CLINICAL DEVELOPMENT IN THE EARLY TREATMENT OF ACUTE ISCHAEMIC STROKE

n ANCROD: A POWERFUL DEFIBRINOGENATING AGENT WITH A UNIQUE MODE OF ACTION

Ancrod is a defibrinogenating enzyme (234-amino acid glycosylated serine protease; MW=35kDa) with selective substrate specificity for fibrinogen.1 The effects of Ancrod are mediated by a rapid and effective removal of normal fibrinogen from the bloodstream, without altering other coagulation factors or platelet turnover.1 The primary mechanism of Ancrod is through its proteolytic effect on the fibrinogen molecule.

Mechanism of action
Ancrod cleaves fibrinopeptides (designated FpA, FpAP, FpAY) from the A-alpha chain of fibrinogen. The resulting fibrin monomers (known as Ancrod-fibrin) cannot be stably cross-linked by factor XIIIa and form unstable, soluble chains 1-2 microm long 1,2_. The selective action of Ancrod on the A-alpha chain of fibrinogen, in the absence of cross-linking by fibrin stabilising factor, explains the increased susceptibility of Ancrod-fibrin to lysis by plasmin and its rapid clearance from the circulation by the reticuloendothelial system (RES) (Figure 1).1,2

Figure 1. The defibrinogenating mechanism of action of Ancrod in comparison to thrombin. After Illig et al.3

Ancrod shares structural similarities with thrombin but unlike thrombin it does not cleave fibrinopeptide B or affect other coagulation factors, factor XIII or platelets.1,2 Therefore, unlike other anticoagulants, Ancrod is rarely associated with significant bleeding.4

Rapid reduction in fibrinogen levels
The administration of Ancrod is followed within minutes by a rapid fall in plasma fibrinogen. Fibrinogen levels remain low so long as Ancrod continues to be administered. Once Ancrod is withdrawn, plasma fibrinogen returns to haemostatic levels within 24 - 48 hours, and to pre-treatment levels within days.

The role of fibrinogen
Fibrinogen plays an important role in the pathogenesis of venous and arterial thromboembolism and is also one of the main determinants of blood viscosity.1

References
1. Wright JG, Geroulakos G. Semin Vasc Surg 1996; 9: 315-328.
2. Soutar RL, Ginsberg JS. Crit Rev Oncol Hematol 1993; 15: 23-33.
3. Illig KA, Ouriel K. Semin Vasc Surg 1996' 9: 315-328.
4. Pollak VE, Glas-Greenwalt P, Olinger CP et al. Am J Med Sci 1990; 299: 319-325.

n ANCROD: A POWERFUL DEFIBRINOGENATING AGENT

Related to its powerful defibrinogenating effect, Ancrod lowers blood viscosity and increases fibrinolytic activity, although it does not directly activate plasminogen.

Reduction of blood and plasma viscosity with increased nutrient blood flow.
Since fibrinogen is a high molecular weight plasma protein, it contributes significantly to blood viscosity. In vivo measurements taken after initiation of Ancrod therapy show significant reductions in blood viscosity which become more pronounced as fibrinogen is progressively removed from the circulation. This may be clinically significant in re-establishing blood flow through stenosed vascular segments and limiting the area of ischaemia or necrosis from arterial thrombosis.1

Ancrod also locally enhances fibrinolysis and clot-specific thrombolysis.
Initiation of treatment with Ancrod is associated with a striking increase in levels of fibrinogen degradation products. These Ancrod-induced fibrinogen degradation products are very similar to those classically generated by plasmin. Evidence suggests that their presence is due partly to defibrinogenation, and partly to activation of the fibrinolytic system.1

Figure 1. The effects of Ancrod on the fibrinolytic pathway (shaded boxes).After Illig et al.2

Ancrod-fibrin triggers the release of t-PA or urokinase from vascular endothelium and is associated with consistently lower levels of plasminogen activator inhibitor-1 (PAI-1) and alpha-2-antiplasmin.6 t-PA binds to plasminogen associated with fibrin in blood clots which results in local digestion of fibrin. Thus, Ancrod-induced t-PA release may enhance clot-specific or internal clot lysis of a primary thrombus.1 However, evidence of "macroscopic" fibrinolysis is lacking.2

Activation of fibrinolysis following Ancrod administration is thought to be rapid and may occur before levels of fibrinogen are measurably reduced. The limited cleavage of plasma fibrinogen to ancrod-fibrin during the first hour of ancrod administration may be sufficient to initiate the process and induce fibrinolysis.

The selective effect of Ancrod on the coagulation pathway
Administration of Ancrod is also associated with an increase in circulating levels of prostacyclin-stimulating factor (PSF). This stimulates the endothelium to produce increased quantities of prostacyclin which has the property, among others, of reducing the propensity of platelets to aggregate and discouraging the formation of secondary clots.

Unlike thrombin, Ancrod does not activate clotting factors other than fibrinogen. In particular, it does not activate factors V, VIII or XIII, and thus does not stimulate coagulation activity. This very limited activity of Ancrod is the reason that haemorrhagic complications are so rare despite its prolonged and highly effective anticoagulant effect, and contrasts with the clinical experience of standard anticoagulants (heparin) and thrombolytics (streptokinase and t-PA), which have been associated with severe haemostatic breakdown and bleeding complications. 1,3-5

References
1. Wright JG, Geroulakos G. Semin Vasc Surg 1996; 9: 315-328.

2. Illig KA, Ouriel K. Semin Vasc Surg 1996' 9: 315-328.

3. Davis JA, Sharp AA, Merrick MV et al. Controlled trial of ancrod and heparin in treatment of deep-vein thrombosis of lower limb. Lancet 1972; i: 113-115.

4. Califf RM, Topol EJ, George BS et al. Hemorrhagic complications associated with the use of intravenous tissue plasminogen activator in treatment of acute myocardial infarction. Am J Med 1988; 85: 353-359.

5. Gore JM, Sloan M, Price TR et al. Intracerebral hemorrhage, cerebral infarction, and subdural hematoma after acute myocardial infarction and thrombolytic therapy in the thrombolysis in myocardial infarction study. Circulation 1991; 83: 448-459.

n FIBRINOGEN REDUCTION WITH ANCROD AMELIORATES FOCAL CEREBRAL ISCHAEMIA IN ANIMAL MODELS

The safety and tolerability of Ancrod and its mode of action have been investigated in a series of preclinical studies in animal models. These experimental investigations on cerebrovascular disorders such as permanent middle cerebral artery occlusion (MCAO), intracerebral haemorrhage and thrombotic cortical infarction in animals have shown pronounced lesion reductions with Ancrod.1,2

In an MCA-occlusion animal model of stroke, Ancrod significantly and dose-dependently reduced the total volume of the thrombotic brain lesion from 142+28 mm3 in controls to 121+28 mm3 with 10 U/kg Ancrod and to 111+20 mm3 with 30U/kg Ancrod) (p<0.05).2

Following stasis-induced thrombosis, there was also a dose-dependent inhibition in thrombus development and a close linear relationship was apparent between plasma fibrinogen levels and thrombus score (r=0.854, p<0.001) or thrombus weight (r=0.767, p<0.001).2

The therapeutic window for efficient post-treatment with Ancrod therapy has been evaluated following focal cerebral ischaemia in animals.3 Brain oedema volumes determined by MRI were reduced in Ancrod-treated animals up to 6 hours after MCAO and there were significant reductions in brain oedema volumes even if Ancrod therapy was delayed for up to 3 hours (p<0.05) (Figure 1 and 2).3

In addition, a close linear relationship between plasma fibrinogen levels and brain oedema volume was observed in animals up to 3 hours after MCAO, showing that there is a correlation between the reduction in plasma fibrinogen levels by Ancrod and reduction in brain oedema volume.3

These animal models give experimental support for the beneficial effects of Ancrod in the treatment of acute ischaemic stroke by demonstrating that the dose-dependent reduction in plasma fibrinogen levels by Ancrod may protect the brain by improving haemorheology. Furthermore, the linear relationship between plasma fibrinogen levels and thrombus score or weight suggest an effect of Ancrod on inhibiting thrombus development.2

References
1. Elger B, Laux V, Horneberger W et al. Effects of Ancrod on intracerebral haemorrhage and haemostatic plug formation in mesenteric arteries of rats. Cerebrovasc Dis 1995; 5(4): 261.

2. Elger B, Laux V, Horneberger W et al. Cerebroprotection by Ancrod in two rat stroke models: diffusion and T2-weighted MRI in vivo studies. Eur J Neurol 1995; 2 (suppl 2): 58.

3. Elger B, Schwarz M, Seega J, Hornberger W. Fibrinogen reduction by delayed Ancrod therapy ameliorates focal cerebral ischaemia in the rat. Fibrinolysis 1996; 10 (suppl 1): 4.

ANCROD

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