Dabigatran versus vitamin k antagonist: an observational across‐cohort comparison in acute coronary syndrome patients with atrial fibrillation

Essentials

• Acute coronary syndrome (ACS) with atrial fibrillation (AF) is a therapeutic challenge.
• Dual and triple antithrombotic therapy showed a similar thrombotic risk in ACS patients with AF.
• The omission of aspirin during the first month did not increase the rate of ischemic events.
• Replacement of vitamin K antagonist by dabigatran leads to an increased thrombotic risk.

Summary

Background

Dual antithrombotic therapy comprising a vitamin K antagonist (VKA) plus clopidogrel reduces the incidence of major bleeding compared with triple therapy (VKA + clopidogrel + aspirin) in acute coronary syndrome (ACS) patients with atrial fibrillation (AF), with a similar thrombotic risk. The oral thrombin inhibitor dabigatran (150 mg twice a day) showed superiority over VKA in non‐valvular AF, but data supporting its use in AF patients presenting with ACS are limited.

Objective

We sought to evaluate the efficacy of dabigatran vs. VKA in the management of AF patients undergoing percutaneous coronary intervention for an ACS.

Methods

In this open‐label study, 133 consecutive patients received dabigatran plus clopidogrel. Another cohort of 133 patients treated with VKA plus clopidogrel was used as the control group.

Results

After propensity score adjustment, the cumulative incidence of major adverse cardiovascular events over 24 months was higher with dabigatran vs. VKA (adjusted hazard ratio, 2.28; 95% confidence interval, 1.46–3.56). Similar rates of major bleeding were found (adjusted hazard ratio, 1.17; 95% confidence interval, 0.46–2.96).

Conclusions

In AF patients presenting with ACS, replacement of VKA by dabigatran concurrently with clopidogrel is associated with an increased thrombotic risk, without a reduction in major bleeding.

     

Extended International Normalized Ratio testing intervals for warfarin‐treated patients

Essentials

• Warfarin typically requires International Normalized Ratio (INR) testing at least every 4 weeks.
• We implemented extended INR testing for stable warfarin patients in six anticoagulation clinics.
• Use of extended INR testing increased from 41.8% to 69.3% over the 3 year study.
• Use of extended INR testing appeared safe and effective.

Summary

Background

A previous single‐center randomized trial suggested that patients with stable International Normalized Ratio (INR) values could safely receive INR testing as infrequently as every 12 weeks.

Objective

To test the success of implementation of an extended INR testing interval for stable warfarin patients in a practice‐based, multicenter collaborative of anticoagulation clinics.

Methods

At six anticoagulation clinics, patients were identified as being eligible for extended INR testing on the basis of prior INR value stability and minimal warfarin dose changes between 2014 and 2016. We assessed the frequency with which anticoagulation clinic providers recommended an extended INR testing interval (> 5 weeks) to eligible patients. We also explored safety outcomes for eligible patients, including next INR values, bleeding events, and emergency department visits.

Results

At least one eligible period for extended INR testing was identified in 890 of 3362 (26.5%) warfarin‐treated patients. Overall, the use of extended INR testing in eligible patients increased from 41.8% in the first quarter of 2014 to 69.3% in the fourth quarter of 2016. The number of subsequent out‐of‐range next INR values were similar between eligible patients who did and did not have an extended INR testing interval (27.3% versus 28.4%, respectively). The numbers of major bleeding events were not different between the two groups, but rates of clinically relevant non‐major bleeding (0.02 per 100 patient‐years versus 0.09 per 100 patient‐years) and emergency department visits (0.07 per 100 patient‐years versus 0.19 per 100 patient‐years) were lower for eligible patients with extended INR testing intervals than for those with non‐extended INR testing intervals.

Conclusions

Extended INR testing for stable warfarin patients can be successfully and safely implemented in diverse, practice‐based anticoagulation clinic settings.

     

Mortality due to bleeding,myocardial infarction and stroke in dialysis patients

Essentials

• Mortality due to bleeding vs. arterial thrombosis in dialysis patients is unknown.
• We compared death causes of 201 918 dialysis patients with the general population.
• Dialysis was associated with increased mortality risks of bleeding and arterial thrombosis.
• Clinicians should be aware of the increased bleeding and thrombosis risks.

Summary

Background

Dialysis has been associated with both bleeding and thrombotic events. However, there is limited information on bleeding as a cause of death versus arterial thrombosis as a cause of death.

Objectives

To investigate the occurrence of bleeding, myocardial infarction and stroke as causes of death in the dialysis population as compared with the general population.

Methods

We included 201 918 patients from 11 countries providing data to the ERA‐EDTA Registry who started dialysis treatment between 1994 and 2011, and followed them for 3 years. Age‐standardized and sex‐standardized mortality rate ratios for bleeding, myocardial infarction and stroke as causes of death were calculated in dialysis patients as compared with the European general population. Associations between potential risk factors and these causes of death in dialysis patients were investigated by calculating hazard ratios (HRs) with 95% confidence intervals (CIs) by the use of Cox proportional‐hazards regression.

Results

As compared with the general population, the age‐standardized and sex‐standardized mortality rate ratios in dialysis patients were 12.8 (95% CI 11.9–13.7) for bleeding as a cause of death (6.2 per 1000 person‐years among dialysis patients versus 0.3 per 1000 person‐years in the general population), 13.4 (95% CI 13.0–13.9) for myocardial infarction (22.5 versus 0.9 per 1000 person‐years), and 12.4 (95% CI 11.9–12.9) for stroke (14.3 versus 0.7 per 1000 person‐years).

Conclusion

Dialysis patients have highly increased risks of death caused by bleeding and arterial thrombosis as compared with the general population. Clinicians should be aware of the increased mortality risks caused by these conditions.

     

The fibrinogen prothrombin time‐derived method is not useful in patients anticoagulated with low molecular weight heparins or rivaroxaban

Essentials

• Fibrinogen prothrombin time‐derived (FIBPT‐d) behavior in anticoagulated patients is under studied.
• FIBPT‐d method overestimates fibrinogen in rivaroxaban and low molecular weight heparin samples.
• Unfractionated heparin and dabigatran samples showed similar bias to the control group.
• Rabbit brain and human recombinant thromboplastin behavior was different in rivaroxaban samples.

Summary

Background

The fibrinogen prothrombin time‐derived (FIBPT‐d) method with photo‐optical coagulometers is easy and economical. However, there are few reports on the behavior of this test on samples from patients anticoagulated with direct oral anticoagulants or low molecular weight heparin (LMWH).

Objective

To compare fibrinogen results obtained with the Clauss (FIB C) method and the FIBPT‐d method with two thromboplastins in anticoagulated patients.

Population

The study population comprised 295 consecutive anticoagulated patients: 99 treated with vitamin K antagonists (VKAs), 49 treated with unfractionated heparin (UFH), 47 treated with LMWH, 50 treated with rivaroxaban, 50 treated with dabigatran, and 100 normal controls (NCs).

Methods

Dabigatran samples were analyzed by the use of FIB C with HemosIL Fibrinogen C or 100 NHI thrombin units mL^?1 reagents; rabbit brain and human recombinant thromboplastins with HemosIL PTFibrinogen HS plus (HS) and Recombiplastin 2G (RP) were used for FIBPT‐d method. Heparin and rivaroxaban levels were assessed with HemosIL Liq antiXa with specific calibrators; dabigatran levels were determined with the HemosIL Direct Thrombin Inhibitor Assay. All assays were performed on the ACL TOP platform in two laboratories. Percentage biases for the FIBPT‐d method versus the FIB C method were calculated by the use of Bland–Altman plots.

Results

Positive biases of the FIBPT‐d method versus the FIB C method with both thromboplastins were seen in NC samples (13.7% and 18.9% for HS and RP, respectively), but biases with HS in rivaroxaban and VKA patient samples were higher than that in NC samples, at 31.9% and 34.0%, respectively. LMWH patient samples showed higher bias than NC samples: 26.5% and 29.3.0% with HS and RP, respectively. UFH and dabigatran patient samples showed similar bias as NC samples.

Conclusion

The FIBPT‐d method should not be used in anticoagulated patients, because the FIBPT‐d mathematical algorithm has been validated only in normal subjects, so overestimation could occur in these patients.

     

A systematic review and Bayesian network meta‐analysis of risk of intracranial hemorrhage with direct oral anticoagulants

Essentials

• Risk of intracranial hemorrhage (ICH) may differ between direct oral anticoagulants (DOACs).
• We compared the risk of ICH between DOACs using network meta‐analysis.
• Dabigatran 110 mg and 150 mg were safer than rivaroxaban on Bayesian analysis.
• Dabigatran 110 mg ranked as the safest DOAC while rivaroxaban ranked last.

Summary

Background

The comparative risk of intracranial hemorrhage (ICH) among direct oral anticoagulants (DOACs) (dabigatran, rivaroxaban, apixaban and edoxaban) remains unclear.

Objective

To determine the difference in risk of ICH between DOACs

Methods

Seventeen randomized controlled trials (RCTs) were selected using PubMed/MEDLINE, EMBASE and CENTRAL (Inception, 31 December 2017). Estimates were reported as odds ratio (OR) with 95% credible interval (CR.I) in Bayesian network meta‐analysis (NMA), and OR with 95% confidence interval (CI) in traditional meta‐analyses. Relative ranking probability of each group was generated based on surface under the cumulative ranking curve (SUCRA).

Results

In NMA of 116 618 patients from 17 RCTs (apixaban = 19 495 patients, rivaroxaban = 14 157 patients, dabigatran = 16 074 patients, edoxaban = 11 652 patients, and comparator = 55 315 patients), all DOACs were safer than warfarin for risk of ICH. Dabigatran 110 mg ranked as the safest drug (SUCRA, 0.85) and reduced the risk of ICH by 56% compared to rivaroxaban (OR, 0.44; 95% Cr.I, 0.22–0.82). Pairwise meta‐analysis validated these findings, showing that DOACs were safer than warfarin (OR, 0.46; 95% CI, 0.35–0.59). Subgroup analysis showed that the benefit was present when DOACs were used in non‐valvular atrial fibrillation (NVAF) (OR, 0.51; 95% CI, 0.38–0.68) or venous thromboembolism (VTE) (OR, 0.32; 95% CI, 0.18–0.58).

Conclusion

Dabigatran 110 mg may be the safest choice among any anticoagulant regarding risk of ICH. Both dabigatran 110 mg and 150 mg were safer than rivaroxaban.

     

Chronic kidney disease and bleeding risk in patients at high cardiovascular risk: a cohort study

Essentials

• The association between chronic kidney disease and bleeding is unknown.
• We followed 10 347 subjects at high cardiovascular risk for bleeding events.
• Chronic kidney disease was associated with a 1.5‐fold increased bleeding risk.
• Especially albuminuria rather than decreased kidney function was associated with bleeding events.

Summary

Background

There are indications that patients with chronic kidney disease have an increased bleeding risk.

Objectives

To investigate the association between chronic kidney disease and bleeding in patients at high cardiovascular risk.

Methods

We included 10 347 subjects referred to the University Medical Center Utrecht (the Netherlands) from September 1996 to February 2015 for an outpatient visit with classic risk factors for arterial disease or with symptomatic arterial disease (Second Manifestation of Arterial disease [SMART] cohort). Patients were staged according to the KDIGO guidelines, on the basis of estimated glomerular filtration rate (eGFR) and albuminuria, and were followed for the occurrence of major hemorrhagic events until March 2015. Hazard ratios (HRs) with 95% confidence intervals (CIs) for bleeding were calculated with Cox proportional hazards analyses.

Results

The incidence rate for bleeding in subjects with chronic kidney disease was 8.0 per 1000 person‐years and that for subjects without chronic kidney disease was 3.5 per 1000 person‐years. Patients with chronic kidney disease (n = 2443) had a 1.5‐fold (95% CI 1.2–1.9) increased risk of bleeding as compared with subjects without chronic kidney disease (n = 7904) after adjustment. Subjects with an eGFR of < 45 mL min^?1 1.73 m^–2 with albuminuria had a 3.5‐fold (95% CI 2.3–5.3) increased bleeding risk, whereas an eGFR of < 45 mL min^?1 1.73 m^–2 without albuminuria was not associated with an increased bleeding risk (HR 1.3, 95% CI 0.7–2.5).

Conclusion

Chronic kidney disease is a risk factor for bleeding in patients with classic risk factors for arterial disease or with symptomatic arterial disease, especially in the presence of albuminuria.

     

Preoperative platelet transfusions to reverse antiplatelet therapy for urgent non‐cardiac surgery: an observational cohort study

Essentials

• An increasing number of patients requiring surgery receive antiplatelet therapy (APT).
• We analyzed 181 patients receiving presurgery platelet transfusions to reverse APT.
• No coronary thrombosis occurred after platelet transfusion.
• This justifies a prospective trial to test preoperative platelet transfusions to reverse APT.

Summary

Background

Patients receiving antiplatelet therapy (APT) have an increased risk of perioperative bleeding and cardiac adverse events (CAE). Preoperative platelet transfusions may reduce the bleeding risk but may also increase the risk of CAE, particularly coronary thrombosis in patients after recent stent implantation.

Objectives

To analyze the incidence of perioperative CAE and bleeding in patients undergoing non‐cardiac surgery using a standardized management of transfusing two platelet concentrates preoperatively and restart of APT within 24–72 h after surgery.

Methods

A cohort of consecutive patients on APT treated with two platelet concentrates before non‐cardiac surgery between January 2012 and December 2014 was retrospectively identified. Patients were stratified by the risk of major adverse cardiac and cerebrovascular events (MACCE). The primary objective was the incidence of CAE (myocardial infarction, acute heart failure and cardiac troponine T increase). Secondary objectives were incidences of other thromboembolic events, bleedings, transfusions and mortality.

Results

Among 181 patients, 88 received aspirin, 21 clopidogrel and 72 dual APT. MACCE risk was high in 63, moderate in 103 and low in 15 patients; 67 had cardiac stents. Ten patients (5.5%; 95% CI, 3.0–9.9%) developed a CAE (three myocardial infarctions, four cardiac failures and three troponin T increases). None was caused by coronary thrombosis. Surgery‐related bleeding occurred in 22 patients (12.2%; 95% CI, 8.2–17.7%), making 12 re‐interventions necessary (6.6%; 95% CI, 3.8–11.2%).

Conclusion

Preoperative platelet transfusions and early restart of APT allowed urgent surgery and did not cause coronary thromboses, but non‐thrombotic CAEs and re‐bleeding occurred. Randomized trials are warranted to test platelet transfusion against other management strategies.

     

Mechanisms and mitigating factors for venous thromboembolism in chronic kidney disease: the REGARDS study

Essentials

• Chronic kidney disease (CKD) is associated with procoagulant and inflammatory biomarkers.
• We studied the association of CKD and venous thromboembolism (VTE) in a case‐cohort study.
• Factor VIII, D‐dimer and C‐reactive protein appeared to explain the association of CKD and VTE.
• Statin use was protective against VTE in those with and without CKD.

Summary

Background

Chronic kidney disease (CKD) is associated with venous thromboembolism (VTE) risk via unknown mechanisms. Whether factors associated with reduced VTE risk in the general population might also be associated with reduced VTE risk in CKD patients is unknown.

Objectives

To determine whether thrombosis biomarkers attenuate VTE risk, and whether factors associated with reduced VTE risk are similarly effective in CKD patients.

Methods

Baseline biomarkers were measured in a cohort (294 VTE cases; 939 non‐cases) from the Reasons for Geographic and Racial Differences in Stroke (REGARDS) study, a nationwide prospective cohort study of 30 239 persons aged ≥45 years with 4.3 years of follow‐up. The hazard ratio (HR) of VTE per 10 mL min^?1 1.73 m^?2 decrease in estimated glomerular filtration rate (eGFR), and the percentage attenuation of this HR by each biomarker, were calculated. Associations of protective factors (physical activity, lower body mass index [BMI], and aspirin, warfarin and statin use) with VTE were estimated in those with and without CKD.

Results

The HR for VTE with lower eGFR was 1.13 (95% confidence interval [CI] 1.02–1.25), and VTE risk was attenuated by 23% (95% CI 5–100) by D‐dimer, by 100% (95% CI 50–100) by factor VIII, and by 15% (95% CI 2–84) by C‐reactive protein. Normal BMI was associated with lower VTE risk in those without CKD (HR 0.47, 95% CI 0.32–0.70), but not in those with CKD (HR 1.07, 95% CI 0.51–2.22). Statin use, physical activity and warfarin use were associated with lower VTE risk in both groups.

Conclusions

Procoagulant and inflammatory biomarkers mediated the association of eGFR with VTE. Higher physical activity, statin use and warfarin use mitigated VTE risk in those with CKD and those without CKD, but normal BMI did not mitigate VTE risk in CKD patients.

     

Abrogating fibrinolysis does not improve bleeding or rFVIIa/rFVIII treatment in a non‐mucosal venous injury model in haemophilic rodents

Essentials

• The efficacy of systemic antifibrinolytics for hemophilic non‐mucosal bleeding is undetermined.
• The effect of systemically inhibiting fibrinolysis in hemophilic mice and rats was explored.
• Neither bleeding nor the response to factor treatment was improved after inhibiting fibrinolysis.
• The non‐mucosal bleeding phenotype in hemophilia A appears largely unaffected by fibrinolysis.

Summary

Background

Fibrinolysis may exacerbate bleeding in patients with hemophilia A (HA). Accordingly, antifibrinolytics have been used to help maintain hemostatic control. Although antifibrinolytic drugs have been proven to be effective in the treatment of mucosal bleeds in the oral cavity, their efficacy in non‐mucosal tissues remain an open question of significant clinical interest.

Objective

To determine whether inhibiting fibrinolysis improves the outcome in non‐mucosal hemophilic tail vein transection (TVT) bleeding models, and to determine whether a standard ex vivo clotting/fibrinolysis assay can be used as a predictive surrogate for in vivo efficacy.

Methods

A highly sensitive TVT model was employed in hemophilic rodents with a suppressed fibrinolytic system to examine the effect of inhibiting fibrinolysis on bleeding in non‐mucosal tissue. In mice, induced and congenital hemophilia models were combined with fibrinolytic attenuation achieved either genetically or pharmacologically (tranexamic acid [TXA]). In hemophilic rats, tail bleeding was followed by whole blood rotational thromboelastometry evaluation of the same animals to gauge the predictive value of such assays.

Results

The beneficial effect of systemic TXA therapy observed ex vivo could not be confirmed in vivo in hemophilic rats. Furthermore, neither intravenously administered TXA nor congenital knockout of the fibrinolytic genes encoding plasminogen or tissue‐type plasminogen activator markedly improved the TVT bleeding phenotype or response to factor therapy in hemophilic mice.

Conclusions

The findings here suggest that inhibition of fibrinolysis is not effective in limiting the TVT bleeding phenotype of HA rodents in non‐mucosal tissues.

     

Heparin–protamine balance after neonatal cardiopulmonary bypass surgery

Essentials

• Heparin‐protamine balance (HPB) modulates bleeding after neonatal cardiopulmonary bypass (CPB).
• HPB was examined in 44 neonates undergoing CPB.
• Post‐operative bleeding occurred in 36% and heparin rebound in 73%.
• Thrombin‐initiated fibrin clot kinetic assay and partial thromboplastin time best assessed HPB.

Summary

Background

Neonates undergoing cardiopulmonary bypass (CPB) are at risk of excessive bleeding. Blood is anticoagulated with heparin during CPB. Heparin activity is reversed with protamine at the end of CPB. Paradoxically, protamine also inhibits blood coagulation when it is dosed in excess of heparin.

Objectives

To evaluate heparin–protamine balance in neonates undergoing CPB by using research and clinical assays, and to determine its association with postoperative bleeding.

Patients/Methods

Neonates undergoing CPB in the first 30 days of life were studied. Blood samples were obtained during and after surgery. Heparin–protamine balance was assessed with calibrated automated thrombography, thrombin‐initiated fibrin clot kinetic assay (TFCK), activated partial thromboplastin time (APTT), anti‐FXa activity, and thromboelastometry. Excessive postoperative bleeding was determined by measurement of chest tube output or the development of cardiac tamponade.

Results and Conclusions

Of 44 neonates enrolled, 16 (36%) had excessive postoperative bleeding. The TFCK value was increased. By heparin in neonatal blood samples, but was only minimally altered by excess protamine. Therefore, it reliably measured heparin in samples containing a wide range of heparin and protamine concentrations. The APTT most closely correlated with TFCK results, whereas anti‐FXa and thromboelastometry assays were less correlative. The TFCK and APTT assay also consistently detected postoperative heparin rebound, providing an important continued role for these long‐established coagulation tests in the management of postoperative bleeding in neonates requiring cardiac surgical repair. None of the coagulation tests predicted the neonates who experienced postoperative bleeding, reflecting the multifactorial causes of bleeding in this population.

     

Factor XI promotes hemostasis in factor IX‐deficient mice

Essentials

• Mice lacking factor IX (FIX) or factor XI (FXI) were tested in a saphenous vein bleeding model.
• FIX‐deficient mice displayed a hemostatic defect and FXI‐deficient mice were similar to wild type mice.
• Infusion of FXI or over‐expression of FXI in FIX‐deficient mice improved hemostasis.
• FXI may affect the phenotype of FIX‐deficiency (hemophilia B).

Summary

Background

In humans, deficiency of coagulation factor XI may be associated with a bleeding disorder, but, until recently, FXI‐deficient mice did not appear to have a hemostatic abnormality. A recent study, however, indicated that FXI‐deficient mice show a moderate hemostatic defect in a saphenous vein bleeding (SVB) model.

Objectives

To study the effect of FXI on bleeding in mice with normal levels of the FXI substrate FIX and in mice lacking FIX (a murine model of hemophilia B).

Methods

Wild‐type mice and mice lacking either FIX (F9^?) or FXI (F11^?/?) were tested in the SVB model. The plasma levels of FXI in F11^?/? mice were manipulated by infusion of FXI or its active form FXIa, or by overexpressing FXI by the use of hydrodynamic tail vein injection.

Results

F9^? mice showed a significant defect in the SVB model, whereas F11^?/? mice and wild‐type mice were indistinguishable. Intravenous infusion of FXI or FXIa into, or overexpression of FXI in, F9^? mice improved hemostasis in the SVB model. Overexpression of a FXI variant lacking a FIX‐binding site also improved hemostasis in F9^? mice.

Conclusions

Although we were unable to demonstrate a hemostatic defect in F11^?/? mice in the SVB model, our results support the premise that supraphysiological levels of FXI improve hemostasis in F9^? mice through FIX‐independent pathways.

     

Rivaroxaban reversal with prothrombin complex concentrate or tranexamic acid in healthy volunteers

Essentials

• Specific reversal agents for managing severe factor Xa inhibitor‐associated bleeding are lacking.
• We assessed 4‐factor‐prothrombin complex concentrate (4F‐PCC) and tranexamic acid (TXA).
• 4F‐PCC, but not TXA, reduced the prothrombin time and increased endogenous thrombin potential.
• These agents may be viable options for reversal of therapeutic doses of rivaroxaban.

Summary

Background

Oral activated factor X inhibitors such as rivaroxaban are widely used, but specific reversal agents are lacking. Although four‐factor prothrombin complex concentrate (4F‐PCC) and tranexamic acid (TXA) are sometimes used to manage serious bleeding, their efficacy is unknown. Prior studies in healthy subjects taking rivaroxaban revealed that 4F‐PCC partially reverses the prolonged prothrombin time (PT), and fully restores the endogenous thrombin potential (ETP). The effect of TXA has not been evaluated.

Methods

In this double‐blind, parallel‐group study, 147 healthy volunteers given rivaroxaban 20 mg twice daily for 3 days were randomized after their morning dose on day 4 to receive intravenous 4F‐PCC (50 IU kg^?1), TXA (1.0 g), or saline. Standardized punch biopsies were performed at baseline and after 4F‐PCC, TXA or saline administration. Reversal was assessed by measuring bleeding duration and bleeding volume at biopsy sites, and by determining the PT and ETP.

Results

As compared with saline, 4F‐PCC partially reversed the PT and completely reversed the ETP, whereas TXA had no effect. Neither 4F‐PCC nor TXA reduced bleeding duration or volume. All treatments were well tolerated, with no recorded adverse events.

Conclusions

Although 4F‐PCC reduced the PT and increased the ETP in volunteers given supratherapeutic doses of rivaroxaban, neither 4F‐PCC nor TXA influenced punch biopsy bleeding.

     

High and long‐term von Willebrand factor expression after Sleeping Beauty transposon‐mediated gene therapy in a mouse model of severe von Willebrand disease

Essentials

• von Willebrand disease (VWD) is the most common inherited bleeding disorder.
• Gene therapy for VWD offers long‐term therapy for VWD patients.
• Transposons efficiently integrate the large von Willebrand factor (VWF) cDNA in mice.
• Liver‐directed transposons support sustained VWF expression with suboptimal multimerization.

Summary

Background

Type 3 von Willebrand disease (VWD) is characterized by complete absence of von Willebrand factor (VWF). Current therapy is limited to treatment with exogenous VWF/FVIII products, which only provide a short‐term solution. Gene therapy offers the potential for a long‐term treatment for VWD.

Objectives

To develop an integrative Sleeping Beauty (SB) transposon‐mediated VWF gene transfer approach in a preclinical mouse model of severe VWD.

Methods

We established a robust platform for sustained transgene murine VWF (mVWF) expression in the liver of Vwf^?/? mice by combining a liver‐specific promoter with a sandwich transposon design and the SB100X transposase via hydrodynamic gene delivery.

Results

The sandwich SB transposon was suitable to deliver the full‐length mVWF cDNA (8.4 kb) and supported supra‐physiological expression that remained stable for up to 1.5 years after gene transfer. The sandwich vector stayed episomal (~60 weeks) or integrated in the host genome, respectively, in the absence or presence of the transposase. Transgene integration was confirmed using carbon tetrachloride‐induced liver regeneration. Analysis of integration sites by high‐throughput analysis revealed random integration of the sandwich vector. Although the SB vector supported long‐term expression of supra‐physiological VWF levels, the bleeding phenotype was not corrected in all mice. Long‐term expression of VWF by hepatocytes resulted in relatively reduced amounts of high‐molecular‐weight multimers, potentially limiting its hemostatic efficacy.

Conclusions

Although this integrative platform for VWF gene transfer is an important milestone of VWD gene therapy, cell type‐specific targeting is yet to be achieved.

     

Polymorphism in dural arteriovenous fistula: matrix metalloproteinase‐2‐1306 C/T as a potential risk factor for sinus thrombosis

Essentials

• Sinus thrombosis may play a crucial role in development of dural arteriovenous fistula (DAVF).
• Little is known about the association between gene polymorphism and the development of DAVF.
• MMP‐2‐1306 C/T showed a higher prevalence rate in DAVF cases with sinus thrombosis.
• MMP‐2‐1306C/T polymorphism is likely a potential risk factor for sinus thrombosis in DAVF.

Summary

Background

Dural arteriovenous fistula (DAVF) is a rare but important cerebrovascular disorder in adults. Little is known about the molecular genetic pathogenesis underlying DAVF development.

Objectives

To investigate the associations of gene polymorphisms and DAVF.

Materials and Methods

By the use of real‐time PCR genotyping, seven single‐nucleotide polymorphisms (SNPs) of angiogenesis‐related genes were analyzed in 72 DAVF patients. Pertinent clinical and imaging data were subgrouped on the basis of location (cavernous sinus versus lateral sinus), lesions (single versus multiple), cerebral venous reflux (CVR) grading (Borden I versus Borden II/III), and sinus thrombosis (with versus without).

Results

We found that individuals carrying the polymorphic allele of matrix metalloproteinase (MMP)‐2‐1306 C/T (rs243865) had a significantly increased risk of sinus thrombosis in DAVF (odds ratio 6.2; 95% confidence interval 1.7–22.9). There was a weak difference in associations of tissue inhibitor of metalloproteinase (TIMP)‐2 (rs2277698) gene polymorphism and DAVF patients subgrouped by CVR grading.

Conclusions

These preliminary results indicate that MMP‐2‐1306 C/T, but not MMP‐9, TIMP‐1, TIMP‐2, and vascular endothelial growth factor A SNP variants, is a risk factor for the development of sinus thrombosis in DAVF patients.

     

Reduced‐dose direct oral anticoagulants in the extended treatment of venous thromboembolism: a systematic review and meta‐analysis

Essentials

• In venous thromboembolism (VTE), benefits of extended treatment are balanced by bleeding risks.
• This is a meta‐analysis of reduced‐dose direct oral anticoagulants (DOACs) in extended treatment.
• Reduced‐dose DOACs are as effective as full anticoagulation with bleeding risks similar to placebo.
• Reduced‐dose DOACs are an attractive option for patients in the extended phase of VTE treatment.

Summary

Background

Extended‐duration anticoagulation is beneficial for preventing recurrent venous thromboembolism (VTE). Reduced‐dose direct oral anticoagulants (DOACs) may be preferable if they preserve efficacy and cause less bleeding. We conducted a systematic review and meta‐analysis of trials comparing reduced‐dose DOACs with full‐dose DOACs and aspirin or placebo in the extended phase of VTE treatment.

Methods

A literature search was conducted by use of the MEDLINE, EMBASE and CINAHL databases, supplemented by hand‐searching. One thousand three hundred and ninety‐nine titles were screened, with data from accepted studies being extracted by two independent reviewers. Major outcomes analyzed included recurrent VTE and major and clinically relevant non‐major bleeding events, presented as risk ratios (RRs) and 95% confidence intervals (CI).

Results

Two trials met the prespecified inclusion criteria. Data from 5847 patients were analyzed for efficacy outcomes, and from 5842 patients for safety outcomes. Reduced‐dose DOACs were as effective as full‐dose treatment in preventing recurrent VTE at 1 year (RR 1.12 [95% CI 0.67–1.87]), and more effective than aspirin or placebo (RR 0.26 [95% CI 0.14–0.46]). Rates of major or clinically relevant non‐major bleeding events were similar between patients receiving reduced‐dose DOACs and and those receiving aspirin or placebo (RR 1.19 [95% CI 0.81–1.77]). There was a trend towards less bleeding when reduced‐dose and full‐dose DOACs were compared (RR 0.74 [95% CI 0.52–1.05]).

Conclusions

Extended‐duration treatment of VTE with reduced‐dose DOACs may be as efficacious as full‐dose treatment, with rates of major bleeding being similar to those in patients receiving treatment with aspirin or placebo, but further long‐term studies are needed.

     

Anti‐C1 domain antibodies that accelerate factor VIII clearance contribute to antibody pathogenicity in a murine hemophilia A model

Essentials

• Inhibitor formation remains a challenging complication of hemophilia A care.
• The Bethesda assay is the primary method used for determining bleeding risk and management.
• Antibodies that block factor VIII binding to von Willebrand factor can increase FVIII clearance.
• Antibodies that increase clearance contribute to antibody pathogenicity.

Summary

Background

The development of neutralizing anti‐factor VIII (FVIII) antibodies remains a challenging complication of modern hemophilia A care. In vitro assays are the primary method used for quantifying inhibitor titers, predicting bleeding risk, and determining bleeding management. However, other mechanisms of inhibition are not accounted for in these assays, which may result in discrepancies between the inhibitor titer and clinical bleeding symptoms.

Objectives

To evaluate FVIII clearance in vivo as a potential mechanism for antibody pathogenicity and to determine whether increased FVIII dosing regimens correct the associated bleeding phenotype.

Methods

FVIII^?/? or FVIII^?/?/von Willebrand factor (VWF)^?/? mice were infused with anti‐FVIII mAbs directed against the FVIII C1, C2 or A2 domains, followed by infusion of FVIII. Blood loss via the tail snip bleeding model, FVIII activity and FVIII antigen levels were subsequently measured.

Results

Pathogenic anti‐C1 mAbs that compete with VWF for FVIII binding increased the clearance of FVIII–mAb complexes in FVIII^?/? mice but not in FVIII^?/?/VWF^?/? mice. Additionally, pathogenic anti‐C2 mAbs that inhibit FVIII binding to VWF increased FVIII clearance in FVIII^?/? mice. Anti‐C1, anti‐C2 and anti‐A2 mAbs that do not inhibit VWF binding did not accelerate FVIII clearance. Infusion of increased doses of FVIII in the presence of anti‐C1 mAbs partially corrected blood loss in FVIII^?/? mice.

Conclusions

A subset of antibodies that inhibit VWF binding to FVIII increase the clearance of FVIII–mAb complexes, which contributes to antibody pathogenicity. This may explain differences in the bleeding phenotype observed despite factor replacement in some patients with hemophilia A and low‐titer inhibitors.

     

Rivaroxaban for thromboprophylaxis among patients recently hospitalized for acute infectious diseases: a subgroup analysis of the MAGELLAN study

Essentials

• Net benefit of venous thromboprophylaxis (VTE) in patients hospitalized for infections is unknown.
• MAGELLAN trial subgroup analysis was performed for patients hospitalized for acute infectious diseases.
• At day 35, prolonged rivaroxaban prophylaxis reduced VTE compared to enoxaparin (4.2% vs. 6.6%).
• Rivaroxaban prophylaxis reduced VTE in patients hospitalized for active lung infections.

Summary

Background

Despite the well‐established association between infection and venous thromboembolism (VTE), there are few data specifically assessing the efficacy and safety of the VTE prophylaxis strategies for patients hospitalized for acute infectious diseases.

Objectives

To estimate the incidence of VTE and bleeding outcomes, comparing prolonged prophylaxis with rivaroxaban 10 mg daily for 35 days with enoxaparin 40 mg daily for 10 days.

Patients/Methods

A subgroup analysis of patients hospitalized for acute infectious diseases in the MAGELLAN trial was performed. The primary efficacy outcome was the composite of asymptomatic proximal or symptomatic VTE at days 10 and 35. The principal safety outcome was the composite of major or clinically relevant non‐major bleeding.

Results

Three thousand one hundred and seventy‐three patients with acute infectious diseases leading to hospitalization were randomized to either rivaroxaban (n = 1585) or enoxaparin/placebo (n = 1588), and received at least one dose of study medication. At day 10, primary composite efficacy outcomes did not differ between prophylaxis strategies (rivaroxaban, 2.7%; and enoxaparin, 3.7%). At day 35, there were fewer VTE events with rivaroxaban (4.2%) than with enoxaparin (6.6%) (relative risk [RR] 0.64; 95% confidence interval [CI] 0.45–0.92). Patients with pulmonary infections randomized to rivaroxaban had a lower incidence of VTE both at 10 days (RR 0.50, 95% CI 0.28–0.90) and at 35 days (RR 0.54, 95% CI 0.33–0.87). Primary safety outcome events were increased with rivaroxaban (RR 2.42, 95% CI 1.60–3.66).

Conclusions

Prolonged rivaroxaban prophylaxis reduced the incidence of VTE in patients hospitalized for acute infectious diseases, particularly those involving the lungs. Efficacy benefits were, in part, offset by bleeding outcomes. ClinicalTrials.gov Number: NCT 00571649.

     

Evaluation of a microfluidic flow assay to screen for von Willebrand disease and low von Willebrand factor levels

Essentials

• von Willebrand factor (VWF) function is shear stress dependent.
• Platelet accumulation in a microfluidic assay correlates with VWF levels.
• The microfluidic assay discriminates type 1 von Willebrand disease from healthy controls.
• The microfluidic flow assay detects responses to therapeutic intervention (DDAVP).

Summary

Background

von Willebrand disease (VWD) is a mucocutaneous bleeding disorder with a reported prevalence of 1 in 10 000. von Willebrand factor (VWF) function and platelet adhesion are regulated by hemodynamic forces that are not integrated into most current clinical assays.

Objective

We evaluated whether a custom microfluidic flow assay (MFA) can screen for deficiencies in VWF in patients presenting with mucocutaneous bleeding.

Methods

Whole blood from individuals with mucocutaneous bleeding was assayed in a custom MFA.

Results

Thirty‐two patients with type 1 VWD (10/32) or reported mucocutaneous bleeding were enrolled. The platelet adhesion velocity (r = 0.5978 for 750 s^?1 and 0.6895 for 1500 s^?1) and the maximum platelet surface area coverage (r = 0.5719 for 750 s^?1 and 0.6633 for 1500 s^?1) in the MFA correlated with VWF levels. Furthermore, the platelet adhesion velocity at 750 s^?1 (type 1 VWD, mean 0.0009761, 95% confidence interval [CI] 0.0003404–0.001612; control, mean 0.003587, 95% CI 0.002455–0.004719) and at 1500 s^?1 (type 1 VWD, mean 0.0003585, 95% CI 0.00003914–0.0006778; control, mean 0.003132, 95% CI 0.001565–0.004699) differentiated type 1 VWD from controls. Maximum platelet surface area coverage at 750 s^?1 (type 1 VWD, mean 0.1831, 95% CI 0.03816–0.3281; control, mean 0.6755, 95% CI 0.471–0.88) and at 1500 s^?1 (type 1 VWD, mean 0.07873, 95% CI 0.01689–0.1406; control, mean 0.6432, 95% CI 0.3607–0.9257) also differentiated type 1 VWD from controls. We also observed an improvement in platelet accumulation after 1‐desamino‐8‐d ‐arginine vasopressin (DDAVP) treatment at 1500 s^?1 (pre‐DDAVP, mean 0.4784, 95% CI 0.1777–0.7791; post‐DDAVP, mean 0.8444, 95% CI 0.7162–0.9726).

Conclusions

These data suggest that this approach can be used as a screening tool for VWD.

     

The chaperone‐like sodium phenylbutyrate improves factor IX intracellular trafficking and activity impaired by the frequent p.R294Q mutation

Essentials

• Missense mutations often impair protein folding, and thus intracellular trafficking and secretion.
• Cellular models of severe type I hemophilia B were challenged with chaperone‐like compounds.
• Sodium phenylbutyrate improved intracellular trafficking and secretion of the frequent p.R294Q.
• The increased coagulant activity levels (～3%) of p.R294Q would ameliorate the bleeding phenotype.

Summary

Background

Missense mutations often impair protein folding and intracellular processing, which can be improved by small compounds with chaperone‐like activity. However, little has been done in coagulopathies, where even modest increases of functional levels could have therapeutic implications.

Objectives

To rescue the expression of factor IX (FIX) variants affected by missense mutations associated with type I hemophilia B (HB) through chaperone‐like compounds.

Methods

Expression studies of recombinant (r)FIX variants and evaluation of secreted levels (ELISA), intracellular trafficking (immunofluorescence) and activity (coagulant assays) before and after treatment of cells with chaperone‐like compounds.

Results

As a model we chose the most frequent HB mutation (p.R294Q, ~100 patients), compared with other recurrent mutations associated with severe/moderate type I HB. Immunofluorescence studies revealed retention of rFIX variants in the endoplasmic reticulum and negligible localization in the Golgi, thus indicating impaired intracellular trafficking. Consistently, and in agreement with coagulation phenotypes in patients, all missense mutations resulted in impaired secretion (< 1% wild‐type rFIX). Sodium phenylbutyrate (NaPBA) quantitatively improved trafficking to the Golgi and dose dependently promoted secretion (from 0.3 ± 0.1% to 1.5 ± 0.3%) only of the rFIX‐294Q variant. Noticeably, this variant displayed a specific coagulant activity that was higher (~2.0 fold) than that of wild‐type rFIX in all treatment conditions. Importantly, coagulant activity was concurrently increased to levels (3.0 ± 0.9%) that, if achieved in patients, would ameliorate the bleeding phenotype.

Conclusions

Altogether, our data detail molecular mechanisms underlying type I HB and candidate NaPBA as affordable ‘personalized’ therapeutics for patients affected by the highly frequent p.R294Q mutation, and with reduced access to substitutive therapy.