Perioperative haemostasis with full-length, PEGylated, recombinant factor VIII with extended half-life (rurioctocog alfa pegol) in patients with haemophilia A: Final results of a multicentre, single-arm phase III trial.

INTRODUCTION: Rurioctocog alfa pegol (BAX 855, TAK-660) is a PEGylated, full-length, recombinant factor VIII (rFVIII) with extended half-life developed from unmodified rFVIII (antihaemophilic factor [recombinant]). AIM: To determine the perioperative haemostatic efficacy and safety of rurioctocog alfa pegol in male previously treated patients (PTPs) with severe haemophilia A. METHODS: This multicentre, single-arm, phase III study included PTPs who were to undergo major or minor elective or minor emergency surgical, dental or other invasive procedures. Rurioctocog alfa pegol dose and frequency were individualized based on patients' pharmacokinetic profiles for major surgeries and by rurioctocog alfa pegol incremental recovery for minor surgeries. Haemostatic efficacy was assessed using the Global Haemostatic Efficacy Assessment score. RESULTS: Twenty-one patients aged 16-61 years underwent 21 major and five minor surgeries. For all 24 evaluable surgeries, overall haemostatic efficacy was rated as excellent and blood loss comparable to that expected in non-haemophilic patients. No blood transfusions were required intraoperatively but were administered postoperatively for four surgeries in three patients. Five injury-related postoperative bleeding episodes occurred in five patients, of which two required additional rurioctocog alfa pegol treatment. Two non-serious adverse events of mild severity (increased ALT level and headache) were considered possibly related to rurioctocog alfa pegol. There were no deaths or treatment-related serious adverse events. No patients developed inhibitory antibodies to FVIII or persistent IgG- or IgM-binding antibodies to FVIII, PEG-FVIII or PEG. CONCLUSION: Rurioctocog alfa pegol was well tolerated and effective for perioperative use in patients with haemophilia A and showed no signs of immunogenicity.

Clinical and laboratory variability in a cohort of patients diagnosed with type 1 VWD in the United States.

von Willebrand disease (VWD) is the most common inherited bleeding disorder, and type 1 VWD is the most common VWD variant. Despite its frequency, diagnosis of type 1 VWD remains the subject of debate. In order to study the spectrum of type 1 VWD in the United States, the Zimmerman Program enrolled 482 subjects with a previous diagnosis of type 1 VWD without stringent laboratory diagnostic criteria. von Willebrand factor (VWF) laboratory testing and full-length VWF gene sequencing was performed for all index cases and healthy control subjects in a central laboratory. Bleeding phenotype was characterized using the International Society on Thrombosis and Haemostasis bleeding assessment tool. At study entry, 64% of subjects had VWF antigen (VWF:Ag) or VWF ristocetin cofactor activity below the lower limit of normal, whereas 36% had normal VWF levels. VWF sequence variations were most frequent in subjects with VWF:Ag <30 IU/dL (82%), whereas subjects with type 1 VWD and VWF:Ag ≥30 IU/dL had an intermediate frequency of variants (44%). Subjects whose VWF testing was normal at study entry had a similar rate of sequence variations as the healthy controls (14%). All subjects with severe type 1 VWD and VWF:Ag ≤5 IU/dL had an abnormal bleeding score (BS), but otherwise BS did not correlate with VWF:Ag. Subjects with a historical diagnosis of type 1 VWD had similar rates of abnormal BS compared with subjects with low VWF levels at study entry. Type 1 VWD in the United States is highly variable, and bleeding symptoms are frequent in this population.

Outcomes in children with deep vein thrombosis managed with percutaneous endovascular thrombolysis.

BACKGROUND: Our center has developed a multidisciplinary approach to percutaneous endovascular thrombolysis with the goal of improving outcomes in children with thrombosis. There is little data describing the safety and efficacy of endovascular thrombolysis and the frequency of post-thrombotic syndrome after thrombolysis in children. OBJECTIVE: Retrospective analysis of children undergoing percutaneous endovascular thrombolysis to determine (1) the safety and efficacy of this procedure and (2) the frequency of the diagnosis of post-thrombotic syndrome after thrombolysis. MATERIALS AND METHODS: We reviewed the medical and imaging databases for children who underwent percutaneous endovascular thrombolysis for deep venous thrombosis (DVT) between November 2008 and June 2013 at our institution. Demographic data were reviewed for the technical success and complications of thrombolysis and the last assigned post-thrombotic syndrome score using standardized scoring tools. RESULTS: Forty-one children ages 3 months to 21 years (median age: 15 years; 44% male) underwent percutaneous endovascular thrombolysis between November 2008 and June 2013. Upper extremity DVT occurred in 13 patients (32%); lower extremity DVT occurred in 28 patients (68%). All 41 patients received thrombolysis grading; 90% of those patients achieved greater than 50% thrombus lysis. Twenty-eight patients received formal post-thrombotic syndrome scoring and 4 (14%) met diagnostic criteria for post-thrombotic syndrome. One major bleeding episode and one pulmonary embolism occurred with no long-term sequelae. CONCLUSION: Endovascular thrombolysis for DVT in children is safe, effective at thrombus removal and may reduce the incidence of post-thrombotic syndrome. Randomized or larger clinical trials would be needed to determine the long-term benefits of endovascular thrombolysis.

Evidence for the transmission of parvovirus B19 in patients with bleeding disorders treated with plasma-derived factor concentrates in the era of nucleic acid test screening.

BACKGROUND: Parvovirus B19 (B19V) is a small, nonenveloped virus that typically causes a benign flu-like illness that occurs most frequently in childhood. The virus is resistant to current viral inactivation steps used in the manufacture of antihemophilic factor concentrates and B19V transmission through these products has been documented. Since 2000, B19V nucleic acid test (NAT) screening of plasma pools has been implemented to further decrease the viral burden in these products, but no study has examined populations using these products to assess the impact of the screening on B19V transmission. STUDY DESIGN AND METHODS: Blood specimens obtained from participants of a surveillance system established in federally supported specialized bleeding disorders clinics were used in a B19V seroprevalence study. RESULTS: A total of 1643 specimens from 1043 participants age 2 to 7 years born after B19V NAT screening was implemented were tested. Age-specific prevalence rates were generally higher for subjects exposed to either plasma-derived products alone or in combination with other products compared to subjects with no exposure to antihemophilic products. Overall, compared to participants unexposed to blood or blood products, those exposed to plasma-derived products alone were 1.7 times more likely to have antibodies to B19V (p = 0.002). CONCLUSION: These results are consistent with continued B19V transmission through plasma-derived factor concentrates. Effective viral inactivation and detection processes are needed to protect users of these products from infection with B19V or other new or emerging viruses.

Developing a pediatric endovascular thrombolysis program: a single-center experience.

Deep venous thrombosis (DVT) is being increasingly recognized as a significant issue in children. Despite the low incidence of DVT, the risks of pulmonary embolism and death in children are significant. Post-thrombotic syndrome, a syndrome of chronic venous insufficiency, can have long-term adverse consequences in children and adolescents. Adult studies have shown that catheter-directed therapy can reduce the incidence of post-thrombotic syndrome. Safety of catheter-directed therapy in adolescents has also been demonstrated. These reasons compelled us to institute a pediatric endovascular thrombolysis program at our institute for management of pediatric DVT. We describe the process of developing a multi-disciplinary thrombolysis program involving interventional radiology (pediatric and adult), pediatric hematology, critical care, anesthesia and vascular surgery, and describe the role of each specialty in the development of the program. We also describe our experience with patient selection, endovascular therapy procedure, pre-, intra- and post-procedure monitoring, and follow-up management for endovascular therapy for DVT.

Atypical Hemolytic Uremic Syndrome.

Atypical hemolytic uremic syndrome is a rare life-threatening disease of unregulated complement activation. Untreated, the prognosis is generally poor; more than one-half of patients die or develop end-stage renal disease within 1 year. Atypical hemolytic uremic syndrome is characterized by thrombotic microangiopathy with evidence of hemolysis, thrombocytopenia, and renal impairment. This systemic disease affects the kidneys, brain, heart, lungs, gastrointestinal tract, pancreas, and skin. Acquired and genetic abnormalities of complement regulation may be identified in approximately 70% of patients. Plasma therapy is generally ineffective. Eculizumab blocks terminal complement activation, prevents complement-mediated organ damage, and is currently recommended as front-line therapy.

Population-specific alleles: the polymorphism (K121Q) of the human glycoprotein PC-1 gene is strongly associated with race but not with insulin resistance in black and white children.

The K121Q polymorphism of the glycoprotein PC-1 gene was recently reported to associate with insulin resistance (IR) in an all-Caucasian, Sicilian population. Given black-white differences in plasma insulin and IR, we compared the prevalence of the KK, KQ, and QQ genotypes and their associations with insulin and IR in 2 large, biracial pediatric samples: 1 hospital-based (n = 301, 137 blacks and 164 whites) and 1 school-based (n = 639, 344 blacks and 295 whites). The Q allele frequencies in the hospital-based and school-based cohorts in black children were 0.80 and 0.77 and in the white children, 0.15 and 0.13. The K allele frequencies in the hospital-based and school-based cohorts in black children were 0.20 and 0.23 and in the white children, 0.85 and 0.87. Differences in allelic frequencies were highly significant (chi square test, P <.0001) for both the hospital-based cohort and the school-based cohort. Both cohorts were in Hardy-Weinberg equilibrium. Within race, after covariance adjusting for age and body mass index (BMI), there were no significant differences (P >/=.10) among the 3 PC-1 genotypes for insulin, glucose, or homeostasis model assessment (HOMA) IR. After covariance adjusting for age and BMI, black girls had higher insulin (P =.0007) and higher HOMA IR (P =.0002) than white girls. The K121Q polymorphism was not associated with insulin, glucose, or HOMA IR measures in black or white children. However, the QQ genotype was population-specific, encompassing most black children versus 1% to 3% of white children. As such, K121Q genotyping should be useful in epidemiology, population genetics, and forensic anthropology.

Legg-Calve-Perthes disease and thrombophilia.

BACKGROUND: Thrombophilia has previously been identified as a potential etiologic factor in Legg-Calve-Perthes disease. We prospectively studied the association between Legg-Calve-Perthes disease and coagulation abnormalities by comparing seventy-two children who had the disease with 197 healthy controls. METHODS: A nonselected, consecutive series of seventy-two patients with Legg-Calve-Perthes disease (mean age [and standard deviation], 6.6 +/- 2.6 years) was studied in their order of referral and compared with 197 healthy controls (mean age, 7.6 +/- 5.1 years). Assays were done for factor-V Leiden, prothrombin G20210A, methylenetetrahydrofolate reductase C677T, and plasminogen activator inhibitor-1 4G/5G gene mutations. Levels of anticardiolipin antibodies immunoglobulin G and M (IgG and IgM), homocysteine, protein C, protein S, antithrombin III, and plasminogen activator inhibitor-1 were also measured. RESULTS: The factor-V Leiden mutation was more common in the patients (eight of seventy-two) than in the controls (seven of 197) (chi-square = 5.7, p = 0.017). After we controlled for the false-discovery rate, the case-control difference remained significant (p = 0.017). The odds ratio for the development of Legg-Calve-Perthes disease in the presence of the factor-V Leiden mutation was 3.39 with a 95% confidence interval of 1.18 to 9.73. A high level of anticardiolipin antibodies (IgG and/or IgM) was found in nineteen of the seventy-two patients compared with twenty-two of the 197 controls (chi-square = 9.5, p = 0.002). After we controlled for the false-discovery rate, the case-control difference remained significant (p = 0.002). The odds ratio of patients with Legg-Calve-Perthes disease having one or more abnormalities in factor V, anticardiolipin antibody IgG, or anticardiolipin antibody IgM as opposed to normal values for all three variables was 3.29 (95% confidence interval, 1.73 to 6.24; p = 0.0003). CONCLUSIONS: Two thrombophilic risk factors, the factor-V Leiden mutation and anticardiolipin antibodies, are associated with Legg-Calve-Perthes disease, an association that may reflect causality. LEVEL OF EVIDENCE: Prognostic study, Level II-1 (retrospective study). See Instructions to Authors for a complete description of levels of evidence.

Integrin αIIb-mediated PI3K/Akt activation in platelets.

Integrin αIIbß3 mediated bidirectional signaling plays a critical role in thrombosis and haemostasis. Signaling mediated by the ß3 subunit has been extensively studied, but αIIb mediated signaling has not been characterized. Previously, we reported that platelet granule secretion and TxA2 production induced by αIIb mediated outside-in signaling is negatively regulated by the ß3 cytoplasmic domain residues R(724)KEFAKFEEER(734). In this study, we identified part of the signaling pathway utilized by αIIb mediated outside-in signaling. Platelets from humans and gene deficient mice, and genetically modified CHO cells as well as a variety of kinase inhibitors were used for this work. We found that aggregation of TxA2 production and granule secretion by ß3Δ724 human platelets initiated by αIIb mediated outside-in signaling was inhibited by the Src family kinase inhibitor PP2 and the PI3K inhibitor wortmannin, respectively, but not by the MAPK inhibitor U0126. Also, PP2 and wortmannin, and the palmitoylated ß3 peptide R(724)KEFAKFEEER(734), each inhibited the phosphorylation of Akt residue Ser473 and prevented TxA2 production and storage granule secretion. Similarly, Akt phosphorylation in mouse platelets stimulated by the PAR4 agonist peptide AYPGKF was αIIbß3-dependent, and blocked by PP2, wortmannin and the palmitoylated peptide p-RKEFAKFEEER. Akt was also phosphorylated in response to mAb D3 plus Fg treatment of CHO cells in suspension expressing αIIbß3-Δ724 or αIIbß3E(724)AERKFERKFE(734), but not in cells expressing wild type αIIbß3. In summary, SFK(s) and PI3K/Akt signaling is utilized by αIIb-mediated outside-in signaling to activate platelets even in the absence of all but 8 membrane proximal residues of the ß3 cytoplasmic domain. Our results provide new insight into the signaling pathway used by αIIb-mediated outside-in signaling in platelets.

Integrated analysis of safety and efficacy of a plasma- and albumin-free recombinant factor VIII (rAHF-PFM) from six clinical studies in patients with hemophilia A.

BACKGROUND: Hemophilia A is an X-linked bleeding disorder that results from insufficient levels of factor VIII (FVIII) coagulant activity. OBJECTIVE: To evaluate the efficacy and safety of ADVATE rAHF-PFM (Baxter Healthcare Corporation), a recombinant FVIII concentrate manufactured without human or bovine blood-derived additives, and to assess the effect of compliance with prophylactic use in preventing bleeding episodes (BEs). METHODS: Clinical data were integrated from six prospective studies. Two hundred thirty-four hemophilia A subjects (FVIII levels < or = 2%) (median age 14.7 (range: 0.02 - 72.7) years) were included. RESULTS: BEs were managed with one or two infusions and nearly all (1953/1956) responded to treatment. Compliance with a prophylactic treatment regimen significantly reduced the incidence of BEs (p = 0.0061) and prevented non-traumatic joint BEs (median annualized BE rate was 0). One previously treated subject developed an inhibitor; no other safety concerns were observed. CONCLUSIONS: These results reinforce the efficacy and safety of rAHF-PFM and suggest that compliance is an essential contributor to the effectiveness of prophylaxis in the treatment of hemophilia A.

Ventilation-perfusion scintigraphy in children and adolescents is associated with a low rate of indeterminate studies.

BACKGROUND: In selected populations, ventilation-perfusion (V/Q) studies are nearly as accurate as CT angiography (CTA) for the diagnosis of pulmonary emboli (PE). This study was performed to determine the percentage of V/Q studies in children and adolescents that are indeterminate for the presence of PE. MATERIALS AND METHODS: V/Q studies performed over a period of 2 years were reviewed. Studies from children and adolescents with chronic lung disease or recent documented severe PE were excluded. There were 37 V/Q studies and 3 perfusion only (Q) studies in 35 patients. Studies were evaluated using modified Biello criteria. Effective doses (EDs) for V/Q and CTA studies of the lung were calculated from administered activities and CT exposure parameters used. RESULTS: Eighteen studies were normal, 4 studies had a very low probability of PE, 6 were low probability, and 2 were high probability for PE. Four studies were negative for new PE when compared with a previous study. Five V/Q studies and 1 Q only study were indeterminate for PE (15%), only slightly higher than the reported percentage of indeterminate CTA in adults. ED from V/Q was about half the ED from CT angiography. Breast dose from V/Q was less than 3% of the breast dose from CT. CONCLUSION: In this selected group of children and adolescents, the percentage of indeterminate V/Q studies is low. V/Q has considerably lower absorbed breast and effective radiation doses than CTA, and is still appropriate for imaging children who are suspected of having PE.

Clinical and positron emission tomography findings of chorea associated with primary antiphospholipid antibody syndrome.

A fourteen-year-old right-handed male with a history of attention deficit hyperactivity disorder (ADHD) presented with alternating hemichorea. Laboratory findings included elevated anticardiolipin IgG and anti-beta(2)-glycoprotein I IgG, which were consistent with primary antiphospholipid antibody syndrome. Positron emission tomography (PET) imaging revealed altered striatal metabolism in his left putamen while he was exhibiting right-sided hemichorea. His symptoms resolved on prednisone; however, his antiphospholipid antibody profile remained markedly abnormal despite being symptom-free for 26 months.

Favorable histology, MYCN-amplified 4S neonatal neuroblastoma.

We report a neonate with 4S neuroblastoma and MYCN amplification, but favorable Shimada histology, successfully treated with chemotherapy and 13-cis-retinoic acid without stem cell transplantation. MYCN amplification in neuroblastoma is usually associated with unfavorable Shimada histology; the presence of these features in infants with 4S disease confers a poor prognosis. A small number of infants with 4S neuroblastoma and MYCN amplification have favorable Shimada histology. In this subgroup of infants, histopathology may be equally important in predicting outcome.

The beta3 subunit of the integrin alphaIIbbeta3 regulates alphaIIb-mediated outside-in signaling.

Bidirectional signaling is an essential feature of alphaIIbbeta3 function. The alphaIIb cytoplasmic domain negatively regulates beta3-mediated inside-out signaling, but little is known about the regulation of alphaIIb-mediated outside-in signaling. We show that alphaIIb-mediated outside-in signaling is enhanced in platelets of a patient lacking the terminal 39 residues of the beta3 cytoplasmic tail. This enhanced signaling was detected as thromboxane A(2) (TxA(2)) production and granule secretion, and required ligand cross-linking of alphaIIbbeta3 and platelet aggregation. This outside-in signaling was specifically inhibited by a palmitoylated version of a beta3 peptide corresponding to cytoplasmic domain residues R724-R734. Unlike the palmitoylated peptide, the nonpalmitoylated beta3 peptide could not cross the platelet membrane and did not inhibit this outside-in signaling. The physiologic relevance of this beta3-mediated negative regulation of alphaIIb outside-in signaling was demonstrated in normal platelets treated with the palmitoylated peptide and a physiologic agonist. Binding of alphaIIbbeta3 complexes to immobilized peptides demonstrated that a peptide corresponding to beta3 residues R724-R734 appears to bind to an alphaIIb cytoplasmic domain peptide containing residues K989-D1002, but not to control peptides. These results demonstrate that alphaIIb-mediated outside-in signaling resulting in TxA(2) production and granule secretion is negatively regulated by a sequence of residues in the membrane distal beta3 cytoplasmic domain sequence RKEFAKFEEER.

Clinical and molecular variability in congenital dyserythropoietic anaemia type I.

Congenital dyserythropoietic anaemia (CDA) type I is a rare, inherited disorder characterised by ineffective erythropoiesis and macrocytic anaemia. Complex bone disease has only occasionally been associated with this disease. CDA I is caused by mutations in the CDAN1 gene encoding for codanin-1. Our aim was to characterise the CDAN1 mutation in eight unrelated patients with sporadic CDA I, three of whom had complex bone disease. Six novel mutations in the CDAN1 gene were identified. In two patients, one mutation and in another, both mutations were elusive. No patient was homozygous for a null-type mutation. However, one patient with complex bone disease was homozygous for a splice-site mutation (IVS-12+5G>A). Western blotting revealed that codanin-1 synthesis was 65% less than the control. Five single nucleotide polymorphisms (SNPs) previously unreported in the literature or the SNP database were also identified. Although the absence of codanin-1 is probably lethal, the presence of 35% of the protein was compatible with life but was associated with severe clinical manifestations. However, in most patients studied, no correlation could be established between the expected levels of codanin-1 or the nature of the mutation and the severity of the clinical manifestations.