Thrombolytic and pharmacokinetic properties of a conjugate of recombinant single-chain urokinase-type plasminogen activator with a monoclonal antibody specific for cross-linked fibrin in a baboon venous thrombosis model.
Thrombolytic and pharmacokinetic properties of a conjugate of recombinant single-chain urokinase-type plasminogen activator with a monoclonal antibody specific for cross-linked fibrin in a baboon venous thrombosis model.
复制标题
重组单链尿激酶型纤溶酶原激活剂与交联纤维蛋白特异性单克隆抗体在狒狒静脉血栓形成模型中的溶栓和药代动力学特性。
作者：
D. Collen;M. Dewerchin;H. Rapold;H. Lijnen;J. Stassen
重组单链尿激酶型纤溶酶原激活剂 (rscu-PA) 与针对交联人纤维蛋白 (MA-15C5) 片段 D-二聚体的鼠单克隆抗体 rscu-PA/MA-15C5 之间以及 rscu-PA 与对照单克隆抗体 (MA-1C8) rscu-PA/MA-1C8 之间的化学缀合物，通过交联产生N-琥珀酰亚胺基 3-(2-吡啶二硫代)丙酸酯 (SPDP)。在由 [125 I] 纤维蛋白标记的狒狒血浆凝块浸入自体柠檬酸血浆中组成的体外系统中，获得了剂量和时间依赖性裂解，游离和结合的 rscu-PA 的效力比率与人血浆中相似：2 小时内 50% 裂解需要 4.3 微克/ml rscu-PA、1.0 微克/ml 尿激酶型纤溶酶原激活剂(u-PA) 等效 rscu-PA/MA-15C5，或 15 微克/ml u-PA 等效 rscu-PA/MA-1C8。在狒狒中将 rscu-PA 和 rscu-PA/MA-15C5 的溶栓和药代动力学特性与股静脉中产生的 0.8-1.0 ml [ 125 I]纤维蛋白标记的自体血凝块进行比较。在 2 小时内连续静脉输注这些化合物导致剂量和时间依赖性裂解。 rscu-PA/MA-15C5 的溶栓效力比通过股静脉段离体同位素回收测量的 rscu-PA 的溶栓效力高 3.0 +/- 0.5 倍（用 0.3 +/- 0.02 mg u-PA 当量/kg 体重溶解 50%）（p 小于 0.001），并且高出 2.7 +/- 0.5 倍（用 0.3 +/- 0.02 mg u-PA 当量/kg 体重溶解 50%）。 0.35 +/- 0.02 mg/kg rscu-PA/MA-15C5)，通过外部放射性同位素计数（p 小于 0.001）。剂量为 0.5 mg/kg 的 rscu-PA/MA-1C8 的活性远低于 rscu-PA。输注结束后，u-PA相关抗原以双相方式从血浆中消失，rscu-PA的初始半衰期为2.7 +/- 0.5，rscu-PA/MA-15C5为24 +/- 1.2，rscu-PA/MA-1C8为21 +/- 0.5分钟，相应的血浆清除率为340 +/- 40、20 +/-分别为 3 和 24 +/- 2 毫升/分钟。总之，rscu-PA/MA-15C5 溶栓效力的增加是由于 rscu-PA 与抗体分子偶联而导致溶栓效力降低的结果，而由于特定独特型的纤维蛋白靶向而导致溶栓效力增强，从而抵消了溶栓效力的降低。
Chemical conjugates between recombinant single-chain urokinase-type plasminogen activator (rscu-PA) and a murine monoclonal antibody directed against fragment D-dimer of cross-linked human fibrin (MA-15C5), rscu-PA/MA-15C5, and between rscu-PA and a control monoclonal antibody (MA-1C8), rscu-PA/MA-1C8, were produced by cross-linking with N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP). In an in vitro system composed of a [125 I]fibrin-labeled baboon plasma clot immersed in autologous citrated plasma, dose- and time-dependent lysis was obtained with a ratio of the potencies of free and conjugated rscu-PA similar to that in human plasma: 50% lysis in 2 hours required 4.3 micrograms/ml rscu-PA, 1.0 microgram/ml urokinase-type plasminogen activator (u-PA) equivalent rscu-PA/MA-15C5, or 15 micrograms/ml u-PA equivalent rscu-PA/MA-1C8. The thrombolytic and pharmacokinetic properties of rscu-PA and of rscu-PA/MA-15C5 were compared in baboons with a 0.8-1.0 ml [125 I]fibrin-labeled autologous blood clot produced in a femoral vein. Continuous intravenous infusion of these compounds during a 2-hour period resulted in dose- and time-dependent lysis. The thrombolytic potency of rscu-PA/MA-15C5 was 3.0 +/- 0.5 times higher (50% lysis with 0.3 +/- 0.02 mg u-PA equivalent/kg body wt) than that of rscu-PA measured by ex vivo isotope recovery from the femoral vein segment (p less than 0.001) and was 2.7 +/- 0.5 times higher (50% lysis with 0.35 +/- 0.02 mg/kg rscu-PA/MA-15C5) by external radioisotope counting (p less than 0.001). A dose of 0.5 mg/kg of rscu-PA/MA-1C8 was much less active than rscu-PA. After the end of the infusion, u-PA-related antigen disappeared from plasma in a biphasic manner with an initial half-time of 2.7 +/- 0.5 for rscu-PA, 24 +/- 1.2 for rscu-PA/MA-15C5, and 21 +/- 0.5 minutes for rscu-PA/MA-1C8 with corresponding plasma clearances of 340 +/- 40, 20 +/- 3, and 24 +/- 2 ml/min, respectively. In conclusion, the increased thrombolytic potency of rscu-PA/MA-15C5 is the result of a reduction of the thrombolytic potency due to coupling of rscu-PA to the antibody molecule, which is counter-balanced by an enhancement of the thrombolytic potency due to fibrin targeting by the specific idiotype.