Abstract 2284: Investigation of notch-1, ErbB-2, and ERα crosstalk in breast cancer

Abstract

Notch-1 is a breast oncogene that is overexpressed with its ligand Jagged-1 in human breast cancers-associated with the poorest overall survival. Notch signaling can be inhibited by a gamma-secretase inhibitor (GSI). We have recently determined that the estrogen receptor-alpha or ErbB-2 inhibits Notch-1 activation in breast cancer cells. In the current study, we sought to uncover the complex crosstalk between ER, ErbB-2, and Notch-1 and ask whether triple combination treatment can disrupt this complex cross-talk model to inhibit growth of ER positive breast cancer cells (MCF-7) and possibly prevent anti-hormone resistance. The results demonstrated that following 1 week of estrogen deprivation, MCF-7 cells develop resistance and regrow. Canonical Notch gene targets: Hes-1, Hey-1, Hes-5, and Deltex-1 are significantly increased at this 1 week time point. Similarly, ErbB-2 mRNA increased significantly after 72 hours of estrogen deprivation. Upon treatment with fulvestrant, a pure ant-estrogen, ER transcriptional activity as measured by PS2 mRNA is inhibited after one week. Interestingly, the combination of fulvestrant and lapatinib, an anti-ErbB-2 tyrosine kinase inhibitor, synergistically increased both Deltex-1 and ErbB-2 mRNAs which were reduced upon GSI treatment. Western blot analysis showed an increase in ER and tyrosine-phosphorylated ErbB-2 and total ErbB-2 proteins. This increase in protein expression was further increased upon dual treatment with fulvestrant and lapatinib. Conversely, treatment with a Notch inhibitor, GSI caused a decrease in both total and tyrosine-phosphorylated ErbB-2. Cell cycle analysis showed that fulvestrant treatment significantly growth arrested cells in G1 while decreasing cells in S-phase. However, lapatnib or GSI or the combination treatments did not further enhance the growth inhibitory effects of fulvestrant. Similar results were observed using Annexin-V analysis by flow cytometry as a measure of early apoptosis. These results suggest that while inhibiting ER and ErbB-2 simultaneously increases Notch activation in a synergistic manner, this dramatic increase in Notch activity in the short term does not sensitize ER positive breast cancer cells to a Notch inhibitor to undergo cell death. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2284. doi:10.1158/1538-7445.AM2011-2284