CO-37 | STEW-EFA STUDY: MULTICENTER EVALUATION OF ONE-STAGE AND CHROMOGENIC ASSAYS FOR EFANESOCTOCOG ALFA MEASUREMENT ON STAGO AND WERFEN PLATFORMS
A.C. Molinari1, M. Marchetti2, A. Catalano3, L. Contino4, R. De Cristofaro5, A.B. Federici6, T. Martini7, I. Quaglia8, R. Rossi9, R. Santoro10, A. Tripodi11, A. Valpreda12 | 1Regional Reference Centre for Hemorrhagic Diseases, Department of Hemato-Oncology, IRCCS Istituto Giannina Gaslini, Genova, Italy; 2Department of Immunohematology and Transfusion Medicine, ASST Papa Giovanni XXIII, Bergamo, Italy; 3Società per la Promozione dell'educazione Medica (S.P.E.Me), Italy; 4Hemostasis and Thrombosis Unit, AOU SS Antonio e Biagio e C Arrigo, Alessandria, Italy; 5Dipartimento di Medicina e Chirurgia Traslazionale, Facoltà di Medicina e Chirurgia "Agostino Gemelli," Università Cattolica S. Cuore, Roma, Italy - Servizio Malattie Emorragiche e Trombotiche, Fondazione Policlinico Universitario "A. Gemelli" Istituto di Ricerca e Cura a Carattere Scientifico, Roma, Italy; 6School of Medicine and Surgery, University of Milan-Bicocca, Milano, Italy; 7Hemophilia Centre, Immune-Hematology and Transfusion Medicine, University Hospital “Città della Salute e della Scienza", Torino, Italy; 8Center for Thrombosis and Hemorrhagic Diseases, IRCCS Humanitas Research Hospital Rozzano, Milan, Italy; 9Laboratory of Clinical Chemistry and Hematology, University Hospital of Parma, Italy; 10Hemophilia, Hemostasis, and Thrombosis Unit, Azienda Ospedaliero Universitaria Dulbecco, Catanzaro, Italy; 11IRCCS Ca' Granda Maggiore Hospital Foundation and Angelo Bianchi Bonomi Hemophilia and Thrombosis Center, Milano, Italy; 12Central Laboratory Baldi and Riberi, AOU Città della Salute e della Scienza, Turin, Italy, reference laboratory of the Regional Center for Hemostasis and Thrombosis.
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Background. Efanesoctocog alfa is an ultra-extended half-life recombinant FVIII enabling once-weekly prophylaxis in hemophilia A. Structural modifications of the molecule may affect FVIII level measurements across different laboratory assays. Previous studies demonstrated relevant variability among activated partial thromboplastin time (aPTT) reagents, with potential under- or overestimation of FVIII activity. Reliable monitoring is essential at low trough levels and in patients previously treated with emicizumab, whose prolonged plasma persistence may interfere with coagulation assays. Limited data are currently available regarding the analytical performance of Stago and Werfen platforms, especially in these settings.
Aims. The STEW-EFA study aims to evaluate the accuracy and reproducibility of efanesoctocog alfa (EFA) measurement using one-stage clotting assays (OSA) and chromogenic substrate assays (CSA) on Stago and Werfen platforms. Secondary objectives include assessment of product-specific calibration strategies and evaluation of assay performance in samples containing both EFA and emicizumab.
Methods. STEW-EFA is a prospective multicenter analytical laboratory study promoted by the AICE Laboratory Quality Working Group. Sixteen Italian hemophilia laboratories will participate, 8 using the Stago platform and 8 using Werfen. FVIII-deficient plasma samples spiked with efanesoctocog alfa at nominal concentrations of 100, 50, 20, 10, and 5 IU/dL will be centrally prepared and distributed by ECAT. Additional samples containing combinations of emicizumab (20 or 50 µg/mL) and efanesoctocog alfa (50 or 100 IU/dL) will be analyzed using CSA. OSA testing will be performed using PTT-A® on Stago analyzers and SynthAFax® on Werfen analyzers. Chromogenic testing will employ TriniCHROM FVIII:C and Coamatic FVIII reagents, respectively. Each laboratory will test samples using both standard commercial calibration and product-specific, generated from serial dilutions of efanesoctocog alfa–spiked plasma (100 IU/dL). All measurements will be performed in triplicate duplicates over three independent analytical sessions using fresh reagents each day. Primary endpoints include assay accuracy, expressed as percentage recovery versus nominal concentration, and intra-/inter-laboratory reproducibility assessed by coefficient of variation (CV%). Secondary endpoints include comparison between standard and product-specific calibration approaches, comparison between analytical platforms, and evaluation of CSA performance in the presence of emicizumab.
Expected Results and Conclusions. The study is expected to provide real-world multicenter data on the analytical performance of Stago and Werfen assays for EFA monitoring. Product-specific calibration may improve assay accuracy, particularly at clinically relevant trough levels. STEW-EFA may contribute to harmonization of laboratory monitoring strategies for efanesoctocog alfa, including complex scenarios involving residual emicizumab interference, and could support future recommendations for coagulation laboratories managing patients with hemophilia A.
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