Masia, Tlangelani Vivian2024-11-182024-11-182024Masia, Tlangelani Vivian . (2024). Verification and implementation of a chromogenic Factor VIII assay to monitor Factor VIII levels in patients with haemophilia A on emicizumab therapy [Master’s dissertation, University of the Witwatersrand, Johannesburg]. WireDSpace.https://hdl.handle.net/10539/42643A research report submitted in partial fulfillment of requirements for the degree of Master of Medicine (Haematology) to the Faculty of Health Sciences, School of Pathology, University of the Witwatersrand, Johannesburg, 2023INTRODUCTION: Hemophilia A is an X-linked inherited bleeding disorder traditionally managed with replacement of the deficient clotting factor. A subset of patients however, develops inhibitors to infused FVIII rendering this treatment ineffective. Emicizumab is a novel, non-factor based therapy for patients suffering from hemophilia A with efficacy in the presence of inhibitors. Assessment of FVIII activity in patients on emicizumab treatment however, requires analysis with a chromogenic FVIII reagent such as the TriniCHROM FVIII:C (Stago) assay. METHODS: STAGO® engineers set-up the TriniChrome FVIII:C assay followed by 20 consecutive analyses of normal and pathological TriniChrome FVIII:C controls on both analyzers to assess precision. FVIII levels were measured in 50 samples from patients with (n 15) and without (n 35) hemophilia with a time-to-clot and the TriniCHROM FVIII:C assays to determine agreement between the 2. Samples from 10 patients with hemophilia A known to have FVIII inhibitors who were not exposed to emicizumab were also analyzed for FVIII inhibitor activity with both assays. FVIII levels in 10 patients with hemophilia A on emicizumab therapy were also determined with both assays. These were all for the accuracy study. Results were analyzed with standard statistical methods. RESULTS: Acceptable SD <8 according to the manufacturer and International Council of Standardization in Hematology and % CV <8.4 according to the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) were obtained on both analyzers. Acceptable agreement of FVIII levels between the 2 assays was obtained on the 50 random plasma samples with a linearity coefficient of determination (R²) of 0.92. FVIII inhibitor levels on the 10 samples from patients with known inhibitors were also within the significance category of more than 0.6 Bethesda Units (BUs) with both assays. FVIII levels of patients with severe hemophilia A on emicizumab therapy analyzed with the time-to-clot assay produced measurable results reflecting the endogenous activity of the drug but FVIII results with the chromogenic assay were similar to the patient’s baseline pre-treatment FVIII levels. CONCLUSION: The TriniCHROM FVIII:C assay is compatible with routine automated coagulation analyzers and can be utilised to assess endogenous FVIII and FVIII inhibitor levels in hemophilia A patients on emicizumab therapyen© 2024 University of the Witwatersrand, Johannesburg. All rights reserved. The copyright in this work vests in the University of the Witwatersrand, Johannesburg. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of University of the Witwatersrand, Johannesburg.Hemophilia AFactor VIII inhibitorsEmicizumabTime-to-clot FVIII assayChromogenic FVIII assayUCTDSDG-3: Good health and well-beingDissertationUniversity of the Witwatersrand, Johannesburg