Differentiation of Acquired Hemophilia a from Lupus Anticoagulant Using Global Coagulation Assays and Magnesium-Dependent Activated Partial Thromboplastin Time Assay

Abstract

CONCLUSION Rapid and reliable differentiation between bleeding and thrombotic tendency in cases with prolonged activated partial thromboplastin time (APTT) remains challenging in clinical practice, in particular, distinguishing cases positive for coagulation factor inhibitor or lupus anticoagulant (LA). To differentiate acquired hemophilia A (AHA) from LA using three global coagulation assays: rotational thromboelastometry (ROTEM), thrombin generation assay (TGA), and clot waveform analysis (CWA); also to validate the effectiveness of magnesium-dependent APTT (APTT-Mg) assay. This method uses APTT-Mg and normal APTT using Ca to screen for LA, and APTT-Mg/APTT ratio is smaller in LA. We included patients with prolonged APTT without congenital coagulation factor deficiency who were referred to our department between November 2019 and September 2021. APTT prolongation was defined as >35 s (normal range: 25-35 s) measured routinely with Coagpia APTT-N (Sekisui Medical). The AHA group had a confirmed diagnosis and underwent treatment of AHA. The LA group was divided into LA1 (possible LA) and LA2 (definite LA). LA1 was defined by any of the following conditions: (1) index of circulation anticoagulant ≥15 in the cross-mixing test performed with Coagpia APTT-N; (2) positive dilute Russell's viper venom time (DRVVT) phospholipid neutralization method using Gladipore LA (Medical & Biological Laboratories); (3) positive APTT phospholipid neutralization test using Hemosil SCT (Instrumentation Laboratory); (4) positive for anti-CL antibody; (5) positive for anti-CLβ2GPI antibody; or (6) positive for phosphatidylserine-dependent anti-PT antibody. LA2 was defined by index of circulation anticoagulant >12.4 in the cross-mixing test performed by Thrombocheck APTT-SLA (Sysmex Corporation), or item (2) or (3) above, in accordance with criteria of LA in the International Society of Thrombosis and Haemostasis. LA1 and LA2 did not overlap, because cases that did not meet the conditions of LA2 were treated as LA1, and global coagulation assays were performed in all LA cases. In the global coagulation assays, 12 healthy volunteer samples were tested and regarded as the normal goroup. For the APTT-Mg assay, equal volumes of 25 mmol MgCl 2 and 25 mmol CaCl 2 were used instead of the normally used CaCl 2, and Thrombocheck APTT-SLA was used to measure APTT-Mg, which was then divided by the normal APTT concentration measured with the same APTT reagent. Ten patients in the AHA, 12 in the LA1, and 32 in the LA2 groups were eligible. ROTEM showed that clotting time in the non-activated rotational thromboelastometry assay (NATEM) mode was >3600 s (minimum 1363 s) for AHA, 501 s (IQR: 430-632 s) for LA1, 533 s (IQR: 443-641 s) for LA2, and 500 s (IQR: 429-645 s) for the normal group. TGA showed that peak thrombin concentration was 16 nM (IQR: 13-29 nM) in the AHA group, 242 nM (IQR: 175-273 nM) in the LA1 group, 174 nM (IQR: 129-259 nM) in the LA2 group, and 190 nM (IQR: 125-288 nM) in the normal group. In the CWA, Ad|min1| was 2.1 (IQR: 1.7-3.2) in the AHA group, 8.7 (IQR: 8.0-9.5) in the LA1 group, 6.7 (IQR: 5.8-8.4) in the LA2 group, and 10.3 (IQR: 9.8-10.6) in the normal group. APTT-Mg/APTT was 1.23 (IQR: 1.18-1.29), 1.26 (IQR: 1.23-1.38), and 1.05 (IQR: 0.95-1.20) in the AHA, LA1, and LA2 groups, respectively. ROTEM, TGA, and CWA effectively differentiated between the LA1/2 and AHA groups. Clotting time in non-activated rotational thromboelastometry assay mode (ROTEM) and peak height (TGA) were better at distinguishing high-titer LA cases from LA-positive AHA cases. APTT-Mg/APTT could be suitable for distinguishing AHA from high-titer LA; however, it may not be useful for differentiating AHA from low-titer or suspected LA.