Accurate evaluation of factor VIII activity of efanesoctocog alfa in the presence of emicizumab

Background: Efanesoctocog is a B-domain-deleted, Fc-fusion factor (F)VIII linked to the D'D3 domain of von Willebrand factor and 2 XTEN polypeptides, designed for an ultra-extended half-life for prophylaxis in hemophilia A, but also aiding in managing acute bleeding or surgery in patients on long-term emicizumab. However, no current laboratory method accurately measures FVIII levels in the presence of emicizumab. Objectives: To test whether the bovine chromogenic FVIII assay, specifically calibrated for efanesoctocog, could provide an accurate assessment of efanesoctocog activity. Methods: Seven centers across 5 countries received 12 plasma samples to measure in triplicate using 2 calibration methods across 3 independent days. Samples (n = 6) contained either only efanesoctocog (FVIII activity [FVIII:C]= 5 to 150 IU/dL), or efanesoctocog (FVIII:C = 5 to 150 IU/dL) in combination with emicizumab (50 μg/mL; n = 5). One sample contained efanesoctocog (FVIII:C = 50 IU/dL) and a high dose of emicizumab (80 μg/mL); another sample contained efanesoctocog (FVIII:C = 50 IU/dL) with a low dose of emicizumab (20 μg/mL). Each center used its own analyzers, along with their usual reagents. Results: Chromogenic assay (CSA) calibrated with standard calibrators highly overestimates FVIII:C. However, specific calibration with efanesoctocog enabled accurate measurement of FVIII:C, with low inter- and intra-laboratory variability, and no interference from emicizumab. All CSA reagents used in the study demonstrated low variability across different laboratories (interlaboratory coefficient of variation ranges between 9% and 20%). Conclusion: Specific calibration of the FVIII CSA using efanesoctocog and bovine reagents allows for accurate measurement of FVIII:C in patients receiving efanesoctocog, even in the presence of emicizumab.