title: 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.
abstract: 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.
authors.affiliation: Center for Thrombosis and Vascular Research, University of Leuven, Belgium.
authors.forename: D
authors.initials: D
authors.lastname: Collen
authors.forename: M
authors.initials: M
authors.lastname: Dewerchin
authors.forename: H J
authors.initials: HJ
authors.lastname: Rapold
authors.forename: H R
authors.initials: HR
authors.lastname: Lijnen
authors.forename: J M
authors.initials: JM
authors.lastname: Stassen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000627:therapeutic use*; D003429:Cross Reactions; D003432:Cross-Linking Reagents; D004792:Enzyme Precursors; D005260:Female; D005338:Fibrin Fibrinogen Degradation Products / Q000276:immunology; D005343:Fibrinolytic Agents / Q000493:pharmacokinetics* / Q000627:therapeutic use*; D008297:Male; D010215:Papio; D010960:Plasminogen Activators / Q000493:pharmacokinetics* / Q000627:therapeutic use*; D011994:Recombinant Proteins / Q000627:therapeutic use; D013388:Succinimides; D013924:Thrombophlebitis / Q000188:drug therapy*; D014568:Urokinase-Type Plasminogen Activator / Q000493:pharmacokinetics* / Q000627:therapeutic use*
chemical_list: D000911:Antibodies, Monoclonal; D003432:Cross-Linking Reagents; D004792:Enzyme Precursors; D005338:Fibrin Fibrinogen Degradation Products; D005343:Fibrinolytic Agents; D011994:Recombinant Proteins; D013388:Succinimides; C036309:fibrin fragment D; C018151:N-succinimidyl 3-(2-pyridyldithio)propionate; D010960:Plasminogen Activators; D014568:Urokinase-Type Plasminogen Activator