Reprogrammed streptokinases develop fibrin-targeting and dissolve blood clots with more potency than tissue plasminogen activator

Abstract

Summary Background: Given the worldwide epidemic of cardiovascular diseases, a more effective means of dissolving thrombi that cause heart attacks, could markedly reduce death, disability and healthcare costs. Plasminogen activators (PAs) such as streptokinase (SK) and tissue plasminogen activator (TPA) are currently used to dissolve fibrin thrombi. SK is cheaper and more widely available, but it appears less effective because it lacks TPA’s fibrin-targeted properties that focus plasminogen activation on the fibrin surface. Objective: We examined whether re-programming SK’s mechanism of action would create PAs with greater fibrin-targeting and potency than TPA. Methods and Results: When fibrinogen consumption was measured in human plasma, reprogrammed molecules SKΔ1 and SKΔ59 were 5-fold and > 119-fold more fibrin-dependent than SK (P < 0.0001), and 2-fold and > 50-fold more fibrin-dependent than TPA (P < 0.001). The marked fibrin-targeting of SKΔ59 was due to the fact that: (i) it did not generate plasmin in plasma, (ii) it was rapidly inhibited by α2-antiplasmin, and (iii) it only processed fibrin-bound plasminogen. To assess the fibrin-targeting and therapeutic potential of these PAs in vivo, a novel ‘humanized’ fibrinolysis model was created by reconstituting plasminogen-deficient mice with human plasminogen. When compared with TPA, SKΔ1 and SKΔ59 were 4-fold (P < 0.0001) and 2-fold (P < 0.003) more potent at dissolving blood clots in vivo, respectively, on a mass-dose basis and 2–3 logs more potent than TPA (P < 0.0001) when doses were calibrated by standard activity assays. Conclusion: These experiments suggest that reprogramming SK’s mechanism of action markedly enhances fibrin-targeting and creates, in comparison with TPA, activators with greater fibrinolytic potency.

Multifunctional Nanoagent for Thrombus-Targeted Fibrinolytic Therapy

“…Native human fibrinogen (50 mkg; American Diagnostica Inc.) was radioiodinated with [ 125 I]NaI (NEN-Dupont, Boston, MA) in pre-coated Pierce iodination tubes (Thermo Scientific Inc.) by the Iodogen method [22, 23]. 125 I-fibrinogen was separated by gel filtration chromatography on a PD-10 column (Pharmacia Biotech.…”

The Role of Streptokinase as a Virulence Determinant of Streptococcus pyogenes – Potential for Therapeutic Targeting

“…Fibrin-independent plasminogen activation by the streptokinase-plasminogen activator complex is thought to be due to the insensitivity of the activator complex to α 2 -antiplasmin inhibition and due to the ability of the activator complex to activate circulating Gluplasminogen in the absence of fibrin [110]. Critical for these characteristics is the α-domain of streptokinase.…”

“…However, the SKΔ59-plasmin complex will readily activate unfolded forms of plasminogen such as Glu-plasminogen in the presence of fibrin [111]. When compared to t-PA, this mutated form of streptokinase displays less fibrinogen degradation in human plasma and has a greater capacity to degrade blood clots in vivo using a murine model of human thrombosis [110].…”

Development and characterization of highly selective target-sensitive liposomes for the delivery of streptokinase: in vitro/in vivo studies

“…a 2 AP) appear to react normally with human plasminogen (Lijnen et al, 1994;Fay et al, 2007). Therefore, in order to simulate human clot model, thrombolysis study was performed in the rats supplemented with physiologic levels of human plasminogen (Sazonova et al, 2009).…”

“…In the animal containing thrombus, various formulations were administered by femoral veins at a dose equivalent to 7500 IU. Simultaneously, human plasminogen was administered to maintain the physiologic level of plasminogen (Sazonova et al, 2009). After 10 min and 60 min of treatment, vessels were dissected and thrombus was blotted using filter paper.…”