Abstract 3018: SHIP2 plays an oncogenic role in breast cancer stem cells through JNK/vimentin activation and its phosphatase activity
Authors: Fu Chiung-Hui , Lin Ruey-Jen , Yu John , Chang Wen-Wei , Liao Guo-Shiou , Chang Wen-Ying , Tseng Ling-Ming , Tsai Yi-Fang , Yu Jyh-Cherng , Yu Alice L. 
Published: May 2015

Abstract
Background and Objectives: Over-expression of SH2-containing-5’-inositol phosphatase-2 (SHIP2) which converts phosphatidylinositol (3,4,5)-trisphosphate [PI(3,4,5)P3] into phosphatidylinositol (3,4)-biphosphate [PI(3,4)P2] has been shown to correlate with decreased disease-free survival in breast cancer. However, its role in breast cancer stem cells (BCSCs) remains unclear.
Results: We analyzed 60 clinical breast cancer specimens and found that the percentage of SHIP2+ cells was positively correlated with that of CD24-CD44+ cells. Among 20 estrogen receptor (ER)-negative samples, 17 had greater SHIP2 expression in CD24-CD44+ subpopulation than the remaining subpopulation. Examination of mouse xenografts derived from primary human breast cancer specimens revealed that SHIP2 protein and its tyrosine 1135 phosphorylation were significantly higher in BCSCs, identified as CD24-CD44+ or ALDH+, than non-BCSCs. SHIP2 silencing or inhibitor of SHIP2 phosphatase significantly decreased mammosphere-forming efficiency, percentage of ALDH+ subpopulation in vitro and tumorigenicity of BCSCs in vivo. SHIP2 shRNA knockdown and SHIP2 phosphatase inhibitor suppressed p-AktSer473 levels. Addition of PI(3,4)P2 could partially rescue Akt phosphorylation and ALDH+ subpopulation in SHIP2-overexpressing AS-B634 cells treated with SHIP2 phosphatase inhibitor. Whereas, PI(3,4)P2 failed to reverse the effects of SHIP2-silencing on Akt activation and ALDH+ subpopulation. Futhermore, over-expression of SHIP2 enhanced the expression of epithelial-mesenchymal transition markers including vimentin (VIM), which was mainly expressed in ER-negative breast cancer cells with higher level in mammospheres than monolayer culture. Ablation of c-Jun N-terminal kinase 1 (JNK1), JNK2 or VIM hampered the increases of both ALDH+ population and tumorigenicity induced by SHIP2 over-expression. BCSCs displayed greater expression of phospho-JNK than non-BCSCs. Futhermore, silencing of JNK suppressed the upregulation of VIM by SHIP2 in breast cancer.
Conclusion: Our study is the first to demonstrate that SHIP2 is crucial for maintaining the properties of BCSCs. Within ER-negative breast cancer, there was a significant correlation of greater expression of SHIP2 in BCSCs than non-BCSCs. We documented a novel functional role of SHIP2 in enhancing mammosphere formation and ALDH+ subpopulation in vitro, promoting tumorigenicity in vivo and inducing the expression of EMT markers including VIM. The upregulation of VIM was mediated by JNK activation. Furthermore, the phosphatase activity of SHIP2 also contributed to its role in BCSCs. In summary, our study has uncovered an oncogenic role of SHIP2 in promoting BCSC features and its underlying molecular mechanisms, providing the impetus for the design of novel BCSC-directed therapeutics by targeting SHIP2.
Citation Format: Chiung-Hui Fu, Ruey-Jen Lin, John Yu, Wen-Wei Chang, Guo-Shiou Liao, Wen-Ying Chang, Ling-Ming Tseng, Yi-Fang Tsai, Jyh-Cherng Yu, Alice L. Yu. SHIP2 plays an oncogenic role in breast cancer stem cells through JNK/vimentin activation and its phosphatase activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3018. doi:10.1158/1538-7445.AM2014-3018