Intracellular Delivery of Diphtheria Toxin Elicits Anti-angiogenic Response in Cells Overexpressing Vascular Endothelial Growth Factor Receptor 2
Intracellular Delivery of Diphtheria Toxin Elicits Anti-angiogenic Response in Cells Overexpressing Vascular Endothelial Growth Factor Receptor 2
Fatemeh Kazemi-Lomedasht,1,*Mahdi Behdani,2
1. Venom and Biotherapeutics Molecules Laboratory, Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran 2. Venom and Biotherapeutics Molecules Laboratory, Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
Introduction: The vascular endothelial growth factor receptors, namely VEGFR1 and VEGFR2, constitute tyrosine kinase receptors prominently expressed on both endothelial cells and tumor vessels, significantly influencing the intricate process of angiogenesis. In this study, a trimeric arrangement of the VEGFR1 and VEGFR2 binding peptide (VGB3) was strategically incorporated through genetic fusion into the truncated diphtheria toxin (TDT). Subsequently, an exhaustive examination of its in vitro activity was conducted to elucidate its functional implications.
Methods: The recombinant construct, denoted as TDT-triVGB3, was synthesized, cloned and expressed within bacterial cells and subsequently purified employing Nickel affinity chromatography. The binding characteristics, as well as the affinity of TDT-triVGB3, were assessed utilizing the enzyme-linked immunosorbent assay (ELISA). To discern its inhibitory effects, the impact of TDT-triVGB3 on the viability, migration, and tube formation of human endothelial cells was evaluated through the employment of MTT assays, migration assays, and tube formation assays, respectively.
Results: In the enzyme-linked immunosorbent assay, TDT-triVGB3 exhibited selective and high-affinity binding to VEGFR1 and VEGFR2. Furthermore, its inhibitory effects on the viability, migration, and tube formation of human endothelial cells were statistically significant.
Conclusion: The synthesized TDT-triVGB3 emerges as a promising novel agent, demonstrating potential efficacy in selectively targeting cancer cells overexpressing VEGFR1/VEGFR2.