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CYP1A1 is overexpressed upon incubation of breast cancer cells with a polyphenolic cocoa extract

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Abstract

Purpose

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.

Methods

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.

Results

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.

Conclusions

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.

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Abbreviations

AhR:

Aryl hydrocarbon receptor,

Arnt:

Aryl hydrocarbon receptor nuclear translocator

BSA:

Bovine serum albumin

CHX:

Cycloheximide

CI:

Combination index

CYP1A1:

Cytochrome P450 family 1 subfamily A polypeptide 1

DEPC:

Diethyl pyrocarbonate

EGCG:

Epigallocatechin-3-gallate

EMSA:

Electrophoretic mobility shift assay

ER:

Estrogen receptor

GS:

Gel shift

NE:

Nuclear extract

OVN:

Overnight

PCE:

Polyphenolic cocoa extract

PI:

Propidium iodide

RT-PCR:

Reverse transcription-polymerase chain reaction

SE:

Standard error

STP:

Staurosporine

TAM:

Tamoxifen

XRE:

Xenobiotic response element

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Acknowledgments

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.

Author disclosure

C. Oleaga, M. García, A. Solé, C.J. Ciudad, M. Izquierdo-Pulido, and V. Noé have no conflicts of interest.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, School of Pharmacy, University of Barcelona, Av. Diagonal 643, 08028, Barcelona, Spain

    Carlota Oleaga, Miriam García, Anna Solé, Carlos J. Ciudad & Véronique Noé

  2. Department of Nutrition and Food Science, School of Pharmacy, University of Barcelona, Av. Diagonal 643, 08028, Barcelona, Spain

    Maria Izquierdo-Pulido

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  1. Carlota Oleaga
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  2. Miriam García
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  3. Anna Solé
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  4. Carlos J. Ciudad
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  6. Véronique Noé
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Correspondence to Véronique Noé.

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Oleaga, C., García, M., Solé, A. et al. CYP1A1 is overexpressed upon incubation of breast cancer cells with a polyphenolic cocoa extract. Eur J Nutr 51, 465–476 (2012). https://doi.org/10.1007/s00394-011-0231-2

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