Hemostatic Challenges Research Articles

Inherited bleeding disorders

Abnormal bleeding is a common clinical presentation in general practice, and a rational approach to this problem is therefore required. Investigation of a suspected bleeding disorder necessitates a comprehensive history, thorough physical examination and systematic laboratory work-up. Inherited bleeding disorders (IBDs) typically manifest in childhood, but may present later in life after a haemostatic challenge (such as trauma, surgery, tooth extraction). This two-part CME series is intended to provide insight to the medical practitioner on the clinical spectrum, diagnosis and management of bleeding disorders. Bleeding due to inherited disorders is the subject of discussion in part 1 (current issue), and in part 2 (forthcoming issue) the focus is on bleeding from acquired causes. Patients diagnosed with an IBD should ideally be referred to a dedicated tertiary healthcare facility, e.g. haemophilia centre, for management and follow-up.

Perinatal lethality of Par4–/– mice delivered by primiparous dams reveals spontaneous bleeding in mice without platelet thrombin receptor function

Protease-activated receptor 4 (PAR4) is a cell surface G protein-coupled receptor for serine proteases, such as thrombin. Par4–/– mice have platelets that are unresponsive to thrombin and thereby allow examination of the importance of thrombin-induced platelet activation in (patho)physiology. Par4–/– mice are protected against arterial thrombosis but show no evidence of spontaneous bleeding. This contrasts with the bleeding experienced by mice with marked thrombocytopenia, such as those with genetic deficiency of the transcription factor, nuclear factor erythroid 2 (Nfe2–/–), that have high rates of perinatal death due to hemorrhage. Given this discrepancy in spontaneous perinatal bleeding between mice without platelets and those without thrombin-induced platelet activation mechanisms, we examined in detail the immediate postnatal survival of Par4–/– pups. We observed significant postpartum loss of Par4–/– pups derived from Par4+/– intercrosses that was restricted to a dam’s first litter; only 9% of surviving pups genotyped as Par4–/– in first litters and this normalized from the second litter onward (26%). A similar perinatal lethality in pups delivered by primiparous dams occurred in mice lacking platelets (Nfe2–/–; 10%) but not in those lacking fibrinogen (Fga–/–; 26%). These data,, provide the first evidence of spontaneous bleeding in Par4–/– mice, suggest that a dam’s first litter provides a greater hemostatic challenge than subsequent litters, and uncovers an important role for platelets—and more specifically thrombin-induced platelet activation—in hemostasis during these more traumatic births.

Challenges of Anticoagulation Therapy in Pregnancy.

Thrombotic complications in pregnancy represent a major cause of morbidity and mortality. Pregnancy is a primary hypercoagulable state due to enhanced production of clotting factors, a decrease in protein S activity, and inhibition of fibrinolysis. These physiologic changes will yield a collective rate of venous thromboembolism (VTE) of about 1-2 in 1000 pregnancies for the general obstetric population, which represents a five- to tenfold increased risk in pregnancy compared to age-matched non-pregnant peers. A select group of women, however, will carry a significantly higher rate of thrombosis due to primary thrombophilia, either inherited or acquired. This introduces a population of women who may benefit from prophylactic anticoagulation, either antepartum or postpartum. The coagulation changes that occur in preparation for the hemostatic challenges of delivery endure for several weeks postpartum. In fact, daily risk for pulmonary embolism (PE) is the highest postpartum. Use of anticoagulation in pregnancy introduces particular risk at the time of delivery, where bleeding and clotting risk collide. Altered metabolism rates of anticoagulants in pregnant women often necessitate closer monitoring than is required outside of pregnancy in order to ensure efficacy and safety. Heparin products are the mainstay of treating VTE in pregnancy, chiefly because they do not cross the placenta. In women with mechanical heart valves, the ideal anticoagulation regimen remains controversial as heparin use has shown inferior outcomes for preventing thromboembolic complications compared to warfarin, but warfarin carries risk for fetal embryopathy. Other populations where a heparin alternative is necessary include women with a history of heparin-associated thrombocytopenia (HIT) or other heparin intolerance. Further challenging the management of anticoagulation in pregnancy is the dearth of randomized clinical trials. The evidence governing treatment recommendations is largely based on expert guidelines, observational studies, or extrapolation from non-pregnant cohorts. A careful critique of a woman's history, as well as the available data, is essential for optimal management of anticoagulation in pregnancy. Such decisions should involve a multidisciplinary team involving obstetrics, hematology, cardiology, and anesthesia.

Factor XI‐deficient mice exhibit increased bleeding after injury to the saphenous vein

Background Factor XI (FXI) is a key component of the intrinsic pathway of coagulation. It can be activated by either FXIIa or thrombin and amplifies thrombin generation during clot formation. Congenital FXI deficiency in humans (known as hemophilia C) is associated with bleeding after hemostatic challenge. However, to date there are no reports of excess bleeding in FXI-deficient mice. Objectives To determine if the absence of FXI in mice prolongs bleeding in different models. Methods We assessed the hemostatic capacity of FXI-/- mice in three different bleeding models: tail bleeding, surgical bleeding and saphenous vein bleeding. Results We found that tail bleeding and surgical bleeding of FXI-/- mice were similar to wild-type mice. However, FXI-/- mice had an impaired hemostatic capacity in the saphenous vein bleeding model compared with wild-type controls. Conclusions Our results indicate that FXI-/- mice have a mild hemostatic defect after injury to the saphenous vein but not after transection of the tail or vessels in the abdominal wall.

Investigating von Willebrand Factor Pathophysiology Using a Flow Chamber Model of von Willebrand Factor-platelet String Formation.

Von Willebrand factor (VWF) is a multimeric glycoprotein coagulation factor that mediates platelet adhesion and aggregation at sites of endothelial damage and that carries factor VIII in the circulation. VWF is synthesized by endothelial cells and is either released constitutively into the plasma or is stored in specialized organelles, called Weibel-Palade bodies (WPBs), for on-demand release in response to hemostatic challenge. Procoagulant and proinflammatory stimuli can rapidly induce WPB exocytosis and VWF release. The majority of VWF released by endothelial cells circulates in the plasma; however, a proportion of VWF is anchored to the endothelial cell surface. Under conditions of physiological shear, endothelial-anchored VWF can bind to platelets, forming a VWF-platelet string that may represent the nidus of thrombus formation. A flow chamber system can be used to visually observe the release of VWF from endothelial cells and the subsequent platelet capture in a manner that is reproducible and relevant to the pathophysiology of VWF-mediated thrombus formation. Using this methodology, endothelial cells are cultured in a flow chamber and are subsequently stimulated with secretagogues to induce WPB exocytosis. Washed platelets are then perfused over the activated endothelium. The platelets are activated and subsequently bind to elongated VWF strings in the direction of fluid flow. Using extracellular histones as a procoagulant and proinflammatory stimulus, we observed increased VWF-platelet string formation on histone-treated endothelial cells compared to untreated endothelial cells. This protocol describes a quantitative, visual, and real-time assessment of the activation of VWF-platelet interactions in models of thrombosis and hemostasis.

Therapeutic plasma exchange for perioperative management of patients with congenital factor XI deficiency.

Factor XI (FXI) deficiency (hemophilia C [HEM-C]) is a bleeding disorder with unpredictable severity that correlates poorly with FXI coagulation activity (FXI:C). It poses a perioperative hemostatic management challenge. For US patients with severe disease, fresh frozen plasma (FFP) or, in current use, thawed plasma is the most readily available option but comes with risk of volume overload. We report our experience of using therapeutic plasma exchange (TPE) as an alternative perioperative management strategy. A retrospective review of all HEM-C patients who underwent surgical procedures. Data were collected, including demographics, bleeding history, surgical site, perioperative hemostatic intervention, and outcome. Between July 1997 and September 2014, 28 HEM-C patients (12 men) were identified, 4 with severe disease (FXI:C <2% or excessive bleeding). Nineteen patients underwent 91 invasive procedures. For nearly 60% of the procedures, no periprocedural hemostatic intervention was provided; before 4 procedures (3 patients), 1 plasma volume TPE preoperatively with FFP was administered. Patient 1, a 28-year-old woman (FXI:C, 35%) with a history of excessive surgical bleeding, underwent 2 TPE procedures before laparoscopic pelvic biopsy and subsequent abdominal hysterectomy with salpingo-oophorectomy that increased her FXI:C to 48%. Patient 2, a 79-year-old man (FXI:C, <2%), had TPE before total hip arthroplasty, increasing his FXI:C to 24%. Patient 3, a 59-year-old man (FXI:C, <2%), had TPE before prostate laser enucleation, increasing his FXI:C to 46%. Patients 1 and 3 had mild reactions during TPE; no patient had evidence of volume overload. All patients had adequate intraoperative surgical hemostatic outcomes. TPE is an effective alternative presurgical hemostatic intervention in HEM-C with potentially lower risk of circulatory volume overload.

A bleeding assessment tool correlates with intraoperative blood loss in children and adolescents undergoing major spinal surgery

Screening laboratory studies for bleeding disorders are of little predictive value for operative bleeding risk in adults. Predicting perioperative bleeding in pediatric patients is particularly difficult as younger patients often have not had significant hemostatic challenges. This issue is distinctly important for high bleeding risk surgeries, such as major spinal procedures. The aim of this study was to determine if the score of a detailed bleeding questionnaire (BQ) correlated with surgical bleeding in pediatric patients undergoing major spinal surgery. A total of 220 consecutive pediatric patients (mean age 14.2years) undergoing major spinal surgery were administered the BQ preoperatively, as well as having routine screening laboratory studies (i.e., PT, aPTT, PFA) drawn. A retrospective analysis was conducted to determine if there was a correlation between either the results of the BQ and/or laboratory studies with operative outcomes. A BQ score>2 showed a strong positive correlation with intraoperative bleeding based on both univariate and multivariate analyses. In contrast, abnormalities in screening laboratory studies showed no significant correlation with operative bleeding outcomes. Relying on screening laboratory studies alone is inadequate. The BQ used here correlated with increased intraoperative hemorrhage, suggesting this tool may be useful for assessing pediatric surgical bleeding risk, and may also be useful in identifying a subset of patients with a very low bleeding risk that may not require laboratory screening.

Management of recurrent thrombotic events in a pregnant patient

Pregnancy is associated with a procoagulant predisposition considered preparatory for the hemostatic challenge of delivery. During pregnancy, deep vein thrombosis (DVT) involves the left leg in 88% of cases. Our patient presents with atypical distribution of DVT. A primipara presented to our service with pain and thrombosed veins of the right calf and thight at 13 and 22 weeks of gestation. Doppler ultrasound excluded DVT. Treatment with topical heparin and elastic support was recommended. Follow-up revealed satisfactory progress. At 34 weeks of gestation, the patient presents to our department with acute pain of her right thigh. Doppler examination confirmed DVT at this atypical site. Systemic heparin therapy and follow-up were initiated. The patient recovered uneventfully, and the fetus was delivered at term via C-section. The awareness of venous conditions during pregnancy may prevent potentialy life-threatening complications for both the mother and fetus through optimal management of high-risk patients.

Factors influencing transfusion requirement in patients undergoing first-time, elective coronary artery bypass graft surgery.

CONTEXT:Coronary artery bypass graft (CABG) operation is associated with high frequency of allogeneic blood transfusion due to the acquired hemostatic challenges in patients undergoing CABG. However, allogeneic blood transfusion carries risks of infection, adverse reaction, and mortality as well as prolonged hospital stay and increased hospital cost. It is important to identify patients who require blood transfusion to mitigate their risk factors and reduce the chance of exposure to allogeneic blood.AIMS:This study was conducted to evaluate factors that influence the decision to transfuse red cell in first-time elective CABG patients.SETTINGS AND DESIGN:This was a cross-sectional study based on a retrospective record review. The study was done in the National Heart Institute.MATERIALS AND METHODS:All patients who underwent first-time elective CABG were included in this study. Variables analyzed include age, gender, body weight, preoperative hemoglobin (Hb) level, patients' comorbidities, and other clinical parameters.STATISTICAL ANALYSIS USED:Data were analyzed using SPSS software version 20.RESULTS:A total of 463 patients underwent first-time elective CABG during the period of the study. Three hundred and eighty-six (83.4%) patients received red cell transfusion. From multiple logistic regression analysis, only age (odds ratio [OR] = 1.040, 95% confidence interval [CI]: 1.003, 1.077, P = 0.032), body weight (OR = 0.951, 95% CI: 0.928, 0.974, P < 0.001), Hb level (OR = 0.500, 95% CI: 0.387, 0.644, P < 0.001), and cardiopulmonary bypass time (OR = 1.013, 95% CI: 1.004, 1.023, P < 0.001) were the significant independent predictors of red cell transfusion.CONCLUSIONS:By stratifying patients according to their risk factor for red cell transfusion, the high-risk patients could be recognized and should be enrolled into effective patient blood management program to minimize their risk of exposure to allogeneic blood transfusion.

Bleeding in Factor XI Deficient Mice

Introduction: Factor XI (FXI) is a key component of the intrinsic or contact pathway of coagulation. It can be activated by either FXIIa or thrombin and amplifies thrombin generation during clot formation. Humans with congenital FXI deficiency, a rare bleeding disorder also referred to as hemophilia C, have a mild hemostatic defect with rare spontaneous bleeding but bleeding after hemostatic challenge, as occurs with some instances of surgery or trauma (e.g. dental surgery, tonsillectomy, prostate surgery or in the postpartum period). In previous studies, FXI deficient mice did not have increased bleeding in a tail transection model. In contrast, we observed increased surgical wound bleeding in FXI deficient mice compared to wild-type mice, which promted us to further investigate the effect of an absence of FXI in mice on bleeding in different models.Methods: We assessed the hemostatic capacity of FXI-/- mice using two quantitative bleeding models (tail transection and saphenous vein injury). In the saphenous vein bleeding model, the saphenous vein was partially transected, clots that formed were removed and the total number of clots formed within a 30 minutes period (i.e. number of hemostatic events) were recorded under the microscope. In this model, a larger number of reforming clots corresponds to a higher hemostatic capacity.Results: We found that tail bleeding times of FXI-/- mice were similar to wild-type mice (median [range]: 63 [42 - 90] vs. 70 [60 - 114] seconds, p=0.119), which is consistent with previous studies. In contrast, we observed an impaired hemostatic capacity in FXI-/- mice compared with wild-type controls in the saphenous vein bleeding model (median number of hemostatic events [range]: (19 [9 - 26] vs. 25 [21 - 30], p=0.003). However, this phenotype was milder than the bleeding observed in FIX-/- mice (median number of hemostatic events [range]: 2 [0 - 4], p<0.001).Conclusions: Our results indicate that FXI-/- mice have a mild hemostatic defect after laparotomy and injury to the saphenous vein but not after tail transection. The mild increase in bleeding in FXI deficient mice after hemostatic challenge is similar to the increased bleeding observed in patients with FXI deficiency. Therefore, these mice may be useful in studying bleeding associated with FXI deficiency and in the presence of FXIa inhibitors. DisclosuresNo relevant conflicts of interest to declare.

Tissue factor deficiency increases alveolar hemorrhage and death in influenza A virus‐infected mice

Essentials H1N1 Influenza A virus (IAV) infection is a hemostatic challenge for the lung. Tissue factor (TF) on lung epithelial cells maintains lung hemostasis after IAV infection. Reduced TF-dependent activation of coagulation leads to alveolar hemorrhage. Anticoagulation might increase the risk for hemorrhages into the lung during severe IAV infection. Background Influenza A virus (IAV) infection is a common respiratory tract infection that causes considerable morbidity and mortality worldwide. Objective To investigate the effect of genetic deficiency of tissue factor (TF) in a mouse model of IAV infection. Methods Wild-type mice, low-TF (LTF) mice and mice with the TF gene deleted in different cell types were infected with a mouse-adapted A/Puerto Rico/8/34 H1N1 strain of IAV. TF expression was measured in the lungs, and bronchoalveolar lavage fluid (BALF) was collected to measure extracellular vesicle TF, activation of coagulation, alveolar hemorrhage, and inflammation. Results IAV infection of wild-type mice increased lung TF expression, activation of coagulation and inflammation in BALF, but also led to alveolar hemorrhage. LTF mice and mice with selective deficiency of TF in lung epithelial cells had low basal levels of TF and failed to increase TF expression after infection; these two strains of mice had more alveolar hemorrhage and death than controls. In contrast, deletion of TF in either myeloid cells or endothelial cells and hematopoietic cells did not increase alveolar hemorrhage or death after IAV infection. These results indicate that TF expression in the lung, particularly in epithelial cells, is required to maintain alveolar hemostasis after IAV infection. Conclusion Our study indicates that TF-dependent activation of coagulation is required to limit alveolar hemorrhage and death after IAV infection.

Heavy menstrual bleeding and health-associated quality of life in women with von Willebrand's disease.

Women with the inherited bleeding disorder von Willebrand's disease (VWD) face gender-specific hemostatic challenges during menstruation. Heavy menstrual bleeding (HMB) can negatively affect their overall life activities and the health-associated quality of life. The purpose of the present study was to investigate whether women with VWD experienced HMB and an impaired health-associated quality of life. The study subjects were recruited from the Coagulation Unit of Karolinska University Hospital. Information was retrieved from various self-administered forms and medical records. Of the 30 women (18-52 years) that were included in the present study, 50% suffered from HMB, although the majority received treatment for HMB. In addition, almost all the included women perceived limitations in the overall life activities due to menstruation. The health-associated quality of life for women with HMB was significantly lower (P<0.10) with regards to 'bodily pain' compared with Swedish women of the general population. In conclusion, women with VWD experienced reduced health-associated quality of life as a result of HMB. Therefore, preventing limitations in overall life activities and improving their health-associated quality of life thorough counseling on menstrual bleeding is important for women with VWD.

Management of bleeding in vascular surgery

Management of acute coagulopathy and blood loss during major vascular procedures poses a significant haemostatic challenge to anaesthetists. The acute coagulopathy is multifactorial in origin with tissue injury and hypotension as the precipitating factors, followed by dilution, hypothermia, acidemia, hyperfibrinolysis and systemic inflammatory response, all acting as a self-perpetuating spiral of events. The problem is confounded by the high prevalence of antithrombotic agent use in these patients and intraoperative heparin administration. Trials specifically examining bleeding management in vascular surgery are lacking, and much of the literature and guidelines are derived from studies on patients with trauma. In general, it is recommended to adopt permissive hypotension with a restrictive fluid strategy, using a combination of crystalloid and colloid solutions up to one litre during the initial resuscitation, after which blood products should be administered. A restrictive transfusion trigger for red cells remains the mainstay of treatment except for the high-risk patients, where the trigger should be individualized. Transfusion of blood components should be initiated by clinical evidence of coagulopathy such as diffuse microvascular bleeding, and then guided by either laboratory or point-of-care coagulation testing. Prophylactic antifibrinolytic use is recommended for all surgery where excessive bleeding is anticipated. Fibrinogen and prothrombin complex concentrates administration are recommended during massive transfusion, whereas rFVIIa should be reserved until all means have failed. While debates over the ideal resuscitative strategy continue, the approach to vascular haemostasis should be scientific, rational, and structured. As far as possible, therapy should be monitored and goal directed.

Assessing Bleeding Risk in Pediatric Patients Undergoing Major Spinal Surgery

Preoperative screening for bleeding disorders in pediatric patients is problematic due to children's limited exposures to significant hemostatic challenges and the inherent difficulty in obtaining blood samples from young patients. Overcoming these challenges is of particular importance for surgical procedures that carry a significant bleeding risk, such as spinal surgeries. Many pediatric surgeons, including the Pediatric Orthopedic team at our Institution, rely on an unfocused history and measurement of general markers of hemostasis for preoperative screening. In order to improve preoperative screening of pediatric patients undergoing spinal procedures, we instituted the use of a detailed semi-quantitative questionnaire based on the ISTH Bleeding Assessment Tool (BAT), in combination with evaluation of PT, aPTT, platelet count, and PFA. The BAT gives positive points for a personal or family history of bleeding, and negative points for significant hemostatic challenges that did not result in bleeding complications. It was decided a priori that a BAT score of ≥3 would result in referral to Pediatric Hematology. A total of 212 patients presenting for major spinal surgeries (e.g., spinal fusion, growth rod placement) ranging in age from 3 to 25 years were prospectively evaluated in this fashion. A total of 41 patients (19.3%) had a prolongation of the PT and/or aPTT, none of which had a high BAT score. The majority of the abnormal PT/aPTT values were minimal prolongations that were not reproducible on repeat testing. Prolongation of the PT and/or aPTT revealed 3 patients with mild deficiencies of either factors VII, X, or XI, none of which were felt to be clinically significant. Prolonged PFAs were observed in 32 patients (16%), 1 of which was diagnosed with type I VWD (BAT score = 1), and the other with "possible VWD" based on a borderline VWF antigen level (BAT score = 0). Both were treated with Humate P. The remainder of the patients with a prolonged PFA were determined not to have a significant bleeding disorder after further testing. A total of 15 patients were referred to Hematology based on a high BAT score. Of these, 2 had a history of thrombocytopenia (1 with known DiGeorge syndrome and 1 with Depakote-related thrombocytopenia). Neither required platelet transfusion. One patient with a high BAT score was known to have type I VWD and was treated with Humate P, another was diagnosed with low expression of glycoprotein GP1b and was treated with Humate P and platelet transfusion. The remainder of the patients with high BAT scores were not felt to have a clinically significant bleeding disorder based on a Hematologist's assessment. None of the 212 patients evaluated were felt to have excessive intraoperative bleeding by the surgical team, suggesting that none of the patients had a significant undiagnosed hemostatic defect. Together, these results suggest that reliance on history or screening labs alone may not be sufficient for many pediatric surgery patients. While the PFA identified 2 patients with mild/possible VWD that would have been missed by the BAT, the PFA also had a significant number of apparent false positives. The combination of a BAT and a platelet count, as well as assessment of VWF activity for patients without previous hemostatic system challenges, may provide a more effective screening methodology for institutions with ready access to VWF activity measurement. DisclosuresNo relevant conflicts of interest to declare.

The Light Chain of Rat Factor VIII Confers Its Superior Cofactor Activity

Hemophilia A is caused by genetic defect of human coagulation factor VIII (hFVIII) and patients have to take lifelong replacement therapy to prevent excessive bleedings upon hemostatic challenges. Due to the short half-life of hFVIII, replacement treatment has to be given frequently and inhibitors against infused hFVIII can be developed in about 20-30% of patients. These shortcomings have generated tremendous interest in developing HA gene therapies which is more efficient and long-lasting. However, early preclinical studies have shown FVIII activities were still limited after vector delivery. A Modified hFVIII with higher specific activity and pharmacodynamics properties is highly desirable to overcome the disadvantages of current protein replacement and gene therapy strategies.In the current study, we successfully constructed a B-domain deleted rat FVIII(rBDDF8) that contained a PACE/furin recognition site (RHQR) within a 14 amino acid linker between A2 and A3 domains. The rBDDF8 displayed significantly higher coagulation activity(~2.5-fold) than hBDDF8 after transfection into HEK 293 cells. In order to explore the mechanism for the observed superior cofactor activity, we constructed heavy chain(rHC) and light chain(rLC) of rFVIII. The rHC and rLC are able to reconstitute 5 times more FVIII activity than their human counter parts. However, when rHC is associated with human FVIII light chain (hLC), the reconstituted FVIII activity is lower that from hHC and hLC, suggesting that high coagulation activity of rFVIII is not mediated by its HC. On the contrary, when FVIII is constituted by hHC with rLC, we found that the activity is increased by 3~5-fold as against hHC and hLC. The hHC antigen level of FVIII reconstituted from hHC and rLC was 1.5-fold higher than that of hHC and hLC, suggesting that higher activity of FVIII with hHC and rLC is not through increased secretion. The specific activity deduced from activity/antigen ratio showed that FVIII with rLC is 3 times higher more than FVIII with hLC. To investigate the potential application of rFVIII in gene therapy, rBDDF8 was delivered in hemophilia A mouse model using AAV8 vectors. The high dose rBDDF8(4*1011 vg/mouse) resulted 2.5U FVIII activity at week 17, which is much higher(about 10-fold) than that of hBDDF8. When the rFVIII was delivered by dual chains strategy, i.e, administering vectors carrying only LC or HC simultaneously, it also showed 2-4 fold increased in FVIII activity. Interestingly, the combination of hHC and rLC also generated similar FVIII activity as rHC and rLC, further proving the rLC is the major contributor to the superior coagulation activity of rFVIII. Our results showed that the rFVIII has higher cofactor activity conferred by its LC. Our results suggest that rFVIII can be further exploited to make an ideal candidate for hemophilia gene therapy using AAV vectors. DisclosuresNo relevant conflicts of interest to declare.

The Class Effect of Tyrosine Kinase Inhibiotrs on Impaired Platelet Aggregation on Stimulation with Collagen and Arachidonic Acid

▪INTRODUCITONTyrosine kinase inhibitors (TKIs), the effective therapy for chronic myeloid leukemia (CML), has been reported to induce bleeding in patients, independent of thrombocytopenia. Some TKIs especially dasatinib and ponatinib can cause platelet function defect. We tested whether platelet dysfunction is associated with other TKIs, including imatinib and nilotinib. The clinical correlation of bleeding and platelet function has been explored.METHODSPatients with CML either newly diagnosed or receiving TKIs including imatinib, nilotinib an dasatinib were enrolled into the study. Those taking antiplatelet medications or having thrombocytopenia less than 100 x 109/μL were excluded. We tested platelet aggregation upon stimulation by 4 agonists, including 2.5 and 20-mmol/L ADP, 10-mmol/L epinephrine, 2-mg/mL collagen and 1-mmol/L arachidonic acid (AA). Abnormal platelet aggregation was defined as less than 60% of platelet aggregation upon stimulation by platelet agonists except single abnormality of platelet aggregation on stimulation with low-dose ADP or epinephrine. Clinical significance of bleeding was evaluated by bleeding scores.ResultsThere were 111 patients with CML, including 14 newly diagnosed patients and those receiving imatinib (n=75), nilotinib (N=18) and dasatinib (n=4). Majority of treatment naïve patients were in chronic phase (n = 11, 79%) whereas 2 and 1 patients were in blast phase and accelerated phase, respectively. Most of the patients on TKIs were in chronic phase (n = 94, 97%) while there were few patients in accelerated (n = 2) and blast phase (n =1). Dose intensity of imatinib was 300 mg (n=15), 400 mg (n=54), 600 mg (n=5) and 800 mg (n=1), while nilotinib dose was 600 mg (n=4) and 800 mg (n=14). Three and 1 patients received 100 mg and 50 mg of dasatinib, respectively. Of those taking TKIs, complete cytogenetic response (CCyR) and major molecular response (MMR) were achieved in 59% and 27% of patients while the minority of them achieved partial cytogenetic response (PCyR) or less (8%). Impaired platelet aggregation on stimulation with 20-mmol/L ADP was observed in 14%, 7%, 17% and 50% of patients who were treatment naïve, on imatinib, nilotinib and dasatinib, respectively (P =0.036). Patients on TKIs had higher proportion of patients with abnormal platelet aggregation on stimulation with collagen compared to treatment naïve patients (72% for imatinib, 56% for nilotinib, 75% for dasatinib and 36% for treatment naïve, P = 0.05). The corresponding figures for proportion of patients with abnormal platelet aggregation on stimulation with AA were 78%, 39%, 50% and 29%, P <0.001). Two out of 3 patients with normal platelet aggregation at diagnosis of CML had abnormal platelet aggregation on stimulation with collagen and AA after imatinib treatment. Combination of abnormal platelet aggregation upon stimulation with collagen and AA was the most common pattern of impaired platelet aggregation for TKIs (63% for imatinib, 33% for nilotinib and 50% for dasatinib). Assessed by bleeding score, none of patients with impaired platelet aggregation had significant clinical bleeding.ConclusionThe effect of TKIS on abnormal platelet aggregation appeared to be class effect, though, the prevalence was more common among patients treated with imatinib and dasatinib. Although, there was no apparent clinical correlation of TKIs induced abnormal platelet aggregation and clinical bleeding, CML patients on TKIS should be closed monitoring when undergoing any hemostatic challenges DisclosuresNo relevant conflicts of interest to declare.

Gynaecological and obstetrical bleeding in women with factor XI deficiency - a systematic review.

Menstrual bleeding, pregnancy and delivery present an intrinsic haemostatic challenge to women with bleeding disorders such as factor XI (FXI) deficiency. To provide a systematic overview of studies on gynaecological and obstetrical bleeding problems in women with FXI deficiency. We searched MEDLINE, EMBASE and the Cochrane library for studies that present original data on the incidence of and treatment options for gynaecological and obstetrical bleeding in FXI-deficient women. We identified 27 studies, including a total of 372 women with FXI deficiency. All studies were observational, no interventional treatment studies were found. Most patients had a mild deficiency (FXI ≥ 20 IU dL-1 ). Heavy menstrual bleeding (HMB) was reported in 7-67%. In 7/19 (37%) women who underwent gynaecological procedures, a bleeding complication occurred, including in 2/7 hysterectomies (29%). About 3-20% of reported pregnancies ended in a miscarriage; of these miscarriages 0-25% (4/23 miscarriages) were complicated by bleeding. Terminations of pregnancies (TOP) were complicated by bleeding in 4 out of 11 cases (36%). In 90 out of 498 (18%) deliveries a postpartum haemorrhage (PPH) was reported, ranging from 0 to 50% in individual studies. In 21% (66/321) of deliveries, prophylaxis was given. This was associated with 9% (6/66) PPH, compared to 19% in deliveries without prophylaxis (84/432). Epidural analgesia was performed without complications in 44 patients. Women with FXI deficiency have a clearly increased risk of HMB, and of bleeding complications after miscarriage, TOP and delivery. No high quality data are available regarding prophylactic treatment.

Targeting factor replacement therapy in severe hemophilia: which level is important?

The original aim of prophylactic replacement therapy was to convert the bleeding pattern of severe hemophilia to that of moderate hemophilia through regular infusions of clotting factor concentrates. However, targeting prophylaxis on minimum trough levels does not prevent all bleeding. At the group level, there is a clear association of factor levels with bleeding and outcome. But bleeding phenotype in individual patients shows large variation, independent of trough levels maintained. The association of peak levels with bleeding on prophylaxis is not established. Experience with surgery suggests that certain peak levels need to be achieved during other hemostatic challenges, such as playing sports. Individualization of prophylaxis should include timing of infusion according to special activities. The clinical relevance of factor levels is even more urgent since the recent introduction of long-acting clotting factor concentrates with their different pharmacokinetic profiles and the prospect of gene therapy resulting in constant factor levels. It should be considered that the success of any prophylactic regimen is also dependent on other factors, such as the age at initiation of prophylaxis, adherence, lifestyle, cartilage susceptibility, and the other components of the clotting system. Factor levels are thus an important but quite small piece in the total picture of treating hemophilia and we currently cannot identify a specific trough or peak level to use for monitoring. At the same time, knowledge of a patients' level during the infusion intervals may help to individualize and adjust treatment according to the clinical symptoms.

Unclassified bleeding disorders: outcome of haemostatic challenges following tranexamic acid and/or desmopressin.

In a minority of patients with a significant bleeding history no cause is found despite extensive testing and we diagnose such cases as unclassified bleeding disorders (UBD). UBDs may have diverse underlying causes and currently no standard management strategy exists in the event of a haemorrhage or to cover surgery. To document the clinical characteristics and response to treatment of UBDs. We performed a retrospective chart review of all patients with UBDs who had an invasive procedure at our centre between 1998 and 2014. The commonest symptoms were menorrhagia (89%) and bleeding at the time of surgery (88%) or dental extraction (85%). A total of 33 patients underwent 78 minor and major haemostatic challenges. Haemostatic cover was provided in 28 procedures with tranexamic acid alone, two with desmopressin and 45 with both agents in combination. A successful haemostatic outcome was observed in 70/78 (90%) cases. No patient required additional surgical intervention to achieve haemostasis, but one patient required a platelet transfusion to control postoperative bleeding. This is the first study to report on the investigation and treatment of UBD. Future studies are needed to further our understanding of the bleeding phenotype and identify any underlying causes.

Von Willebrand Disease: diagnosis and management

Abstract Von Willebrand Disease is a common cause of excessive bruising and bleeding in children. Its diagnosis can be challenging, not only because of the various subtypes of the disorder but because of the considerable overlap between VWD and normal individuals. Laboratory diagnosis requires a range of quantitative and qualitative tests of the VWF protein, with targeted gene analysis increasingly used to confirm the diagnosis of type 2 and type 3 VWD. Bleeding Assessment Tools may be helpful in directed laboratory testing but are often less so in young children who have had limited haemostatic challenges. The diagnosis and management of affected children is best undertaken by specialist Haemophilia Centres and, as with all bleeding disorders, will often require a multidisciplinary approach. Treatment for VWD may require the use of antifibrinolytic drugs, vasopressin or VWF-containing plasma concentrates: treatment is often on-demand for individual bleeding episodes but there are specific indications for the use of prophylactic treatment in some children. Given its prevalence and presenting symptoms, VWD should always be considered in the assessment of children suspected of non-accidental injury.