Cytotoxicity of pomegranate polyphenolics in breast cancer cells in vitro and vivo: potential role of miRNA-27a and miRNA-155 in cell survival and inflammation
- 1.2k Downloads
Several studies have demonstrated that polyphenolics from pomegranate (Punica granatum L.) are potent inhibitors of cancer cell proliferation and induce apoptosis, cell cycle arrest, and also decrease inflammation in vitro and vivo. There is growing evidence that botanicals exert their cytotoxic and anti-inflammatory activities, at least in part, by decreasing specificity protein (Sp) transcription factors. These are overexpressed in breast tumors and regulate genes important for cancer cell survival and inflammation such as the p65 unit of NF-κB. Moreover, previous studies have shown that Pg extracts decrease inflammation in lung cancer cell lines by inhibiting phosphatidylinositol-3,4,5-trisphosphate (PI3K)-dependent phosphorylation of AKT in vitro and inhibiting the activation of NF-kB in vivo. The objective of this study was to investigate the roles of miR-27a–ZBTB10–Sp and miR-155–SHIP-1–PI3K on the anti-inflammatory and cytotoxic activity of pomegranate extract. Pg extract (2.5-50 μg/ml) inhibited growth of BT-474 and MDA-MB-231 cells but not the non-cancer MCF-10F and MCF-12F cells. Pg extract significantly decreased Sp1, Sp3, and Sp4 as well as miR-27a in BT474 and MDA-MB-231 cells and increased expression of the transcriptional repressor ZBTB10. A significant decrease in Sp proteins and Sp-regulated genes was also observed. Pg extract also induced SHIP-1 expression and this was accompanied by downregulation of miRNA-155 and inhibition of PI3K-dependent phosphorylation of AKT. Similar results were observed in tumors from nude mice bearing BT474 cells as xenografts and treated with Pg extract. The effects of antagomirs and knockdown of SHIP-1 by RNA interference confirmed that the anti-inflammatory and cytotoxic effects of Pg extract were partly due to the disruption of both miR-27a–ZBTB10 and miR-155–SHIP-1. In summary, the anticancer activities of Pg extract in breast cancer cells were due in part to targeting microRNAs155 and 27a. Both pathways play an important role in the proliferative/inflammatory phenotype exhibited by these cell lines.
KeywordsBreast cancer Polyphenolics Pomegranate Xenografts Inflammation Cytotoxicity
We would like to thank Dr. Weston Porter Department Veterinary Integrated Bioscience, at Texas A&M University, College Station, and Texas for providing imaging equipment. Lastly we would like to thank Stefan Wypyszyk at Stiebs LLC (Kirkland, WA) for kindly supplying the pomegranate juice. Financial support for this research has been provided by the National Institutes of Health (KOIATOO 4597 to SMT).
Conflict of interest
The authors have no conflicts of interest to declare.
- 1.Prakobwong SG, Gupta SC, Kim JH, Sung B, Pinlaor P, Hiraku Y, Wongkham S, Sripa B, Pinlaor S, Aggarwal BB (2011) Curcumin suppresses proliferation and induces apoptosis in human biliary cancer cells through modulation of multiple cell signaling pathways. Carcinogenesis 32:1372–1380PubMedCrossRefGoogle Scholar
- 2.Kim ND, Mehta R, Yu W, Neeman I, Livney T, Amichay A, Poirier D, Nicholls P, Kirby A, Jiang W, Mansel R, Ramachandran C, Rabi T, Kaplan B, Lansky E (2002) Chemopreventive and adjuvant therapeutic potential of pomegranate (Punica granatum) for human breast cancer. Breast Cancer Res Treat 71:203–217PubMedCrossRefGoogle Scholar
- 15.Noratto GD, Kim Y, Talcott ST, Mertens-Talcott SU (2011) Flavonol-rich fractions of yaupon holly leaves (Ilex vomitoria, Aquifoliaceae) induce microRNA-146a and have anti-inflammatory and chemopreventive effects in intestinal myofibroblast CCD-18Co cells. Fitoterapia 82:557–569PubMedCrossRefGoogle Scholar
- 20.Seeram NP, Adams LS, Henning SM, Niu Y, Zhang Y, Nair MG, Heber D (2005) In vitro antiproliferative, apoptotic and antioxidant activities of punicalagin, ellagic acid and a total pomegranate tannin extract are enhanced in combination with other polyphenols as found in pomegranate juice. J Nutr Biochem 16:360–367PubMedCrossRefGoogle Scholar
- 26.Kumar AP, Bhaskaran S, Ganapathy M, Crosby K, Davis MD, Kochunov P, Schoolfield J, Yeh IT, Troyer DA, Ghosh R (2007) Akt/cAMP-responsive element binding protein/cyclin D1 network: a novel target for prostate cancer inhibition in transgenic adenocarcinoma of mouse prostate model mediated by Nexrutine, a Phellodendron amurense bark extract. Clin Cancer Res 13:2784–2794PubMedCrossRefGoogle Scholar
- 28.Yao JC, Wang L, Wei D, Gong W, Hassan M, Wu TT, Mansfield P, Ajani J, Xie K (2004) Association between expression of transcription factor Sp1 and increased vascular endothelial growth factor expression, advanced stage, and poor survival in patients with resected gastric cancer. Clin Cancer Res 10:4109–4117PubMedCrossRefGoogle Scholar
- 31.Mertens-Talcott SU, Noratto G, Li X, Angel-Morales G, Bertoldi MC, Safe S (2012) Betulinic acid decreases ER-negative breast cancer cell growth in vitro and in vivo: role of sp transcription factors and MicroRNA-27a:ZBTB10. Mol Carcinog 10. doi: 10.1002/mc.21893
- 33.Chintharlapalli S, Papineni S, Abdelrahim M, Abudayyeh A, Jutooru I, Chadalapaka G, Wu F, Mertens-Talcott S, Vanderlaag K, Cho SD et al (2009) Oncogenic microRNA-27a is a target for anticancer agent methyl 2-cyano-3,11-dioxo-18 beta-olean-1,12-dien-30-oate in colon cancer cell. Int J Cancer 125:1965–1974PubMedCrossRefGoogle Scholar
- 40.Boesch-Saadatmandia C, Loboda A, Wagnera AE, Stachurskab A, Jozkowiczb A, Dulakb J, Döringa F, Wolfframc S, Rimbacha G (2011) Effect of quercetin and its metabolites isorhamnetin and quercetin-3-glucuronide on inflammatory gene expression: role of miR-155. J Nutr Biochem 22:293–299CrossRefGoogle Scholar
- 48.Wang V, Wu W (2007) MicroRNA: a new player in breast cancer development. J Cancer Mol 3:133–138Google Scholar
- 53.Ozbaya T, Nahta R (2011) Delphinidin inhibits HER2 and Erk1/2 signaling and suppresses growth of HER2-overexpressing and triple negative breast cancer cell lines. Breast Cancer 5:143–154Google Scholar
- 54.Pandey PR, Okuda H, Watabe M, Pai SK, Liu W, Kobayashi A, Xing F, Fukuda K, Hirota S, Sugai T, Wakabayashi G, Koeda K, Kashiwaba M, Suzuki K, Chiba T, Endo M, Fujioka T, Tanji S, Mo YY, Cao D, Wilber AC, Watabe K (2011) Resveratrol suppresses growth of cancer stem-like cells by inhibiting fatty acid synthase. Breast Cancer Res Treat 130:387–398PubMedCrossRefGoogle Scholar
- 57.Jutooru I, Chadalapaka G, Abdelrahim M, Basha MR, Samudio I, Konopleva M, Andreeff M, Safe S (2010) Methyl 2-cyano-3,12-dioxooleana-1,9-dien-28-oate decreases specificity protein transcription factors and inhibits pancreatic tumor growth: role of MicroRNA-27a. Mol Pharmacol 78:226–236PubMedCrossRefGoogle Scholar
- 58.Wang Y, Keogh RJ, Hunter MG, Mitchell CA, Frey RS, Javaid K, Malik AB, Schurmans S, Tridandapani S, Marsh CB (2004) SHIP2 is recruited to the cell membrane upon macrophage colony-stimulating factor (M-CSF) Stimulation and regulates M-CSF-induced signaling. J Immunol 173:6820–6830PubMedGoogle Scholar