Blockade of CD40/CD40 ligand interactions prevents induction of factor VIII inhibitors in hemophilic mice but does not induce lasting immune tolerance.
Patients with severe hemophilia A frequently develop neutralizing anti-factor VIII antibodies after replacement therapy with factor VIII (FVIII). In a search for new strategies to induce immune tolerance against FVIII in these patients, we used a murine model of hemophilia A to investigate the importance of CD40/CD40 ligand (CD40L) interactions for the initiation of the anti-FVIII immune response. We focused our attention in particular on the induction of neutralizing anti-FVIII antibodies and the Th1/Th2 polarization of FVIII-specific T cells. The development of anti-FVIII antibodies was analyzed by ELISA systems (detection of total anti-FVIII antibodies) and Bethesda assays (determination of neutralizing anti-FVIII antibodies). Factor VIII-specific T cells were characterized by multiparameter flow cytometry and cytokine ELISAs for the detection of cytokine production in splenic CD4+ T cells after in vitro restimulation with FVIII. Hemophilic mice received four doses of FVIII and anti-CD40L antibody MR1 (24 h before FVIII). Subsequently mice received four doses of FVIII only. The induction of neutralizing anti-FVIII antibodies in hemophilic mice after treatment with human FVIII could be prevented completely by a blockade of CD40/CD40L interactions using MR1. Furthermore, FVIII-specific T-cell responses that included both Th1 and Th2 cells were suppressed when mice were treated with FVIII and MR1. The initial blockade of CD40/CD40L interactions was, however, not sufficient to induce a lasting immune tolerance against FVIII. The immune suppression was abolished and both neutralizing anti-FVIII antibodies and FVIII-specific T cells developed when treatment with FVIII was continued after the omission of MR1. In addition, there were no alterations in the Th1/Th2 polarization induced by the initial blockade of CD40/CD40L interactions.
Reipert
B M
BM
Baxter BioScience, Vienna, Austria.
Sasgary
M
M
Ahmad
R U
RU
Auer
W
W
Turecek
P L
PL
Schwarz
H P
HP
D000818:Animals; D000910:Antibodies, Heterophile / Q000096:biosynthesis / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D001402:B-Lymphocytes / Q000276:immunology; D019013:CD40 Antigens / Q000276:immunology*; D023201:CD40 Ligand / Q000276:immunology*; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005169:Factor VIII / Q000008:administration & dosage / Q000276:immunology*; D006467:Hemophilia A / Q000276:immunology*; D006801:Humans; D007108:Immune Tolerance; D007115:Immunization Schedule; D007117:Immunization, Secondary; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000276:immunology; D007165:Immunosuppression Therapy; D008211:Lymphocyte Cooperation; D051379:Mice; D018345:Mice, Knockout; D009500:Neutralization Tests; D011993:Recombinant Fusion Proteins / Q000276:immunology; D013154:Spleen / Q000276:immunology; D018417:Th1 Cells / Q000276:immunology* / Q000378:metabolism; D018418:Th2 Cells / Q000276:immunology / Q000378:metabolism
D000910:Antibodies, Heterophile; D000911:Antibodies, Monoclonal; D019013:CD40 Antigens; D016207:Cytokines; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D023201:CD40 Ligand; D005169:Factor VIII