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US20110129526A1 - Compositions and methods for treating vascular disease - Google Patents

Compositions and methods for treating vascular disease Download PDF

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Publication number
US20110129526A1
US20110129526A1 US12/918,726 US91872609A US2011129526A1 US 20110129526 A1 US20110129526 A1 US 20110129526A1 US 91872609 A US91872609 A US 91872609A US 2011129526 A1 US2011129526 A1 US 2011129526A1
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tpa
annexin
medium
stroke
dose
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Xiaoying Wang
Eng Lo
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General Hospital Corp
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General Hospital Corp
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Assigned to THE GENERAL HOSPITAL CORPORATION reassignment THE GENERAL HOSPITAL CORPORATION ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: LO, ENG, WANG, XIAOYING
Publication of US20110129526A1 publication Critical patent/US20110129526A1/en
Assigned to NATIONAL INSTITUTES OF HEALTH (NIH), U.S. DEPT. OF HEALTH AND HUMAN SERVICES (DHHS), U.S. GOVERNMENT reassignment NATIONAL INSTITUTES OF HEALTH (NIH), U.S. DEPT. OF HEALTH AND HUMAN SERVICES (DHHS), U.S. GOVERNMENT CONFIRMATORY LICENSE (SEE DOCUMENT FOR DETAILS). Assignors: THE GENERAL HOSPITAL CORPORATION
Assigned to NATIONAL INSTITUTES OF HEALTH (NIH), U.S. DEPT. OF HEALTH AND HUMAN SERVICES (DHHS), U.S. GOVERNMENT reassignment NATIONAL INSTITUTES OF HEALTH (NIH), U.S. DEPT. OF HEALTH AND HUMAN SERVICES (DHHS), U.S. GOVERNMENT CONFIRMATORY LICENSE (SEE DOCUMENT FOR DETAILS). Assignors: THE GENERAL HOSPITAL CORPORATION
Assigned to NATIONAL INSTITUTES OF HEALTH - DIRECTOR DEITR reassignment NATIONAL INSTITUTES OF HEALTH - DIRECTOR DEITR CONFIRMATORY LICENSE (SEE DOCUMENT FOR DETAILS). Assignors: PARTNERS HEALTHCARE INNOVATION
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/46—Hydrolases (3)
    • A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
    • A61K38/49—Urokinase; Tissue plasminogen activator
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention is related to the field of vascular disorders.
  • the present invention is related to the treatment and management of diseases including, but not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • diseases including, but not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • a patient having suffered a vascular disorder may be administered a composition comprising tPA and Annexin A2.
  • the tPA dose may be reduced such that the risk of hemorrhagic side effects are minimal.
  • Thrombolytic therapy with tissue plasminogen activator is the only FDA-approved medicine for achieving both vascular reperfusion and clinical benefit, but only 2-5% of stroke patients receive tPA in the US. In part, this because tPA therapy unfortunately increases the risk of intracerebral hemorrhage by approximately 10-fold. Perhaps even more importantly, there is accumulating evidence from experimental models and clinical studies that tPA can have neurotoxic actions separate from its beneficial clot lysis properties. tPA neurotoxicity may further exacerbate ischemic brain damage, particularly in the 50% of patients who have no perfusion improvement after receiving intravenous tPA.
  • tPA-based thrombolytic therapy is the only FDA-approved treatment for achieving vascular reperfusion and clinical benefit, this agent is given to only about 2-5% of stroke patients (25, 26). This may be related, in part, to the elevated risks of symptomatic intracranial hemorrhage, and a short therapeutic time window in order to decrease the clinical risk of tPA's limitations.
  • tPA therapy limitations include: (1) short 3 hr treatment time window, (2) risk of intracerebral hemorrhage, and (3) neurotoxicity. Others have tried to find other lytics with equal thrombolysis properties for safe and effective reperfusion at longer times after stroke onset.
  • compositions and methods that increases the thrombolytic efficacy of tPA, while reducing neurotoxicity and the risk of hemorrhagic transformation.
  • the present invention is related to the field of vascular disorders.
  • the present invention is related to the treatment and management of diseases including, but not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • diseases including, but not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • a patient having suffered a vascular disorder may be administered a composition comprising Annexin A2 and tPA.
  • the tPA dose may be reduced such that the risk of hemorrhagic side effects are minimal.
  • the present invention contemplates a method comprising: a) providing: i) a patient exhibiting symptoms associated with a vascular disorder, and ii) a medium comprising Annexin A2 and tissue plasminogen activator (tPA); and b) administering said medium to said patient under conditions such that said symptoms are reduced.
  • the vascular disorder is selected from the group comprising stroke, myocardial infarction, pulmonary embolism, deep vein thrombosis or intracerebral hematoma.
  • the Annexin A2 and the tPA have a dose ratio of 2:1.
  • the medium comprises a carrier.
  • the Annexin A2 and the tPA are attached to the carrier.
  • the carrier is selected from the group comprising a liposome or a microparticle.
  • the medium comprises a liquid.
  • the administering is intravenous.
  • the patient is a human. In one embodiment, the patient is a non-human.
  • the present invention contemplates a method comprising: a) providing: i) a patient exhibiting symptoms associated with a recently incurred stroke, ii) a medium comprising Annexin A2 and tissue plasminogen activator (tPA), wherein Annexin A2 and tPA have a dose ratio of 2:1; and, b) administering said medium to said human subject under conditions such that said symptoms are reduced.
  • the administering occurred less than three hours after said stroke.
  • the administering occurred less than six hours after said stroke.
  • the administering occurred less than twelve hours after said stroke.
  • the tPA dose is at least two-fold lower than the currently recommended dose.
  • the tPA dose is at least three-fold lower than the currently recommended dose.
  • the tPA dose is at least four-fold lower than the currently recommended dose.
  • the present invention contemplates a medium comprising Annexin A2 and tissue plasminogen activator (tPA).
  • the Annexin A2 and the tPA have a dose ratio of 2:1.
  • the medium further comprises a carrier.
  • the Annexin A2 and the tPA are attached to the carrier.
  • the carrier comprises a liposome population.
  • the carrier comprises a microparticle population.
  • the medium comprises a liquid.
  • the present invention contemplates a kit comprising a medium comprising Annexin A2 and tissue plasminogen activator (tPA).
  • the medium further comprises a carrier.
  • the tPA and the Annexin A2 are attached to said carrier.
  • the kit further comprises a sheet of instructions regarding administration of said medium following a vascular disorder.
  • the vascular disorder is selected from the group comprising stroke, myocardial infarction, pulmonary embolism, deep vein thrombosis or intracerebral hematoma.
  • the kit further comprises a syringe.
  • the kit further comprises an intravenous catheter.
  • the kit further comprises an intravenous drip bag capable of fluid communication with said intravenous catheter.
  • Attachment refers to any interaction between a medium (or carrier) and a drug. Attachment may be reversible or irreversible. Such attachment includes, but is not limited to, covalent bonding, ionic bonding, Van der Waals forces or friction, and the like.
  • a drug is attached to a medium (or carrier) if it is impregnated, incorporated, coated, in suspension with, in solution with, mixed with, etc.
  • a medium refers to any material, or combination of materials, which may serve as vehicle for delivering of a drug, or carrier, to a treatment point (e.g., a thrombosis, a stenosis etc.).
  • a medium is selected from the group including, but not limited to, liquids, foams, or gels (including, but not limited to, hydrogels).
  • a medium constitutes a drug delivery system that provides a controlled and sustained release of drugs over a period of time lasting approximately from 1 day to 6 months.
  • the controlled and sustained drug release is from a carrier mixed within the medium.
  • carrier refers to any material capable of attaching a drug or composition wherein a medium facilitates delivery of the carrier to a treatment point.
  • a carrier is selected from the group including, but not limited to, liposomes, xerogels, or microparticles (i.e., microspheres, liposomes, microcapsules etc.). Any carrier contemplated by this invention may comprise a controlled release formulation.
  • xerogel refers to any device comprising a combination of silicone and oxygen having a plurality of air bubbles and an entrapped drug.
  • the resultant glassy matrix is capable of a controlled release of an entrapped drug during the dissolution of the matrix.
  • foam refers to a dispersion in which a large proportion of gas, by volume, is in the form of gas bubbles and dispersed within a liquid, solid or gel.
  • the diameter of the bubbles are usually relatively larger than the thickness of the lamellae between the bubbles.
  • gel refers to any material forming, to various degrees, a medium viscosity liquid or a jelly-like product when suspended in a solvent.
  • a gel may also encompass a solid or semisolid colloid containing a certain amount of water. These colloid solutions are often referred to in the art as hydrosols.
  • One specific type of gel is a hydrogel.
  • hydrogel refers to any material forming, to various degrees, a jelly-like product when suspended in a solvent, typically water or polar solvents comprising such as, but not limited to, gelatin and pectin and fractions and derivatives thereof.
  • a hydrogel is capable of swelling in water and retains a significant portion of water within its structure without dissolution.
  • the present invention contemplates a gel that is liquid at lower than body temperature and forms a firm gel when at body temperature.
  • drug refers to any pharmacologically active substance capable of being administered which achieves a desired effect.
  • Drugs or compounds can be synthetic or naturally occurring, non-peptide, proteins or peptides, oligonucleotides or nucleotides, polysaccharides or sugars.
  • Drugs or compounds may have any of a variety of activities, which may be stimulatory or inhibitory, such as antibiotic activity, antiviral activity, antifungal activity, steroidal activity, cytotoxic, cytostatic, anti-proliferative, anti-inflammatory, analgesic or anesthetic activity, or can be useful as contrast or other diagnostic agents.
  • Drugs or compounds may be capable of reducing thromboses and/or adhesions.
  • administering refers to any method of providing a drug or compound to a patient such that the drug or compound has its intended effect on the patient.
  • one method of administering is by an indirect mechanism using a medical device such as, but not limited to a syringe, an intravenous catheter, etc.
  • a second exemplary method of administering is by a direct mechanism such as, local tissue administration (i.e., for example, extravascular placement), oral ingestion, transdermal patch, topical, inhalation, suppository etc.
  • biocompatible refers to any material does not elicit a substantial detrimental response in the host. There is always concern, when a foreign object is introduced into a living body, that the object will induce an immune reaction, such as an inflammatory response that will have negative effects on the host.
  • biocompatibility is evaluated according to the application for which it was designed: for example; a bandage is regarded a biocompatible with the skin, whereas an implanted medical device is regarded as biocompatible with the internal tissues of the body.
  • biocompatible materials include, but are not limited to, biodegradable and biostable materials.
  • tissue surface includes, but is not limited to, the external skin or any internal tissue (i.e., for example, the periadvential blood vessel) and/or organ surface.
  • an antiplatelet drug refers to any drug that prevents aggregation of platelets or fibrin formation (i.e., for example as a prior event to adhesion formation).
  • an antiplatelet drug comprises an inhibitor of glycoprotein IIb/IIIa (GPIIb/IIIa).
  • GPIIb/IIIa inhibitor includes, but is not limited to, xemilofiban, abciximab (ReoPro®.) cromafiban, elarofiban, orbofiban, roxifiban, sibrafiban, RPR 109891, tirofiban (Aggrastat®), eptifibatide (Integrilin®), UR-4033, UR-3216 or UR-2922.
  • antithrombins or “antithrombin drug” as used herein, refers to any drug that inhibits or reduces thrombi formation and include, but are not limited to, bivalirudin, ximelagatran, hirudin, hirulog, argatroban, inogatran, efegatran, or thrombomodulin.
  • anticoagulants refers to any drug that inhibits the blood coagulation cascade.
  • a typical anticoagulant comprises heparin, including but not limited to, low molecular weight heparin (LMWH) or unfractionated heparin (UFH).
  • LMWH low molecular weight heparin
  • UHF unfractionated heparin
  • Other anticoagulants include, but are not limited to, tinzaparin, certoparin, parnaparin, nadroparin, ardeparin, enoxaparin, reviparin or dalteparin.
  • Specific inhibitors of the blood coagulation cascade include, but are not limited to, Factor Xa (FXa) inhibitors (i.e., for example, fondaparinux), Factor IXa (FIXa) inhibitors, Factor XIIIa (FXIIIa) inhibitors, and Factor VIIa (FVIIa) inhibitors.
  • FXa Factor Xa
  • FIXa Factor IXa
  • FXIIIa Factor XIIIa
  • FVIIa Factor VIIa
  • patient is a human or animal (i.e., for example, a dog, cat, horse, cow, pig etc.) and need not be hospitalized.
  • out-patients persons in nursing homes are “patients.”
  • a patient may comprise any age of a human or non-human animal and therefore includes both adult and juveniles (i.e., children and/or offspring). It is not intended that the term “patient” connote a need for medical treatment, therefore, a patient may voluntarily or involuntarily be part of experimentation whether clinical or in support of basic science studies.
  • medical device refers broadly to any apparatus used in relation to a medical procedure. Specifically, any apparatus that contacts a patient during a medical procedure or therapy is contemplated herein as a medical device. Similarly, any apparatus that administers a drug or compound to a patient during a medical procedure or therapy is contemplated herein as a medical device.
  • Direct medical implants include, but are not limited to, urinary and intravascular catheters, dialysis catheters, wound drain tubes, skin sutures, vascular grafts and implantable meshes, intraocular devices, implantable drug delivery systems and heart valves, and the like.
  • “Wound care devices” include, but are not limited to, general wound dressings, non-adherent dressings, burn dressings, biological graft materials, tape closures and dressings, surgical drapes, sponges and absorbable hemostats.
  • “Surgical devices” include, but are not limited to, surgical instruments, endoscope systems (i.e., catheters, vascular catheters, surgical tools such as scalpels, retractors, and the like) and temporary drug delivery devices such as drug ports, injection needles etc. to administer the medium.
  • a medical device is “coated” when a medium comprising a tPA and Annexin A2 becomes attached to the surface of the medical device (either directly or indirectly). Such an indirect attachment may result from coating a medical device with a polymer comprising the medium. This attachment may be permanent or temporary. When temporary, the attachment may result in a controlled release of a tPA/rA2 combination.
  • dialysis/apheresis catheter refers to any multi-lumen catheter (i.e., for example, a triple lumen catheter) capable of providing a simultaneous withdrawal and return of blood to a patient undergoing a blood treatment process.
  • Apheresis (called also pheresis) comprises a blood treatment process involving separation of blood elements that can remove soluble drugs or cellular elements from the circulation.
  • pheresis comprises a blood treatment process involving separation of blood elements that can remove soluble drugs or cellular elements from the circulation.
  • pheresis comprises a blood treatment process involving separation of blood elements that can remove soluble drugs or cellular elements from the circulation.
  • Deisseroth et al. “Use Of Blood And Blood Products”, Cancer: Principles And Practice Of Oncology, Devita, V. T. Jr. et al. Editors, Philadelphia: J. B. Lippincott Company 1989, p. 2045-2059.
  • blood is withdrawn from a donor, some blood elements (
  • dialysis catheter refers to any device capable of removing toxic substances (impurities or wastes) from the body when the kidneys are unable to do so.
  • a dialysis catheter may comprise a single catheter having at least a dual lumen (i.e., one lumen withdraws arterial blood and a second lumen returns the dialyzed blood to the venous system) or involve placing two catheters—one that is placed in an artery, and one in an adjacent vein. Dialysis catheters are most frequently used for patients who have kidney failure, but may also be used to quickly remove drugs or poisons in acute situations.
  • peritoneal dialysis catheter refers to any continuous flow catheters with at least two lumens, one of which is a short lumen (used to infuse a dialysis solution into the peritoneum), and the other of which is a long coiled lumen having a plurality of openings, generally located on the inside of the coil. It is believed that peritoneal solutes enter into the coiled lumen openings and are thereby removed from the peritoneum.
  • peritoneal dialysis works by using the peritoneal membrane inside the abdomen as the semipermeable membrane. Special solutions that facilitate removal of toxins may be infused in, remain in the abdomen for a time, and then drained out.
  • fixed split-tip dialysis catheter refers to any catheter having at least two distinct elongated end portions that extend substantially parallel to the longitudinal axis of the catheter and are flexible to the lateral displacement of an infused fluid. It is believed that this flexibility prevents a permanent catheter tip splay that is known to injure tissue.
  • a fixed-tip dialysis catheter provides indwelling vascular access for patients undergoing long-term renal dialysis care (i.e., for example, end-stage renal disease).
  • femoral catheter refers to any catheter that is inserted into the femoral vein.
  • Femoral catheters are typically provided for intermediate term blood access because the superior vena cava is relatively close to the right atrium of the heart, the minimal range of shape changes of these veins with natural movements of the patient (to minimize the damage to the vessel intima), and because of good acceptance by the patients of the skin exit on the thoracic wall. Further, the femoral veins are easy to cannulate, so that catheters of this invention may be inserted into the femoral veins at the bed side.
  • endoscope refers to any medical device that is capable of being inserted into a living body and used for tasks including, but not limited to, observing surgical procedures, performing surgical procedures, or applying medium to a surgical site.
  • An endoscope is illustrated by instruments including, but not limited to, an arthroscope, a laparoscope, hysteroscope, cytoscope, etc. It is not intended to limit the use of an endoscope to hollow organs. It is specifically contemplated that endoscopes, such as an arthroscope or a laparoscope is inserted through the skin and courses to a closed surgical site.
  • microparticle refers to any microscopic carrier to which a drug or compound may be attached.
  • microparticles contemplated by this invention are capable of formulations having controlled release properties.
  • PLGA refers to mixtures of polymers or copolymers of lactic acid and glycolic acid.
  • lactide polymers are chemically equivalent to lactic acid polymer and glycolide polymers are chemically equivalent to glycolic acid polymers.
  • PLGA contemplates an alternating mixture of lactide and glycolide polymers, and is referred to as a poly(lactide-co-glycolide) polymer.
  • stenosis is defined herein as referring to any narrowing of the internal diameter of a biological tissue, such as a vessel.
  • narrowing is caused by phenomenon including, but not limited to, arteriosclerosis, scar tissue and/or adhesions.
  • restenosis is defined herein as referring to any condition wherein “stenosis”, having been treated and at least partially reversed, recurs.
  • vascular access site is defined herein as referring to any percutaneous insertion of a medical device into the vasculature.
  • a hemodialysis catheter placement comprises a vascular access site.
  • Such sites may be temporary (i.e., placed for a matter of hours) or permanent (i.e., placed for days, months or years).
  • syringe refers to any device or apparatus designed for liquid administration, as defined herein.
  • a syringe or catheter may comprise at least one storage vessel (i.e., for example, a barrel) wherein a single medium resides prior to administration.
  • a syringe or catheter comprising two or more barrels, each containing a separate medium, may mix the media from each barrel prior to administration or the media of each barrel may be administered separately.
  • any catheter designed to perform dialysis, as defined herein may also administer liquids.
  • vascular graft refers to any conduit or portion thereof intended as a prosthetic device for conveying blood and, therefore, having a blood contacting surface (i.e., “luminal”). While usually in a tubular form, the graft may also be a sheet of material useful for patching portions of the circumference of living blood vessels (these materials are generally referred to as surgical wraps). Likewise, the term vascular graft includes intraluminal grafts for use within living blood vessels. The inventive grafts as such may also be used as a stent covering on the exterior, luminal or both surfaces of an implantable vascular stent.
  • anti-thrombotic drug combination refers to any composition comprising at least one plasminogen activator (i.e., for example, tPA) and at least one Annexin protein (i.e., for example, recombinant Annexin A2; rA2).
  • plasminogen activator i.e., for example, tPA
  • Annexin protein i.e., for example, recombinant Annexin A2; rA2
  • Other drugs including, but not limited to, antithrombin drugs, anticoagulant drugs or antiinflammatory drugs may also be in this combination.
  • controlled release drug elution refers to any stable and quantifiable drug release from a polymer-based medium as contemplated herein.
  • synthetic vascular graft refers to any artificial tube or cannula designed for insertion into a blood vessel. Such grafts may be constructed from polytetrafluoroethylene (PTFE).
  • PTFE polytetrafluoroethylene
  • fibrin sheath refers to any encapsulation of a medical device subsequent to implantation.
  • platelets and white blood cells respond to foreign substances in much the same way as an injured tissue (i.e., for example, a blood vessel) and that platelet adherence, followed by fibrin encapsulation, is involved in fibrin sheath formation.
  • non-adhesive luminal surface refers to any vascular graft having been constructed, or treated, that prevents platelet attachment and subsequent thrombosis formation.
  • the Food & Drug Administration FDA
  • the FDA has approved the administration to humans of no more than 100 mg of tissue plasminogen activator (tPA: Activase®, Genentech) over the period of one hour.
  • vascular disorder refers to any biochemical, physiological, structural, or anatomical abnormality occurring within the cardiovascular system.
  • vascular disorders include, but are not limited to, stroke, myocardial infarction, deep vein thrombosis, pulmonary embolism, thrombophlebitis, or intracerebral hematoma.
  • annexin refers to a family of highly homologous antithrombotic proteins believed to prevent both cellular and humoral amplification of platelet aggregation. It also may act as a “plasmin activator” such that when in contact with a plasminogen activator, the production of plasmin is increased.
  • plasminogen activator refers to any compound (i.e., usually a protein) that is capable of converting plasminogen into plasmin (i.e., for example, tissue plasminogen activator (tPA) or urokinase-type plasminogen activator (uPA)).
  • tissue plasminogen activator tPA
  • uPA urokinase-type plasminogen activator
  • population refers to any mixture of particles (i.e., for example, liposomes or microparticles) having a distribution in diameter size.
  • a population of microparticles may range is particle diameter from between approximately 1-5000 ⁇ m, preferably between approximately 350-3500 ⁇ m, more preferably between approximately 1000-2000 ⁇ m.
  • symptoms are reduced refers to a qualitative or quantitative reduction in detectable symptoms, including, but not limited to, a detectable impact on the rate of recovery from disease (e.g. rate of thrombus regression) or a detectable impact on the rate of development of disease (e.g., rate of thrombus growth).
  • cently incurred refers to the onset of a particular vascular disorder within the past twelve hours. Preferably onset within the past six hours, but more preferably onset within the past four hours.
  • a sheet of instructions refers to any method of means of storing and retrieving written or spoken information. Such instructions are preferably related to the use of a kit containing a therapeutic method contemplated herein. Such instructions may also provide information regarding interpretation of the results of the therapeutic method such that a clinical diagnosis can be reached.
  • FIG. 1 presents exemplary data showing an effect of rA2 on tPA-dependent plasmain generation in vitro.
  • FIG. 2 presents examplary data showing an effect of treating rats at 2 hrs after initiation of cerebral ischemia.
  • FIG. 2A demonstrates that two hours after initiation of ischemia, animals were treated intravenously with either saline, high dose tPA (10 mg/kg, H-tPA), intermediate dose tPA (5 mg /kg, M-tPA), low dose tPA (2.5 mg/kg, L-tPA), rA2 alone (5 mg/kg), or a combination of low dose tPA (2.5 mg/kg) plus rA2 (5 mg/kg).
  • Laser doppler flowmetry was used to monitor regional cerebral blood flow (rCBF) for up to 1 h after treatment.
  • FIG. 3 presents exemplary data showing an effect of treating rats at 4 hrs after initiation of ischemia.
  • FIG. 3A At 4 h after stroke onset, three groups of rats were treated intravenously with either saline, high dose tPA (10 mg/kg, H-tPA), or low dose tPA (2.5 mg/kg, L-tPA) plus rA2 (5 mg/kg). Shown here are representative images of brain sections after TTC staining at 24 h after initiating ischemia. Ischemic infarctions (white color area) were detected in all three groups, however large visible hemorrhage appeared only on the brain sections of H-tPA treated rats pointed by arrows.
  • FIG. 3B At 24 h after stroke, brain infarction was quantified using computer-assisted image analysis.
  • FIG. 3C Volumes of intracerebral hemorrhage ware quantified with hemoglobin assay at 24 h after stroke.
  • FIG. 4 present exemplary data showing an effect of tPA along or in combination with rA2 on fibrinolysis, and reperfusion improvement.
  • FIG. 4A Effects of rA2 plus low dose tPA in fibrinolysis.
  • Plasma samples were collected before ischemia (Pre-ischemia), just before thrombolytic therapy (Ischemia), and 1 h after treatment (Thrombolysis).
  • FIG. 4B Representative MR angiograms (MRA). rA2 (5 mg/kg) plus low dose tPA (2.5 mg/kg) was IV injected at 4 hr after stroke onset in embolic stroke rats. Time of flight (TOF) technique was used to assess cerebral vessel MRA imaging. It showed clear MCA occlusion examined at 3 hr after stroke (Before Thrombolysis), and clear recanalization at 1 hr after thrombolysis by the combination.
  • TOF Time of flight
  • FIG. 5 presents one embodiment of a human Annexin II protein sequence (A) and corresponding nucleotide sequence (B).
  • the present invention is related to the field of vascular disorders.
  • the present invention is related to the treatment and management of diseases including, but not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • diseases including, but not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • a patient having suffered a vascular disorder may be administered a composition comprising tPA and Annexin A2.
  • the tPA dose may be reduced such that the risk of hemorrhagic side effects are minimal.
  • Hemorrhagic transformations, neurotoxicities and a short treatment time window include, but not limited to, some limitations for effective tPA stroke therapy. These limitations are related to the high dose of tPA required to generate the necessary amounts of plasmin sufficient for clot lysis.
  • the present invention contemplates that soluble Annexin A2 unexpectedly potentiates tPA-mediated plasmin generation in vitro, thereby resulting in improved reductions in in vivo thrombus formation.
  • a combination of Annexin A2 with tPA can significantly enhance thrombolysis efficacy, such that lower doses of tPA can be administered that avoid neurotoxic and hemorrhagic complications.
  • Annexin A2-tPA induced in vitro amplification of plasmin generation.
  • a tPA-recombinant Annexin A2 protein combination was administered 2 hours after the ischemia-induced embolic stroke.
  • the effective dose required for tPA to restore cerebral blood flow was reduced by 4-fold, also resulting in reduced brain infarctions.
  • the Annexin A2-tPA combination also prolonged the effective treatment time window to prevent thrombolysis (the currently accepted therapeutic window is limited to three hours post stroke event).
  • the Annexin A2 5 mg/kg/tPA (2.5 mg/kg) (i.e., a 2:1 Annexin A2-tPA ratio) combination significantly enhanced fibrinolysis and reperfusion; attenuated mortality, brain infarctions, and hemorrhagic transformations, when administered at 4 hours post stroke event.
  • the effective thrombolytic dose of tPA can be decreased.
  • brain hemorrhage and infarction can be reduced, and the therapeutic time window for stroke reperfusion prolonged.
  • tPA-Annexin A2-plasminogen amplifies tPA-converted plasmin generation in vitro and inhibits clot formation in vivo.
  • a combination treatment at 2 hrs after stroke onset with low dose (2.5 mg/kg) plus Annexin A2 (5 mg/kg) showed similar therapeutic benefits as treatment with high dose (10 mg/kg) tPA alone in terms of restoring cerebral perfusion and reducing infarct volumes at 24 hrs.
  • a tPA plus Annexin A2 combination treatment improved cerebral perfusion and reduced infarction without hemorrhagic transformation.
  • ischemic stroke patients may be able to receive tPA treatments beyond the currently recommended three hour post-stroke event time window.
  • patients receiving a tPA/Annexin A2 combination treatment will have lower risk for intracerebral hemorrhage, and better clinic outcomes.
  • Tissue plasminogen activator is a fibrin-specific activator that converts plasminogen to plasmin.
  • tPA stimulates thrombolysis and rescues the ischemic brain by restoring blood flow.
  • exogenous tPA may have: i) pleiotropic actions in the brain (14); ii) direct vasoactivity (27); iii) cleavage of the N-methyl-D-aspartate (NMDA) NR1 subunit (28); and iv) activation of other extracellular proteases such as matrix metalloproteinases (MMPs) (3, 13, 28, 29).
  • MMPs matrix metalloproteinases
  • the present invention contemplates a composition capable of being administered using an effective tPA dose that is four times lower than currently recommended. Although it is not necessary to understand the mechanism of an invention it is believed that such a reduced dose reduces tPA-mediated side effects resulting from intracerebral hemorrhage and/or direct neurotoxicity.
  • the present invention contemplates a composition comprising a combination of tPA and recombinant human Annexin A2, wherein plasmin generation is amplified. Although it is not necessary to understand the mechanism of an invention it is believed that such plasmin generation will improve perfusion and provide greater clinical benefits.
  • the present invention contemplates a method comprising administering a composition comprising tPA and Annexin A2 to a patient more than three hours after a stroke event, and under conditions such that effective treatment is provided.
  • a composition comprising tPA and Annexin A2
  • Annexin A2 is an endogenous human protein
  • tPA may reduce the risk of an immune response.
  • the present embodiments identify new strategies that can increase the thrombolytic efficacy of tPA, while reducing its associated neurotoxicity and hemorrhagic transformation (1-6).
  • Successful clot lysis for stroke therapy has required high doses of tPA, but it is well accepted that this practice increases the risk of intracerebral hemorrhage (7-9).
  • Emerging evidence from experimental models and clinical studies demonstrates that tPA administered systemically can enter both normal and injured brain parenchyma (10, 11). Moreover, exogenous tPA can have neurotoxic actions that are not related to any clot lysis properties (12-15). Also, it has been reported that tPA mediated neurotoxicity may exacerbate ischemic damage, particularly to the about 50% of patients who have poor improvement in perfusion after receiving intravenous tPA (14).
  • Annexin A2 (also known as Annexin II) is a calcium- and phospholipid-binding protein that serves as a pro-fibrinolytic co-receptor for tPA and plasminogen on endothelial cells (16, 17).
  • Annexin A2 accelerates the activation of the clot-dissolving protease plasmin by complexing with tPA and with the plasmin precursor plasminogen which binds on endothelial cell surface and exist in the clot (18, 19).
  • Annexin A2 is also transported to the extracellular environment, and exists as both soluble and membrane-bound protein.
  • Annexin A2 can be detected in human plasma and can be transported to the cell surface in response to cellular stress (33). Recent reports have demonstrated that soluble Annexin A2 not only dramatically increases tPA-dependent plasmin generation in vitro, but also reduces thrombus formation in rat carotid arteries and middle cerebral arteries in vivo (20, 21, respectively).
  • the present invention contemplates a recombinant human Annexin A2 having a tPA binding site. In one embodiment, the present invention contemplates a tPA peptide is bound to the Annexin A2 peptide. In one embodiment, the binding site is modified to provide a high affinity tPA binding site.
  • the present invention contemplates a recombinant human Annexin A2 having a plasmin binding site. In one embodiment, the present invention contemplates a plasmin peptide is bound to the Annexin A2 peptide. In one embodiment, the binding site is modified to provide a high affinity plasmin binding site.
  • Annexin A2 annexin proteins
  • Annexin A2 may be linked with the Fc portion of an immunoglobulin.
  • this administration of Annexin A2 is advocated for administration in a “heterotetramer” form.
  • Modified Annexin A2 proteins, including recombinant proteins, were suggested to prevent transient ischemic attacks from developing into full blown strokes (i.e., a preventative therapy).
  • Combinations of Annexin A2 with tPA were not suggested, and in fact the disadvantages of tPA (i.e., increased bleeding risk) were pointed out tending to teach away from the presently contemplated embodiment.
  • Annexin A2 has also been proposed as an effective therapeutic to reduce hemorrhaging when administered following retinal trauma and other ophthalmological disorders. Rich, “Methods of Stabilizing Hyaluronidase with Annexin A2,” U.S. Pat. No. 7,115,408 (herein incorporated by reference).
  • the present invention contemplates that Annexin A2 may be therapeutically administered with tissue plasminogen activator (tPA) for treatment following a stroke.
  • tissue plasminogen activator tPA
  • the present invention addresses current problems associated with the fact that tPA is currently the only FDA approved medicine for the generation of vascular reperfusion in stroke patients but relatively few patients actually receive tPA due to the increased risk of intracerebral hemorrhaging.
  • the present invention contemplates that Annexin A2, an endogenous protein of human plasma, may be combined in treatment regimens with tPA such that lower, non-neurotoxic dosages of tPA may be administered.
  • the dose of WA may be reduced at least four-fold from currently recommended dosages when administered in combination with Annexin A2 a still achieve similar therapeutic effectiveness.
  • tPA is broadly used in treating a range of thrombotic disorders, besides acute ischemic stroke, this new combination approach for lowering the minimum effective tPA dose may potentially improve tPA treatments for other conditions such as acute myocardial infarction, pulmonary embolism, deep venous thrombosis, thrombosed medical devices (i.e., for example, intravenous catheters), and lysis of intracerebral hematomas.
  • tPA's associated risks when administered alone and at a high dose may be alleviated and/or avoided.
  • tPA/Annexin A2 combination therapies may further improve treatment outcomes for other medical conditions including, but not limited to, myocardial infarction, pulmonary embolism, deep vein thrombosis and lysis for intracerebral hematoma.
  • Administered doses for a WA and rA2 in vivo combination was based on in vitro plasmin generation data.
  • the in vitro data indicated that an rA2/tPA combination at a 2:1 ratio increases the plasmin-generating capability of tPA by almost four-fold.
  • the thrombolytic profiles may be evaluated by examining plasma levels of a fibrin degradation product, D-dimer.
  • the assay is based upon degradation of cross-lined fibrin (enriched in fibrin thrombi) that produces a number of fragments containing the D-dimer epitope.
  • the released D-dimer epitope is then utilized as an indicator the extent of fibrin clot lysis by ELISA.
  • ELISA data showed that low dose WA plus rA2 significantly increased D-dimer levels by about 2.9 fold compared to either saline or tPA alone.
  • Annexin A2 has been reported to be the putative human endothelial tPA cell surface receptor. This relationship has resulted in broad speculation that the Annexin A2-tPA relationship may play a role in hemostasis and thrombosis. Hajjar et al., “An Endothelial Cell Receptor for Plasminogen/Tissue Plasminogen Activator” Journal of Biological Chemistry 269:21191-21197 (1994). Other, in vitro studies, have shown that Annexin A2 is capable of binding both plasmin and plasminogen in human endothelial cells and suggest that Annexin A2 may act as a catalyst for the activation of plasminogen by tPA.
  • Annexin A2-plasminogen activator combinations have been used for the treatment of other diseases. For example, an anti-angiogenic plasmin fragment and an Annexin A2 heterotetramer was reported to affect angiogenesis. In one combination used to effect angiogenesis, an Annexin A2 heterotetramer was combined with a urokinase-type plasminogen activator; an enzyme that interacts with tPA. Novokhatny et al., “Tissue-type plasminogen activator (tPA) interacts with urokinase-type plasminogen activator (uPA) via tPA's lysine binding site.
  • tPA tissue-type plasminogen activator
  • uPA urokinase-type plasminogen activator
  • rA2 Recombinant human Annexin A2 protein
  • a combination of rA2 and tPA significantly amplified tPA-converted plasmin generation in a dose-dependent manner.
  • the data demonstrate that equal plasmin activity levels can be reached by either: i) a high dose tPA alone (clinically associated with cerebral hemorrhage); or ii) different rA2/tPA combinations having lower doses of tPA, such as: i) tPA 5 ⁇ g/ml (alone) versus a tPA (1 ⁇ g/ml)/rA2 (2.5 ⁇ g/ml) combination; or ii) tPA 10 ⁇ g/ml (alone) versus a tPA (2.5 ⁇ g/ml)/rA2 (5 ⁇ g/ml) combination.
  • rats were intravenously administered with either: i) saline; ii) a high dose tPA (10 mg/kg); iii) an intermediate dose tPA (5 mg /kg); iv) a low dose tPA (2.5 mg/kg); v) rA2 alone (5 mg/kg); or vi) a combination of low dose tPA (2.5 mg/kg) plus rA2 (5 mg/kg).
  • Annexin A2 accelerates the activation of plasmin by complexing with tPA and with the plasmin precursor plasminogen, which binds the endothelial cell surface and is enriched in the clot (19, 31). Therefore, a combination of tPA and recombinant Annexin A2 might locally bind plasminogen and consequently amplify plasmin generation in the vicinity of the clot, thereby resulting in more locally effective fibrinolysis.
  • the present invention contemplates a method wherein a combination therapy with tPA plus rA2 prolongs the conventional thrombolytic time window.
  • the presently contemplated invention demonstrates that addition of a “plasmin amplifier” such as rA2 can decrease the effective thrombolytic dose of tPA, thereby reducing hemorrhage and brain infarction, and prolonging the reperfusion time window for stroke.
  • a “plasmin amplifier” such as rA2 can decrease the effective thrombolytic dose of tPA, thereby reducing hemorrhage and brain infarction, and prolonging the reperfusion time window for stroke.
  • Plasma levels were examined of the fibrin degradation product D-dimer, a biomarker for fibrinolysis.
  • ELISA data showed both high dose tPA and low dose tPA plus rA2 significantly increased D-dimer after thrombolysis for 2.2-fold and 3.2-fold, respectively, and the increase in low dose tPA plus rA2 combination was significantly greater compared to high dose tPA-only treatment. See, FIG. 4A . These data indicate that this combination thrombolytic therapy was more effective and specific for fibrinolysis than tPA along in delayed 4 hrs treatment (24).
  • the present invention contemplates administering a rA2/tPA composition for the treatment of a vascular disorder.
  • the disorder may comprise a stroke.
  • the disorder may comprise a thrombosis.
  • the thrombosis may be attached to a medical device.
  • the disorder may comprise an embolism.
  • Thrombolytic therapy with tPA is the only FDA-approved medicine for achieving both vascular reperfusion and clinical benefit, but only 2-5% of stroke patients receive tPA in the US. In part, this because tPA therapy unfortunately increases the risk of intracerebral hemorrhage by approximately 10-fold. Perhaps even more importantly, there is accumulating evidence from experimental models and clinical studies that tPA can have neurotoxic actions separate from its beneficial clot lysis properties. tPA neurotoxicity may further exacerbate ischemic brain damage, particularly in the 50% of patients who have no perfusion improvement after receiving intravenous tPA.
  • the present invention contemplates increasing the thrombolytic efficacy of tPA, while reducing neurotoxicity and the risk of hemorrhagic transformation. Although it is not necessary to understand the mechanism of an invention, it is believed that Annexin A2 will lower the dose of tPA required to generate plasmin thereby allowing the use of lower non-neurotoxic doses of tPA and also extending the treatment time window without incurring risks of brain hemorrhage.
  • ischemic stroke This type of stroke can sometimes lead to a brain hemorrhage because the affected brain tissue softens and this can lead to breaking down of small blood vessels.
  • brain hemorrhage can occur when people have problems forming blood clots.
  • Clots which are the body's way of stopping any bleeding, are formed by proteins called coagulation factors and by sticky blood cells called platelets. Whenever the coagulation factors or platelets do not work well or are insufficient in quantity, people may develop a tendency to bleed excessively.
  • Ischemic strokes may be preceded by transient ischemic attacks (TIA), and it is estimated that about 300,000 persons suffer a TIA every year in the United States. It would be desirable to have a safe and effective agent that could be administered as a bolus and would for several days prevent recurrence of cerebral thrombosis without increasing the risk of cerebral hemorrhage. Thrombosis also contributes to peripheral arterial occlusion in diabetics and other patients, and an efficacious and safe antithrombotic agent for use in such patients is needed.
  • TIA transient ischemic attacks
  • Hemorrhagic stroke occurs when a blood vessel bursts inside the brain.
  • the brain is very sensitive to bleeding and damage can occur very rapidly, either because of the presence of the blood itself, or because the fluid increases pressure on the brain and harms it by pressing it against the skull.
  • Bleeding irritates the brain tissue, causing swelling.
  • the surrounding tissues of the brain resist the expansion of the bleeding, which is finally contained by forming a mass (i.e., for example, an intracerebral hematoma). Both swelling and hematoma will compress and displace normal brain tissue.
  • hemorrhagic stroke is associated with high blood pressure, which stresses the artery walls until they break.
  • Aneurysm This is a weak spot in an artery wall, which balloons out because of the pressure of the blood circulating inside the affected artery. Eventually, it can burst and cause serious harm. The larger the aneurysm is, the more likely it is to burst.
  • Stroke symptoms are typically of sudden onset and may quickly become worse. Stroke symptoms may include, but are not limited to: i) Weakness or inability to move a body part; ii) Numbness or loss of sensation; iii) Decreased or lost vision (may be partial); iv) Speech difficulties; v) Inability to recognize or identify familiar things; vi) Sudden headache; vii) Vertigo; viii) Dizziness; xi) Loss of coordination; x) Swallowing difficulties; and xi) Sleepy, stuporous, lethargic, comatose, and/or unconscious.
  • a stroke event may be detected by using a neurologic exam, which would be expected to show abnormal results. Further, a patient may look drowsy and confused. An eye examination may show abnormal eye movements, and changes may be seen upon retinal examination (examination of the back of the eye with an instrument called ophthalmoscope). The patient may also have abnormal reflexes.
  • a computerized tomography scan will confirm the presence of a brain hemorrhage by providing pictures of the brain.
  • a brain magnetic resonance imaging (MRI) scan can also be obtained later to better understand what caused the bleeding.
  • a conventional angiography i.e., for example, an X-ray of the arteries using dye
  • Other tests may include, but are not limited to: complete blood count, bleeding time, prothrombin/partial thromboplastin time (PT/PTT), and CSF (cerebrospinal fluid) examination.
  • Thrombosis may be defined as the formation, development, or presence of a blood clot (i.e. for example, a thrombus) in a blood vessel and is believed to be a common severe medical disorder.
  • Thromboses may be involved in the generation of a variety of vascular disorders including, but not limited to, myocardial infarctions, cardiac ischemia, and/or deep vein thrombosis.
  • Unstable angina caused by inadequate oxygen delivery to the heart due to coronary occlusion, is the most common cause of admission to hospital, with 1.5 million cases a year in the United States alone.
  • patients with occlusion of coronary arteries are treated with angioplasty and stenting, the use of an antibody against platelet GP IIb/IIIa decreases the likelihood of restenosis.
  • the same antibody has shown no benefit in unstable angina without angioplasty, and a better method for preventing coronary occlusion in these patients is needed.
  • Deep venous thrombosis is a frequent complication of surgical procedures such as hip and knee arthroplasties. It would be desirable to prevent thrombosis without increasing hemorrhage into the field of operation. Similar considerations apply to venous thrombosis associated with pregnancy and parturition. Some persons are prone to repeated venous thrombotic events and are currently treated by antithrombotic agents such as coumarin-type drugs. The dose of such drugs must be titrated in each patient, and the margin between effective antithrombotic doses and those increasing hemorrhage is small. Having a treatment with better separation of antithrombotic activity from increased risk of bleeding is desirable.
  • Deep vein thrombosis may be detected by tests including, but not limited to: i) Doppler ultrasound exam of an extremity blood flow studies; ii) Venography of the legs; or iii) Plethysmography of the legs.
  • Platelet microaggregates which plug capillaries and accumulate over damaged or activated endothelial cells in small blood vessels.
  • Inhibitors of platelet aggregation including agents suppressing the formation or action of thromboxane A 2 , ligands of GP IIa/IIIb, and drugs acting on ADP receptors such as clopidogrel (Hallopeter, Nature 409:202-207 (2001), incorporated herein by reference), interfere with this process and therefore increase the risk of bleeding (Levine et al., 2001).
  • occlusion by an arterial or venous thrombus requires the continued recruitment and incorporation of platelets into the thrombus.
  • platelets To overcome detachment by shear forces in large blood vessels, platelets must be bound tightly to one another and to the fibrin network deposited around them.
  • a prothrombinase complex is formed on the surface of activated platelets and microvesicles. This generates thrombin and fibrin.
  • Thrombin is itself a potent platelet activator and inducer of the release of Gas6 (Ishimoto and Nakano, FEBS Lett. 446:197-199 (2000), incorporated herein by reference). Fully activated platelets bind tightly to the fibrin network deposited around them.
  • antibodies against Gas6 inhibited platelet aggregation in vitro as well as thrombosis induced in vivo by collagen and epinephrine.
  • such antibodies, or ligands competing for Gas6 binding to receptor tyrosine kinases might be used to inhibit thrombosis.
  • such an inhibitor would also have additional suppressive activity on the Gas6-mediated cellular amplification mechanism.
  • Annexins A strategy for preventing both cellular and humoral amplification of platelet aggregation is provided by the annexins, a family of highly homologous antithrombotic proteins of which ten are expressed in several human tissues (Benz and Hofmann, Biol. Chem. 378:177-183 (1997), incorporated herein be reference).
  • Annexins share the property of binding calcium and negatively charged phospholipids, both of which are required for blood coagulation. Under physiological conditions, negatively charged phospholipid is mainly supplied by phosphatidylserine (PS) in activated or damaged cell membranes. In intact cells, PS is confined to the inner leaflet of the plasma membrane bilayer and is not accessible on the surface.
  • PS phosphatidylserine
  • Proteins involved in the blood coagulation cascade (factors X, Xa, and Va) bind to membranes bearing PS on their surfaces, and to one another, forming a stable, tightly bound prothrombinase complex.
  • annexins including II, V, and VIII, bind PS with high affinity, thereby preventing the formation of a prothrombinase complex and exerting antithrombotic activity.
  • Platelets are also known to release growth factors, in particular, platelet-derived growth factor (PGDF) which promote smooth muscle cell proliferation.
  • PGDF platelet-derived growth factor
  • Schwartz et al. “Common Mechanisms Of Proliferation Of Smooth Muscle In Atherosclerosis And Hypertension” Hum Pathol. 18:240-247 (1987).
  • PGDF platelet-derived growth factor
  • the bound platelets release growth factors that result in restenosis.
  • Restenosis is a condition where smooth muscle cells accumulate within an injured blood vessel such that vessel blockage occurs within 3-6 months (i.e., such as following an intravascular stent placement).
  • Stent technology is attempting to solve this problem using antiplatelet drug-eluting stents or grafts, but its efficacy is as yet unknown.
  • Falotico, R. “Coated Medical Devices For The Prevention And Treatment Of Vascular Disease” United States Patent Application 2003/0216699 A1. Filed: May 7, 2003. Published; Nov. 20, 2003.
  • the present invention contemplates administering a drug combination comprising tPA and Annexin A2 at, or near, an intravascular stent placement.
  • Platelet-mediated thrombosis is also known to complicate successful native and synthetic graft implantation.
  • Vascular neointimal formations are known to occur in native and synthetic grafts, particularly in the venous outflow tracts. Walles et al., “Functional Neointima Characterization Of Vascular Prostheses In Human” Ann Thorac Surg. 77:864-868 (2004).
  • Vascular neointimal formations are composed primarily of smooth muscle cells, and ultimately lead to a decreased blood flow within the grafts.
  • Platelet-released growth factors may, in part, stimulate vascular neointimal formations.
  • blood flow becomes more turbulent and further injury occurs, resulting in additional platelet recruitment.
  • fibrin deposition may result with complete graft failure as a probable consequence.
  • a drug combination comprising tPA and Annexin A2 may have distinct advantages over currently recommended antiproliferative and/or anticoagulant therapies.
  • the present invention contemplates devices and methods to administer a drug combination to a graft venous outflow tract.
  • a drug combination is administered comprising tPA and Annexin A2.
  • the medium or carrier may be wrapped or draped around the exterior graft surface such that the drug combination diffuses to an intraluminal blood vessel surface (i.e., for example, the vaso vasorum).
  • Another embodiment of the present invention contemplates coating a medical device with a medium or carrier comprising tPA and Annexin A2.
  • a medical device is “coated” when a medium comprising tPA and Annexin A2 becomes attached to the surface of the medical device.
  • attachment includes, but is not limited to, surface adsorption, impregnation into the material of manufacture, covalent or ionic bonding and simple friction adherence to the surface of the medical device.
  • Carriers or mediums contemplated by this invention may comprise a polymer including, but not limited to, gelatin, collagen, cellulose esters, dextran sulfate, pentosan polysulfate, chitin, saccharides, albumin, fibrin sealants, synthetic polyvinyl pyrrolidone, polyethylene oxide, polypropylene oxide, block polymers of polyethylene oxide and polypropylene oxide, polyethylene glycol, acrylates, acrylamides, methacrylates including, but not limited to, 2-hydroxyethyl methacrylate, poly(ortho esters), cyanoacrylates, gelatin-resorcin-aldehyde type bioadhesives, polyacrylic acid and copolymers and block copolymers thereof.
  • a polymer including, but not limited to, gelatin, collagen, cellulose esters, dextran sulfate, pentosan polysulfate, chitin, saccharides, albumin, fibrin sealants,
  • a pulmonary embolus is a blockage of an artery in the lungs by fat, air, blood clot, or tumor cells. Pulmonary emboli are most often caused by blood clots in the veins, especially veins in the legs or in the pelvis (hips). More rarely, air bubbles, fat droplets, amniotic fluid, or clumps of parasites or tumor cells may obstruct the pulmonary vessels.
  • DVT deep vein thrombosis
  • Risk factors for a pulmonary embolus may include, but are not limited to: i) Prolonged bed rest or inactivity (including long trips in planes, cars, or trains); ii) Oral contraceptive use; iii) Surgery (especially pelvic surgery); iv) Childbirth; v) Massive trauma; vi) Burns; vii) Cancer; viii) Stroke; ix) Heart attack; x) Heart surgery; or xi) Fractures of the hips or femur.
  • persons with certain clotting disorders and/or autoimmune diseases i.e., for example, anti-cardiolipin antibody syndrome
  • Symptoms of pulmonary embolism may be vague, or they may resemble symptoms associated with other diseases. Symptoms can include, but are not limited to: i) Sudden cough; ii) Bloody sputum (significant amounts of visible blood or lightly blood streaked sputum); iii) Sudden onset of shortness of breath at rest or with exertion; iv) splinting of ribs with breathing (bending over or holding the chest); v) chest pain; vi) rapid breathing; or vii) rapid heart rate (tachycardia)
  • Pulmonary emboli may be identified using tests including, but not limited to: i) Arterial blood gases; ii) Pulse oximetry; iii) Chest x-ray; iv) Pulmonary ventilation/perfusion scan; v) Pulmonary angiogram; vi) electrocardiogram; and v) computerized tomographic chest angiogram.
  • Thrombophlebitis is swelling (inflammation) of a vein caused by a blood clot. Such conditions are usually a result of sitting for a long period of time (such as on a long airplane trip). Disorders that increase a person's chance for blood clots also lead to thrombophlebitis. Superficial thrombophlebitis affects veins near the skin surface.
  • Symptoms often associated with superficial thrombophlebitis may include but are not limited to: i) Warmth and tenderness over the vein; ii) Pain in the part of the body affected; iii) Skin redness (not always present); or iv) Inflammation (swelling) in the part of the body affected.
  • Objective tests may be performed to detect thrombophlebitis including, but not limited to: i) Doppler ultrasound; ii) Venography; and iii) Blood coagulation studies.
  • the present invention further provides pharmaceutical compositions (e.g., comprising the Annexin A2/tPA combinations described above).
  • the pharmaceutical compositions of the present invention may be administered in a number of ways depending upon whether local or systemic treatment is desired and upon the area to be treated. Administration may be topical (including ophthalmic and to mucous membranes including vaginal and rectal delivery), pulmonary (e.g., by inhalation or insufflation of powders or aerosols, including by nebulizer; intratracheal, intranasal, epidermal and transdermal), oral or parenteral. Parenteral administration includes intravenous, intraarterial, subcutaneous, intraperitoneal or intramuscular injection or infusion; or intracranial, e.g., intrathecal or intraventricular, administration.
  • compositions and formulations for topical administration may include transdermal patches, ointments, lotions, creams, gels, drops, suppositories, sprays, liquids and powders.
  • Conventional pharmaceutical carriers, aqueous, powder or oily bases, thickeners and the like may be necessary or desirable.
  • compositions and formulations for oral administration include powders or granules, suspensions or solutions in water or non-aqueous media, capsules, sachets or tablets. Thickeners, flavoring agents, diluents, emulsifiers, dispersing aids or binders may be desirable.
  • compositions and formulations for parenteral, intrathecal or intraventricular administration may include sterile aqueous solutions that may also contain buffers, diluents and other suitable additives such as, but not limited to, penetration enhancers, carrier compounds and other pharmaceutically acceptable carriers or excipients.
  • compositions of the present invention include, but are not limited to, solutions, emulsions, and liposome-containing formulations. These compositions may be generated from a variety of components that include, but are not limited to, preformed liquids, self-emulsifying solids and self-emulsifying semisolids.
  • the pharmaceutical formulations of the present invention may be prepared according to conventional techniques well known in the pharmaceutical industry. Such techniques include the step of bringing into association the active ingredients with the pharmaceutical carrier(s) or excipient(s). In general the formulations are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product.
  • compositions of the present invention may be formulated into any of many possible dosage forms such as, but not limited to, tablets, capsules, liquid syrups, soft gels, suppositories, and enemas.
  • the compositions of the present invention may also be formulated as suspensions in aqueous, non-aqueous or mixed media.
  • Aqueous suspensions may further contain substances that increase the viscosity of the suspension including, for example, sodium carboxymethylcellulose, sorbitol and/or dextran.
  • the suspension may also contain stabilizers.
  • the pharmaceutical compositions may be formulated and used as foams.
  • Pharmaceutical foams include formulations such as, but not limited to, emulsions, microemulsions, creams, jellies and liposomes. While basically similar in nature these formulations vary in the components and the consistency of the final product.
  • compositions of the present invention may additionally contain other adjunct components conventionally found in pharmaceutical compositions.
  • the compositions may contain additional, compatible, pharmaceutically-active materials such as, for example, antipruritics, astringents, local anesthetics or anti-inflammatory agents, or may contain additional materials useful in physically formulating various dosage forms of the compositions of the present invention, such as dyes, flavoring agents, preservatives, antioxidants, opacifiers, thickening agents and stabilizers.
  • additional materials useful in physically formulating various dosage forms of the compositions of the present invention such as dyes, flavoring agents, preservatives, antioxidants, opacifiers, thickening agents and stabilizers.
  • such materials when added, should not unduly interfere with the biological activities of the components of the compositions of the present invention.
  • the formulations can be sterilized and, if desired, mixed with auxiliary agents, e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings and/or aromatic substances and the like which do not deleteriously interact with the nucleic acid(s) of the formulation.
  • auxiliary agents e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings and/or aromatic substances and the like which do not deleteriously interact with the nucleic acid(s) of the formulation.
  • Dosing is dependent on severity and responsiveness of the disease state to be treated, with the course of treatment lasting from several days to several months, or until a cure is effected or a diminution of the disease state is achieved.
  • Optimal dosing schedules can be calculated from measurements of drug accumulation in the body of the patient. The administering physician can easily determine optimum dosages, dosing methodologies and repetition rates.
  • Optimum dosages may vary depending on the relative potency of individual peptide combinations, and can generally be estimated based on EC 50 s found to be effective in in vitro and in vivo animal models or based on the examples described herein. In general, dosage is from 0.01 ⁇ g to 100 g per kg of body weight, and may be given once or more daily, weekly, monthly or yearly.
  • the treating physician can estimate repetition rates for dosing based on measured residence times and concentrations of the drug in bodily fluids or tissues. Following successful treatment, it may be desirable to have the subject undergo maintenance therapy to prevent the recurrence of the disease state, wherein the peptide combination is administered in maintenance doses, ranging from 0.01 ⁇ g to 100 g per kg of body weight, once or more daily, to once every 20 years.
  • the present invention contemplates several drug delivery systems that provide for the administration of a roughly uniform distribution, having controllable rates of release, of a combination of Annexin A2 and tPA.
  • a variety of different media are described below that are useful in creating such drug delivery systems. It is not intended that any one medium or carrier is limiting to the present invention. Note that any medium or carrier may be combined with another medium or carrier; for example, in one embodiment a polymer microparticle carrier attached to a compound may be combined with a gel medium.
  • Carriers or mediums contemplated by this invention comprise a material selected from the group comprising gelatin, collagen, cellulose esters, dextran sulfate, pentosan polysulfate, chitin, saccharides, albumin, fibrin sealants, synthetic polyvinyl pyrrolidone, polyethylene oxide, polypropylene oxide, block polymers of polyethylene oxide and polypropylene oxide, polyethylene glycol, acrylates, acrylamides, methacrylates including, but not limited to, 2-hydroxyethyl methacrylate, poly(ortho esters), cyanoacrylates, gelatin-resorcin-aldehyde type bioadhesives, polyacrylic acid and copolymers and block copolymers thereof.
  • One embodiment of the present invention contemplates a medical device comprising several components including, but not limited to, a reservoir comprising tPA/Annexin A2, a catheter, a sprayer or a tube.
  • said medical device administers either an internal or external spray to a patient.
  • said medical device administers either an internal or external gel to a patient.
  • microparticles comprise liposomes, nanoparticles, microspheres, nanospheres, microcapsules, and nanocapsules.
  • some microparticles contemplated by the present invention comprise poly(lactide-co-glycolide), aliphatic polyesters including, but not limited to, poly-glycolic acid and poly-lactic acid, hyaluronic acid, modified polysacchrides, chitosan, cellulose, dextran, polyurethanes, polyacrylic acids, psuedo-poly(amino acids), polyhydroxybutrate-related copolymers, polyanhydrides, polymethylmethacrylate, poly(ethylene oxide), lecithin and phospholipids.
  • Liposomes capable of attaching and releasing tPA and Annexin A2.
  • Liposomes are microscopic spherical lipid bilayers surrounding an aqueous core that are made from amphiphilic molecules such as phospholipids.
  • a liposome may comprise tPA and Annexin A2 trapped between hydrophobic tails of the phospholipid micelle.
  • water soluble drugs can be entrapped in the core and lipid-soluble drugs can be dissolved in the shell-like bilayer. Liposomes have a special characteristic in that they enable water soluble and water insoluble chemicals to be used together in a medium without the use of surfactants or other emulsifiers.
  • Liposomes form spontaneously by forcefully mixing phospholipids in aqueous media. Water soluble compounds are dissolved in an aqueous solution capable of hydrating phospholipids. Upon formation of the liposomes, therefore, these compounds are trapped within the aqueous liposomal center.
  • the liposome wall being a phospholipid membrane, holds fat soluble materials such as oils. Liposomes provide controlled release of incorporated compounds.
  • liposomes can be coated with water soluble polymers, such as polyethylene glycol to increase the pharmacokinetic half-life.
  • One embodiment of the present invention contemplates an ultra high-shear technology to refine liposome production, resulting in stable, unilamellar (single layer) liposomes having specifically designed structural characteristics. These unique properties of liposomes, allow the simultaneous storage of normally immiscible compounds and the Capability of their controlled release.
  • cationic liposomes comprise negatively-charged materials by mixing the materials and fatty acid liposomal components and allowing them to charge-associate.
  • cationic liposomes include lipofectin, lipofectamine, and lipofectace.
  • liposomes that are capable of controlled release i) are biodegradable and non-toxic; ii) carry both water and oil soluble compounds; iii) solubilize recalcitrant compounds; iv) prevent compound oxidation; v) promote protein stabilization; vi) control hydration; vii) control compound release by variations in bilayer composition such as, but not limited to, fatty acid chain length, fatty acid lipid composition, relative amounts of saturated and unsaturated fatty acids, and physical configuration; viii) have solvent dependency; iv) have pH-dependency and v) have temperature dependency.
  • compositions of liposomes are broadly categorized into two classifications.
  • Conventional liposomes are generally mixtures of stabilized natural lecithin (PC) that may comprise synthetic identical-chain phospholipids that may or may not contain glycolipids.
  • Special liposomes may comprise: i) bipolar fatty acids; ii) the ability to attach antibodies for tissue-targeted therapies; iii) coated with materials such as, but not limited to lipoprotein and carbohydrate; iv) multiple encapsulation and v) emulsion compatibility.
  • Liposomes may be easily made in the laboratory by methods such as, but not limited to, sonication and vibration.
  • compound-delivery liposomes are commercially available.
  • Collaborative Laboratories, Inc. are known to manufacture custom designed liposomes for specific delivery requirements.
  • Microspheres and microcapsules are useful due to their ability to maintain a generally uniform distribution, provide stable controlled compound release and are economical to produce and dispense.
  • an associated delivery gel or the compound-impregnated gel is clear or, alternatively, said gel is colored for easy visualization by medical personnel.
  • microspheres, microcapsules and microparticles i.e., measured in terms of micrometers
  • nanospheres, nanocapsules and nanoparticles i.e., measured in terms of nanometers.
  • nanoparticles i.e., measured in terms of nanometers
  • Microspheres can be obtainable commercially (Prolease®, Alkerme's: Cambridge, Mass.) wherein tPA and Annexin A2 may be homogenized in a suitable solvent and sprayed to manufacture microspheres in the range of 20 to 90 ⁇ m. Techniques are then followed that maintain sustained release integrity during phases of purification, encapsulation and storage. Scott et al., Improving Protein Therapeutics With Sustained Release Formulations, Nature Biotechnology, Volume 16:153-157 (1998).
  • Modification of the microsphere composition by the use of biodegradable polymers can provide an ability to control the rate of tPA/Annexin A2 release.
  • Miller et al. Degradation Rates of Oral Resorbable Implants (Polylactates and Polyglycolates: Rate Modification and Changes in PLA/PGA Copolymer Ratios, J. Biomed. Mater. Res., Vol. II:711-719 (1977).
  • a sustained or controlled release microsphere preparation is prepared using an in-water drying method, where an organic solvent solution of a biodegradable polymer metal salt is first prepared. Subsequently, a dissolved or dispersed medium of tPA/Annexin A2 is added to the biodegradable polymer metal salt solution.
  • the weight ratio of tPA/Annexin A2 to the biodegradable polymer metal salt may for example be about 1:100000 to about 1:1, preferably about 1:20000 to about 1:500 and more preferably about 1:10000 to about 1:500.
  • the organic solvent solution containing the biodegradable polymer metal salt and tPA/Annexin A2 is poured into an aqueous phase to prepare an oil/water emulsion. The solvent in the oil phase is then evaporated off to provide microspheres. Finally, these microspheres are then recovered, washed and lyophilized. Thereafter, the microspheres may be heated under reduced pressure to remove the residual water and organic solvent.
  • phase separation during a gradual addition of a coacervating agent; ii) an in-water drying method or phase separation method, where an antiflocculant is added to prevent particle agglomeration and iii) by a spray-drying method.
  • the present invention contemplates a medium comprising a microsphere or microcapsule capable of delivering a controlled release of a tPA/Annexin A2 composition for a duration of approximately between 1 day and 6 months.
  • Controlled release microcapsules may be produced by using known encapsulation techniques such as centrifugal extrusion, pan coating and air suspension.
  • Microspheres/microcapsules can be engineered to achieve particular release rates.
  • Oliosphere® Macromed
  • These particular microsphere's are available in uniform sizes ranging between 5-500 ⁇ m and composed of biocompatible and biodegradable polymers.
  • Specific polymer compositions of a microsphere may control the drug release rate such that custom-designed microspheres are possible, including effective management of the burst effect.
  • ProMaxx® (Epic Therapeutics, Inc.) is a protein-matrix drug delivery system. The system is aqueous in nature and is adaptable to standard pharmaceutical drug delivery models. In particular, ProMaxx® are bioerodible protein microspheres that deliver both small and macromolecular drugs, and may be customized regarding both microsphere size and desired drug release characteristics.
  • a microsphere or microparticle comprises a pH sensitive encapsulation material that is stable at a pH less than the pH of the internal mesentery.
  • the typical range in the internal mesentery is pH 7.6 to pH 7.2. Consequently, the microcapsules should be maintained at a pH of less than 7.
  • the pH sensitive material can be selected based on the different pH criteria needed for the dissolution of the microcapsules. The encapsulated compound, therefore, will be selected for the pH environment in which dissolution is desired and stored in a pH preselected to maintain stability.
  • lipids comprise the inner coating of the microcapsules.
  • these lipids may be, but are not limited to, partial esters of fatty acids and hexitiol anhydrides, and edible fats such as triglycerides. Lew C. W., Controlled-Release pH Sensitive Capsule And Adhesive System And Method. U.S. Pat. No. 5,364,634 (herein incorporated by reference).
  • a microparticle contemplated by this invention comprises a gelatin, or other polymeric cation having a similar charge density to gelatin (i.e., poly-L-lysine) and is used as a complex to form a primary microparticle.
  • gelatin or other polymeric cation having a similar charge density to gelatin (i.e., poly-L-lysine) and is used as a complex to form a primary microparticle.
  • a primary microparticle is produced as a mixture of the following composition: i) Gelatin (60 bloom, type A from porcine skin), ii) chondroitin 4-sulfate (0.005% -0.1%), iii) glutaraldehyde (25%, grade ‘1), and iv) 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC hydrochloride), and ultra-pure sucrose (Sigma Chemical Co., St. Louis, Mo.).
  • the source of gelatin is not thought to be critical; it can be from bovine, porcine, human, or other animal source.
  • the polymeric cation is between 19,000-30,000 daltons. Chondroitin sulfate is then added to the complex with sodium sulfate, or ethanol as a coacervation agent.
  • a tPA/Annexin A2 composition is directly bound to the surface of the microparticle or is indirectly attached using a “bridge” or “spacer”.
  • the amino groups of the gelatin lysine groups are easily derivatized to provide sites for direct coupling of a composition.
  • spacers i.e., linking molecules and derivatizing moieties on targeting ligands
  • avidin-biotin are also useful to indirectly couple targeting ligands to the microparticles.
  • Stability of the microparticle is controlled by the amount of glutaraldehyde-spacer crosslinking induced by the EDC hydrochloride.
  • a controlled release medium is also empirically determined by the final density of glutaraldehyde-spacer crosslinks.
  • kits for the practice of the methods of this invention.
  • the kits preferably include one or more containers containing a vascular disorder treatment method of this invention.
  • the kit can include a medium comprising tPA and Annexin A2.
  • the kit can include a container comprising the medium.
  • the medium can optionally be a liquid.
  • the kit can optionally include medical devices including, but not limited to, injection syringes, intravenous drip bags, intravenous catheters, tubing comprising connector (i.e., for example, Leur Lock connectors) capable of attaching a drip bag to a catheter.
  • the kit can optionally include a pharmaceutically acceptable excipient and/or a drug delivery vehicle (e.g., a liposome).
  • the medium may be provided suspended in the excipient and/or delivery vehicle or may be provided as a separate component which can be later combined with the excipient and/or delivery vehicle.
  • kits may also optionally include appropriate systems (e.g. opaque containers) or stabilizers (e.g. antioxidants) to prevent degradation of the reagents by light or other adverse conditions.
  • appropriate systems e.g. opaque containers
  • stabilizers e.g. antioxidants
  • kits may optionally include instructional materials containing directions (i.e., protocols) providing for the use of the mediums for treatment of vascular disorders.
  • the disorders can include, but are not limited to, stroke, myocardial infarction, deep vein thrombosis, or pulmonary embolism.
  • instructional materials typically comprise written or printed materials they are not limited to such. Any material capable of storing such instructions and communicating them to an end user is contemplated by this invention. Such material include, but are not limited to electronic storage material (e.g., magnetic discs, tapes, cartridges, chips), optical material (e.g., CD ROM), and the like. Such material may include addresses to internet sites that provide such instructional materials.
  • Histidine-tagged recombinant Annexin A2 was produced in E. coli. using a known bacterial expression vector containing full-length human Annexin A2 cDNA (20). Briefly, A full-length human Annexin II cDNA (See, FIG. 5B ) was inserted into a histidine-tagged bacterial expression vector, pQE (purchased from Qiagen), between the KpnI and SalI sites (pQE-A2). A mutant A2 (mA2) expression vector that lacks 224 amino acids of the C-terminal was created as follows.
  • a stop codon was introduced at 114 amino acids using an oligonucleotide-based mutagenesis kit (Quick Change, Clontech) and two complement oligonucleotides 5′-GCTTCTGAGCTATA-GGCTTCCATGAAG-3′ and 5′-CTTCATGGAAGCCTATAGCT-CAGAAGC-3′ according to the manufacturer's instructions.
  • the mutated sequence was confirmed by DNA sequencing.
  • a cell lysate prepared from overnight cultures of pQE-AN II-transformed JM109 was pelleted (8000 rpm, 15 minutes), resuspended in a buffer containing 50 mmol/L sodium phosphate and 300 mmol/L NaCl (pH8), and sonicated for 2 minutes.
  • the lysate was purified using a nickel-affinity column (His-Trap, purchased from Amersham Pharmacia). Protein purity of rA2 protein was confirmed by SDS-PAGE followed by Coomassie blue staining, and its identity was verified by Western Blot analysis.
  • Focal embolic strokes were induced as previously described (2), except that two blood clots, 20-mm in length, were infused.
  • human recombinant tPA (Activase®, Genentech Inc, San Francisco, Calif., U.S.A.) and/or rA2 protein were administered intravenously. An initial 10% bolus dose was followed by continuous infusion of the remaining drug over a 30 minute period.
  • the relatively high dose of tPA was chosen based on the approximately ten-fold difference in fibrin-specific enzyme activity between human and rodent systems (22). Only animals surviving 24 h after stroke were included in the study. Numbers of dead animals were counted in calculating the overall mortality rate for each group of animals. All drug treatments were performed by an investigator blinded to the surgical groups.
  • rCBF Regional cerebral blood flow
  • LDF laser-doppler flowmetry
  • the volume of intracerebral hemorrhage was quantified using a previously described spectrophotometric hemoglobin assay (2).
  • Plasma samples were collected, anticoagulated upon addition of one-ninth volume of 3.2% (0.109M) trisodium citrate, and centrifuged at 400 g for 15 min. Plasma samples were stored at ⁇ 80° C. until assayed. D-dimer was assayed on plasma samples diluted 1:4 in an ELISA kit (American Diagnostica Inc, Stamford, Conn., USA) according to the manufacturer's instructions.

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2015089268A1 (fr) 2013-12-11 2015-06-18 University Of Massachusetts Compositions et procédés de traitement d'une maladie au moyen d'une protéine effectrice t3ss de salmonella (sipa)
US9314500B2 (en) 2011-10-12 2016-04-19 The General Hospital Corporation Non-N-glycosylated recombinant human annexin A2

Families Citing this family (15)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US10213187B1 (en) 2012-01-25 2019-02-26 Mubin I. Syed Method and apparatus for percutaneous superficial temporal artery access for carotid artery stenting
WO2014081947A1 (fr) 2012-11-21 2014-05-30 Syed Mubin I Système pour le placement intravasculaire d'un dispositif médical
US9636244B2 (en) 2015-04-09 2017-05-02 Mubin I. Syed Apparatus and method for proximal to distal stent deployment
US10327929B2 (en) 2015-10-30 2019-06-25 Ram Medical Innovations, Llc Apparatus and method for stabilization of procedural catheter in tortuous vessels
US9980838B2 (en) 2015-10-30 2018-05-29 Ram Medical Innovations Llc Apparatus and method for a bifurcated catheter for use in hostile aortic arches
US10779976B2 (en) 2015-10-30 2020-09-22 Ram Medical Innovations, Llc Apparatus and method for stabilization of procedural catheter in tortuous vessels
US10492936B2 (en) 2015-10-30 2019-12-03 Ram Medical Innovations, Llc Apparatus and method for improved access of procedural catheter in tortuous vessels
US11020256B2 (en) 2015-10-30 2021-06-01 Ram Medical Innovations, Inc. Bifurcated “Y” anchor support for coronary interventions
US10173031B2 (en) 2016-06-20 2019-01-08 Mubin I. Syed Interchangeable flush/selective catheter
WO2019055589A1 (fr) * 2017-09-13 2019-03-21 The Regents Of The University Of Michigan Inhibition de la protéine kinase c pour prolonger le traitement de l'activateur tissulaire du plasminogène pour une maladie ischémique
US11007075B2 (en) 2018-02-18 2021-05-18 Ram Medical Innovations, Inc. Vascular access devices and methods for lower limb interventions
US20230190657A1 (en) * 2020-05-22 2023-06-22 Trustees Of Boston University Methods and compositions for treating a fibrotic disease
JP2023538673A (ja) * 2020-08-24 2023-09-08 アブレキサ ファーマスーティカルズ，インコーポレイテッド 出血および血管系不安定症の予防および処置用化合物
US12171885B2 (en) * 2021-07-15 2024-12-24 University Of Maryland, Baltimore County Hemostatic nanocapsules for stopping bleeding, visualizing injury, and delivering drugs
US20250242055A1 (en) * 2022-04-11 2025-07-31 The Board Of Trustees Of The Leland Stanford Junior University Neuroprotection and Axon Regeneration Therapies for CNS Axonopathies by Modulating Membrane Structure, Cytoskeleton and Signaling Molecules

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20020155160A1 (en) * 1999-06-10 2002-10-24 Leach Jonathan K. Microencapsulated plasminogen activators

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5364634A (en) 1991-11-08 1994-11-15 Southwest Research Institute Controlled-release PH sensitive capsule and adhesive system and method
US7419678B2 (en) 2000-05-12 2008-09-02 Cordis Corporation Coated medical devices for the prevention and treatment of vascular disease
US6776796B2 (en) 2000-05-12 2004-08-17 Cordis Corportation Antiinflammatory drug and delivery device
EP1839670A3 (fr) 2001-02-21 2009-11-11 Alavita Pharmaceuticals, Inc. Protéines d'annexine modifiées et leur utilisation contre la thrombose
US7115408B1 (en) 2001-05-02 2006-10-03 Ista Pharmaceuticals, Inc. Method of stabilizing hyaluronidase with annexin II
AU2002357017A1 (en) 2001-11-28 2003-06-10 Mediomics, Llc Anti-angiogenesis methods, compositions and uses therefor

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20020155160A1 (en) * 1999-06-10 2002-10-24 Leach Jonathan K. Microencapsulated plasminogen activators

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
Burggraf et al., Eur J Neurosci. 2004; 20: 2903-8. *
Clark et al., JAMA, 1999; 282: 2019-2026. *
Ishii et al., Circ Res., 2001; 89: 1240-1245. *
Kimata et al., Chem Pharm. Bull., 1991; 39: 3327-3330. *
Roda et al., JBC, 2003; 278: 5702-5709. *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9314500B2 (en) 2011-10-12 2016-04-19 The General Hospital Corporation Non-N-glycosylated recombinant human annexin A2
WO2015089268A1 (fr) 2013-12-11 2015-06-18 University Of Massachusetts Compositions et procédés de traitement d'une maladie au moyen d'une protéine effectrice t3ss de salmonella (sipa)
US10201589B2 (en) 2013-12-11 2019-02-12 University Of Massachusetts Compositions and methods for treating disease using Salmonella T3SS effector protein (SipA)
US10835575B2 (en) 2013-12-11 2020-11-17 University Of Massachusetts Compositions and methods for treating disease using Salmonella T3SS effector protein (SipA)
EP3779446A1 (fr) 2013-12-11 2021-02-17 University of Massachusetts Compositions et procédés de traitement d'une maladie au moyen d'une protéine effectrice t3ss de salmonella (sipa)

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