Lupus Anticoagulant Screening Research Articles

Novel Ex Vivo DOAC Removal Methods Reduce Interference in Lupus Anticoagulant Testing.

Direct oral anticoagulants (DOAC) interfere in laboratory coagulation testing. The aim here was to study how commercial DOAC removal methods, DOAC Filter® and DOAC-Stop™, perform to eliminate DOAC concentrations and false positive results in lupus anticoagulant (LAC) testing. We acquired 50 patient samples with high concentrations of DOACs: apixaban (n = 18, range 68–572 ng/mL), dabigatran (n = 8, range 47–154 ng/mL), edoxaban (n = 8, range 35–580 ng/mL) and rivaroxaban (n = 16, range 69–285 ng/mL). DOACs were removed ex vivo with either DOAC Filter® (n = 28) or DOAC-Stop™ (n = 22). Additionally, commercial control and calibrator samples were studied (n = 13 for DOAC Filter®, n = 14 for DOAC-Stop™). LAC screening was performed before and after DOAC removal. Both DOAC Filter® and DOAC-Stop™ were effective in removing DOAC concentrations in samples: DOAC concentrations decreased to median of 0 ng/mL (range 0–48 ng/mL). Only one sample had more than residual 25 ng/mL of DOAC (apixaban). Before DOAC removal, 96% (48/50) of patient samples and over 90% (12/13 DOAC Filter®, 13/14 DOAC-Stop™) of control/calibrator samples were positive in the LAC screening. In patient samples, LAC screening turned negative in 61% (17/28) after DOAC Filter® and 45% (10/22) after DOAC-Stop™ treatment. All control samples became negative after DOAC removal. In conclusion, DOAC removal ex vivo reduces false positives in LAC screening. DOAC removal halved the need for confirmation or mixing tests- Although a subset of patients would require further testing, DOAC removal reduces unnecessary repeated LAC testing.

International multicenter, multiplatform study to validate Taipan snake venom time as a lupus anticoagulant screening test with ecarin time as the confirmatory test: Communication from the ISTH SSC Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibodies

BackgroundLupus anticoagulant (LA) assays are compromised in anticoagulated patients, and existing strategies to overcome the interferences have limitations. The prothrombin‐activating Taipan snake venom time (TSVT) screening test and ecarin time (ET) confirmatory test are innately insensitive to vitamin K antagonists (VKA) and direct factor Xa inhibitors (DFXaI). ObjectivesValidate standardized TSVT/ET reagents for LA detection, in a multicenter, multiplatform study. Patients/MethodsSix centers from four countries analyzed samples with TSVT/ET from 81 nonanticoagulated patients with LA, patients with established antiphospholipid syndrome (APS), and proven persistent LA who were either not anticoagulated (n = 120) or were anticoagulated with VKAs (n = 180) or DFXaIs (n = 71). Additionally, 339 nonanticoagulated LA‐negative patients, and 575 anticoagulated non‐APS patients (172 VKA, 403 DFXaI) were tested. Anticoagulant spiking experiments were performed and 112 samples containing potential interferences (i.e., direct thrombin inhibitors) were tested. Results were evaluated against locally derived cutoffs. Imprecision was evaluated. ResultsCutoffs were remarkably similar despite use of different analyzers and donor populations. Cutoffs for TSVT ratio, ET ratio, percent correction, and normalized TSVT ratio/ET ratio ranged between 1.08 and 1.10, 1.09 and 1.12, 9.3% and 14.8%, and 1.10 and 1.15, respectively. Coefficients of variation for TSVT and ET ratios were ≤5.0%. TSVT/ET exhibited sensitivity, specificity, and negative and positive predictive values of 78.2%/95.0%/86.3%/91.5%, respectively, with established APS as the LA‐positive population, and 86.9%/95.0%/76.8%/97.4%, respectively, with triple‐positive APS. Interference was seen with direct thrombin inhibitors, unfractionated heparin, and low molecular weight heparins, but not VKAs or DFXaIs. ConclusionsTSVT/ET are validated for LA detection in nonanticoagulated patients and those on VKAs or DFXaIs.

A SYSTEMATIC LITERATURE REVIEW OF BUDD-CHIARI SYNDROME IN PATIENTS WITH SYSTEMIC LUPUS ERYTHEMATOSUS

Antiphospholipid syndrome is associated with multiple and recurrent episodes of arterial and venous thrombosis, thrombocytopenia, spontaneous fetal losses, and high levels of antiphospholipid antibodies. These antibodies can also be present in systemic lupus erythematosus (SLE), and predispose patients with Lupus to a hypercoagulable state. Budd-Chiari syndrome (BCS) is characterized by hepatic venous outflow obstruction that can lead to hepatic congestion and it is deeply associated with acquired or inherited hypercoagulable states. Therefore, the aim of this study was to conduct a systematic literature review to analyze the association between Budd-Chiari syndrome and SLE. A systematic literature review was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. Articles were selected from PubMed/Medline, SciELO and LILACS databases in August 2021 using the search terms [Lupus] OR [SLE] AND [Budd-Chiari syndrome]. The inclusion criterion was limited to observational studies, case reports or case series that reported an association between Budd-Chiari syndrome and SLE. Language was restricted to English, Spanish or Portuguese. Among 76 articles initially identified, 31 were eligible for this review after full texts were read. The vast majority of the studies have linked the antibodies lupus anticoagulant and anticardiolipin as etiological factors of BCS in patients with lupus. One study has reported six cases in which factor V Leiden was strongly related to SLE, suggesting that this mutation should also be investigated in patients with lupus and BCS. Few studies have mentioned isolated deficiency of protein C and/or protein S as common etiologies of BCS in patients with lupus. Studies suggest that Lupus anticoagulant can be sought in patients with idiopathic hepatic venous thrombosis. Possible mechanisms by which antiphospholipid antibodies induce thrombotic events in patients with SLE have been suggested: it can bind to platelet membrane phospholipids and accentuate their adhesion and aggregation ability, it can interfere in the interactions of proteins C and S and impair the formation of the coagulation control complex (activated protein C, protein S and factor V). Budd-Chiari syndrome can be a rare complication in patients with SLE. A methodical search for hepatic vein occlusion in patients with SLE who develop unexplained hepatomegaly should be performed. In addition, screening for lupus anticoagulant and anticardiolipin antibodies should take place to reduce the percentage of apparently idiopathic BCS cases.

DIFFUSE ALVEOLAR HEMORRHAGE WITH LATE PRESENTING RIGHT VENTRICULAR THROMBI

TOPIC: Critical Care TYPE: Medical Student/Resident Case Reports INTRODUCTION: Diffuse alveolar hemorrhage is a life threatening emergency that can be caused by multiple etiologies. Pulmonary capillaritis, one common etiology, is particularly associated with autoimmune or inflammatory diseases. We present a challenging case of diffuse alveolar hemorrhage with associated acute hypoxic respiratory failure due to isolated pulmonary capillaritis with multiple late presenting intracardiac thrombi. CASE PRESENTATION: A 20 year-old woman presented with four days of myalgias, fever, and shortness of breath. Testing for pulmonary embolism and SARS-CoV2 was negative. She was diagnosed with pneumonia and treated with ceftriaxone and azithromycin. She then developed blood tinged mucous which was concerning for diffuse alveolar hemorrhage and received three days of pulse dose steroids. Seven days after initial presentation, she required escalation to intensive care. Her oxygen saturation was 96% on 70% FiO2 and 50 liters of pressure and subsequently required intubation for hypoxic respiratory failure. Imaging revealed diffuse bilateral infiltrates and bronchoscopy with lavage yielded mild endobronchial bleeding, but no overt hemorrhage. Pathology demonstrated neutrophilic capillaritis with diffuse alveolar damage. Prior outpatient autoimmune workup was significant only for mildly positive anti-nuclear antibody (speckled pattern 1:80). Transthoracic echocardiogram performed after intubation revealed two right ventricular thrombi that were not present on prior echocardiogram from admission. We initiated a heparin infusion in the setting of her alveolar bleeding and continued treatment with methylprednisolone. She then developed acute hemodynamic changes concerning for pulmonary embolism but stabilized without further intervention. The patient's oxygenation improved and she was successfully extubated. An antiphospholipid syndrome screen returned positive for a Dilute Russell Viper Venom Time (though notably value was 1.25 and upper limit of normal is 1.24) and anticoagulation was transitioned to warfarin prior to discharge home. DISCUSSION: Concurrent hemorrhage and hypercoagulability is a difficult management dilemma that many clinicians will face in the hospital setting. This case highlights the degree of suspicion required to evaluate for thromboembolic phenomena even in a patient with hemorrhage. In this young patient with no significant past medical history, autoimmune etiologies must be considered for her presentation of diffuse alveolar hemorrhage, which can also cause a concurrent hypercoagulable state. Despite mildly positive lupus anticoagulant screen, remainder of autoimmune workup was negative and suspicion low for systemic autoimmune disease. CONCLUSIONS: Bedside clinical judgement must drive decision making in the balance between worsening bleeding with anticoagulation or considering intervention for hemodynamically significant thromboembolic events. REFERENCE #1: Stoots SA, Lief L, Erkan D. Clinical Insights into Diffuse Alveolar Hemorrhage in Antiphospholipid Syndrome. Curr Rheumatol Rep. 2019;21(10):56. Published 2019 Sep 6. doi:10.1007/s11926-019-0852-7 REFERENCE #2: Park MS. Diffuse alveolar hemorrhage. Tuberc Respir Dis (Seoul). 2013;74(4):151-162. doi:10.4046/trd.2013.74.4.151 DISCLOSURES: No relevant relationships by Micah Fisher, source=Web Response No relevant relationships by David Wilhoite, source=Web Response

Applying index of circulating anticoagulant to mixing tests with lupus anticoagulant screen and confirm reagents can distinguish with high specificity between lupus anticoagulants and direct factor Xa inhibitors.

Lupus anticoagulants (LA) are detected by prolongation of clotting times for dilute Russell's viper venom time (dRVVT) and activated partial thromboplastin time (APTT) screening tests. Direct oral anticoagulants (DOACs) can interfere with both screening and confirmatory tests. The present study aimed to investigate the influence of direct factor Xa inhibitors (DiXaIs) on screen, confirm and mixing tests and establish a method for differentiation from other sample types. A total of 257 samples including nonanticoagulated LA positive, LA positive with DiXaIs, factor deficiency, FVIII inhibitors, warfarin and non-APS DiXaIs were tested. APTT reagents Cephen LS/Cephen and dRVVT reagents LA1/LA2 were used, respectively, to screen/confirm the study group. Index of circulating anticoagulant (ICA) was calculated from clotting times based on the following formula as ICA screening and ICA confirmation. ICA= (1:1 Mix sample - Normal pooled plasma) / Screen patient x 100. An ICA matrix was established which suggested the presence of a DiXaI when both ICA screening and confirmation were above the cut-off. When only ICA screening is elevated, LA is suspected. Sensitivity and specificity of the ICA matrix were 52.2% and 92.8% for DiXaIs and 38.1% and 96.7% for LA in APTT, and 61.2% and 92.9% for DiXaIs and 22.2% and 88.4% for LA in dRVVT, respectively. The ICA matrix achieved high specificity with a lower apparent sensitivity for DiXaI samples comparatively to other devices but due only to less interferences: the matrix could contribute to differentiating DiXaIs from LA in samples where anticoagulation status is unknown.

Testing lupus anticoagulants in a real-life scenario - a retrospective cohort study.

IntroductionLupus anticoagulant (LAC) testing is challenging. Most data are derived from a well-controlled study environment with potential alterations to daily routines. The aim of this retrospective cohort study was to assess the capacity of various LAC screening tests and derived mixing tests to predict a positive result in subsequent confirmation tests in a large cohort of patients.Materials and methodsIn 5832 individuals, we retrospectively evaluated the accuracy of the aPTT-A, aPTT-LAscreen, aPTT-FS and dRVVTscreen and of their derived mixing tests in detecting a positive confirmation test result within the same blood specimen. The group differences, degree of correlation and the predictive accuracy of LAC coagulation tests were analysed using the Mann-Whitney U test, the Spearman-rank-correlation and by area under the receiver operating characteristic curve (ROC-AUC) analysis. ROC-AUCs were compared with the Venkatraman´s permutation test.ResultsThe pre-test probability of patients with clinically suspected LAC was 36% in patients without factor deficiency or anticoagulation therapy. The aPTT-LAscreen showed the best diagnostic accuracy with a ROC-AUC of 0.84 (95% CI: 0.82 – 0.86). No clear advantage of the dRVVT-derived mixing test was detectable when compared to the dRVVTscreen (P = 0.829). Usage of the index of circulating anticoagulant (ICA) did not improve the diagnostic power of respective mixing tests.ConclusionsAmong the parameters evaluated, aPTT-LAscreen and derived mixing test parameters were the most accurate tests. In our study cohort, neither other mixing test nor the ICA presented any further advantage in LAC diagnostics.

Screening for lupus anticoagulants in patients treated with vitamin K antagonists.

The dRVVT test for detecting lupus anticoagulants (LA) is difficult to interpret when patients are treated with vitamin K antagonists (VKA). We performed LA testing in 33 VKA-treated patients with definite antiphospholipid syndrome (APS) and compared the results with 100 controls subjects not receiving VKA and 110 APL-negative patients anticoagulated for reasons other than APS. Compared with the dRVVT ratio before the initiation of VKA therapy, a higher cutoff value, defined as the 99th percentile, was established for VKA-treated patients with INR values between 2.0 and 3.5. A dRVVT ratio of >1.7 yielded a sensitivity of 81.3%, specificity of 99.1%, and positive and negative predictive values of 98.7% and 85.8%, respectively, for detecting LA. Cohen's kappa coefficient indicated good agreement for the dRVVT ratio obtained from testing with and without VKA treatment (κ = 0.813; 95% CI: 0.773-0.853), which was higher (κ = 0.941; 95% CI: 0.917-0.965) when the LA diagnosis was based on the results of both the dRVVT and a second test system (i.e., Mixcon-LA assay). Lupus anticoagulants testing in VKA-treated patients with APS according to current guidelines appears to be possible for the majority of patients without discontinuing anticoagulant therapy.

Genetic thrombophilia in patients with ischemic heart disease

In premature IHD, the role of genetic risk factors should be more important than environmental factors. Genetic thrombophilias represent a group of inherited conditions that predispose to thrombosis. Although the association between inherited thrombophilia and some thrombotic diseases as deep venous thrombosis is well-established, controversial data are present on the association between inherited thrombophilia and IHD. To study some genetic thrombophilic variants in patients suffering from ischemic heart disease. 72 subjects: 52 patients with premature ischemic heart disease <40 years. classified into: • Acute M.I. group:(31 patients). • Unstable angina group: (21 patients), and 20 apparently healthy individuals (of matched age and sex as a control group History taking, clinical and E.C.G. and laboratory investigations (cardiac markers, lipogram, lupus anticoagulants screening, antiphospholipids IgG/lgM and anticardiolipin IgG/lgM antibodies). Also, samples were taken for DNA extraction and typing with cardiovascular disease (CVD) StripAssay for the identification of mutations associated with CVD based on polymerase chain reaction (PCR) and reverse-hybridization molecular technique. The assay covers 12 polymorphisms. In UA patients group, the heterozygous (GA) genotype, and the (A) for UA, compared with the control group In addition, there were significant decreases in the frequencies of heterozygous (GT), and T allele of FXIII Val34Leu in UA patients group, compared with the control group assuming a protective role of FXIII 34Leu allele against unstable angina development. The prevalence of other studied polymorphisms; factor V H1299R (R2), Glycoprotein IIIa Leu33Pro,PAI-1 4G/5G, MTHFR C677T polymorphisms, MTHFR A1298C, apo B R3500Q and apo E polymorphisms; did not differ between IHD patients and control subjects. There was a significant positive association between FVL and family history of IHD in AMI patients. In addition, a highest prevalence of Pl A2 in patients <35 years of age surviving a first AMI, so that age-at-onset of AMI was significantly affected by PlA2 (<35 years vs. 35–40). Apo E3/E4 genotype was associated with both high total Cholesterol (TC) and high TC/HDL ratio in UA group, while in AMI patients, Apo E3/E4 genotype was associated with high TC/HDL ratio, and high triglycerides levels. This study has identified a positive association for FV Leiden, Prothrombin G20210A heterozygosity, and ACE (I/D) polymorphism with premature AMI; a positive association for β-fibrinogen 455-GA polymorphism in UA patients; and a negative association (protective role) for FXIII Val34Leu in UA patients. This study may open a dooor to promised goals of new therapeutics and personalised medicine for premature IHD based on single nucleotide polymorphism in well defined groups of high risk patients (<40 years); and their families.

Pulmonary Thromboembolism Associated With Clozapine

Pulmonary Thromboembolism Associated With Clozapine

Screening for lupus anticoagulant: improving the performance of the lupus-sensitive PTT-LA

The aim of the present work was to verify whether calculating a ratio between clotting times obtained with the sensitive PTT-LA and a less sensitive activated partial thromboplastin time (aPTT)-reagent may represent a valuable aPTT-based screening strategy for lupus anticoagulants (LA). For the pilot study, plasma samples from normal subjects (n = 15) and from patients with LA (n = 10), therapeutic anticoagulation with vitamin K-antagonists (VKA) (n = 15) or unfractionated heparin (n = 15), coagulation factors deficiency (n = 16), and inhibitory antibodies against factor VIII or IX (n = 11) were studied. For the evaluation study, 1553 consecutive plasma samples from nonanticoagulated patients investigated for LA between January 2005 and December 2007 at our institution were studied. Following screening strategies were employed: Pathromtin-SL (aPTT-SL), PTT-LA (aPTT-LA), ratio aPTT-LA/aPTT-SL (aPTT-ratio), and Russell's viper venom (RVV) based LA-Check. LA positive samples were identified by mixing studies and diluted RVV confirmation test (LA-Check/LA-Sure). Pilot study: All screening strategies had a 100% sensitivity, and the aPTT-ratio reached the highest specificity (82%; 95%CI: 74-90%). Within the evaluation study, following sensitivities for LA screening were observed: aPTT-SL 59.0% (95%CI: 57-61%), aPTT-LA 82.1% (95%CI: 80-84%), aPTT-ratio 92.3% (95%CI: 91-94), and LA-Check 83.3% (95%CI: 82-85%). Calculating a ratio between the LA-sensitive PTT-LA and the less sensitive Pathromtin-SL improves the performance of the PTT-LA itself and represents a simple and sensitive aPTT-based integrated strategy for LA screening.

Increased Thrombophilic Tendency in Pediatric Cystic Fibrosis Patients

Thrombophilia has recently been reported to be increased in patients with cystic fibrosis (CF). We wanted to determine whether this was applicable to our population with CF and how our patients compared to the previously reported groups. Seventy one pediatric CF patients were assessed for a thrombophilic tendency, using a lupus anticoagulant screen, protein C, protein S, antithrombin assay, and activated protein C resistance (APCR) screen. The incidence of activate protein C resistance (4.2%) was within expected limits for the general population as was the incidence of antithrombin deficiency. However there was a marked increase in the incidence of lupus anticoagulants (18%) and 14% and 19.7% of the patients showed a reduced protein C and protein S, respectively, far in excess of the general population. This increased incidence of thrombophilia was not related to any specific CF phenotype and while perturbed liver function cannot be entirely ruled out, it appeared unlikely to be responsible for all the abnormal coagulation findings. Despite the apparent thrombophilic tendency, no clinically evident thrombotic episodes were noted during the study period. Thrombophilia is of concern because of the increasingly frequent placement of indwelling catheters in CF patients. The precise cause for the thrombophilic tendency in CF patients is unknown at this stage.

Further evidence of false negative screening for lupus anticoagulants

Introduction Accurate and timely detection of lupus anticoagulants (LA) is of diagnostic and prognostic importance due to the association of persistent LA with thrombotic disease. Antibody heterogeneity and assay variability complicate LA detection and weak antibodies can go undetected. Methods Screen and confirm assays on equal volume mixing studies were performed on known LA using dilute Russell's viper venom time (DRVVT), dilute activated partial thromboplastin time (DAPTT) and activated seven lupus anticoagulant (ASLA) assay. Two established calculations for phospholipid dependence were applied to ascertain whether any antibodies diluted to within screening test reference ranges maintained a significant difference between screen and confirm results sufficient to imply LA activity. We then performed confirmatory tests on neat plasma samples from patients with thrombotic disease whose screening tests were within reference ranges. Results Forty nine of 155 DRVVT positive LA were conventionally positive in the mixing studies and 8 of the 106 negative mixing studies showed significant screen and confirm test discordance. This was the case for 21 of 56 negative DAPTT mixing studies and 2 of 39 negative ASLA mixing studies. Performance of confirm assays on the neat plasma samples with screen results within reference ranges revealed possible LA activity in 19 of 166 DRVVT results, 63 of 184 DAPTT results and 9 of 117 ASLA results. Conclusions LA activity can be demonstrated by assessment of screen and confirm data irrespective of screening test elevation above a reference range. Other workers have demonstrated this phenomenon in APTT using different study designs and it may be that standard interpretation criteria warrant re-assessment.

Cutaneous and systemic effects of varying doses of brown recluse spider venom in a rabbit model.

To ascertain whether a dose response exists between the dose of brown recluse spider venom (BRSV) and the cutaneous and coagulation effects in a rabbit model. Cutaneous necrosis is a serious complication of brown recluse spider envenomation (spider bite with venom). Disseminated intravascular coagulation (DIC) is a dreaded complication of brown recluse envenomation in humans. New Zealand white (NZW) rabbits have proved to be a model for the study of therapeutic regimens to prevent skin necrosis after spider bites. We studied the venom's effects on the skin and the coagulation mechanism in this rabbit model to determine if a clear dose-response relationship could be established. Establishment of a dose-response relationship is an important first step in determining if the NZW rabbit is a suitable model to study both cutaneous and systemic effects of the venom. Thirty-six NZW rabbits were divided into three groups. One group received a saline injection, and the other two groups received a 4.0 microg or a 10.0 microg dose of purified BRSV intradermally into the skin on the dorsum of the back. Blood was collected at baseline, 24, 48, and 72 hours. Tissue specimens were obtained after seven days during the animal necropsy and gross and microscopic pathology examination was conducted to assess tissue damage. Measurements included complete blood count (CBC); platelets; PT; activated partial thromboplastin time (APTT); fibrinogen (clottable, immunological); coagulation factors II, V, VII, VIII, IX, X, XI, XII; anti-thrombin (AT); alpha-2 antiplasmin (AP); Protein C (PC); mixing studies; lupus anticoagulant screening; plasminogen; thrombin-antithrombin; fibrin degradation products (FDP); d-dimer; and thrombin time. Gross pathology results were consistent with previous studies that used higher doses of BRSV. The WBC and platelet counts decreased at 24 hours in the two groups receiving the BRSV (p < 0.05). BRSV produced a dose related prolongation in the APTT (p < 0.05). Levels of fibrinogen as well as factors V, VII, VIII, IX, X, AT, and AP (p < 0.05) were increased in response to the BRSV. Protein C decreased at 24 hours (p < 0.05) and remained low in other time points. Mixing studies corrected the prolonged APTTs to normal ranges. Factor IIXI and XII showed no significant alteration in response to the BRSV. In the model, both the size and depth of the eschar were dose-related. We also observed a dose related elevation in the APTT that corrected with mixing studies. The dose-response relationship suggests direct interference by a component of the venom, rather than an idiosyncratic response. We did not detect a deficiency of commonly measured coagulation factors or evidence of a lupus anticoagulant. Protein C demonstrated a decrease. Although DIC did not occur in this rabbit model, a dose-related elevation in APTT was noted. The finding that the elevation corrected with mixing studies suggests that a plasma factor is essential in the coagulopathy associated with brown recluse envenomation. Further studies to identify this factor could shed light on human coagulopathy following envenomation.

Thrombophilic Patients with Migraines Have a High Incidence of Antiphospholipid Antibodies.

Migraines are associated with an increased risk of stroke and are recognized as a non-stroke neurologic complicaton in the antiphospholipid syndrome (aPls). Traditional tests for antiphospholipid antibodies (APA) include PTT assays [aPTT, dRVV, and Hexagonal phospholipid (Hex PL)], a dilute PT assay (Tissue Thromboplastin Inhibition) for lupus anticoagulants (LAC), and ELISA assays for anticardiolipin antibodies (ACA). These assays have not correlated with migraines. It is now recognized that antibodies to a phospholipid binding protein called beta-2-GPI are important in autoimmune aPls. Some patients (pts) with aPls are sero-negative, eg. negative in the routine assays, but have + anti-beta-2-GPI antibodies. Animal studies have shown an association of the latter antibodies to neurologic dysfunction. This study is a clinical retrospective medical record methodology, examining the relationship between migraines and APA results in pts referred to the Hemostasis & Thrombosis Clinic at the Institute for Transfusion Medicine from 7/1/03-7/15/05. Inclusion criteria were any pt. referred to the PI, who had a LAC screening panel and anti-beta-2-GPI testing. Exclusion criteria were any pt who did not have this testing or were + without repeat testing >2 mos later. Abstracted data included history of migraines (with or without aura), opthalmic migraines, referral reason, age, sex and APA results. Results were considered to be + (in non-anticoagulated pts), if any of the clotting assays were repeatedly +, did not correct on a mix, and shortened with phospholipid. Patients with only a + clotting assay, were taken off coumadin (OAC) and retested. Patients with + ELISA assays on OAC Rx, required an abnormal dRVV mix and/or a + Hex PL assay to be considered as + for a LAC. ELISA results were considered as +, if they were low titer or higher, e.g. ≥ mean ± 3 SD. The cohort consisted of 258 pts (69M: 189F) with a median age of 48 (range=13–85). Of these, 76 (29%) had migraines. Thirty-five/258 (13.6%) of the pts were referred for risk assessment and 10/35 (28.6%) had migraines. The remaining 223 pts were referred for clinical events: arterial or venous thrombosis [n= 133/233 (59.6%)], neurologic reasons [n=62/223 (28.2%)] and other [n=27/233 (12.1%)]. Of this group, migraines were present in 66 (29.6%). None of the 10 risk pts with migraines had + results. One of the 25 risk pts had a + anti-beta-2-GPI antibody (1/25=4.0%). Positive APA were detected in 58/233 (24.9%) pts with clinical events. Table 1 is a breakdown of the APA results vs migraines in pts with events vs those evaluated for risk issues. It appears that pts with thrombotic events with migraines have a higher incidence of +APA than those who do not have migraines. This data supports the suspected association between migraines and APA in the thrombphilic pt population. The same does not appear to be true in pts (without a thrombotic event) undergoing risk assessment.Incidence of Positive APA in Patients with regard to Migraine StatusGroupTotal PosTotal NegPORCIRisk-no migraine1/25 (4%)24/25 (96%)Risk-migraines0/10 (0%)10/10 (100%)1.000.8100.03–21.5Events-ALL pts58/233 (25%)175/233 (75%)0.00211.271.51 to 84.2Events-no migraine30/157 (19%)127/157 (81%)0.0585.830.77 to 44.22Events-migraines28/66 (42%)38/66 (58%)<0.00125.053.23 to 194.2APA = LAC/ACA and/or anti-beta-2-GPI

Is It Time To Reconfigure Lupus Anticoagulant Panels?.

Traditional lupus anticoagulant (LAC) panels are designed to test the presence/absence of antiphospholipid antibodies (APA). These usually include testing for a LAC with clot based assays (dRVV, Hex PL, aPTT) or with the tissue thromboplastin inhibition assay (TTI) and ELISA assays for anticardiolipin antibodies (ACA). It is now recognized that antibodies to a phospholipid binding protein, beta-2-GPI are important in the autoimmune form of the antiphospholipid syndrome (aPls). Moreover, some patients (pts.) may only be positive in this latter testing and have what is termed sero-negative aPls. The present study is a retrospective medical record review methodology to analyze the results of APA tests in 258 consecutive pts. referred to the PI at the Hemostasis & Thrombosis Clinic at the Institute For Transfusion Medicine. Inclusion criteria were any pt. who had a lupus panel and anti-beta-2-GPI (IgG, IgM and IgA) testing performed. Exclusion criteria were any pts. who did not have this testing or pts. with positive results which were not repeated > 2 months later. The cohort consisted of 258 pts (69M:189F) with a median age of 48 (range=13–85). Of these pts., 35 (13%) were referred for risk assessment, 133 (52%) for arterial or venous thrombosis, 62 (24%) for neurologic reasons, and 27 (11%) for other reasons. Patients on no anticoagulation (OAC) were considered to be LAC+, if any of the clot based assays was repeatedly positive, did not correct on a mix, and shortened with the additon of phospholipid. OAC pts. with positive ELISA assays were considered to have a LAC if the dRVV was positive on a mixing study and/or if the Hex PL test was positive. OAC pts. with only evidence for a LAC, were taken off OAC and then studied. ELISA assays were considered to be positive if low titer positive or higher, e.g. ≥ 3 STD above the mean. Positive results were found in 59/258 (22.8%) of these pts. The antibody results are listed in the table below in the pts. referred for risk assessment and in the pts. referred with events. The results are listed in categories of APA positivity.Antiphospholipid Antibody Results In Pts With Risk Assessment and Pts With Clinical EventsPt. GroupLAC+beta-2-GPILAC OnlyBeta-2-GPI OnlyACA OnlyTotalRisk Assessment0/35 (0%)0/35 (0%)1/35 (3%)0/35 (0%)1/35 (3%)Pts. With Events26/233 (11%)12/233 (5%)20/233 (9%)0/35 (0%)58/233 (25%)Total Positive Results26/59 (44%)12/59 (20%)21/59 (36%)059NOTE: 36% of the positive pts. would be missed by the traditional LAC panel.These results indicate that one-third of the pts. with phospholipid antibodies were sero-negative, e.g. negative in the presently constructed panels but positive for anti-beta-2-GPI antibodies. Skeptism exists about the meaning of isolated anti-beta-2GPI positivity. Of the pts. with only + beta-2-GPI antibodies, one male was asymptomatic (5%). The remaining pts. in this group were females. Ten/twenty-one (48%) of the pts. with isolated beta-2-GPI positivity were referred for neurologic reasons. These consisted of TIA's in 6, complex migraines with TIA's and acral cyanosis (1), recurrent fetal loss and TIA's in 3. Eight pts. (38%) had recurrent venous thromboembolism and 2 (9%) had both arterial and venous thromboembolism. We suggest that it may be time to consider constructing LAC screening panels containing tests for both LAC and anti-beta-2-GPI antibodies (all three isotypes) as sero-negative APls appear to be fairly common in females referred for thrombophilic testing.

Case 1: The girl who couldn't sit still

A seven-year-old Caucasian girl was referred to the hospital from the community with concerns about abnormal movements of her upper and lower extremities and slurring of speech. Before this, she had a two-week history of low grade fever, difficulty with feeding herself, deterioration in handwriting and trouble concentrating in school. She was seen by her family doctor at the onset and was believed to have had a viral infection. There was no history of a preceding sore throat or any concurrent symptoms such as headache, vomiting or fluctuating level of consciousness. Her past medical history was unremarkable. She was not taking any medications and there was nothing to suggest any intentional or accidental drug or poison ingestion. On examination, she was afebrile with stable vital signs. She was alert and cooperative but would not speak to hospital staff. She displayed facial grimacing, thrusting of the tongue and bilateral writhing nonpurposeful movements of her arms and legs, as well as the ‘milk maid’ sign when asked to squeeze the examiner’s fingers. She had a mild fleeting erythematous macular rash over her forearms. Cardiac auscultation revealed normal heart sounds, with no audible murmur or rub. The remainder of her examination was normal. Laboratory investigations showed a white blood cell count of 11.1×109/L (polymorphs 4.8×109/L, bands 1.1×109/L, lymphocytes 4.0×109/L), hemoglobin of 128 g/L, and platelets of 536×109/L. Her erythrocyte sedimentation rate was elevated at 30 mm/h (normal <10 mm/h). Throat swab, cerebrospinal fluid culture, blood and urine toxicology screens and a head CT scan were normal. Antinuclear antibody, anti-double stranded DNA antibody, and 24 h urine copper level and serum ceruloplasmin were normal. Anticardiolipin antibody and the lupus anticoagulant screen were both negative. A further laboratory test confirmed the diagnosis.

High prevalence of antiphospholipid antibodies in Asian cancer patients with thrombosis.

Thrombotic complications are a significant cause of morbidity and mortality in cancer patients. Studies in Caucasian populations have shown that up to one-third of such patients test positive to antiphospholipid antibodies. Our aim was to determine the prevalence and serotypes of antiphospholipid antibodies in an unselected group of Asian cancer patients with thrombosis. All patients with cancer-related thrombosis seen in the Department of Hematology-Oncology and Radiation Oncology were enrolled in this study. The study period was from April 2000 to May 2001. Antiphospholipid antibodies tests were performed, namely lupus anticoagulant screen, anticardiolipin antibodies (IgG and IgM) and anti-beta-2 glycoprotein I antibodies (B2 GPI) IgG, IgM and IgA. Thirty-three patients were recruited. There were 14 males and 19 females, with an age range of 35-78 years of age. Of those enrolled, there were 25 Chinese, five Malays and three Indians. The patients had several cancer types: 11 (36.7%) patients had adenocarcinoma as the histological cell type. Of the 33 patients, 75.8% had stage IV disease. Arterial thrombosis was seen in eight patients (24.2%), and venous thrombosis occurred in 29 patients (87.9%). Antiphospholipid antibodies were positive in 60.6% of the patients, of which anti-B2GPI IgA antibody was the most prevalent antiphospholipid present (46.9%). The presence of anti-beta-2 glycoprotein I IgA antibody was associated with strokes, extensive and recurrent venous thrombosis, and coincident arterial and venous thrombosis. A high prevalence of antiphospholipid antibodies (60.6%) was found in Asian patients with cancer-related thrombosis. The presence of antiphospholipid antibodies, particularly anti B2GPI IgA, may identify a subset of cancer patients who are at high risk of developing thrombotic complications, and further studies are warranted.

Lupus Anticoagulatiuon African Women With Recurrent Abortions

Objective: The presence of the lupus anticoagulant (LA) an auto antibody has been implicated as a marker for first trimester spontaneous abortions as well as second and third trimester foetal death in the Caucasians. There is paucity of information on LA in African women where recurrent abortion and obstetrics complications are still common. Our aim therefore is to determine the prevalence of lupus anticoagulant in Nigerian women with recurrent abortion Subjects and methods: A total of seventy – three pregnant women were studied. Twenty three of them had history of recurrent abortions; they were compared with 50 without obstetrics complications as controls. Coagulation studies using the kaolin clotting time were carried out on their plasma sample. Mixing experiments were conducted on samples with prolonged clotting time in order to detect the presence of the lupus anticoagulant. The anticoagulant was considered present if the kaolin clotting time ratio is greater than or equal to 1.2. Result: Nineteen {82.62%} of the patients with recurrent abortion had normal kaolin clotting time, one {4.35%} had subnormal clotting time. The kaolin clotting time ration was greater than 1.2 in one (4.35%) of the patients with recurrent abortions signifying the presence of the lupus anticoagulant. Four (8%) of the pregnant controls had subnormal kaolin clotting time, 44 (88%) had normal clotting time while 2 (4%) had prolonged clotting time. The lupus anticoagulant was present in one (2%) of the pregnant controls. Conclusion: A 4.35% prevalence of LA among Nigerians with recurrent foetal loss may necessitate the screening of pregnant women with history of recurrent abortion for the lupus anticoagulant. We recommend that screening for lupus anticoagulant should be part of the routine investigation for women with recurrent spontaneous abortions KEY WORDS: Recurrent Abortions, Pregnant Women, Lupus Anticoagulant, Kaolin clotting time Nigerian Journal of Clinical Practice Vol.6(1) 2003:26-29

Lupus anticoagulant testing: improvements in performance in a UK NEQAS proficiency testing exercise after dissemination of national guidelines on laboratory methods.

Laboratory screening for lupus anticoagulant (LA) has been shown to be suboptimal in several studies. Guidelines have recently been published by an expert group for the British Committee for Standards in Haematology, in an attempt to standardize and improve screening procedures. The value of using screening tests conforming with these guidelines was investigated in a United Kingdom National External Quality Assessment Scheme (UK NEQAS) proficiency testing exercise. The correct diagnosis was achieved by 97% of laboratories for a LA-negative sample. However, 18.3% of centres reported a false-negative result for a sample from a LA-positive subject. A significantly higher proportion of centres that used methods conforming with the published guidelines achieved the correct diagnosis for this sample (P < 0.002, chi-square test). A wide variety of screening tests were used by laboratories in this study. Within-method agreement could be improved by the use of a common normal pooled plasma to determine ratios. However, between-method agreement was not improved by this procedure. We conclude that adoption of methods compliant with national guidelines may improve the diagnosis of LA. There is a need, however, for reference and standardization materials to ensure further improvement in the accuracy of LA methods.

Lack of association between antiphospholipid antibodies and thrombocytopenia in patients with Wegener's granulomatosis

Background and Objective: In patients with Wegener's granulomatosis (WG), thrombocytopenia is less common than thrombocytosis. An increased prevalence of antiphospholipid antibodies (aPL), which is associated with thrombocytopenia, has been noted in patients with WG. The aim of this study was to examine the relationship between thrombocytopenia and aPL in patients with WG. Methods: Thrombocytopenic episodes were searched for in a random sample of 83 patients with WG. Stored sera obtained during thrombocytopenia, which was defined as platelet count below 130 × 109/L, were examined by 2 different enzyme-linked immunosorbent assays (ELISA) for IgG and IgM anticardiolipin antibodies (aCL) and for IgG antiphosphatidylserine antibodies (aPS). Screening for lupus anticoagulant was performed by use of activated partial thromboplastin time (aPTT). Results were compared with the prevalence of aPL in 20 consecutive nonthrombocytopenic patients with WG. Results: Six cases with thrombocytopenic episodes were found in the group of 83 patients with WG. Increased IgG and IgM aCL were detected in 1 patient, who also had elevated IgG aPS. A positive test result solely for IgM aCL was found in another patient. These findings were consistent in both ELISA for aPL. Five patients were being treated with cyclophosphamide when thrombocytopenia occurred. In the group of nonthrombocytopenic patients with WG, elevated IgG aCL and IgG aPS were consistently detected in 1 patient in both ELISA. Three other patients had positive results in single tests, which were not confirmed by the second assay. In all patients, aPTT was normal. Conclusions: Thrombocytopenia is a rare finding in patients with WG. A similar prevalence of aPL in thrombocytopenic and nonthrombocytopenic patients with WG provides no evidence that aPL play a major role in the pathogenesis of these events. Thrombocytopenia in WG is more likely caused by the myelotoxic effect of preceding cyclophosphamide treatment. We found a frequency of aPL in WG that exceeds frequencies seen in the general population but does not approximate those detected in systemic lupus erythematosus and closely related disorders. Semin Arthritis Rheum 31:4-11. Copyright © 2001 by W.B. Saunders Company