Data from Anastellin, the Angiostatic Fibronectin Peptide, Is a Selective Inhibitor of Lysophospholipid Signaling

Abstract

<div>Abstract<p>Angiogenesis is regulated by integrin-dependent cell adhesion and the activation of specific cell surface receptors on vascular endothelial cells by angiogenic factors. Lysophosphatidic acid (LPA) and sphingosine-1 phosphate (S1P) are bioactive lysophospholipids that activate G protein–coupled receptors that stimulate phosphatidylinositol 3-kinase (PI3K), Ras, and Rho effector pathways involved in vascular cell survival, proliferation, adhesion, and migration. Previous studies have shown that anastellin, a fragment of the first type III module of fibronectin, functions as an antiangiogenic peptide suppressing tumor growth and metastasis. We have previously shown that anastellin blocks serum-dependent proliferation of microvessel endothelial cells (MVEC) by affecting extracellular signal-regulated kinase (ERK)–dependent G<sub>1</sub>-S transition. However, the mechanism by which anastellin regulates endothelial cell function remains unclear. In the present study, we mapped several lysophospholipid-mediated signaling pathways in MVEC and examined the effects of anastellin on LPA- and S1P-induced MVEC proliferation, migration, and cytoskeletal organization. Both LPA and S1P activated PI3K, Ras/ERK, and Rho/Rho kinase pathways, leading to migration, G<sub>1</sub>-S cell cycle progression, and stress fiber formation, respectively. Stimulation of proliferation by LPA/S1P occurred through a G<sub>i</sub>-dependent Ras/ERK pathway, which was independent of growth factor receptors and PI3K and Rho/Rho kinase signaling. Although LPA and S1P activated both PI3K/Akt and Ras/ERK signaling through G<sub>i</sub>, anastellin inhibited only the Ras/ERK pathway. Stress fiber formation in response to LPA was dependent on Rho/Rho kinase but independent of G<sub>i</sub> and unaffected by anastellin. These results suggest that lysophospholipid mediators of G<sub>i</sub> activation leading to PI3K/Akt and Ras/ERK signaling bifurcate downstream of G<sub>i</sub> and that anastellin selectively inhibits the Ras/ERK arm of the pathway. (Mol Cancer Res 2009;7(2):255–65)</p></div>