Abstract
Purpose :
TIE2/ANGPT signaling has important roles in angiogenesis. Ang-1 is a critical player in vessel maturation, and it mediates migration, adhesion and survival of endothelial cell. We have used phage display to identify agonistic TIE-2 binding peptides. Here, we evaluate their potential to inhibit angiogenesis.
Methods :
For affinity selection, panning phage-display was performed against immobilized TIE-2 with a 9mer peptide library from New England BioLabs. To study the in vitro effects of peptides on angiogenesis, human umbilical vein endothelial cell (HUVECs) were treated to detect stimulation of TIE2 downstream signaling by pAKT western blot with accompanying inhibitory effects on VEGF driven cell proliferation. For in vivo experiments, different doses of peptides were tested in the corneal micropocket assay with FGF pellets (80 ng) surgically implanted into cornea micropockets in C57BL/6J mice. The mice were treated daily after surgery through intraperitoneal injections. On day 5, new blood vessel growth area (VA in mm2) was quantified.
Results :
Among the three candidates from phage display, we have identified a peptide which we have named P2 as the most active. Western blot analysis showed that P2 treatment significantly increased the level of pAKT (Ser473). Interestingly, we found that P2 has synergistic effect with Ang-1. In vivo studies with mice revealed that P2 significantly reduced VA in the cornea (32% reduction, P < 0.006) with a dose of 20mg/kg per day.
Conclusions :
Herein we show that a TIE 2 binding peptide significantly modulated angiogenesis in cornea, suggesting that it may be useful for the treatment of angiogenesis dependent diseases.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.