Abstract 1248: The Gemini vitamin D analogue BXL0124 inhibits Notch signaling via HES1, resulting in the reduction of CD44+/CD24-/low subpopulation and proliferation of MCF10DCIS cells

Activation of Notch signaling is correlated with poor prognosis and decreased survival of breast cancer patients. Recent studies reported that Notch signaling plays an important role for the maintenance of tumor-initiating cells in breast cancer. Using MCF10DCIS.com human breast cancer cells (MCF10DCIS), which contain tumor-initiating subpopulation (CD44+/CD24-/low), we demonstrated that the Gemini vitamin D analog, BXL0124, reduced the tumor-initiating subpopulation in vitro and repressed the growth of MCF10DCIS xenograft tumors in vivo. In the present study, we investigated Notch signaling in CD44+/CD24-/low and CD44+/CD24+ subpopulations of MCF10DCIS cells and determined whether BXL0124 represses tumor-initiating subpopulation of breast cancer by targeting Notch signaling. CD44+/CD24-/low subpopulation showed higher Notch1 activation and cell proliferation than CD44+/CD24+ subpopulation in MCF10DCIS cells. To investigate the effects of BXL0124 on Notch signaling, the protein levels of Notch receptors and their ligands were determined. BXL0124 decreased activated Notch1 (c-Notch1) as well as its ligands, Jagged1, Jagged2 and DLL1. However, total Notch1 was not affected by BXL0124, suggesting that BXL0124 inhibits Notch signaling by decreasing Notch ligands. A downstream target of Notch signaling, cMyc, and cell proliferation were reduced by BXL0124. The inhibition of Notch1 activation by BXL0124 was blocked with knock-down of the vitamin D receptor (VDR), indicating that it is a VDR-dependent event. In a time course study, the effects of BXL0124 on the mRNA expression of Notch1, Notch ligands and downstream targets were investigated. BXL0124 decreased the mRNA levels of Jagged1, Jagged2 and DLL1 starting at 8 h which followed by the repression of cMyc mRNA at 16 h. The mRNA level of a transcriptional repressor HES1, which is a key downstream target of Notch signaling, was rapidly induced by BXL0124 as early as 30 min. To determine the involvement of HES1 in Notch signaling, both knock-down and overexpression of HES1 were performed by using HES1 siRNA and HES1-expression vector, respectively. The inhibition of Notch signaling by BXL0124 was reversed by the knock-down of HES1, while the overexpression of HES1 inhibited Notch activation, reduced CD44+/CD24-/low population and decreased cell proliferation of MCF10DCIS cells. These results suggest that HES1 functions as a repressor of Notch signaling, and BXL0124 inhibits Notch signaling in a HES1-dependent manner. The present study demonstrates that BXL0124 inhibits Notch signaling by rapid induction of HES1 which results in the reduction of CD44+/CD24-/low subpopulation and proliferation of MCF10DCIS cells. Our study suggests the Gemini vitamin D analog, BXL0124, as a potential agent to repress the tumor-initiating subpopulation of breast cancer by targeting Notch signaling. Citation Format: Jae Young So, David M. Salerno, Hubert Maehr, Milan Uskokovic, Nanjoo Suh. The Gemini vitamin D analogue BXL0124 inhibits Notch signaling via HES1, resulting in the reduction of CD44+/CD24-/low subpopulation and proliferation of MCF10DCIS cells. [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 1248. doi:10.1158/1538-7445.AM2014-1248