Factor VIII Activity and Inhibitor Assays in the Diagnosis and Treatment of Hemophilia A

Abstract

The treatment of a patient with a factor VIII (FVIII) deficiency can be complicated. The mainstay of therapy is factor replacement. Replacement therapy can be given prophylactically, with the goal of decreasing hemarthroses and spontaneous hemorrhage, or on-demand for the bleeding patient. Intra- and interindividual variability in a patient's response to treatment has been well documented by the differences in observed half-lives of infused product. Although weight-based dosing nomograms are most often used, personalized therapies are coming into use to ease the burden of therapy and cost. The most significant complication of treatment is the formation of inhibitors to FVIII. The role of the laboratory is to provide results for FVIII activity that accurately reflects a patient's baseline level and response to treatment. However, factor activity assays have many components that can contribute to result variability. These include the methodology and reagent components used to measure the FVIII activity, the reference standard employed, algorithm used to interpret the dilutions, and the replacement factor being measured. An understanding of assay variables and their impact will assist in providing accurate factor activity results and appropriate patient care.

Keywords

hemophilia - treatment - factor VIII - inhibitor - one stage - chromogenic

• References

• 1 Mannucci PM, Tuddenham EGD. The hemophilias—from royal genes to gene therapy. N Engl J Med 2001; 344 (23) 1773-1779
• 2 Blanchette VS, Key NS, Ljung LR, Manco-Johnson MJ, ven den Berg HM, Srivastava A. For the Subcommittee on factor VIII, factor IX and rare coagulation disorders: Definitions in hemophilia: communication from the SSC of the ISTH. J Thromb Haemost 2014; 12: 1935-1939
• 3 Nilsson IM, Hedner U, Ahlberg A. Haemophilia prophylaxis in Sweden. Acta Paediatr Scand 1976; 65 (2) 129-135
• 4 Peyvandi F, Garagiola I, Seregni S. Future of coagulation factor replacement therapy. J Thromb Haemost 2013; 11 (Suppl. 01) 84-98
• 5 Collins PW, Fischer K, Morfini M, Blanchette VS, Björkman S ; International Prophylaxis Study Group Pharmacokinetics Expert Working Group. Implications of coagulation factor VIII and IX pharmacokinetics in the prophylactic treatment of haemophilia. Haemophilia 2011; 17 (1) 2-10
• 6 Oldenburg J. Optimal treatment strategies for hemophilia: achievements and limitations of current prophylactic regimens. Blood 2015; 125 (13) 2038-2044
• 7 Verbruggen B, Meijer P, Novákova I, Van Heerde W. Diagnosis of factor VIII deficiency. Haemophilia 2008; 14 (Suppl. 03) 76-82
• 8 Björkman S, Folkesson A, Berntorp E. In vivo recovery of factor VIII and factor IX: intra- and interindividual variance in a clinical setting. Haemophilia 2007; 13 (1) 2-8
• 9 Eich S, Kusch M, Grundmann C, Hanker C, Seitz R, König H. Factor VIII determination in patient's plasma and concentrates: a novel test equally suited for both matrices. Blood Coagul Fibrinolysis 2003; 14 (4) 347-353
• 10 Lundblad RL, Kingdon HS, Mann KG, White GC. Issues with the assay of factor VIII activity in plasma and factor VIII concentrates. Thromb Haemost 2000; 84 (6) 942-948
• 11 Favaloro EJ, Meijer P, Jennings I , et al. Problems and solutions in laboratory testing for hemophilia. Semin Thromb Hemost 2013; 39 (7) 816-833
• 12 Preston FE, Kitchen S, Jennings I, Woods TAL, Makris M. SSC/ISTH classification of hemophilia A: can hemophilia center laboratories achieve the new criteria?. J Thromb Haemost 2004; 2 (2) 271-274
• 13 Kitchen S, Gray E, Mertens K. Monitoring of modified factor VIII and IX products. Haemophilia 2014; 20 (Suppl. 04) 36-42
• 14 Favaloro EJ, Bonar R, Kershaw G, Mohammed S, Duncan E, Marsden K ; RCPA Haematology QAP Haemostasis Committee. Laboratory identification of factor VIII inhibitors in the real world: the experience from Australasia. Haemophilia 2010; 16 (4) 662-670
• 15 Peerschke EI, Castellone DD, Ledford-Kraemer M, Van Cott EM, Meijer P ; NASCOLA Proficiency Testing Committee. Laboratory assessment of factor VIII inhibitor titer: the North American Specialized Coagulation Laboratory Association experience. Am J Clin Pathol 2009; 131 (4) 552-558
• 16 World Health Organization. Delivery of treatment for hemophilia. Report of a joint WHO/WFH/ISTH meeting. London, United Kingdom, 11–13 February 2002. Available at: http://www1.wfh.org/publication/files/pdf-1451.pdf . Accessed December 10, 2015
• 17 Stachnik J. Hemophilia: Etiology, complications, and current options in management. Formulary 2010; 45: 218-227
• 18 Stobart K, Iorio A, Wu JK. Clotting factor concentrates given to prevent bleeding and bleeding-related complications in people with hemophilia A or B. Cochrane Database Syst Rev 2005; CDOO3429 (2) CD003429
• 19 Ar MC, Vaide I, Berntorp E, Björkman S. Methods for individualising factor VIII dosing in prophylaxis. Eur J Haematol Suppl 2014; 76 (Suppl. 76) 16-20
• 20 Tang L, Leong L, Sim D , et al. von Willebrand factor contributes to longer half-life of PEGylated factor VIII in vivo. Haemophilia 2013; 19 (4) 539-545
• 21 Collins PW. Personalized prophylaxis. Haemophilia 2012; 18 (Suppl. 04) 131-135
• 22 Denvigl IE, Maeser Bonschoren EP, Rosendaal G , et al. Clinical Severity of Hemophilia A: Does the classification of the 1950's still stand?. Heamophilia 2011; 17: 349-353
• 23 Björkman S, Folkesson A, Jönsson S. Pharmacokinetics and dose requirements of factor VIII over the age range 3-74 years: a population analysis based on 50 patients with long-term prophylactic treatment for haemophilia A. Eur J Clin Pharmacol 2009; 65 (10) 989-998
• 24 Björkman S, Blanchette VS, Fischer K , et al; Advate Clinical Program Group. Comparative pharmacokinetics of plasma- and albumin-free recombinant factor VIII in children and adults: the influence of blood sampling schedule on observed age-related differences and implications for dose tailoring. J Thromb Haemost 2010; 8 (4) 730-736
• 25 Björkman S, Collins P ; Project on Factor VI I I/Factor IX Pharmacokinetics of the Factor VIII/Factor IX Scientific and Standardization Committee of The Isth. Measurement of factor VIII pharmacokinetics in routine clinical practice. J Thromb Haemost 2013; 11 (1) 180-182
• 26 Determination of factor Coagulant Activities, Approved Guideline . NCCLS document H48A, NCCLS, 940 West Valley Road Suite 1400, Wayne, PA 19087-1898, 1997
• 27 Keeling DM, Sukhu K, Kemball-Cook G, Waseem N, Bagnall R, Lloyd JV. Diagnostic importance of the two-stage factor VIII:C assay demonstrated by a case of mild haemophilia associated with His1954—>Leu substitution in the factor VIII A3 domain. Br J Haematol 1999; 105 (4) 1123-1126
• 28 Potgieter JJ, Damgaard M, Hillarp A. One-stage vs. chromogenic assays in haemophilia A. Eur J Haematol 2015; 94 (Suppl. 77) 38-44
• 29 Trossaert M, Lienhart A, Nougier C , et al. Diagnosis and management challenges in patients with mild haemophilia A and discrepant FVIII measurements. Haemophilia 2014; 20 (4) 550-558
• 30 Hakeos WH, Miao H, Sirachainan N , et al. Hemophilia A mutations within the factor VIII A2-A3 subunit interface destabilize factor VIIIa and cause one-stage/two-stage activity discrepancy. Thromb Haemost 2002; 88 (5) 781-787
• 31 ECAT Survey. A European external quality assessment programme for laboratories in the field of haemostasis performed. ECAT Foundation 2013; 1: 1-109
• 32 Hubbard AR, Dodt J, Lee T , et al; Factor VIII and Factor IX Subcommittee of The Scientific and Standardisation Committee of The International Society on Thrombosis and Haemostasis. Recommendations on the potency labelling of factor VIII and factor IX concentrates. J Thromb Haemost 2013; 11 (5) 988-989
• 33 Barrowcliffe TW. Standardization of FVIII & FIX assays. Haemophilia 2003; 9 (4) 397-402
• 34 Barrowcliffe TW. Standardization of assays of factor VIII and factor IX. Ric Clin Lab 1990; 20 (2) 155-165
• 35 Chandler WL, Ferrell C, Lee J, Tun T, Kha H. Comparison of three methods for measuring factor VIII levels in plasma. Am J Clin Pathol 2003; 120 (1) 34-39
• 36 Hayes TE, Brandt JT, Chandler WL , et al. External peer review quality assurance testing in von Willebrand disease: the recent experience of the United States College of American Pathologists proficiency testing program. Semin Thromb Hemost 2006; 32 (5) 499-504
• 37 One-stage Prothrombin Time and Activated Partial Thromboplastin Time; Approved Guideline-Second Edition . CLSI document H-47 A2 Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19057-1898, 2008
• 38 Plug I, Mauser-Bunschoten EP, Bröcker-Vriends AH , et al. Bleeding in carriers of hemophilia. Blood 2006; 108 (1) 52-56
• 39 Lawrie AS, Kitchen S, Purdy G, Mackie IJ, Preston FE, Machin SJ. Assessment of Actin FS and Actin FSL sensitivity to specific clotting factor deficiencies. Clin Lab Haematol 1998; 20 (3) 179-186
• 40 Bowyer A, Kitchen S, Makris M. The responsiveness of different APTT reagents to mild factor VIII, IX and XI deficiencies. Int J Lab Hematol 2011; 33 (2) 154-158
• 41 Ruiz JA, Limerick KK. Utility of the activated partial thromboplastin time: evaluation of contact activation and other clinical conditions. CHR 1994; 8 (2) 11-12
• 42 Griffin JH. Role of surface in surface-dependent activation of Hageman factor (blood coagulation factor XII). Proc Natl Acad Sci U S A 1978; 75 (4) 1998-2002
• 43 Shetty S, Ghosh K, Mohanty D. Comparison of four commercially available activated partial thromboplastin time reagents using a semi-automated coagulometer. Blood Coagul Fibrinolysis 2003; 14 (5) 493-497
• 44 Okuda M, Yamamoto Y. Usefulness of synthetic phospholipid in measurement of activated partial thromboplastin time: a new preparation procedure to reduce batch difference. Clin Lab Haematol 2004; 26 (3) 215-223
• 45 Kitchen S, Cartwright I, Woods TAL, Jennings I, Preston FE. Lipid composition of seven APTT reagents in relation to heparin sensitivity. Br J Haematol 1999; 106 (3) 801-808
• 46 Morfini M. Pharmacokinetics of factor VIII and factor IX. Haemophilia 2003; 9 (Suppl. 01) 94-99 , discussion 100
• 47 van den Besselaar AM, Neuteboom J, Meeuwisse-Braun J, Bertina RM. Preparation of lyophilized partial thromboplastin time reagent composed of synthetic phospholipids: usefulness for monitoring heparin therapy. Clin Chem 1997; 43 (7) 1215-1222
• 48 Hubbard AR. International biological standards for coagulation factors and inhibitors. Semin Thromb Hemost 2007; 33 (3) 283-289
• 49 Raut S, Daniels S, Heath AB ; SSC Sub-Committee on Factor. Value assignment of the WHO 8th International Standard for factor VIII, concentrate (07/350). J Thromb Haemost 2012; 10 (6) 1175-1176
• 50 Arkin CF, Bovill EG, Brandt JT, Rock WA, Triplett DA. Factors affecting the performance of factor VIII coagulant activity assays. Results of proficiency surveys of the College of American Pathologists. Arch Pathol Lab Med 1992; 116 (9) 908-915
• 51 Ofosu F, Cassidy K, Blajchman MA, Hirsh J. Immunodepletion of human plasma factor VIII. Blood 1980; 56 (4) 604-607
• 52 van Geffen M, Dardikh M, Vebruggen B. Factor VIIII inhibitor Assays:methodology, shortcomings, and challenges. Journal of Coagulation Disorders 2010; 2 (2) 1-7
• 53 Bai B, Christie DJ, Gorman RT, Wu JR. Comparison of optical and mechanical clot detection for routine coagulation testing in a large volume clinical laboratory. Blood Coagul Fibrinolysis 2008; 19 (6) 569-576
• 54 Meijer P, Verbruggen HW, Spannagl M. Clotting Factors and Inhibitors: Assays and Interpretation. , in Kottke-Marchant K, Davis BH. Laboratory Hematology Practice, Wiley Blackwell; 2012: 435-446
• 55 Kitchen S, Preston FE. Assay of factor VIII and other clotting factors. In: Kitchen S, Olson JD, Preston FE, eds. Quality in Laboratory Hemostasis and Thrombosis. Chichester, United Kingdom: Wiley-Blackwell; 2013: 105-114
• 56 Jennings I, Kitchen DP, Woods TA, Kitchen S, Walker ID. Emerging technologies and quality assurance: the United Kingdom National External Quality Assessment Scheme perspective. Semin Thromb Hemost 2007; 33 (3) 243-249
• 57 Kaufman RJ, Powell JS. Molecular approaches for improved clotting factors for hemophilia. Blood 2013; 122 (22) 3568-3574
• 58 Viuff D, Barrowcliffe T, Saugstrup T, Ezban M, Lillicrap D. International comparative field study of N8 evaluating factor VIII assay performance. Haemophilia 2011; 17 (4) 695-702
• 59 Sommer JM, Moore N, McGuffie-Valentine B , et al. Comparative field study evaluating the activity of recombinant factor VIII Fc fusion protein in plasma samples at clinical haemostasis laboratories. Haemophilia 2014; 20 (2) 294-300
• 60 Robinson M, Tiefenbacher S, Ross EL , et al. Comparison of FVIII activity of select novel recombinant FVIII replacement products in commonly used FDA-approved one-stage clot assay systems. In: Proceedings of the Congress of the International Society of Thrombosis and Haemostasis; June 20–25, 2015; Toronto, Canada. J Thromb Haem 2015; 13 (S2): 566
• 61 Dodt J, Hubbard AR, Wicks SJ , et al. Potency determination of factor VIII and factor IX for new product labelling and postinfusion testing: challenges for caregivers and regulators. Haemophilia 2015; 21 (4) 543-549
• 62 Franchini M, Mannucci PM. Past, present and future of hemophilia: a narrative review. Orphanet J Rare Dis 2012; 7: 24-31
• 63 Collins PW, Chalmers E, Hart DP , et al; UK Haemophilia Centre Doctors. Diagnosis and treatment of factor VIII and IX inhibitors in congenital hemophilia: (4th edition). UK Haemophilia Centre Doctors Organization. Br J Haematol 2013; 160 (Suppl. 02) 153-170
• 64 van den Berg HM, Chalmers EA. Clinical prediction models for inhibitor development in severe hemophilia A. J Thromb Haemost 2009; 7 (Suppl. 01) 98-102
• 65 Witmer C, Young G. Factor VIII inhibitors in hemophilia A: rationale and latest evidence. Ther Adv Hematol 2013; 4 (1) 59-72
• 66 Blanchette VS, Key NS, Ljung LR, Manco-Johnson MJ, van den Berg HM, Srivastava A ; Subcommittee on Factor VIII, Factor IX and Rare Coagulation Disorders of the Scientific and Standardization Committee of the International Society on Thrombosis and Hemostasis. Definitions in hemophilia: communication from the SSC of the ISTH. J Thromb Haemost 2014; 12 (11) 1935-1939
• 67 Kasper CK, Aledort LM, Counts BB , et al. Letter: A more uniform measurement of factor VIII inhibitors. Thromb Diath Haemorrh 1975; 34 (3) 869-872
• 68 Verbruggen B, Novakova I, Wessels H, Boezeman J, van den Berg M, Mauser-Bunschoten E. The Nijmegen modification of the Bethesda assay for factor VIII:C inhibitors: improved specificity and reliability. Thromb Haemost 1995; 73 (2) 247-251
• 69 Dardikh M, Meijer P, van der Meer F, Favaloro EJ, Verbruggen B. Acquired functional coagulation inhibitors: review on epidemiology, results of a wet-workshop on laboratory detection, and implications for quality of inhibitor diagnosis. Semin Thromb Hemost 2012; 38 (6) 613-621
• 70 Favaloro EJ, Bonar R, Duncan E , et al. Mis-identification of factor inhibitors by diagnostic haemostasis laboratories: recognition of pitfalls and elucidation of strategies. A follow up to a large multicentre evaluation. Pathology 2007; 39 (5) 504-511
• 71 Favaloro EJ, Bonar R, Duncan E , et al; RCPA QAP in Haematology Haemostasis Committee. Identification of factor inhibitors by diagnostic haemostasis laboratories: a large multi-centre evaluation. Thromb Haemost 2006; 96 (1) 73-78
• 72 Kitchen S, Jennings I, Preston FE, Kitchen DP, Woods TAL, Walker ID. Interlaboratory variation in factor VIII:C inhibitor assay results is sufficient to influence patient management: data from the UK national quality external assessment scheme for blood coagulation. Semin Thromb Hemost 2009; 35 (8) 778-785
• 73 Bonar RA, Favaloro EJ, Marsden K. External quality assessment of factor VIII inhibitor assays. Semin Thromb Hemost 2013; 39 (3) 320-326
• 74 Miller CH, Platt SJ, Rice AS, Kelly F, Soucie JM ; Hemophilia Inhibitor Research Study Investigators. Validation of Nijmegen-Bethesda assay modifications to allow inhibitor measurement during replacement therapy and facilitate inhibitor surveillance. J Thromb Haemost 2012; 10 (6) 1055-1061
• 75 Giles AR, Verbruggen B, Rivard GE, Teitel J, Walker I ; Association of Hemophilia Centre Directors of Canada. Factor VIII/IX Subcommittee of Scientific and Standardization Committee of International Society on Thrombosis and Haemostasis. A detailed comparison of the performance of the standard versus the Nijmegen modification of the Bethesda assay in detecting factor VIII:C inhibitors in the haemophilia A population of Canada. Thromb Haemost 1998; 79 (4) 872-875
• 76 Shetty S, Ghosh K, Mohanty D. An ELISA assay for the detection of factor VIII antibodies - comparison with the conventional Bethesda assay in a large cohort of haemophilia samples. Acta Haematol 2003; 109 (1) 18-22
• 77 Towfighi F, Gharagozlou S, Sharifian RA , et al. Comparative measurement of anti-factor VIII antibody by Bethesda assay and ELISA reveals restricted isotype profile and epitope specificity. Acta Haematol 2005; 114 (2) 84-90
• 78 Lindgren A, Wadenvik H, Tengborn L. Characterization of inhibitors to FVIII with an ELISA in congenital and acquired haemophilia A. Haemophilia 2002; 8 (5) 644-648
• 79 Adcock DM, Favaloro EJ. Pearls and pitfalls in factor inhibitor assays. Int J Lab Hematol 2015; 37 (Suppl. 01) 52-60
• 80 Mackie I, Cooper P, Laurie A , et al; British Committee for Standards in Haematology. Guidelines in the laboratory aspects of assays used in hemostasis and thrombosis. Int J Lab Hematol 2013; 35 (Suppl. 01) 1-13
• 81 College of American Pathologists. Hematology and Coagulation Checklist, CAP Accreditation Process. Chicago, IL: College of American Pathologists; 2014: 1-46