In vitro studies show synergistic effects of a procoagulant bispecific antibody and bypassing agents

Background: Investigational non-factor products such as emicizumab offer a treatment option for patients with hemophilia and inhibitors. However, their mechanism of action raises questions regarding safety when they are combined with treatments for breakthrough bleeding. Objectives: To evaluate in vitro thrombin generation (TG) and clot formation for combinations of activated prothrombin complex concentrate (aPCC), recombinant activated factor VII (rFVIIa), and a sequence-identical analog of emicizumab (SIA). Methods: Therapeutic concentrations of SIA (20-600 nM) alone or with aPCC (0.05-1 U mL(-1)), isolated aPCC components or rFVIIa (0.885.25 mu g mL(-1)) were tested for TG and compared with reference ranges for healthy donor plasma. Coagulation of FVIII-inhibited blood was determined with a widely established method, i.e. rotational thromboelastometry (ROTEM), and confirmed with the Total Thrombusformation Analysis System. Results and conclusions: SIA (600 nM) or aPCC (0.5 U mL(-1)) alone resulted in peak thrombin levels of 21.4 nM and 38.6 nM, respectively, both of which are lower than normal (83.7 +/- 29.8 nM). SIA plus aPCC (0.5 U mL(-1)) increased the peak thrombin level 17-fold over SIA alone, exceeding the reference plasma value by 4.2-fold. This hypercoagulable effect occurred with 600 nM SIA combined with as little as 0.25 U mL(-1) aPCC, confirmed by ROTEM. FIX was the main driver for enhanced TG. SIA plus rFVIIa (1.75 mu g mL(-1)) induced a 1.8-fold increase in the peak thrombin level in platelet-rich plasma, but it did not reach the normal range. These in vitro experiments demonstrate excessive TG after administration of a combination of aPCC and SIA at clinically relevant doses. Careful judgement may be required when breakthrough bleeding is treated in patients receiving emicizumab.