微波消融联合温敏载药胶束用于乳腺癌局部协同治疗的研究

The residual lesion after microwave ablation (MWA) is an important problem in the clinical application of MWA in breast cancer. The local temperature of tumor lesion is higher during the treatment of MWA, which can be applied to trigger responsive drug release from thermosensitive nanocarriers resulting in effective suppression of the residual lesion. MWA can also induce intracellular production of reactive oxygen species which facilitates the translocation of drugs into cytoplasm and thus enhances the cytotoxicity to tumor cells. Moreover, Doxorubicin is able to inhibit the expression of heat shock protein 70 and therefore improve the ablation efficiency of MWA. Taking into account these properties, thermosensitive polymer micelles loaded with Doxorubicin are designed for synergistic therapy of MWA and chemotherapeutics. In this work, responsive release behavior of Doxorubicin triggered by MWA is evaluated by the drug release experiment; The effect of reactive oxygen species on the translocation of doxorubicin is observed by subcellular localization experiment; The inhibition of HSP70 is evaluated by Western Blotting; CCK-8 assay and in vivo anticancer experiment are carried out to verify the synergistic effect of the combination therapies on breast cancer at the cellular and animal levels, respectively. This work is highly valuable to explore efficient anticancer method to inhibit residual lesion after the treatment of MWA.

微波消融（Microwave Ablation，MWA）后的残留病灶是其应用于乳腺癌临床亟待解决的重要问题。MWA治疗过程中肿瘤病灶局部温度增高，该特性可用于设计温敏纳米载体使其在MWA的高温刺激下响应性释放化疗药物，实现对残留病灶的高效抑制。MWA本身还可诱导胞内产生活性氧自由基促进药物胞浆分布，进一步提高化疗效果；化疗药物阿霉素可抑制热休克蛋白70的表达，从而反向增强MWA热疗效应。基于此，本项目结合MWA和化疗各自的优势，构建温敏聚合物胶束负载阿霉素进行MWA和化疗的联合治疗研究。通过释放实验明确MWA在温敏胶束响应性释药中的关键作用，亚细胞定位实验观察活性氧对阿霉素胞浆分布的影响，以及分子生物学实验考察阿霉素对热休克蛋白的抑制情况，并分别通过CCK-8和抑瘤实验在细胞、动物水平验证联合治疗对乳腺癌的协同抑制效果，从而为解决MWA治疗后的残留病灶问题提供帮助。

乳腺癌是女性最常见的恶性肿瘤，增强乳腺癌局部治疗效果是临床面临的重要问题。本项目构建了放化疗一体的纳米载药体系，通过调控化疗药物在肿瘤组织富集、细胞摄取、释放及胞浆分布并利用Au进行放疗增敏，增强乳腺癌局部协同治疗效果。具体研究内容如下：以天然氨基酸为原料合成了生物相容性好、安全性高两亲性多肽（SPP），细胞实验证实多肽纳米材料溶液对鼠源乳腺癌细胞(4T1)以及正常细胞（成纤维细胞）的安全性均较好。通过溶剂透析法成功负载上了化疗药物阿霉素，并通过静电相互作用负载上放疗增敏剂Au制备出Au@SPP@DOX载药胶束，表征实验证实胶束粒径较好（~150 nm）、载药量较高，偏中性电荷有助于胶束体系在肿瘤部位聚集，体外实验证实胶束具有缓释的作用，在肿瘤部位酸性环境下药物释放增多。胶束可被4T1乳腺癌细胞有效摄取并定位于溶酶体酸性环境中。X射线辐照时Au对X射线的增敏比为1:2.95，通过诱导ROS产生增多显示出增强的抗肿瘤效果。溶酶体染色实验证实ROS不仅可以导致细胞凋亡，而且可以促进溶酶体膜破裂，实现化疗药物溶酶体逃逸，更多地作用于细胞核、线粒体等靶点。动物实验证实Au@SPP@DOX胶束的肿瘤靶向性较好，在48 h时肿瘤部位信号最强，且该联合治疗模式局部抗肿瘤效果最佳。该项目为乳腺癌局部高效协同治疗进行了有益的探索。