Pro-oxidant and pro-apoptotic activity of polyphenol extract from Annurca apple and its underlying mechanisms in human breast cancer cells

D'Angelo,Stefania; Martino,Elisa; Ilisso,Concetta  Paola; Bagarolo,Maria  Libera; Porcelli,Marina; Cacciapuoti,Giovanna

doi:10.3892/ijo.2017.4088

Pro-oxidant and pro-apoptotic activity of polyphenol extract from Annurca apple and its underlying mechanisms in human breast cancer cells

Authors:
- Stefania D'Angelo
- Elisa Martino
- Concetta Paola Ilisso
- Maria Libera Bagarolo
- Marina Porcelli
- Giovanna Cacciapuoti
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Affiliations: Department of Motor Sciences and Wellness, Parthenope University, Naples, Italy, Department of Biochemistry, Biophysics, and General Pathology, University of Campania ‘Luigi Vanvitelli’, Naples, Italy
Published online on: July 31, 2017 https://doi.org/10.3892/ijo.2017.4088
Pages: 939-948

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Abstract

Among nutraceuticals, polyphenols represent the most intriguing and studied class of compounds that can be therapeutics for a large spectrum of the most common diseases, including cancer. Although polyphenols are well known as potent antioxidants, a pro-oxidant effect has been associated with a pro-apoptotic function of these compounds in various types of tumor cells. Annurca apple, a southern Italian variety, is characterized by an extremely high content of polyphenols and displays a stronger antioxidant activity compared with other varieties. In the present study we explored the antiproliferative effect of Annurca apple polyphenol extract (APE) in human breast cancer MCF-7 cells and we investigated the underlying mechanisms. Results showed that at 500 µM catechin equivalent (EqC) APE acts as a pro-oxidant increasing thiobarbituric acid-reactive species cell content of approximately 6-fold more than the untreated cells. We found that APE strongly inhibits the proliferation of MCF-7 cells by inducing G2/M cell cycle arrest and apoptosis. Immunoblot analysis demonstrated that APE treatment increases the levels of p53 and p21, downregulates the expression of the cell cycle regulatory protein cyclin D1, and inhibits ERK1/2 phosphorylation. Moreover, APE treatment caused a marked increase of pro-apoptotic Bax/Bcl-2 ratio paralleled by caspase-9, -6, -7, and poly(ADP ribose) polymerase cleavage. Altogether our data indicate that APE, at elevated concentrations, acts as a potent pro-oxidant and antiproliferative agent able to downregulate ERK1/2 pathway leading to cell cycle inhibition and apoptosis and provides a rationale for its potential use in the development of novel therapeutics towards breast cancer.

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1	Reczek CR and Chandel NS: The two faces of reactive oxygen species in cancer. Annu Rev Cancer Biol. 1:79–98. 2017. View Article : Google Scholar
2	Wang P, Yang HL, Yang YJ, Wang L and Lee SC: Overcome cancer cell drug resistance using natural products. Evid Based Complement Alternat Med. 2015:7671362015. View Article : Google Scholar : PubMed/NCBI
3	Li L and Leung PS: Use of herbal medicines and natural products: An alternative approach to overcoming the apoptotic resistance of pancreatic cancer. Int J Biochem Cell Biol. 53:224–236. 2014. View Article : Google Scholar : PubMed/NCBI
4	Hemaiswarya S and Doble M: Potential synergism of natural products in the treatment of cancer. Phytother Res. 20:239–249. 2006. View Article : Google Scholar : PubMed/NCBI
5	León-González AJ, Auger C and Schini-Kerth VB: Pro-oxidant activity of polyphenols and its implication on cancer chemo-prevention and chemotherapy. Biochem Pharmacol. 98:371–380. 2015. View Article : Google Scholar
6	Rengarajan T and Yaacob NS: The flavonoid fisetin as an anticancer agent targeting the growth signaling pathways. Eur J Pharmacol. 789:8–16. 2016. View Article : Google Scholar : PubMed/NCBI
7	Batra P and Sharma AK: Anti-cancer potential of flavonoids: Recent trends and future perspectives. 3 Biotech. 3:439–459. 2013. View Article : Google Scholar : PubMed/NCBI
8	Khan HY, Zubair H, Ullah MF, Ahmad A and Hadi SM: A prooxidant mechanism for the anticancer and chemopreventive properties of plant polyphenols. Curr Drug Targets. 13:1738–1749. 2012. View Article : Google Scholar : PubMed/NCBI
9	Crozier A, Jaganath IB and Clifford MN: Dietary phenolics: Chemistry, bioavailability and effects on health. Nat Prod Rep. 26:1001–1043. 2009. View Article : Google Scholar : PubMed/NCBI
10	Vauzour D, Rodriguez-Mateos A, Corona G, Oruna-Concha MJ and Spencer JP: Polyphenols and human health: Prevention of disease and mechanisms of action. Nutrients. 2:1106–1131. 2010. View Article : Google Scholar : PubMed/NCBI
11	Boyer J and Liu RH: Apple phytochemicals and their health benefits. Nutr J. 3:52004. View Article : Google Scholar : PubMed/NCBI
12	Kern M, Tjaden Z, Ngiewih Y, Puppel N, Will F, Dietrich H, Pahlke G and Marko D: Inhibitors of the epidermal growth factor receptor in apple juice extract. Mol Nutr Food Res. 49:317–328. 2005. View Article : Google Scholar : PubMed/NCBI
13	Sun J, Chu YF, Wu X and Liu RH: Antioxidant and anti-proliferative activities of common fruits. J Agric Food Chem. 50:7449–7454. 2002. View Article : Google Scholar : PubMed/NCBI
14	Tow WW, Premier R, Jing H and Ajlouni S: Antioxidant and antiproliferation effects of extractable and nonextractable polyphenols isolated from apple waste using different extraction methods. J Food Sci. 76:T163–T172. 2011. View Article : Google Scholar
15	Lo Scalzo R, Testoni A and Genna A: 'Annurca' apple fruit, a southern Italy apple cultivar: Textural properties and aroma composition. Food Chem. 73:333–343. 2001. View Article : Google Scholar
16	Napolitano A, Cascone A, Graziani G, Ferracane R, Scalfi L, Di Vaio C, Ritieni A and Fogliano V: Influence of variety and storage on the polyphenol composition of apple flesh. J Agric Food Chem. 52:6526–6531. 2004. View Article : Google Scholar : PubMed/NCBI
17	Tenore GC, Campiglia P, Stiuso P, Ritieni A and Novellino E: Nutraceutical potential of polyphenolic fractions from Annurca apple (M. pumila Miller cv Annurca). Food Chem. 140:614–622. 2013. View Article : Google Scholar : PubMed/NCBI
18	Fini L, Piazzi G, Daoud Y, Selgrad M, Maegawa S, Garcia M, Fogliano V, Romano M, Graziani G, Vitaglione P, et al: Chemoprevention of intestinal polyps in Apc^Min/+ mice fed with western or balanced diets by drinking Annurca apple polyphenol extract. Cancer Prev Res (Phila). 4:907–915. 2011. View Article : Google Scholar
19	Fini L, Selgrad M, Fogliano V, Graziani G, Romano M, Hotchkiss E, Daoud YA, De Vol EB, Boland CR and Ricciardiello L: Annurca apple polyphenols have potent demethylating activity and can reactivate silenced tumor suppressor genes in colorectal cancer cells. J Nutr. 137:2622–2628. 2007.PubMed/NCBI
20	D'Angelo S, La Porta R, Napolitano M, Galletti P, Quagliuolo L and Boccellino M: Effect of Annurca apple polyphenols on human HaCaT keratinocytes proliferation. J Med Food. 15:1024–1031. 2012. View Article : Google Scholar : PubMed/NCBI
21	Wang S, Bai L, Lu J, Liu L, Yang CY and Sun H: Targeting inhibitors of apoptosis proteins (IAPs) for new breast cancer therapeutics. J Mammary Gland Biol Neoplasia. 17:217–228. 2012. View Article : Google Scholar : PubMed/NCBI
22	Dou QP: Molecular mechanisms of green tea polyphenols. Nutr Cancer. 61:827–835. 2009. View Article : Google Scholar
23	Singleton VL, Orthofer R and Lamuela-Raventos RM: Analysis of total phenols and other oxidation substrates and antioxidants by means of Folin-Ciocalteu reagent. Methods Enzymol. 299:152–178. 1999. View Article : Google Scholar
24	D'Angelo S, Cimmino A, Raimo M, Salvatore A, Zappia V and Galletti P: Effect of reddening-ripening on the antioxidant activity of polyphenol extracts from cv. 'Annurca' apple fruits. J Agric Food Chem. 55:9977–9985. 2007. View Article : Google Scholar : PubMed/NCBI
25	D'Angelo S and Sammartino D: Protective effect of Annurca apple extract against oxidative damage in human erythrocytes. Curr Nutr Food Sci. 11:248–256. 2015. View Article : Google Scholar
26	Vermes I, Haanen C, Steffens-Nakken H and Reutelingsperger C: A novel assay for apoptosis. Flow cytometric detection of phosphatidylserine expression on early apoptotic cells using fluorescein labelled Annexin V. J Immunol Methods. 184:39–51. 1995. View Article : Google Scholar : PubMed/NCBI
27	Gago-Dominguez M, Jiang X and Castelao JE: Lipid peroxidation, oxidative stress genes and dietary factors in breast cancer protection: A hypothesis. Breast Cancer Res. 9:201–211. 2007. View Article : Google Scholar : PubMed/NCBI
28	Fantini M, Benvenuto M, Masuelli L, Frajese GV, Tresoldi I, Modesti A and Bei R: In vitro and in vivo antitumoral effects of combinations of polyphenols, or polyphenols and anticancer drugs: Perspectives on cancer treatment. Int J Mol Sci. 16:9236–9282. 2015. View Article : Google Scholar : PubMed/NCBI
29	Min NY, Kim JH, Choi JH, Liang W, Ko YJ, Rhee S, Bang H, Ham SW, Park AJ and Lee KH: Selective death of cancer cells by preferential induction of reactive oxygen species in response to (−)-epigallocatechin-3-gallate. Biochem Biophys Res Commun. 421:91–97. 2012. View Article : Google Scholar : PubMed/NCBI
30	Casanova F, Quarti J, da Costa DC, Ramos CA, da Silva JL and Fialho E: Resveratrol chemosensitizes breast cancer cells to melphalan by cell cycle arrest. J Cell Biochem. 113:2586–2596. 2012. View Article : Google Scholar : PubMed/NCBI
31	Lee HP, Li TM, Tsao JY, Fong YC and Tang CH: Curcumin induces cell apoptosis in human chondrosarcoma through extrinsic death receptor pathway. Int Immunopharmacol. 13:163–169. 2012. View Article : Google Scholar : PubMed/NCBI
32	Etienne-Selloum N, Dandache I, Sharif T, Auger C and Schini-Kerth VB: Polyphenolic compounds targeting p53-family tumor suppressors: current progress and challenges. Future Aspects of Tumor Suppressor Gene. Cheng Y: InTech; pp. 129–166. 2013, https://doi.org/10.5772/56102.
33	Taylor WR and Stark GR: Regulation of the G2/M transition by p53. Oncogene. 20:1803–1815. 2001. View Article : Google Scholar : PubMed/NCBI
34	Zhang X, Min KW, Wimalasena J and Baek SJ: Cyclin D1 degradation and p21 induction contribute to growth inhibition of colorectal cancer cells induced by epigallocatechin-3-gallate. J Cancer Res Clin Oncol. 138:2051–2060. 2012. View Article : Google Scholar : PubMed/NCBI
35	Sherr CJ: D-type cyclins. Trends Biochem Sci. 20:187–190. 1995. View Article : Google Scholar : PubMed/NCBI
36	Yang K, Hitomi M and Stacey DW: Variations in cyclin D1 levels through the cell cycle determine the proliferative fate of a cell. Cell Div. 1:322006. View Article : Google Scholar : PubMed/NCBI
37	Alao JP: The regulation of cyclin D1 degradation: Roles in cancer development and the potential for therapeutic invention. Mol Cancer. 6:242007. View Article : Google Scholar : PubMed/NCBI
38	Cagnol S and Chambard JC: ERK and cell death: Mechanisms of ERK-induced cell death - apoptosis, autophagy and senescence. FEBS J. 277:2–21. 2010. View Article : Google Scholar
39	Chaparro M, González Moreno L, Trapero-Marugán M, Medina J and Moreno-Otero R: Review article: Pharmacological therapy for hepatocellular carcinoma with sorafenib and other oral agents. Aliment Pharmacol Ther. 28:1269–1277. 2008. View Article : Google Scholar : PubMed/NCBI
40	Briviba K, Pan L and Rechkemmer G: Red wine polyphenols inhibit the growth of colon carcinoma cells and modulate the activation pattern of mitogen-activated protein kinases. J Nutr. 132:2814–2818. 2002.PubMed/NCBI
41	Granado-Serrano AB, Martín MA, Bravo L, Goya L and Ramos S: Quercetin induces apoptosis via caspase activation, regulation of Bcl-2, and inhibition of PI-3-kinase/Akt and ERK pathways in a human hepatoma cell line (HepG2). J Nutr. 136:2715–2721. 2006.PubMed/NCBI
42	Hardwick JM and Soane L: Multiple functions of BCL-2 family proteins. Cold Spring Harb Perspect Biol. 5:pii: a008722. 2013. View Article : Google Scholar : PubMed/NCBI
43	Babich H, Schuck AG, Weisburg JH and Zuckerbraun HL: Research strategies in the study of the pro-oxidant nature of polyphenol nutraceuticals. J Toxicol. 2011:4673052011. View Article : Google Scholar : PubMed/NCBI
44	Trachootham D, Lu W, Ogasawara MA, Nilsa RD and Huang P: Redox regulation of cell survival. Antioxid Redox Signal. 10:1343–1374. 2008. View Article : Google Scholar : PubMed/NCBI
45	Burch PM and Heintz NH: Redox regulation of cell-cycle re-entry: Cyclin D1 as a primary target for the mitogenic effects of reactive oxygen and nitrogen species. Antioxid Redox Signal. 7:741–751. 2005. View Article : Google Scholar : PubMed/NCBI
46	Fasanaro P, Magenta A, Zaccagnini G, Cicchillitti L, Fucile S, Eusebi F, Biglioli P, Capogrossi MC and Martelli F: Cyclin D1 degradation enhances endothelial cell survival upon oxidative stress. FASEB J. 20:1242–1244. 2006. View Article : Google Scholar : PubMed/NCBI
47	Budanov AV: The role of tumor suppressor p53 in the antioxidant defense and metabolism. Subcell Biochem. 85:337–358. 2014. View Article : Google Scholar : PubMed/NCBI
48	Wu GS: The functional interactions between the p53 and MAPK signaling pathways. Cancer Biol Ther. 3:156–161. 2004. View Article : Google Scholar : PubMed/NCBI