Abstract
Following repeated administration of factor VIII (FVIII), a significant number of hemophilia A patients develop antibodies (Abs), inhibiting the procoagulant activity of infused FVIII. We have designed an approach based on the blocking of the deleterious activity of these Abs by peptide decoys mimicking the anti-FVIII Ab epitopes. Here, the well characterized inhibitory monoclonal Ab ESH8 served as a model. Several phage peptide libraries were screened for specific binding to ESH8. Seven constrained dodecapeptide sequences were obtained. Six sequences carried the consensus motif, hydrophobic-(Y/F)GKTXL. This motif showed a certain similarity with the (2231)QVDFQKTMKV(2240) sequence of the C(2) domain. In the seventh sequence, YCNPSIGDKNCR, the residues GDKN are similar to the sequence (2267)DGHQ(2270). Upon inspection of the C(2) domain crystallographic structure, the two stretches QVDFQKTMKV and DGHQ appeared close together in space and might constitute a discontinuous epitope. Corresponding synthetic peptides were able to inhibit the binding of ESH8 to FVIII in a specific and dose-dependent manner. Moreover, the ability of the selected peptides to neutralize the inhibitory activity of ESH8 was demonstrated in functional tests as well as in vivo in a murine model of hemophilia A. This study demonstrates the potential of this approach to neutralize the activity of potent inhibitory Abs.
Highlights
Coagulation defects attributed to the absence or dysfunction of blood coagulation factor VIII (FVIII)1 are observed in 0.01– 0.02% of the male population [1]
Its epitope is mapped to residues 2248 –2285 on the C2 domain of FVIII, a domain frequently recognized by human anti-FVIII Abs [31]
The peptides binding to mAb ESH8 were selected by the phage display technology, a method allowing the identification of peptides with significant mimicry potential for a broad spectrum of applications [14]
Summary
Coagulation defects attributed to the absence or dysfunction of blood coagulation factor VIII (FVIII) are observed in 0.01– 0.02% of the male population [1]. A number of different therapeutic approaches have been developed to circumvent complications arising from inhibitors, such as the administration of porcine FVIII, which is less antigenic than its human counterpart [6], the administration of activated human FVII to bypass the intrinsic pathway [7], and the activation of the coagulation cascade by using prothrombin complex concentrates [8] These treatments have greatly improved the medical management of the disease, the ultimate goal for therapy, the specific neutralization of the immune response to FVIII, has not yet been achieved. We present here evidence both in vitro and in vivo that mimotopes of the C2 domain can efficiently neutralize the inhibitory activity of mAb ESH8 and as such could represent an efficient method for the treatment of hemophilia A patients with inhibitory antibodies
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