Antigenic Peptides Capable of Inducing Specific Antibodies for Detection of the Major Alterations Found in Type 2B Von Willebrand Disease

Marina De Oliveira Paro,Marcos Aurélio De Santana,Maria Sueli Silva Namen-Lopes,Milton Hércules Guerra De Andrade,Fernanda Silva Vieira,William Castro-Borges,Cibele Velloso-Rodrigues,Cyntia Silva Ferreira,Sophie Yvette Leclercq

doi:10.1155/2013/590329

Abstract

Von Willebrand disease (VWD) is an inherited hemorrhagic disorder promoted by either quantitative or qualitative defects of the von Willebrand factor (VWF). The disease represents the most common human coagulopathy afflicting 1.3% of the population. Qualitative defects are subdivided into four subtypes and classified according to the molecular dysfunction of the VWF. The differential diagnosis of the VWD is a difficult task, relying on a panel of tests aimed to assess the plasma levels and function of the VWF. Here, we propose biochemical approaches for the identification of structural variants of the VWF. A bioinformatic analysis was conducted to design seven peptides among which three were representatives of specific amino acid sequences belonging to normal VWF and four encompassed sequences found in the most common VWD subtype 2B. These peptides were used to immunize mice, after which, peptide-specific immunoglobulins were purified. This resulted in four Ig preparations capable of detecting alterations in the subtype 2B VWD plus additional three antibody fractions targeting the normal VWF. The panel of antibodies could serve many applications among them (1) assessment of VWF: antigen interaction, (2) VWF multimer analysis, and (3) production of monoclonal antibodies against VWF for therapeutic purposes as in thrombotic thrombocytopenic purpura.

Highlights

Von Willebrand disease (VWD) is an inherited hemorrhagic disturbance related to quantitative and/or qualitative defects of the von Willebrand factor (VWF) [1, 2]
Alterations on the plasma levels of VWF are associated with VWD types 1 and 3, whereas structural and functional defects of VWF result in VWD type 2 [3, 6,7,8,9,10,11]
Whilst some patients with type 2B VWD can be treated with desmopressin, patients who do not show a satisfactory response to this drug should receive VWF/factor VIII (FVIII)-containing products [13, 14]

Summary

Introduction

Von Willebrand disease (VWD) is an inherited hemorrhagic disturbance related to quantitative and/or qualitative defects of the von Willebrand factor (VWF) [1, 2]. Variants of the VWF in the subtype 2N display reduced affinity by coagulation factor VIII (FVIII) These six VWD types correlate with diverse clinical outcomes each requiring adequate therapeutic interventions [4, 12]. The possibility of a precise and direct diagnosis of qualitative defects in the VWF would certainly permit application of a better oriented medical approach to afflicted patients In this context, it is worth emphasizing that antibodies capable of detecting structural variants of the VWF are not commercially available. It has previously been shown that amino acid substitutions R1306W, R1308C, V1316M, and R1341Q account for 90% prevalence of the subtype 2B VWD [6, 8] These mutations in the A1 domain of VWF are responsible for a “gain-offunction” defect allowing for an increased affinity of large multimers to platelets in the circulation [15].

Materials and Methods

Results and Discussion

Conclusions