• blood clotting is a fundamental homeostatic mechanism
  • the participation of different systems is necessary:
    • vascular wall (vasoconstriction)
    • platelets (adhesion → activation → aggregation) with the formation of a primary plug
    • plasma coagulation factors → thrombin formation → fibrin formation → definitive plug
    • plasma inhibitors
    • fibrinolytic system

Timing of successive steps of hemostasis:

  • primary hemostasis:
    • vasoconstriction (fractions of seconds)
    • platelet adhesion and activation (tenths to units of seconds)
    • platelet aggregation and primary plug formation (seconds to minutes)
  • coagulation:
    • activation of coagulation factors (seconds to minutes)
    • formation of a solid fibrin coagulum (minutes)
  • fibrinolysis:
    • activation of fibrinolysis (minutes)
    • dissolution of fibrin coagulum (hours to tens of hours)

Fibrinolysis

  • the fibrinolytic system has two functions in the process of hemostasis:
    • dissolves the fibrin plug (within hours)
    • limits coagulum formation
  • the fibrinolytic system comprises a group of activators and inhibitors that are linked by a series of positive and negative feedback loops and has four main components:
    • plasminogen
    • plasmin
    • plasminogen activators
    • plasminogen inhibitors
  • the main component is the inactive proenzyme plasminogen, which is a precursor of plasmin protease
  • plasmin hydrolytically cleaves fibrin to form degradation products
  • several plasminogen activators are required to activate plasminogen to plasmin:
    • tissue-type (tPA) is mainly involved in the dissolution of thrombi in circulation
    • urokinase-type (uPA) is involved in extravascular proteolysis
    • plasminogen activation can also be mediated by streptokinase (a product of bacteria) or other fibrinolytics, kallikrein, and factor XIIa
  • Inhibition of fibrinolysis occurs at the level of inhibition of activators (PAI 1,2) and the level of inhibition of plasminogen (α2-antiplasmin and non-specifically α2-macroglobulin)
  • fibrinolysis is fibrin specific
    • the high affinity of tPA for plasminogen in the presence of fibrin allows efficient activation of plasminogen on the fibrin barrier, whereas in plasma plasminogen activation by tPA is ineffective
    • at the same time, fibrin-bound plasmin is protected against rapid inhibition by α2-plasminogen. In contrast, free plasmin is rapidly inhibited.
    • the fibrinolytic system is triggered and conditioned by fibrin in the above ways
    • streptokinase and urokinase are non-specific fibrinolytics and activate both circulating and fibrin-bound plasminogen, leading to widespread systemic activation of the fibrinolytic system, leading to degradation of other plasma proteins, including fibrinogen, factor V or factor VII
Fibrinolysis

Components of the fibrinolytic system

molecular weight (Da) effect
plasminogen 88000 proenzyme
plasmin 88000 active enzyme
tPA 70000 tissue enzyme
uPA 54000 urokinase-type
α2-antiplasmin 70000 specific fast-acting plasma inhibitor
PAI-1 43000 endothelium-produced rapid inhibitor of both t-PA and u-PA
  • glycoprotein present in plasma at a concentration of 1.5-2 pmol/l
  • contains “lysine-binding surfaces” that specifically react with amino acids such as lysine and 6-aminohexanoic acid. They mediate the specific binding of plasminogen to fibrin and the interaction of plasmin with α2-antiplasmin and play a key role in the control of fibrinolysis
  • tissue plasminogen activator (tPA) – a serine protease (alteplase, tenecteplase)
  • urokinase-type activator (uPA)
  • plasmin as a proteolytic enzyme cleaves not only fibrin and fibrinogen but also factor V, factor VIII, and prothrombin
  • Inhibition of fibrinolysis may occur at the level of inhibition of plasmin or inhibition of plasminogen activators
  • α2 – antiplasmin is the main physiological inhibitor of plasmin in human plasma
    • a glycoprotein belonging to the serine protease inhibitor proteins (serpins)
    • forms a complex with plasmin without protease activity
  • the main inhibitor of tPA and uPA is the plasminogen activator inhibitor (PAI 1,2)
    • glycoprotein belonging to serine protease inhibitors (serpins)
    • is the primary inhibitor of tPA and uPA in human plasma
    • reacts with tPA and urokinase, but not with pro-urokinase
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Overview of plasminogen activators (fibrinolytics)

  • all fibrinolytic agents act as plasminogen activators

1st generation thrombolytics

  • urokinase
  • streptokinase
    • no specificity to fibrin
    • streptokinase activates plasminogen to plasmin indirectly – in the first stage, streptokinase forms a complex with plasminogen, then plasminogen-streptokinase is converted to plasmin-streptokinase complex – in stroke, it is contraindicated due to a higher incidence of intracranial bleeding

2nd generation thrombolytics

  • alteplase (ACTILYSE)
    • recombinant human tissue plasminogen activator (tPA)
    • short half-life (3-6 minutes), administration by infusion (with initial bolus) is necessary
    • the most widely used thrombolytic in stroke therapy to date → intravenous thrombolysis in acute stroke
    • the high affinity of tPA for plasminogen in the presence of fibrin allows effective activation on fibrin barrier, whereas in plasma, plasminogen activation by tPA is ineffective
      • theoretically, alteplase should therefore only induce thrombolysis of the fibrin clot and not a hypocoagulable state and hypofibrinogenemia – we know from practice that even here we have to take into account the increased risk of bleeding and a decrease in fibrinogen levels

3rd generation thrombolytics
selective binding to fibrin, no systemic fibrinolysis, faster onset of action than generation II

  • tenecteplase (METALYSE)
    • genetically engineered tPA
    • longer half-life (17 ± 7 min) allows bolus administration
    • higher affinity for fibrin, higher resistance to PAI-1
    • significant trials:
      • EXTEND-IA TNK  demonstrated superior outcomes of TNK compared to tPA in patients with LVO and subsequent EVT
      • NOR-TEST   – similar results to tPA
    • up to 4.5h can be given as an alternative to tPA (ESO guidelines 2021
  • reteplase (RAPILYSIN / RETAVASE)
    • recombinant plasminogen activator, which has a longer half-life than tPA (15-18 min), allowing bolus administration
    • after administration of reteplase, fibrinogen depletion is less pronounced than after streptokinase but more than after tPA
    • approved for thrombolysis in myocardial infarction

4th generation thrombolytics

  • desmoteplase
    • genetically engineered thrombolytic protein from the saliva of the bat Desmodus rotundus
    • about 180 times more fibrin-selective than tPA, so it does not significantly affect systemic coagulation
    • no effect on HEB, no neurotoxicity, and longer plasma half-life than tPA (4 h)
    • DEDAS, DIAS, DIAS2 studies
Comparison of alteplase,reteplase, and tenecteplase

Synthetic antifibrinolytics

  • antifibrinolytics block the binding of plasmin to fibrin
  • aminocaproic acid and tranexamic acid are the commonly used antifibrinolytic drugs in the EU
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