Comparative Study on the Anticoagulant Effects of Low Molecular Weight Heparins as Measured by Whole Blood Act, Anti-Xa and a Newly Developed Prothrombinase Induced Clotting Time (PiCT) Assay.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4148-4148
Author(s):  
D. Hoppensteadt ◽  
M. Tobu ◽  
R. Wahi ◽  
O. Iqbal ◽  
J. M. Walenga ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Whole Blood ◽  
Low Molecular Weight ◽  
Plasma Samples ◽  
Clotting Time ◽  
Low Molecular Weight Heparins ◽  
Blood Samples ◽  
Good Relationship ◽  
Surgical Conditions ◽  
Better Than

Abstract Low Molecular Weight Heparins (LMWHs) are currently developed for anticoagulation in various intravenous indications such as DVT treatment and percutaneous interventions (PCI). Traditionally, these drugs are administered in anti-Xa units. The purpose of this study was to compare the relative levels of anticoagulation produced by different LMWHs in whole blood ACT (Hemochron), an amidolytic anti-Xa method and a plasma-based global anticoagulant assay, the PiCT test (Pentapharm, Basel, Switzerland). Various LMWHs such as dalteparin, enoxaparin, reviparin and a synthetic pentasaccharide were supplemented to freshly drawn native blood samples to simulate patient samples containing 1 U/ml anti-Xa levels. Three generic versions of enoxaparin were also included in this study. In addition to the ACT measurement in whole blood, citrated plasma samples were also tested for APTT, Heptest, PiCT, amidolytic anti-Xa and IIa activities, and protease generation assays. All agents produced assay-based effects on different tests, which were not proportional to the adjusted anti-Xa activity. Correlation of the ACT data resulted in a good relationship with PiCT for the commercially available branded products(r2=0.97). ACT and Heptest (r2=0.51), ACT and anti-Xa (r2<0.3), ACT and anti-IIa (r2= 0.93). The correlation of PiCT and Heptest was (r2=0.73) better than ACT and Heptest. However, the generic versions of enoxaparin did not provide similar results. PiCT and Heptest also gave the good correlation. These results clearly suggest that for the global anticoagulant actions of LMWHs, anti-Xa measurements are of limited value and should not be used to asses the anticoagulant efficacy of LMWHs in PCI and other surgical conditions. These differences may be more obvious in the generic versions of LMWHs, which are apparently produced using pharmacopeial specifications. Thus, it is important to include a clot-based method in the cross referencing and standardization of LMWHs.

Defibrotide Augments the Anticoagulant Actions of Heparin and Low Molecular Weight Heparins

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4086-4086
Author(s):  
Jawed Fareed ◽  
Omer Iqbal ◽  
Debra Hoppensteadt ◽  
Cafer Adiguzel ◽  
Massimo Iacobelli ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Whole Blood ◽  
Thrombin Generation ◽  
Synergistic Effects ◽  
Low Molecular Weight ◽  
Inhibitory Effects ◽  
Clotting Time ◽  
Low Molecular Weight Heparins ◽  
Significant Prolongation ◽  
Antithrombotic Effects

Abstract Defibrotide represents a polydeoxyribonucleotide derived antithrombotic and antiischemic drug, which has been used in the management of vascular disorders and is currently being developed in other clinical indications. Defibrotide is a polyelectrolyte-based agent with target effects on endothelium, platelets, and blood cells. In addition, the aptameric consensus sequences in the nucleotides exhibit inhibitory effects towards thrombin and related proteases. In the anticoagulant assays defibrotide exhibits relatively weak effects (<5 USP U/mg). These studies were undertaken to study whether there is an interaction between defibrotide and unfractionated heparin (UFH) in various systems of anticoagulation. The interaction of defibrotide with commercially available low molecular weight heparins (LMWHs), enoxaparin and dalteparin, was also studied. For the first investigation, to evaluate the effect of defibrotide on the anticoagulant effects of UFH, native whole blood freshly drawn from human volunteers (n = 20) was supplemented with UFH at a fixed concentration of 5 μg/mL (0.8 U/mL), and graded amounts of defibrotide were added in a concentration range of 12.5 – 100 μg/mL. The whole blood celite Activated Clotting Time test (ACT) and the thrombin generation markers fibrinopeptide A (FPA), thrombin-antithrombin complex (TAT), and prothrombin fragment 1.2 (F1.2) were measured. Parallel controls with saline were included. While defibrotide did not produce a significant prolongation of the ACT compared to saline (128 ± 9 s vs 132 ± 7 s), it produced a concentration-dependent increase in the heparinized whole blood leading to an almost doubling of the anticoagulant action of UFH (248 ± 19 s vs 418 ± 21 s). Additional studies carried out by varying the concentrations of the two agents also revealed supraadditive to synergistic effects. Defibrotide also augmented the inhibitory effects of UFH on thrombin generation markers in a concentration-dependent fashion. Similar studies carried out with the two LMWHs did not reveal a similar interaction in the anticoagulant assays such as the ACT; however, significant interactions between defibrotide and the LMWHs were observed in the thrombin generation studies. For the second investigation, studies were carried out using plasma samples collected from heparinized patients (aPTT of 50 – 100 s). These studies also revealed that supplementation of defibrotide augmented the anticoagulant effects of UFH in a concentration-dependent fashion. While defibrotide at 12.5 μg/mL did not significantly increase the aPTT of normal plasma, when supplemented to heparinized plasmas (n = 50 with aPTT of 64.6 ± 14.0 s) it produced a strong prolongation of the clotting time (96.1 ± 20.6 s). In the third investigation, animal models of thrombosis including the rat jugular vein clamping model, demonstrated an augmentation of the antithrombotic effects of intravenously administered UFH by defibrotide. However, no augmentation of the hemorrhagic effect was observed in the rat tail bleeding model. These studies demonstrate that defibrotide exhibits a strong anticoagulant interaction with UFH and to a lesser degree LMWH. While the combination of defibrotide and UFH exhibits enhanced anticoagulant/antithrombotic activities, it does not exhibit any alteration of the hemorrhagic profile. These studies clearly suggest that defibrotide can be combined with UFH to achieve a superior anticoagulant approach with better safety/efficacy profile.

A NEW ONE STAGE CLOTTING ASSAY FOR HEPARIN AND LOW MOLECULAR WEIGHT HEPARINS

1987 ◽  
Author(s):  
Ch Giese ◽  
A Knodler ◽  
R Zimmermann ◽  
J Harenberg
Keyword(s):  
Molecular Weight ◽  
Factor Xa ◽  
Low Molecular Weight ◽  
Chromogenic Substrate ◽  
Clotting Time ◽  
Low Molecular Weight Heparins ◽  
One Stage ◽  
High Preference ◽  
Coagulation Tests ◽  
Plasma Assay

Heparin and its low molecular weight (LMW) derivatives are usually measured by chromogenic or fluorogenic synthetic substrate assays and by coagulation tests. Since the activated partial thromboplastin time (aPTT) and thrombin clotting time (TCT) are insensitive to LMW heparins, we report here of data obtained with heptest, a new one stage modification of the original heparin in plasma assay of Yin. The assay was compared with the antifactor Xa chromogenic substrate S2222 method, the TCT and aPTT tests in 100 patients receiving unfractionated pig intestinal mucosa heparin and 100 patients treated with low molecular weight heparin Kabi 2165. The results indicate a high correlation between the heptest and the anti Xa chromogenic substrate method, whereas the correlations were lower for the aPTT and TCT. correlations with LMW heparinThe lowest detection limit of the heptest is 0,005 heparin units per ml plasma. The test is very sensitive, simple, highly reproducable and reliable clotting assay for unfractionated and low molecular weight heparins in human plasma. The test detects with high preference the inhibition on factor Xa but also the other anticoagulant effects on die coagulation tractors.

Comparative Studies on An Anti-Xa Enriched Ultra Low Molecular Weight Heparin with Bemiparin

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4394-4394
Author(s):  
Debra Hoppensteadt ◽  
Angel Gray ◽  
Josephine Cunanan ◽  
Walter Jeske ◽  
Jeanine M. Walenga ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Platelet Aggregation ◽  
Whole Blood ◽  
Anticoagulant Activity ◽  
The Other ◽  
Positive Interactions ◽  
Low Molecular Weight ◽  
Significant Activity ◽  
Thrombin Time ◽  
Low Molecular Weight Heparins

Abstract Abstract 4394 Most low molecular weight heparins (LMWHs) have a mean molecular weight in the range of 4–6 kDa and anti-Xa/IIa ratios of 3–6. Further depolymerization of porcine mucosal heparin results in the generation of Ultra low molecular weight heparins (ULMWHs) with a molecular weight range of 2–4 kDa with proportionately decreased anti-Xa and anti-IIa activities. Bemiparin (Rovi, Madrid, Spain) represents one such ULMWH. AVE 5026 (Sanofi-Aventis, Paris, France) is a unique ULMWH (2.5 kDa) which exhibits higher affinity to antithrombin (AT) and therefore, enhanced anti-Xa activity. Because of the compositional differences between these two agents, it was hypothesized that each of these agents will have distinct anticoagulant, antiprotease and thrombin generation effects. Each of these agents was supplemented to native whole blood. Anticoagulant activity was measured using ACT, TEG, PT, APTT, thrombin time and Heptest assays. Similar studies were carried out in plasma. Amidolytic assays were used to determine the anti-Xa and anti-IIa activities. Both agents were also tested for the interactions with heparin cofactor II (HC II) and AT and were compared in the HIT antibody screening assay using platelet aggregation. In whole blood clotting assays bemiparin showed a strong anticoagulant activity in comparison to AVE 5026. Both agents also exhibited assay dependent differences in the APTT, heptest and thrombin time assays. AVE 5026 exhibited a higher anticoagulant activity in the heptest whereas bemiparin showed a stronger anticoagulant effect in the other clot based assays. In the amidolytic anti-Xa assay, AVE 5026 showed an activity of 156U/mg compared to 86 U/mg for bemiparin. In the anti-IIa assay bemiparin showed a higher activity (10 U/mg) in comparison to AVE 5026 (3.2 U/mg). The calculated Xa/IIa ratio of AVE 5026 was > 48, whereas it was 8.6 for bemiparin. While bemiparin exhibited interactions with HC II, AVE 5026 did not show significant activity in the tested concentrations (anti-IIa – IC50: 1.10±.45 μ M and >3.44±.00 μ M, respectively). On the other hand, AVE 5026 exhibited stronger interactions with AT in comparison to bemiparin (anti-FXA – IC50: .223±.03 μ M and .894±.06 μ M, respectively). Interestingly, heparinase digestion of the two products resulted in a complete loss of anti-IIa activity, but residual anti-Xa activity was found. AVE 5026 exhibited stronger anti-Xa interactions even after heparinase digestion. In the heparin induced platelet aggregation assay at 2.5 μ g/ml, bemiparin showed a relatively higher prevalence of positive interactions with HIT antibodies, whereas AVE 5026 showed a much lower prevalence (slope; AVE 5026 compared bemiparin, p=0.012). Bemiparin exhibited greater platelet factor 4 neutralization in comparison to AVE 5026. These studies clearly demonstrate that while bemiparin behaves like a typical ULMWH, AVE 5026 behaves differently in the different assays. Moreover, the oligosaccharide composition of the two products, in terms of distribution profile structure, is also different. Therefore, AVE 5026 does not represent a typical depolymerized ULMWH and is expected to exhibit a distinct pharmacologic and clinical profile. Disclosures: Hoppensteadt: Sanofi-Aventis: Research Funding.

Anti-Xa Potentiating Effect Of Low Molecular Weight Heparin

1981 ◽  
Author(s):  
W Junker ◽  
J Harenberg ◽  
F Fussi ◽  
K Mattes ◽  
R Zimmermann ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Intestinal Mucosa ◽  
Low Molecular Weight Heparin ◽  
Bleeding Complications ◽  
Low Molecular Weight ◽  
Chromogenic Substrate ◽  
Clotting Time ◽  
Blood Samples ◽  
Increased Risk

Recently special attention has been drawn to bleeding complications of commercial heparins in patients with increased risk for haemorrhages. Alternative heparin preparations with high antithrombotic and low haemostaseological properties have been developed. We now report on a new low molecular weight (LMW) heaprin (mean MW 5000, 85 USP/mg), which has been obtained by depolymerisation of a heparin from pig intestinal mucosa (mean MW 15000, 154 USP/mg).In vitro the anti-Xa-activity (chromogenic substrate S2222) was 15% higher for the LMW heparin in a range of 0.01-2.0 USP/ml plasma. No difference was seen on the anti-IIa-activity (thrombin clotting time)and the aPTT for both heparins in the same range. Both Heparins were injected s.c. in a dose of 100, 50 and 25 USP/kg bodyweight into each of six volunteers randomly at weekly intervalIs. The pharmacodynamic effects were controlled for 6-10 hrs by 8-12 blood samples in relation to the dose applied. Increasing dosis the effects of each heparin increased in all test systems. The anti- Xa-activity of LMW heparin was somewhat higher at 100 and 25 USP/kg. At 50 USP/kg the effect of LMW heaprin was in the same range as 100 USP/kg of the original preparation (MW 15000). The factor Ila activity and aPTT were not influenced differently by the two heparins at each dose.The data indicate, that the LMW heparin presented here may have a more pronounced antithrombotic property by a specific anti-Xa-activity than the compaired commercial heparin. This effect is most pronounced at doses, which have only small haemostaseological effects.

COMPARATIVE CLINICAL PHARMACOLOGY OF LOW MOLECULAR WEIGHT HEPARINS IN MAN

1987 ◽  
Author(s):  
Y Ordu ◽  
J Augustin ◽  
E V Hodenberg ◽  
V Bode ◽  
J Harenberg
Keyword(s):  
Molecular Weight ◽  
Clinical Pharmacology ◽  
Ex Vivo ◽  
Average Molecular Weight ◽  
Factor Xa ◽  
Low Molecular Weight ◽  
Pharmacological Properties ◽  
Clotting Time ◽  
Activity Time ◽  
Low Molecular Weight Heparins

Low molecular weight (LMW) heparins are obtained by diffent chemical procedures from conventional pig intestinal mucosa heparin. The LMW heparins differ in their molecular weight distribution and physicochemical properties. Therefore, we report of comparative studies on the anticoagulant and lipolytic effects of low molecular weight heparins in man.The following LMW heparins were used: BM 21-23 (Braun, Melsungen, FRG), CY 216 (Choay Laboratories, Paris, France), Heparin NM (Sandoz, Niimberg, FRG), Kabi 2165 (Kabi Vitrum AB, Stockholm, Sweden), RD Heparin (Hepar Industries, Franklin, US A), normal heparin (Braun). All heparins were administered intravenously and subcutaneously to six volunteers each.The data show considerable differences in the anticoagulant and lipolytic effects between the different low molecular weight heparins. From the area under the activity time curves (AUC) of the clotting assays for factor Xa (heptest), aPTT and thrombin clotting time the aXa/aPTT ratio ex vivo and aXa/alla ratio ex vivo were determined (table, average values)It can be seen that there are clear differences in the ex vivo ratios of the LMW heparins. There is a good correlation between the average molecular weight of the LMW heparins and the aXa/aPTT ratio after s.c. administration and of the aXa/alla ratio ex vivo after s.c. administration. Therefore, LMW heparins differ significantly in their clinical pharmacological properties.

Heptest-STAT, a new assay for monitoring of low-molecular-weight heparins, is not influenced by pregnancy-related changes of blood plasma

2009 ◽  
Vol 102 (11) ◽  
pp. 1001-1006 ◽  
Author(s):  
Ulyana Zharkowa ◽  
Elif Elmas ◽  
Parviz Ahmad-Nejad ◽  
Michael Neumaier ◽  
Martin Borggrefe ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Pregnant Women ◽  
Factor Xa ◽  
Low Molecular Weight ◽  
Plasma Samples ◽  
Third Trimester ◽  
Low Molecular Weight Heparins ◽  
Individual Calibration ◽  
Standard Calibration ◽  
In Pregnancy

SummaryMonitoring of anti-factor Xa activity is often performed during treatment with low-molecular-weight heparins (LMWHs) in pregnancy because the anticoagulant effect may decrease as pregnancy progresses, but assays for anti-factor Xa activity are unavailable in many clinical institutions caring for pregnant women. Heptest-STAT is a new clotting assay for monitoring of LMWHs, which has been optimised for use in near-patient laboratory instrumentation. It has been suggested that monitoring of LMWHs requires the use of individual calibration curves for each LMWH.We compared the dose response of four conventional LMWHs and fondaparinux in normal plasma, and plasma from women in first, second and third trimester of pregnancy. Three concentrations of LMWHs, fondaparinux, or unfractionated heparin were added to pooled plasma samples from nonpregnant women (n=10), and pregnant women in first (n=10), second (n=10) and third (n=10) trimester of pregnancy. Heptest results are not influenced by the stage of pregnancy. In contrast, dose-related aPTT prolongation declines during pregnancy. All LMWHs tested, as well as fondaparinux, display a similar doseresponse in Heptest compared to the chromogenic anti-factor Xa assay. Heptest-STAT can be used with the same standard calibration for non-pregnant and pregnant patients and for all LMWHs under investigation, including fondparinux. No individual calibrations are necessary.

Measurement of whole blood factor Xa-activated clotting time during hemodialysis with low-molecular-weight heparin

1998 ◽  
Vol 12 (3) ◽  
Author(s):  
Masakazu Mori ◽  
Shigenori Yoshitake ◽  
Takaaki Kitano ◽  
Shunsuke Oda ◽  
Takayuki Noguchi
Keyword(s):  
Molecular Weight ◽  
Whole Blood ◽  
Low Molecular Weight Heparin ◽  
Factor Xa ◽  
Low Molecular Weight ◽  
Clotting Time ◽  
Activated Clotting Time
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