Wogonin suppresses tumor growth in vivo and VEGF-induced angiogenesis through inhibiting tyrosine phosphorylation of VEGFR2 Na Lu a,1 , Ying Gao a,1 , Yun Ling a , Yan Chen a , Yong Yang a , Hong-Yan Gu a , Qi Qi a , Wei Liu a , Xiao-Tang Wang b , Qi-Dong You a, ⁎, Qing-Long Guo a, ⁎ a Jiangsu Key Laboratory of Carcinogenesis and Intervention (China Pharmaceutical University), Nanjing 210009, People's Republic of China b Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA article info abstract Article history: Received 4 December 2007 Accepted 20 February 2008 Previous studies revealed that wogonin, a naturally occurring monoflavonoid extracted from Scutellariae radix, possessed anticancer activity both in vitro and in vivo. However, the molecular mechanism of its potent anticancer activity remains poorly understood and warrants further investigations. In this study, we found for the first time that wogonin inhibited the growth and tumor angiogenesis of human gastric carcinoma in nude mice. We explored the inhibitory effect of wogonin on angiogenesis stimulated by vascular endothelial growth factor (VEGF) in vitro. Wogonin suppressed the VEGF-stimulated migration and tube formation of human umbilical vein endothelial cells (HUVECs). It also restrained VEGF-induced tyrosine phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2). This inhibition of receptor phosphorylation was correlated with a significant decrease in VEGF-triggered phosphorylated forms of ERK, AKT and p38. Taken together, these findings strongly suggest that wogonin might be a promising antitumor drug. © 2008 Elsevier Inc. All rights reserved. Keywords: Wogonin Human gastric carcinoma Tumor Angiogenesis VEGF VEGFR2 Introduction Angiogenesis, the formation of new blood vessels, is a fundamental step in physiologic processes, such as wound healing, organ growth and reproduction, as well as in pathological conditions like tumor progression, metastasis, chronic inflammation and arthritis (Folkman, 1971, 1995; Folkman and Chesney, 1997; Hanahan and Folkman, 1996; Risau, 1997). To some extent, tumor growth is dependent on angiogenesis. An avascular tumor can rarely increase in size N 2– 3 mm 3 . Once vascularized, a tumor grows rapidly and nearly exponentially. The newly generated blood vessels are required to supply adequate oxygen and nutrition to the growing tumor mass, and for initiation of metastatic spread (Folkman and Klagsbrun, 1987; Kerbel and Folkman, 2002). The angiogenic process is a highly complex, dynamic process regulated by a number of pro- and antiangiogenic molecules. The process of tumor angiogenesis involves recruitment of sprouting vessels from existing blood vessels and incorporation of endothelial progenitors into the growing vascular bed (Rafii et al., 2002). Events included in this process are the proliferation, migration and invasion of endothelial cells, organization of endothelial cells into functional tubular structures, maturation of vessels, and vessel regression. Studies of the molecular basis of angiogenesis have identified a number of growth factor receptor pathways that promote tumor angiogenesis. One of the major pathways involved in this process is the vascular endothelial growth factor (VEGF) family of proteins and receptors. Vascular endothelial growth factor (VEGF) is one of the most important proangiogenic factors, which acts as a mitogen for vascular endothelial cells in vitro and as an angiogenic factor in vivo (Ferrara, 1996). It is overexpressed in various human cancers (Macchiarini et al., 1992; Paley et al., 1997; Weidner et al., 1991, 1993). VEGF ligands mediate their angiogenic effects via several different receptors. Two receptors were originally identified on endothelial cells and characterized as the specific tyrosine kinase receptors VEGFR-1 (also referred to a fms-like tyrosine kinase 1 [Flt- 1]) (Shibuya et al., 1990) and VEGFR-2 (also referred to as KDR (Terman et al., 1992), and the murine homologue, Flk-1) (Matthews et al., 1991). Inhibition of VEGF activity by neutralizing antibodies or the introduc- tion of dominant negative VEGF receptors into endothelial cells of tumor-associated blood vessels often results in inhibition of tumor growth or even tumor regression (Kim et al., 1993). Wogonin (C 16 H 12 O 5 )(Fig. 1), a naturally occurring monoflavonoid extracted from Scutellariae radix (Chi et al., 2001), has been shown to be a promising candidate for selective and effective management of human cancers (Tai et al., 2005). For example, wogonin inhibited the growth of human ovarian cancer cell A2780 (Li et al., 2003), human promyeloleukemic cell HL-60 (Lee et al., 2002; Yu et al., 2005), human hepatocellular carcinoma cell SK-HEP-1 (Chen et al., 2002) and human hepatoma cell SMMC-7721 (Wang et al., 2006a). Our previous study also showed that wogonin treatment resulted in significant inhibition Life Sciences 82 (2008) 956–963 ⁎ Corresponding authors. Q.-L. Guo is to be contacted at Tel./fax: +86 25 83271055. Q.-D. You, Tel./fax: +86 25 83271351. E-mail addresses: youqidong@gmail.com (Q.-D. You), anticancer_drug@yahoo.com.cn (Q.-L. Guo). 1 These two authors contributed equally to this article. 0024-3205/$ – see front matter © 2008 Elsevier Inc. All rights reserved. doi:10.1016/j.lfs.2008.02.013 Contents lists available at ScienceDirect Life Sciences journal homepage: www.elsevier.com/locate/lifescie