Poly(L-histidine) Based Triblock Copolymers: pH Induced Reassembly of Copolymer Micelles and Mechanism Underlying Endolysosomal Escape for Intracellular Delivery
Poly(L-histidine) Based Triblock Copolymers: pH Induced Reassembly of Copolymer Micelles and Mechanism Underlying Endolysosomal Escape for Intracellular Delivery
复制标题
基于聚（L-组氨酸）的三嵌段共聚物：pH 诱导的共聚物胶束重组以及细胞内溶酶体逃逸的机制
DOI：
10.1021/bm5010756
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发表时间：
2014-11-01
影响因子：
6.2
通讯作者：
Qiao, Mingxi
中科院分区：
文献类型：
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作者：
Zhang, Xiaojun;Chen, Dawei;Qiao, Mingxi
由于pH响应特性和从内溶酶体途径逃逸，各种基于聚（l-组氨酸）的两亲性共聚物已被开发用于细胞内药物递送。然而，pH诱导的共聚物胶束重组以及共聚物在促进内溶酶体逃逸过程中假定的内溶酶体膜破裂尚未得到阐明。为了解决这些问题，合成了一系列聚乙二醇-聚（d，l-丙交酯）-聚（l-组氨酸）（mPEG-PLA-PHis），它们具有不同程度的聚乳酸和聚乳酸嵌段聚合。利用透射电镜（TEM）、核磁共振（H-1 NMR）、荧光探针技术和动态光散射（DLS）表征了共聚物的自组装和重组行为。在pH值为8.5 ~ 4.5的范围内，TEM图像、pH峰H-1 NMR谱、芘荧光光谱、粒径及粒径分布的变化表明，在酸性环境下，共聚物胶束重新组装成以PLA为疏水核、质子化的PHis和PEG为亲水壳的胶束。pH诱导的重组触发了合并的阿霉素（DOX）释放，表明体外加速药物释放和增强细胞毒性。通过共聚焦激光扫描显微镜（CLSM）观察共聚物促进DOX内溶酶体逃逸过程中内溶酶体膜的完整性，并通过溶血试验和计算内溶酶体膜的临界尺寸进一步评估。结果表明，在共聚物过程中，内溶酶体膜保持完整，促进了DOX的内溶酶体逃逸。更合理的解释是，在不破坏内溶酶体膜的情况下，基于PHis的共聚物促进内溶酶体逃逸为质子海绵假说。
Various poly(l-histidine) based amphiphilic copolymers have been developed for intracellular drug delivery due to the pH responsive properties and the escape from endolysosomal pathway. However, the pH induced reassembly of copolymer micelles and the assumed endolysosome membrane rupture during the copolymer facilitated endolysosomal escape have never been elucidated. To address these issues, a series of poly(ethylene glycol)-poly(d,l-lactide)-poly(l-histidine) (mPEG-PLA-PHis) with different degrees of polymerization of PLA and PHis block were synthesized. The self-assembly and reassembly behaviors of the copolymers were characterized using transmission electron microscopy (TEM), H-1 NMR, fluorescence probe technique, and dynamic light scattering (DLS). The copolymers self-assembled into micelles with PLA and unprotonated PHis blocks as hydrophobic core and PEG as hydrophilic shell at neutral pH. The changes in TEM images, H-1 NMR spectrum of PHis peak, pyrene fluorescene spectrum, and particle size as well as size distribution over the pH range from pH 8.5 to 4.5 suggest that the copolymer micelles reassembled into micelles with PLA as hydrophobic core and protonated PHis and PEG as hydrophilic shell under acidic environment. The pH induced reassembly triggered the incoporated doxorubicin (DOX) release, as indicated by the in vitro accelerated drug release and enhanced cytotoxicity. The integrity of endolysosome membrane during the copolymer facilitated DOX endolysosomal escape was observed by confocal laser scan microscopy (CLSM) and further evaluated by hemolysis test and calculation of the critical size of endolysosomal membrane. The results indicate that the endolysosomal membrane remained intact during the copolymer facilitated endolysosomal escape of DOX. It is more reasonable to ascribe the PHis based copolymer facilitation endolysosomal escape to the proton sponge hypothesis without rupturing the endolysosomal membrane.