assembling by ultrasonic and stirring treatment. The morphology and size of the nanorods were characterized by scanning electron microscope(SEM) and transmission electron microscope(TEM), the successful combination of the two drugs was characterized by UV-vis. Then, acid-dependent drug release curves were monitored in fluid environments with different pH values(7.4 and 5.0), and the cell entry of nanocomposites was characterized in real time using confocal laser scanning microscopy(CLSM). Finally, the synergistic inhibition of tumor cells and biocompatibility of functional polymers were evaluated by tetrazolium salt colorimetry(MTT). Results After strict control of experimental conditions, PTX-DOX core-shell nanorods with uniform morphology and size were successfully prepared in large quantities(500 nm in length and 40 nm in diameter). The characteristic absorption peak at 505 nm verified the successful assembly of DOX. The nanocomposites were stably dispersed in biological fluids for more than 6 months without agglomeration. After 250 h release, 30% of DOX was released in a humoral environment with pH=7.4 and 75% released in an acidic environment with pH=5.0. The nanorods were promptly internalized into cancer cells and showed plateau after 2 h incubation. Furthermore, the dual-drug nanorods exhibited significantly better synergistic anti-tumor activity than single drug or the mixture of two drugs. Conclusion The novel pH-responsive PTX-DOX core-shell nanorods show good dispersibility and stability, and it can be rapidly internalized by tumor cells, thus displaying acid-stimulated drug release and synergistically enhanced anti-tumor activity. These promising results will provide ingenious drug design ideas and guidance for targeted therapy of malignant tumors.