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Forkhead Box Q1 Is a Novel Target of Breast Cancer Stem Cell Inhibition by Diallyl Trisulfide.

ABSTRACT: Diallyl trisulfide (DATS), a metabolic byproduct of garlic, is known to inhibit the growth of breast cancer cells in vitro and in vivo This study demonstrates that DATS targets breast cancer stem cells (bCSC). Exposure of MCF-7 and SUM159 human breast cancer cells to pharmacological concentrations of DATS (2.5 and 5 ?m) resulted in dose-dependent inhibition of bCSC, as evidenced by a mammosphere assay and flow cytometric analysis of aldehyde dehydrogenase 1 (ALDH1) activity and the CD44(high)/CD24(low)/epithelial specific antigen-positive fraction. DATS-mediated inhibition of bCSC was associated with a decrease in the protein level of FoxQ1. Overexpression of FoxQ1 in MCF-7 and SUM159 cells increased ALDH1 activity and the CD49f(+)/CD24(-) fraction. Inhibition of ALDH1 activity and/or mammosphere formation upon DATS treatment was significantly attenuated by overexpression of FoxQ1. In agreement with these results, stable knockdown of FoxQ1 using small hairpin RNA augmented bCSC inhibition by DATS. Expression profiling for cancer stem cell-related genes suggested that FoxQ1 may negatively regulate the expression of Dachshund homolog 1 (DACH1), whose expression is lost in invasive breast cancer. Chromatin immunoprecipitation confirmed recruitment of FoxQ1 at the DACH1 promoter. Moreover, inducible expression of DACH1 augmented DATS-mediated inhibition of bCSC. Expression of FoxQ1 protein was significantly higher in triple-negative breast cancer cases compared with normal mammary tissues. Moreover, an inverse association was observed between FoxQ1 and DACH1 gene expression in breast cancer cell lines and tumors. DATS administration inhibited ALDH1 activity in vivo in SUM159 xenografts. These results indicate that FoxQ1 is a novel target of bCSC inhibition by DATS.

SUBMITTER: Kim SH

PROVIDER: S-EPMC4919436 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

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Forkhead Box Q1 Is a Novel Target of Breast Cancer Stem Cell Inhibition by Diallyl Trisulfide.

Kim Su-Hyeong SH Kaschula Catherine H CH Priedigkeit Nolan N Lee Adrian V AV Singh Shivendra V SV

The Journal of biological chemistry 20160429 26

Diallyl trisulfide (DATS), a metabolic byproduct of garlic, is known to inhibit the growth of breast cancer cells in vitro and in vivo This study demonstrates that DATS targets breast cancer stem cells (bCSC). Exposure of MCF-7 and SUM159 human breast cancer cells to pharmacological concentrations of DATS (2.5 and 5 μm) resulted in dose-dependent inhibition of bCSC, as evidenced by a mammosphere assay and flow cytometric analysis of aldehyde dehydrogenase 1 (ALDH1) activity and the CD44(high)/CD ...[more]

PMID: 27129776

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Project description:Diallyl sulfide, diallyl disulfide, and daillyl trisulfide (DATS) are major volatile components of garlic oil. In this study, we assessed their relative potency in inducing antioxidant enzyme expression. Among the three organosulfur compounds, DATS was found to be most potent in inducing heme oxygenase-1 (HO-1) andNad(p)hquinone oxidoreductase-1 (NQO1) in human gastric epithelial (AGS) cells. Furthermore, DATS administration by gavage increased the expression of HO-1 and NQO1 in C57BL/6 mouse stomach. Treatment with DATS increased the accumulation of nuclear factor-erythroid-2-related factor-2 (Nrf2) in the nucleus of cultured AGS cells and in mouse stomach in vivo. The DATS-induced expression of HO-1 and NQO1 was abrogated in the cells transiently transfected with Nrf2-siRNA or in the embryonic fibroblasts from Nrf2-null mice, indicating that Nrf2 is a key mediator of the cytoprotective effects of DATS. Pretreatment of AGS cells with N-acetylcysteine or dithiothreitol attenuated DATS-induced nuclear localization of Nrf2 and the expression of HO-1 and NQO1. Cysteine-151, -273 and -288 of Kelch-like ECH-associated protein-1 (Keap1), a cytosolic repressor of Nrf2, have been considered to act as a redox sensor and play a role in Nrf2 activation. To determine whether DATS could inactivate Keap1 through thiol modification, we established cell lines constitutively expressing wild type-Keap1 or three different mutant constructs in which cysteine-151, -273, or -288 of Keap1 was replaced with serine by retroviral gene transfer. DATS failed to activate Nrf2, and to induce expression of HO-1 and NQO1 only in Keap1-C288S mutant cells. LC-ESI-MS/MS analysis of recombinant Keap1 treated with DATS revealed that the peptide fragment containing Cys288 gained a molecular mass of 72.1 Da equivalent to the molecular weight of mono-allyl mono-sulfide. Taken together, these findings suggest that DATS may directly interact with the Cys288 residue of Keap1, which partly accounts for its ability to induce Nrf2 activation and upregulate defensive gene expression.

Project description:BackgroundColorectal cancer (CRC) is an extremely malignant tumor with a high mortality rate. Little is known about the mechanism by which forkhead Box q1 (FOXQ1) causes CRC invasion and metastasis through the epidermal growth factor receptor (EGFR) pathway.AimTo illuminate the mechanism by which FOXQ1 promotes the invasion and metastasis of CRC by activating the heparin binding epidermal growth factor (HB-EGF)/EGFR pathway.MethodsWe investigated the differential expression and prognosis of FOXQ1 and HB-EGF in CRC using the Gene Expression Profiling Interactive Analysis (GEPIA) website (http://gepia.cancer-pku.cn/index.html). Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used to detect the expression of FOXQ1 and HB-EGF in cell lines and tissues, and we constructed a stable low-expressing FOXQ1 cell line and verified it with the above method. The expression changes of membrane-bound HB-EGF (proHB-EGF) and soluble HB-EGF (sHB-EGF) in the low-expressing FOXQ1 cell line were detected by flow cytometry and ELISA. Western blotting was used to detect changes in the expression levels of HB-EGF and EGFR pathway-related downstream genes when exogenous recombinant human HB-EGF was added to FOXQ1 knockdown cells. Proliferation experiments, transwell migration experiments, and scratch experiments were carried out to determine the mechanism by which FOXQ1 activates the EGFR signaling pathway through HB-EGF, and then to evaluate the clinical relevance of FOXQ1 and HB-EGF.ResultsGEPIA showed that the expression of FOXQ1 in CRC tissues was relatively high and was related to a lower overall survival rate. PCR array results showed that FOXQ1 is related to the HB-EGF and EGFR pathways. Knockdown of FOXQ1 suppressed the expression of HB-EGF, and led to a decrease in EGFR and its downstream genes AKT, RAF, KRAS expression levels. After knockdown of FOXQ1 in CRC cell lines, cell proliferation, migration and invasion were attenuated. Adding HB-EGF restored the migration and invasion ability of CRC, but not the cell proliferation ability. Kaplan-Meier survival analysis results showed that the combination of FOXQ1 and HB-EGF may serve to predict CRC survival.ConclusionBased on these collective data, we propose that FOXQ1 promotes the invasion and metastasis of CRC via the HB-EGF/EGFR pathway.

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Forkhead Box Q1 Is a Novel Target of Breast Cancer Stem Cell Inhibition by Diallyl Trisulfide.

Publications

Forkhead Box Q1 Is a Novel Target of Breast Cancer Stem Cell Inhibition by Diallyl Trisulfide.

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