The combination of emicizumab and recombinant factor VIII in plasma: Which assays can we use for accurate measurement?

Abstract

The bispecific antibody, emicizumab, is a prophylactic therapy used for the treatment of haemophilia A (HA). Patients may require additional replacement factor VIII (FVIII) to ensure adequate haemostasis. This study investigated the laboratory measurement of severe HA (SHA) plasma spiked with 36 combinations of emicizumab plus recombinant (r) FVIII concentrates. FVIII assays were performed by one stage assay (OSA) using eight APTT reagents from three manufacturers and chromogenic assays (CSA) using seven kits. CSA kits comprised a range of bovine FX/FIXa, bovine FX/human FIXa or human FX/FIXa. Thrombin generation (TG) was assessed by CAT and ST-Genesia. Emicizumab-calibrated modified OSA and human FX CSA both overestimated rFVIII in the presence of emicizumab; median FVIII:C of up to 89% higher was observed in plasma spiked with both drugs compared to just rFVIII. In bovine FX CSA assays, there was a FVIII:C increase of up to 11% in plasmas spiked with both drugs compared to rFVIII alone. TG parameters were not all normalized by the presence of emicizumab however addition of rFVIII increased TG. ETP and peak thrombin were normalized at 50 μg/ml emicizumab using ST-Genesia but were still reduced at 75 μg/ml with CAT. Addition of rFVIII further normalized results. Modified OSA and human FX CSA could not distinguish between rFVIII or emicizumab. The presence of both emicizumab and rFVIII increased thrombin generation to normal levels compared to each drug alone. Bovine FX CSA can be used to accurately determine FVIII activity of rFVIII in plasma which also contains emicizumab.