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Phase-II metabolism limits the antiproliferative activity of urolithins in human colon cancer cells

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Abstract

Purpose

Urolithins, gut microbiota metabolites derived from ellagic acid and ellagitannins, reach micromolar concentrations in the colon lumen where can have anti-inflammatory and anticancer effects. The antiproliferative activity of urolithins (Uro-A, Uro-B, Uro-C and Uro-D) and their most relevant in vivo glucuronides were evaluated in three human colon cancer cell lines (Caco-2, SW480 and HT-29).

Methods

Cell proliferation was evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide and Trypan blue exclusion assays. Cell cycle was evaluated by flow cytometry and urolithins metabolism by HPLC–MS/MS.

Results

Urolithins inhibited cell proliferation and cell cycle progression in a time- and dose-dependent manner and arrested the cells at S and G2/M phases, depending on the urolithin. Uro-A exerted the highest antiproliferative activity, followed by Uro-C, Uro-D and Uro-B. Unlike Caco-2 and SW480 cells, HT-29 cells partially overcame the effects after 48 h, which was related to the complete glucuronidation of urolithins. Uro-A or Uro-B glucuronides did not affect cell cycle and showed lower antiproliferative activity than their aglycone counterparts. Uro-A or Uro-B plus inhibitors of drug efflux ABC transporters partially prevented the glucuronidation of urolithins in HT-29 cells which became more sensitive.

Conclusions

Uro-A, Uro-B, Uro-C and Uro-D exerted different antiproliferative effects depending on the colon cancer cell line. We also report here, for the first time, the role of ABC transporters and Phase-II metabolism in HT-29 cells as a mechanism of cancer resistance against urolithins due to their conversion to glucuronide conjugates that exerted lower antiproliferative activity.

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Abbreviations

ABC:

ATP-binding cassette

ACN:

Acetonitrile

ATP:

Adenosine-5′-triphosphate

BCRP:

Breast cancer resistance protein

DMEM:

Dulbecco’s modified Eagle’s medium

DMSO:

Dimethyl sulfoxide

DNA:

Deoxyribonucleic acid

EDTA:

Ethylenediaminetetraacetic acid

ESI:

Electrospray interface

Glur:

Glucuronide

HPLC:

High-performance liquid chromatography

IT:

Ion trap

MDCKII:

Mardin–Darby canine kidney

MEM:

Minimal essential medium

MeOH:

Methanol

MRP:

Multidrug resistant protein

MS:

Mass spectrometry

MTT:

3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide

OH-:

Hydroxyl groups

PBS:

Phosphate-buffered saline

P-gp:

P-glycoprotein

RNA:

Ribonucleic acid

SD:

Standard deviation

TNF-α:

Tumor necrosis factor alpha

UGTs:

UDP-glucuronosyltransferases

Uro:

Urolithins

UV:

Ultraviolet

μM:

Micromolar

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Acknowledgments

This work was funded by the Projects CICYT AGL2011-22447 (MINECO, Spain), Consolider Ingenio 2010 (CSD2007-00063, Fun-C-Food), and Fundación Seneca de la Región de Murcia, Spain (Grupo de Excelencia GERM 06 04486 and 05556/PI/04).

Conflict of interest

Authors declare no conflict of interests.

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Authors and Affiliations

  1. Research Group on Quality, Safety and Bioactivity of Plant Foods, Department of Food Science and Technology, CEBAS-CSIC, P.O. Box 164, Campus de Espinardo, 30100, Murcia, Spain

    Antonio González-Sarrías, Juan Antonio Giménez-Bastida, María Ángeles Núñez-Sánchez, Mar Larrosa, María Teresa García-Conesa, Francisco A. Tomás-Barberán & Juan Carlos Espín

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  1. Antonio González-Sarrías
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  2. Juan Antonio Giménez-Bastida
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  5. María Teresa García-Conesa
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  6. Francisco A. Tomás-Barberán
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  7. Juan Carlos Espín
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Correspondence to Antonio González-Sarrías.

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González-Sarrías, A., Giménez-Bastida, J.A., Núñez-Sánchez, M.Á. et al. Phase-II metabolism limits the antiproliferative activity of urolithins in human colon cancer cells. Eur J Nutr 53, 853–864 (2014). https://doi.org/10.1007/s00394-013-0589-4

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  • DOI: https://doi.org/10.1007/s00394-013-0589-4

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