pH和还原双重生物响应氨基酸聚氨酯纳米药物载体的设计合成

In this project, novel pH and reduction dual-bioresponsive micelles will be developed based on poly(ethylene glycol)-poly(amino acid-urethane)-poly(ethylene glycol) (PEG-b-SSPAAU-b-PEG) triblock copolymers for efficient intracellular delivery of anti-cancer drugs such as DOX and PTX. SSPAAU is prepared from condensation polymerization of disulfide-containing diserine or dithreonine diol with L-lysine methyl ester diisocyanate or 1,4-diisocyanatobutane followed by ammonolysis reaction to introduce pH-sensitive functional groups like imidazole. The molecular weight, composition, disulfide content as well as pH-sensitive group content of triblock copolymer can be well controlled. Using heterodifunctional PEG, targeting ligands such as RGD peptides, folic acid, and galactose can be easily introduced to achieve tumor-specific cellular uptake. The polyurethanes developed by this project are based on amino acids and should therefore possess good biocompatibility. The pH sensitivity will help micelles to partially release drugs in endosomal compartment as well as facilitate endosomal escape of micelles via proton-sponge effect. The reduction sensitivity will lead to rapid disruption of micelles and complete release of drugs in the cytoplasm, achieving potent anticancer effect. In this project, synthesis of SSPAAU, preparation of drug-loaded micelles, pH and redox-responsive drug release, in vivo biodistribution and antitumor efficacy of drug-loaded micelles will be investigated.

本项目将设计合成pH和还原双重生物响应性聚乙二醇-氨基酸聚氨酯-聚乙二醇（PEG-b-SSPAAU-b-PEG）纳米胶束载体，用于临床抗癌药物如阿霉素和紫杉醇等的高效肿瘤细胞内释放。其中，SSPAAU由含双硫键的二丝氨酸或二苏氨酸二醇单体与L-赖氨酸乙酯二异氰酸甲酯或1,4-二异氰酸丁酯缩聚后通过胺解反应引入咪唑等pH响应基团制备得到。聚合物的分子量、组成、双硫键和pH响应基团含量等可控。采用异官能化PEG可引入多肽、叶酸和半乳糖等生物靶向分子，提高胶束的肿瘤特异性内吞。本项目设计的聚氨酯基于氨基酸，可望拥有优异的生物相容性；其pH响应性不但促使部分药物在内涵体释放，且可望通过质子海绵效应帮助胶束逃离内涵体进入细胞质；其还原响应性可望实现高效细胞质内药物释放，产生高抗癌效果。我们将具体研究氨基酸聚氨酯的合成、载药胶束的制备、响应性药物释放、体内生物分布和肿瘤治疗效果。

为了加快药物在肿瘤组织或者细胞内的释放，本项目设计合成了pH 和还原生物响应性聚乙二醇-聚氨酯-聚乙二醇纳米胶束载体，用于临床抗癌药物阿霉素的高效肿瘤细胞内释放。主要结果如下：（1）还原敏感PEG-AAPU(SS)-PEG胶束的制备及其用于DOX在细胞内的递送。AAPU(SS)由含双硫键的二丝氨酸二醇单体与L-赖氨酸乙酯二异氰酸甲酯缩聚后制备得到。聚合物的分子量、组成和双硫键含量可控。这是一种完全由氨基酸单元组成的聚氨酯，具有优良的生物相容性。在此基础上合成的PEG-AAPU(SS)-PEG胶束可高效包载DOX，并在细胞内还原环境快速释放DOX。表面修饰了c(RGDfK)多肽的PEG-AAPU(SS)-PEG胶束对U87 MG细胞表现出显著的特异性内吞，细胞杀伤力显著增强；（2）pH 和还原双重敏感PEG-AAPU(SS)(-g-his)-PEG的制备及其用于DOX在细胞内的递送。AAPU(SS)(-g-his)是在AAPU(SS)基础上通过侧链胺解反应引入组胺得到的。PEG-AAPU(SS)(-g-his)-PEG胶束具有明显的pH和还原响应性，能快速从内涵体逃离到细胞质还原环境中，并在细胞质中快速释放出包载的DOX。与仅具有还原敏感的载药胶束相比，对RAW 264.7和MCF-7/ADR细胞均表现出了更强的细胞毒性。此外，我们还研究了pH和还原双重敏感的PEG-SSPU(TMBPE)-PEG胶束对DOX 的包载、靶向释放和抗肿瘤效果。该项目的研究结果表明pH和还原响应聚氨酯胶束体系可实现药物在细胞内的高效释放，进而产生高抗癌效果，该研究结果为刺激响应聚氨酯胶束体系的临床前试验奠定了坚实的基础。