Baudys M, Letourneur D, Liu F, Mix D, Jozefonvicz J, Kim S W
Department of Pharmaceutics and Pharmaceutical Chemistry/Center for Controlled Chemical Delivery, University of Utah, Salt Lake City 84112, USA.
Bioconjug Chem. 1998 Mar-Apr;9(2):176-83. doi: 10.1021/bc970180a.
The biochemical and pharmacological properties of bioactive peptides and proteins can be altered by conjugation with polymers. This report describes site-specific attachment of insulin to activated carboxyl groups of carboxymethyl dextran (CMD, MW=51000) through the GlyA1 insulin amino group. On average, three or four insulin molecules were grafted to a CMD linear chain. Coupled insulin molecules were properly folded, and the bioactivity of conjugated insulin in the blood glucose depression assay was 9.6 IU/mg, which was only 2.6 times less than that for native insulin. The cell growth study indicated that the CMD-insulin conjugate was as mitogenic as insulin on vascular smooth muscle cells, whereas the starting CMD polymer was not. The insulin receptor binding constant of the conjugate (3.6 x 10[9] M[-1]) compared well with that of native insulin (7.6 x 10[9] M[-1]), indicating that the CMD chain does not present any major constraints to binding. Plasma clearance of CMD-insulin obeyed a two-compartment pharmacokinetic (PK) model with a CMD-insulin conjugate plasma elimination half-life of 114.1 min, which was significantly longer than that of soluble Zn-insulin (12.4 min). In contrast, pharmacodynamic (PD) profiles (blood glucose lowering effects) after intravenous (iv) administration of the conjugate or insulin in rats were not different. Subcutaneous (sc) administration of the conjugate resulted in a significantly prolonged plasma profile with a noncompartmental PK parameter mean residence time (MRT) of 103.5 min which was significantly longer than that of soluble Zn-insulin (40.5 min). This was reflected in the protracted PD effect of sc administered conjugate with time needed to reach minimum glucose concentration Tnadir of 95.7 min, which was significantly longer than that of insulin (62 min). We conclude that the conjugation of insulin to CMD leads to a bioactive conjugate with a delayed sc PD profile showing prolonged response, resembling intermediate acting insulin preparations.
生物活性肽和蛋白质的生化及药理特性可通过与聚合物结合而改变。本报告描述了胰岛素通过甘氨酸A1胰岛素氨基与羧甲基葡聚糖（CMD，分子量 = 51000）的活化羧基进行位点特异性连接。平均而言，三到四个胰岛素分子被接枝到一条CMD线性链上。偶联的胰岛素分子正确折叠，且共轭胰岛素在血糖降低试验中的生物活性为9.6 IU/mg，仅比天然胰岛素低2.6倍。细胞生长研究表明，CMD - 胰岛素共轭物在血管平滑肌细胞上与胰岛素一样具有促有丝分裂作用，而起始的CMD聚合物则没有。共轭物的胰岛素受体结合常数（3.6×10⁹ M⁻¹）与天然胰岛素的结合常数（7.6×10⁹ M⁻¹）相当，表明CMD链对结合没有任何重大限制。CMD - 胰岛素的血浆清除符合二室药代动力学（PK）模型，CMD - 胰岛素共轭物的血浆消除半衰期为114.1分钟，明显长于可溶性锌胰岛素（12.4分钟）。相比之下，大鼠静脉注射（iv）共轭物或胰岛素后的药效学（PD）曲线（降血糖作用）并无差异。皮下（sc）注射共轭物导致血浆曲线显著延长，非房室PK参数平均驻留时间（MRT）为103.5分钟，明显长于可溶性锌胰岛素（40.5分钟）。这反映在皮下注射共轭物的持久PD效应上，达到最低葡萄糖浓度Tnadir所需的时间为95.7分钟，明显长于胰岛素（62分钟）。我们得出结论，胰岛素与CMD的结合产生了一种具有延迟皮下PD曲线的生物活性共轭物，显示出延长的反应，类似于中效胰岛素制剂。
