Will the ‘true’ factor level make itself known? Measuring factor therapy for treatment of hemophilia in the era of enhanced half‐life products

Factor (F)VIII N8GP, turoctocog alfa pegol, is a 40kDa glycopegylated version of the B domain truncated recombinant FVIII Novoeight (N8; Novo Nordisk). Variability in the predicted and measured factor level of N8GP between onestage clotting assay (OSA) and chromogenic factor VIII assay, and between various OSA depending on activated partial thromboplastin time (aPTT) reagents, notably those with silicabased activators has been observed.1,2 In Research and Practice in Thrombosis and Haemostasis, Persson and colleagues investigated the mechanisms underlying these differences.3 By measuring FXIa generated on contact activation, measuring activation of N8GP by thrombin and measuring FXa generation in the presence of various OSA aPTT reagents, the authors showed that the variable results are, at least in part, the result of the differing times of contact activation within each assay. Longer incubation times for contact activation in the OSA resulted in relatively higher levels of FXIa accumulation. This, together with an overall slower rate of activation of N8GP by endogenous thrombin formed in the OSA, resulted in underestimation of the N8GP level. Interestingly, shortening the contact activation time diminished the underestimation in reagents associated with a discrepancy, whereas prolonging the time resulted in underestimation in OSA previously not associated with a discrepancy. The underestimation was not the result of silica based activators per se, but a variability in assay conditions. The study highlights the problems involved in ensuring accurate measurement of enhanced halflife (EHL) product treatment where even modest differences in assays reaction conditions can give rise to apparently discrepant assay results. EHL replacement factor concentrates have increased the available treatment options for hemophilia A and hemophilia B. This new generation of concentrates promises effective treatment by increasing the treatment halflife while conserving the coagulant ability of FVIII or FIX. Prolonging the period between treatment infusions reduces the burden of treatment and will improve the quality of life for many patients. However, due to the relative novelty of the treatments, close monitoring, including laboratory monitoring, of EHL treatment is recommended for each patient.4 This may appear to be relatively straightforward, but it is increasingly apparent that EHL products may not behave in factor assays in an identical manner to the corresponding native plasma factor. Discrepancies between assays for recombinant factor products has been previously recognized, however, EHL factor concentrates are specifically designed to behave both differently and identically to the corresponding parent plasma factor. EHL factors are modified by attachment of moieties to the protein by chemical or recombinant means, to alter the life cycle of the protein. The modification either shields the protein from clearance receptors (PEGylation) or induces alternative clearance and recycling mechanisms (immunoglobulin FC fusion and albumin fusion). However, once activated, FVIIIa cofactor or FIXa enzyme supports coagulation near identically to the corresponding activated plasma factor. Given the nature of the designed modifications, it is perhaps then less surprising that the modified factors may behave differently to native plasma derived factor VIII or IX in onestage and/or chromogenic factor assays.