Organotropism is one of the most distinct properties of cancer metastasis which indicates that metastatic cancer cells can only thrive in permissive microenvironment (Hu et al, 2009
; Lu & Kang, 2007
). Glial cells constitute a major part of the brain cells and astrocytes are the most abundant glial cells (Grosche et al, 1999
; Ventura & Harris, 1999
). Fitzgerald et al previously found that a large amount of glial cells were trapped within the inner-tumour mass in surgically resected brain samples and demonstrated that reactive glial cells can be recruited by cancer cells to promote tumour growth in the brain (Fitzgerald et al, 2008
). Furthermore, reactive astrocytes are known to protect cancer cells from chemotherapy by activating signalling pathway related to cell survival (Langley et al, 2009
; Lin et al, 2010
). It is also noteworthy that brain-metastatic lung cancer cells were shown to stimulate the production of pro-inflammatory cytokines in astrocytes, which significantly promoted the growth of cancer cells (Seike et al, 2011
). In our study, we have shown that reactive astrocytes appeared abundantly around the brain metastatic regions and that the activated astrocytes were indeed able to promote self-renewal of CSCs by direct interaction. We have also shown that brain-metastatic cancer cells secrete excessive amounts of IL-1β and activate astrocytes which in turn promote Notch signalling in CSCs. Therefore, our results indicate that CSCs establish their niche in the brain through reciprocal interaction with astrocytes, which plays a pivotal role in pathogenesis of brain-specific metastasis of breast cancer.
Interleukin-1 (IL-1) is one of the most well studied cytokines that play key roles in cancer progression, and two forms of IL-1 have been identified, namely IL-1 α and IL-1 β (Elaraj et al, 2006
; Voronov et al, 2003
). IL-1β is processed by interleukin-1β-converting enzyme (ICE) before it becomes functional as a secreted cytokine, while IL-1α can localize in the cytosol and mediate intracellular signalling (Aotsuka et al, 1991
; Debets et al, 1995
; Miller et al, 1993
). The secreted IL-1β induces inflammatory response and alters tumour microenvironment; however, it was also shown to enhance the growth and invasion abilities of cancer cells in an autocrine fashion (Aotsuka et al, 1991
; Kawakami et al, 1997
). IL-1β is also known to promote cancer progression by upregulating pro-metastatic genes such as matrix metalloproteinases and stimulate adjacent cells to produce angiogenic proteins or growth factors including VEGF, IL-8, IL-6, TNF-α and TGF-β (Lewis et al, 2006
). Many solid tumours are known to express a high level of IL-1β which is shown to correlate with patient survival (Elaraj et al, 2006
; Lee et al, 2006
; Liu et al, 2006
). Notably, we have shown that the expression levels of IL-1β in the primary tumours of breast cancer patients were significantly associated with their brain metastatic statuses, suggesting that IL-1β may serve as a potential prognostic marker and a therapeutic target for brain metastasis. Interestingly, treatment with IL-1RA, a potent IL-1 inhibitor, was shown to significantly decrease the growth and metastases of colon and lung cancer cells in mouse models (Lewis et al, 2006
). However, BBB permeability of IL-1RA is still unknown and it has a relatively short half-life (4–6 h), therefore, developing a more effective small molecule mimicking IL-1RA is needed.
Metastatic growth is believed to be initiated by CSCs at the distant organs that constitute totally different microenvironment from the primary tumour sites. Similar to embryonic stem cells, CSCs also require specific niche which provides factors to activate various pathways for the maintenance of stemness of CSCs through direct cell–cell interaction or by secreting growth factors. In this context, it is noteworthy that Karnoub et al reported that bone mesenchymal stem cells (BMSC) generate a ‘pre-metastatic niche’ at the distant organs even before metastatic cells arrive at the site (Karnoub et al, 2007
). Interestingly, Li et al recently found that prostaglandin E2 (PGE2) was secreted by BMSCs in response to cancer cell-derived IL-1 and that the BMSC-derived PGE2 significantly enhanced the CSCs population via Akt/GSK-3/β-catenin signalling axis (Li et al, 2012
). However, the ‘pre-metastatic niche’ hypothesis may not be applicable to brain metastasis because the brain is a highly specialized organ and also due to the brain-blood barrier, it is unlikely that BMSC reach the brain before metastasis, although this possibility cannot be totally excluded.
Increasing lines of evidence suggest that the Notch pathway plays a crucial role in maintaining the stemness of CSCs in a particular microenvironment (Charles et al, 2010
; McGowan et al, 2011
). A hallmark of Notch signalling is the requirement of the ligand–receptor interaction through direct cell–cell contact, which may occur between tumour cells or tumour cell–stroma interactions (Sethi et al, 2011
; Xing et al, 2011
). Butler et al have recently shown that bone marrow endothelial cells which express Notch ligands were indeed required for the self-renewal of haematopoietic stem cells in a Notch dependent manner (Butler et al, 2010
). We have shown that direct interaction of CSCs and activated astrocytes is essential for up-regulating Notch signalling and the following self-renewal of CSCs in the brain. Our data also indicate that this activated Notch signalling up-regulated the HES5, which significantly augmented self-renewal of CSCs. It has been reported that HES5-expressing telencephalic cells are maintained as neural stem cells during embryogenesis, indicating a possible role of HES5 in maintaining self-renewal of CSCs (Ohtsuka et al, 2001
). In this report, we have discovered a novel pathological mechanism by which breast CSCs establish a niche in the metastasized brain through interaction with activated astrocytes. Our results have revealed a vicious paracrine loop of IL-1β and Notch signalling through direct interaction of CSCs and astrocytes, which in turn promotes the growth of metastasized CSCs in the brain. Importantly, we have also shown that a BBB-permeable Notch inhibitor can serve as an effective therapeutic drug to suppress metastatic growth of breast cancer in the brain. These discoveries open a window of opportunity to identify a novel therapeutic target for brain metastasis.