Download PDF
Thromb Haemost 2015; 114(06): 1127-1135
DOI: 10.1160/TH14-06-0535
Coagulation and Fibrinolysis
Schattauer GmbH

Deletion or inhibition of Fc gamma receptor 2B (CD32) prevents FVIII-specific activation of memory B cells in vitro

Sonja Werwitzke
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Nadine Vollack
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Marcus von Hornung
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Katy Kalippke
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Julia Kutzschbach
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Arne Trummer
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Arnold Ganser
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
,
Andreas Tiede
1   Department of Haematology, Haemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
› Author Affiliations Financial support: This work was supported by the Hans Egli Grant of the German, Austrian and Swiss Thrombosis and Haemostasis Society (GTH), and a non-restricted grant from Biotest AG. Recombinant human factor VIII (Kogenate®) was provided by Bayer HealthCare.
Further Information

Publication History

Received: 23 June 2014

Accepted after major revision: 19 June 2015

Publication Date:
30 November 2017 (online)

Also available at
    Permissions and Reprints

Summary

Development of inhibitory antibodies against factor VIII (FVIII) is a severe complication of replacement therapy in haemophilia A. Patients with inhibitors are treated with high FVIII doses in the context of immune tolerance therapy (ITT). Data from haemophilia A mouse model suggest that high FVIII concentrations prevent the formation of antibody secreting cells (ASCs) from memory B cells (MBCs) by inducing apoptosis. Fc gamma receptor 2B (CD32) is an important regulator of B cell function, mediating inhibitory signals after cross-linking with the B cell receptor. Here, the role of CD32 in the regulation of FVIII-specific MBCs was investigated using F8-/- and F8-/-CD32-/- knockout mice and monoclonal antibodies (mAbs). The initial immune response was similar between F8-/- and F8-/-CD32-/- mice, including concentration of anti-FVIII antibodies and number of FVIII-specific ASCs in spleen and bone marrow. In contrast, formation of ASCs from MBCs upon rhFVIII re-stimulation in vitro was abolished in F8-/-CD32-/- mice, whereas FVIII/anti-FVIII immune complexes significantly enhanced ASC formation in F8-/- mice. Inhibition of CD32 by mAbs or F(ab)2 fragments prevented ASC formation in a dose-dependent manner. Transfer of B cell-depleted splenocytes using CD45R (B220) depletion from CD32-competent mice did not restore ASC formation in F8-/-CD32-/- cells confirming that CD32 is required on B cells. We conclude that CD32 is a crucial regulator of FVIII-specific B cells and is required for the differentiation of MBCs into ASCs. Inhibition of CD32 could potentially improve the efficacy of FVIII in the context of ITT.

Keywords

Haemophilia A mouse model - CD32 - inhibitor - memory B cells - recall response

 

  • References

  • 1 Witmer C, Young G. Factor VIII inhibitors in hemophilia A: rationale and latest evidence. Ther Adv Hematol 2013; 04: 59-72.
    CrossrefPubMedSearch in Google Scholar
  • 2 Pratt KP. Inhibitory antibodies in hemophilia A. Curr Opin Hematol 2012; 19: 399-405.
    CrossrefPubMedSearch in Google Scholar
  • 3 Hay CR, DiMichele DM. International Immune Tolerance S. The principal results of the International Immune Tolerance Study: a randomized dose comparison. Blood 2012; 119: 1335-1344.
    CrossrefPubMedSearch in Google Scholar
  • 4 DiMichele DM. Immune tolerance in haemophilia: the long journey to the fork in the road. Br J Haematol 2012; 159: 123-134.
    CrossrefPubMedSearch in Google Scholar
  • 5 Kempton CL, White 2nd GC. How we treat a hemophilia A patient with a factor VIII inhibitor. Blood 2009; 113: 11-17.
    CrossrefPubMedSearch in Google Scholar
  • 6 Hausl C, Ahmad RU, Schwarz HP. et al. 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.
    CrossrefPubMedSearch in Google Scholar
  • 7 Hausl C, Ahmad RU, Sasgary M. et al. High-dose factor VIII inhibits factor VIII-specific memory B cells in hemophilia A with factor VIII inhibitors. Blood 2005; 106: 3415-3422.
    CrossrefPubMedSearch in Google Scholar
  • 8 Reipert BM, Allacher P, Hausl C. et al. Modulation of factor VIII-specific memory B cells. Haemophilia 2010; 16: 25-34.
    PubMedSearch in Google Scholar
  • 9 Berard M, Casamayor-Palleja M, Billian G. et al. Activation sensitizes human memory B cells to B-cell receptor-induced apoptosis. Immunology 1999; 98: 47-54.
    CrossrefPubMedSearch in Google Scholar
  • 10 Gessner JE, Heiken H, Tamm A. et al. The IgG Fc receptor family. Ann Hematol 1998; 76: 231-248.
    CrossrefPubMedSearch in Google Scholar
  • 11 Qin D, Wu J, Vora KA. et al. Fc gamma receptor IIB on follicular dendritic cells regulates the B cell recall response. J Immunol 2000; 164: 6268-6275.
    CrossrefPubMedSearch in Google Scholar
  • 12 Nimmerjahn F, Ravetch JV. Fcgamma receptors as regulators of immune responses. Nat Rev Immunol 2008; 08: 34-47.
    CrossrefPubMedSearch in Google Scholar
  • 13 Takai T, Ono M, Hikida M. et al. Augmented humoral and anaphylactic responses in Fc gamma RII-deficient mice. Nature 1996; 379: 346-349.
    CrossrefPubMedSearch in Google Scholar
  • 14 Bolland S, Ravetch JV. Spontaneous autoimmune disease in Fc(gamma)RIIB-deficient mice results from strain-specific epistasis. Immunity 2000; 13: 277-285.
    CrossrefPubMedSearch in Google Scholar
  • 15 Clynes R, Maizes JS, Guinamard R. et al. Modulation of immune complex-induced inflammation in vivo by the coordinate expression of activation and inhibitory Fc receptors. J Exp Med 1999; 189: 179-185.
    CrossrefPubMedSearch in Google Scholar
  • 16 Li J, Tuzun E, Wu XR. et al. Inhibitory IgG receptor FcgammaRIIB fails to inhibit experimental autoimmune myasthenia gravis pathogenesis. J Neuroimmunol 2008; 194: 44-53.
    CrossrefPubMedSearch in Google Scholar
  • 17 Bi L, Lawler AM, Antonarakis SE. et al. Targeted disruption of the mouse factor VIII gene produces a model of haemophilia A. Nat Genet 1995; 10: 119-121.
    CrossrefPubMedSearch in Google Scholar
  • 18 Bi L, Sarkar R, Naas T. et al. Further characterisation of factor VIII-deficient mice created by gene targeting: RNA and protein studies. Blood 1996; 88: 3446-3450.
    PubMedSearch in Google Scholar
  • 19 Qian J, Borovok M, Bi L. et al. Inhibitor antibody development and T cell response to human factor VIII in murine hemophilia A. Thromb Haemost 1999; 81: 240-244.
    Thieme ConnectPubMedSearch in Google Scholar
  • 20 Reipert BM, Ahmad RU, Turecek PL. et al. Characterisation of antibodies induced by human factor VIII in a murine knockout model of hemophilia A. Thromb Haemost 2000; 84: 826-832.
    Thieme ConnectPubMedSearch in Google Scholar
  • 21 Hausl C, Maier E, Schwarz HP. et al. 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.
    Thieme ConnectPubMedSearch in Google Scholar
  • 22 Sasgary M, Ahmad RU, Schwarz HP. et al. Single cell analysis of factor VIII-specific T cells in hemophilic mice after treatment with human factor VIII. Thromb Haemost 2002; 87: 266-272.
    Thieme ConnectPubMedSearch in Google Scholar
  • 23 Kalippke K, Werwitzke S, von Hornung M. et al. DNA analysis from stool samples: a fast and reliable method avoiding invasive sampling methods in mouse models of bleeding disorders. Lab Anim 2009; 43: 390-393.
    CrossrefPubMedSearch in Google Scholar
  • 24 Sondermann P, Huber R, Oosthuizen V. et al. The 3.2-A crystal structure of the human IgG1 Fc fragment-Fc gammaRIII complex. Nature 2000; 406: 267-273.
    CrossrefPubMedSearch in Google Scholar
  • 25 Werwitzke S, Trick D, Sondermann P. et al. Treatment of lupus-prone NZB/ NZW F1 mice with recombinant soluble Fc gamma receptor II (CD32). Ann Rheum Dis 2008; 67: 154-161.
    CrossrefPubMedSearch in Google Scholar
  • 26 Tomayko MM, Steinel NC, Anderson SM. et al. Cutting edge: Hierarchy of maturity of murine memory B cell subsets. J Immunol 2010; 185: 7146-7150.
    CrossrefPubMedSearch in Google Scholar
  • 27 Zuccarino-Catania GV, Sadanand S, Weisel FJ. et al. CD80 and PD-L2 define functionally distinct memory B cell subsets that are independent of antibody isotype. Nat Immunol 2014; 15: 631-637.
    PubMedSearch in Google Scholar
  • 28 Hausl C vHP, Ahmad RU, Muchitsch EM. et al. The Relevance of the Inhibitory Receptor FcgIIB for the Regulation of Anti-Factor VIII Antibody Response in Murine Models. Blood (ASH Annual Meeting Abstracts). November 2007 110. Abstract 785.
    PubMedSearch in Google Scholar
  • 29 Nakamura A, Yuasa T, Ujike A. et al. Fcgamma receptor IIB-deficient mice develop Goodpasture’s syndrome upon immunisation with type IV collagen: a novel murine model for autoimmune glomerular basement membrane disease. J Exp Med 2000; 191: 899-906.
    CrossrefPubMedSearch in Google Scholar
  • 30 Boross P, Arandhara VL, Martin-Ramirez J. et al. The inhibiting Fc receptor for IgG, FcgammaRIIB, is a modifier of autoimmune susceptibility. J Immunol 2011; 187: 1304-1313.
    CrossrefPubMedSearch in Google Scholar
  • 31 Williams EL, Tutt AL, French RR. et al. Development and characterisation of monoclonal antibodies specific for the murine inhibitory FcgammaRIIB (CD32B). Eur J Immunol 2012; 42: 2109-2120.
    CrossrefPubMedSearch in Google Scholar
  • 32 Zhang AH, Skupsky J, Scott DW. Effect of B-cell depletion using anti-CD20 therapy on inhibitory antibody formation to human FVIII in hemophilia A mice. Blood 2011; 117: 2223-2226.
    CrossrefPubMedSearch in Google Scholar