Optimal utilization of thrombophilia testing.

The global incidence of venous thromboembolism is high so laboratory testing for hereditary thrombophilia and selected acquired thrombophilia is common. Given the costs associated with testing and multiple pre-analytic and analytic variables affecting the assays, careful patient selection and timing of testing and diligent application to patient management are critical to providing high-value clinical care. Collaboration between the ordering providers and performing laboratories has the potential to achieve these goals. Herein, utility of thrombophilia testing, variables that affect the assays, and impact on patient management are reviewed. Where available, information on cost-effectiveness is discussed.

A single-centre study of haemostatic outcomes of joint replacement in von Willebrand disease and control patients and an analysis of the literature.

INTRODUCTION: Haemostatic assessments of patients with von Willebrand disease (VWD) who undergo total knee arthroplasty (TKA) and total hip arthroplasty (THA) have mainly relied on subjective parameters. AIMS: To compare objective haemostatic outcomes of TKA/THA in VWD patients and controls without bleeding disorders. METHODS: We retrospectively analysed haemostatic outcomes in VWD patients undergoing TKA/THA from 1993 to 2011 and compared them with two matched controls per operation. Using one-way analysis of variance, we tested the effect of VWD on bleeding risk after TKA and THA. RESULTS: Twelve VWD patients (6 type 1, 3 type 2M, 1 each of types 2A/2B/3) undergoing 19 operations (12 TKA, 7 THA) were matched to 38 controls. One (5%) of 19 operations in VWD patients and none of the control operations met clinical criteria for major bleeding. Baseline and postoperative day 1 haemoglobin levels, postoperative blood loss, transfused red blood cells (RBCs) and mean hospitalization days were not significantly different. More VWD patients than controls received RBC transfusions [12 (63%) vs. 12 (32%)]. Only 9 (47%) VWD patients vs. 38 (100%) controls received pharmacologic VTE prophylaxis. No postoperative symptomatic VTE occurred in either group. CONCLUSION: In this largest, single-institutional study, von Willebrand factor replacement based on daily levels resulted in low frequency of major bleeding in VWD patients after TKA/THA. RBC transfusion was more frequent compared with matched controls, but other objective measures of haemostasis were similar. Lack of sufficient details in published reports precluded comparison of haemostatic outcomes.

Do incident and recurrent venous thromboembolism risks truly differ between heterozygous and homozygous Factor V Leiden carriers? A retrospective cohort study.

INTRODUCTION: While Factor V Leiden (F5 rs6025 A allele) is a known venous thromboembolism (VTE) risk factor, VTE risk among heterozygous vs. homozygous carriers is uncertain. MATERIALS AND METHODS: In a retrospective cohort study of Mayo Clinic patients referred for genotyping between 1996 and 2013, we tested Factor V Leiden genotype as a risk factor for incident and recurrent VTE. RESULTS: Among heterozygous (n=268) and homozygous (n=111) carriers, the prevalence of VTE was 54% and 68%, respectively (p=0.016). While mean patient age at first VTE event (43.9 vs. 42.9years; p=0.70) did not differ significantly, median VTE-free survival was modestly shorter for homozygous carriers (56.8 vs 59.5 years; p=0.04). Sixty-nine (48%) and 31 (42%) heterozygous and homozygous carriers had ≥1 VTE recurrence (p=0.42). In a multivariable model, idiopathic incident VTE and a second thrombophilia were associated with increased and anticoagulation duration >6months with reduced hazards of VTE recurrence; Factor V Leiden genotype was not an independent predictor of recurrence. CONCLUSIONS: Aside from a higher VTE prevalence and modestly reduced VTE-free survival, VTE penetrance and phenotype severity did not differ significantly among homozygous vs. heterozygous carriers, suggesting that VTE prophylaxis and management should not differ by Factor V Leiden genotype.

Validation of an automated latex particle-enhanced immunoturbidimetric von Willebrand factor activity assay.

BACKGROUND: Laboratory diagnosis of von Willebrand disease (VWD) requires accurate measurement of plasma von Willebrand factor (VWF) activity. OBJECTIVES: To evaluate laboratory characteristics, diagnostic accuracy and testing utilities of an automated latex particle-enhanced immunoturbidimetric VWF assay (VWF:Lx) based on a monoclonal antibody recognizing the VWF-platelet glycoprotein (GP) Ib binding domain. METHODS: Laboratory characteristics including lower detection limit, linearity, precision, sample stability, and method comparison between VWF:Lx and VWF ristocetin cofactor activity by platelet aggregometry (VWF:RCo) were examined. To assess VWF:Lx diagnostic accuracy, 492 patient plasma samples, including 40 previously characterized VWD patient samples, were tested for VWF antigen (VWF:Ag) and VWF:RCo by either aggregometry or flow cytometry, and VWF:Lx with supplemental VWF multimer analysis when indicated. Based on results of VWF:Ag, VWF:RCo and VWF multimer analysis, and available clinical information, samples were categorized as: normal; VWD types 1, 2A/B, 2M, or severe 1 vs. 2M; or acquired VWF abnormalities (AVWA) due to subtle loss of highest molecular weight multimers. RESULTS: VWF:Lx had excellent laboratory characteristics and linear correlation with VWF:RCo (R(2) = 0.93). VWF:Lx accurately classified virtually all normal and VWD patient samples. Compared with VWF:RCo, VWF:Lx had superior sensitivity and specificity for distinguishing severe type 1 vs. 2M VWD and identifying AVWA. A proposed screening panel comprising VWF:Ag and VWF:Lx had 100% and 83% sensitivity for detecting VWD and AVWA, respectively. CONCLUSIONS: VWF:Lx has excellent laboratory characteristics and diagnostic accuracy compared with VWF:RCo, and can be used as part of an initial VWD screening panel and as a supplementary test.

Low risk of symptomatic venous thromboembolic events during growth factor administration for PBSC mobilization.

The use of erythropoietic agents has been associated with an increased risk of venous thromboembolic events (VTEs), especially in patients with underlying malignancies. However, it is not known whether there is an increased risk of VTE associated with granulocyte growth factors. We reviewed 621 patients undergoing PBSC mobilization using granulocyte growth factors, alone or in combination with CY. Patients with a diagnosis of AL amyloidosis (AL: 114; 18%), multiple myeloma (MM: 278; 44%) Hodgkin lymphoma (HL: 20; 3%) or non-Hodgkin lymphoma (NHL: 209; 33%) were included. Symptomatic VTE occurred in six (0.97%) patients: two AL, two MM and two NHL. Of the six patients, two had pulmonary embolism, one developed deep vein thrombosis and three developed symptomatic catheter related thrombosis. Two patients with AL had heparin-induced thrombocytopenia and thrombosis. We found a low incidence of VTE among patients undergoing PBSC mobilization.

To circumcise or not to circumcise? Circumcision in patients with bleeding disorders.

SUMMARY: Circumcision is one of the most common procedures performed in male neonates, but few published reports have described circumcision in patients with bleeding disorders. The aim of this study was to analyse outcomes of circumcision among children evaluated at our institution to determine the extent of complications and to provide guidelines for circumcision management. We searched our patient database for records of children who were followed up at the Mayo Clinic Comprehensive Hemophilia Center from 2000 through 2007 and who had been circumcised. We retrospectively reviewed the medical records to document complications and determine management strategies in this patient population. Of 55 children and young adults identified (median [range] age, 15 years [11 months to 21 years]), 48 patients were circumcised. Indications for circumcision were parental request (n = 45) and medical recommendation (n = 3). Twelve of 21 patients with a known bleeding disorder at the time of circumcision received factor replacement before the procedure. Three of these 21 patients had bleeding complications. Of the other 27 patients, who were diagnosed later in life as having a bleeding disorder, 8 had bleeding complications. The overall incidence of bleeding after circumcision was 23% (11/48). The 23% overall incidence of bleeding complications in our patients with bleeding disorders is comparable to that reported for patients without a bleeding disorder (0.1-35%). Some of our patients had significant bleeding despite adequate factor replacement before and after the procedure. Parents and patients must be aware that bleeding risk is a possibility despite adequate factor replacement for hemostasis.

Ror2, a developmentally regulated kinase, promotes tumor growth potential in renal cell carcinoma.

Inappropriate kinase expression and subsequent promiscuous activity defines the transformation of many solid tumors including renal cell carcinoma (RCC). Thus, the expression of novel tumor-associated kinases has the potential to dramatically shape tumor cell behavior. Further, identifying tumor-associated kinases can lend insight into patterns of tumor growth and characteristics. Here, we report the identification of the RTK-like orphan receptor 2 (Ror2), a new tumor-associated kinase in RCC cell lines and primary tumors. Ror2 is an orphan receptor tyrosine kinase with physiological expression normally seen in the embryonic kidney. However, in RCC, Ror2 expression correlated with expression of genes involved at the extracellular matrix, including Twist and matrix metalloprotease-2 (MMP2). Expression of MMP2 in RCC cells was suppressed by Ror2 knockdown, placing Ror2 as a mediator of MMP2 regulation in RCC and a potential regulator of extracellular matrix remodeling. The suppression of Ror2 not only inhibited cell migration, but also inhibited anchorage-independent growth in soft agar and growth in an orthotopic xenograft model. These findings suggest a novel pathway of tumor-promoting activity by Ror2 within a subset of renal carcinomas, with significant implications for unraveling the tumorigenesis of RCC.

Central venous access devices for paediatric patients with haemophilia: a single-institution experience.

Use of a central venous access device (CVAD) can facilitate early introduction of home-based infusion of factor concentrate for long-term prophylaxis or immune tolerance therapy in children with bleeding disorders. The aim was to review outcomes associated with use of CVAD. Retrospective review of paediatric patients with bleeding disorders was observed at the Mayo Clinic Comprehensive Hemophilia Center. Thirty-seven CVAD were placed in 18 patients (haemophilia A [n = 15], type 3 von Willebrand disease [n = 2] and haemophilia B [n = 1]). Follow-up was for 45 952 CVAD days, and median time that CVAD remained in place was 1361 days per device. Factor VIII (FVIII) inhibitors were present in 4 of the 15 patients. Ten CVAD-related infections occurred (median, 672 days; range, 72-1941 days), of which six were in one patient with FVIII inhibitors. Overall infection rate was 0.22 (95% confidence interval [CI], 0.10-0.40) per 1000 CVAD days, with 0.11 infections in patients without FVIII inhibitors compared with a pooled incidence of 0.66 (95% CI, 0.44-0.97) reported in the literature. Indications for removal of 27 CVAD were blockage, change to peripheral venous access, catheter displacement, infection, leak in the port septum, short catheter and skin erosion. No clinically apparent thrombosis or sequelae of thrombosis were observed. Infection is the most common complication associated with CVAD use and is increased in patients who have inhibitors. The low rate of clinically apparent thrombosis reflects our practice of not screening for thrombosis. The low infection rate reflects our practice of using and reinforcing the aseptic technique.

A highly-sensitive plasma von Willebrand factor ristocetin cofactor (VWF:RCo) activity assay by flow cytometry.

BACKGROUND: Assays of plasma von Willebrand factor (VWF) ristocetin cofactor activity (VWF:RCo) are essential for the laboratory diagnosis of von Willebrand disease (VWD) and for monitoring therapy. However, current manual or automated VWF:RCo assay methods have relatively poor operating characteristics. Our goal was to develop and validate a simple, accurate, specific and sensitive platelet-based VWF:RCo assay. METHODS: Using green or red fluorochrome-labeled, fixed normal platelets and normal or patient plasma, ristocetin-dependent and VWF-mediated platelet aggregation was detected by flow cytometry. VWF:RCo activity was assayed as the number of double-positive events (green and red) among all green or red events, relative to the calibrator plasma signal (6-150% or IU dL(-1)), and reported as percent or IU dL(-1). We tested plasma samples from normal donors (n = 51) and known VWD patients (type 1, n = 16; type 2, n = 17) based on clinical history, levels of plasma VWF antigen (VWF:Ag), VWF:RCo activity (manual platelet aggregometry/agglutination assay), factor (F) VIII activity and VWF multimer analysis. RESULTS: For normal donors and type 1 VWD patients, VWF:RCo activity by flow cytometry vs. manual platelet aggregation correlated closely (R2 = 0.74), and VWF:RCo/VWF:Ag ratios did not differ significantly. In contrast, VWF:RCo/VWF:Ag ratios for type 2 VWD subtypes were significantly lower using VWF:RCo by flow cytometry vs. manual platelet aggregation assay (P < 0.01), especially for type 2A VWD patients. CONCLUSIONS: This new flow cytometry-based VWF:RCo assay is simple, accurate, specific and sensitive, particularly for type 2 VWD.