发光/磁性杂合靶向纳米胶束联合阿霉素和miR-34a治疗肿瘤协同增敏和双模态成像研究

The ultimate objective of nanomedicine is to develop targeted nanocarriers for site-specific delivery of bioactives to achieve high therapeutic efficacy as well as excellent molecular imaging. It is great significant how to reverse the multidrug resistance (MDR) and promote the synergistic sensitivity of cancer cells to chemotherapeutic drugs, and resulting in efficient anti-cancer therapy. In the presence, theranostics is more attractive in cancer treatment and diagnosis. In this research proposal, we will design a novel type of multifunctional nanomicelles doped Fe3O4 and doxorubicin (Dox) by self-assembly of mPEG-PCL-PEI, FA-PEG-PCL and Cy7-PEG-PCL. The nanomicelles will be further modified to entrap the miR-34a by electrostatic absorption. This novel multifunctional nanomicelles (FA/Cy7-miR-34a/(Dox/Fe3O4)-nanomicelles) could be realized their functions, i.e., co-delivery of chemotherapeutic drug and miRNA, and dual-modality imaging (fluorescence imaging and MR imaging) in vitro and in vivo. The nanomicelles uptake, intracellular distribution, cell viability, apoptosis, and some key targeted proteins expression will be investigated. On the other hand, the mechanisms of synergistic sensitivity and MDR downregulation will further explored. Finally, we will test the feasibility of FA/Cy7-miR-34a/(Dox/Fe3O4)-nanomicelles as a new type of dual-modality contrast agent. This proposed research will expand the knowledge on theranostic nanomedicine, and also significant to realize the synchronous therapy and diagnosis by one nanocarriers for cancer treatment. The expected outcomes will provide more information for future clinic application of the multifunctional nanomicelles.

提高肿瘤细胞对化疗药物增敏效果，降低其多耐药性，同时进行成像与诊断，是肿瘤防治的一个重要发展趋势。本项目围绕如何实现肿瘤诊疗一体化这一中心目标，在前期研究基础上，以生物相容性好的聚乙二醇-聚己内酯-聚乙烯亚胺(mPEG-PCL-PEI)为材料，通过修饰叶酸和近红外成像荧光分子Cy7，采用分子自组装方式将超顺磁性Fe3O4纳米粒子和抗肿瘤药物阿霉素包埋在胶束中，然后静电吸附miR-34a，构建一种可以实现Dox/miR-34a共输送并兼具荧光/MR双模态成像的功能化靶向纳米胶束，深入分析此功能化靶向纳米胶束输送(内化)规律，探讨其协同增敏和降低肿瘤细胞耐药性的细胞分子机制和体内药效学分析，并在离体和在体水平上研究此功能化靶向纳米胶束对肿瘤的治疗效果、及其荧光/MR双模态成像规律。本课题的开展将加深对协同增敏机制的理解与认识，为开发诊疗一体化的多功能纳米载体及其临床应用提供实验证据。

提高肿瘤细胞对化疗药物增敏效果，降低其多耐药性，同时进行成像与诊断，是肿瘤防治的一个重要发展趋势。本项目围绕如何实现肿瘤诊疗一体化这一中心目标，以嵌段聚合物为骨架材料，携载化疗药物DOX、miR-34a，构建了一种可以实现Dox/miR-34a共输送并兼具荧光/MR双模态成像的功能化靶向纳米胶束，即Gd-CM-Dox/34a。通过TEM、核磁共振等确定其化学组成和形貌特征；荧光成像、MR成像、3D细胞球渗透实验研究其渗透性和肿瘤部位聚集特性；体内外实验研究其安全性和抗肿瘤效果。结果表明，该纳米胶束粒径50nm左右，形貌均一，DOX载药率6.2%。包封率81.3%。其MRI弛豫率比对照高出1.4倍，该纳米胶束能帮助miR-34a进行溶酶体逃逸以及DOX释放入核。miR-34a下调了Bcl-2, cyclin D1, CDK6, 和Bax等下游靶基因的表达，抑制了乳腺癌细胞的增殖和转移。纳米胶束的粒径有利于其渗透进入实体肿瘤内部，在3D肿瘤球及实体瘤模型中，与对照相比，该纳米胶束在肿瘤部位聚集更多，MRI成像及联合治疗的能力最强。在治疗第19天，与对照相比，纳米胶束实验组对肿瘤抑制率达到了80%。该纳米胶束能增加药物/基因和成像分子在肿瘤部位的渗透和积累，实现对肿瘤部位和药物分布的实时监控，有望成为一种应用于临床的多功能纳米复合药物，实现对肿瘤高效诊断和治疗。