Human CD4+ T-cell epitope repertoire on the C2 domain of coagulation factor VIII

Abstract

Summary. Approximately 25% of severe hemophilia A patients develop antibodies (Ab) that neutralize the procoagulant function of factor (F)VIII (inhibitors). Autoimmune FVIII inhibitors may develop in individuals without congenital FVIII deficiency and cause acquired hemophilia. Low titers of anti‐FVIII Ab may be present in hemophilia A patients without inhibitors and in healthy blood donors. FVIII‐specific CD4+ T‐cells drive the synthesis of anti‐FVIII Ab. We examined the epitope repertoire of CD4+ T‐cells from 15 healthy subjects, 10 hemophilia A patients without inhibitors, 11 hemophilia A patients with inhibitors, and six acquired hemophilia patients. Blood CD4+ T‐cells were challenged in proliferation assays with a panel 16 overlapping synthetic peptides, spanning the sequence of the FVIII C2 domain. The sequence region 2291–2330 contained the most frequently and strongly recognized peptides in each of the four subject groups. Crystallographic B factor data and the location of these peptides within the three‐dimensional structure of the C2 domain confirm that this region has a high degree of solvent exposure and flexibility within the peptide backbone, which are structural features typical of immunodominant universal CD4+ epitopes. Furthermore, this sequence region overlaps inhibitor‐binding sites, suggesting that CD4+ T‐cells recognizing peptide sequences within this region might be involved in inhibitor synthesis. The sequence regions 2191–2210 (recognized strongly by each study group except hemophilia A patients with inhibitors) and 2241–2290 (recognized primarily by acquired hemophilia patients and healthy subjects) share the same structural features, and also overlap inhibitor‐binding sites. Although similar, there appear to be important differences in the CD4+ epitope repertoires of congenital and acquired hemophilia patients.

Phosphatidylinositol Containing Lipidic Particles Reduces Immunogenicity and Catabolism of Factor VIII in Hemophilia A Mice

“…The A2, A3, and C2 domains of FVIII participate in vWF (9-13), LRP (14,15), and phospholipid binding (16)(17)(18)(19)(20) that contributes to catabolism of FVIII, and these domains also contain epitopes that can lead to inhibitor development (24)(25)(26). Therefore, it will be of interest to investigate the influence of macromolecular interactions on catabolism and immunogenicity.…”

“…However, we speculate that the PI reduces FVIII immunogenicity by multiple mechanisms. Anti-FVIII antibodies targeted against epitopes in A2, A3, and C2 domains of FVIII (24)(25)(26). As the lipid binding involve these domains, these immunogenic epitopes were shielded resulting in reduction of antibody titer levels by immune ignorance.…”

“…Unfortunately, 15-35% of patients develop FVIII neutralizing antibodies that abrogate the activity of the protein, thus complicating FVIII replacement therapy and clinical manage-ment of the disease (22,23). Based on systematic epitope mapping experiments, the anti-FVIII antibodies have been found to mainly target defined regions in the A2 (heavy chain) and A3 and C2 domains (light chain) of FVIII (24)(25)(26). Thus, macromolecular interactions of FVIII involve A2, A3, and C2 domains, and these domains also contain epitope regions that can lead to inhibitor development in FVIII replacement therapy.…”

Lower Inhibitor Development in Hemophilia A Mice following Administration of Recombinant Factor VIII-O-Phospho-L-serine Complex

“…The immunoprecipitation and inhibitor neutralization assays of the inhibitor plasma from hemophilia A patients clearly indicated that the anti-light chain antibody titer was the highest (12). More recently, Reding et al (13) and Pratt et al (14) have identified several universal epitopes for CD4ϩ T-cells in the 2291-2330 region of the C2 domain using proliferation assays with CD4ϩ cells from normal humans, hemophilia A patients (13), and mice (14). Three-dimensional models proposed based on crystallographic studies and mutational analysis show that the C2 domain also contains 2-4 hydrophobic loops and other charged residues that promote lipid binding ( Fig.…”

Desenvolvimento de inibidores do fator VIII na hemofilia A

“…9 Os inibidores do FVIII são imunoglobulinas policlonais principalmente da subclasse IgG4, cuja síntese é direcionada por células T CD4+ específicas para o FVIII. 10 Entretanto, subclasses IgG1 e IgG2 estão também usualmente presentes nas populações de anticorpos anti-FVIII. Estes dados sugerem que os inibidores do FVIII estão associados com uma combinação de respostas Th1/Th2.…”