American Association for Cancer Research
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Supplemental Figures 1 - 8, Tables 1 - 2, Methods from Akt1 and Akt3 Exert Opposing Roles in the Regulation of Vascular Tumor Growth

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posted on 2023-03-30, 22:29 authored by Thuy L. Phung, Wa Du, Qi Xue, Sriram Ayyaswamy, Damien Gerald, Zeus Antonello, Sokha Nhek, Carole A. Perruzzi, Isabel Acevedo, Rajesh Ramanna-Valmiki, Paul Rodriguez-Waitkus, Ladan Enayati, Marcelo L. Hochman, Dina Lev, Sandaruwan Geeganage, Laura E. Benjamin

Supplementary Figure S1. Phosphorylated-Akt staining in human vascular tumors. Immunohistochemical stains for phospho-Akt (S473) in normal human skin and human vascular tumors. Representative areas of "low" and "high" levels of staining within a tumor are shown. Arrows in normal skin indicate immuno-reactive blood vessels. Scale bar, 100 μm. Supplementary Figure S2. Isolation of primary infantile hemangioma endothelial cells, and effects of Akt1 knockdown on vascular tumor cell apoptosis. Supplementary Figure S3. Expression levels of Akt1, Akt2 and Akt3 in double transgenic myrAkt1 mice, and the development of hemangioma in myrAkt1 skin grafts in syngeneic immunocompetent FVB recipients. Supplementary Figure S4. Akt1, Akt2 and Akt3 expression in human vascular tumors. Supplementary Figure S5. Akt3 expression in vascular tumor cell lines. Supplementary Figure S6. Knockdown of Akt1, Akt2 and Akt3 in tumor cells, and the effects of loss of Akt1, Akt2 and Akt3 on vascular tumor growth in vivo. Supplementary Figure S7. Loss of S6-Kinase rescues the effects of Akt3 on tumor cell migration. Supplementary Figure S8. Loss of Rictor increases S6K pathway activation. Supplementary Table S1. In vitro properties of the S6K inhibitor LY2584702. *Kinases related to p70 S6K. Supplementary Table S2. Sequences of lentiviral short-hairpin RNA (shRNA) constructs.

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ARTICLE ABSTRACT

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) through modulation 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, our findings 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.

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