PhD Scientific Days 2025

Poster Session I. - I: Theoretical and Translational Medicine

Modified Tyrosine Kinase Inhibitors Become Targetable and Exert Anti-Angiogenic Effects by Inhibiting the VEGF-A Induced VEGFR2 Signaling In Vitro

Name of the presenter

Antal Réka

Institute/workplace of the presenter

Translational Retina Research Group, Institute of Translational Medicine, Semmelweis University

Authors

Réka Antal¹, Balázs Besztercei¹, Attila Szappanos¹, Krisztián András Kovács¹

1: Translational Retina Research Group, Institute of Translational Medicine, Semmelweis University

Text of the abstract

Introduction: Ocular neovascularization-based diseases such as neovascular age-related macular degeneration (nAMD) and proliferative diabetic retinopathy (PDR) are among the leading causes of blindness. PDR and nAMD can be caused by abnormal blood vessel growth due to elevated levels of vascular endothelial growth factor (VEGF) in the eye.
Aims: Here, we investigate the inhibitory potential of novel, chemically modified tyrosine kinase inhibitors (TKIs) and hypothesize that a light-based approach enables targeting the molecules into specific tissues by permanently binding them to vascular endothelial growth factor receptor 2 (VEGFR2) and helps increase their inhibitory effect on neovascularization in vitro.
Methods: In this study, TKIs EYE1118 and EYE1090 were tested in the presence and absence of light via assays. We performed ADP-Glo Kinase assay followed by Western Blot on recombinant VEGFR2 kinase. To determine the efficacy of the inhibitors in vitro, VEGFR2/NFAT reporter HEK293 cells were treated with the inhibitors (0.1 pM – 10 μM), and luciferase reporter assay was performed. Human retinal microvascular endothelial cells (HRMECs) were pre-incubated with compounds (0.1 nM - 10 μM) to test their inhibitory effect on in vitro angiogenesis.
Results: The kinase activity decreased significantly (p<0.0001) in the presence of the inhibitors in the dark, and the presence of light increased the inhibitory effect of EYE1118 (p<0.01) and EYE1090 (p<0.001) even more. Both TKIs strongly decreased the phosphorylation levels of Tyr951, Tyr1175, and Tyr1214 on recombinant VEGFR2 both with and without illumination. In HEK293 cells, VEGFR2 signaling was strongly inhibited by EYE1118 and EYE1090, and the inhibition increased significantly (p<0.0001) after illumination. Both molecules strongly inhibited the in vitro angiogenic potential of HRMECs in the presence of light, and this inhibition was significantly more potent than in the dark.
Conclusion: Our results demonstrate that the new TKIs exhibit potent inhibitory effects in vitro and in biochemical tests. It was confirmed that photoactivation enables them to be targeted and bound permanently to VEGFR2; thus, the inhibitory potential of these molecules can be increased when exposed to light.
Funding: Funded by the European Commission via an EIC Pathfinder grant (Acronym: NeoVasculoStop, Grant No: 101047120).