Multimerization of peptide mimotopes for blocking of factor VIII neutralizing antibodies.
About 30 % of patients with severe hemophilia A develop neutralizing antibodies (inhibitors) to coagulation factor VIII (FVIII) upon treatment with exogenous factor preparations. Two peptides, C6 (NPVENMMDRDSQ) and H10 (QSPWQTWFTRAL), that mimic putative inhibitor epitopes (mimotopes), were previously selected by phage display screening of plasma samples from patients with inhibitors. Synthetic peptide mimotopes inhibited IgG binding to FVIII (IC(50): 30-50 microM). This effect was increased by an equimolar combination of both mimotopes. Mimotopes were fused to the C-terminal multimerization domain of the C4bp alpha-chain and expressed as multimers in 293T cells. Multimerized mimotopes showed improved binding to anti-FVIII IgG and prolonged in vitro half-life relative to synthetic peptides. The two mimotopes were combined in heteromultimers by co-transfection of 293T cells with respective vectors, resulting in bi-specific molecules that almost completely blocked polyclonal antibody binding to FVIII (IC(50): 2-3 microM). This strategy is capable of functionally improving synthetic peptides by multimerization and could provide a basis for novel therapeutic approaches for patients with hemophilia A and inhibitors.
Kessel
Christoph
C
Department of Pediatrics III, JW Goethe University, 60596 Frankfurt am Main, Germany.
Kreuz
Wolfhart
W
Klich
Katharina
K
Becker-Peters
Karin
K
Vorpahl
Frank
F
Dietrich
Ursula
U
Klingebiel
Thomas
T
Königs
Christoph
C
D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology / Q000378:metabolism*; D002460:Cell Line; D000939:Epitopes / Q000737:chemistry; D005169:Factor VIII / Q000276:immunology* / Q000378:metabolism; D006207:Half-Life; D006467:Hemophilia A / Q000188:drug therapy; D006801:Humans; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology
D000906:Antibodies; D000939:Epitopes; D019151:Peptide Library; D010455:Peptides; D011994:Recombinant Proteins; D005169:Factor VIII