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Research Articles: Therapeutics, Targets, and Development

Antimigratory effect of TK1-2 is mediated in part by interfering with integrin ₂β₁

Cancer Research Institute and Department of Biomedical Science, College of Medicine, The Catholic University of Korea, Seoul, Korea

Requests for reprints: Young Ae Joe, Cancer Research Institute and Department of Biomedical Science, College of Medicine, The Catholic University of Korea, Banpo-dong 505, Seocho-ku, Seoul 137-701, Korea. Phone: 82-2-590-2404; Fax: 82-2-532-0575. E-mail: youngjoe{at}catholic.ac.kr

Abstract

The recombinant two kringle domain of human tissue-type plasminogen activator (TK1-2) has been shown to inhibit endothelial cell proliferation, angiogenesis, and tumor cell growth despite of sharing a low amino acid sequence homology with angiostatin. Here, we explored a possible inhibitory mechanism of action of TK1-2 by focusing on antimigratory effect. TK1-2 effectively inhibited endothelial cell migration induced by basic fibroblast growth factor or vascular endothelial growth factor in a dose-dependent manner and tube formation on Matrigel. It blocked basic fibroblast growth factor–induced or vascular endothelial growth factor–induced phosphorylation of extracellular signal-regulated kinase 1/2 and formation of actin stress fibers and focal adhesions. Interestingly, TK1-2 alone induced the weak phosphorylation of focal adhesion kinase, whereas it inhibited focal adhesion kinase phosphorylation induced by growth factors. When immobilized, TK1-2 promoted adhesion and spreading of endothelial cells compared with bovine serum albumin. However, treatment with anti-₂β₁ blocking antibody markedly diminished endothelial cell adhesion to immobilized TK1-2 compared with anti-_vβ₃ or anti-₅β₁ antibody. Pretreatment of soluble TK1-2 also altered the binding level of anti-₂β₁ antibody to endothelial cells in fluorescence-activated cell sorting analysis. Indeed, a blocking antibody against integrin ₂β₁ or knocking down of integrin ₂ expression prevented the inhibitory effect of TK1-2 in cell migration. Therefore, these results suggest that TK1-2 inhibits endothelial cell migration through inhibition of signaling and cytoskeleton rearrangement in part by interfering with integrin ₂β₁. [Mol Cancer Ther 2008;7(7):2133–41]

Grant support: Korea Research Foundation Grant funded by the Korean Government (MOEHRD; KRF-2003-015-C00446).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Supplementary material for this article is available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

² In preparation.

Received 12/27/07; revised 4/ 8/08; accepted 4/15/08.