Phosphorothioate Oligodeoxyribonucleotides Antisense to PAI-1 mRNA Increase Fibrinolysis and Modify Experimental Thrombosis in Rats

Summary The effect of systemic inhibition of PAI-1 expression in rats by PS-16R, a phosphorothioate analogue of hexadecadeoxyribonucleotide complementary to a signal peptide coding sequence of rat PAI-1 mRNA, on PAI-1 activity in blood plasma and thrombus formation was studied in rat models for experimental thrombosis. In previous in vitro studies, oligonucleotides of PS-16R family have been shown to inhibit efficiently PAI-1 synthesis in endothelial cells by antisense mechanism. When PS-16R was administered intravenously as a single bolus injection (1 to 5 mg per rat), it produced a significant reduction in PAI-1 activity of blood plasma. This effect was both time- and concentration-dependent. Under the same conditions, three groups of rats were treated with control oligodeoxynucleotides such as PS-16R with double mismatches, with scrambled sequence, and an oligodeoxynucleotide with sense sequence (complementary to PS-16R), respectively. Based on these preliminary experiments, a low dose of 1.5 mg per rat was selected to produce approximately 20-30% reduction of PAI-1 activity in blood plasma and the effect of such a decrease in PAI-1 expression was tested on thrombus formation in two rat models for experimentally induced thrombosis. Such a limited decrease in PAI-1 activity produced a significant antithrombotic effect in the arterial thrombosis model. There was a profound delay in the occlusion time in rats treated with PS-16R when compared to control animals (80 ± 3 and 55 ± 3 h, respec tively), although blood plasma activity of PAI-1 in the same groups of rats differed only by 20%. There was also a tendency to reduce both an incidence of venous thrombosis (58.33 and 68.11%, respectively) and thrombus weight (2.1 ± 0.4 and 2.9 ± 0.9 mg, respectively) in the animals treated with PS-16R. However, this effect was not significant. Thus, low dose of PS-16R through inhibition of PAI-1 synthesis in targeted cells in rats reduced PAI-1 activity in blood plasma and protected against arterial thrombus formation in the rat.

[1]  W. Stec,et al.  Stability of stereoregular oligo(nucleoside phosphorothioate)s in human plasma: diastereoselectivity of plasma 3'-exonuclease. , 1997, Antisense & nucleic acid drug development.

[2]  J T Arnold,et al.  A single injection of pegylated murine megakaryocyte growth and development factor (MGDF) into mice is sufficient to produce a profound stimulation of megakaryocyte frequency, size, and ploidization. , 1997, Blood.

[3]  L. Iacoviello,et al.  Antithrombotic Activity of Dermatan Sulphates, Heparins and their Combination in an Animal Model of Arterial Thrombosis , 1996, Thrombosis and Haemostasis.

[4]  W. Stec,et al.  Internalization of oligodeoxynucleotide antisense to type-1 plasminogen activator inhibitor mRNA in endothelial cells: a three-dimensional reconstruction by confocal microscopy. , 1996, Biology of the cell.

[5]  H. Pannekoek,et al.  Structure of plasminogen activator inhibitor 1 (PAI-1) and its function in fibrinolysis: an update , 1995 .

[6]  D. Lawrence,et al.  Peptide-mediated inactivation of recombinant and platelet plasminogen activator inhibitor-1 in vitro. , 1995, The Journal of clinical investigation.

[7]  J. T. ten Cate,et al.  Thrombolysis and reocclusion in experimental jugular vein and coronary artery thrombosis. Effects of a plasminogen activator inhibitor type 1-neutralizing monoclonal antibody. , 1995, Circulation.

[8]  W. Stec,et al.  Inhibition by modified oligodeoxynucleotides of the expression of type-1 plasminogen activator inhibitor in human endothelial cells. , 1995, European journal of biochemistry.

[9]  B. Sobel,et al.  Inhibition of type-1 plasminogen activator inhibitor production by antisense oligonucleotides in human vascular endothelial and smooth muscle cells. , 1994, The Journal of biological chemistry.

[10]  W. Stec,et al.  Bis(O,O-diisopropoxy phosphinothioyl) disulfide - a highly efficient sulfurizing reagent for cost-effective synthesis of oligo(nucleoside phosphorothioate)s , 1993 .

[11]  T. Sakamoto,et al.  Plasma plasminogen activator inhibitor activity and tissue plasminogen activator levels in patients with unstable angina and those with coronary spastic angina. , 1992, American heart journal.

[12]  D. Loskutoff,et al.  Increased type 1 plasminogen activator inhibitor gene expression in atherosclerotic human arteries. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[13]  D. Vaughan,et al.  Reactivated Recombinant Plasminogen Activator Inhibitor-1 (rPAI-1) Effectively Prevents Thrombolysis In Vivo , 1992, Thrombosis and Haemostasis.

[14]  A. V. van Zonneveld,et al.  Inhibition of Plasminogen Activator Inhibitor‐1 Activity Results in Promotion of Endogenous Thrombolysis and Inhibition of Thrombus Extension in Models of Experimental Thrombosis , 1992, Circulation.

[15]  B. Alving,et al.  Plasminogen Activator Inhibitor Type 1: Biochemistry and Evidence for Modulation of Fibrinolysis In Vivo , 1992, Seminars in thrombosis and hemostasis.

[16]  M. L. Lewis,et al.  A miniaturized fibrinolytic assay for plasminogen activators. , 1991, Thrombosis research.

[17]  D. Belin,et al.  The plasminogen activator/plasmin system. , 1991, The Journal of clinical investigation.

[18]  M. Prins,et al.  A critical review of the relationship between impaired fibrinolysis and myocardial infarction. , 1991, American heart journal.

[19]  B. Eriksson,et al.  Impaired fibrinolysis and postoperative thromboembolism in orthopedic patients. , 1991, Thrombosis research.

[20]  Y. Yamori,et al.  Indirect systolic and mean blood pressure determination by a new tail cuff method in spontaneously hypertensive rats , 1991, Laboratory animals.

[21]  K. Marotti,et al.  Development of venous occlusions in mice transgenic for the plasminogen activator inhibitor-1 gene , 1990, Nature.

[22]  J. Schellekens,et al.  Plasminogen Activator Inhibitor 1: A New Prognostic Marker in Septic Shock , 1989, Thrombosis and Haemostasis.

[23]  B. Bennett,et al.  Plasminogen activator inhibitor (PAI‐1) in plasma and platelets , 1988, British journal of haematology.

[24]  M. Alessi,et al.  Measurement of plasminogen activator inhibitor 1 in biologic fluids with a murine monoclonal antibody-based enzyme-linked immunosorbent assay. , 1988, Blood.

[25]  M. Blombäck,et al.  PLASMINOGEN ACTIVATOR INHIBITOR IN PLASMA: RISK FACTOR FOR RECURRENT MYOCARDIAL INFARCTION , 1987, The Lancet.

[26]  G. de Gaetano,et al.  Aspirin and the Prevention of Experimental Arterial Thrombosis: Difficulty in Establishing Unequivocal Effectiveness , 1985, Thrombosis and Haemostasis.

[27]  D. Loskutoff,et al.  Endothelial cells produce a latent inhibitor of plasminogen activators that can be activated by denaturants. , 1985, The Journal of biological chemistry.

[28]  F. Werf,et al.  Plasminogen activator inhibitor in the blood of patients with coronary artery disease. , 1985, British medical journal.

[29]  E. Kruithof,et al.  Demonstration of a fast-acting inhibitor of plasminogen activators in human plasma. , 1984, Blood.

[30]  E. Dejana,et al.  Bleeding Time in Rats: A Comparison of Different Experimental Conditions , 1982, Thrombosis and Haemostasis.

[31]  G. de Gaetano,et al.  Failure of aspirin at different doses to modify experimental thrombosis in rats. , 1980, Thrombosis research.

[32]  G. de Gaetano,et al.  Decreased plasminogen activator but normal prostacyclin activity in rat veins during development of experimental thrombosis. , 1980, Thrombosis research.

[33]  L. Poller.  Pathogenesis of Thrombosis , 1960, Nature.