Immunodominant T-cell epitopes in the factor VIII C2 domain are located within an inhibitory antibody binding site

Summary

Formation of inhibitor antibodies to factor VIII (FVIII) is a major complication of FVIII replacement therapy for hemophilia A patients, and it occurs through a T-cell dependent process. The C2 domain of FVIII contains epitopes that are recognized by antibody inhibitors. We have examined regions of the C2 domain that form epitopes for T cells in mice congenitally deficient in FVIII. We obtained CD4⁺ T cells from mice immunized by intravenous infusion of therapeutic doses of recombinant human FVIII (rFVIII), or by subcutaneous injections of rFVIII or recombinant human C2 domain in adjuvant. In all cases, the T cells recognized most strongly and consistently two overlapping peptides that spanned residues 2191 to 2220 of the C2 domain. Analysis of the crystal structure of human factor VIII C2 bound to a human monoclonal antibody, BO2C11, showed these residues also constitute part of a human alloimmune B-cell epitope (Spiegel et al., Blood 2001; 98: 13-19).This region includes one of the “hydrophobic spike” protrusions, consisting of M2199 and F2200, as well as the basic residues R2215 and R2220. These residues contribute to membrane binding and to association with von Willebrand factor (vWF).These findings suggest that a major T-cell epitope in the C2 domain recognized by hemophilic mice is located within the same region that binds to inhibitors, vWF, and activated membranes.

Keywords

T-cell epitopes - factor VIII - C2 domain

^* The first three authors shared equally in this work.

^¶ Current address: Jiahua Qian, Navy Bldg 8 Room 5101, NCI, NIH, Bethesda, MD 20982, USA

• References

• 1 van Dieijen G. et al. The role of phospholipid and factor VIIIa in the activation of bovine factor X. J Biol Chem 1981; 256: 3433-42.
• 2 Pemberton S. et al. A molecular model for the triplicated A domains of human factor VIII based on the crystal structure of human ceruloplasmin. Blood 1997; 89: 2413-21.
• 3 Liu ML. et al. A domain mutations in 65 haemophilia A families and molecular modeling of dysfunctional factor VIII proteins. Br J Haematol 1998; 103: 1051-60.
• 4 Fuentes-Prior P. et al. New insights into binding interfaces of coagulation factors V and VIII and their homologues. Lessons from high resolution crystal structures. Curr Protein Pept Sci 2002; 03: 313-39.
• 5 Pratt KP. et al. Structure of the C2 domain of human factor VIII at 1.5 Å resolution. Nature 1999; 402: 439-42.
• 6 Gilbert GE. et al. Four hydrophobic amino acids of the factor VIII C2 domain are constituents of both the membrane-binding and von Willebrand factor-binding motifs. J Biol Chem 2002; 277: 6374-81.
• 7 Liu ML. et al. Hemophilic factor VIII C1-and C2-domain missense mutations and their modeling to the 1.5-angstrom human C2-domain crystal structure. Blood 2000; 96: 979-87.
• 8 Thompson AR. Structure and function of the factor VIII gene and protein. Semin Thromb Hemost 2003; 29: 11-22.
• 9 Antonarakis SE. et al. Factor VIII gene inversions in severe hemophilia A: results of an international consortium study. Blood 1995; 86: 2206-12.
• 10 Schwaab R. et al. Haemophilia A: mutation type determines risk of inhibitor formation. Thromb Haemost 1995; 74: 1402-6.
• 11 Scandella D. et al. Epitope specificity and functional characterization of factor VIII inhibitors. Adv Exp Med Biol 1995; 386: 47-63.
• 12 Goodeve AC. et al. Relationship between factor VIII mutation type and inhibitor development in a cohort of previously untreated patients treated with recombinant factor VIII (Recombinate). Recombinate PUP Study Group. Thromb Haemost 2000; 83: 844-8.
• 13 Goodeve AC, Peake IR. The molecular basis of hemophilia a: genotype-phenotype relationships and inhibitor development. Semin Thromb Hemost 2003; 29: 23-30.
• 14 Tuddenham EG, Schwaab R, Seehafer J. et al. Haemophilia A: database of nucleotide substitutions, deletions, insertions and rearrangements of the factor VIII gene, second edition. Nucleic Acids Res 1994; 22: 3511-33.
• 15 Scandella D. et al. Epitope mapping of human factor VIII inhibitor antibodies by deletion analysis of factor VIII fragments expressed in Escherichia coli. Proc Natl Acad Sci USA 1988; 85: 6152-6.
• 16 Scandella D. et al. Localization of epitopes for human factor VIII inhibitor antibodies by immunoblotting and antibody neutralization. Blood 1989; 74: 1618-26.
• 17 Scandella D. et al. Some factor VIII inhibitor antibodies recognize a common epitope corresponding to C2 domain amino acids 2248 through 2312, which overlap a phospholipidbinding site. Blood 1995; 86: 1811-9.
• 18 Scandella DH. et al. In hemophilia A and autoantibody inhibitor patients: the factor VIII A2 domain and light chain are most immunogenic. Thromb Res 2001; 101: 377-85.
• 19 Saenko EL. et al. Haemophilia A: effects of inhibitory antibodies on factor VIII functional interactions and approaches to prevent their action. Haemophilia 2002; 08: 1-11.
• 20 Shima M. et al. A factor VIII neutralizing monoclonal antibody and a human inhibitor alloantibody recognizing epitopes in the C2 domain inhibit factor VIII binding to von Willebrand factor and to phosphatidylserine. Thromb Haemost 1993; 69: 240-6.
• 21 Shima M. et al. Common inhibitory effects of human anti-C2 domain inhibitor alloantibodies on factor VIII binding to von Willebrand factor. Br J Haematol 1995; 91: 714-21.
• 22 Jacquemin MG, Desqueper BG, Benhida A. et al. Mechanism and kinetics of factor VIII inactivation: study with an IgG4 monoclonal antibody derived from a hemophilia A patient with inhibitor. Blood 1998; 92: 496-506.
• 23 Newton-Nash DK, Tollerud D, Guevarra L. et al. Interferon-gamma secretion defects in haemophilia A patients receiving highly purified plasma-derived or recombinant factor VIII. Br J Haematol 1996; 95: 554-60.
• 24 Singer ST. et al. T lymphocyte proliferative responses induced by recombinant factor VIII in hemophilia A patients with inhibitors. Thromb Haemost 1996; 76: 17-22.
• 25 Reding MT, Wu H, Krampf M. et al. CD4+ Tcell response to factor VIII in hemophilia A, acquired hemophilia, and healthy subjects. Thromb Haemost 1999; 82: 509-15.
• 26 Reding MT, Wu H, Krampf M. et al. Sensitization of CD4+ T cells to coagulation factor VIII: Response in congenital and acquired hemophilia patients and in healthy subjects. Thromb Haemost 2000; 84: 643-52.
• 27 Reding MT, Lei S, Lei H. et al. Distribution of Th1-and Th2-induced anti-factor VIII IgG subclasses in congenital and acquired hemophilia patients. Thromb Haemost 2002; 88: 568-75.
• 28 Reding MT, Okita DK, Diethelm-Okita BM. et al. Human CD4+ T-cell epitope repertoire on the C2 domain of coagulation factor VIII. J Thromb Haemost 2003; 01: 1777-84.
• 29 Hu GL, Okita DK, Diethelm-Okita B. et al. Recognition of coagulation factor VIII by CD4+ T cells of healthy humans. J Thromb Haemost 2003; 01: 2159-66.
• 30 Wu H, Reding M, Qian J. et al. Mechanism of the immune response to human factor VIII in murine hemophilia A. Thromb Haemost 2001; 85: 125-33.
• 31 Spiegel Jr PC, Jacquemin M, Saint-Remy JM. et al. Structure of a factor VIII C2 domainimmunoglobulin G4kappa Fab complex: identification of an inhibitory antibody epitope on the surface of factor VIII. Blood 2001; 98: 13-19.
• 32 Takeshima K, Smith C, Tait J. et al. The preparation and phospholipid binding property of the C2 domain of human factor VIII. Thromb Haemost 2003; 89: 788-94.
• 33 Stern LJ, Brown JH, Jardetzky TS. et al. Crystal structure of the human class II MHC protein HLA-DR1 complexed with an influenza virus peptide. Nature 1994; 368: 215-21.
• 34 Moiola L, Karachunski P, Protti M. et al. Epitopes on the ß subunit of human muscle acetylcholine receptor recognized by CD4+ cells of myasthenia gravis patients and healthy subjects. J Clin Invest 1994; 93: 1020-8.
• 35 Manfredi AA, Protti MP, Dalton MWM. et al. Myasthenia Gravis: T helper cell recognition of muscle acetylcholine receptor in myasthenia gravis. Epitopes on the gamma and delta subunits. J Clin Invest 1993; 92: 1055-67.
• 36 Raju R, Navaneetham D, Okita DK. et al. Epitopes for human CD4⁺ T cells on diphtheria toxin: structural features of sequence segments forming epitopes recognized by most subjects. Eur J Immunol 1995; 25: 3207-14.
• 37 Diethelm-Okita B. et al. Universal epitopes for human CD4⁺ cells on tetanus and diphtheria toxins. J Infect Dis 2000; 181: 1001-9.
• 38 Wang ZY, Okita DK, Howard JF. et al. T cell recognition of muscle acetylcholine receptor subunits in generalized and ocular myasthenia gravis. Neurology 1998; 50: 1045-54.
• 39 Healey JF. et al. Residues Glu2181-Val2243 contain a major determinant of the inhibitory epitope in the C2 domain of human factor VIII. Blood 1998; 92: 3701-9.
• 40 Barrow RT, Healey JF, Gailani D. et al. Reduction of the antigenicity of factor VIII toward complex inhibitory antibody plasmas using multiply-substituted hybrid human/porcine factor VIII molecules. Blood 2000; 95: 564-8.
• 41 Barrow RT, Healey JF, Jacquemin MG. et al. Antigenicity of putative phospholipid membrane-binding residues in factor VIII. Blood 2001; 97: 169-74.
• 42 Rawle FEM, Labelle AD, Postma L. et al. Oral administration of factor VIII contributes to the prevention of anti-factor VIII antibody development after protein infusion, and prolongs factor VIII transgene expression post-adenoviral gene transfer. Blood. 2003 102. abstract #566.
• 43 Brown BD, Shi CX, Rawle FE. et al. Factors influencing therapeutic efficacy and the host immune response to helper-dependent adenoviral gene therapy in hemophilia A mice. J Thromb Haemost 2004; 02: 111-8.