Kinetic analysis of novel mono- and multivalent VHH-fragments and their application for molecular imaging of brain tumors

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DOIResolve DOI: http://doi.org/10.1111/j.1476-5381.2010.00742.x
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TypeArticle
Journal titleBritish Journal of Pharmacology
Volume160
Issue4
Pages10161028; # of pages: 13
Subjectmolecular imaging; multivalency; epidermal growth factor receptor; single domain antibody; brain cancer; surface plasmon resonance
AbstractBackground and purpose:? The overexpression of epidermal growth factor receptor (EGFR) and its mutated variant EGFRvIII occurs in 50% of glioblastoma multiforme. We developed antibody fragments against EGFR/EGFRvIII for molecular imaging and/or therapeutic targeting applications. Experimental approach:? An anti–EGFR/EGFRvIII llama single-domain antibody (EG2) and two higher valency format constructs, bivalent EG2-hFc and pentavalent V2C-EG2 sdAbs, were analysed in vitro for their binding affinities using surface plasmon resonance and cell binding studies, and in vivo using pharmacokinetic, biodistribution, optical imaging and fluorescent microscopy studies. Key results:? Kinetic binding analyses by surface plasmon resonance revealed intrinsic affinities of 55 nM and 97 nM for the monovalent EG2 to immobilized extracellular domains of EGFR and EGFRvIII, respectively, and a 10- to 600-fold increases in apparent affinities for the multivalent binders, V2C-EG2 and EG2-hFc, respectively. In vivo pharmacokinetic and biodistribution studies in mice revealed plasma half-lives for EG2, V2C-EG2 and EG2-hFc of 41 min, 80 min and 12.5 h, respectively, as well as a significantly higher retention of EG2-hFc compared to the other two constructs in EGFR/EGFRvIII-expressing orthotopic brain tumours, resulting in the highest signal in the tumour region in optical imaging studies. Time domain volumetric optical imaging fusion with high-resolution micro-computed tomography of microvascular brain network confirmed EG2-hFc selective accumulation/retention in anatomically defined tumour regions. Conclusions:? Single domain antibodies can be optimized for molecular imaging applications by methods that improve their apparent affinity and prolong plasma half-life and, at the same time, preserve their ability to penetrate tumour parenchyma.
Publication date
LanguageEnglish
AffiliationNRC Genomics and Health Initiative; NRC Biotechnology Research Institute; NRC Institute for Biological Sciences; NRC Institute for Biodiagnostics; National Research Council Canada
Peer reviewedYes
NPARC number17673441
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Record identifier0f07dc17-4db4-4365-a582-b3d95afd1c68
Record created2011-04-03
Record modified2016-05-09
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