Microenvironment and Immunology Akt1 and Akt3 Exert Opposing Roles in the Regulation of Vascular Tumor Growth

Vascular tumors are endothelial cell neoplasms whose mechanisms of tumorigenesis are poorly understood. Moreover, current therapies, particularly those for malignant lesions, have little beneficial effect on clinical outcomes. In this study, we show that endothelial activation of the Akt1 kinase is sufficient to drive de novo tumor formation. Mechanistic investigations uncovered opposing functions for different Akt isoforms in this regulation, where Akt1 promotes and Akt3 inhibits vascular tumor growth. Akt3 exerted negative effects on tumor endothelial cell growth and migration by inhibiting activation of the translation regulatory kinase S6-Kinase (S6K) throughmodulation of Rictor expression. S6K in turn acted through a negative feedback loop to restrain Akt3 expression. Conversely, S6K signaling was increased in vascular tumor cells where Akt3 was silenced, and the growth of these tumor cells was inhibited by a novel S6K inhibitor. Overall, ourfindings offer a preclinical proof of concept for the therapeutic utility of treating vascular tumors, such as angiosarcomas, with S6K inhibitors. Cancer Res; 75(1); 40–50. 2014 AACR. Introduction Vascular tumors are endothelial cell neoplasms with a wide spectrum of clinical presentations, ranging from benign infantile hemangiomas in children to low-grade malignant hemangioendotheliomas and highly aggressive angiosarcomas in adults. To date, the molecular pathogenesis of vascular tumors is poorly understood and current therapies, particularly those for malignant vascular tumors, do not significantly improve patient outcome (1). Akt is a major signaling pathway activated by VEGF that regulates endothelial cell survival (2). In infantile hemangioma, hemangioma-derived endothelial cells (hemeEC) have constitutively active VEGF receptor-2 signalingwith highphosphorylation levels of ERK1/2 and Akt (3). Human angiosarcoma expresses VEGF-A and the VEGF receptors (4). We have shown increased phosphorylation of Akt and 4E-BP1 in angiosarcoma (5). Hyperactivation of PI3K results in hemangiosarcoma formation in chicken chorioallantoic membrane (6). Akt1, Akt2, and Akt3 are isoforms that have shared as well as distinct functions in cancer cells. BothAkt1 andAkt2 promote cancer cell survival and growth. However, in breast and ovarian cancer, Akt1 decreases cell motility and metastasis and blocks epithelial-to-mesenchymal phenotype, whereas Akt2 enhances these processes (7–9). Akt3 is preferentially required for the growth of triple-negative breast cancer (10), and a gene fusion of Akt3 with MAGI3 leads to constitutive Akt3 activation and is enriched in these tumors (11). Interestingly, there is some evidence suggesting that Akt3 exerts inhibitory effects in cancer. N-Cadherin promotes breast cancer metastasis by inhibiting Akt3, andAkt3 has been shown to inhibit lung tumor growth in mice (12–14). Studies of animal models of breast cancer with simultaneous deletion or overexpression of Akt1, Akt2, and Akt3 lend further support to Akt isoform-specific roles in cancer (8, 9). mTOR complex-1 (mTORC1) and complex-2 (mTORC2) are composed of multiple subunits, includingmTOR and Raptor (in mTORC1), andmTOR and Rictor (in mTORC2; refs. 15, 16). Akt activates mTORC1, which phosphorylates the translational regulators 4E-BP1 and p70 S6-Kinase (S6K). S6K in turn activates S6 ribosomal protein (S6; refs. 17, 18). mTORC2 directly activates Akt by phosphorylating it at serine 473, thereby exerting feedback regulation on the Akt signaling pathway (15). The S6K pathway is important in protein synthesis and cell growth, and acts as a regulator of actin cytoskeleton dynamics in cell migration (19, 20). In this study, we showed that endothelial Akt1 drives vascular tumor growth. Importantly, we have uncovered the opposing functions of Akt1 and Akt3 in the regulation of tumor growth, which is mediated through S6K, and found a novel negative feedback regulation on Akt3 by S6K. We also demonstrated the clinical utility of a novel S6K inhibitor in the treatment of vascular lesions. Department of Pathology, Texas Children's Hospital and Baylor College of Medicine, Houston,Texas. Eli Lilly and Company, Indianapolis, Indiana. Department of Pathology, Beth Israel Deaconess Medical Center, Boston, Massachusetts. Hemangioma International Treatment Center, Charleston, South Carolina. Department of Cancer Biology, MD Anderson Cancer Center, Houston, Texas. Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Corresponding Authors: Laura E. Benjamin, Eli Lilly and Company, 450 East 29th Street, 12th Floor, New York, NY 10016. Phone: 646-638-6381; Fax: 212-2134785; E-mail: [email protected]; and Thuy L. Phung, Texas Children's Hospital, 1102 Bates Avenue, Suite 830, Houston, TX 77030. Phone: 832-8245202; Fax: 832-825-1165; E-mail: [email protected] doi: 10.1158/0008-5472.CAN-13-2961 2014 American Association for Cancer Research. Cancer Research Cancer Res; 75(1) January 1, 2015 40 on April 18, 2017. © 2015 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from Published OnlineFirst November 11, 2014; DOI: 10.1158/0008-5472.CAN-13-2961