Overview
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Background
Vascular Endothelial Growth Factor (VEGF)-A is a key regulator of angiogenesis and vascular permeability. It activates VEGFR1-3 on endothelial cells, thereby promoting tumor growth, cell survival, and increased vessel permeability to meet the tumor’s metabolic demands. VEGF-A is also elevated in conditions like diabetic retinopathy and macular edema. Anti-VEGF drugs such as bevacizumab and aflibercept bind VEGF-A, blocking receptor interaction and preventing VEGFR-2 dimerization and downstream signaling. This inhibits endothelial cell proliferation and vascular leakage, suppressing both angiogenesis and tumor growth.
PathHunter KDR/KDR Cell Line models this molecular MOA in a simple, homogenous format, suitable for screening, target validation, characterization, rank ordering, and potency applications. The assay is also available in a ready-to-use bioassay format for lot release programs.
Product Highlights
- Cell-based functional assay for anti-VEGF drugs, antibodies or biosimilar based on their mechanism of action
- Simple, homogenous assay protocol with results in less than 24 hours
- Significantly better assay performance compared to difficult HUVEC assay
- Suitable for rank ordering, comparability studies, potency measurement, stability studies and neutralizing antibody detection
- Applicable across various stages from screening to characterization and QC lot-release
Assay Principle
The PathHunter KDR/KDR assay utilizes EFC-based platform and is designed to detect VEGF-A-induced functional homodimerization of the kinase insert domain (KDR), also known as the VEGFR2 receptor. Activation of the KDR receptor through VEGF-A leads to receptor dimerization, which is an essential event in the receptor's signaling cascade. A. The PathHunter KDR/KDR cells have been engineered to co-express KDR fused to PK, and another KDR construct fused to EA. B. Upon activation by VEGF-A, the VEGFR2 receptors naturally dimerize forcing the two fragments to complement, creating an active β-gal enzyme, which hydrolyzes its substrate and produces a chemiluminescent signal, indicating receptor activation C. Anti-VEGF antibodies like bevacizumab or aflibercept, inhibit VEGF-A’s ability to activate the receptors resulting in a dose-dependent reduction of the chemiluminescent signal.
