An IP-10 (CXCL10)-Derived Peptide Inhibits Angiogenesis

Cecelia C Yates-Binder,Timothy Turner,Alan Wells,Richard J Bodnar,Jesse Jaynes,Margaret Rodgers,Paul Proost

doi:10.1371/journal.pone.0040812

Abstract

Angiogenesis plays a critical role in processes such as organ development, wound healing, and tumor growth. It requires well-orchestrated integration of soluble and matrix factors and timely recognition of such signals to regulate this process. Previous work has shown that newly forming vessels express the chemokine receptor CXC receptor 3 (CXCR3) and, activation by its ligand IP-10 (CXCL10), both inhibits development of new vasculature and causes regression of newly formed vessels. To identify and develop new therapeutic agents to limit or reverse pathological angiogenesis, we identified a 21 amino acid fragment of IP-10, spanning the α-helical domain residues 77–98, that mimic the actions of the whole IP-10 molecule on endothelial cells. Treatment of the endothelial cells with the 22 amino acid fragment referred to as IP-10p significantly inhibited VEGF-induced endothelial motility and tube formation in vitro, properties critical for angiogenesis. Using a Matrigel plug assay in vivo, we demonstrate that IP-10p both prevented vessel formation and induced involution of nascent vessels. CXCR3 neutralizing antibody was able to block the inhibitory effects of the IP-10p, demonstrating specificity of the peptide. Inhibition of endothelial function by IP-10p was similar to that described for IP-10, secondary to CXCR3-mediated increase in cAMP production, activation of PKA inhibiting cell migration, and inhibition of VEGF-mediated m-calpain activation. IP-10p provides a novel therapeutic agent that inhibits endothelial cell function thus, allowing for the modulation of angiogenesis.

Highlights

The formation of new blood vessels, whether by angiogenesis or vasculogenesis, is critical for several physiological processes including embryogenesis, organogenesis and vascular remodeling
These small oligo peptides have been shown to interact with various membrane receptors. Using this as the modality for our analysis, we identified the a-helical region of IP-10 as a possible agonist for CXC receptor 3 (CXCR3) (Figure 1)
To determine whether IP10p binds to CXCR3 on human dermal microvascular endothelial cells (HMEC-1) [4], we analyzed the ability of Biotin-tagged IP-10 and IP10p to compete for binding to human microvascular endothelial cells HMEC-1 using flow cytometery

Summary

Introduction

The formation of new blood vessels, whether by angiogenesis or vasculogenesis, is critical for several physiological processes including embryogenesis, organogenesis and vascular remodeling. Researchers have sought to better understand the signaling pathways of these angiogenesis regulators to provide new therapies to modulate these and other pathological conditions. Recent evidence demonstrates that members of the CXC chemokine family can act as either angiogenic or angiostatic factors, depending on the presence of the ELR (Glu-Leu-Arg) motif in their NH2 terminus [1]. Among this family, the chemokines (IP-9/ITAC (CXCL11) IP-10 (CXCL10), MIG (CXCL9) and CXCL4 (PF4) lack the canonical N-terminal ELR sequence [4] and bind in common to the ubiquitous CXCR3 chemokine receptor. IP-10 and PF4 have been reported to be angiostatic and have anti-tumor activity via its signaling through CXCR3 [7,9,10]

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Results

Conclusion