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Title: Rational humanization of the powerful antithrombotic anti-GPIb alpha antibody: 6B4
Authors: Fontayne, Alexandre ×
Vanhoorelbeke, Karen
Pareyn, Inge
Van Rompaey, Isabel
Meiring, Muriel
Lamprecht, Seb
Roodt, Jan
Desmet, Johan
Deckmyn, Hans #
Issue Date: Nov-2006
Publisher: Schattauer gmbh-verlag medizin naturwissenschaften
Series Title: Thrombosis and haemostasis vol:96 issue:5 pages:671-684
Abstract: Fab-fragments of the monoclonal antibody 6134, raised against human glycoprotein lb alpha (GPIb alpha), have a powerful antithrombotic effect in baboons by blocking the GPIb alpha binding site for von Willebrand factor (VWF), without significant prolongation of the skin bleeding time. In order to bring this antibody to the clinic,we here humanized for the first time an anti-human GPIb alpha by variable-domain resurfacing guided by computer modeling. First, the genes coding for the variable regions of the heavy and light chains of 6134 were cloned and sequenced. Based on this, a three-dimensional structure of the Fv-fragment was constructed by using homology-based modeling, and with this and comparison with antibodies with known structure,"murine" putative immunogenic residues which are exposed, were changed for "human-like" residues. The humanized Fab-fragment, h6B4-Fab, was constructed in the pKaneo vector system, expressed and purified and showed in vitro an unaltered, even slightly higher binding affinity for its antigen than the murine form as determined by different ELISA set-ups and surface plasmon resonance. Finally, injection of doses of 0.1 to 1.5 mg/kg of h6B4-Fab in baboons showed that both pharmacokinetics and ex-vivo bio-activity of the molecule were to a large extent preserved. In conclusion,the method used hereto humanize 6134 by resurfacing resulted in a fully active derivative, which is now ready for further development.
URI: 
ISSN: 0340-6245
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Interdisciplinary Research Facility Life Sciences, Campus Kulak Kortrijk
Chemistry, Campus Kulak Kortrijk
× corresponding author
# (joint) last author

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