What Is Urokinase?

This product is an enzyme protein isolated from healthy human urine or obtained from human kidney tissue culture. It consists of two parts with molecular weights of 33000 (LMW-tcu-PA) and 54000 (HMW-tcu-PA). This product directly acts on the endogenous fibrinolytic system and can catalyze the lysis of plasminogen into plasmin, which can not only degrade fibrin clots, but also degrade fibrinogen, coagulation factor and coagulation in the blood circulation Factor and so on, thus exerting a thrombolytic effect. This product has fast onset and good effect on newly formed thrombus. This product can also increase vascular ADP enzyme activity, inhibit ADP-induced platelet aggregation, and prevent thrombosis. After intravenous infusion of this product, the patient's plasmin activity significantly increased; after a few hours of discontinuation, the plasmin activity returned to its original level.

Chinese name: Urokinase
English name: Urokinase
nickname: Adrenaline, Urea, Human Plasmin, Human Plasmin
Chemical formula: C21H25BrN2O3

Molecular weight: 433.35
CAS Registry Number: 9039-53-6
EINECS registration number: 232-917-9
Exterior: White powder
Application: Thrombolytic therapy for thromboembolic diseases

Basic information of urokinase

Urokinase Chinese name: urokinase

Chinese alias: akinin; urin; human plasmin; human plasmin; urokinase 6000; temploxin; power urokinase; akinin; urate oxidase

English name: Urokinase

English alias: Urokinase [USAN: INN: BAN: JAN]; Actosolv; Breokinase; EC 3.4.21.31; EC 3.4.21.73; EC 3.4.99.26; Kinase (enzyme-activating), uro-urokinase; Kinlytic; Plasminokinase, urinary; TCUK; Tissue culture urokinase; Two-chain urokinase; UK; UNII-83G67E21XI; Ukidan; Urochinasi; Urochinasi [DCIT]; Urokinasum; Urokinasum [INN-Latin]; Uroquinasa; Uroquinasa [INN-Spanish]; WIN 22005; Win-Kinase ; Kinase (enzyme-activating), uro-; Abbokinase; Breokinase; Purochin; Ukidan; Urokinasum

CAS: 9039-53-6

EINECS: 232-917-9

Molecular formula: C ₂₁ H ₂₅ BrN ₂ O ₃

Molecular weight: 433.35

Properties: This product is white powder

Urokinase Pharmacopoeia Standard

Urokinase source content

This strain is an enzyme that can activate plasminogen from fresh human urine. It is a mixture of high molecular weight urokinase (Mw 54000) and low molecular weight urokinase (Mw 33000). The content of high molecular weight urokinase should not be less than 90%, and the activity of urokinase per 1mg of protein should not be less than 120,000 units.

Urokinase method requirements

This product should be extracted from the urine of healthy people, and the production process should meet the requirements of the current edition of the Good Manufacturing Practice for Drugs. This product needs to be heated at 60 ° C for 10 hours during the production process to inactivate the virus.

Urokinase traits

This product is white or off-white powder.

Urokinase identification

Take the solution of the test product under the titer measurement, dilute it with a barbiturate-sodium chloride buffer solution (pH 7.8) to a solution containing 20 units per 1 ml, draw 1 ml, add 0.3 ml of bovine fibrinogen solution, and then Add 0.2ml of bovine plasminogen solution and 0.2ml of bovine thrombin solution in order, shake them quickly, immediately incubate in a constant temperature water bath at 37 ° C ± 0.5 ° C, and immediately count the time. It should coagulate within 30 to 45 seconds, and the clot redissolves within 15 minutes. A 0.9% sodium chloride solution was used as a blank, and the same operation was performed, and the clot was insoluble within 2 hours (reagent preparation was determined by titer).

Urokinase test

1 Clarity and color of the solution

Take this product, add 0.9% sodium chloride solution to make a solution containing 3000 units per 1ml, and check it according to law (Appendix A and B of Part Two of the 2010 Pharmacopoeia). It should be clear and colorless.

2 Molecular composition ratio

Take this product, add water to dissolve and make a solution containing 2mg per 1ml, then add an equal volume of buffer solution (take 2.5ml of concentrated gel buffer, 2.5ml of 20% sodium lauryl sulfate solution, 0.1% bromophenol blue 1.0ml of solution and 3.5ml of 87% glycerol solution, add water to 10ml), place in a water bath for 3 minutes, let cool, as the test solution; take 10l of the test solution, add it to the sample well, according to the electrophoresis method (2010 edition Pharmacopoeia (Appendix F, Coomassie Brilliant Blue Staining of the fifth method), the relative content (%) of high molecular weight urokinase was calculated by the following formula.

3 Loss on drying

Take this product, use phosphorus pentoxide as a desiccant, and dry under reduced pressure to constant weight at 60 ° C, and the weight loss should not be greater than 5.0% (Appendix L of the second edition of the Pharmacopoeia of 2010 Edition).

4 Hepatitis B surface antigen

Take this product and add 0.9% sodium chloride solution to make a solution containing 10mg per 1ml. It should be negative according to the instructions in the kit instructions.

5 Abnormal toxicity

Take this product, add sodium chloride injection to make a solution containing 5000 units per 1ml, check according to law (Appendix C of Part Two of the 2010 Pharmacopoeia), and administer it by intravenous injection, which should meet the requirements.

6 Bacterial endotoxin

Take this product and check it according to the law (Appendix E of Part Two of the Pharmacopoeia 2010), the amount of endotoxin per 10,000 units of urokinase should be less than 1.0EU.

7 Coagulation-like active substances

(1) Preparation of blood plasma Take fresh rabbit blood, add 3.8% sodium citrate solution (add 3.8% sodium citrate solution 1ml for every 9ml rabbit blood), mix well, at 2 8 , 5000 per minute Centrifuge for 20 minutes. The supernatant was frozen and stored at -20 ° C for future use, and thawed at 25 ° C before use.

(2) Determination method: Take this product, dissolve and dilute gababitur buffer (pH 7.4) to make a test solution containing 5000 units, 2500 units, 1250 units, 625 units, and 312 units per 1ml. If the test product contains ethylenediamine tetraacetate or phosphate, it must be removed by dialysis at 2 ° C with barbitur's buffer (pH 7.4), and then prepared into a solution with the above concentration.

Take 7 small test tubes (12mm × 75mm), add 0.1ml of barbitur's buffer (pH 7.4) to the first tube and the 7th tube as a blank control, and add the above-mentioned diluted test solution to the remaining 5 tubes 0.1ml each, and then add 6-aminocaproic acid solution [take 1.97g of 6-aminocaproic acid, add barbiturate buffer (pH 7.4) to dissolve, and dilute to 50ml] and 0.1ml each of plasma, gently shake Evenly, place in a 25 ° C water bath for 3 minutes, add 0.1ml of calcium chloride solution (taken 1.84g of calcium chloride, add water to dissolve and dilute to 500ml), and mix into the water bath. And immediately time it. Pay attention to observe the coagulation of the plasma. The end point is judged to gently tilt the test tube to set it horizontally. The solution is inclined but does not flow. Record the coagulation time (seconds). Each concentration is measured 3 times and the average value is obtained (the difference between the maximum value and the minimum value in the 3 determinations shall not exceed 10% of the average value). Take the logarithm of the solution concentration of the test sample as the ordinate, and shorten the time for recalcification (the solidification time of the blank tube minus the coagulation time of the test tube) as the abscissa. Connect the points of the test product with different dilutions, which should form a straight line. The intersection of the extension line and the ordinate axis is the concentration of the test product, that is, the enzyme activity of the test product when the coagulation-like activity is zero. The unit of product solution indicates that each 1ml should not be less than 150 units.

Urokinase titer

1 Enzyme activity

(1) Reagent Bovine Fibrinogen Solution Take bovine fibrinogen and gababital sodium chloride buffer solution (pH 7.8) to make a solution containing 6.67 mg of coagulable protein per 1 ml.

Bovine thrombin solution Take bovine thrombin and gababital sodium chloride buffer solution (pH 7.8) to make a solution containing 6.0 units per 1 ml.

Bovine plasminogen solution Take bovine plasminogen, add trimethylolaminomethane buffer (pH 9.0) to make a solution containing 1 to 1.4 casein units per 1ml (if the solution is turbid, centrifuge, take Supernatant is ready for use).

Just before use, the same volume of bovine thrombin solution and bovine plasminogen solution were taken and mixed.

(2) Preparation of the standard solution Take the urokinase standard solution, gababital-sodium chloride buffer solution (pH 7.8) and dissolve it quantitatively to make a solution containing 60 units per 1ml.

(3) Preparation of test product solution Take an appropriate amount of this product, dissolve in gababitur-sodium chloride buffer (pH 7.8), mix well, and quantitatively dilute to the same concentration as the standard solution.

(4) Assay method: Take 4 test tubes, each add 0.3ml of bovine fibrinogen solution, put in 37 ± 0.5 water bath, and add barbitur-sodium chloride buffer solution (pH 7.8) 0.9ml, 0.8ml, 0.7ml, 0.6ml, add the standard solution 0.1ml, 0.2ml, 0.3ml, 0.4ml in order, and then add 0.4ml of the mixed solution, shake immediately, and time separately. The reaction system should coagulate within 30 to 40 seconds. When the small bubbles in the clot rise to half the volume of the reaction system, it is regarded as the end point of the reaction, and the time is immediately recorded. Each concentration is measured 3 times and the average value is obtained (the difference between the maximum value and the minimum value in the 3 determinations shall not exceed 10% of the average value). Linear regression was performed with the logarithm of the urokinase concentration as the abscissa and the logarithm of the reaction end time as the ordinate. The test article was measured according to the above method, and the titer was obtained using a linear regression equation, and the titer (unit) of the test article per 1 mg was calculated.

2 protein content

Take about 10mg of this product, weigh it accurately, and measure it according to the protein content determination method (Appendix M of the Pharmacopoeia Part II of the 2010 edition, M first method).

3 specific activity

Each 1mg protein contains the number of urokinase activity units ^[1] .

Urokinase category

Thrombolytic drugs.

Urokinase storage

Shaded, sealed and stored below 10 ° C.

Urokinase preparation

Urokinase for injection

Urokinase drug description

Urokinase classification

Circulatory Drugs> Antithrombotic Drugs> Thrombolytic Drugs

Urokinase dosage form

Injection (powder injection): 500U, 1,000U, 5,000U, 10,000U, 20,000U, 50,000U, 100,000U, 200,000U, 250,000U, 500,000U, 1.5 million U, 2.5 million U .

Pharmacodynamics of urokinase

The product directly acts on the endogenous fibrinolytic system and can catalyze the lysis of plasminogen into plasmin,

Urokinase The latter can not only degrade fibrin clot, but also degrade fibrinogen, coagulation factor and coagulation factor in the blood circulation, thus exerting thrombolytic effect. The product has fast onset and good effect on newly formed thrombus. The product can also increase vascular ADP enzyme activity, inhibit ADP-induced platelet aggregation, and prevent thrombosis. After intravenous infusion of the product, the plasmin activity in the patient was significantly increased; after a few hours of discontinuation, the plasmin activity recovered to the original level, but the plasma fibrin or fibrinogen level decreased and their degradation products increased Can last 12-24 hours. This product shows a clear correlation between the thrombolytic effect and the dose of the drug and the time window of administration. The product is very toxic, and the mouse has a lethal dose of more than 1 million IU / kg body weight. There is no obvious antigenicity, teratogenicity, carcinogenicity and mutagenicity. There are few reports of allergic reactions in clinical applications; however, in view of the increased plasmin activity of this product and the reduction of unbound plasminogen and plasminogen bound to fibrin in the blood circulation, there may be a serious risk of bleeding.

Urokinase pharmacokinetics

After intravenous injection of urokinase, the activity of plasmin increased rapidly, peaked at 15 minutes, and still increased after 6 hours; fibrinogen decreased to about 1000 mg / L, and slowly returned to normal after 24 hours. Urokinase is metabolized in the liver, and its half-life in vivo is about 20 minutes. The half-life of patients with liver dysfunction has been extended. A small amount of drug is excreted with bile and urine ^[2] .

Urokinase indication

This product is mainly used for thrombolytic therapy of thromboembolic diseases. Including acute extensive pulmonary embolism, chest pain 6-12

Urokinase Coronary arterial embolism and myocardial infarction within hours, acute cerebrovascular embolism, retinal arterial embolism, and other peripheral arterial embolism symptoms of skeletal-femoral vein thrombosis who are shorter than 3-6 hours. It is also used to prevent thrombosis after artificial heart valve surgery, to keep the vascular cannula and the drainage of the thoracic and pericardial drainage tubes open. The efficacy of thrombolysis should be maintained by subsequent heparin anticoagulation.

This product is an enzyme thrombolytic drug, which can activate the body to convert plasminogen to plasmin, thereby hydrolyzing fibrin and dissolving freshly formed thrombus. For acute myocardial infarction, acute cerebral thrombosis and cerebrovascular embolism, arteriovenous thrombosis around the limb, central retinal arteriovenous thrombosis and other fresh thrombo-occlusive diseases. This product has no obvious effect on old thrombus. It is also suitable for the treatment of fresh thromboembolic diseases such as cerebral thrombosis, peripheral vascular embolism, central retinal vascular embolism, acute myocardial infarction, and the occurrence of thrombosis in kidney transplantation and orthopedic surgery.

Urokinase dosage

Intravenous bolus or intravenous drip, 40,000 to 60,000 u per day, dissolved in 20 to 40 ml of physiological saline, 1 or 2 to 3 boluses; or dissolved in 5% glucose physiological saline or low molecular dextran 250 ml . Usually 7 to 10 days for a course of treatment, or increase or decrease as appropriate.

Urokinase 1. Cerebrovascular disease: within 6 hours to 6 days after the onset of stroke symptoms of acute cerebral thrombosis, 60,000 u intravenous injection or infusion.

2. Acute venous thrombosis: the first dose can be 60,000 to 180,000 u per day, and later changed to 60,000 u, twice a day for 7 to 10 days.

3. For acute arterial embolization: 60,000 u is injected, and 60,000 u is continued after operation, twice a day, for 5 to 7 days.

4. Acute myocardial infarction: 500,000 u dissolved in 25% glucose solution 20ml intravenous injection, and then 500,000 u added to 5% glucose solution 500ml intravenous injection.

5. Ophthalmological application: 10,000 to 20,000 u intravenous drip or bolus injection, or 200 to 500 u dissolved in 0.5ml of water for injection for subconjunctival or post-ball injection.

6. Coronary arterial infusion: 200,000 1 million units dissolved in sodium chloride injection or 5% glucose injection in 20-60ml intracoronary infusion, input at the rate of 12,000 units per minute, the dose can be Adjust according to the patient's weight, physical condition and thrombolytic effect.

Use of urokinase with caution

Disabled in the following cases:

14 days of active bleeding (gastric and duodenal ulcers, hemoptysis, hemorrhoids, bleeding

Urokinase Etc.), had surgery, biopsy, cardiopulmonary resuscitation (extracorporeal cardiac massage, intracardiac injection, tracheal intubation), vascular puncture in areas where compression could not be performed, and history of trauma;

Control of unsatisfactory hypertension (blood pressure> 21.3 / 14.7kPa) or those who cannot exclude aortic dissection aneurysm;

People with a history of hemorrhagic stroke (including transient ischemic attack); Shock that does not respond to volume expansion and vasopressor;

Pregnancy, bacterial endocarditis, mitral valve disease, atrial fibrillation, and highly suspected thrombosis in the left ventricle;

Diabetic patients with retinopathy;

bleeding disease or bleeding tendency, severe liver and kidney dysfunction and progressive disease;

Patients with unconsciousness.

Patients with severe liver dysfunction, hypofibrinogenemia and hemorrhagic quality should not be used.

Patients with severe liver dysfunction and severe hypertension, hypofibrinogenemia, and patients with bleeding disorders are contraindicated.

Elderly and severe atherosclerosis should be used with caution.

Urokinase adverse reactions

(1) When using a large dose, a small number of patients may have bleeding, mild bleeding such as skin, mucous membranes, naked eyes

Urokinase And under the microscope hematuria, blood sputum or a small amount of hemoptysis, vomiting blood, etc., take appropriate measures to relieve symptoms. If severe bleeding occurs, such as massive hemoptysis or major gastrointestinal bleeding, retroperitoneal bleeding, intracranial, spinal cord, mediastinal, or pericardial bleeding, etc., the use should be discontinued. Blood loss can be transfused with whole blood (preferably fresh blood, not plasma) Effective control can be obtained, and aminocaproic acid and aminotoluic acid can be considered to combat urokinase in emergency situations.

(2) A small number of patients may have allergic reactions: generally mild, such as bronchospasm and rash. Occasionally seen anaphylactic shock.

(3) Fever: About 2 to 3% of patients can see different degrees of fever. Paracetamol can be used as an antipyretic. Do not use aspirin or other anti-platelet antipyretics.

(4) Others: Nausea, vomiting, loss of appetite, fatigue, and elevated ALT may still be seen. Can cause bleeding, a few have allergic reactions, headache, nausea, vomiting, loss of appetite, etc. should be discontinued immediately.

Urokinase considerations

1. (1) Alanine aminotransferase (ALT) may be elevated; (2) Some patients may have a moderate decrease in hematocrit, but there is no bleeding.

2. Bleeding time, partial thromboplastin generation time, prothrombin time, thrombin time, platelet count, hemoglobin, hematocrit, etc. should be tested before use to exclude bleeding constitution.

3. Urokinase is easy to decompose and degrade in acidic liquid, so the diluent used should be close to neutral. When diluting with a glucose injection, a product with a pH greater than or equal to 4.5 should be selected.

4. Sufficient initial amount of urokinase must be given within a short time (15-30min) to neutralize urokinase antibodies in the body. However, an excessively large initial amount can deplete the plasminogen pool and coagulation factors , , and in the body and affect the thrombolytic effect.

5. Urokinase is only for intravenous and intracardiac injection, not intramuscular or local injection.

6. The urokinase solution must be prepared fresh immediately before use and used with the preparation. Urokinase is dissolved with 5ml of sterile water for injection (cannot be dissolved with other solutions), and the prepared solution can be light straw yellow (not applicable if the color is deep or cannot be completely dissolved). When dissolving, the bottle should be rotated gently. Do not shake it vigorously (because insoluble materials can be generated). The prepared liquid should pass a 0.45 m terminal filter or a small celluloid filter to remove insoluble particles. Ask for dilution.

7. The dissolved solution is easily inactivated, and the unused solution should be discarded, and it should not be stored and reused.

8. After thrombolytic therapy, antiplatelet and anticoagulant drugs must be given to inhibit the tendency of potential thrombotic recurrence. Therefore, at the beginning of thrombolytic therapy, urokinase should be combined with low-dose aspirin (160mg). After thrombolysis, continue to use aspirin for 1 month to reduce the mortality rate in the acute phase and 15 months, but bleeding Inclinations will increase slightly.

9. When using urokinase and sending out blood, mild bleeding can be relieved by taking corresponding measures; severe bleeding should be discontinued immediately, and whole blood transfusion (preferably fresh blood, not plasma replacement) can be effectively controlled during blood loss, under emergency conditions Consider using aminocaproic acid and aminotoluic acid to fight the effect of urokinase.

10. If fever occurs during medication, acetaminophen can be used for symptomatic antipyretic treatment, but aspirin or other antipyretic drugs with antiplatelet effect can not be used.

11. In the process of thrombolysis, if the thrombus is not completely dissolved, the thrombolysis should be continued to dissolve the thrombus.

12. Urokinase can cause needle hole bleeding at the injection site, and it is generally not appropriate to perform puncture and other operations during the medication.

13. The dissolution of pulmonary embolism is often accompanied by hemodynamic changes, and care should be taken to maintain blood pressure.

14. Thrombolytic therapy for deep venous thrombosis can accelerate thrombolysis and promote rapid blood flow reconstruction. In theory, it is a better treatment than anticoagulation. High, so thrombolytic therapy is not a routine treatment for patients with deep vein thrombosis. In recent years, patients with severe venous thrombosis can also be directly inserted into the emboli through a catheter, and a large dose of urokinase is used for local thrombolysis. Renal veins, superior vena cava veins, subclavian veins, and intracranial veins (including sigmoid sinus veins) thrombi, which were inserted into the thrombus site with a catheter and thrombolytic with urokinase have all been successfully reported. However, care must be taken in the treatment of intracranial venous thrombosis. Because of the presence of venous valves in the lower extremity veins, local infusion of thrombus in the lower extremities is often limited. Patients with chronic abdominal venous thrombosis (such as superior vena cava or hepatic vein thrombosis), after infusion of thrombolytic drugs through the hepatic vein, the drugs are inactivated in the liver, so the efficacy is poor. But there are also a few successful reports.

15. Arterial thrombolysis combined with surgical treatment can reduce the scope of surgery, so thrombolysis should be done in time. And inserting a porous catheter for thrombolysis can improve the success rate.

Drugs for pregnant and lactating women: Animal experiments have shown that the product is 1,000 times the amount used in humans without harming the reproductive capacity of the female mice and rats and the fetus. No carcinogenicity has been reported for long-term use. There have been no reports of medications in pregnant women with a strict control group. Therefore, unless the product is urgently needed, it is not used by pregnant women. Whether the product can be excreted from milk has not been reported. Therefore, lactating women use this product with caution.

Medication for children: The safety and effectiveness of this product in children have not been reported.

Medication for elderly patients: The safety and effectiveness of this product in elderly patients have not been reported. However, use it with caution if you are over 70 years old.

Urokinase overdose

This product is usually given intravenously with a significant effect of 2500 units / fraction. The total adult medication should not exceed 3 million units. Thrombolytic effect is necessarily accompanied by a certain risk of bleeding. Once bleeding occurs, the drug should be stopped immediately, and fresh whole blood should be replenished according to the bleeding and blood loss. Fibrinogen plasma levels <100mg / dl with bleeding tendency should be supplemented with fresh frozen plasma or cryoprecipitate. It is not appropriate to use dextran hydroxyl Ethyl starch. The rescue effect of aminocaproic acid has not been reported, but it can be used in emergency situations ^[3] .

Urokinase drug interactions

When urokinase is used in combination with heparin, the former activity is inhibited, and alternate administration can be taken every 2 to 3 hours to avoid ^[2] .

Urokinase poisoning

Clinical manifestation

Mainly manifested as bleeding, which may include venipuncture sites, wounds and nosebleeds, hematuria, and sometimes skin or mucous membrane bleeding. Severe visceral bleeding, especially intracranial bleeding, can be life threatening. May have allergic reactions, such as fever, headache, nausea, vomiting, facial flushing, eyelid edema, urticaria, etc. In severe cases, anaphylactic shock occurs. Arrhythmias may occur during coronary perfusion, such as accelerated ventricular autonomic rhythm and frequent pre-ventricular contractions.

Laboratory inspection

Blood clotting time, prothrombin time, bleeding time and other indicators were prolonged.

Key points for diagnosis

1. Medical history with a clear history of medication.

2. Possess clinical manifestations of poisoning such as bleeding.

3. Laboratory inspection support.

Essentials of treatment

1. Patients with bleeding reactions should stop taking the drug immediately.

2. Local bleeding can stop bleeding at local bleeding compression.

3. Severe systemic bleeding can be treated with anti-fibrinolytic drugs such as 6-aminocaproic acid or aminotoluic acid.

4. Transfusion of fresh plasma, cryoprecipitate, fibrinogen and fresh blood if necessary.

5. Anti-allergic treatment.

6. Others, anti-shock, anti-arrhythmia, etc. ^[4] .

Urokinase expert review

Urokinase is a proteolytic enzyme isolated from the urine of healthy humans and can also be prepared from human kidney cell culture without antigenicity. UK can directly activate plasminogen into active plasmin and play a thrombolytic effect. Because activated plasmin can simultaneously degrade fibrinogen in blood, its clinical application is limited. Compared with SK, urokinase has similar therapeutic effect to two drugs. It has obvious curative effect on cerebral embolism, but it is expensive ^[1] .