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Published on: December 10, 2010
A vascular endothelial growth factor high affinity receptor 1-specific peptide with antiangiogenic activity
Mayada El-Mousawi1, Lioudmila Tchistiakova, Ludmila Yurchenko
1Supratek Pharma Inc., 215 Bouchard Street, Dorval, Quebec H9S 1A9, Canada.
Abstract:
Vascular endothelial growth factor (VEGF) is known to play a predominant role in tumor angiogenesis and metastasis formation that is mediated by its interactions with two tyrosine kinase receptors, VEGFRI (Flt-1) and VEGFRII (KDR). Inhibition of VEGF-dependent events in tumor tissues is known to enhance apoptosis and to suppress tumor growth. A novel peptide, SP5.2, which selectively binds Flt-1 and inhibits a broad range of VEGF-mediated events, was identified using a phage-display library screening. The fluorescein-labeled SP5.2 specifically bound to VEGF-stimulated primary human cerebral endothelial cells (HCECs), whereas non-stimulated HCECs, as well as human neuroblastoma cells (ShyY) did not show any interaction with the peptide. SP5.2 prevented proliferation of cultured primary human umbilical vein endothelial cells induced by recombinant human VEGF165 with an IC50 of 5 microm. SP5.2 was also shown to antagonize VEGF- and PLGF-induced, but not basic fibroblast growth factor-induced proliferation of HCECs. In contrast to "scrambled" peptide, SP5.2 was also found to selectively inhibit VEGF-stimulated migration of HCECs. The in vitro analysis of antiangiogenic activity of SP5.2 using a capillary-like tube formation assay showed that VEGF-induced angiogenesis of HCECs grown on Matrigel was completely inhibited in the presence of 10 microm SP5.2. Further studies demonstrated that SP5.2 prevented VEGF-induced permeability increase in HCECs monolayers. To explore whether SP5.2 can be used as a targeting agent, chemical and recombinant conjugates of SP5.2 with reporter proteins (peroxidase and beta-galactosidase) were produced. The resulting products showed significant increases (200-fold for SP5.2-beta-gal and 400-fold for SP5.2-peroxidase) in binding affinity to recombinant Flt-1 compared with the original synthetic SP5.2, suggesting that conjugate with therapeutic activity in nanomolar range could potentially be developed based on SP5.2 structure.
Insights
A novel peptide, SP5.2, selectively targets VEGF receptor Flt-1, inhibiting tumor angiogenesis and metastasis. This peptide shows potential for developing targeted anti-cancer therapies by blocking VEGF-mediated cell proliferation and migration.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Vascular endothelial growth factor (VEGF) drives tumor angiogenesis and metastasis by activating tyrosine kinase receptors Flt-1 and KDR.
- Inhibiting VEGF signaling can enhance apoptosis and suppress tumor growth.
Purpose of the Study:
- To identify and characterize a novel peptide, SP5.2, that selectively targets Flt-1 and inhibits VEGF-mediated events.
- To evaluate the anti-angiogenic and anti-metastatic potential of SP5.2 in vitro.
- To explore SP5.2 as a potential targeting agent for therapeutic development.
Main Methods:
- Phage-display library screening to identify SP5.2.
- In vitro assays including cell binding, proliferation, migration, and capillary-like tube formation using human endothelial cells (HCECs).
- VEGF-induced permeability assays and development of SP5.2-reporter protein conjugates.
Main Results:
- SP5.2 selectively binds to VEGF-stimulated HCECs and inhibits VEGF-induced proliferation and migration.
- SP5.2 completely inhibits VEGF-induced angiogenesis in vitro and prevents VEGF-induced permeability.
- SP5.2 conjugates show significantly enhanced binding affinity to Flt-1, suggesting potential for nanomolar therapeutic activity.
Conclusions:
- SP5.2 is a potent inhibitor of VEGF-mediated angiogenesis and metastasis.
- SP5.2 demonstrates selective binding to Flt-1 and effective inhibition of key angiogenic processes.
- SP5.2-based conjugates hold promise for developing targeted anti-cancer therapies.
