PB0991 - Assessment of Interference by Emicizumab in the Measurement of Susoctocog Alfa Factor VIII Activity Using a Chromogenic Assay

Background: Recombinant porcine factor VIII (rpFVIII, susoctocog alfa) is indicated for the treatment of bleeding episodes (BEs) in adults with acquired hemophilia A (AHA). A key benefit of rpFVIII is the ability to measure factor VIII (FVIII) activity in plasma samples using the activated partial thromboplastin time (aPTT) clotting assay. Emicizumab, an anti-FIXa/FX bispecific antibody, is being studied in patients with AHA to prevent or reduce the frequency of BEs, but it interferes with aPTT-based clotting assays.

Aims: To evaluate whether the FVIII activity of rpFVIII can be measured in the presence of emicizumab using a chromogenic assay.

Methods: The FVIII activity of rpFVIII was measured using the Coatest SP4 FVIII chromogenic assay (Chromogenix) on the BCS XP analyzer (Siemens). Samples (rpFVIII 0, 0.05, or 1.20 U/mL; emicizumab 0, 30, or 100 μg/mL) were made by mixing intermediate solutions prepared in pooled congenital FVIII-deficient plasma. High (1.500–0.150 IU/mL) and low (0.200–0.0075 IU/mL) calibration curves were prepared with normal reference plasma containing a known concentration of human FVIII (calibrated vs WHO 6th International Plasma Standard for FVIII). Samples were tested in duplicate at 3 dilutions either on the high curve (1:12, 1:24, and 1:48), prepared in SP4 assay buffer containing 1% bovine serum albumin, or on the low curve (1:5, 1:10, and 1:20), prepared in congenital FVIII-deficient plasma.

Results: FVIII activity in unspiked plasma (zero rpFVIII) was below the lower limit of quantitation ( < 0.030 IU/mL) at all tested emicizumab concentrations. The FVIII activity of samples containing rpFVIII (0.05 or 1.20 U/mL) and emicizumab (30 or 100 μg/mL) fell within ±20% of the FVIII activity in corresponding samples containing rpFVIII only. Emicizumab concentrations ≤100 μg/mL did not interfere with the assay.

Conclusion(s): The FVIII activity of rpFVIII can be measured accurately in the presence of emicizumab (≤100 μg/mL) using the chromogenic Coatest SP4 FVIII assay.