Effects of direct oral anticoagulants on lupus anticoagulant assays in a real-life setting.

Laboratory diagnosis of lupus anticoagulant (LA) is based on prolongation in at least one coagulation assay (diluted Russell's viper venom time - dRVVT or activated partial thromboplastin time - aPTT), which normalises after addition of phospholipids. Both assays may be influenced by anticoagulants and therefore LA should not be tested during warfarin or heparin treatment. It has been shown (primarily in vitro) that direct oral anticoagulants (DOACs - dabigatran [DAB], rivaroxaban [RIV] and apixaban [API]) may also influence LA testing. We tested the effects of DOACs on assays routinely used for the diagnosis of LA in patients treated with these drugs in a real-life setting. Plasma from patients with atrial fibrillation treated with DAB (n=30), RIV (n=20) and API (n=17) and not known to have LA were tested using dRVVT (LA-screen and LA-confirm, Life Diagnostics) and aPTT (PTT-LA, Diagnostica Stago and aPTT Actin FS, Siemens Healthcare Diagnostics) assays. According to the diagnostics algorithm, dRVVT and aPTT ratios of <1.2 were considered negative, ratios of >1.4 positive, while if the ratios were 1.2-1.4 LA could not be ruled out. Plasma concentrations varied between 8-172 µg/l for DAB, 8-437 µg/l for RIV and 36-178 µg/l for API. LA diagnosis was negative in only eight (27 %) plasma samples from patients treated with DAB, and in five (25 %) and four samples (24 %) from patients treated with RIV and API, respectively. LA Positivity (dRVVT and aPTT ratios >1.4) was found in 5 cases (17 %) among patients treated with DAB, in 10 cases (50 %) treated with RIV and in 7 cases (41 %) treated with API. A concentration-dependent effect of DOACs on dRVVT-based parameters was observed, particularly as regards DAB. At lower concentrations, RIV and API had only minor effects on the confirmatory tests (below 100 µg/l and 70 µg/l, respectively). Our results suggest that a risk of overestimation of LA detection is present in samples from patients treated with DOACs. Therefore, LA testing should not be performed during treatment with DOACs. Prolongation in confirmatory assays may be helpful for the recognition of false positivity, especially as regards DAB.

Evaluation of a diagnostic algorithm for Heparin-Induced Thrombocytopenia.

INTRODUCTION: Heparin-Induced Thrombocytopenia (HIT) is a rare but serious immune-mediated complication of heparin treatment. HIT is characterized by an acute, transient prothrombotic state combined with thrombocytopenia and is caused by platelet-activating IgG antibodies that bind to complexes of heparin and platelet factor 4. The diagnosis of HIT relies on clinical presentation and the demonstration of HIT antibodies. One approach to improve the efficacy of laboratory analysis is to use a diagnostic algorithm. AIM: To evaluate one diagnostic algorithm for HIT where the 4 T's clinical risk score is combined with immunochemical and/or functional assays. MATERIALS AND METHODS: The quality of the diagnostic algorithm was retrospectively evaluated in 101 patients with suspected HIT. Laboratory results obtained from the diagnostic algorithm were compared to Heparin-Induced Platelet Aggregation (HIPA) and clinico-pathological evaluation of patients' medical records. RESULTS: We found that the algorithm had a diagnostic efficacy of 94%, with specificity of 94% and sensitivity 94%. Positive likelihood ratio (LR+) was 16.0, and the negative likelihood ratio (LR-) 15.5. The efficacy of PaGIA (n=95) was 0.46, and IgG-specific HPF4-abELISA (n=54) was 0.87. CONCLUSIONS: The diagnostic algorithm for HIT is sufficiently accurate and leads to in overall faster results and decreased cost of analysis. The weakest link of the algorithm is the risk of miscalculated 4T's scores, which is inevitably exacerbated by the insufficient experience most clinicians have with HIT. This highlights the importance of clear instructions from the laboratory and coagulation clinic.

Evaluation of coagulation assays versus LC-MS/MS for determinations of dabigatran concentrations in plasma.

BACKGROUND: Dabigatran is an oral direct thrombin inhibitor for which routine laboratory monitoring is currently not recommended. However, there are situations in which measurements of the drug and its effect are desirable. We therefore compared and validated different coagulation methods for assessments of dabigatran in clinical samples in relation to measurements of plasma dabigatran, without the purpose of establishing effective and safe concentrations of dabigatran in plasma. METHODS: Samples were obtained from 70 atrial fibrillation patients treated with dabigatran etexilate. Plasma concentrations were measured using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and were compared with coagulation methods Hemoclot thrombin inhibitors (HTI) and Ecarin clotting assay (ECA), as well as with prothrombin time-international normalized ratio (PT-INR) and activated partial thromboplastin time (aPTT). RESULTS: A wide range of dabigatran concentrations was determined by LC-MS/MS (<0.5-586 ng/mL). Correlations between LC-MS/MS results and estimated concentrations were excellent for both HTI and ECA overall (r(2) = 0.97 and 0.96 respectively, p < 0.0001), but the precision and variability of these assays were not fully satisfactory in the low range of dabigatran plasma concentrations, in which ECA performed better than HTI. aPTT performed poorly, and was normal (<40 s) even with dabigatran levels of 60 ng/mL. PT-INR was normal even at supratherapeutic dabigatran concentrations. CONCLUSION: LC-MS/MS is the gold standard for measurements of dabigatran in plasma. Alternatively, either HTI or ECA assays may be used, but neither of these assays is dependable when monitoring low levels or to infer total absence of dabigatran. The aPTT assay is relatively insensitive to dabigatran, and normal aPTT results may be observed even with therapeutic dabigatran concentrations.