Anti-idiotype monoclonal antibodies against emicizumab enable accurate procoagulant and anticoagulant assays, irrespective of the test base, in the presence of emicizumab.

Abstract

Emicizumab markedly shortens the activated partial thromboplastin time (aPTT), resulting in inaccurate measurements of procoagulant and anticoagulant factor activities. We have recently reported that mixtures of two different anti-idiotype monoclonal antibodies against emicizumab (anti-emicizumab-mAbs) allow measurement of factor (F)VIII activity (FVIII:C) and FVIII inhibitor in emicizumab-containing plasmas. It is unknown whether anti-emicizumab mAbs can work for other aPTT-based procoagulant and anticoagulant assays. To investigate whether anti-emicizumab mAbs were measured by all of the aPTT-based assays tested. Two anti-emicizumab-mAbs (300μg/mL each) were preincubated with emicizumab (200μg/mL)-spiked FVIII-deficient plasma; we then measured FVIII:C, FIX:C, FXI:C, FXII:C, protein (P)C:C, PS:C, global PC-FV (aPTT-based), and prothrombin time (PT), diluted Russel's viper venom time (dRVVT), chromogenic-based FVIII:C, FIX:C and PC:C (non-aPTT-based). Emicizumab (100μg/mL)-spiked haemophilia (H)A plasmas from patients (n=23) were also measured. Emicizumab shortened the clotting time in all aPTT-based assays, resulting in high levels of FVIII:C, FIX:C, FXI:C and FXII:C; low levels of PC:C and PS:C; and false-positive results for activated PC resistance. The addition of anti-emicizumab-mAbs to emicizumab-added plasma restored all factors to the initial levels without emicizumab. Chromogenic FVIII:C measurement by human FIXa/FX was affected by emicizumab, but anti-emicizumab mAbs cancelled this effect. PT-based assays and dRVVT, chromogenic FIX:C and PC:C assays showed no effect with emicizumab. Twenty-three plasma samples from HA patients also showed similar patterns. Anti-emicizumab mAbs in vitro could cancel the effect of emicizumab, irrespective of the test base, resulting in accurate measurements of procoagulant and anticoagulant factor activity.