A new parameter in the thrombin generation assay, mean velocity to peak thrombin, reflects factor VIII activity in patients with haemophilia A

Haemophilia ◽  
2016 ◽  
Vol 22 (5) ◽  
pp. e474-e477 ◽  
Author(s):  
M. Takeyama ◽  
K. Nogami ◽  
M. Shima
Keyword(s):  
Factor Viii ◽  
Thrombin Generation ◽  
Haemophilia A ◽  
Factor Viii Activity ◽  
Mean Velocity ◽  
Thrombin Generation Assay

scholarly journals Thrombin generation assay using factor IXa as a trigger to quantify accurately factor VIII levels in haemophilia A

2011 ◽  
Vol 9 (8) ◽  
pp. 1549-1555 ◽  
Author(s):  
M. NINIVAGGI ◽  
Y. DARGAUD ◽  
R. Van OERLE ◽  
B. De LAAT ◽  
H. C. HEMKER ◽  
...  
Keyword(s):  
Factor Viii ◽  
Thrombin Generation ◽  
Haemophilia A ◽  
Thrombin Generation Assay ◽  
Factor Ixa

Clinical Profile of Haemophilia in a Tertiary Care Hospital in Pune, Maharashtra, India

2021 ◽  
Vol 8 (15) ◽  
pp. 968-971
Author(s):  
Sadiq Yunus Mulla ◽  
Sachin Sitaram Pandit ◽  
Sachin Kisan Shivnitwar
Keyword(s):  
Factor Viii ◽  
Tertiary Care Hospital ◽  
Tertiary Care ◽  
Factor Ix ◽  
Clinical Profile ◽  
Factor Viii Inhibitor ◽  
Haemophilia A ◽  
Care Hospital ◽  
Factor Viii Activity ◽  
Viii Inhibitor

BACKGROUND Haemophilia’s are X-linked hereditary blood clotting disorders due to deficiency of factor VIII (haemophilia A) or factor IX (haemophilia B) & also has identical clinical manifestations, screening tests abnormalities and sex-linked genetic transmission. Haemophilia’s result from defects in the factor VIII / IX gene that lead to decreased amount of factor VIII / IX protein, the presence of a functionally abnormal protein, or combination of both. Haemophilia A is a classic example of an X-linked recessive trait. The severity of their bleeding depends on their factor VIII activity level; and, rarely, a woman can have very low factor VIII activity, and present with symptoms of moderate or even severe haemophilia. We wanted to study the clinical profile of patients of haemophilia admitted in a tertiary care hospital. METHODS This is a cross-sectional study enrolling 60 known cases of haemophilia A & B admitted in wards & ICU / attending OPD of a tertiary care hospital. History was obtained in detail & thorough clinical examination was carried out. Precipitating factors for bleeding (spontaneous / minor trauma / major trauma / surgical operation / dental procedure / others), family h / o bleeding were studied in detail. RESULTS Of the total 60 cases of haemophilia, majority (49) of cases were of haemophilia A and 11 cases were of haemophilia B. In the study, majority (28.33 %) of cases belonged to 12 - 20 years age group and the most common presentation was haemarthrosis (61.67 %). 6 patients had factor VIII inhibitor antibodies and all of them were of haemophilia A. CONCLUSIONS Haemarthrosis is the most common clinical presentation of haemophilia and most common cause for haemarthrosis is spontaneous bleeding. Most common joint involved in bleeding was knee joint (target joint). Presence of factor VIII inhibitor antibodies specially in haemophilia A patients is not uncommon. KEYWORDS Haemophilia, Factor VIII, Factor IX

scholarly journals Mild hemophilia A with factor VIII assay discrepancy: using thrombin generation assay to assess the bleeding phenotype

2008 ◽  
Vol 6 (3) ◽  
pp. 486-493 ◽  
Author(s):  
M. TROSSAËRT ◽  
V. REGNAULT ◽  
M. SIGAUD ◽  
P. BOISSEAU ◽  
E. FRESSINAUD ◽  
...  
Keyword(s):  
Factor Viii ◽  
Thrombin Generation ◽  
Hemophilia A ◽  
Thrombin Generation Assay

Binding To Phospholipid Protects Factor VIII From Inhibition By Human Antibodies

1981 ◽  
Author(s):  
T W Barrowcliffe ◽  
E Gray ◽  
G Kemball-Cook
Keyword(s):  
Factor Viii ◽  
Thrombin Generation ◽  
Total Phospholipid ◽  
Fluid Phase ◽  
Clinical Applications ◽  
Factor Ix ◽  
Two Stage ◽  
Human Antibodies ◽  
Thrombin Generation Assay ◽  
Detectable Antigen

Previous studies with activated Factor IX concentrates have suggested that they may contain a form of Factor VIII clotting activity (VIII:C) which is partly protected from inactivation by antibodies. A possible mechanism for such protection is binding to phospholipid. The interaction between Factor VIII, phospholipid and human antibodies to Factor VIII was studied by a two-stage clotting assay, and by a fluid-phase immunoradiometric assay for Factor VIII clotting antigen (VIII C:Ag).In the two-stage thrombin generation assay, Factor VIII:C was rapidly destroyed by human antibodies, even in the presence of optimal phospholipid. However, preincubation of Factor VIII with phospholipid before addition of antibody protected the Factor VIII from inactivation, resulting in the production of much more thrombin.In assays of VIII C:Ag, pre-incubation of Factor VIII with phospholipid before addition of labelled antibody reduced the amount of detectable antigen. The reduction was greater with increasing phospholipid concentration, up to 60% of the original antigen being ‘lost’ at a total phospholipid concentration of around 250 μg/i.u.These results suggest that human antibodies to Factor VIII are directed largely at its phospholipid binding site. The protection of Factor VIII from inactivation by complexing with phospholipid could have important clinical applications in treatment of haemophiliacs with inhibitors.

Relevance of Thrombophilic Risk Factors on Clinical Phenotype in Patients with Haemophilia A.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3981-3981
Author(s):  
Manuela Krause ◽  
Anika Hahn ◽  
Ferrial Peyvandi ◽  
Inge Scharrer
Keyword(s):  
Factor Viii ◽  
Healthy Subjects ◽  
Clinical Phenotype ◽  
Treatment Center ◽  
Study Group ◽  
Haemophilia A ◽  
Bleeding Tendency ◽  
Factor Viii Activity ◽  
On Demand ◽  
Bleeding Episodes

Abstract Introduction: By assuming the hypothesis, that a high coinheritance of genetic haemostatic abnormalities influences the clinical phenotype in patients with coagulation disorders, we analysed the bleeding tendency in adult patients with haemophilia A. Patients: A total of 49 patients with haemophilia A (severe:20 patients, moderate:12 patients, mild:17 patients; median age:44 years, range:22–78 years) and 80 sex-matched healthy subjects (median age:26 years, range:16–58 years) were studied in our haemophilia treatment center. Two of this patients have an inhibitor against factor VIII. 10 of 49 patients receive the factor VIII replacement therapy on demand. In addition to factor VIII activity, FV G1691A mutation and the FII G20210A variant were investigated. Results: In our study the prevalence of FV G1691A was significantly higher among patients 6/49 (severe:5, moderate:1) than among control subjects 2/80 [12% vs 2.5%; p=0.03 OR 4.9]. We found no differences in the FII G20210A variant in 2/49 patient (moderate:1, mild:1) as compared to 2/80 among controls [4% vs 2.5%; p=0.62 OR 1.63 ]. In none of our patients both defects together were identified. Within the last year the onset of bleeding was not different in thrombophilic haemophilia patients (n=8) 64 bleeding episodes/year compared with non-carriers (n=41) 284 bleeding episodes/year [8 vs 7 bleeding episodes/year/patient; p=0.72; OR 1.15]. Conclusion: The FV G1691A was found significantly more frequent in the thrombophilic haemophiliacs than in the general population. In our smal study group we could not demonstrate a relevant influence of the FV G1691A and FII G20210A on bleeding tendency. Further studies are needed to confirm whether FV G1691A and FII G20210A plays a role on the influence of bleeding tendency in haemophilia A patients.

scholarly journals Comparison of platelet-derived and plasma factor VIII efficacy using a novel native whole blood thrombin generation assay

2015 ◽  
Vol 13 (12) ◽  
pp. 2210-2219 ◽  
Author(s):  
C. K. Baumgartner ◽  
G. Zhang ◽  
E. L. Kuether ◽  
H. Weiler ◽  
Q. Shi ◽  
...  
Keyword(s):  
Factor Viii ◽  
Whole Blood ◽  
Thrombin Generation ◽  
Thrombin Generation Assay ◽  
Plasma Factor

scholarly journals Thrombelastography (TEG) and Thrombin Generation Assay (TGA) in Severe Hemophilia Following Factor Replacement Therapy

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4666-4666 ◽  
Author(s):  
Tania T. Sarker ◽  
Donald Brophy ◽  
Meera B. Chitlur
Keyword(s):  
Factor Viii ◽  
Half Life ◽  
Thrombin Generation ◽  
Factor Activity ◽  
Treatment Frequency ◽  
Thrombin Generation Assay ◽  
Fviii Activity ◽  
The Mean ◽  
Life Factor ◽  
Time Point

Abstract Background: Monitoring therapy in hemophilia is a major challenge. Measurement of factor levels is time consuming and not available in time to make clinical decisions. With the introduction of extended half-life factor products, determination of treatment frequency becomes important. Global hemostatic assays such as Thrombelastography (TEG) and Thrombin Generation Assay (TGA) may improve monitoring. Focused toward individualizing therapy, these assays may help determine treatment frequency based not just on Factor VIII PK (pharmacokinetic), but also on total hemostatic potential. Objective: To determine the correlation between TGA and TEG parameters, and Factor activity and half-life (t1/2). Design/Methods: With IRB approval and participant consent baseline FVIII activity was obtained at enrollment, 15minutes, 1, 4, 8, 24 and 48 hours post factor replacement in patients who had not received replacement factor for a minimum of 72 hours and were not bleeding. FVIII:C, TEG, and TGA at each time point were measured. Non-compartmental PK analysis was performed on each individual patient profile to measure Factor VIII terminal half-life (t 1/2), mean normalized factor clearance rate and volume of distribution at steady-state (Vdss). Pearson correlation statistical analyses on other variables were performed using JMP ¨ Pro version 12.0.1 (SAS Institute, Cary, NC, USA) Results: 27 patients with hemophilia have enrolled, with a median age of 14 years (range: 2-24 years). 9 patients were eliminated from analysis because of a diagnosis of inhibitors (n=1), factor activity >1% (n=4), inadequate sample collection (n=2), patient on episodic factor replacement (n=1), and inaccurate TGA time point (n=1). The mean Factor level prior to factor administration, after elimination of the subjects (n=18) was 0.4%. As expected, our results indicate a rise in ETP and Factor activity following factor replacement, peaking at 15 minutes post infusion. The mean normalized factor clearance rate was 3.3 ± 1.2ml/h/kg. The Vdss was 2.3 ± 1 L and Factor VIII t½ was 11.5 ± 3 hours. There were strong correlations between ETP and FVIII:C (R2=0.65; p<0.0001), Peak and FVIII:C (R2=0.6; p<0.0001), R Time and Factor VIII:C (R2=0.71; p<0.0001), Peak and R Time (R2=0.59; p<0.0001), ETP and R Time (R2=0.51; p<0.0001) as shown in table 1. Table 1. Correlation data on Factor VIII:C with TGA & TEG Parameters; and TGA parameters with TEG R time R2 P-value TGA Parameters (Peak & ETP) ETP and Factor VIII:C 0.65 p<0.0001 Peak and Factor VIII:C 0.60 p<0.0001 TEG Parameter (R Time) R Time and Factor VIII:C 0.71 p<0.0001 TEG and TGA Parameters Peak and R Time 0.59 p<0.0001 ETP and R Time 0.51 p<0.0001 Conclusions: Global hemostatic assays are less expensive than traditional PK testing and are available at the time of care decisions. Results of global coagulation assays (TEG and TGA) correlated closely with FVIII activities. Global assays may predict breakthrough bleeding independent of factor levels, representing an improvement in monitoring over traditional PK. With the emergence of the bioengineered extended half-life factor products, there is a renewed interest in pharmacokinetic analysis and individualization of therapy. Assays like TEG provide the opportunity to receive feed back in real time that corresponds to FVIII activity, and enable us to make treatment decisions rapidly for each individual patient. Since these assays measure more than just the factor activity, the parameters such as ETP on TGA may be more prognostic of bleeding tendency, as has been shown previously. Pharmacokinetic and pharmacodynamics analysis of this data is ongoing. Our small sample size precludes us from making global predictions. Larger multi center trials would assist in confirming these findings. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Relationship between Thrombin Generation Assay and FVIII Levels in Patients with Mild to Moderate Haemophilia (A)

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2454-2454
Author(s):  
Pu-Lin Luo ◽  
Steven K. Austin ◽  
Kiran Parmar ◽  
Dan P Hart ◽  
Michael Laffan
Keyword(s):  
Research Funding ◽  
Thrombin Generation ◽  
Haemophilia A ◽  
Contact Activation ◽  
Extrinsic Pathway ◽  
Two Stage ◽  
One Stage ◽  
Thrombin Generation Assay ◽  
The Relationship

Abstract Introduction Haemophilia A (HA) phenotypes (mild, moderate and severe) are based on the baseline FVIII levels, however considerable variation in the bleeding phenotype exists between patients with similar FVIII level. Moreover, approximately 40% of patients with mild HA have large discrepancies between FVIII measured by one stage (FVIII:C1) and two stage methods (FVIII:Chr2) and it is unclear which method correlates best with in vivo FVIII function and bleeding phenotype. The Thrombin Generation assay (TGA), a global measure of haemostasis may be a better predictor of bleeding phenotype but pre-analytical factors such as contact activation can confound the results. Choice of initiating conditions may also be critical in determining sensitivity: recent studies have suggested that initiation with FIXa rather than tissue factor (TF) in detecting low levels of FVIII:C in severe HA, however its utility in mild to moderate HA patients has yet to be determined. The aim of this study is to establish the relationship between FVIII:C and TGA and the influence of contact factor activation in TF and FIXa triggered TGA in patients with mild to moderate HA. Methods This is a prospective cohort study. Patients aged >18 with known congenital HA and FVIII:C 0.01- 0.2 iu/ml were recruited from 3 Haemophilia Comprehensive Care Centres in London. Peripheral blood was drawn into citrate Vacutainer tubes containing 0.106M trisodium citrate (1:9 volume) and Vacutainer tubes preloaded with CTI (50µg/ml). Samples underwent double centrifugation (2500g) to obtain platelet free plasma. Thrombin generation assay, using a standard calibrated automated thrombogram was triggered with either TF (1pmol) or FIXa (5nM). Factor FVIII levels were assessed by one stage APTT based (FVIII:C1) and two stage chromogenic (FVIII:Chr2) methods. Mutation analysis was carried out in all patients. Results 40 patients were recruited in the study. Five patients (13%) had standard FVIII discrepancy (FVIII:C1/FVIII:Chr2>1.5) with 4 different FVIII mutations located on the inter-domain surface of the A2 domain (p.Tyr683Ser, p.Arg550Cys, p.Gly498Arg, p.MET681.Le). One patient had reverse FVIII discrepancy. In TF triggered TGA, the presence of CTI resulted in significant reduction in mean ETP (nmol .min)(455. vs 278, p<0.01, 95% CI 104-243), mean Peak thrombin (nM) (37.81 vs 16.54, t(6.6) p<0.01 95%CI 14.7-27.3), and mean Velindex (nM/min) (4.86 vs 1.29 t(7.0), p<0.01, 95% CI2.3-4.19) and a longer mean ttPeak (min) (14.26 vs 16.22, t(-3.2) p=0.02 95% CI-3.1- -0.76). In contrast, the presence of CTI did not affect ETP (1143 vs 1042, p=0.19 95% CI -54-256), mean Peak thrombin (252 vs 251, p=0.6 95%CI 27-46) or Velindex (118.54 vs 119.15 p= 0.95, 95%CI -23-12.9) in FIXa triggered TGA. There was a good correlation between FVIII:Chr2 and ETP (r=0.56, p=<0.001) Peak (r=0.6, p=<0.001) and Velindex (r=0.7, p=<0.001) in TF(CTI-) triggered TGA, however no relationship was seen between FVIII:C and TG parameters (ETP r=-0.01 p=0.9, Peak r=-0.003, p=0.97 and Velindex r=0.018, p=0.9) in TF(CTI+) triggered TGA. In both FIXa(CTI-) and FIXa (CTI+) triggered TGA, there was a good correlation seen between Lagtime (r=-0.6 p=<0.01), Peak (r=0.4-0.6, p=<0.01) ttpeak (r= -0.6, p=<0.01) and Velindex (r=0.69 <0.01) with FVIII:Chr2 but not with ETP. In patients with standard FVIII discrepancy (n=5), their ETP and Peak levels in TF and FIXa triggered TGA were in keeping with the ETP and Peak levels of non-discrepant patients with similar FVIII:C2 and significantly lower than that of non-discrepant patients with similar FVIII:C1. Conclusions Our study confirms that at low TF triggered TG, contact factor activation in vitro is an important preanalytical variable. Curiously any TG correlation with FVIII level is lost once the contact pathway is inhibited suggesting that TG remains largely determined by the extrinsic pathway in this system. In contrast, factor FIXa triggered TG is unaffected by inhibition of contact activation and demonstrates a good correlation to FVIII:C with or without CTI. This can be explained by suggesting that the supply of FIXa negates any effect of XIa from contact activation and that TG by this route is more completely dependent on FVIII. Therefore a FIXa triggered TGA may offer a better alternative in the assessment of haemophilia and further studies are underway to determine whether this is a better predictor of bleeding phenotypes. Disclosures Luo: Pfizer: Research Funding. Austin:Pfizer: Research Funding. Laffan:Pfizer: Honoraria; Roche: Consultancy, Speakers Bureau.

Thrombin Generation Assay: Use in Monitoring Treatment of Patients with Coagulation Factor Inhibitors.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3087-3087
Author(s):  
Aine N. McCormick ◽  
Peter L. Turecek ◽  
Katalin Varadi ◽  
Geoffrey F. Savidge
Keyword(s):  
Factor Viii ◽  
Thrombin Generation ◽  
Coagulation Factor ◽  
Rational Approach ◽  
Factor Viii Inhibitor ◽  
Specific Time ◽  
Time Intervals ◽  
Monitoring Method ◽  
Standard Curve ◽  
Thrombin Generation Assay

Abstract Treatment of severe Haemophilia A relies upon the infusion of plasma derived or recombinant Factor VIII concentrates. The development of antibodies to these products is approximately 25–30% in severe cases and remains a major clinical challenge to management. Treatment in inhibitor cases with high antibody levels is based on the effect of products that bypass Factor VIII in the cascade and activate the common pathway either directly or through the extrinsic pathway. These include recombinant Factor VIIa and Factor VIII inhibitor bypassing activity (FEIBA®). At present, the use and dosage of bypassing products is severely constrained by the current inability to measure directly their beneficial effect using a standardised assay, and clinical management rests exclusively upon clinical assessment. The thrombin generation assay (TGA) provides a convenient and reproducible method for quantifying thrombin produced following activation of the cascade, and is measured by means of monitoring a fluorescent residue that arises following cleavage of an artificial thrombin substrate. Through sequential measurements quantifiable kinetic data may be collated and assessed. We have used the TGA to assess thrombin generation in inhibitor patients using samples taken at specific time intervals from patients after administration of FEIBA (n=4) or rFVIIa (n=3) following informed consent. In a number of experiments, platelets from the patient’s pre-infusion platelet rich plasma (PRP) were added. Peakthrombin (nM), peaktime (min) and maximum initial rates of thrombin generation (nM/min) from each sample were quantified, and the maximum initial rates of thrombin generation were further processed using PKAnalyst® Pharamacokinetic Data Analysis software program to define one and two PK compartment elimination models following bolus injection. Thrombin generation parameters were found to persist at levels greater than those measured for the pre infusion samples for periods beyond 6 hours post FEIBA and for 2 hours post rFVIIa, prior to reinfusion of rFVIIa. In PK evaluation studies FEIBA was found to have a relative thrombin generation T1/2 of 1–3.7 hr using the one compartment model, while rFVIIa was found to have a relative thrombin generation T1/2 of 0–2.8 hr following a single infusion. A second infusion of rFVIIa gave a relative thrombin generation T1/2 of 1.2–1.8 hr suggesting a cumulative effect. The addition of platelets from pre-infusion PRP to the assay enhanced thrombin generation by approximately 30% in all samples studied. The TGA was also used to quantify the thrombin generation produced by FEIBA and rFVIIa in samples obtained at specific time intervals post treatment, using a pre infusion sample spiked with a range of FEIBA/rFVIIa concentrations to generate a standard curve. This standard curve was then used to estimate the relative FEIBA/rFVIIa concentrations in the post infusion samples with and without the addition of platelets. We consider that the use of the TGA in this clinical setting demonstrates its value as a monitoring method highly effective for providing a rational approach to dosage adjustments of FEIBA/rFVIIa in the treatment of patients with FVIII inhibitors. This valuable analytical means for monitoring FVIII bypassing treatment is of considerable relevance in relation to surgery and the management of acute bleeds in this patient category.

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