多柔比星纳米胶束抗肿瘤作用与机制：Pluronic的影响

Due to the toxicity and low efficiency of cytotoxic pharmaceuticals, the therapeutic effect of chemotherapy is still unsatisfied. Using Nanometer carrier, it is possible to improve the efficiency of clinical available cytotoxic pharmaceuticals if the drugs could be delivered to the tumor cells far away from the vessels, then bind to the DNA in the cellular nucleus. It will also be helpful if the nanometer carrier could overcome the multidrug resistance (MDR) to somehow. The molecular weight of Pluronic have great impact on the characteristics of nano-micelles. In this study, based on our former research, doxorubicin-loaded nanomicelles will be prepared using Pluronic of different molecular weights. Cellular nucleus target of the drugs will be studied using Laser confocal scanning microscopy and spectrometry. The penetration and retention of the loaded drugs will be measured via multicellular spheroids and tumor bearing nude mice. Multidrug resistance studies will also be performed. Finally, the anticancer effect of prepared doxorubicin-loaded nanomicelles will be exammed in the levels of monolayer cellular culture, multicellular spheroids model and tumor bearing nude mice. We are investigating a novel route to improve the efficiency of clinical available cytotoxic pharmaceuticals through more efficient tumor penetration, nucleus target and overcome of multidrug resistance.

化疗的疗效仍有较大的提升空间。利用纳米载体，改善药物在肿瘤无血管区中的穿透与滞留能力，使药物更容易进入远离微血管的肿瘤细胞；实现化疗药的细胞核DNA靶向；减少或逆转肿瘤多药耐药就可能使目前已有的化疗药发挥更大的抗肿瘤作用。Pluronic是制备纳米胶束的主要材料，其分子量影响纳米胶束的特性。本课题在前期工作基础上，以不同分子量（PEO长度）的Pluronic构建多柔比星纳米胶束，激光共聚焦显微镜和光谱技术从定位定量两方面评价细胞核靶向；采用三维多细胞球体技术和移植瘤模型，研究多柔比星渗入肿瘤实质的深度及瘤内滞留时间；进而评价多柔比星纳米胶束克服多药耐药情况；再从单层细胞培养MTT试验、克隆形成能力、多细胞球体生长与迁移以及对移植瘤的抑制作用等方面研究多柔比星纳米胶束的抗肿瘤效应。本研究从克服多药耐药、改善化疗药的细胞核靶向和瘤内无血管区穿透与滞留能力的角度，探讨提升化疗药抗肿瘤作用的新途径

肿瘤耐药是临床上常见的问题，是导致化疗失败的常见原因。许多研究表明，普郎尼克胶束具有逆转肿瘤细胞耐药的效应。本研究以PCL-PEG和不同分子量的普郎尼克为胶束主材，使用溶剂挥发法制备一系列多柔比星纳米胶束。激光共聚焦显微镜和光谱技术从定位定量两方面评价多柔比星的亚细胞分布与过程；采用三维多细胞球体技术和移植瘤模型，研究多柔比星渗入肿瘤实质的深度及在瘤内的滞留；溴化四氮唑蓝(MTT)法测定阿霉素和阿霉素载药胶束对肺癌A549细胞、乳腺癌耐阿霉素细胞MCF7/Adr及普通的MCF7细胞的抑制作用。从克隆形成能力、多细胞球体生长与迁移等方面研究多柔比星纳米胶束的抗肿瘤效应。Western blot及RT-PCR检测细胞多药耐药蛋白p-糖蛋白（p-gp）、多药耐药相关蛋白(MRP1)、乳腺癌耐药蛋白（BCRP）表达的变化，评价多柔比星纳米胶束克服多药耐药情况。研究发现，Dox-PEG-PCL-P105比Dox-PEG-PCL-F68和Dox-PEG-PCL-P85更容易进入实验肿瘤细胞，且滞留时间明显更长。与线粒体共定位，与溶酶体部分定位。以P105 制备的纳米胶束Dox-PEG-PCL-P105比以F68制备的Dox-PEG-PCL-F68更易穿透瘤体，且在瘤内滞留时间更长。阿霉素胶束可能通过降低MRP1表达逆转MCF7/Adr细胞的耐药性。