Department of Materials Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
Department of Bio-Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
J Control Release. 2014 Aug 28;188:67-77. doi: 10.1016/j.jconrel.2014.05.048. Epub 2014 Jun 2.
The ubiquitin-proteasome system is central in the regulation of cellular proteins controlling cell cycle progression and apoptosis, drawing much interest for developing effective targeted cancer therapies. Herein, we developed a novel pH-responsive polymeric-micelle-based carrier system to effectively deliver the proteasome inhibitor MG132 into cancer cells. MG132 is covalently bound to the block copolymer composed of polyethylene glycol (PEG) and polyaspartate through an acid-labile hydrazone bond. This bond is stable at physiological condition, but hydrolytically degradable in acidic compartments in the cell, such as late-endosomes and lysosomes, and thus, it was used for controlled release of MG132 after EPR-mediated preferential accumulation of the micelles into the tumor. MG132-loaded micelles have monodispersed size distribution with an average diameter of 45nm, and critical micelle concentration is well below 10(-7)M. In vitro studies against several cancer cell lines confirmed that MG132-loaded micelles retained the cytotoxic effect, and this activity was indeed due to the inhibition of proteasome by released MG132 from the micelles. Real-time in vitro confocal-microscopy experiments clearly indicated that MG132-conjugated micelles disintegrated only inside the target cells. By intravital confocal micro-videography, we also confirmed the prolonged circulation of MG132 loaded micelles in the bloodstream, which lead to tumor specific accumulation of micelles, as confirmed by in vivo imaging 24h after injection. These micelles showed significantly lower in vivo toxicity than free MG132, while achieving remarkable antitumor effect against a subcutaneous HeLa-luc tumor model. Our findings create a paradigm for future development of polymeric-micelle-based carrier system for other peptide aldehyde type proteasome inhibitors to make them effective cohort of the existing cancer therapeutic regiments.
泛素-蛋白酶体系统在调节细胞周期进程和细胞凋亡的细胞蛋白中起着核心作用，这为开发有效的靶向癌症治疗方法提供了很大的兴趣。在此，我们开发了一种新型的 pH 响应型聚合物胶束载体系统，有效地将蛋白酶体抑制剂 MG132 递送到癌细胞中。MG132 通过酸不稳定的腙键共价结合到由聚乙二醇（PEG）和聚天冬氨酸组成的嵌段共聚物上。该键在生理条件下稳定，但在细胞内的酸性隔室（如晚期内体和溶酶体）中可水解降解，因此，它可用于在 EPR 介导的优先积累胶束进入肿瘤后控制 MG132 的释放。载有 MG132 的胶束具有单分散的粒径分布，平均直径为 45nm，临界胶束浓度远低于 10(-7)M。对几种癌细胞系的体外研究证实，载有 MG132 的胶束保留了细胞毒性作用，而这种活性实际上是由于从胶束中释放的 MG132 抑制了蛋白酶体。实时体外共聚焦显微镜实验清楚地表明，只有在靶细胞内，MG132 缀合的胶束才会解体。通过活体共聚焦显微镜视频，我们还证实了载有 MG132 的胶束在血液中的循环时间延长，这导致了胶束在肿瘤中的特异性积累，这在注射后 24 小时的体内成像中得到了证实。与游离 MG132 相比，这些胶束的体内毒性显著降低，同时对皮下 HeLa-luc 肿瘤模型也表现出显著的抗肿瘤作用。我们的研究结果为基于聚合物胶束的载体系统的未来发展创造了范例，用于将其他肽醛类蛋白酶体抑制剂作为现有癌症治疗方案的有效组成部分。
