Glycosylation improves α-chymotrypsin stability upon encapsulation in poly(lactic-co-glycolic)acid microspheres.
Glycosylation improves α-chymotrypsin stability upon encapsulation in poly(lactic-co-glycolic)acid microspheres.
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DOI：
10.1016/j.rinphs.2012.08.001
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发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Griebenow K
中科院分区：
文献类型：
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作者：
Flores-Fernández GM;Griebenow K
在包封和从聚合物释放时增强蛋白质稳定性是持续释放应用中的关键问题。此外，聚合物颗粒中的最佳药物分散对于实现具有低的不需要的初始“突释”释放的释放曲线是关键的。在本文中，我们通过将模型酶α-糜蛋白酶（α-CT）配制为纳米颗粒以改善药物分散，并通过用聚糖共价修饰它以在聚（乳酸-乙醇酸）（PLGA）微球中包封期间提供改善的稳定性来解决这两个问题。用活化乳糖（500 Da）对α-CT进行化学修饰，使乳糖与蛋白质的摩尔比达到4.5和7.1。将生物缀合物与甲基-β-环糊精共冻干，然后悬浮于乙酸乙酯中以提供纳米颗粒。纳米颗粒的形成没有显著影响蛋白质的稳定性;小于5%的蛋白质聚集，并且对于所有制剂，残留活性保持在90%以上。使用我们实验室开发的用于纳米颗粒的固体-油-水（s/o/w）方法，我们获得了61%的最大包封效率。糖基化完全防止了在未修饰的酶的包封期间的否则大量的蛋白质聚集和活性损失。此外，糖基化制剂的体外蛋白质释放得到改善。这些结果突出了化学糖基化在持续释放应用中改善药物蛋白质的稳定性的潜力。
Enhancing protein stability upon encapsulation and release from polymers is a key issue in sustained release applications. In addition, optimum drug dispersion in the polymer particles is critical for achieving release profiles with low unwanted initial “burst” release. Herein, we address both issues by formulating the model enzyme α-chymotrypsin (α-CT) as nanoparticles to improve drug dispersion and by covalently modifying it with glycans to afford improved stability during encapsulation in poly(lactic-co-glycolic) acid (PLGA) microspheres. α-CT was chemically modified with activated lactose (500 Da) to achieve molar ratios of 4.5 and 7.1 lactose-to-protein. The bioconjugates were co-lyophilized with methyl-β-cyclodextrin followed by suspension in ethyl acetate to afford nanoparticles. Nanoparticle formation did not significantly impact protein stability; less than 5% of the protein was aggregated and the residual activity remained above 90% for all formulations. Using a solid-in-oil-in-water (s/o/w) methodology developed in our laboratory for nanoparticles, we obtained a maximum encapsulation efficiency of 61%. Glycosylation completely prevented otherwise substantial protein aggregation and activity loss during encapsulation of the non-modified enzyme. Moreover, in vitro protein release was improved for glycosylated formulations. These results highlight the potential of chemical glycosylation to improve the stability of pharmaceutical proteins in sustained release applications.