Delivery of Anti-Platelet-Endothelial Cell Adhesion Molecule Single-Chain Variable Fragment-Urokinase Fusion Protein to the Cerebral Vasculature Lyses Arterial Clots and Attenuates Postischemic Brain Edema

Danielyan, Kristina; Ding, Bi-Sen; Gottstein, Claudia; Cines, Douglas B.; Muzykantov, Vladimir R.

doi:10.1124/jpet.107.120535

Cited by 43 publications

(57 citation statements)

References 38 publications

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“…34 However, PECAM-1 targeting is not lung specific; in fact, local arterial infusion of anti-PECAM conjugates and fusion proteins via the coronary or carotid artery permits enhanced drug delivery to their respective organs. 33,34 These observations extend the potential utility of the proposed approach to diverse vascular beds. Nevertheless, the pulmonary vasculature collects the approximately 30% of injected dose after systemic intra-arterial or intravenous injection, consistent with its contribution to the total endothelial surface of the body; the lungs receive 50% of the total cardiac output and the slow rate of perfusion to the high-capacity, low-resistance pulmonary vasculature further promotes binding of PECAM targeted compounds.…”

Section: Discussionsupporting

confidence: 57%

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Prophylactic thrombolysis by thrombin-activated latent prourokinase targeted to PECAM-1 in the pulmonary vasculature

Ding

Hong

Murciano

et al. 2008

Blood

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A recombinant prodrug, single-chain urokinase-type plasminogen activator (scuPA) fused to an anti-PECAM-1 antibody single-chain variable fragment (anti-PECAM scFv/scuPA) targets endothelium and augments thrombolysis in the pulmonary vasculature. 1 To avoid premature activation and inactivation and to limit systemic toxicity, we replaced the native plasmin activation site in scFv/lowmolecular-weight (lmw)-scuPA with a thrombin activation site, generating anti-PECAM scFv/uPA-T that (1) is latent and activated by thrombin instead of plasmin; (2) binds to PECAM-1; (3) does not consume plasma fibrinogen; (4) accumulates in mouse lungs after intravenous injection; and (5) resists PA inhibitor PAI-1 until activated by thrombin. In mouse models of pulmonary thrombosis caused by thromboplastin and ischemiareperfusion (I/R), scFv/uPA-T provided more potent thromboprophylaxis and greater lung protection than plasminsensitive scFv/uPA. Endothelium-targeted thromboprophylaxis triggered by a prothrombotic enzyme illustrates a novel approach to time-and site-specific regulation of proteolytic reactions that can be modulated for therapeutic benefit. IntroductionPlasminogen activators (PAs) used in the treatment of thrombosis convert the zymogen plasminogen into plasmin, which lyses fibrin and restores perfusion. [2][3][4] Unfortunately, the utility of PAs is limited by inadequate delivery (rapid clearance, inactivation, and ineffective penetration of occlusive clots) and side effects, including hemorrhage and extravascular toxicity. [5][6][7] Further, postevent thrombolytic therapy is marred by inevitable delays (time needed for diagnosis, transportation, injection, and lysis) causing ischemia-reperfusion (I/R) injury that perpetuates thrombosis and worsens outcome. 8,9 Attempts to improve the effectiveness of PAs by increasing clot affinity have further diminished permeation into occluding thrombi due to retention on the clot surface 5 and have yet to provide decisively better clinical outcomes. [10][11][12][13][14] Thrombi often recur due to underlying procoagulation states, vascular damage, disturbance in blood flow, inflammation, or/and patients' immobility. 15 Rethromboses often occur within hours to days after acute myocardial infarction, transient ischemic attack, ischemic stroke, and pulmonary embolism. The effectiveness of secondary prevention using existing anticoagulants and antiplatelet agents is limited and dosing is constrained by the risk of bleeding. 16 In theory, prophylactic delivery of a PA into nascent thrombi, which are more susceptible to dissolution than mature occlusive clots, would expedite thrombolysis and minimize ischemiareperfusion injury in patients at high risk of imminent thrombosis. Unfortunately, PAs are not suitable for thromboprophylaxis due to unfavorable pharmacokinetics and toxicity.Conceivably, the failure to exploit PAs for thromboprophylaxis could be overcome by optimization of drug and targeting strategies. Single-chain urokinase-type plasminogen activator (scuPA) seems suitable for t...

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Section: Discussionsupporting

confidence: 57%

“…Indeed, uPA targeting to PECAM-1 prevents clot accretion in a model of acute pulmonary embolism 1 and cerebrovascular thrombosis. 33 However, for use as thromboprophylaxis, endothelium-targeted PAs must resist premature inactivation by inhibitors and be activated preferentially at sites of incipient thrombosis.…”

Section: Discussionmentioning

confidence: 99%

Prophylactic thrombolysis by thrombin-activated latent prourokinase targeted to PECAM-1 in the pulmonary vasculature

Ding

Hong

Murciano

et al. 2008

Blood

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“…This was previously shown as an efficient approach to the targeted delivery of medication in the form of the fusion proteins. As an example, the urokinase-type plasminogen activator (uPA) fused with a single-chain antibody directed to PECAM-1 was shown to accumulate in the brain after the intravenous application in a mouse model of cerebrovascular thrombosis [74] . Compared to the unconjugated drug, the PECAM-targeting uPA efficiently lysed the clots blocking the cerebral arterial vasculature, thus providing a rapid and stable cerebral reperfusion and alleviating the post-thrombotic brain edema.…”

Section: Targets For Therapy: Coagulation Cascadementioning

confidence: 99%

Nanomedicine in diagnostics and therapy of cardiovascular diseases: beyond atherosclerotic plaque imaging

Cicha

Lyer

Garlichs

2013

Nanotechnology Reviews

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Atherosclerosis results from the accumulation of the modified lipoproteins within the artery walls, which triggers complex vascular inflammatory processes. Although the pharmacologic agents for the treatment of clinical manifestations of atherosclerosis are available, their systemic delivery has serious disadvantages, such as considerable side effects or low efficacy at tolerated doses. Moreover, the treatment of atherosclerosis using the interventional techniques bears further shortcomings: the implanted stents require a lengthy antiplatelet therapy and carry the risk of in-stent restenosis. In the surgical approach to atherosclerosis, apart from the overall risk of open heart surgery, the lack of adequate venous material for bypasses constitutes a common problem. The nanotechnology has the potential to overcome the disadvantages of the current therapy of atherosclerosis, e.g., by the formation of nanosized assemblies for the earlier detection of atherosclerotic lesions and for cell-specific delivery of therapeutics. Replacing the current systemic pharmacological approach by a locally targeted treatment of plaques can substantially minimize the adverse effects, by lowering the drug cytotoxicity and reducing the required dosage. Moreover, a new generation of nanotechnological approaches to the revascularization procedures is now emerging, e.g., vascular tissue engineering utilizing the magnetic nanoparticles or the design of stents with the reduced risk of thrombosis and restenosis. This review discusses the possible applications of the nanomedical approaches in the treatment of cardiovascular diseases.

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“…[40][41][42][43] Platelet endothelial cell adhesion molecule-1 fused with urokinase in a model of cerebrovascular thromboembolism mediated almost complete clot lysis without exacerbating the characteristic side effects such as intracerebral hemorrhage. 44 Targeting RBCs was shown to have effective prophylaxis against arterial and venous thrombosis for ≈24 hours. 42 Although targeting platelet endothelial cell adhesion molecule-1 or RBCs has shown successful thrombolysis, they do not serve as exclusive targets because of ubiquitous expression.…”

Section: Discussionmentioning

confidence: 99%

Towards Effective and Safe Thrombolysis and Thromboprophylaxis

Wang¹,

Palasubramaniam²,

Gkanatsas³

et al. 2014

Circulation Research

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Rationale: Fibrinolysis is a valuable alternative for the treatment of myocardial infarction when percutaneous coronary intervention is not available in a timely fashion. For acute ischemic stroke, fibrinolysis is the only treatment option with a very narrow therapeutic window. Clinically approved thrombolytics have significant drawbacks, including bleeding complications. Thus their use is highly restricted, leaving many patients untreated. Objective: We developed a novel targeted fibrinolytic drug that is directed against activated platelets. Methods and Results: We fused single-chain urokinase plasminogen activator (scuPA) to a small recombinant antibody (scFv SCE5 ), which targets the activated form of the platelet–integrin glycoprotein IIb/IIIa. Antibody binding and scuPA activity of this recombinant fusion protein were on par with the parent molecules. Prophylactic in vivo administration of scFv SCE5 –scuPA (75 U/g body weight) prevented carotid artery occlusion after ferric chloride injury in a plasminogen-dependent process compared with saline ( P <0.001), and blood flow recovery was similar to high-dose nontargeted urokinase (500 U/g body weight). Tail bleeding time was significantly prolonged with this high dose of nontargeted urokinase, but not with equally effective targeted scFv SCE5 –scuPA at 75 U/g body weight. Real-time in vivo molecular ultrasound imaging demonstrates significant therapeutic reduction of thrombus size after administration of 75 U/g body weight scFv SCE5 –scuPA as compared with the same dose of a mutated, nontargeting scFv–scuPA or vehicle. The ability of scFv SCE5 –scuPA to lyse thrombi was lost in plasminogen-deficient mice, but could be restored by intravenous injection of plasminogen. Conclusions: Targeting of scuPA to activated glycoprotein IIb/IIIa allows effective thrombolysis and the potential novel use as a fibrinolytic agent for thromboprophylaxis without bleeding complications.

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Delivery of Anti-Platelet-Endothelial Cell Adhesion Molecule Single-Chain Variable Fragment-Urokinase Fusion Protein to the Cerebral Vasculature Lyses Arterial Clots and Attenuates Postischemic Brain Edema

Cited by 43 publications

References 38 publications

Prophylactic thrombolysis by thrombin-activated latent prourokinase targeted to PECAM-1 in the pulmonary vasculature

Prophylactic thrombolysis by thrombin-activated latent prourokinase targeted to PECAM-1 in the pulmonary vasculature

Nanomedicine in diagnostics and therapy of cardiovascular diseases: beyond atherosclerotic plaque imaging

Towards Effective and Safe Thrombolysis and Thromboprophylaxis

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