Potentiation by Lys-Plasminogen of Clot Lysis by Single or Two Chain Urokinase-Type Plasminogen Activator or Tissue-Type Plasminogen Activator

Summary

Study has been made of the influence of addition of human NH₂ terminal glutamic acid plasminogen (Glu-Plg) or human NH₂ terminal lysine plasminogen (Lys-Plg) to normal citrated plasma upon the rate of lysis of fully crosslinked plasma clots in the presence of single or two chain urokinase type plasminogen activator (scu-PA/tcu-PA) or tissue plasminogen activator (t-PA). The Specificity of any thrombolytic property was evaluated by measurement of plasma fibrinogen levels. Lys-plgadded to a concentration of 20% of normal plasma plasminogen caused 5 to 6 fold increase in the extent of lysis observed at 6 hours by 100 units/ml of scu-PA and with a small increase in fibrinogenolysis. Glu-Plg added at 20% of normal level had no influence on thrombolysis but at 50% of normal caused increased thrombolysis with rapid depletion of plasma fibrinogen. An apparently synergistic effect of addition of tcu-PA on scu-PA activity was increased by addition of plasminogen (e.g. addition of 20% Lys-Plg increased the lysis rate 4 to 5 fold over the first hour equivalent to an increase of potency of approximately three to four fold). Addition of plasminogen up to double the normal plasma concentration was observed to have no influence on clot lysis in the presence of t-PA. Plasminogen potentiated the rate of lysis by scu-Pr/t-PA synergic mixtures with an approximately 1.5 to 1.9 fold increase in potency. Potentiation occurred without increase in the depletion of plasma fibrinogen. It is concluded that Lys-Plg may be a suitable agent to improve the thrombolytic efficacy of scu-PA, scu-PA/tcu-PA or scu-PA/t-PA therapeutic regimen.

Keywords

Single chain urokinase-type plasminogen activator (scu-PA) - Urokinase - Tissue-type plasminogen activator (tPA) - Gluplasminogen - Lys-plasminogen - Clot lysis

• Reference

• 1 Gurewich V, Pannell R, Louie S, Kelley P, Suddith PL, Greenlee R. Effective and fibrin specific clot lysis by a zymogen precursor form of urokinase (UK). A study in vitro and in two animal species J Clin Invest 1984; 73: 1731-1739
• 2 Collen D, Stassen JM, Blaber M, Winkler M, Verstraete M. Biological and thrombolytic properties of proenzyme and active forms of human urokinase, Ill-Thrombolytic properties of natural and recombinant urokinase in rabbits with experimental jugular vein thrombosis. Thromb Haemostas 1984; 52: 27-30
• 3 Collen D, Stump D, Van de Werf F, Jang I K, Nobuhara M, Lijnen HR. Coronary thrombolysis in dogs with intravenously administered human pro-urokinase. Circulation 1984; 72: 384-388
• 4 Kominger C, Collen D. Studies on the specific fibrinolytic effect of human extrinsic (tissue-type) plasminogen activator in human blood and in various species in vitro. Thromb Haemostas 1981; 46: 561-566
• 5 Collen D, Stassen JM, Verstraete M. Thrombolysis with human extrinsic (tissue-type) plasminogen activator in rabbits with experimental jugular vein thrombosis. Effect of molecular form and dose of activator, age of the thrombus and route of administration J Clin Invest 1983; 71: 368-376
• 6 Gurewich V, Pannell R. A comparative study of the efficacy and specificity of tissue plasminogen activator and pro-urokinase: demonstration of synergism and of different thresholds of non-selectivity. Thromb Res 1986; 44: 217-228
• 7 Collen D, De Cock F, Demarsin E, Lijnen HR, Stump DC. Absence of synergism between tissue plasminogen activator (t-PA), single chain urokinase type plasminogen activator (scu-PA) and urokinase on clot lysis in a plasma milieu in vitro. Thromb Haemostas 1986; 56: 35-39
• 8 Collen D, Stassen JM, Stump DC, Verstraete M. Synergism of thrombolytic agents in vivo. Circulation 1986; 774: 838-842
• 9 Collen D, Stassen JM, De Cock F. Synergistic effect on thrombolysis of sequential infusion of tissue-type plasminogen activator (t-PA) single-chain urokinase-type plasminogen activator (scu-PA) and urokinase in the rabbit jugular vein thrombosis model. Thromb Haemostas 1987; 58: 943-946
• 10 Kakkar VV, Sagar S, Lewis M. Treatment of deep vein thrombosis with intermittent streptokinase and plasminogen infusion. Lancet 1975; 2: 6174-6176
• 11 Kakkar VV, Sagar S, Scully MF, Lane DA. Intermittent plasminogen-streptokinase treatment of deep vein thrombosis. Aktuel Probl Angiol 1978; 37: 142-154
• 12 Scully MF, Strachan CJ L, Kakkar VV. Binding of iodinated proteins to forming and preformed thrombi. Biochem Soc Trans 1973; 1: 1204-1206
• 13 Strachan CJ L, Scully MF, Kakkar VV. The behaviour of isotopic labelled blood proteins in thrombosis. Thromb Res 1974; 4: 303-318
• 14 Scully MF, Lane DA, Sagar S, Thomas DP, Kakkar VV. Intermittent plasminogen-streptokinase treatment of deep vein thrombosis. Thromb Haemostas 1977; 37: 162-169
• 15 Welzel D, Wolf H. Clinical research on single chain urokinase-type plasminogen activator (scu-PA) in Germany. Results in patients with acute myocardial infarction (AMI) Thromb Haemostas 1987; 58: 47 Abstr
• 16 Collen D. On the regulation and control of fibrinolysis. Thromb Haemostas 1980; 43: 77-89
• 17 Gurewich V, Pannell R. Synergism of tissue type plasminogen activator (t-PA) and single chain urokinase type plasminogen activator (scu-PA) on clot lysis in vitro and the mechanism of its effect. Thromb Haemostas 1987; 57: 372
• 18 Collen D. Rebuttal. Synergism of tissue-type plasminogen activator (t- PA) and single chain urokinase type plasminogen activator (scu-PA) on clot lysis in vitro and a mechanism for this effect Thromb Haemostas 1987; 57: 373
• 19 Claeys H, Molla A, Verstraete M. Conversion of NH₂-terminal glutamic acid to NH₂-terminal lysine human plasminogen by plasmin. Thromb Res 1973; 3: 515-523
• 20 Claeys H, Vermylen J. Physico-chemical and pro-enzyme properties of NH₂-terminal lysine human plasminogen. Biochim Biophys Acta 1973; 342: 351-358
• 21 Scully MF, Kakkar VV. Measurement of levels of degraded plasminogen during thrombolytic therapy. In: Chromogenic Peptide Substrate. Scully MF, Kakkar VV. eds Churchill Livingstone; Edinburgh: 1979. pp 141-153
• 22 Takada A, Takada Y. Effects of w-amino acids and clot formation on the activation by urokinase of various plasminogen preparations. Thromb Res 1980; 18: 167-176
• 23 Collen D, Zamarron E, Lijnen HR, Hoylaerts M. Activation of plasminogen by pro-urokinase. J Biol Chem 1986; 261: 1259-1266
• 24 Ellis V, Scully MF, Kakkar VV. Plasminogen activation by single chain urokinase in functional isolation. A kinetic study J Biol Chem 1987; 262: 14998-15003
• 25 Thorsen S. Differences in binding to fibrin of native plasminogen and plasminogen modified by proteolytic degradation. Influence of aminocarboxylic acids Biochim Biophys Acta 1975; 393: 55-62
• 26 Scully MF, Kakkar VV. Levels of degraded plasminogen during intermittent thrombolytic therapy. In: Progress in Chemical Fibrinolysis and Thrombolysis, Vol IV. Davidson JF, Cepalak V, Samama MM, Desnoyers PC. eds Churchill Livingstone; Edinburgh: 1979. pp 469-473
• 27 Holvoet P, Lijnen HR, Collen D. A monoclonal antibody specific for Lys-plasminogen. Application to the study of the activation pathways of plasminogen in vivo J Biol Chem 1985; 260: 12106-12111
• 28 Gurewich V, Black J, Pannell R. A mechanism for the potentiating effect of urokinase or tissue plasminogen activator on clot lysis by prourokinase. Thromb Haemostas 1987; 58: 439 Abstr
• 29 Pannell R, Black J, Gurewich V. Complementary models of action of tissue-type plasminogen activator and pro-urokinase by which their synergistic effect on clot lysis may be explained. J Clin Invest 1988; 81: 853-859
• 30 Urano J, Sator de Serrano V, Gaffney PJ, Castellino FJ. The activation of human [Glu] plasminogen by human single-chain urokinase. Arch Biochem Biophys 1988; 264: 222-230
• 31 Scully MF, Ellis V, Watahiki Y, Kakkar VV. Activation of prourokinase by plasmin: Non-Michaelian kinetics indicates a mechanism of negative cooperativity. Arch Biochem Biophys. 1989 in press
• 32 Ellis V, Scully MF, Kakkar VV. Plasminogen activation initiated by single-chain urokinase-type plasminogen activator. Potentiation by U937 monocytes. J Biol Chem 1989 in press
• 33 Collen D. Synergism, mutants and hybrids of tissue-type plasminogen activator (t-PA) and single chain urokinase type plasminogen activator (scu-PA): Potential for thrombolytic therapy. Thromb Haemostas 1987; 58: 254 Abstr
• 34 Collen D, Ong EB, Johnson AJ. Human plasminogen: in vitro and in vivo evidence for the biological integrity of NH₂-terminal glutamic acid plasminogen. Thromb Res 1975; 7: 515-529