Performance Characteristics of DOAC-Remove for Neutralization of the Effects of Apixaban and Rivaroxaban in Lupus Anticoagulant Assays.

OBJECTIVES: This study established the performance characteristics of DOAC-Remove for neutralization of the effects of rivaroxaban and apixaban in lupus anticoagulant (LAC) testing. METHODS: Normal donor, LAC control, and patient samples were spiked with rivaroxaban or apixaban to simulate their effects on the dilute Russell's viper venom time (dRVVT), activated partial thromboplastin time (APTT), and dilute prothrombin time (dPT). Anti-Xa activity was measured after spiking and after DOAC-Remove neutralization. Accuracy, complex precision, and reference interval verification were evaluated. RESULTS: DOAC-Remove neutralized rivaroxaban and apixaban concentrations as high as 415 ng/mL and 333 ng/mL, respectively. Percentage positive and negative agreement between the baseline and postneutralization interpretations were 75% or higher for the dRVVT and APTT methods but not for the dPT method. Coefficients of variation (CVs) were 10% or less for all assays except the Staclot-LA delta, which had a standard deviation of 2.5 seconds or CV of 25% or less depending on the level. The laboratory's reference intervals were verified for the dRVVT and APTT assays after DOAC-Remove treatment but not for the dPT assays. CONCLUSIONS: DOAC-Remove appears to have acceptable performance characteristics for neutralizing the effects of rivaroxaban and apixaban in the dRVVT and APTT methods but not in the dPT method.

Plasma thrombospondin-1 is increased during acute sickle cell vaso-occlusive events and associated with acute chest syndrome, hydroxyurea therapy, and lower hemolytic rates.

Platelets are activated in sickle cell disease (SCD), and particularly during vaso-occlusive episodes (VOE). Thrombospondin-1 (TSP1), a major secretory product of activated platelets, is increased in the circulation in VOE and binds to sickle red blood cells (RBC) promoting vascular adhesion. Thus, we hypothesized that TSP1 may represent a plasma biomarker of disease severity in SCD. We tested the plasma collected from patients in steady state (n = 27) and VOE (n = 14), as well as healthy controls (n = 17) at the University of Pittsburgh Medical Center (UPMC), and from patients in steady state enrolled in the walk-PHaSST clinical trial (n = 483). We found that TSP1 levels were increased in VOE in the UPMC cohort. Among steady-state patients at UPMC, TSP1 values correlated positively with lifetime history of acute chest syndrome (r = 0.72, P < 0.0001) and hemoglobin concentration (r = 0.49, P = 0.01), and negatively with markers of hemolysis, such as LDH (r = -0.50, P = 0.009). Analysis of the walk-PHaSST cohort also showed a positive association between TSP1 levels and hydroxyurea use (r = 0.14, P = 0.003), and confirmed the negative associations with the severity of hemolysis. Our results suggest that TSP1 levels are associated with more VOE, hydroxyurea use and lower rates of hemolysis. High TSP1 concentrations may indicate higher risk of the viscosity/vaso-occlusion phenotype of SCD.

Hepatic function after genetically engineered pig liver transplantation in baboons.

BACKGROUND: If "bridging" to allo-transplantation (Tx) is to be achieved by a pig liver xenograft, adequate hepatic function needs to be assured. METHODS: We have studied hepatic function in baboons after Tx of livers from alpha1,3-galactosyltransferase gene-knockout (GTKO, n=1) or GTKO pigs transgenic for CD46 (GTKO/CD46, n=5). Monitoring was by liver function tests and coagulation parameters. Pig-specific proteins in the baboon serum/plasma were identified by Western blot. In four baboons, coagulation factors were measured. The results were compared with values from healthy humans, baboons, and pigs. RESULTS: Recipient baboons died or were euthanized after 4 to 7 days after internal bleeding associated with profound thrombocytopenia. However, parameters of liver function, including coagulation, remained in the near-normal range, except for some cholestasis. Western blot demonstrated that pig proteins (albumin, fibrinogen, haptoglobin, and plasminogen) were produced by the liver from day 1. Production of several pig coagulation factors was confirmed. CONCLUSIONS: After the Tx of genetically engineered pig livers into baboons (1) many parameters of hepatic function, including coagulation, were normal or near normal; (2) there was evidence for production of pig proteins, including coagulation factors; and (3) these appeared to function adequately in baboons although interspecies compatibility of such proteins remains to be confirmed.

Immunomodulatory derivatives induce PU.1 down-regulation, myeloid maturation arrest, and neutropenia.

The immunomodulatory drugs (IMiDs) lenalidomide and pomalidomide yield high response rates in patients with multiple myeloma, but the use of IMiDs in multiple myeloma is associated with neutropenia and increased risk for venous thromboembolism (VTE) by mechanisms that are unknown. We show that IMiDs down-regulate PU.1, a key transcription factor involved in granulocyte differentiation in vitro and in patients treated with lenalidomide. Loss of PU.1 results in transient maturation arrest with medullary accumulation of immature myeloid precursors and subsequent neutropenia. Accumulation of promyelocytes leads to high levels of the platelet aggregation agonist, cathepsin G stored in the azurophilic granules of promyelocytes. High levels of cathepsin G subsequently may increase the risk of VTE. To our knowledge, this is the first report investigating the underlying mechanism of IMiD-induced neutropenia and increased risk of VTE in multiple myeloma.

Cryoprecipitate prepared from plasma frozen within 24 hours after phlebotomy contains acceptable levels of fibrinogen and VIIIC.

BACKGROUND: As blood centers prepare fewer plasma products from female donors to avoid HLA antibodies, the standard starting material for cryoprecipitate (cryo), fresh-frozen plasma (FFP), might become increasingly scarce. A unit of cryo must contain at a minimum 80 IU of VIIIC and 150 mg of fibrinogen. Plasma frozen within 24 hours after phlebotomy (FP24) contains nearly equivalent levels of clotting factors as FFP at the time of thawing and, although not AABB/FDA approved for this purpose, might be a suitable starting material for cryo (cryo24) manufacture. STUDY DESIGN AND METHODS: FP24 was collected from 20 male donors and cryo24 was subsequently prepared according to standard blood center protocols. The cryo24 was then thawed and VIIIC, von Willebrand factor (VWF) antigen, and fibrinogen levels were measured in these cryo24 units and compared to the corresponding levels in 20 randomly selected units of standard cryo (prepared from FFP). RESULTS: The mean (+/-SD) levels in cryo and cryo24 at the time of thawing were as follows: fibrinogen (mg/unit) 455.8 (+/-172.6) and 575.8 (+/-185.9; p = 0.04), VIIIC (IU/unit) 216.1 (+/-52.3) and 252.4 (+/-70.1; p = 0.07), and VWF (IU/unit) 448.1 (+/-118.9) and 505.9 (+/-135.1; p = 0.16), respectively. All units of standard cryo and cryo24 met the current minimum content standards. CONCLUSION: All units of cryo24 in this pilot study met the current minimum content standards for cryo. While clinical studies of its efficacy are warranted, cryo24's in vitro characteristics are at least equivalent to that of standard cryo.

Intensive plasma exchange increases a disintegrin and metalloprotease with thrombospondin motifs-13 activity and reverses organ dysfunction in children with thrombocytopenia-associated multiple organ failure.

BACKGROUND: Thrombocytopenia-associated multiple organ failure (TAMOF) is a poorly understood syndrome in critically ill children. A disintegrin and metalloprotease with thrombospondin motifs (ADAMTS-13), formerly known as von Willebrand factor (VWF) cleaving protease, is decreased in adults with VWF-mediated thrombotic microangiopathy, and intensive plasma exchange (PEx) both replenishes ADAMTS-13 and improves outcome in these patients. OBJECTIVES: To determine whether: 1) critically ill children with TAMOF syndrome have decreased ADAMTS-13 activity, 2) ADAMTS-13 activity correlates with platelet counts and VWF antigen, 3) the autopsies from patients who died with reduced ADAMTS-13 activity have VWF-rich microthrombi, and 4) intensive PEx will restore ADAMTS-13 activity and facilitate organ failure resolution. DESIGN: First study: observational. Second study: randomized control trial. SETTING: Single center university pediatric intensive care unit. PATIENTS: First study: thirty-seven consecutive children (17 males and 20 females; ages ranging from 9 days to 23 years) identified with > or = 2 organs dysfunction were enrolled. Seventy-six percent of these children had thrombocytopenia (platelet counts < 100,000/mm3). Five additional critically ill children without MOF were also enrolled. In the second study, children with severe TAMOF (platelet counts < 100,000/mm3 and > 3 organ failure) were randomized to PEx or standard therapy. Primary physicians and parents agreed to enrollment in 10 of the 20 eligible patients with ages ranging from 1 year to 18 years. Five patients received PEx and 5 patients received standard therapy. RESULTS: First study: children with TAMOF (n = 28) had decreased ADAMTS-13 activity, but similar plasminogen activator inhibitor-1 activity and prothrombin time compared to children with MOF without thrombocytopenia (n = 9, p < 0.05). All non-survivors (n = 7) had TAMOF, reduced ADAMTS-13 activity, and VWF-rich microvascular thromboses at autopsy. In the second study, PEx (n = 5, median 12 days, 4-28 days) restored ADAMTS-13 activity and organ function, compared to standard therapy (n = 5, p < 0.05). CONCLUSIONS: Children with TAMOF syndrome can have VWF-mediated thrombotic microangiopathy. Similar to adult experience, PEx can replenish ADAMTS-13 activity and reverse organ failure.

von Willebrand's disease and psychotic disorders: co-segregation and genetic associations.

OBJECTIVES: To evaluate co-segregation and genetic associations between von Willebrand's disease (vWD) and psychotic disorders. METHODS: The study was initiated following ascertainment of a nuclear family in which four members were diagnosed with vWD and psychotic/mood disorders. As co-segregation was uncertain in the extended pedigree, we also investigated population-based linkage and association using polymorphisms of vWF, the gene conferring susceptibility to vWD. Three common vWF polymorphisms were investigated among 194 patients with psychotic disorders (bipolar I disorder, BD I; schizoaffective disorder, SZA and schizophrenia, SZ) and their parents. The cases were also compared with unrelated population-based controls (n = 183). RESULTS: The transmission disequilibrium test and related analyses suggested nominally significant transmission distortion of one allele and related haplotypes to the probands from their parents. The most significant results were obtained among patients with BD I, and similar trends were also evident in the SZ sample. Comparisons between the cases and population-based controls did not reveal associations, though marginally significant case-control differences were obtained in the BD I sample. CONCLUSIONS: These studies are consistent with association and linkage between vWF and BD I. However, given the relatively small sample, stochastic variation is an alternative explanation.