CYP1A1 is overexpressed upon incubation of breast cancer cells with a polyphenolic cocoa extract
- 578 Downloads
To evaluate the effect of cocoa flavonoids in breast cancer cells at the molecular level, a functional genomic analysis was performed using a polyphenolic cocoa extract (PCE) in MCF-7 and SKBR3 cell lines.
The expression profile of 84 genes included in the Stress & Toxicity PathwayFinder™ PCR Array was analyzed after PCE incubation for 24 h. mRNA and protein levels were analyzed by RT-PCR and western blot, respectively. Gel shift assays were used to evaluate DNA–protein complexes. Protein complexes were identified by co-immunoprecipitation. Cell viability was evaluated by MTT assays.
Upon PCE incubation, 7 genes were overexpressed and 1 underexpressed in MCF-7 cells, whereas 9 genes were overexpressed in SKBR3 cells. Among the differentially expressed genes in both cell lines, cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) was chosen for further study. CYP1A1 mRNA and protein levels and enzymatic activity increased upon PCE incubation. CYP1A1 transcriptional activation by PCE was mediated through AhR binding to XRE elements within the CYP1A1 promoter in MCF-7 cells. A protein complex including AhR and ERα was detected. The combination of PCE with tamoxifen caused a synergistic cytotoxicity in both cell lines and was due to an increase in apoptosis in MCF-7 cells.
The interaction between ERα and AhR upon incubation with PCE leads to CYP1A1 induction in breast cancer cells. The synergy between PCE and non-cytotoxic tamoxifen concentrations opens the possibility for a combination therapy based on polyphenols from cocoa that increased tamoxifen efficacy.
KeywordsAhR Cocoa CYP1A1 ERα Tamoxifen Breast cancer
Aryl hydrocarbon receptor,
Aryl hydrocarbon receptor nuclear translocator
Bovine serum albumin
Cytochrome P450 family 1 subfamily A polypeptide 1
Electrophoretic mobility shift assay
Polyphenolic cocoa extract
Reverse transcription-polymerase chain reaction
Xenobiotic response element
This work was supported by grants by Nutrexpa SA (CDTI 050618), SAF08-0043 (Ministerio de Educación y Ciencia de España), and ISCIII-RTICc RD06/0020 (Redes Temáticas de Investigación Cooperativa en Salud) RD06/0020/0046. Our research group holds the “quality distinction” from the “Generalitat de Catalunya” SGR2009-00118. CO. was a recipient of a fellowship from the FEC (Federación Española del Café). The authors wish to thank to Dr. Lamuela-Raventós for the analysis of the polyphenol contents in the cocoa samples and Dr. Cascante for her help in the analyses of synergism by statistical methods.
C. Oleaga, M. García, A. Solé, C.J. Ciudad, M. Izquierdo-Pulido, and V. Noé have no conflicts of interest.
- 1.Ferlay J, Bray F, Pisani P et al (2004) GLOBOCAN 2002: cancer incidence, mortality and prevalence worldwide. IARC CancerBase no. 5, Version 2.0. International Agency for Research on Cancer, LyonGoogle Scholar
- 7.Cavalieri E, Chakravarti D, Guttenplan J, Hart E, Ingle J, Jankowiak R, Muti P, Rogan E, Russo J, Santen R, Sutter T (2006) Catechol estrogen quinones as initiators of breast and other human cancers: implications for biomarkers of susceptibility and cancer prevention. Biochim Biophys Acta 1766:63–78Google Scholar
- 11.Rutqvist LE, Cedermark B, Fornander T, Glas U, Johansson H, Nordenskjold B, Rotstein S, Skoog L, Somell A, Theve T (1989) The relationship between hormone receptor content and the effect of adjuvant tamoxifen in operable breast cancer. J Clin Oncol 7:1474–1484Google Scholar
- 16.Gupta S, Ahmad N, Nieminen AL, Mukhtar H (2000) Growth inhibition, cell-cycle dysregulation, and induction of apoptosis by green tea constituent (-)-epigallocatechin-3-gallate in androgen-sensitive and androgen-insensitive human prostate carcinoma cells. Toxicol Appl Pharmacol 164:82–90CrossRefGoogle Scholar
- 22.Andrés-Lacueva C, Lamuela-Raventós RM, Jauregui O, Casals I, Izquierdo-Pulido M, Permanyer J (2000) An LC method for the analysis of cocoa phenolics. LCoGC Eur 13:902–904Google Scholar
- 23.Swain T, Hillis WE (1969) The total phenolic constituents of Prunus domestica. J Sci Food Agric 10:4292–4296Google Scholar
- 26.Tapias A, Ciudad CJ, Noe V (2008) Transcriptional regulation of the 5′-flanking region of the human transcription factor Sp3 gene by NF-1, c-Myb, B-Myb, AP-1 and E2F. Biochim Biophys Acta 1779:318–329Google Scholar
- 30.Blasco F, Penuelas S, Cascallo M, Hernandez JL, Alemany C, Masa M, Calbo J, Soler M, Nicolas M, Perez-Torras S, Gomez A, Tarrason G, Noe V, Mazo A, Ciudad CJ, Piulats J (2004) Expression profiles of a human pancreatic cancer cell line upon induction of apoptosis search for modulators in cancer therapy. Oncology 67:277–290CrossRefGoogle Scholar
- 33.Trapani V, Patel V, Leong CO, Ciolino HP, Yeh GC, Hose C, Trepel JB, Stevens MF, Sausville EA, Loaiza-Perez AI (2003) DNA damage and cell cycle arrest induced by 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203, NSC 703786) is attenuated in aryl hydrocarbon receptor deficient MCF-7 cells. Br J Cancer 88:599–605CrossRefGoogle Scholar
- 36.Stuart EC, Larsen L, Rosengren RJ (2007) Potential mechanisms for the synergistic cytotoxicity elicited by 4-hydroxytamoxifen and epigallocatechin gallate in MDA-MB-231 cells. Int J Oncol 30:1407–1412Google Scholar
- 39.Huang Z, Fasco MJ, Figge HL, Keyomarsi K, Kaminsky LS (1996) Expression of cytochromes P450 in human breast tissue and tumors. Drug Metab Dispos 24:899–905Google Scholar
- 47.Yoshida T, Maeda A, Horinaka M, Shiraishi T, Nakata S, Wakada M, Yogosawa S, Sakai T (2005) Quercetin induces gadd45 expression through a p53-independent pathway. Oncol Rep 14:1299–1303Google Scholar
- 49.Schlittenhardt D, Schober A, Strelau J, Bonaterra GA, Schmiedt W, Unsicker K, Metz J, Kinscherf R (2004) Involvement of growth differentiation factor-15/macrophage inhibitory cytokine-1 (GDF-15/MIC-1) in oxLDL-induced apoptosis of human macrophages in vitro and in arteriosclerotic lesions. Cell Tissue Res 318:325–333CrossRefGoogle Scholar
- 50.Golkar L, Ding XZ, Ujiki MB, Salabat MR, Kelly DL, Scholtens D, Fought AJ, Bentrem DJ, Talamonti MS, Bell RH, Adrian TE (2007) Resveratrol inhibits pancreatic cancer cell proliferation through transcriptional induction of macrophage inhibitory cytokine-1. J Surg Res 138:163–169CrossRefGoogle Scholar
- 53.Katz DL, Doughty K, Ali A (2011) Cocoa and chocolate in human health and disease. Antioxid Redox Signal. doi: 10.1089/ars.2010.3697
- 55.Fletcher N, Wahlstrom D, Lundberg R, Nilsson CB, Nilsson KC, Stockling K, Hellmold H, Hakansson H (2005) 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) alters the mRNA expression of critical genes associated with cholesterol metabolism, bile acid biosynthesis, and bile transport in rat liver: a microarray study. Toxicol Appl Pharmacol 207:1–24CrossRefGoogle Scholar
- 56.Ferlini C, Scambia G, Distefano M, Filippini P, Isola G, Riva A, Bombardelli E, Fattorossi A, Benedetti Panici P, Mancuso S (1997) Synergistic antiproliferative activity of tamoxifen and docetaxel on three oestrogen receptor-negative cancer cell lines is mediated by the induction of apoptosis. Br J Cancer 75:884–891CrossRefGoogle Scholar
- 57.Shen F, Xue X, Weber G (1999) Tamoxifen and genistein synergistically down-regulate signal transduction and proliferation in estrogen receptor-negative human breast carcinoma MDA-MB-435 cells. Anticancer Res 19:1657–1662Google Scholar
- 58.McDougal A, Wormke M, Calvin J, Safe S (2001) Tamoxifen-induced antitumorigenic/antiestrogenic action synergized by a selective aryl hydrocarbon receptor modulator. Cancer Res 61:3902–3907Google Scholar