Pre-clinical studies of Notch signaling inhibitor RO4929097 in inflammatory breast cancer cells. Breast Cancer Res Treat

Department of Radiation Oncology, Unit 1202, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Blvd, Houston, TX 77030, USA.
Breast Cancer Research and Treatment (Impact Factor: 3.94). 05/2012; 134(2):495-510. DOI: 10.1007/s10549-012-2075-8
Source: PubMed


Basal breast cancer, common among patients presenting with inflammatory breast cancer (IBC), has been shown to be resistant to radiation and enriched in cancer stem cells. The Notch pathway plays an important role in self-renewal of breast cancer stem cells and contributes to inflammatory signaling which promotes the breast cancer stem cell phenotype. Herein, we inhibited Notch signaling using a gamma secretase inhibitor, RO4929097, in an in vitro model that enriches for cancer initiating cells (3D clonogenic assay) and conventional 2D clonogenic assay to compare the effect on radiosensitization of the SUM149 and SUM190 IBC cell lines. RO4929097 downregulated the Notch target genes Hes1, Hey1, and HeyL, and showed a significant reduction in anchorage independent growth in SUM190 and SUM149. However, the putative self-renewal assay mammosphere formation efficiency was increased with the drug. To assess radiosensitization of putative cancer stem cells, cells were exposed to increasing doses of radiation with or without 1 μM RO4929097 in their standard (2D) and self-renewal enriching (3D) culture conditions. In the conventional 2D clonogenic assay, RO4929097 significantly sensitized SUM190 cells to ionizing radiation and has a modest radiosensitization effect in SUM149 cells. In the 3D clonogenic assays, however, a radioprotective effect was seen in both SUM149 and SUM190 cells at higher doses. Both cell lines express IL-6 and IL-8 cytokines known to mediate the efficacy of Notch inhibition and to promote self-renewal of stem cells. We further showed that RO429097 inhibits normal T-cell synthesis of some inflammatory cytokines, including TNF-α, a potential mediator of IL-6 and IL-8 production in the microenvironment. These data suggest that additional targeting agents may be required to selectively target IBC stem cells through Notch inhibition, and that evaluation of microenvironmental influences may shed further light on the potential effects of this inhibitor.

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    • "In this study, we chose to evaluate the Notch3 pathway, and we assessed the effects of Notch3 suppression on ALDH1A1expression, clonogenicity, and the cell cycle in A549s cells. Expression of ALDH1A1 and the Notch genes were confirmed to be higher in A549s than in A549 cells, as reported previously [30]. When ALDH1A1 was downregulated in A549s cells, the expression levels of all Notch genes increased significantly, especially those of Notch1 (by over 15-fold). "
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    ABSTRACT: To evaluate the expression of ALDH1A1 in lung adenoma stem cells (LASCs) and maintenance of their stemness through the Notch pathway. LASCs (A549s) were isolated from lung adenoma cells (A549) and identified by their coexpression of CD133 and CD326 and their capacity formulti-directional differentiation. Expression of ALDH1A1 in A549 and A549s cells were evaluated by Real-time PCR. Effects of ALDH1A1 upregulation in A549 cells and its downregulation in A549s cells on the clonogenicity and cell cycle were assessed by colony-forming unit assay. Moreover, the effects of ALDH1A1 on the Notch pathway, and thus on the cell cycle, were studied. A549s cells were successfully isolated and identified.ALDH1A1expression was significantly higher in A549s than in A549 cells. Clonogenicity was significantly decreased in A549s cells treated with ALDH1A1 siRNA. Duration of the G1 stage of the cell cycle increased after ALDH1A1 was overexpressed, or decreased with ALDH1A1 siRNA. ALDH1A1, Notch1, -2, and -3, CDK2, and CCNE1 expression levels were higher in A549s cells than in A549 cells. Expression of Notch1, -2, and -3, CDK2, and CCNE1 was significantly decreased by upregulation of ALDH1A1 in A549 cells, but increased by its interruption in A549s cells. When Notch3 or CDK2 expression was downregulated, the expression levels of ALDH1A1, Notch1, -2, and -3, CDK2, and CCNE1 were reduced in all cell types. ALDH1A1 expression improved clonogenicity and inhibited the cell cycle, maintaining the stemness of the A549s cells; this may involve suppression of the Notch/CDK2/Cyclin pathway.
    Full-text · Article · Mar 2014 · PLoS ONE
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    • "IL-1 is a potential inducer of IL-8 production by breast cancer cells in vitro [130] IL-6 Contributes to the tumor proliferation via up-regulating antiapoptotic and angiogenic response [31] [94] [131] Produced by IBC cell lines (SUM149 and SUM190) stimulate Notch signaling that induces self-renewal pathways of cancer stem cell [93] IL-8 Has been identified as an angiogenic stimulator [132] Promotes invasion and motility of IBC carcinoma cells by inducing of PI3 k/Akt signaling pathway and increasing the expression of the mesenchymal marker fibronectin [42] IBC cell lines (SUM149 and SUM190) secrets IL-8 that promotes cancer stem cell self-renewal pathways through Notch signaling [93] IL-10 Production of IL-10 has been linked to chronicinfection with Mouse Mammary Tumor Virus (MMTV), which related to IBC aggressiveness and etiopathogenesis [92] [109] [133] TNF-a Contributes to epithelial mesenchymal transition (EMT) in breast tumor cells [134] [135] Act as a mediator for IL-6 and IL-8 production [93] Induce NF-B signaling pathway activation in stem-like phenotype [136] MCP-1 or CCL2 Promotes breast tumor growth and metastasis [137] CCL2 and CCL5 are up-regulated by TNF-a and IL-1bin breast cancer cells [138] 6 M.M. Mohamed et al. Please cite this article in press as: Mohamed MM et al., Inflammatory breast cancer: New factors contribute to disease etiology: "
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