Thrombin (Ki = 5-39 nM)

Argatroban is a potent and selective synthetic thrombin inhibitor with Ki values against thrombin ranging from 5 nM to 39 nM. In numerous animal models of thrombosis that is both platelet-rich and erythrocyte-rich, argatroban exhibits antithrombotic characteristics. Argatroban inhibits thrombus formation in a dose-dependent manner; in this test, its estimated ED50 was 125 μg/kg. Argatroban increases thrombin time in a dose-dependent manner; at the highest dose used, this increase was 511%, but the APTT increased only by 73%.[1] Argatroban has the ability to directly cause vascular smooth muscle cells to change phenotype, which leads to an increase in the mRNAs for SMemb, PAI-1, and beta-actin.[2]

Argatroban prevents the growth of microthrombi for three hours following middle cerebral artery (MCA) occlusion; after that, it has no further effect. Six hours following MCA occlusion, argatroban also significantly reduces the size of ischemic cerebral lesions. In [3] In the rat model of distal middle cerebral artery occlusion, Argatroban (0.3 mg/h/rat) significantly reduces the number of microthrombi 1 day after distal middle cerebral artery (dMCA) occlusion. One day after distal middle cerebral artery (dMCA) occlusion, a reduction in regional cerebral blood flow (rCBF) is significantly reversed by argatroban (0.1 and 0.3 mg/h/rat). Argatroban (0.3 mg/h/rat) also greatly diminishes the cerebral infarction's size.[4]

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1. The antithrombotic action of Argatroban, a synthetic thrombin inhibitor, was studied in three models of thrombosis in the rat, and in the tail transection bleeding time test. Heparin was studied as a reference anticoagulant. 2. In the model of venous thrombosis induced by thromboplastin followed by stasis of the abdominal vena cava, argatroban had an ED50 of 125 micrograms kg-1, when administered as an i.v. bolus 5 min prior to the thromboplastin injection: the ED50 of heparin was 42 micrograms kg-1, where ED50 is the dose which reduces the weight of the thrombus by 50% compared with that of the control animals. When the two compounds were administered by continuous i.v. infusion, argatroban (ED50 = 1.5 micrograms kg-1 min-1) had the same potency as heparin (ED50 = 1.2 micrograms kg-1 min-1). 3. Argatroban was active in the arterio-venous shunt model with an ED50 of 0.6 mg kg-1 when the compound was given as a bolus. The ED50 of heparin was 0.04 mg kg-1 under the same conditions. The two compounds had ED50 values of 6 micrograms kg-1 min-1 (argatroban) and 3 micrograms kg-1 min-1 (heparin), when administered by continuous i.v. infusion. 4. When tested against occlusive arterial thrombus formation by electrical stimulation of the left carotid artery, both compounds given as either an i.v. bolus or a continuous infusion led to dose-dependent increases in the duration of post-lesion vessel patency. [1]

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Ischemic cerebral infarcts induce hypercoagulation and microthrombosis in the surrounding region, thus leading to vascular occlusion. We determined whether microthrombi contribute to the spreading of ischemic lesions following thrombotic middle cerebral artery (MCA) occlusion and also determined whether Argatroban, a selective thrombin inhibitor, reduces the formation of the microthrombi and the area of the ischemic lesions. The rat left MCA was occluded by a platelet-rich thrombus formed following the photochemical reaction between rose bengal and green light. Microthrombi were histologically identified in the left hemisphere. The extent of ischemic lesions and microthrombi containing fibrin increased in a time-dependent manner after MCA occlusion. Argatroban inhibited the formation of microthrombi up to 3 hr after MCA occlusion; beyond 3 hr, it was ineffective. Argatroban also significantly (P < 0.01) reduced the size of ischemic cerebral lesions at 6 hr after MCA occlusion. It is concluded that the formation of microthrombi contributes to the progression of ischemic lesions in the early stage. It is likely that thrombin generated following thrombotic MCA occlusion contributes to the progression of ischemic lesions by promoting the formation of microthrombi. Argatroban can reduce the formation of microthrombi and ischemic lesions in the early stage. [3]

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To elucidate the role of thrombin in brain damage during focal cerebral ischemia, we investigated the effects of a selective thrombin inhibitor, Argatroban, on microthrombi formation, regional cerebral blood flow (rCBF), infarct areas and neurological deficits using a rat thrombotic distal middle cerebral artery (dMCA) occlusion model. The rat dMCA was occluded by a platelet-rich thrombus formed after photochemical reaction between rose bengal and green light. One day after dMCA occlusion, the number of microthrombi were counted. In the separate animals, rCBF was measured by using the iodoantipyrine method 1 day after dMCA occlusion. Three days after dMCA occlusion, behavioral tests were performed and the size of the cerebral infarction was determined. In the present study, argatroban was administered i.p. by continuous infusion after dMCA occlusion. Argatroban (0.3 mg/h/rat) significantly (P < .05) decreased the number of microthrombi 1 day after dMCA occlusion. Argatroban (0.1 and 0.3 mg/h/rat) significantly (P < .01) reversed a decrease in rCBF 1 day after dMCA occlusion. Argatroban (0.3 mg/h/rat) also significantly (P < .01) reduced the size of the cerebral infarction. Administration of argatroban (0.1 and 0.3 mg/h/rat) resulted in a significant improvement in neurological deficits 3 days after dMCA occlusion (P < .01 and P < .05, respectively). Argatroban decreased the size of the cerebral infarction and improved neurological deficits in the rat thrombotic dMCA occlusion model. These effects were thought to be due to the improvement of rCBF and to the reduction in secondary thrombus formation after dMCA occlusion[4].

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T2DM was associated with cardiac structural and functional disturbances as evidenced from impaired cardiac functional parameters and increased fibrosis. There was a significant increase in PAR expression after 20 weeks of diabetes induction. Four weeks argatroban treatment ameliorated metabolic alterations (reduced plasma glucose and cholesterol), ventricular dysfunctions (improved systolic and diastolic functions), cardiac fibrosis (reduced percentage area of collagen in picro-sirius red staining), and apoptosis (reduced TUNEL positive nuclei). Reduced expression of PAR1 and PAR4 in the argatroban-treated group indicates a response towards inhibition of thrombin. In addition, AKT (Ser-473), GSK-3β (Ser-9), p-65 NFĸB phosphorylation, TGF-β, COX-2, and caspase-3 expression were reduced significantly along with an increase in SERCA expression in argatroban-treated diabetic rats which indicated the anti-fibrotic, anti-inflammatory, and anti-apoptotic potential of argatroban in DCM. Conclusion: This study suggests the ameliorative effects of argatroban in diabetic cardiomyopathy by improving ventricular functions and reducing fibrosis, inflammation, apoptosis, and PAR expression [5].

Effect of argatroban on the expression of myosin heavy-chain isoform mRNA [2]
The effect of specific thrombin inhibitor, Argatroban, on phenotype conversion of cultured vascular smooth muscle cells was tested. The cultured vascular smooth muscle cells were detached by incubating with 0.01% trypsin–EDTA solution for 5 min and replated on 25-cm2 flask with 1.5×105 cells. After the cells were cultured in 10% fetal bovine serum–DMEM for 9 h, the medium was changed to serum-free DMEM containing PDGF-BB (10 and 50 ng/ml) or argatroban (10 and 50 μg/ml), and then incubated for 3 or 24 h. After incubation, total RNA was extracted to perform RT-PCR and the cells were fixed with 10% formalin–PBS for in situ hybridization to detect mRNA expressions of myosin heavy chain isoform.

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Effect of argatroban on the expression of SMemb protein [2]
To measure the protein level of SMemb after stimulation of the cultured vascular smooth muscle cells by Argatroban, the Western blots were performed according to the method by Towbin et al. (1979). Three and twenty-four hours after stimulations, crude myosin extracts were prepared according to the method by Rovner et al. (1986) with a minor modification. In brief, cultured vascular smooth muscle cells (1.5×105 cells) were removed from flasks by Rubber Policeman, rinsed with PBS, and were collected by centrifugation (1600 rpm, 4 °C) for 5 min. 50 μl of extraction buffer (50 mM NaH2PO4, 1 mM EGTA, 0.125 mM α-phenylmethanesulfonyl fluoride, pH 7.0) was added to cell pellet and homogenized by disposable pestles with Mini Cordless Grinder. After centrifugation (10,000 rpm, 4 °C) for 10 min, the pellets were dissolved in Gubba–Straub solution (150 mM NaH2PO4, 300 mM NaCl, 1 mM EGTA, 0.125 mM α-phenylmethanesulfonyl fluoride, 1 mM 2-mercaptoethanol, 10 mM ATP, pH 6.7), and then gently mixed at 4 °C for 1 h. After centrifugation (10,000 rpm, 4 °C) for 10 min, the supernatants were colleted as crude myosin extracts. To separate the proteins in the crude myosin extracts, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) with 8–25% gradient gel was performed using a Phast System. The proteins in the gel were electrically blotted onto polyvinylidene difluoride (PVDF) membrane by PhastTransfer at 15 °C, 20 V, and 25 mA for 90 min. After blotting, the membrane was blocked with 3% non-fat milk in PBS for 2 h at room temperature, and then the membrane was incubated at 4 °C overnight with monoclonal anti-nonmuscle myosin heavy chain, which was diluted to 1:1000 by 3% non-fat milk in PBS. After washing five times with 0.1% Tween 20 in PBS (PBST) for 10 min each, the membrane was incubated with 1:3000 diluted sheep anti-mouse IgG2b in 1% non-fat milk in PBST at room temperature for 1 h. After washing four times by PBST and two times by PBS, substrate solution (0.1 mg/ml 3-amino-9-ethylcarbazole in 0.1 M sodium phosphate buffer) was added. The intensity of colored SMemb protein bands were analyzed by NIH image.

Experimental Design and Pharmacological Interventions [5]
Argatroban is a direct thrombin inhibitor that binds to the catalytic site of thrombin reversibly and thereby inhibits its action. Argatroban was used and the doses (0.3 and 1 mg/kg) were selected based on previous literature. Argatroban was dissolved in dimethyl sulfoxide (DMSO) and administered daily through i.p. route. Similarly, DMSO was administered through i.p. route in the HFD + STZ group and control group to negate the effect of the vehicle. After the 16th week of STZ injection, presence of diabetes was again reconfirmed and animals with plasma glucose ≥250 mg/dl were considered for further study. Argatroban and vehicle were administered at the dose of 1 ml/kg of body weight.

Animals were randomly assigned to the following groups.
1. Normal control group (Ctrl): Animals treated with vehicle dimethyl sulfoxide (DMSO) from the 16th to 20th week for 4 weeks to control animals on normal pellet diet (NPD).
2. High-fat diet + low-dose STZ 35 mg/kg group (T2DM): Vehicle (DMSO) was administered to T2DM animals for 4 weeks.
3.Low dose argatroban-treated diabetic animals group (T2DM + Arg 0.3 mg/kg): Argatroban was administered at the dose of 0.3 mg/kg, i.p. once daily for 4 weeks to T2DM animals.
4. High dose argatroban-treated diabetic animals group (T2DM + Arg 1 mg/kg): Argatroban was administered at the dose of 1 mg/kg, i.p. once daily for 4 weeks to T2DM animals.
5. Argatroban-treated per se group (Ctrl + Arg 1 mg/kg): Argatroban was administered at the dose of 1 mg/kg i.p., once daily for 4 weeks to control animals on NPD diet.

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Argatroban is suitable for constructing animal models to induce endogenous intestinal thrombosis. In hepatectomized rats, the pharmacokinetic parameters of Argatroban, such as area under the curve, volume of distribution, and elimination half-life, are significantly increased‌4. Argatroban is converted from the 21-(R) isomer to the 21-(S) diastereomer in the kidneys, with the latter exhibiting stronger antithrombotic activity and potentially higher hepatic clearance‌.

‌Thrombolysis Induction‌
‌Pathogenic Mechanism‌
Argatroban is a specific thrombin inhibitor that establishes thrombolysis models by inhibiting thrombin activity in animals.
‌Modeling Protocol‌
‌Animal‌: Wistar ST • Male • 8 weeks old
‌Administration‌: 2 mg/kg/h • IV • 1 h
‌Note‌: Allow free access to water and fast overnight before thrombolysis experiments.
‌Success Criteria‌
Relative rate of thrombus dissolution ↓

Absorption, Distribution and Excretion
Bioavailability is 100% (intravenous).
Argatroban is excreted primarily in the feces (65%), presumably through biliary secretion; 22% is eliminated via urine.
174 mL/kg
12.18 L [70-kg adult]
5.1 L/kg/hr [infusion doses up to 40 mcg/kg/min]
Argatroban is excreted primarily in the feces, presumably through biliary secretion. In a study in which (14)C-argatroban (5 ug/kg/min) was infused for 4 hours into healthy subjects, approximately 65% of the radioactivity was recovered in the feces within 6 days of the start of infusion with little or no radioactivity subsequently detected. Approximately 22% of the radioactivity appeared in the urine within 12 hours of the start of infusion. Little or no additional urinary radioactivity was subsequently detected. Average percent recovery of unchanged drug, relative to total dose, was 16% in urine and at least 14% in feces.
It is not known if argatroban crosses the human placenta. the molecular weight (about 527 for the hydrated from), low metabolism, and moderate serum protein binding suggest that exposure of the embryo-fetus should be expected, especially sine the drug is given as a continuous infusion.
Argatroban distributes mainly in the extra cellular fluid as evidenced by an apparent steady-state volume of distribution of 174 mL/kg (12.18 L in a 70 kg adult). Argatroban is 54% bound to human serum proteins, with binding to albumin and a1 - acid glycoprotein being 20% and 34%, respectively.
/MILK/ Argatroban is detected in rat milk.

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Metabolism / Metabolites
Liver via hydroxylation and aromatization of the 3-methyltetrahydroquinoline ring. Age and gender do not substantially affect the pharmacodynamic or pharmacokinetic profile of argatroban.
The main route of argatroban metabolism is hydroxylation and aromatization of the 3-methyltetrahydroquinoline ring in the liver. The formation of each of the 4 known metabolites is catalyzed in vitro by the human liver microsomal cytochrome P450 enzymes CYP3A4/5. The primary metabolite (M1) exerts 3- to 5-fold weaker anticoagulant effects than argatroban. Unchanged argatroban is the major component in plasma. The plasma concentrations of M1 range between 0% and 20% of that of the parent drug. The other metabolites (M2 to M4) are found only in very low quantities in the urine and have not been detected in plasma or feces. These data, together with the lack of effect of erythromycin (a potent CYP3A4/5 inhibitor) on argatroban pharmacokinetics, suggest that CYP3A4/5-mediated metabolism is not an important elimination pathway in vivo.
Argatroban is metabolized principally by the liver via hydroxylation and aromatization of the 3-methyltetrahydroquinoline ring.

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Biological Half-Life
39 and 51 minutes
The terminal elimination half life is 39-51 minutes.

Toxicity Summary
IDENTIFICATION AND USE: Argatroban is antithrombin, platelet aggregation inhibitor. Direct thrombin inhibitors including argatroban are commonly used anticoagulants in patients with known or suspected heparin-induced thrombocytopenia. HUMAN STUDIES: There are two case reports documenting safe use of argatroban during human pregnancy. Argatroban was not genotoxic in the WI-38 human fetal lung cell unscheduled DNA synthesis (UDS) test. ANIMAL STUDIES: Single intravenous doses of argatroban at 200, 124, 150, and 200 mg/kg were lethal to mice, rats, rabbits, and dogs, respectively. The symptoms of acute toxicity were loss of righting reflex, tremors, clonic convulsions, paralysis of hind limbs, and coma. Developmental studies performed in rats (during gestation Days 7 to 17) with argatroban at intravenous doses up to 27 mg/kg/day and in rabbits (during gestation Days 6 to 18) at intravenous doses up to 10.8 mg/kg/day have revealed no evidence of harm to the fetus. Argatroban was not genotoxic in the Ames test, the Chinese hamster ovary cell (CHO/HGPRT) forward mutation test, the Chinese hamster lung fibroblast chromosome aberration test, the rat hepatocyte or the mouse micronucleus test.

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Because argatroban is poorly absorbed orally, it might not adversely affect a breastfeeding infant. However, no information is available on the use of argatroban during breastfeeding, so an alternate drug is preferred.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Protein Binding
54%

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Interactions
A combined effect on the INR occurs with coadministration of argatroban and warfarin, and the relationship between INR and bleeding risk is altered. Daily INR determinations are recommended during concomitant use of argatroban and warfarin. Continue to monitor the effects of argatroban using aPTT during conversion to warfarin. Argatroban therapy can be discontinued when the INR exceeds 4 with combined therapy. Repeat INR determinations 4-6 hours after discontinuance of the argatroban infusion should be within the desired therapeutic range for warfarin monotherapy.
Potential pharmacologic interaction (increased risk of hemorrhage) with concomitant use of thrombolytics, antiplatelet agents, or other anticoagulants. No pharmacokinetic or pharmacodynamic interaction demonstrated with low-dose oral aspirin (162.5 mg given 26 and 2 hours prior to argatroban infusion) or oral acetaminophen (1 g given every 6 hours for 5 doses beginning 12 hours prior to argatroban infusion). The manufacturer states that the safety and efficacy of concomitant therapy with argatroban and platelet glycoprotein (GP) IIb/IIIa-receptor antagonists has not been established.
Heparin is contraindicated in patients with heparin-induced thrombocytopenia (HIT). Prior to initiation of argatroban therapy, allow sufficient time for effect of heparin on activated partial thromboplastin time (aPTT) to decrease.
Pharmacodynamic interaction (increased prothrombin time (PT) and international normalized ratio (INR) relative to warfarin alone).

[1]. Br J Pharmacol. 1994 Dec;113(4):1209-14.

[2]. Eur J Pharmacol. 2003 Feb 7;461(1):9-17.

[3]. Jpn J Pharmacol. 1995 Oct;69(2):143-8.

[4]. J Pharmacol Exp Ther. 1996 Aug;278(2):780-5.

[5]. Cardiovasc Drugs Ther. 2017 Jun;31(3):255-267.

(2R,4R)-1-[(2S)-5-(diaminomethylideneamino)-2-[(3-methyl-1,2,3,4-tetrahydroquinolin-8-yl)sulfonylamino]-1-oxopentyl]-4-methyl-2-piperidinecarboxylic acid is a peptide.
Argatroban is a direct, selective thrombin inhibitor. The American College of Cardiologists (ACC) recommend using bivalirudin or argatroban in patients who have had, or at risk for, heparin induced thrombocytopenia (HIT) and are undergoing percutaneous coronary intervention. Argatroban is a non-heparin anticoagulant shown to both normalize platelet count in patients with HIT and prevent the formation of thrombi. Parental anticoagulants must be stopped and a baseline activated partial thromboplastin time must be obtained prior to administering argatroban.
Argatroban anhydrous is an Anti-coagulant and Direct Thrombin Inhibitor. The mechanism of action of argatroban anhydrous is as a Thrombin Inhibitor.
Argatroban is a synthetic derivative of L-arginine with antithrombotic activity. Argatroban is a univalent and direct inhibitor of fibrin-bound thrombin. This agent reversibly binds to the thrombin active site thereby preventing the thrombin-dependent reactions, which include conversion of fibrinogen to fibrin; the activation of factors V, VIII and XI; the activation of protein C; and platelet aggregation. Argatroban is highly selective for thrombin and is able to inhibit the action of both free and clot-associated thrombin. As a result, stabilization of blood clots and coagulation is inhibited.
Argatroban Anhydrous is the anhydrous form of argatroban, a synthetic derivative of L-arginine with antithrombotic activity. Argatroban is a univalent and direct inhibitor of fibrin-bound thrombin. This agent reversibly binds to the thrombin active site, thereby preventing the thrombin-dependent reactions, which include the conversion of fibrinogen to fibrin; the activation of factors V, VIII and XI; the activation of protein C; and platelet aggregation. Argatroban is highly selective for thrombin and is able to inhibit the action of both free and clot-associated thrombin. As a result, stabilization of blood clots occurs and coagulation is inhibited.

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Drug Indication
Argatroban is indicated for prevention and treatment of thrombosis caused by heparin-induced thrombocytopenia (HIT). It is also indicated for use in patients with, or at risk for, HIT who are undergoing percutaneous coronary intervention.
FDA Label

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Mechanism of Action
Argatroban exerts its anticoagulant effects by inhibiting thrombin-catalyzed or -induced reactions, including fibrin formation; activation of coagulation factors V, VIII, and XIII; protein C; and platelet aggregation.

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Therapeutic Uses
Antithrombins; Platelet Aggregation Inhibitors
/CLINICAL TRIALS/ ClinicalTrials.gov is a registry and results database of publicly and privately supported clinical studies of human participants conducted around the world. The Web site is maintained by the National Library of Medicine (NLM) and the National Institutes of Health (NIH). Each ClinicalTrials.gov record presents summary information about a study protocol and includes the following: Disease or condition; Intervention (for example, the medical product, behavior, or procedure being studied); Title, description, and design of the study; Requirements for participation (eligibility criteria); Locations where the study is being conducted; Contact information for the study locations; and Links to relevant information on other health Web sites, such as NLM's MedlinePlus for patient health information and PubMed for citations and abstracts for scholarly articles in the field of medicine. Argatroban is included in the database.
Argatroban injection is indicated as an anticoagulant in adult patients with or at risk for heparin-induced thrombocytopenia (HIT) undergoing percutaneous coronary intervention (PCI). /Included in US product label/
Argatroban injection is indicated for prophylaxis or treatment of thrombosis in adult patients with heparin-induced thrombocytopenia (HIT). /Included in US product label/
For more Therapeutic Uses (Complete) data for Argatroban (10 total), please visit the HSDB record page.

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Drug Warnings
Other nonhemorrhagic adverse effects occurring in at least 2% of argatroban-treated patients with HIT/HITTS undergoing PCI include chest pain, back pain, headache, bradycardia, and myocardial infarction.
Adverse hemorrhagic effects reported in 2% or more of nonsurgical patients with HIT/HITSS receiving argatroban include major or minor GI bleeding, minor genitourinary bleeding or hematuria, minor decrease in hemoglobin/hematocrit, minor groin or brachial bleeding (e.g., catheter insertion site), and hemoptysis;9 nonhemorrhagic effects include dyspnea, hypotension, fever, diarrhea, sepsis, cardiac arrest, nausea, ventricular tachycardia, pain, urinary tract infection, vomiting, infection, pneumonia, atrial fibrillation, coughing, abnormal renal function, abdominal pain, and cerebrovascular disorder.
Safety and efficacy of argatroban not fully established in pediatric patients; however, the drug has been evaluated in a limited number of seriously ill pediatric patients younger than 16 years of age with HIT or HITTS. In a small, multicenter open-label study, 18 seriously ill pediatric patients with a clinical condition requiring alternative nonheparin anticoagulation received argatroban at an initial dosage of 1 ug/kg per minute titrated to maintain a target aPTT of 1.5-3 times the baseline value. During the 30-day study period, thrombotic events occurred in 5 patients and major bleeding (intracranial hemorrhage) was reported in 2 patients. All of the patients had serious comorbid conditions and were receiving multiple concomitant medications; most were diagnosed with documented or suspected HIT. Pharmacokinetic analysis of the data indicated that argatroban clearance was reduced by 50% in seriously ill pediatric patients compared with healthy adults and by approximately 80% in pediatric patients with elevated bilirubin concentrations compared to pediatric patients with normal bilirubin concentrations. Based on these results, reduced dosages of argatroban are recommended in pediatric patients.
There are no data on the presence of argatroban in human milk, or its effects on milk production. Argatroban is present in rat milk. The developmental and health benefits of breastfeeding should be considered along with the mother's clinical need for Argatroban and any potential adverse effects on the breastfed infant from Argatroban or from the underlying maternal condition.
For more Drug Warnings (Complete) data for Argatroban (16 total), please visit the HSDB record page.

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Pharmacodynamics
Argatroban is a synthetic direct thrombin inhibitor derived from L-arginine indicated as an anticoagulant for prophylaxis or treatment of thrombosis in patients with heparin-induced thrombocytopenia. Argatroban is a direct thrombin inhibitor that reversibly binds to the thrombin active site. Argatroban does not require the co-factor antithrombin III for antithrombotic activity. Argatroban exerts its anticoagulant effects by inhibiting thrombin-catalyzed or -induced reactions, including fibrin formation; activation of coagulation factors V, VIII, and XIII; protein C; and platelet aggregation. Argatroban is highly selective for thrombin with an inhibitory constant (K_i) of 0.04 µM. At therapeutic concentrations, Argatroban has little or no effect on related serine proteases (trypsin, factor Xa, plasmin, and kallikrein). Argatroban is capable of inhibiting the action of both free and clot-associated thrombin.

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Purpose: Chronic diabetes is associated with cardiovascular dysfunctions. Diabetic cardiomyopathy (DCM) is one of the serious cardiovascular complications associated with diabetes. Despite significant efforts in understanding the pathophysiology of DCM, management of DCM is not adequate due to its complex pathophysiology. Recently, involvement of protease-activated receptors (PARs) has been postulated in cardiovascular diseases. These receptors are activated by thrombin, trypsin, or other serine proteases. Expression of PAR has been shown to be increased in cardiac diseases such as myocardial infarction, viral myocarditis, and pulmonary arterial hypertension. However, the role of PAR in DCM has not been elucidated yet. Therefore, in the present study, we have investigated the role of PAR in the condition of DCM using a pharmacological approach. We used argatroban, a direct thrombin inhibitor for targeting PAR. Methods: Type-2 diabetes mellitus (T2DM) was induced by high-fat feeding along with low dose streptozotocin (STZ 35 mg/kg, i.p. single dose) in male Sprague-Dawley rats. After 16 weeks of diabetes induction, animals were treated with argatroban at 0.3 and 1 mg/kg dose daily for 4 weeks. After 20 weeks, ventricular functions were measured using ventricular catheterization. Cardiac histology, TUNEL staining, and immunoblotting were performed to evaluate cardiac fibrosis, DNA fragmentation, and expression level of different proteins, respectively. Results: T2DM was associated with cardiac structural and functional disturbances as evidenced from impaired cardiac functional parameters and increased fibrosis. There was a significant increase in PAR expression after 20 weeks of diabetes induction. Four weeks argatroban treatment ameliorated metabolic alterations (reduced plasma glucose and cholesterol), ventricular dysfunctions (improved systolic and diastolic functions), cardiac fibrosis (reduced percentage area of collagen in picro-sirius red staining), and apoptosis (reduced TUNEL positive nuclei). Reduced expression of PAR1 and PAR4 in the argatroban-treated group indicates a response towards inhibition of thrombin. In addition, AKT (Ser-473), GSK-3β (Ser-9), p-65 NFĸB phosphorylation, TGF-β, COX-2, and caspase-3 expression were reduced significantly along with an increase in SERCA expression in argatroban-treated diabetic rats which indicated the anti-fibrotic, anti-inflammatory, and anti-apoptotic potential of argatroban in DCM. Conclusion: This study suggests the ameliorative effects of argatroban in diabetic cardiomyopathy by improving ventricular functions and reducing fibrosis, inflammation, apoptosis, and PAR expression.5]

C23H36N6O5S

508.63

508.246

C, 54.31; H, 7.13; N, 16.52; O, 15.73; S, 6.30

74863-84-6

Argatroban monohydrate;141396-28-3;Argatroban-d3;1356847-56-7

92722

White to off-white solid powder

1.5±0.1 g/cm3

801.3±75.0 °C at 760 mmHg

188-1890C

438.4±37.1 °C

0.0±3.0 mmHg at 25°C

1.674

2.56

5

8

9

35

887

3

O=C([C@@H]1N(C([C@@H](NS(=O)(C2=CC=CC3=C2NCC(C)C3)=O)CCCNC(N)=N)=O)CC[C@@H](C)C1)O

KXNPVXPOPUZYGB-IOVMHBDKSA-N

InChI=1S/C23H36N6O5S/c1-14-8-10-29(18(12-14)22(31)32)21(30)17(6-4-9-26-23(24)25)28-35(33,34)19-7-3-5-16-11-15(2)13-27-20(16)19/h3,5,7,14-15,17-18,27-28H,4,6,8-13H2,1-2H3,(H,31,32)(H4,24,25,26)/t14-,15?,17+,18-/m1/s1

(2R,4R)-1-[(2S)-5-(diaminomethylideneamino)-2-[(3-methyl-1,2,3,4-tetrahydroquinolin-8-yl)sulfonylamino]pentanoyl]-4-methylpiperidine-2-carboxylic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

2% DMSO+40% PEG 300+2% Tween 80+ddH2O: 5mg/mL (Please use freshly prepared in vivo formulations for optimal results.)