ABSTRACT

Polymersomes are self-assembled nanostructures with high loading capacity, possibility to deliver hydrophilic as well as hydrophobic drugs and stealth characteristic resulting in low immunogenicity. These vesicles can be used to deliver enzyme drugs such as L-asparaginase (ASNase), a first line drug for acute lymphoblastic leukemia treatment. Here, polymersomes based on three semi-crystalline copolymers of poly(ethylene glycol)-b-poly(ε-caprolactone), namely PEG₄₅PCL₄₄, PEG₁₁₄PCL₉₈, and PEG₁₁₄PCL₁₁₄, were investigated for the encapsulation of ASNase, as well as of bovine serum albumin (BSA) as a model protein. Critical aggregation concentration (CAC) of the copolymers was determined by fluorescence spectroscopy and the values varied from 0.6 to 1.26 mg/L. Using film hydration, polymersomes of 200–400 nm and narrow size distribution (polydispersity index values of 0.2–0.3) were obtained when centrifugation was used as a post-film technique. The encapsulation efficiency (EE %) was determined either after centrifugation of the suspension, followed by the proteins measurement in the supernatant or after Size Exclusion Chromatography (SEC) purification, in which the quantification was performed in the eluted fractions corresponding to the free protein. Higher encapsulation efficiency values were obtained after centrifugation (EE% ≈ 20%) in comparison to the measurements after SEC (EE% ≈ 1–5%), indicating that the protein could be partially entrapped in the polymeric network when centrifugation is used as separation method. Nonetheless, this system could provide an initial effect of the free ASNase followed by a long-term effect based on the encapsulated enzyme, leading to decreased dose administration.

摘要

聚合物小体是具有高负载能力的自组装纳米结构，具有传递亲水性和疏水性药物的可能性，并且具有隐形性，从而导致免疫原性低。这些囊泡可用于递送酶药物，例如L-天冬酰胺酶（ASNase），这是用于急性淋巴细胞白血病治疗的一线药物。在此，基于聚（乙二醇）-b-聚（ε-己内酯）的三种半结晶共聚物的聚合物囊泡，即PEG ₄₅ PCL ₄₄，PEG ₁₁₄ PCL ₉₈和PEG ₁₁₄ PCL ₁₁₄研究了ASNase的封装，以及作为模型蛋白的牛血清白蛋白（BSA）的封装。共聚物的临界聚集浓度（CAC）通过荧光光谱法测定，其值在0.6至1.26mg / L之间变化。使用膜水合作用时，将离心作为膜后技术，可获得200–400 nm的聚合物囊泡和窄尺寸分布（多分散指数值为0.2–0.3）。在离心悬浮液之后，接着在上清液中测量蛋白质，或者在尺寸排阻色谱法（SEC）纯化之后确定包封效率（EE％），其中在定量中对与游离蛋白质相对应的洗脱级分进行定量。与SEC后的测量值（EE％≈1-5％）相比，离心后获得的包封效率值更高（EE％≈20％），这表明当离心分离时，蛋白质可能会部分包埋在聚合物网络中方法。但是，该系统可以提供游离ASNase的初始作用，然后基于封装的酶产生长期作用，从而导致剂量减少。