Emicizumab and its Impact on aPTT-based Assays and FVIII Inhibitor Measurement

Objectives: Emicizumab (Emi) is a humanised bispecific antibody with a novel mechanism of action (MoA) - it bridges factor X (FX) and activated factor IX (FIXa) to restore haemostatic function; due to its biochemical properties, Emi interferes with various standard coagulation assays. Emi is approved for prophylactic treatment in people with haemophilia A (PwHA) and FVIII inhibitors. The aim herein is to provide a closer view of available data concerning the questions: how does Emi interfere with coagulation assays? Which assays can be applied in clinical routine?

Methods: A review was undertaken of the results and developments in the literature concerning the effects of Emi on aPTT-based coagulation assays and FVIII inhibitor measurement.

Results: How does emicizumab interfere with coagulation assays?

As Emi does not require activation by thrombin, it exhibits strong interference with aPTT-based assays. As such, even at low concentrations, Emi shortens aPTT, which affects several assays including aPTT-based single factor (FVIII, FIX, FXI, FXII) one-stage assays, protein C activity, protein S activity, and activated protein C resistance.[1] This aPTT shortening correlates with the normalisation of aPTT that occurred after the first dose of Emi at sub-therapeutic plasma concentrations, in the HAVEN 1 and 2 studies,[2] thereby suggesting higher protection levels than were actually present at the time. Moreover, Emi interferes with the standard one-stage, aPTT-based Bethesda assay, which is used to monitor FVIII inhibitor levels in PwHA. In addition to its interference with aPTT, Emi is not inactivated by heat treatment or FVIII inhibitors, thus leading to false-negative results with the standard Bethesda assay.[3]

Which assays can be reliably applied in clinical routine?

To permit clinical monitoring of Emi levels without interference, a modified aPTT-based FVIII assay, which uses a dedicated calibrator and controls for Emi, can be used. The results of this assay correlate with the results of an enzyme-linked immunosorbent assay (R²=0.98), allowing quantification of Emi across clinically expected concentration ranges.[4] As Emi does not bind to bovine forms of FIX and FX, a chromogenic Bethesda assay employing bovine components can be used to monitor FVIII inhibitor titres in the presence of Emi. This assay permitted the quantification of FVIII inhibitors in PwHA receiving Emi in HAVEN 1.[5]

Conclusions: Emi is the first haemophilia treatment product that can be quantified. Since Emi has a half-life of 4-5 weeks and exhibits small peak-trough fluctuations, it does not require routine monitoring as compared with other treatment options. However, for special clinical situations, measurement of its haemostatic active concentration or anti-FVIII inhibitor titres is possible. Emi interference should be considered when selecting and interpreting coagulation assays for PwHA receiving Emi.

• References

• 1 Calatzis A. , et al. Society of Thrombosis and Haemostasis Research Annual Meeting, 2018 (oral 5.4)
• 2 Schmitt C. , et al. European Association for Haemophilia and Allied Disorders, 11th Annual congress, 2018 (oral 9)
• 3 Adamkewicz JI. , et al. Hemostasis & Thrombosis Research Society, Scientific Symposium, 2017 (poster)
• 4 Calatzis A. , et al. Society of Thrombosis and Haemostasis Research Annual Meeting, 2018 (poster no. 238)
• 5 Adamkewicz JI. , et al. International Society on Thrombosis and Haemostasis Annual Meeting, 2017 (poster no. PB 954)