Abstract LB-242: MicroRNA-125b, a key mediator for Snail-induced stem cell propagation and chemoresistance.
Authors: Liu Zixing , Liu Hao , Desai Shruti , Schmitt David C. , Zhou Ming , Khong Hung T. , Klos Kristine S. , McClellan Steven , Fodstad Oystein , Tan Ming 
Published: August 2014

MicroRNAs (miRNAs) are short ribonucleic acid molecules commonly deregulated in various tumors. Our previous studies have shown that miR-125b plays a major role in the chemoresistance of breast cancer. Snail, a transcription factor important in epithelial-mesenchymal transition, has been reported to be responsible for tumor metastasis and recurrence, however, the mechanism by which Snail exerts these effects is not understood. We report a novel mechanism in which upregulation of miR-125b through Wnt signaling by Snail enriches cancer stem cells and confers chemoresistance. We found that Snail is upregulated in chemoresistant cancer cells. Overexpressing Snail increases miR-125b expression by activating the Wnt/beta-catenin/TCF4 axis, while silencing Snail decreases miR-125b expression. Snail activates Wnt signaling by interacting with beta-catenin/TCF4 to activate the expression of target genes. We discovered 5 TCF4 binding sites in the miR-125b promoter. The full length miR-125b promoter showed higher activity with Snail overexpression, and deletion of TCF4 binding sites decreased the promoter activity of miR-125b in a dose-dependent manner. Knockdown of TCF4 decreased miR-125b expression, which confirms that TCF4 is required for Snail to fully activate the miR-125b promoter.

Bak1 is a pro-apoptosis molecule and confirmed target of miR-125b. Deregulation of Bak1 is important in cancer development and drug resistance. Since overexpression of Snail confers chemotherapeutic resistance, and Snail was upregulated in Paclitaxel-resistant cells, we examined whether Snail induces drug resistance through Bak1. We found low Bak1 expression in Paclitaxel-resistant cells, which have high expression of Snail. To verify whether Snail regulates Bak1 expression, we expressed Snail in Snail-low expressing cells and saw repression of Bak1 expression. To confirm, we knocked-down Snail in Snail-high expressing cells, which resulted in increased Bak1 expression. Overexpression of Snail or miR-125b increased the resistance of cancer cells to chemotherapy. Inhibiting miR-125b expression or restoring Bak1 expression re-sensitized cells to chemotherapy. These results indicate that Snail increases chemoresistance by repressing Bak1 through miR-125b.

Overexpression of miR-125b significantly increases the cancer stem cell population, while depletion of miR-125b or rescue of Bak1 expression increases the non-stem cell population. These findings support that miR-125b functions as a key mediator in Snail-induced cancer stem cells and chemoresistance. This novel mechanism for Snail-induced stem cell propagation and chemoresistance may have important implications in the development of strategies for overcoming chemotherapy resistance.

Work supported by: Vincent F. Kilborn Jr. Cancer Research Foundation, NIH Grant R01CA149646, and the Norwegian Hospitalet Legater Project 334003.

Citation Format: Zixing Liu, Hao Liu, Shruti Desai, David C. Schmitt, Ming Zhou, Hung T. Khong, Kristine S. Klos, Steven McClellan, Oystein Fodstad, Ming Tan. MicroRNA-125b, a key mediator for Snail-induced stem cell propagation and chemoresistance. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-242. doi:10.1158/1538-7445.AM2013-LB-242