Abstract
Overexpression of human epidermal growth factor receptor 3 (HER3) is involved in resistance to several therapies for malignant tumours. Currently, several anti-HER3 monoclonal antibodies are under clinical development. We introduce an alternative approach to HER3-targeted therapy based on engineered scaffold proteins, i.e. affibody molecules. We designed a small construct (22.5 kDa, denoted 3A3), consisting of two high-affinity anti-HER3 affibody molecules flanking an albumin-binding domain ABD, which was introduced for prolonged residence in circulation. In vitro, 3A3 efficiently inhibited growth of HER3-expressing BxPC-3 cells. Biodistribution in mice was measured using 3A3 that was site-specifically labelled with 111In via a DOTA chelator. The residence time of 111In-DOTA-3A3 in blood was extended when compared with the monomeric affibody molecule. 111In-DOTA-3A3 accumulated specifically in HER3-expressing BxPC-3 xenografts in mice. However, 111In-DOTA-3A3 cleared more rapidly from blood than a size-matched control construct 111In-DOTA-TAT, most likely due to sequestering of 3A3 by mErbB3, the murine counterpart of HER3. Repeated dosing and increase of injected protein dose decreased uptake of 111In-DOTA-3A3 in mErbB3-expressing tissues. Encouragingly, growth of BxPC-3 xenografts in mice was delayed in an experimental (pilot-scale) therapy study using 3A3. We conclude that the 3A3 affibody format seems promising for treatment of HER3-overexpressing tumours.
Highlights
Overexpression of human epidermal growth factor receptor 3 (HER3) is involved in resistance to several therapies for malignant tumours
The high-affinity HER3-binding affibody molecule is of interest since it was shown to efficiently inhibit ligand-induced proliferation of HER3-positive breast cancer cells in vitro[31], and the bivalent format was chosen since it was the most potent in blocking ligand-induced phosphorylation of HER receptors in a comparison of different bispecific and bivalent constructs[30]
Monoclonal antibodies are used for cancer therapy in clinics to interrupt ligand binding and, in this way, inhibit downstream proliferation-promoting signalling of such receptors as EGFR41 and VEGFR42
Summary
Overexpression of human epidermal growth factor receptor 3 (HER3) is involved in resistance to several therapies for malignant tumours. We introduce an alternative approach to HER3-targeted therapy based on engineered scaffold proteins, i.e. affibody molecules. Different so called non-immunoglobulin scaffold proteins have been developed as tumour targeting agents, mostly for medical imaging applications, and for therapy purposes[12,13]. Affibody molecules[15] constitute a type of alternative scaffold proteins that has demonstrated excellent properties for tumour targeting purposes[16]. They have a molecular weight around 6.5 kDa, forming a three-helix bundle fold with very fast folding kinetics and capacity to re-fold after denaturing[17]. GMP-production of affibody-ABD fusion proteins is possible by prokaryotic cells, which makes it less expensive compared with the GMP-production of monoclonal antibodies[19]
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