A novel combined strategy for the physical PEGylation of polypeptides
A novel combined strategy for the physical PEGylation of polypeptides
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DOI：
10.1016/j.jconrel.2016.02.009
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发表时间：
2016-03-28
影响因子：
10.8
通讯作者：
Salmaso, Stefano
中科院分区：
文献类型：
--
作者：
Ambrosio, Elena;Barattin, Michela;Salmaso, Stefano
聚乙二醇(PEG)可以共价连接到多肽药物上，以克服它们的快速清除，但这样做可能会失去它们的效力。在这里，使用非共价方法将含有末端胆碱部分的聚乙二醇酯(mpeg(5 KDa)-胆烷)偶联到28个氨基酸的多肽-血管活性肠肽(VIP)上。多肽的棕榈酰化对于通过两个亲和常数类似于3.10(4)M-1的mpeg(5 KDa)-胆烷分子的单一结合部位促进物理相互作用是必不可少的；这些量热数据证实了Scatchard对溶出度数据的分析。多肽/聚合物复合体(直径小于10-12 nm)在pH 7.4时提供了高达5000倍的多肽溶解度(4微克/毫升)，并显著提高了多肽溶液在25℃下30天的稳定性。甘露醇使该复合体能够被冷冻干燥以产生冻干制剂，该制剂被有效地重组，尽管溶解度下降了10%左右。当pH为7.4时，仅在pH 5-6.5时，多肽的主要α-螺旋构象消失，而当用2摩尔的mPEG5 kDa胆烷时，该构象完全恢复。在冷冻干燥和重组后，观察到类似于10%的α-螺旋构象的损失，这可能反映了相当于溶解度的下降。皮下注射多肽(0.1 mg/kg)和2摩尔当量聚合物后的药代动力学研究表明，mpeg(5 KDa)-胆烷显著增加了体内循环中的多肽浓度。这是首次证明酰化多肽的非共价聚乙二醇化，这是一种重要的生物类别，它改善了体外和体内的性质，从而可能被证明是共价聚乙二醇化策略的替代方案。(C)2016爱思唯尔B.V.保留所有权利。
Poly(ethylene glycol) (PEG) may be covalently conjugated to peptide drugs to overcome their rapid clearance but in doing so their potency can be lost. Here, a non-covalent approach was used to conjugate PEG bearing a terminal cholanic moiety (mPEG(5kDa)-cholane) to a 28 amino acid peptide, vasoactive intestinal peptide (VIP). Palmitoylation of the peptide was essential to facilitate physical interaction via a single binding site involving two mPEG(5kDa)-cholane molecules with an affinity constant of similar to 3.10(4) M-1; these calorimetry data corroborating Scatchard analysis of dissolution data. The peptide/polymer complex (below 10-12 nm diameter) provided for up to 5000-fold greater solubility of the peptide at pH 7.4 (4 mu g/mL) and markedly increased peptide solution stability at 25 degrees C over 30 days. Mannitol enabled the complex to be lyophilized to yield a freeze-dried formulation which was efficiently reconstituted albeit with an similar to 10% decrease in solubility. The predominantly a-helical conformation of the peptide alone at pH 5-6.5 was lost at pH 7.4 but fully recovered with 2 molar equivalents of mPEG5kDa-cholane. After lyophilization and reconstitution an similar to 10% loss of a-helical conformation was observed, which may reflect the equivalent decrease in solubility. Pharmacokinetic studies following subcutaneous administration of the peptide (0.1 mg/Kg) alone and with 2 molar equivalents of polymer showed that mPEG(5kDa)-cholane dramatically increased peptide concentration in the systemic circulation. This is the first demonstration of non-covalent PEGylation of acylated peptides, an important biologic class, which improves in vitro and in vivo properties, and thereby may prove an alternative to covalent PEGylation strategies. (C) 2016 Elsevier B.V. All rights reserved.