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Targeting angiogenesis with small antibody fragments and antibody-like molecules

Avramović, Dragana. Targeting angiogenesis with small antibody fragments and antibody-like molecules. 2016, Doctoral Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_12236

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Abstract

Angiogenesis is the formation of new blood vessels from pre-existing capillaries. It occurs throughout the life of higher organisms, in both health and disease. Meta- bolically active tissues in the body are found in the proximity of the blood capillaries, formed by the process of angiogenesis. The idea that control of angiogenesis could have a therapeutic potential has raised great interest during the past 40 years. Stim- ulation of angiogenesis can be therapeutic for ischemic heart disease, peripheral arterial disease and wound healing. Decreasing or inhibiting angiogenesis can be a therapeutic goal in cancer, some ophthalmic pathological conditions, rheumatoid arthritis, and other diseases.
Vascular Endothelial Growth Factors (VEGFs) and their receptors (VEGFRs) are major players in both physiological and pathological angiogenesis. VEGFs constitute a fam- ily of proteins that play important roles in blood and lymphatic vessel development. VEGFs bind VEGFR-1, -2, and -3, promoting cell survival, proliferation, di erentiation, and migration. VEGFR-2 is the major mediator of angiogenic signaling in endothelial cells, and its activity is regulated at multiple levels. Ligand binding to the extracellular domain (ECD) of VEGFR-2 leads to receptor dimerization, followed by intracellular kinase domain activation, receptor internalization and downstream signaling.
The goal of this study was to develop novel antiangiogenic drugs targeting VEGFR-2, as an addition or alternative to existing cancer therapies. As for targeting tools, we used di erent antibody formats and antibody-like molecules. In this study, we gen- erated and tested three types of ECD binders, single chain fragment variable anti- bodies (scFvs), fragment antigen-binding antibodies (Fabs) and full-length Immuno- globulins G (IgGs), speci cally interacting with single Ig-homology domains, located in the receptor ECD. We identi ed several promising antibodies, interacting with the ECD of VEGFR-2 and blocking ligand-stimulated receptor activation. Di erent formats of antibodies showed similar e ects with signi cant inhibition of VEGFR-2 phosphorylation at Tyr1175 and phosphorylation of PLC-γ, as well as inhibition of in vitro angiogenesis in Human Umbilical Vein Endothelial cell (HUVEC) tube formation and HUVEC migration assays. Our ndings provide the proof-of-principle that highly speci c anti-VEGFR-2 agents targeting the membrane-proximal Ig-domains D4 and D7 inhibit receptor activity.
Interestingly, the binding of antibody fragments to VEGFR-2 led to receptor internal- ization as demonstrated by an increase in total volume of intracellular receptor-pos- itive vesicles. Internalization independent of ligand binding represents a new, prom- ising property of these VEGFR-2 antibodies. The new agents will be useful for in vivo studies aimed at vessel imaging or at inhibiting VEGFR-2 signaling.
Additionally, we tested the already characterized Designed Ankyrin Repeat Proteins (DARPins) for tumor targeting in vivo.
Advisors:Ballmer-Hofer, Kurt and Christofori, Gerhard M.
Faculties and Departments:03 Faculty of Medicine > Departement Biomedizin > Division of Biochemistry and Genetics > Tumor Biology (Christofori)
UniBasel Contributors:Ballmer-Hofer, Kurt and Christofori, Gerhard M.
Item Type:Thesis
Thesis Subtype:Doctoral Thesis
Thesis no:12236
Thesis status:Complete
Bibsysno:Link to catalogue
Number of Pages:1 Online-Ressource (127 Seiten)
Language:English
Identification Number:
Last Modified:05 Apr 2018 17:36
Deposited On:22 Aug 2017 14:26

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