Clinical and Applied Thrombosis/Hemostasis

Heparin has been conventionally used as an anticoagulant for medical and surgical indications. Because factor Xa is an essential component of the prothrombinase complex and leads to the generation of thrombin, its inhibition has become a focus of newer antithrombotic drug development. The in vitro anticoagulant profile of DX-9065a, a synthetic direct factor Xa inhibitor, was studied using activated clotting time assay, thrombelastography, and global clotting tests, such as prothrombin time (PT), activated partial thromboplastin time (aPTT), diluted aPTT, Heptest, Heptest-HI, dilute Russell's viper venom time (dRVVIT), thrombin time, ecarin clotting time, and amidolytic anti-Xa assay. In addition, the effect of DX-9065a on platelet aggregation and inhibition of thrombin generation markers (FPA, Fl + 2, and TAT) were studied. The pharmacokinetic and pharmacodynamic profiles of DX-9065a were also studied in a non-human primate (Macaca mulatta) model. DX-9065a produced a concentration-dependent increase in the Hemochron celite ACT and HemoTec ACT. Clotting times of 538 ± 19 and 401 ± 12, respectively, were reached at a concentration of 25 μg/mL signifying that DX-9065a may be useful in interventional cardiological procedures. DX-9065a prolonged the r-time on thrombelastography. DX-9065a did not show any effect on adenosine diphosphate (ADP)-, collagen-, epinephrine-, and arachidonic acid-induced platelet aggregation at concentrations up to 10 μg/mL. DX-9065a exhibited a concentration-dependent prolongation of the PT, aPTT, diluted aPTT, Heptest, dRVVT, and reached the clotting times of 51.6, 132, 193, 47.9, 129.9 seconds, respectively, at a final concentration of 12.5,tg/mL; compared to a control value of 10.6, 30.2, 41.9, 14, 32.2 seconds, respectively. DX9065a did not affect the ecarin clotting time and thrombin time at concentrations up to 12.5 μg/mL. Because DX-9065a prolonged the dRVVT, this may impact diagnostic screening of patients with systemic lupus erythematosus.

Because of the unsatisfactory options available for safe and effective antithrombotic therapy, recent, intense research and development efforts have been focused on direct thrombin inhibitors, also known as site-directed thrombin inhibitors. The intravenous agent Novastan (argatroban) is a small-molecule, reversible, direct thrombin inhibitor that is selective for the catalytic site of the thrombin molecule. Argatroban's molecular properties (small molecule; fast, selective, and reversible inhibition of the thrombin catalytic site; and similar in vitro potency for inhibiting both clot-bound and soluble thrombin) offer the potential for significant antithrombotic efficacy with minimal systemic anticoagulant ef fects. Its clinical pharmacologic properties offer the potential for minimal risk of bleeding, very rapid achievement of therapeutic antithrombotic efficacy, predictable dose-response, and rapid restoration of the hemostatic systems to normal upon termination of intravenous infusion. Argatroban is currently approved for clinical use in Japan for the treatment of peripheral arterial occlusive disease. It is in advanced clinical development in North America, South America, and Western Europe for several clinical indications, including (1) adjunctive therapy to thrombolytic agents in the treatment of acute myocardial infarction and (2) antithrombotic therapy for patients with heparin-induced thrombocytopenia and heparin-induced thrombocytopenia and thrombosis syndrome. Key Words: Molecular properties—Novastan (argatroban)—Pharmacology—Thrombin inhibitor.

Our purpose was to study the peculiarities of changing the hemostasis in children with chronic constipation. in the course of our investigation of hemostasis in children with chronic constipation, we found chronometric hypocoagulation th impairment of internal (XII, XI, IX, VIII) and external (II, VII, X) mechanisms of blood clotting, at the base of which a deficit of vitamin K-dependent factors (II, VII, IX, X) and slight impairment of the final stage of coagulation. In thrombocytovascular hemostasis, thrombocytopathy demonstrated increased adenosine-5-diphosphate aggregation and suppression of the internal route of fibrinolysis and endotheliosis. We must point out that all these changes lead to impairment of microcirculation in the form of thromboses, particularly in the submucous and mucous membrane vessels, which is confirmed the morphologic findings. In the compensated form, chronometric hypocoagulation with a deficit of II, IX, and X factors is noted. In the subcompensated form, hypocoagulation increases, and the deficit of V and VII factors is in part the cause. Endotheliosis and the increase of the thrombogenesis dynamics are noted in the threombocytovascular section. In the decompensated form, there is marked hypocoagulation. Low activity of antithrombin III leads to high content of fibrinmonomeasured complexes in combination with reduction of the thrombocytes quantity and to hypercoagulation. The present changes are confirmed by the morphologic research methods of studying the vascular channel of the large intestine wall, where organized thrombi and a marked plethora of neutrophil tissue infiltration are often revealed. Key Words: Chronic constipation— Hemostasis—Thrombosis.

Deficiency of any of the two coagulation in hibitors antithrombin (AT) and tissue factor pathway in hibitor (TFPI) lowers the resistance to thrombosis. He reditary deficiency of AT leads to a high risk of throm bosis, which occasionally responds poorly to heparin therapy. Experimental deficiency of TFPI lowers the re sistance to infusion of both tissue factor and endotoxin, both regarding microvascular thrombosis and fatality. Administration of either AT or TFPI protects against mi cro- and macrovascular thrombosis. Injection of heparin and some other glycosaminoglycans releases intima bound TFPI to the blood. Heparin accelerates the inhib itory effects of both inhibitors, in particular the effect of AT. The influence of the two inhibitors on the various anticoagulant reactions have been studied using blocking antibodies. It is suggested that the anticoagulant and an tithrombotic effects of heparin are mainly mediated by the accelerated inactivation of thrombin, factor IXa and factor X by AT, and augmented inactivation of tissue factor-factor VIIa by TFPI released to the blood.

Three-hundred fifty-one women were referred for thrombosis and hemostasis evaluation after suffering recurrent miscarriages. All patients were referred by a high-risk obstetrician or reproductive medicine specialist after anatomic, hormonal or chromosomal defects had been ruled out. These patients were assessed over a three year period. The mean patient age at referral was 34 years and the mean number of miscarriages was 2.9 (2-9). All patients underwent a thorough evaluation for thrombophilia and, when indicated, a hemorrhagic disorder. Of the 351 patients, 29 (8%) had no defect. Of the remaining 322 patients, 7 (2%) had a bleeding disorder: 3 with platelet dysfunction, 1 with Factor XIII deficiency, 3 with von Willebrand’s and 3 with Osler-Weber-Rendu. The remainder of the patients had a thrombophilia as follows: 195 (60%) had antiphospholipid syndrome, 64 (20%) had Sticky Platelet Syndrome, 38 (12%) had MTHFR mutation, 23 (7.1%) had PAI-1 polymorphism, 12 (3.7%) had Protein S deficiency, 12 (3.7%) had Factor V Leiden, 3 (1%), had AT deficiency, 3 (1%) had Heparin-Cofactor II deficiency, 3 (1%) had TPA deficiency, and 6 (2%) had Protein C deficiency. There were a total of 364 defects found in the 312 patients harboring thrombophilia; thus, several harbored two and a few harbored three separate defects. All patients with thrombophilia were treated with preconception ASA at 81 mg/day with the immediate post-conception addition of heparin or LMW heparin (Dalteparin). Both ASA and heparin/LMW heparin were used to term. The first 120 patients were treated with unfractionated heparin at 5,000 U every 24 hours, subcutaneously and the last 192 have been treated with Dalteparin at 5,000 U/day subcutaneously. The patients with MTHFR were also treated with folate at 5 mg/day + pyridoxine at 50 mg/day. All patients were carefully monitored with CBC and platelet counts, anti-Xa levels, frequent ultrasounds and physical exams. Only 2 of the thrombophilia patients suffered another miscarriage; all others had a normal term delivery. There were no pregnancy-related thromboses, no delivery complications and no episodes of post-partum thrombosis. The only bleeding consisted of 1-4 cm bruises at injection sites. No episodes of thrombocytopenia (HIT) were noted. In our experience, thrombophilia is a common cause of recurrent miscarriage and all patients with no anatomical, hormonal or chromosomal defect should be evaluated for thrombophilia or a bleeding disorder. The success rate of normal term delivery in these 312 patients was 94% using ASA + heparin or Dalteparin. In addition, side effects of therapy were minimal.

A regimen of prophylaxis of thromboembolism with low molecular weight heparin (LMWH) and unfractionated heparin (UFH) in orthopedic surgery is presented. Over a 5-year period, 11,905 patients have been operated in all fields of orthopedic surgery in a big trauma center. In 498 cases, deep vein thrombosis (DVT) was clinically suspected and phlebography was performed: 122 patients had confirmed DVT (rate 0.98%); 59.8% of the DVT were limited to the calf. Nineteen pulmonary embolisms (rate 0.16%) were diagnosed (3 lethal). Heparin-induced thrombocytopenia type II was diagnosed in 12 of 122 patients. Only one of these had received LMWH. The related rates of DVT were highest for tibial head fractures (12.4%), total knee replacement (6.7%), and multiple trauma (6.4%). The extent of DVT was most severe in multiple trauma. The results proved the regimen to be of good clinical use and showed that more attention has to be paid to fractures, especially of the knee; and to patients with multiple injuries.

Excessive bleeding is one of the major complica tions of cardiac surgery with cardiopulmonary bypass (CPB). This complication is related to the operation and the defects in hemostasis induced by extracorporeal circulation. The system atic effects of CPB are called whole body inflammatory reac tion. Heparin, platelet dysfunctions, and fibrinolysis are the major causes of bleeding problems associated with CPB. Dif ferent pharmacological approaches have been used to reduce bleeding and the need for blood transfusions in patients under going cardiac surgery. The most effective is aprotinin, a serum protease inhibitor that is an antifibrinolytic with a platelet- preserving action. It inhibits the activation of the intrinsic co agulation system. Aprotinin therapy effectively reduces blood loss and donor blood requirements. According to most reports, it does not increase the risk of acute myocardial infarction, renal dysfunction, and mortality.

Thirty-five patients diagnosed to have ovarian cancer (early FIGO stage I/II n = 11, advanced FIGO stage III/IV, n = 24) were evaluated for hemostatic parameters relating to survival outcome by 36 months of disease. Systemic plasminogen activators and inhibitors were evaluated and we found no significant association with survival outcome and eventually only fibrinogen, von Willebrand Factor (vWF), antithrombin III (ATIII), and D-dimer levels were determined for their association with disease outcome by 12 months, 24 months, and 36 months. Twenty-four patients succumbed to the disease by 36 months (early n = 2, advanced n = 22). The 11 surviving patients (advanced n = 3, including one deceased at 52 months) is still living past 36 months and 82 months at the time of analysis. Elevated fibrinogen, vWF, and D-dimer together with reduced ATIII levels were found to be associated with poor survival outcome by 12 months of disease. Moreover, elevated vWF and D-dimer with reduced ATIII levels was strongly implicated with poor survival outcome by 36 months from ovarian cancer. The overall survival rate at 36 months from ovarian cancer was 31.4%. It is therefore suggested that fibrinogen, vWF, ATIII, and D-dimer levels be used together as prognostic markers for disease outcome especially in patients with advanced ovarian cancer within 36 months of disease. An expanded study is required to confirm these findings.

Venous thromboembolism (VTE), encompassing deep venous thrombosis and pulmonary embolism, is a potentially fatal but preventable complication of stroke. Reported rates of VTE after stroke have decreased over the last four decades, possibly due to the implementation of stroke units, early mobilization and hydration, and increased early use of antiplatelets. Additional means of thromboprophylaxis in stroke include mechanical methods (ie, compression stockings) to prevent venous stasis and medical therapy including antiplatelets, heparins, and heparinoids. Risk of VTE must be balanced by potential risk of hemorrhagic complications from pharmacotherapy. Unfractionated heparin, low-molecular-weight heparin (LMWH), and danaparoid are acceptable options for chemoprophylaxis though none have shown superior efficacy for VTE prevention without an associated increase in major hemorrhage. The efficacy and timing of pharmacological thromboprophylaxis in hemorrhagic stroke are not well defined. Graduated compression stockings are associated with an increased rate of adverse events and are not recommended and intermittent pneumatic compression stockings require further investigation.

Factor X Padua, first described a few years ago, is characterized by a defect only in the extrinsic system. In this present paper, the molecular basis for this peculiar defect is investigated. Polymerase chain reaction amplification and direct sequencing of the entire FX coding sequence and of exon-intron junctions detected in the proposita a C-to-T translocation in exon 8 of nucleotide 875 at the homozygous level. This resulted in the substitution of tryptophan for arginine 251. A niece of the proposita was shown to be heterozygous for the abnormality. Molecular modeling suggested that the mutation does not alter significantly folding and stability of the protein but may be involved in the Ca²⁺ binding site.