Evaluation of the activated partial thromboplastin time assay for clinical monitoring of <scp>PEG</scp>ylated recombinant factor <scp>VIII</scp> (<scp>BAY</scp> 94‐9027) for haemophilia A

TLDR

Hemophilia patients are monitored by aPTT assays, but different aPTT activators can alter clotting time and FVIII activity measurements for PEGylated FVIII. The study aimed to assess PEGylated FVIII (BAY 94‑9027) performance in various aPTT assays and identify optimal reagents for monitoring its activity in patients. The authors spiked WHO 8th FVIII standards and BAY 94‑9027 into severe hemophilia A plasma and compared five widely used commercial aPTT reagents for their ability to measure FVIII activity. Ellagic‑acid reagents (SynthAFax, Dade Actin, Cephascreen) produced clotting times and precision comparable to WHO standards, whereas silica‑based reagents (APTT‑SP, STA PTT 5) yielded prolonged clotting times and poor precision, and PEG alone dose‑dependently prolonged clotting time only in the APTT‑SP assay, leading the authors to recommend ellagic‑acid reagents as the preferred choice for monitoring BAY 94‑9027.

Abstract

Summary Patients with haemophilia ( PWH ) are usually monitored by the one‐stage activated partial thromboplastin time (aPTT) factor VIII (FVIII) assay. Different aPTT activators may affect clotting time ( CT ) and FVIII:C levels in patients treated with PEGylated FVIII. To evaluate the characteristics of PEGylated FVIII (BAY 94‐9027) in various aPTT clotting assays, and to identify suitable aPTT reagents for monitoring BAY 94‐9027 during the treatment of PWH , BAY 94‐9027 and World Health Organization ( WHO ) 8th FVIII standards ( WHO ‐8) were spiked into pooled and individual severe haemophilia A plasma at 1.0, 0.25 and 0.05 IU mL −1 . Five commercial aPTT reagents widely used in clinical laboratories were compared and evaluated for BAY 94‐9027 activity in plasma from PWH . BAY 94‐9027 and WHO ‐8 bestowed similar CT and excellent precision when ellagic acid (SynthAFax, Dade Actin, and Cephascreen) aPTT reagents were used. In contrast, BAY 94‐9027 showed significantly prolonged CT and poor precision compared with WHO ‐8 using silica aPTT reagents (APTT‐SP and STA PTT 5). Furthermore, free 60‐kDa polyethylene glycol ( PEG ), used for the conjugation of FVIII, showed a dose‐dependent prolongation of CT in the APTT‐SP assay. There was no effect on the SynthAFax‐APTT, prothrombin time, or FXIa‐initiated thrombin generation assay, demonstrating that the PEG moiety on FVIII has no general effect on the coagulation cascade. In summary, ellagic aPTT reagents (SynthAFax, Dade Actin, and Cephascreen) are most suitable for evaluating potency of BAY 94‐9027 and should be the preferred aPTT reagents used in clinical laboratories for monitoring FVIII activity after infusion of BAY 94‐9027 to PWH .