Abstract 1413: Interleukin-1β promotes brain metastasis of breast cancer by activating Notch signaling through reactive astrocytes

Abstract

Abstract Brain metastasis is found in around 20% of advanced breast cancer patients whose survival rates are generally less than one year after the diagnosis. Despite this clinical importance, the molecular basis of breast tumor metastasis to the brain is still virtually unknown. The goal of this project is to elucidate the role of brain microenvironment, particularly astrocytes, in breast tumor cell growth and invasion. Our analysis of existing microarray database indicate that interleukin-1β (IL-1β) is highly expressed in MDA-MB231BrM2a (231BrM) cell line which is capable of specifically metastasizing to the brain. IL-1β is known to have an ability of “activating” astrocytes that are often found in the metastatic niche, and therefore, it is speculated that the reactive astrocytes can promote growth and invasion of metastasized cells in the brain. To explore this possibility, we examined the effect of IL-1β and the conditioned medium of 231BrM cells on rat primary astrocytes in vitro. Our results of qRT-PCR and Western blot analysis indicate that IL-1β and the conditioned medium significantly up-regulated the Notch ligand, JAG1, in astrocytes, and that this up-regulation was abrogated by the NF-kB inhibitor, PDTC. To further examine the interaction of astrocytes and tumor cells in the brain, we intracardially injected 231BrM cells in nude mice, and tumor growth was monitored by Xenogen bioimager. After 4 weeks, metastatic tumor growth was observed at multiple sites of the brain, and the results of immunohistochemical analysis indicate that astrocytes adjacent to the metastatic lesions strongly expressed JAG1 and the reactive astrocyte marker, GFAP (Glial fibrillary acidic protein). These results strongly suggest that IL-1β secreted from 231BrM activates astrocytes and induces the expression of the Notch ligand, which in turn promotes the growth and invasion of metastasized breast tumor cells. To further test this hypothesis, we performed in vitro invasion assay by culturing astrocytes with or without the treatment of IL-1β on top of matrigel, followed by overlaying MDA-MB231 cells. We found that the IL-1β treatment of astrocytes significantly augments the invasive ability of MDA-MB231. We also introduced DNMAML (dominant negative mastermind) gene by lentivirus infection into 231BrM and found that the blocking of Notch signal by DNMAML failed to inhibit tumor growth but significantly down-regulated the expression of the MMPs (Matrix metallopeptidase) genes and decreased the invasive ability of 231BrM. These data strongly support our notion that a metastasized breast tumor cell in the brain secretes IL-1β and activates astrocytes which in turn up-regulates JAG1 and that the interaction of the activated astrocytes and tumor cells turns on Notch signaling in the tumor cells followed by promotion of tumor invasion. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1413.