In Vitro Effects of Mim8 and Combined Mim8-Bypassing Therapy on Thrombin Generation and Thromboelastography
Authors: Josset Laurie , Rezigue Hamdi , Lund Jacob , Dargaud Yesim 
Published: December 2024

Introduction: Primary prophylaxis is the gold standard of care for severe hemophilia. The development of subcutaneous Factor VIII (FVIII) mimetics has the potential to significantly reduce the disease burden and improve protection against bleeding episodes. Mim8, a fully human bispecific IgG4 antibody, mimics the function of activated FVIII (FVIIIa) by bridging activated factor IX (FIXa) and factor X (FX). This enhances the proteolytic activity of FIXa and allows effective activation of FX, making it suitable for prophylaxis in patients with hemophilia A. While Mim8 and emicizumab have similar modes of action, differences in their respective anti-FIXa and anti-FX arms influence their FVIIIa-like function.

Aim: This study investigates the in vitro hemostatic activity of Mim8 in blood samples from six patients with severe hemophilia A using a thrombin generation assay (TGA) and thromboelastography. The results were compared to those obtained with emicizumab. In addition, the combined hemostatic efficacy of Mim8 at 6 µg/mL was studied in combination with rFVIII at 100 IU/dL, rFVIIa at 25 mM (equivalent to a plasma concentration of a clinical dose of 90 µg/kg), and activated prothrombin complex concentrate (APCC) at doses of 20 and 30 U/kg.

Materials and Methods: Thrombin generation (TG) was measured in both platelet-poor plasma (PPP) and platelet-rich plasma (PRP) using the calibrated automated thrombogram (CAT, Stago®, France). For PPP, the PPP-low reagent containing tissue factor at a final concentration of 1 pM and phospholipids at 4 µM (Stago®, France) was used, with or without the addition of activated factor XI (FXIa) at 1.5 pM. For PRP, the PRP reagent containing tissue factor at a final concentration of 1 pM and without exogenous phospholipids (Stago®, France) was used. Whole blood samples were analyzed using NATEM (which operates without an exogenous coagulation activator) on a ROTEM delta device (Werfen®).

A previous study investigated the combined effect of Mim8 with rFVIIa and APCC. However, since the mechanism of action of rFVIIa is highly platelet-dependent, this study evaluated the combined in vitro effect of Mim8 and rFVIIa in PRP. The usual clinical dose of 90 µg/kg did not induce hypercoagulability in the TGA model (p&gt;0.05). As expected, no synergistic effect was observed when FVIII concentrate was combined with Mim8 at therapeutic doses due to the higher affinity of FVIII for FIXa and FX compared to Mim8. The peak thrombin values, which represent the maximum concentration of generated thrombin, correlate with the ETP results, reflecting the activity of thrombin throughout its lifespan.

ROTEM-NATEM analysis of whole blood from the same patients with severe hemophilia A confirmed the TGA results. Mim8 at 6 µg/mL significantly improved coagulation capacity as evidenced by a shorter coagulation time (CT) (p=0.0022) and a higher alpha angle (p=0.0043). The hemostatic activity of Mim8 at 6 µg/mL was also higher than that of emicizumab at 50 µg/mL (p=0.05).

Conclusions: Mim8 significantly improves TG in vitro in both PPP and PRP from patients with severe hemophilia A, with ETP levels comparable to those of FVIII at 100 IU/dL. The combination of Mim8 with rFVIIa at 90 µg/kg does not induce hypercoagulability, whereas the combination of Mim8 with APCC may carry a significant risk of hypercoagulability. The TGA can effectively monitor the combined treatment with Mim8 and either rFVIIa or APCC, which is not possible with currently available routine laboratory tests.