Mansfield Kathryn M, Maynard Heather D
Department of Chemistry and Biochemistry, University of California, Los Angeles, 607 Charles E. Young Drive East, Los Angeles, California 90095-1569.
California NanoSystems Institute, University of California, Los Angeles, 570 Westwood Plaza, Los Angeles, California 90095-1569.
ACS Macro Lett. 2018 Mar 20;7(3):324-329. doi: 10.1021/acsmacrolett.7b00974. Epub 2018 Feb 20.
Insulin is an important therapeutic protein for the treatment of diabetes, but it is unstable and aggregates upon exposure to environmental stressors encountered during storage and transport. To prevent degradation of the protein in this manner and retain as much bioactivity as possible, a well-defined insulin-trehalose glycopolymer conjugate was synthesized. To accomplish this, a strategy was employed to site-specifically modify insulin with a polymerization initiator at a particular conjugation site; this also facilitated purification and characterization. Lysine of the B chain was preferentially modified by conducting the reaction at high pH, taking advantage of its higher nucleophilicity than the N-terminal amines. Trehalose monomer was polymerized directly from this macroinitiator to form a well-defined conjugate. Bioactivity of the site-specific conjugate was shown to be higher compared to the non-specific conjugate and the same as the analogous site-specific polyethylene glycol (PEG) conjugate as confirmed by the insulin tolerance test (ITT) in mice. The conjugated trehalose glycopolymer also stabilized insulin to heat as measured by high-performance liquid chromatography (HPLC).
胰岛素是治疗糖尿病的一种重要治疗性蛋白质，但它不稳定，在储存和运输过程中遇到环境应激源时会发生聚集。为了防止蛋白质以这种方式降解并尽可能保留其生物活性，合成了一种结构明确的胰岛素-海藻糖糖聚合物共轭物。为此，采用了一种策略，在特定的共轭位点用聚合引发剂对胰岛素进行位点特异性修饰；这也便于纯化和表征。通过在高pH值下进行反应，利用B链赖氨酸比N端胺具有更高的亲核性，优先对其进行修饰。海藻糖单体直接从这种大分子引发剂聚合形成结构明确的共轭物。通过小鼠胰岛素耐量试验（ITT）证实，位点特异性共轭物的生物活性高于非特异性共轭物，且与类似的位点特异性聚乙二醇（PEG）共轭物相同。通过高效液相色谱（HPLC）测量，共轭海藻糖糖聚合物也使胰岛素对热稳定。
