Lysophosphatidic acid stimulates cell migration, invasion, and colony formation as well as tumorigenesis/metastasis of mouse ovarian cancer in immunocompetent mice Hui Li, 1 Dongmei Wang, 1 Hong Zhang, 1,2 Kashif Kirmani, 1 Zhenwen Zhao, 1 Rosemary Steinmetz, 1 and Yan Xu 1 1 Department of Obstetrics and Gynecology, Indiana University, Indianapolis, Indiana and 2 2nd Hospital of Jilin University, Jilin University, Changchun, Jilin, People's Republic of China Abstract We have already established human xenographic models for the effect of lysophosphatidic acid (LPA) on tumor metastasis in vivo.Thepurposeofthisworkistoestablish a preclinical LPA effect model in immunocompetent mice. We first characterized the mouse epithelial ovarian cancer (EOC) cell line ID8 for its responsiveness to LPA in cell proliferation, migration, and invasion and compared these properties with those of human EOC. The signaling path- ways related to cell migration were further investigated using pharmacologic and genetic approaches. The effects of LPA on the tumorigenesis of ID8 cells and mouse survival were then examined using two different mouse models (i.p. and orthotopic injections). LPA stimulated cell proliferation, migration, and invasion of mouse EOC ID8 cells in a manner closely resembling its activity in human EOC cells. The signaling pathways involved in LPA- induced cell migration in ID8 cells were also similar to those identified in human EOC cells. We have identified cyclooxygenase-1 and 15-lipoxygenase as two new sig- naling molecules involved in LPA-induced cell migration in both human and mouse EOC cells. In addition, LPA enhanced the tumorigenesis/metastasis of ID8 cell in vivo as assessed by increased tumor size, early onset of asci- tes formation, and reduced animal survival. We have established the first LPA-EOC preclinical model in immuno- competent mice. Because ID8 cells respond to LPA similar to human EOC cells, this model is very valuable in devel- oping and testing therapeutic reagents targeting LPA in EOC. [Mol Cancer Ther 2009;8(6):1692–701] Introduction Epithelial ovarian cancer (EOC) is the most deadly gyneco- logic disease mainly due to the lack of highly sensitive and specific methods for early detection and effective treatments for late-stage diseases. Since our first report of elevated lysophosphatidic acid (LPA) levels in human ovarian cancer ascites and the identification of the stimulatory role of LPA in EOC more than 13 years ago (1), extensive information has accumulated showing that LPA plays an important role in EOC development. LPA regulates almost every aspect of EOC cell biology, including cell proliferation, apoptosis, morphology, drug resistance, adhesion, migration, and in- vasion (2–4). Most of the studies on the mechanisms of LPA in ovarian cancer have been conducted in vitro. Only recently, animal studies showing a role for LPA and/or its receptors in the development of cancers in vivo have been published (5–9). We have recently provided the first direct evidence that LPA stimulates tumor metastasis in vivo, which is inhibited by LY294002, a phosphatidylinositol 3-kinase inhibitor, and by 17-dimethylaminoethylamino- 17-demethoxygeldanamycin, an inhibitor of the heat shock protein 90, which stabilizes hypoxia-inducible factor-1α (10, 11). However, these experiments were conducted using nude or severe combined immunodeficient mice, which are immunocompromised. It will be important to test the same concepts in immunocompetent mice. The two most common and critical processes in metasta- sis are cell migration and invasion. We and others have shown that LPA stimulates the migration and invasion of ovarian cancer cells (10–22). Although most, if not all, of these studies were conducted in established ovarian cancer cell lines, we have recently conducted LPA- and S1P- induced cell migration and invasion experiments in primary EOC and nonmalignant human ovarian surface epithelial cells (11, 23). All three LPA receptors have been implicated in the mi- gration of different cell types and under different assay con- ditions (24, 25). Whereas LPA 1 plays a very important role in migration of breast, pancreatic, and prostate cancer cells (25, 26), our data suggest that LPA 2 and LPA 3 are more important in cell adhesion, migration, and invasion of EOC cells, which are consistent with reports showing that LPA 1 is involved in the negative growth regulation in ovar- ian cancer cells and that LPA 2 and LPA 3 , but not LPA 1 , ex- pression is up-regulated in last-stage EOC (27). In addition, we have shown that either inhibitors and/or dominant- negative forms of G i protein, phosphatidylinositol 3-kinase, or cytosolic phospholipase A 2 (cPLA 2 ) completely or nearly completely block LPA-induced cell migration in human EOC cells, suggesting that these three molecules play a pi- votal role in this process (2, 10, 11, 24). The PLA 2 family of Received 11/20/08; revised 2/19/09; accepted 3/13/09; published OnlineFirst 6/9/09. Grant support: RO1 CA095042 and CA-89228 (Y. Xu). 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. Requests for reprints: Yan Xu, Indiana University-Purdue University Indianapolis, 975 West Walnut Street, IB355A, Indianapolis, IN 46202. Phone: 317-274-3972; Fax: 317-278-4828. E-mail: xu2@iupui.edu Copyright © 2009 American Association for Cancer Research. doi:10.1158/1535-7163.MCT-08-1106 Mol Cancer Ther 2009;8(6). June 2009 1692 on June 19, 2020. © 2009 American Association for Cancer Research. mct.aacrjournals.org Downloaded from Published OnlineFirst June 9, 2009; DOI: 10.1158/1535-7163.MCT-08-1106