Main Article Content

Davide Matino
Paul Tieu, Dr.
Antony Chan


Keywords: inhibitors, hemophilia, anti-FVIII antibodies, anti-FIX antibodies


The development of neutralizing antibodies in hemophilia is a serious complication of factor replacement therapy. These antibodies, also known as “inhibitors”, significantly increase morbidity within the hemophilia population and lower the quality of life for these patients. People with severe hemophilia A have an overall 25-40% lifetime risk of inhibitor development, compared to that of 5-15% lifetime risk in those with moderate/mild hemophilia A. The risk is lower in hemophilia B population (about 1-5%) and occurrence of inhibitors is almost only seen in patients with severe hemophilia B. The understanding of the pathophysiological mechanism leading to the development of inhibitors in patients with hemophilia has improved considerably over the last 2 decades. Identification of early biomarkers which predict inhibitor development in previously untreated patients with hemophilia will assist in risk identification and possible early intervention strategies. In this review, we aim to summarize the molecular mechanisms of inhibitor development in hemophilia and to identify potential areas in need of further investigation.


Download data is not yet available.
Abstract 1420 | PDF Downloads 775 HTML Downloads 137


[1] Soucie JM, Evatt B, Jackson D. Occurrence of hemophilia in the United States. The Hemophilia Surveillance System Project Investigators. Am J Hematol 1998; 59: 288-294.
[2] Blanchette VS, Key NS, Ljung LR, Manco-Johnson MJ, van den Berg HM, Srivastava A. Definitions in hemophilia: communication from the SSC of the ISTH. J Thromb Haemost 2014; 12: 1935-1939.
[3] Antonarakis SE, Rossiter JP, Young M, Horst J, de MP, Sommer SS, Ketterling RP, Kazazian HH, Jr., Negrier C, Vinciguerra C, Gitschier J, Goossens M, Girodon E, Ghanem N, Plassa F, Lavergne JM, Vidaud M, Costa JM, Laurian Y, Lin SW, Lin SR, Shen MC, Lillicrap D, Taylor SA, Windsor S, Valleix SV, Nafa K, Sultan Y, Delpech M, Vnencak-Jones CL, Phillips JA, III, Ljung RC, Koumbarelis E, Gialeraki A, Mandalaki T, Jenkins PV, Collins PW, Pasi KJ, Goodeve A, Peake I, Preston FE, Schwartz M, Scheibel E, Ingerslev J, Cooper DN, Millar DS, Kakkar VV, Giannelli F, Naylor JA, Tizzano EF, Baiget M, Domenech M, Altisent C, Tusell J, Beneyto M, Lorenzo JI, Gaucher C, Mazurier C, Peerlinck K, Matthijs G, Cassiman JJ, Vermylen J, Mori PG, Acquila M, Caprino D, Inaba H. Factor VIII gene inversions in severe hemophilia A: results of an international consortium study. Blood 1995; 86: 2206-2212.
[4] Schwaab R, Brackmann HH, Meyer C, Seehafer J, Kirchgesser M, Haack A, Olek K, Tuddenham EG, Oldenburg J. Haemophilia A: mutation type determines risk of inhibitor formation. Thromb Haemost 1995; 74: 1402-1406.
[5] Hay CR. Factor VIII inhibitors in mild and moderate-severity haemophilia A. Haemophilia 1998; 4: 558-563.
[6] Gomez K, Klamroth R, Mahlangu J, Mancuso ME, Mingot ME, Ozelo MC. Key issues in inhibitor management in patients with haemophilia. Blood Transfus 2014; 12 Suppl 1: s319-s329.
[7] Astermark J, Oldenburg J, Carlson J, Pavlova A, Kavakli K, Berntorp E, Lefvert AK. Polymorphisms in the TNFA gene and the risk of inhibitor development in patients with hemophilia A. Blood 2006; 108: 3739-3745.
[8] Astermark J, Oldenburg J, Pavlova A, Berntorp E, Lefvert AK. Polymorphisms in the IL10 but not in the IL1beta and IL4 genes are associated with inhibitor development in patients with hemophilia A. Blood 2006; 107: 3167-3172.
[9] Astermark J, Wang X, Oldenburg J, Berntorp E, Lefvert AK. Polymorphisms in the CTLA-4 gene and inhibitor development in patients with severe hemophilia A. J Thromb Haemost 2007; 5: 263-265.
[10] Hollestelle MJ, Thinnes T, Crain K, Stiko A, Kruijt JK, van Berkel TJ, Loskutoff DJ, van Mourik JA. Tissue distribution of factor VIII gene expression in vivo--a closer look. Thromb Haemost 2001; 86: 855-861.
[11] Madoiwa S, Yamauchi T, Kobayashi E, Hakamata Y, Dokai M, Makino N, Kashiwakura Y, Ishiwata A, Ohmori T, Mimuro J, Sakata Y. Induction of factor VIII-specific unresponsiveness by intrathymic factor VIII injection in murine hemophilia A. J Thromb Haemost 2009; 7: 811-824.
[12] Vremec D, Pooley J, Hochrein H, Wu L, Shortman K. CD4 and CD8 expression by dendritic cell subtypes in mouse thymus and spleen. J Immunol 2000; 164: 2978-2986.
[13] Mebius RE, Kraal G. Structure and function of the spleen. Nat Rev Immunol 2005; 5: 606-616.
[14] Navarrete A, Dasgupta S, Delignat S, Caligiuri G, Christophe OD, Bayry J, Nicoletti A, Kaveri SV, Lacroix-Desmazes S. Splenic marginal zone antigen-presenting cells are critical for the primary allo-immune response to therapeutic factor VIII in hemophilia A. J Thromb Haemost 2009; 7: 1816-1823.
[15] Qian J, Burkly LC, Smith EP, Ferrant JL, Hoyer LW, Scott DW, Haudenschild CC. Role of CD154 in the secondary immune response: the reduction of pre-existing splenic germinal centers and anti-factor VIII inhibitor titer. Eur J Immunol 2000; 30: 2548-2554.
[16] Feuerer M, Beckhove P, Garbi N, Mahnke Y, Limmer A, Hommel M, Hammerling GJ, Kyewski B, Hamann A, Umansky V, Schirrmacher V. Bone marrow as a priming site for T-cell responses to blood-borne antigen. Nat Med 2003; 9: 1151-1157.
[17] Stern LJ, Brown JH, Jardetzky TS, Gorga JC, Urban RG, Strominger JL, Wiley DC. Crystal structure of the human class II MHC protein HLA-DR1 complexed with an influenza virus peptide. Nature 1994; 368: 215-221.
[18] Celli S, Lemaitre F, Bousso P. Real-time manipulation of T cell-dendritic cell interactions in vivo reveals the importance of prolonged contacts for CD4+ T cell activation. Immunity 2007; 27: 625-634.
[19] Lipscomb MF, Masten BJ. Dendritic cells: immune regulators in health and disease. Physiol Rev 2002; 82: 97-130.
[20] Pfistershammer K, Stockl J, Siekmann J, Turecek PL, Schwarz HP, Reipert BM. Recombinant factor VIII and factor VIII-von Willebrand factor complex do not present danger signals for human dendritic cells. Thromb Haemost 2006; 96: 309-316.
[21] Peerlinck K, Arnout J, Gilles JG, Saint-Remy JM, Vermylen J. A higher than expected incidence of factor VIII inhibitors in multitransfused haemophilia A patients treated with an intermediate purity pasteurized factor VIII concentrate. Thromb Haemost 1993; 69: 115-118.
[22] Peerlinck K, Arnout J, Di GM, Gilles JG, Laub R, Jacquemin M, Saint-Remy JM, Vermylen J. Factor VIII inhibitors in previously treated haemophilia A patients with a double virus-inactivated plasma derived factor VIII concentrate. Thromb Haemost 1997; 77: 80-86.
[23] Qadura M, Waters B, Burnett E, Chegeni R, Bradshaw S, Hough C, Othman M, Lillicrap D. Recombinant and plasma-derived factor VIII products induce distinct splenic cytokine microenvironments in hemophilia A mice. Blood 2009; 114: 871-880.
[24] Bovenschen N, Mertens K, Hu L, Havekes LM, van Vlijmen BJ. LDL receptor cooperates with LDL receptor-related protein in regulating plasma levels of coagulation factor VIII in vivo. Blood 2005; 106: 906-912.
[25] Lenting PJ, Neels JG, van den Berg BM, Clijsters PP, Meijerman DW, Pannekoek H, van Mourik JA, Mertens K, van Zonneveld AJ. The light chain of factor VIII comprises a binding site for low density lipoprotein receptor-related protein. J Biol Chem 1999; 274: 23734-23739.
[26] Bovenschen N, Rijken DC, Havekes LM, van Vlijmen BJ, Mertens K. The B domain of coagulation factor VIII interacts with the asialoglycoprotein receptor. J Thromb Haemost 2005; 3: 1257-1265.
[27] Dasgupta S, Navarrete AM, Bayry J, Delignat S, Wootla B, Andre S, Christophe O, Nascimbeni M, Jacquemin M, Martinez-Pomares L, Geijtenbeek TB, Moris A, Saint-Remy JM, Kazatchkine MD, Kaveri SV, Lacroix-Desmazes S. A role for exposed mannosylations in presentation of human therapeutic self-proteins to CD4+ T lymphocytes. Proc Natl Acad Sci U S A 2007; 104: 8965-8970.
[28] Dasgupta S, Repesse Y, Bayry J, Navarrete AM, Wootla B, Delignat S, Irinopoulou T, Kamate C, Saint-Remy JM, Jacquemin M, Lenting PJ, Borel-Derlon A, Kaveri SV, Lacroix-Desmazes S. VWF protects FVIII from endocytosis by dendritic cells and subsequent presentation to immune effectors. Blood 2007; 109: 610-612.
[29] Goudemand J, Rothschild C, Demiguel V, Vinciguerrat C, Lambert T, Chambost H, Borel-Derlon A, Claeyssens S, Laurian Y, Calvez T. Influence of the type of factor VIII concentrate on the incidence of factor VIII inhibitors in previously untreated patients with severe hemophilia A. Blood 2006; 107: 46-51.
[30] Gouw SC, van den Berg HM, le CS, van der Bom JG. Treatment characteristics and the risk of inhibitor development: a multicenter cohort study among previously untreated patients with severe hemophilia A. J Thromb Haemost 2007; 5: 1383-1390.
[31] Delignat S, Repesse Y, Navarrete AM, Meslier Y, Gupta N, Christophe OD, Kaveri SV, Lacroix-Desmazes S. Immunoprotective effect of von Willebrand factor towards therapeutic factor VIII in experimental haemophilia A. Haemophilia 2012; 18: 248-254.
[32] Herczenik E, van Haren SD, Wroblewska A, Kaijen P, van den Biggelaar M, Meijer AB, Martinez-Pomares L, ten BA, Voorberg J. Uptake of blood coagulation factor VIII by dendritic cells is mediated via its C1 domain. J Allergy Clin Immunol 2012; 129: 501-509.e5.
[33] Meems H, Meijer AB, Cullinan DB, Mertens K, Gilbert GE. Factor VIII C1 domain residues Lys 2092 and Phe 2093 contribute to membrane binding and cofactor activity. Blood 2009; 114: 3938-3946.
[34] Wroblewska A, van Haren SD, Herczenik E, Kaijen P, Ruminska A, Jin SY, Zheng XL, van den Biggelaar M, ten BA, Meijer AB, Voorberg J. Modification of an exposed loop in the C1 domain reduces immune responses to factor VIII in hemophilia A mice. Blood 2012; 119: 5294-5300.
[35] Bayry J, Lacroix-Desmazes S, Kazatchkine MD, Kaveri SV. Monoclonal antibody and intravenous immunoglobulin therapy for rheumatic diseases: rationale and mechanisms of action. Nat Clin Pract Rheumatol 2007; 3: 262-272.
[36] Bayry J, Siberil S, Triebel F, Tough DF, Kaveri SV. Rescuing CD4+CD25+ regulatory T-cell functions in rheumatoid arthritis by cytokine-targeted monoclonal antibody therapy. Drug Discov Today 2007; 12: 548-552.
[37] Marcucci M, Mancuso ME, Santagostino E, Kenet G, Elalfy M, Holzhauer S, Bidlingmaier C, Escuriola EC, Iorio A, Nowak-Gottl U. Type and intensity of FVIII exposure on inhibitor development in PUPs with haemophilia A. A patient-level meta-analysis. Thromb Haemost 2015; 113: 958-967.
[38] Davis SJ, Ikemizu S, Evans EJ, Fugger L, Bakker TR, van der Merwe PA. The nature of molecular recognition by T cells. Nat Immunol 2003; 4: 217-224.
[39] Waters B, Lillicrap D. The molecular mechanisms of immunomodulation and tolerance induction to factor VIII. J Thromb Haemost 2009; 7: 1446-1456.
[40] Qian J, Collins M, Sharpe AH, Hoyer LW. Prevention and treatment of factor VIII inhibitors in murine hemophilia A. Blood 2000; 95: 1324-1329.
[41] Reipert BM, Sasgary M, Ahmad RU, Auer W, Turecek PL, Schwarz HP. Blockade of CD40/CD40 ligand interactions prevents induction of factor VIII inhibitors in hemophilic mice but does not induce lasting immune tolerance. Thromb Haemost 2001; 86: 1345-1352.
[42] Rossi G, Sarkar J, Scandella D. Long-term induction of immune tolerance after blockade of CD40-CD40L interaction in a mouse model of hemophilia A. Blood 2001; 97: 2750-2757.
[43] Ewenstein BM, Hoots WK, Lusher JM, DiMichele D, White GC, Adelman B, Nadeau K. Inhibition of CD40 ligand (CD154) in the treatment of factor VIII inhibitors. Haematologica 2000; 85: 35-39.
[44] Kawai T, Andrews D, Colvin RB, Sachs DH, Cosimi AB. Thromboembolic complications after treatment with monoclonal antibody against CD40 ligand. Nat Med 2000; 6: 114.
[45] Koyama I, Kawai T, Andrews D, Boskovic S, Nadazdin O, Wee SL, Sogawa H, Wu DL, Smith RN, Colvin RB, Sachs DH, Cosimi AB. Thrombophilia associated with anti-CD154 monoclonal antibody treatment and its prophylaxis in nonhuman primates. Transplantation 2004; 77: 460-462.
[46] LeBien TW, Tedder TF. B lymphocytes: how they develop and function. Blood 2008; 112: 1570-1580.
[47] Slifka MK, Antia R, Whitmire JK, Ahmed R. Humoral immunity due to long-lived plasma cells. Immunity 1998; 8: 363-372.
[48] Manz RA, Hauser AE, Hiepe F, Radbruch A. Maintenance of serum antibody levels. Annu Rev Immunol 2005; 23: 367-386.
[49] Hausl C, Maier E, Schwarz HP, Ahmad RU, Turecek PL, Dorner F, Reipert BM. Long-term persistence of anti-factor VIII antibody-secreting cells in hemophilic mice after treatment with human factor VIII. Thromb Haemost 2002; 87: 840-845.
[50] Hausl C, Ahmad RU, Sasgary M, Doering CB, Lollar P, Richter G, Schwarz HP, Turecek PL, Reipert BM. High-dose factor VIII inhibits factor VIII-specific memory B cells in hemophilia A with factor VIII inhibitors. Blood 2005; 106: 3415-3422.
[51] Hausl C, Ahmad RU, Schwarz HP, Muchitsch EM, Turecek PL, Dorner F, Reipert BM. Preventing restimulation of memory B cells in hemophilia A: a potential new strategy for the treatment of antibody-dependent immune disorders. Blood 2004; 104: 115-122.
[52] Reipert BM, Gangadharan B, Hofbauer CJ, Scheiflinger F, Bowen J, Donnachie E, Fijvandraat K, Gruppo RA, Klintman J, Male C, McGuinn CE, Meeks SL, Recht M, Ragni MV, Yaish HM, Santagostino E, Brown DL. Appearance of high-affinity antibodies precedes clinical diagnosis of FVIII inhibitors - Preliminary analysis from the Hemophilia Inhibitor PUP Study (HIPS). Blood 128[22], 328. 2016. (Abstract)
[53] Matino D, Gargaro M, Santagostino E, Di Minno MN, Castaman G, Morfini M, Rocino A, Mancuso ME, Di MG, Coppola A, Talesa VN, Volpi C, Vacca C, Orabona C, Iannitti R, Mazzucconi MG, Santoro C, Tosti A, Chiappalupi S, Sorci G, Tagariello G, Belvini D, Radossi P, Landolfi R, Fuchs D, Boon L, Pirro M, Marchesini E, Grohmann U, Puccetti P, Iorio A, Fallarino F. IDO1 suppresses inhibitor development in hemophilia A treated with factor VIII. J Clin Invest 2015; 125: 3766-3781.
[54] Palmer DS, Dudani AK, Drouin J, Ganz PR. Identification of novel factor VIII inhibitor epitopes using synthetic peptide arrays. Vox Sang 1997; 72: 148-161.
[55] Huang CC, Shen MC, Chen JY, Hung MH, Hsu TC, Lin SW. Epitope mapping of factor VIII inhibitor antibodies of Chinese origin. Br J Haematol 2001; 113: 915-924.
[56] Lacroix-Desmazes S, Bayry J, Misra N, Horn MP, Villard S, Pashov A, Stieltjes N, d'Oiron R, Saint-Remy JM, Hoebeke J, Kazatchkine MD, Reinbolt J, Mohanty D, Kaveri SV. The prevalence of proteolytic antibodies against factor VIII in hemophilia A. N Engl J Med 2002; 346: 662-667.
[57] Scandella D, Gilbert GE, Shima M, Nakai H, Eagleson C, Felch M, Prescott R, Rajalakshmi KJ, Hoyer LW, Saenko E. Some factor VIII inhibitor antibodies recognize a common epitope corresponding to C2 domain amino acids 2248 through 2312, which overlap a phospholipid-binding site. Blood 1995; 86: 1811-1819.
[58] Saenko EL, Shima M, Rajalakshmi KJ, Scandella D. A role for the C2 domain of factor VIII in binding to von Willebrand factor. J Biol Chem 1994; 269: 11601-11605.
[59] Zhong D, Saenko EL, Shima M, Felch M, Scandella D. Some human inhibitor antibodies interfere with factor VIII binding to factor IX. Blood 1998; 92: 136-142.
[60] Lollar P, Parker ET, Curtis JE, Helgerson SL, Hoyer LW, Scott ME, Scandella D. Inhibition of human factor VIIIa by anti-A2 subunit antibodies. J Clin Invest 1994; 93: 2497-2504.
[61] Lacroix-Desmazes S, Moreau A, Sooryanarayana, Bonnemain C, Stieltjes N, Pashov A, Sultan Y, Hoebeke J, Kazatchkine MD, Kaveri SV. Catalytic activity of antibodies against factor VIII in patients with hemophilia A. Nat Med 1999; 5: 1044-1047.
[62] Lavigne-Lissalde G, Lacroix-Desmazes S, Wootla B, Tarrade C, Schved JF, Kaveri SV, Granier C, Villard-Saussine S. Molecular characterization of human B domain-specific anti-factor VIII monoclonal antibodies generated in transgenic mice. Thromb Haemost 2007; 98: 138-147.
[63] Klintman J, Hillarp A, Berntorp E, Astermark J. Long-term anti-FVIII antibody response in Bethesda-negative haemophilia A patients receiving continuous replacement therapy. Br J Haematol 2013; 163: 385-392.
[64] Butenas S, Krudysz-Amblo J, Rivard GE, Mann G. Product-dependent anti-factor VIII antibodies. Haemophilia 2013; 19: 619-625.
[65] Hofbauer CJ, Whelan SF, Hirschler M, Allacher P, Horling FM, Lawo JP, Oldenburg J, Tiede A, Male C, Windyga J, Greinacher A, Knobl PN, Schrenk G, Koehn J, Scheiflinger F, Reipert BM. Affinity of FVIII-specific antibodies reveals major differences between neutralizing and nonneutralizing antibodies in humans. Blood 2015; 125: 1180-1188.
[66] Prescott R, Nakai H, Saenko EL, Scharrer I, Nilsson IM, Humphries JE, Hurst D, Bray G, Scandella D. The inhibitor antibody response is more complex in hemophilia A patients than in most nonhemophiliacs with factor VIII autoantibodies. Recombinate and Kogenate Study Groups. Blood 1997; 89: 3663-3671.
[67] Whelan SF, Hofbauer CJ, Horling FM, Allacher P, Wolfsegger MJ, Oldenburg J, Male C, Windyga J, Tiede A, Schwarz HP, Scheiflinger F, Reipert BM. Distinct characteristics of antibody responses against factor VIII in healthy individuals and in different cohorts of hemophilia A patients. Blood 2013; 121: 1039-1048.
[68] White GC, Kempton CL, Grimsley A, Nielsen B, Roberts HR. Cellular immune responses in hemophilia: why do inhibitors develop in some, but not all hemophiliacs? J Thromb Haemost 2005; 3: 1676-1681.
[69] Miao CH, Ye P, Thompson AR, Rawlings DJ, Ochs HD. Immunomodulation of transgene responses following naked DNA transfer of human factor VIII into hemophilia A mice. Blood 2006; 108: 19-27.
[70] Peng B, Ye P, Blazar BR, Freeman GJ, Rawlings DJ, Ochs HD, Miao CH. Transient blockade of the inducible costimulator pathway generates long-term tolerance to factor VIII after nonviral gene transfer into hemophilia A mice. Blood 2008; 112: 1662-1672.
[71] Bray GL, Kroner BL, Arkin S, Aledort LW, Hilgartner MW, Eyster ME, Ragni MV, Goedert JJ. Loss of high-responder inhibitors in patients with severe hemophilia A and human immunodeficiency virus type 1 infection: a report from the Multi-Center Hemophilia Cohort Study. Am J Hematol 1993; 42: 375-379.
[72] Matzinger P. The danger model: a renewed sense of self. Science 2002; 296: 301-305.
[73] Pradeu T, Cooper EL. The danger theory: 20 years later. Front Immunol 2012; 3: 287.
[74] Pordes AG, Baumgartner CK, Allacher P, Ahmad RU, Weiller M, Schiviz AN, Schwarz HP, Reipert BM. T cell-independent restimulation of FVIII-specific murine memory B cells is facilitated by dendritic cells together with toll-like receptor 7 agonist. Blood 2011; 118: 3154-3162.
[75] Reding MT. Immunological aspects of inhibitor development. Haemophilia 2006; 12 Suppl 6: 30-35.
[76] Reding MT, Lei S, Lei H, Green D, Gill J, Conti-Fine BM. Distribution of Th1- and Th2-induced anti-factor VIII IgG subclasses in congenital and acquired hemophilia patients. Thromb Haemost 2002; 88: 568-575.
[77] Reding MT, Okita DK, Diethelm-Okita BM, Anderson TA, Conti-Fine BM. Human CD4+ T-cell epitope repertoire on the C2 domain of coagulation factor VIII. J Thromb Haemost 2003; 1: 1777-1784.
[78] Reding MT, Okita DK, Diethelm-Okita BM, Anderson TA, Conti-Fine BM. Epitope repertoire of human CD4(+) T cells on the A3 domain of coagulation factor VIII. J Thromb Haemost 2004; 2: 1385-1394.
[79] Reipert BM, van Helden PM, Schwarz HP, Hausl C. Mechanisms of action of immune tolerance induction against factor VIII in patients with congenital haemophilia A and factor VIII inhibitors. Br J Haematol 2007; 136: 12-25.
[80] Cao O, Loduca PA, Herzog RW. Role of regulatory T cells in tolerance to coagulation factors. J Thromb Haemost 2009; 7 Suppl 1: 88-91.
[81] Tang Q, Bluestone JA. The Foxp3+ regulatory T cell: a jack of all trades, master of regulation. Nat Immunol 2008; 9: 239-244.
[82] Key NS. Inhibitors in congenital coagulation disorders. Br J Haematol 2004; 127: 379-391.