Increased Thrombin Generation in Severe Hemophiliacs with Mild Clinical Phenotype

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 515-515
Author(s):  
Elena Santagostino ◽  
Maria Elisa Mancuso ◽  
Armando Tripodi ◽  
Veena Chantarangkul ◽  
Gianluigi Pasta ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Clinical Phenotype ◽  
Platelet Rich Plasma ◽  
Gene Mutations ◽  
Clinical Score ◽  
Bleeding Tendency ◽  
Spontaneous Bleeding ◽  
Chromogenic Assay ◽  
Bleeding Episodes ◽  
The Impact

Abstract Introduction: Some severe hemophiliacs (FVIII/FIX<1%) exhibit a mild bleeding tendency, but the basis for this clinical heterogeneity is poorly understood. This study investigated the relationship between the values of endogenous thrombin potential (ETP) and clinical phenotype in severe hemophiliacs. The impact of FVIII/FIX gene mutations and thrombophilic polymorphisms was also evaluated. Methods: severe hemophiliacs older than 18 years without inhibitor history and treated on demand were eligible. Mild bleeders (MB) and severe bleeders (SB) were defined as follows: spontaneous bleeding episodes per year ≤2 (MB) or 25 (SB) and concentrate consumption <500 (MB) or >2000 (SB) IU/Kg/year. Patients who did not fit these criteria were considered as intermediate bleeders (IB). FVIII was measured by chromogenic assay and ETP was measured in platelet-rich plasma after addition of tissue factor. Results: 22MB, 22SB and 28IB were enrolled. MB had lower clinical and radiological scores when compared with both IB and SB (p<0.005). MB showed an older age at first bleed compared to SB (p < 0.005) and p for trend among the 3 groups was also significant (p < 0.05). The prevalence of severe FVIII/FIX gene defects (null mutations) was lower and ETP values were higher in MB compared with both IB and SB (p<0.05; table 1). Conclusions: our results indicate an extremely low prevalence of null mutations in severe hemophiliacs with mild bleeding diathesis. The measurement of thrombin generation in platelet-rich plasma may allow to identify this subgroup of patients, not otherwise distinguishable by conventional functional assays. SB (#22) IB (#28) MB (#22) p Age (yr) 38 (21–76) 38 (23–62) 32 (22–73) NS Age 1st bleed (yr) 1 (0–4) 2 (0–6) 3 (1–10) < 0.005 Bleeding episodes/yr 36 (25–60) 10 (3–20) 0 (0–2) < 0.0005 Factor use (IU/Kg/yr) 2207 (2040–8696) 1068 (207–2400) 60 (25–487) < 0.0005 Clinical score 18 (10–35) 10 (0–34) 3 (0–17) < 0.005 Pettersson score 44 (14–62) 28 (0–48) 17 (3–40) < 0.0005 Null mutations (%) 59 70 6 < 0.005 PTG20210A (%) 0 7 5 NS FV Leiden (%) 5 7 0 NS Median ETP (nM) 414 478 850 < 0.05

Thrombin Generation in Severe Hemophiliacs with Different Clinical Phenotype.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1154-1154
Author(s):  
Elena Santagostino ◽  
Maria E. Mancuso ◽  
Armando Tripodi ◽  
Veena Chantarangkul ◽  
Mariagrazia Clerici ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Clinical Phenotype ◽  
Platelet Rich Plasma ◽  
Gene Mutations ◽  
Bleeding Tendency ◽  
Spontaneous Bleeding ◽  
Chromogenic Assay ◽  
Bleeding Episodes ◽  
Fv Leiden ◽  
The Impact

Abstract Introduction: Some severe hemophiliacs (FVIII/FIX<1%) exhibit a mild bleeding tendency, but the basis for this heterogeneous clinical expression is poorly understood. This study investigated the relationship between the values of endogenous thrombin potential (ETP) and clinical phenotype in severe hemophiliacs. The impact of FVIII/FIX gene mutations and thrombophilic polymorphisms into the modulation of the phenotype was also evaluated. Methods: severe hemophiliacs older than 18 years without inhibitor history and treated on demand were eligible for inclusion in the study. Mild bleeders (MB) and severe bleeders (SB), representing the extremes of the clinical spectrum, were defined according to the following criteria: spontaneous bleeding episodes per year ≤2 (MB) or ≥25 (SB) and concentrate consumption <500 (MB) or >2000 (SB) IU/Kg/year. Patients who did not fit these criteria were considered as intermediate bleeders (IB). Plasma samples were obtained after a minimum wash-out period of 5 days. FVIII was measured by chromogenic assay. ETP was measured in platelet-rich plasma after addition of tissue factor. Results: 22MB, 22SB and 28IB were enrolled. MB showed an older age at first bleed compared to SB (p < 0.005) and p for trend among the 3 groups was also significant (p < 0.05). The prevalence of severe FVIII/FIX gene defects (null mutations) was extremely low in MB (6%). ETP values were higher in MB compared with both IB and SB (p<0.05); p for trend among the 3 groups was also significant (p <0.05). Conclusions: this study shows that the measurement of thrombin generation in platelet-rich plasma may allow to identify patients with mild bleeding diathesis among severe hemophiliacs, in contrast with the features of conventional functional assays. SB (#22) IB (#28) MB (#22) P Age (yr) 38 (21–76) 38 (23–62) 32 (22–73) NS Age 1st bleed (yr) 1 (0–4) 2 (0–6) 3 (1–10) < 0.005 Bleeding episodes/yr 36 (25–60) 10 (3–20) 0 (0–2) < 0.0005 Factor use (IU/Kg/yr) 2207 (2040–8696) 1068 (207–2400) 60 (25–487) < 0.0005 Null mutations (%) 59 70 6 < 0.005 PTG20210A (%) 0 7 5 NS FV Leiden (%) 5 7 0 NS Median ETP (nM) 414 478 850 < 0.05

Thrombin Generation in Severe Hemophiliacs with Different Clinical Phenotype.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1000-1000 ◽  
Author(s):  
Elena Santagostino ◽  
Maria E. Mancuso ◽  
Armando Tripodi ◽  
Veena Chantarangkul ◽  
Marigrazia Clerici ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Clinical Phenotype ◽  
Bleeding Tendency ◽  
Bleeding Diathesis ◽  
Severe Hemophilia ◽  
Spontaneous Bleeding ◽  
Radiological Score ◽  
Bleeding Episodes ◽  
Fv Leiden ◽  
The Impact

Abstract Introduction: based on the plasma levels of factor VIII or IX (FVIII, FIX) hemophiliacs usually experience bleeding episodes of different clinical relevance. Some hemophiliacs classified as severe, having undetectable coagulant activity of FVIII or FIX (FVIII/FIX:C &lt;1 U/dL), exhibit a mild bleeding tendency, but the basis for this heterogeneous clinical expression of the disease is still poorly understood. The thrombin generation measured as endogenous thrombin potential (ETP) in patients with hemophilia A has shown a significant correlation with FVIII levels while a wide range of ETP values were found in patients with FVIII:C &lt;1 U/dL (Chantarangkul et al. Haematologica88: 547–54, 2003) suggesting the potential for distinguishing different coagulation profiles among severe hemophiliacs. Objectives: this case-control study was designed to investigate the relationship between ETP values and clinical phenotype in severe hemophiliacs, in order to evaluate the use of this measurement as an indicator of the bleeding tendency. The impact of thrombophilic mutations PTG20210A and FV Leiden into the modulation of the hemophilia phenotype was also evaluated. Patients: severe hemophiliacs (FVIII/FIX:C &lt; 1 IU/dL) older than 18 years, without a history of inhibitors and treated on demand were eligible. Two groups of patients with severe hemophilia, representing the extremes of the clinical spectrum of bleeding symptoms, were defined. Patients with extremely mild bleeding diathesis (mild bleeders, MB) should have had 2 or less spontaneous bleeding episodes per year; a concentrate consumption lower than 120 U/Kg/year; a total radiological score ≤ 10 points and a total orthopaedic score ≤ 4 points (scoring system recommended by the Orthopaedic Advisory Committee of the World Federation of Hemophilia). Patients with markedly severe bleeding history (severe bleeders, SB) should have had 25 or more spontaneous bleeding episodes per year; a concentrate consumption greater than 3000 U/Kg/year; a total radiological score ≥ 40 points and a total orthopaedic score ≥ 25 points. Patients who did not fit the definition criteria of neither MB nor SB were considered as “intermediate” bleeders (IB). Methods: plasma samples were obtained after a minimum wash-out period of 5 days. FVIII levels were measured by chromogenic assay. ETP was measured in platelet-rich plasma after addition of tissue factor. Results: 22 MB, 22 SB and 28 IB were enrolled. No statistically significant differences were found in the age of patients included in the three groups (median: 32, 38 and 38 years). PTG20210A was detected in 5% MB and 4% IB; FV Leiden in 7% IB and 5% SB. ETP values were significantly higher in MB (median: 850 nM) compared with both IB (median: 478 nM; p&lt;0.05) and SB (median: 414 nM; p&lt;0.05). Also p for trend was statistically significant (p&lt;0.05). Conclusions: this study shows that thrombin generation measurements may allow to identify patients with mild bleeding diathesis among severe hemophiliacs, in contrast with the features of conventional functional assays. On this basis, ETP measurement in the setting of severe hemophilia may be useful to improve the selection of candidates for early prophylaxis.

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 One-Stage FVIII Activity Is Inversely Correlated with Heavy Menstrual Bleeding in Hemophilia a Carriers

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3542-3542
Author(s):  
Robert F. Sidonio ◽  
Shannon L. Meeks ◽  
Hilary Baker Whitworth
Keyword(s):  
Thrombin Generation ◽  
Hemophilia A ◽  
Menstrual Bleeding ◽  
Bleeding Tendency ◽  
One Stage ◽  
Chromogenic Assay ◽  
Fviii Activity ◽  
Coagulation Assay ◽  
Bleeding Score ◽  
Risk For Bleeding

Abstract Introduction: The majority of carriers of hemophilia A have historically been considered to have normal hemostasis (FVIII:C >50%) and thus not have an increased bleeding tendency. However, our research group has demonstrated that some hemophilia A carriers can experience increased bleeding compared to normal women despite this normal FVIII activity. In our recently published study in adult hemophilia A carriers there was no correlation between FVIII:C, as measured by a one-stage coagulation assay, and bleeding phenotype, as measured by the MCMDM-1 VWD bleeding score. In this follow up study we sought to determine which of these hemophilia A carriers with normal FVIII:C are at risk for bleeding. The goal of this study is to determine the relationship between FVIII assays (one-stage and chromogenic) and thrombin generation with both menstrual bleeding and an overall bleeding tendency. Methods: We recruited mothers of children with hemophilia A in the Emory University pediatric bleeding disorders clinic. We included only adult (>18 year of age) obligate hemophilia A carriers that did not have a concomitant bleeding disorder or chronic disease that would increase their bleeding tendency. We gathered basic demographic information and evaluated the overall bleeding tendency using the ISTH Bleeding Assessment Tool (ISTH BAT), a semi-quantitative assessment of bleeding scored from 0 to 56. We considered a score of 6 or higher consistent with pathologic bleeding. In addition, we inventoried menstrual bleeding with the Pictorial Bleeding Assessment Chart (PBAC), a visual representation of menstrual blood loss. We considered a PBAC greater than 100 consistent with heavy menstrual bleeding. Plasma samples were collected from each subject at the time of the survey in sodium citrate tubes without corn trypsin inhibitor. Samples were double spun at 2,500 rpm for 15 minutes and stored at -80°C.Laboratory evaluation for each subject included FVIII:C, measured by one-stage coagulation assay (aPTT reagent with micronized silica) and chromogenic assay (COATEST SP4 FVIII kit, Chromogenix), as well as thrombin generation (Calibrated Automated Thrombogram, PPP-Reagent Low, FluCa Kit) to evaluate endogenous thrombin potential (ETP, nM*min), peak thrombin (nM) and lag time (min). We performed linear regression using GraphPad Prism; p<0.05 was considered significant. Results: Over a three-month period, we approached 32 adult obligate hemophilia A carriers; 23 agreed to participate in the survey, 16 consented to blood draw. One carrier was excluded in extreme outlier analysis and due to an autoimmune disorder, leaving 15 evaluable subjects. Our cohort is relatively young with a median age of 41 years (range 23-51), predominantly Caucasian (53%, 8/15) and the majority carry a severe mutation (10/15 severe, 2/15 moderate, 3/15 mild). Reproductive bleeding (post-partum hemorrhage and/or menstrual bleeding) was commonly reported (93%, 14/15). The median ISTH BAT bleeding score for subjects was 2 (range 1-8). The median PBAC score was 148 (range 10-608). The carriers had a relatively normal FVIII:C by one-stage assay with a median of 0.78 U/mL (range 0.30 - 1.06) and by chromogenic assay with a median of 1.15 U/mL (range 0.66 - 1.9). FVIII:C by one-stage assay was inversely correlated to PBAC (r2 =0.4; p=0.01, figure 1A). Conversely, FVIII:C by chromogenic assay did not correlate with PBAC, however trended toward significance (r2 = 0.19; p=0.10, figure 1A). Additionally, there was no correlation found between FVIII:C (one-stage or chromogenic assay) and ISTH BAT bleeding score (figure 1B). There was also no correlation found between bleeding score or PBAC and the parameters of the thrombin generation assay or between severity of the closest male relative's hemophilia and any of the assays performed. Conclusions: In our cohort of adult obligate hemophilia A carriers, as the FVIII:C (one-stage assay) decreased, the menstrual bleeding tendency increased as measured by the PBAC. We were unable to find a correlation between other measures of hemostasis, including the chromogenic assay and thrombin generation, and commonly used bleeding assessment tools. Further larger studies are warranted to help determine which hemophilia A carriers would be at risk for bleeding. Disclosures No relevant conflicts of interest to declare.

Coagulation Functions with Clinical Phenotype and Inhibitory Mechanisms in Acquired Factor V Inhibitors.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2232-2232
Author(s):  
Tomoko Matsumoto ◽  
Keiji Nogami ◽  
Kenichi Ogiwara ◽  
Midori Shima
Keyword(s):  
Light Chain ◽  
Thrombin Generation ◽  
Clinical Phenotype ◽  
Waveform Analysis ◽  
Severe Bleeding ◽  
Functional Evaluation ◽  
Factor V ◽  
Bleeding Tendency ◽  
Coagulation Function ◽  
Apc Resistance

Abstract Abstract 2232 Development of acquired factor V (aFV) inhibitors rarely occurs, but its clinical phenotype varies from asymptomatic to life-threatening bleeding. A recent systematic report describes that little bleeding symptom is present in 20% at the diagnosis for patients with acquired FV (aFV) inhibitors. However, the coagulation function and its mechanism(s) on the different clinical phenotype are poorly understood. In this study, we examined the coagulation function on aFV inhibitors by using comprehensive coagulation assays, thrombin generation test (TGT) and clot waveform analysis (CWA). TGT was performed using tissue factor (0.5 pM), phospholipid (PL; 4 μM) and ellagic acid (0.3 μM). CWA, that evaluates the parameters of min1 as maximum coagulation velocity and min2 as maximum coagulation acceleration, was performed on MDA-II® system. We tested 7 cases with aFV inhibitors. Four cases were asymptomatic (FV:C; 3.6±3.4 IU/dl, inhibitor 5.8±3.3 BU/ml: non-B group), and 3 cases had severe bleeding tendency (2.9 ± 4.5 IU/dl, 66 ±51 BU/ml; B group). In TGT, all cases in both groups little showed the thrombin generation within 60 min, independently of FV:C level and clinical phenotype, showing little informative in functional evaluation for aFV inhibitors. However, in a PT-based CWA, the clotting time observed in non-B group was markedly shorted compared to that in B group (62.2±17.0/112±15 sec; p=0.006). In addition, both parameters in non-B group were significantly greater than those in B group ( min1 ; 2.89±1.10/0.98±0.29 dT/dt; p=0.014) and ( min2 ; 0.75±0.40/0.15±0.07 d2T/dt2; p=0.028), suggesting that CWA was useful for the prediction and monitoring of hemorrhagic symptoms in patients with aFV inhibitors. To confirm the distinct mechanism(s) on both groups, the IgGs from aFV plasmas were immune-purified using protein G-Sepharose. In the reactant mixtures with normal plasma and aFV IgGs, all parameters obtained in CWA were similar to those obtained in patients' plasmas. SDS-PAGE and western blotting revealed that 2 cases in B group reacted the light chain of FV(a). However, 2 cases in non-B group reacted the heavy chain, and other 2 cases were reacted with both chains (heavy>light), indicative of the distinct epitopes of IgGs in both groups. Since the light chain contains the PL-binding site(s), the effects of aFV IgGs were examined on the FV-PL binding in an ELISA. All IgGs in B group inhibited this binding (by 40–90%) dose-dependently, whilst little affected in non-B group. Since FV acts as a cofactor of activated protein C (APC) on inactivation of FVIIIa, the effects of aFV IgGs on the ability of APC on FVIIIa inactivation were examined using intrinsic FXa generation assay. The APC sensitivity ratio (APCsr) was expressed as ratio of amounts of generated FXa in the absence of APC relative to its presence. A low level of APCsr indicates the reduction in FVIIIa inactivation, consequently APC resistance. APCsr values in B group were >2.0 within normal range, whilst those in non-B group were decreased to 1.5, supportive of APCR in non-B group. Based on these findings, we propose that severe bleeding tendency in B group would be appeared through negligible prothrombinase activity, since aFV IgGs blocked the FV(a)-PL binding. While, clinical phenotype in non-B group would be asymptomatic, since aFV IgGs unaffect the FV(a)-PL binding and further cause the APC resistance. In addition, various clinical phenotypes in aFV inhibitors appear to be dependent on the recognizing epitope of these IgGs. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The Irish Personalized Approach to the Treatment of Haemophilia (iPATH) - Determinants of Inter-Individual Variation in FVIII Pharmacokinetics

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1190-1190
Author(s):  
Michelle Lavin ◽  
Niamh Larkin ◽  
Anjali Patel ◽  
Evelyn Singleton ◽  
Mary Byrne ◽  
...  
Keyword(s):  
Bayesian Analysis ◽  
Blood Group ◽  
Research Funding ◽  
Clinical Phenotype ◽  
Employment Equity ◽  
Limited Sampling ◽  
Bleeding Episodes ◽  
Equity Ownership ◽  
The Impact ◽  
Antihemophilic Factor

Abstract Introduction: Standard factor VIII (FVIII) prophylaxis aims to minimise bleeding episodes in people with haemophilia A (PWHA) using a weight based dosing strategy. However, it is well recognised that FVIII half-life (FVIII t1/2) varies markedly between individual PWHA. Consequently, some PWHA continue to experience repeated bleeding episodes even after commencing standard dose prophylaxis therapy. In essence, these patients 'bleed their way' to eventually reaching their required optimal FVIII prophylactic regimen. Given that even a small number of joint bleeds have been shown to represent a risk factor for hemophilic arthropathy, there is a clear unmet need for the development of personalised treatment approaches for PWHA that include consideration of individual FVIII pharmacokinetics (PK) to guide prophylaxis dosing. With the introduction of limited plasma FVIII sampling and Bayesian analysis, evaluation of FVIII pharmacokinetics (PK) are now possible in routine clinical practice. However, real life clinical experience to date has been limited to small cohorts of PWHA. Critically, the influence of PK in modulating clinical phenotype has not been systematically studied. As part of the Irish Personalised Approach to the Treatment of Haemophilia (iPATH) study, we integrated extensive clinical data and individual PK profiles to investigate modulators of FVIII PK, the role of limited versus extended sampling FVIII PK and the impact of PK on FVIII prophylaxis and clinical phenotype. Methods: Written informed consent was obtained prior to recruitment to the iPATH study. All PK samples were obtained following administration of a single rFVIII product (antihemophilic factor [recombinant]; ADVATE®, Shire, Lexington, MA, USA) and using myPKFiT™ software (Shire, Lexington, MA, USA). Utilising limited sampling (2 timed FVIII:C levels, taken at 3‒6 and 24‒32 hours post rFVIII administration) and Bayesian analysis, PK curves were constructed using myPKFiT™. For each individual the FVIII t1/2, Von Willebrand Factor antigen (VWF:Ag), blood group, weight, age at PK and Haemophilia Joint Health Score (HJHS) was recorded. rFVIII prescribed over the preceding 5 years was analysed; all patients used antihemophilic factor (recombinant) throughout this time period. Mann-Whitney analyses were used for non-parametric comparisons and correlations were performed using the Pearson test on Prism 7.0c for Mac OSX. Results: Complete PK profiles were available for 53 patients with FVIII deficiency recruited to the iPATH study (3 moderate, 50 severe FVIII deficiency). FVIII t1/2 varied considerably, ranging from 7.7‒20.1 hours (median=11.4 hours). Patients with blood group O had a significantly shorter FVIII t1/2 (n=31, median=10.9 hours) than non-O patients (n=22, median=12.2 hours, p=0.014). Both increased age (r2=0.2932, p<0.0001) and higher plasma VWF:Ag levels (r2=0.3, p<0.0001) at time of PK were associated with a significantly longer FVIII t1/2. HJHS significantly increased with age (r2=0.494, p<0.0001) but no significant correlation between FVIII t1/2 and HJHS was observed, likely reflecting the multifactorial nature of arthropathy in PWHA. In a subset of 10 PWHA we compared the FVIII t1/2 results generated from extended (10 point) to those from limited sampling (2 point) PK. Each limited sampling study included a 24 hour time point as well as either a 3, 4 or 6 hour sample. The 10 point results significantly correlated with limited sampling results using either the 3/24 hour (r2=0.9801, p<0.0001), 4/24 hour (r2=0.9762, p<0.0001) or 6/24 hour sampling combination (r2=0.9693, p<0.0001), underscoring the clinical utility of limited PK sampling. Prior to the introduction of routine PK-guided dosing in 2018, prescribed rFVIII dose was altered according to bleed rate at clinical review. Analysing the prescribed rFVIII dose over the past 5 years, a clear relationship to rFVIII t1/2 was identified (r2=0.3517, p<0.0001), suggesting that bleeding events in PWHA guided them towards their PK dose. Conclusions: This study highlights the accuracy of simplified vs extended sampling for generation of PK profiles on antihemophilic factor (recombinant) using myPKFiT™. Bleed rate adjusted prophylaxis was found to correlate with FVIII t1/2 in an Irish population of PWHA, suggesting that early evaluation of PK and implementation of PK guided dosing of prophylaxis may be of benefit in minimising bleeds in PWHA. Disclosures Lavin: Shire: Honoraria, Research Funding, Speakers Bureau. Reipert:Shire: Employment, Equity Ownership. Pipe:Shire: Consultancy, Research Funding; HEMA Biologics: Consultancy; Pfizer: Consultancy; Nove Nordisk: Consultancy; Bioverativ: Consultancy; CSL Behring: Consultancy; Genentech: Consultancy; Ainylam: Consultancy; F. Hoffmann-La Roche Ltd: Consultancy; Apcintex: Consultancy; Catalyst Biosciences: Consultancy; Spark Therapeutics: Consultancy; Freeline: Consultancy; Bayer: Consultancy; uniQure: Consultancy; Biomarin: Consultancy; R2 Diagnostics: Research Funding; Siemens: Research Funding. Turecek:Shire: Employment, Equity Ownership. O'Donnell:Bayer: Research Funding, Speakers Bureau; Novo Nordisk: Research Funding, Speakers Bureau; Leo Pharma: Speakers Bureau; Octapharma: Speakers Bureau; CSL Behring: Consultancy; Daiichi Sankyo: Consultancy; Pfizer: Consultancy, Research Funding; Baxter: Research Funding, Speakers Bureau; Shire: Research Funding, Speakers Bureau.

scholarly journals Platelets as Modifiers of Clinical Phenotype in Hemophilia

2006 ◽  
Vol 6 ◽  
pp. 661-668 ◽  
Author(s):  
Donald L. Yee
Keyword(s):  
Vascular Injury ◽  
Thrombin Generation ◽  
Clinical Phenotype ◽  
Phenotypic Variability ◽  
Fibrin Clot ◽  
Bleeding Tendency ◽  
Related Factors ◽  
Plasma Factor ◽  
Life Threatening ◽  
Biologic Factors

Platelets occupy a central role in the maintenance of hemostasis by adhering to sites of vascular injury and facilitating thrombin generation, which leads to the formation of a fibrin clot. Patients with hemophilia exhibit defective thrombin generation secondary to reduced plasma factor concentrations, which can lead to excessive and sometimes life-threatening bleeding. Individuals differ greatly with respect to platelet function and platelets from different individuals differ inherently in their ability to enact thrombin generation, the key coagulative process that is deficient in hemophilia. Similarly, some patients with hemophilia seem to bleed less often than others despite exhibiting similar plasma factor levels. The biologic factors that underlie this phenotypic variability remain poorly understood, but evidence is reviewed supporting a role for platelets and platelet-related factors in modifying bleeding tendency in patients with hemophilia and potential directions for further clinical research in this area are discussed.

scholarly journals A Chromogenic Test to Determine the Procoagulant Phospholipids in Platelet-rich Plasma and Whole Blood

1994 ◽  
Vol 72 (04) ◽  
pp. 582-587 ◽  
Author(s):  
R J Wagenvoord ◽  
H H Hendrix ◽  
Hu Kai ◽  
H C Hemker
Keyword(s):  
Whole Blood ◽  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
Product Formation ◽  
Phosphatidyl Serine ◽  
Linear Increase ◽  
Control Group ◽  
Chromogenic Substrate ◽  
Chromogenic Assay ◽  
Two Parameters

SummaryWe have developed a chromogenic assay to measure the phospholipid-related procoagulant activity (PPA) in whole blood, or platelet-rich plasma. The test is based upon thrombin formation from prothrombin by prothrombinase and is designed in such a way that procoagulant lipids are rate limiting for the prothrombinase activity. In the chromogenic test PPA concentrations equivalent to 0-10 nM phospholipid vesicles containing 75% phosphatidyl choline (PC) and 25% phosphatidyl serine (PS) can be measured.The thrombin, which develops during the test, is measured with a chromogenic substrate. By the action of thrombin on this chromogenic substrate p-nitroaniline is liberated, which causes an increase in absorbance. Thrombin formed in the assay mixture activates the present platelets. This causes a linear increase of the velocity of thrombin generation during the test, i. e. a parabolic increase of product formation. For that reason the thrombin generation in time is characterized by two parameters, the basal PPA (PPA-B) of the original mixture and the increase in PPA due to platelet activation (PPA-A). To determine these figures the absorbency-data were fitted to parabolas. In most cases the contribution of PPA-A to the total amount of formed thrombin becomes considerable already after 30 s.Preliminary tests show that PPA-B activity in whole blood or platelet-rich plasma of patients with a thrombotic disorder is significantly higher than the activity of a control group of the same age.

Thrombin Generation Assessment in Platelet Rich Plasma Offers Additional Criteria for Low Molecular Weight Heparin Antithrombotic Fingerprint Characterization.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4177-4177
Author(s):  
Grigorios T. Gerotziafas ◽  
Galea Vasso ◽  
Jeanine M. Walenga ◽  
Evi Kalodiki ◽  
Mourad Chaari ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Physicochemical Properties ◽  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
Inhibitory Effect ◽  
Low Molecular Weight ◽  
Generation Process ◽  
Health Authorities ◽  
The Impact

Abstract Abstract 4177 Introduction National and international organizations recognize that low molecular weight heparins (LMWHs) are distinct entities and that they should not be used interchangeably in clinical practice. Physicochemical properties, such as molecular weight anti-Xa/anti-IIa ratios, are used by health authorities to establish LMWHs differentiation. LMWHs are multitargeted drugs in which small differences in physicochemical properties may lead to significantly different inhibition of blood coagulation. In the present in vitro study we have investigated if thrombin generation assay, reflecting the overall coagulation process, is a suitable tool for the LMWH variability evalution. Materials and methods LMWHs (bemiparin, enoxaparin, nadroparin, dalteparin and tinzaparin) and unfractionated heparin (UFH) were added in vitro in normal citrated platelet rich plasma from 10 healthy individuals, at clinically relevant concentrations (ranging from 0.2 to 1 anti-Xa IU/ml). Thrombin generation was studied with Thrombogram-Thrombinoscope® assay using diluted thromboplastin (Innovin®, 1/1000 final dilution, Siemens France). The concentrations doubling the lag-time, the time to Peak, the IC50 for Peak, the endogenous thrombin potential (ETP) and the mean rate index of propagation phase (MRI) were determined for each compound. Results LMWHs compared on their anti-Xa activity basis showed variable inhibitory effect on thrombin generation. At equivalent anti-Xa concentrations tinzaparin was significantly more potent than the other LMWHs, being almost similar to UFH profile. Enoxaparin, nadroparin and dalteparin showed a similar inhibitory activity. Bemiparin had the lesser pronounced inhibitory effect on thrombin generation. The impact of anti-Xa and anti-IIa activities on each phase of thrombin generation process is different. Thrombogram chronometric parameters are mainly influenced by the anti-IIa activity. Similarly, ETP inhibition depends more on anti-IIa rather than anti-Xa activity. The MRI of the propagation phase is more sensitive to the anti-Xa activity of LMWHs. Conclusion Thrombin generation assessment in platelet rich plasma allows to evaluate the anticoagulant fingerprint of LMWHs and to differentiate them on global and functional criteria. Each LMWH has a particular inhibitory impact on each phase of thrombin generation process. These functional criteria need to be standardized and probably required for a better characterization of LMWHs heterogeneity by health authorities. They could be used also to evaluate bioequivalence of generic and original LMWHs. Disclosures: No relevant conflicts of interest to declare.

A central role of GPIb-IX in the procoagulant function of platelets that is independent of the 45-kDa GPIbα N-terminal extracellular domain

Blood ◽  
2010 ◽  
Vol 116 (7) ◽  
pp. 1157-1164 ◽  
Author(s):  
Catherine Ravanat ◽  
Catherine Strassel ◽  
Béatrice Hechler ◽  
Simone Schuhler ◽  
Gaëtan Chicanne ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
Phospholipid Composition ◽  
Extracellular Domain ◽  
Human Platelets ◽  
Normal Platelet ◽  
Activated Platelets ◽  
The Impact ◽  
Normal Calcium

Abstract Activated platelets become procoagulant and efficiently promote the conversion of prothrombin to thrombin. A role of the GPIb-V-IX complex has long been postulated in view of the decreased prothrombin consumption in Bernard-Soulier patients. We evaluated the impact of GPIb-V-IX deficiency and the requirement for the GPIbα extracellular domain. In GPIbβ−/− mice, thrombin generation was profoundly decreased in tissue factor– or collagen-related peptide (CRP)–activated platelet-rich plasma and in washed platelets supplemented with normal plasma or with FVa, FXa, and prothrombin. Phosphatidylserine (PS) exposure was similarly decreased in response to thrombin, CRP, or CRP + PAR4 peptide despite a normal platelet phospholipid composition. The hypothesis that these defects originate from lack of the GPIbα N-terminal domain was evaluated after its removal from normal mouse and human platelets with Nk protease or O-sialoglycoprotein endopeptidase. Unexpectedly, the treated platelets exhibited normal thrombin generation and PS exposure, indicating that GPIb-V-IX regulates procoagulant activity independently of its GPIbα-binding region. These results suggested a more general structuring role through intracellular cytoskeleton-anchoring portions regulating responses leading to PS exposure. This hypothesis was supported by the decreased calcium mobilization observed in GPIbβ−/− platelets in response to several agonists, some acting independently of GPIb, in contrast to the normal calcium responses in Nk protease–treated platelets.

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