The role of polycomb group protein Bmi-1 and Notch4 in breast cancer stem cell inhibition by benzyl isothiocyanate
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We showed previously that garden cress constituent benzyl isothiocyanate (BITC) inhibits self-renewal of breast cancer stem cells (bCSC) in vitro and in vivo. The present study offers novel insights into the mechanism by which BITC inhibits bCSC. Flow cytometry and mammosphere assay were performed to quantify bCSC fraction. Protein expression was determined by western blotting. Apoptosis was assessed by flow cytometry using Annexin V-propidium iodide method. Cell migration was determined by Boyden chamber assay. BITC treatment resulted in a marked decrease in protein level of polycomb group protein B-lymphoma Moloney murine leukemia virus insertion region-1 (Bmi-1) in cultured human breast cancer cells (MCF-7, SUM159, MDA-MB-231, and MDA-MB-361) and MDA-MB-231 xenografts in vivo. Overexpression (MCF-7) or knockdown (SUM159, and MDA-MB-231) of Bmi-1 protein had no meaningful impact on the BITC’s ability to inhibit cell viability and cell migration and/or induce apoptosis. On the other hand, inhibition of bCSC markers (aldehyde dehydrogenase 1 activity and mammosphere frequency) resulting from BITC exposure was significantly altered by Bmi-1 overexpression and knockdown. BITC was previously shown to cause activation of Notch1, Notch2, and Notch4 in association with induction of γ-secretase complex component Nicastrin, which are also implicated in maintenance of cancer stemness. BITC-mediated inhibition of bCSC was augmented by knockdown of Notch4 and Nicastrin, but not by RNA interference of Notch1 or Notch2. The present study highlights important roles for Bmi-1 and Notch4 in BITC-mediated suppression of bCSC.
KeywordsBenzyl isothiocyanate Bmi-1 Notch4 bCSC
Aldehyde dehydrogenase 1
Breast cancer stem cells
B-lymphoma Moloney murine leukemia virus insertion region-1
Small hairpin RNA
Small interfering RNA
This study was supported by the National Cancer Institute, National Institutes of Health Grant RO1 CA129347-07. This study used the Flow Cytometry Facility supported in part by Cancer Center Support Grant P30 CA047904.
Conflict of interest
None of the authors has any conflict of interest.
- 1.Fisher B, Costantino JP, Wickerham DL, Redmond CK, Kavanah M, Cronin WM, Vogel V, Robidoux A, Dimitrov N, Atkins J, Daly M, Wieand S, Tan-Chiu E, Ford L, Wolmark N (1998) Tamoxifen for prevention of breast cancer: report of the National Surgical Adjuvant Breast and Bowel Project P-1 study. J Natl Cancer Inst 90(18):1371–1388CrossRefPubMedGoogle Scholar
- 2.Goss PE, Ingle JN, Ales-Martinez JE, Cheung AM, Chlebowski RT, Wactawski-Wende J, McTierman A, Robbins J, Johnson KC, Martin LW, Winquist E, Sarto GE, Garber JE, Fabian CJ, Pujol P, Maunsell E, Farmer P, Gelmon KA, Tu D, Richardson H, NCIC CTG MAP.3 Study Investigators (2011) Exemestane for breast-cancer prevention in postmenopausal women. N Eng J Med 364(25):2381–2391CrossRefGoogle Scholar
- 22.Xiao D, Srivastava SK, Lew KL, Zeng Y, Hershberger P, Johnson CS, Trump DL, Singh SV (2003) Allyl isothiocyanate, a constituent of cruciferous vegetables, inhibits proliferation of human prostate cancer cells by causing G2/M arrest and inducing apoptosis. Carcinogenesis 24(5):891–897CrossRefPubMedGoogle Scholar
- 25.Finn RS, Dering J, Conklin D, Kalous O, Cohen DJ, Desai AJ, Ginther C, Atefi M, Chen I, Fowst C, Los G, Slamon DJ (2009) PD0332991, a selective cyclin D1 kinase 4/6 inhibitor, preferentially inhibits proliferation and luminal estrogen receptor-positive human breast cancer cell lines in vitro. Breast Cancer Res 11(5):R77CrossRefPubMedCentralPubMedGoogle Scholar
- 32.Lombardo Y, Filipović A, Molyneux G, Periyasamy M, Giamas G, Hu Y, Trivedi PS, Wang J, Yagüe E, Michel L, Coombes RC (2012) Nicastrin regulates breast cancer stem cell properties and tumor growth in vitro and in vivo. Proc Natl Acad Sci USA 109(41):16558–16563CrossRefPubMedCentralPubMedGoogle Scholar
- 36.Rustighi A, Zannini A, Tiberi L, Sommaggio R, Piazza S, Sorrentino G, Nuzzo S, Tuscano A, Eterno V, Benvenuti F, Santarpia L, Aifantis I, Rosato A, Bicciato S, Zambelli A, Del Sal G (2014) Prolyl-isomerase Pin1 controls normal and cancer stem cells of the breast. EMBO Mol Med 6(1):99–119CrossRefPubMedCentralPubMedGoogle Scholar