Georges E. Rivard

Evaluation of the profile of thrombin generation during the process of whole blood clotting as assessed by thrombelastography

Summary.  The objective of this study was to evaluate the possibility of linking the tracing of whole blood clotting in a thrombelastograph® (TEG®) hemostasis system with the generation of thrombin assessed by thrombin/antithrombin complex (TAT). Citrated whole blood containing corn trypsin inhibitor from volunteers was clotted in the presence of CaCl2 and tissue factor. Clotting was monitored with the eight channels of a TEG® system. At different time points, the whole blood TEG® reaction cups were kept in a cold quenching solution, centrifuged, and the supernatants were kept at −80 °C until assayed for TAT by ELISA. The total thrombus generation (TTG) was calculated from the first derivative of the TEG® waveform and was compared with thrombin generation measured by TAT. The two vector values – the TAT thrombin generation data and the corresponding TEG® TTG – were analyzed using Pearson correlation coefficients (r) and linear, non‐linear and natural log (ln) transformation of TAT values for least‐squares goodness‐of‐fit curves. The best least‐squares fit is an exponential curve. Linearizing using the ln of the TAT thrombin generation variable produces the same r (0.94) as of the exponential curve. The prediction equation is y = 8.0465 + 0.0005x (P ≤ 0.0001), where y is the TAT thrombin generation variable in the ln transformation and x is the TEG® TTG variable. The high magnitude of r and the high significance of the prediction equation demonstrate the high efficacy of the prediction of TAT thrombin generation by the use of TEG® TTG.

Tailored prophylaxis in severe hemophilia A: interim results from the first 5 years of the Canadian Hemophilia Primary Prophylaxis Study.

Summary.  Background: Prophylactic treatment for severe hemophilia A is likely to be more effective than treatment when bleeding occurs, however, prophylaxis is costly. We studied an inception cohort of 25 boys using a tailored prophylaxis approach to see if clotting factor use could be reduced with acceptable outcomes. Methods: Ten Canadian centers enrolled subjects in this 5‐year study. Children were followed every 3 months at a comprehensive care hemophilia clinic. They were initially treated with once‐weekly clotting factor; the frequency was escalated in a stepwise fashion if unacceptable bleeding occurred. Bleeding frequency, target joint development, physiotherapy and radiographic outcomes, as well as resource utilization, were determined prospectively. Results: The median follow‐up time was 4.1 years (total 96.9 person‐years). The median time to escalate to twice‐weekly therapy was 3.42 years (lower 95% confidence limit 2.05 years). Nine subjects developed target joints at a rate of 0.09 per person‐year. There was an average of 1.2 joint bleeds per person‐year. The cohort consumed on average 3656 IU kg−1year−1 of factor (F) VIII. Ten subjects required central venous catheters (three while on study); no complications of these devices were seen. One subject developed a transient FVIII inhibitor. End‐of‐study joint examination scores – both clinically and radiographically – were normal or near‐normal. Conclusions: Most boys with severe hemophilia A will probably have little bleeding and good joint function with tailored prophylaxis, while infusing less FVIII than usually required for traditional prophylaxis.

Validation of a new pediatric joint scoring system from the International Hemophilia Prophylaxis Study Group: Validity of the hemophilia joint health score

Repeated hemarthrosis in hemophilia causes arthropathy with pain and dysfunction. The Hemophilia Joint Health Score (HJHS) was developed to be more sensitive for detecting arthropathy than the World Federation of Hemophilia (WFH) physical examination scale, especially for children and those using factor prophylaxis. The HJHS has been shown to be highly reliable. We compared its validity and sensitivity to the WFH scale.

Persons with Quebec platelet disorder have a tandem duplication of PLAU, the urokinase plasminogen activator gene

Quebec platelet disorder (QPD) is an autosomal dominant bleeding disorder linked to a region on chromosome 10 that includes PLAU, the urokinase plasminogen activator gene. QPD increases urokinase plasminogen activator mRNA levels, particularly during megakaryocyte differentiation, without altering expression of flanking genes. Because PLAU sequence changes were excluded as the cause of this bleeding disorder, we investigated whether the QPD mutation involved PLAU copy number variation. All 38 subjects with QPD had a direct tandem duplication of a 78-kb genomic segment that includes PLAU. This mutation was specific to QPD as it was not present in any unaffected family members (n = 114), unrelated French Canadians (n = 221), or other persons tested (n = 90). This new information on the genetic mutation will facilitate diagnostic testing for QPD and studies of its pathogenesis and prevalence. QPD is the first bleeding disorder to be associated with a gene duplication event and a PLAU mutation.

Platelet disorders in children: A diagnostic approach†‡

The investigation of children with suspected inherited platelet disorders is challenging. The causes of mucocutaneous bleeding are many, and specialized testing for platelet disorders can be difficult to access or interpret. An algorithm developed for the investigation of suspected platelet disorders provides a sequential approach to evaluating both platelet function abnormalities and thrombocytopenia. Investigation begins with a clinical evaluation and laboratory testing that is generally available, including platelet counting, peripheral blood cell morphology, and aggregometry. Based on results of initial investigations, the algorithm recommends specialized testing for specific diagnoses, including flow cytometry, immunofluorescence microscopy, electron microscopy, and mutational analysis. Pediatr Blood Cancer 2011;56:975–983.

Magnetic resonance imaging and joint outcomes in boys with severe hemophilia A treated with tailored primary prophylaxis in Canada.

Summary.  Background/objectives: Tailored primary prophylaxis (TPP) is a reduced‐intensity treatment program for hemophiliacs with the goal of preventing arthropathy. Our primary aim was to evaluate the joint outcomes of treated subjects using magnetic resonance imaging (MRI) and physical examination as outcome measures. Methods: Ankles, elbows and knees (index joints) of 24 subjects (median [range] age at start of therapy, 1.6 [1–2.5] years) with severe hemophilia A enrolled in the Canadian Hemophilia Primary Prophylaxis Study (CHPS) were examined by MRI at a median age of 8.8 years (range 6.2–11.5 years). Subjects were treated with TPP using a recombinant factor VIII concentrate, starting once weekly and escalating in frequency and dose according to frequency of bleeding. Results: Osteochondral changes (cartilage loss/subchondral bone damage) were detected in 9% (13/140) of the index joints and 50% (12/24) of study subjects. Osteochondral changes were restricted to joints with a history of clinically reported joint bleeding. Soft tissue changes were detected in 31% (20/65) of index joints with no history of clinically reported bleeding (ankles 75% (12/16); elbows 19% (6/32); and knees 12% (2/17)). In these apparently ‘bleed free’ index joints hemosiderin deposition was detected by MRI in 26% (17/65) of joints (ankles 63% (10/16); elbows 16% (5/32), and knees 12% (2/17)). Conclusion: TPP did not completely avoid the development of MRI‐detected structural joint changes in hemophilic boys in this prospective study. A longer period of follow‐up is required for assessment of the longitudinal course of these early changes in hemophilic arthropathy, detected using a sensitive imaging technique (MRI).

Recommendations for performing thromboelastography/thromboelastometry in hemophilia: communication from the SSC of the ISTH

M. CHITLUR ,* G . E . R IVARD ,† D. L I LL ICRAP ,‡ K . MANN,§ M. SH IMA, ¶ G. YOUNG** and ON BEHALF OF THE FACTOR V I I I , FACTOR IX , AND RARE COAGULAT ION DISORDERS SUBCOMMITTEE OF THE SC I ENT IF IC AND STANDARDISAT ION COMMITTEE OF THE INTERNAT IONAL SOCIETY ON THROMBOS IS AND HAEMOSTAS I S *Children’s Hospital of Michigan, Detroit, MI, USA; †CHU Sainte-Justine, Montr eal, QC; †Queens University, Kingston, ON, Canada; §University of Vermont, Colchester, VT, USA; ¶Nara Medical University, Kashihara, Nara, Japan; and **Children’s Hospital Los Angeles, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA

Quebec platelet disorder: features, pathogenesis and treatment.

Quebec platelet disorder (QPD) is a rare, autosomal-dominant, inherited bleeding disorder that is associated with unique abnormalities in fibrinolysis. Its hallmark features are delayed-onset bleeding following hemostatic challenges that responds to fibrinolytic inhibitor therapy and increased expression and storage of the fibrinolytic enzyme urokinase plasminogen activator in platelets, without increased plasma urokinase plasminogen activator or systemic fibrinolysis. The increased urokinase plasminogen activator in QPD platelets is only partially inhibited, and, as a result, there is intraplatelet generation of plasmin, and secondary degradation of many platelet α-granule proteins. During clot formation, the urokinase plasminogen activator released by QPD platelets leads to platelet-dependent increased fibrinolysis, and this is postulated to be a major contributor to QPD bleeding. The focus of the present review is to summarize the current state of knowledge on QPD, including the history of this disorder, its clinical and laboratory features, and recommended approaches for its diagnosis and treatment.

Management of von Willebrand disease: a survey on current clinical practice from the haemophilia centres of North America.

The optimal treatment of patients with von Willebrand’s disease (vWD) remains to be defined. Moreover, it has not been firmly established which, if any, commonly measured parameters of von Willebrand factor (vWF) protein in the plasma are useful in guiding treatment. To better understand what guidelines physicians follow in the management of vWD, we surveyed 194 North American physicians who are members of the Hemophilia Research Society. Ninety‐nine per cent of responding physicians depend on factor VIII (FVIII):C, vWF:RCo activity and vWF:AG to diagnose vWD, while only 49% use the bleeding time. The minimal goals of treatment for patients undergoing major surgery/trauma or central nervous system haemorrhage were FVIII:C and vWF:RCo activity greater than 80% while levels of more than 50% for minor surgery and dental extractions were considered adequate. Treatment of vWD was based on the type of vWD with type 1 patients being treated most often with desmopressin acetate (DDAVP) alone, types 2A and 2B patients with a combination of DDAVP and a vWF‐containing FVIII product, type 3 patients with vWF‐containing concentrate. Viral infections, including human immunodeficiency virus, hepatitis A, B and C viruses, and parvovirus have been seen in vWD and the efficacy of viral attenuation processes is a major criterion for the selection of treatment by physicians. Based on this survey, prospective studies need to be designed to address the clinical efficacy, safety and predictive value of laboratory monitoring of patients with vWD.

Quebec platelet disorder is linked to the urokinase plasminogen activator gene (PLAU) and increases expression of the linked allele in megakaryocytes.

Quebec platelet disorder (QPD) is an autosomal dominant disorder with high penetrance that is associated with increased risks for bleeding. The hallmark of QPD is a gain-of-function defect in fibrinolysis due to increased platelet content of urokinase plasminogen activator (uPA) without systemic fibrinolysis. We hypothesized that increased expression of uPA by differentiating QPD megakaryocytes is linked to PLAU. Genetic marker analyses indicated that QPD was significantly linked to a 2-Mb region on chromosome 10q containing PLAU with a maximum multipoint logarithm of the odds (LOD) score of +11 between markers D10S1432 and D10S1136. Analysis of PLAU by sequencing and Southern blotting excluded mutations within PLAU and its known regulatory elements as the cause of QPD. Analyses of uPA mRNA indicated that QPD distinctly increased transcript levels of the linked PLAU allele with megakaryocyte differentiation. These findings implicate a mutation in an uncharacterized cis element near PLAU as the cause of QPD.