pH敏感型热休克笼形蛋白纳米载体的构建及其理化性质评价

Preparation and Evaluation of Physicochemical Properties of pH Sensitive Nanocarriers Based on Cage-like Heat Shock Proteins

关键词:  热休克蛋白  聚乙二醇  Tat肽  pH敏感  纳米载体

Key words:  heat shock proteins(HSP)  PEG  Tat  pH-sensitive  nanocarriers

摘要:

目的 设计并制备具有靶向肿瘤且pH敏感的热休克蛋白（heat shock proteins，HSP）笼形蛋白纳米递药系统，并对其理化性质进行表征。方法 采用基因全合成与蛋白质重组表达技术纯化HSP为母版，通过表面官能团功能化制备得到修饰穿膜肽Tat、聚乙二醇包衣的热休克笼形蛋白纳米载体（PT-HSP）。通过透射电镜、纳米粒度与Zeta电位测定仪对其形态、粒径及Zeta电位进行表征，并建立HPLC测定其载药量与包封率。考察载紫杉醇（paclitaxel，PTX）的PT-HSP在生理pH条件（pH 7.4）与肿瘤pH条件（pH 6.5）下的体外释药行为。结果 形态学结果表明，PT-HSP是呈现典型双层结构的均一球体，平均粒径为（154.4±23.6） nm，Zeta电位为（-2.6±0.7） mV。HPLC测得载PTX的PT-HSP的包封率为（75.3±3.6）%，载药量为（7.0±0.2）%。体外释药试验结果表明PT-HSP在pH 7.4条件下的释放速率显著慢于pH 6.5条件下的释放速率（P<0.01）。结论 本研究制备得到的pH敏感的HSP笼形蛋白智能纳米递药系统具有载药量高、稳定性强及智能靶向等优点，有望成为一种安全、有效、智能的抗肿瘤药物载体。

Abstract:

OBJECTIVE To design and prepare a tumor-targeted pH-sensitive delivery system based on heat shock proteins(HSP) nanocages, and to evaluate its physicochemical properties. METHODS Using HSP as the matrix, which were prepared by total gene synthesis and protein recombination expression technology, HSP nanocages modified with cell-penetrating peptides Tat and PEG(PT-HSP) were produced via the functionalization of surface groups. Morphology was examined by TEM, particle size and Zeta potential were determined by nanometer particle size and zeta potential measuring instrument. Encapsulation efficiency and loading capacity were determined by HPLC. The in vitro release behavior of paclitaxel-loaded PT-HSP under pH 7.4 and pH 6.5 was investigated. RESULTS Morphological results showed that PT-HSP presented uniform spherical with a typical two-layer structure, the average size of PT-HSP was (154.4±23.6)nm, Zeta potential was (−2.6±0.7)mV. Encapsulation efficiency and loading capacity of PT-HSP detected by HPLC were (75.3±3.6)% and (7.0±0.2)%. The results of in vitro release test showed that the release rate of PT-HSP at pH 6.5 was faster than pH 7.4(P<0.01). CONCLUSION The pH-sensitive intelligent HSP nanocages drug delivery system constructed in this study, is expected to become a safe, effective and intelligent antitumor drug delivery carriers by its high loading capacity, good stability and intelligent tumor-targeting.