Cancer Letters

Cancer Letters

Volume 230, Issue 2, 18 December 2005, Pages 239-247
Cancer Letters

Chemopreventive properties of the ethanol extract of chinese licorice (Glycyrrhiza uralensis) root: induction of apoptosis and G1 cell cycle arrest in MCF-7 human breast cancer cells

https://doi.org/10.1016/j.canlet.2004.12.038Get rights and content

Abstract

Much of the interest on the chemopreventive properties of licorice has been focused on the plant genius Glycyrrhiza glabra. In this study the ethanol extract of Chinese licorice root, Glycyrrhiza uralensis (G. uralensis) was investigated for its estrogenic effect and the ability to inhibit cell proliferation in the MCF-7 human breast cancer cell line. The extract of the root of G. uralensis was fractionated in EtOH:H2O (80:20) (80% ethanol). The extract exhibited estrogenic effects similar to 17β- estradiol (E2) and induced apoptosis at the same dose level (100 μg/ml) in MCF-7 breast cancer cells, results were associated with up-regulation of tumor suppressor gene p53 and pro-apoptotic protein Bax. G. uralensis extract caused the up-regulation of p21waf1/cip1 and down-regulation of cdk 2 and cyclin E and most significantly, induced G1 cell cycle arrest. This is the first study to show that the ethanolic extract of the root of G. uralensis has an estrogen-like activity and anti-cancer effects against MCF-7 human breast cancer cells. Whilst the use of phytoestrogens to protect against hormone-dependent cancers or as a ‘natural’ alternative to hormone replacement therapy remains controversial, the data in this paper support the suggestion that extracts of root of the Chinese licorice G. uralensis might be of importance in this debate.

Introduction

The incidence of cancers in particular those of the breast and prostate continue to be focus of worldwide attention. Breast cancer is the most common form of cancer affecting women with cases in North America and Northern Europe being considerably higher than in Asia, the Far East, Africa and South America. Hulka and Moorman [1] have noted that ‘the international differences in breast-cancer incidence and mortality rates indicate important differences in the endogenous hormonal milieu, lifestyle and environmental factors, genetic susceptibility, and mammographic screening activities among countries’. Breast cancer incidence and mortality rates may be explained by differences in the relative risk or prevalence of risk factors including dietary factors but this remain inconclusive. Indeed the extent to which diet is involved in the etiology of breast cancer as opposed to its mediation in part through hormonal mechanisms is being borne out by several studies [2], [3], [4] has yet to be fully defined. Although dietary phytoestrogens in edible plants including lignans and isoflavonoids, used in a wide variety of products, can mimic the effects of the main natural estrogen, 17β-estradiol, by binding to the estrogen receptor and influencing the expression of estrogen-dependent genes, the effect of the intake of phytoestrogens in decreasing breast cancer risk remain controversial [3], [4]. Nevertheless research in this area is attractive. Most studies on phytoestrogens and breast cancer have addressed the effect of phytoestrogens on the growth of breast tumor cells was biphasic [5], [6], [7].

Licorice root is also used as flavoring and sweetening agents for tobaccos, chewing gums, candies, toothpaste and beverages [8]. The genius of licorice widely discussed in the western part of the world is the Glycyrrhiza glabra which is indigenous to Turkey, Spain, Iraq, Turkey, Russia and North China. Glycyrrhiza uralensis is indigenous to Northern China, Mongolia and Siberia. The content of G. uralensis is totally different from that of G. glabra in result of HPLC profiles [9]. Most work has been concentrated on the active components and extracts of G. glabra in terms of chemoprevention activities [10], [11], [12], [13]. Recent research has focused on the activities of extracts of the roots of Chinese licorice G. uralensis. The CHCl3, EtOAc, C6H14 and CH3OH–H2O (70:30) extracts of G. uralensis induced apoptosis in a dose- and time dependent manner by possibly by mediating cleavage of PARP, up-regulation of Bax and cleavage of Bcl-2 in MCF-7 human breast cancer cells [14].

Licochalcone-A of licorice root of G. glabra induced apoptosis in MCF-7 and HL-60 cell lines, as demonstrated by PARP cleavage, the substrate of ICE-like protease [10]. Isoliquiritigenin of licorice root of G. glabra is widely suggested as a candidate agent for the treatment of prostate cancer [11]. The DNA damage response gene GADD153 may play an important role in isoliquiritigenin-induced cell cycle arrest and cell growth inhibition [11]. Tamir et al. [12] studied the effect of glabridin from G. glabra on the growth of breast tumor cells and found that there was a biphasic. Glabridin showed an estrogen receptor-dependent, growth-promoting effect at low concentration (10 nM–10 μM) [12]. Low and intermediate concentrations of G. glabra derived isoliquiritigenin induced substantial transcriptional activity stimulate the proliferation of MCF-7 cells [13]. In the present study, the unknown properties of Chinese licorice root G. uralensis were investigated by comparing their ability to the human ER and their effect on estrogen-responsive human breast cancer MCF-7 cells over a broad range of concentrations. Our results show that the 80% ethanol extract of licorice root of the genius G. uralensis has estrogenic activity and anti-proliferative activity on MCF-7 human breast cancer cell.

Section snippets

Materials

Licorice roots were purchased from Kyungdong oriental medicine market, Seoul, Korea. The fresh fruiting bodies of licorice root (Glycyrrhiza uralensis) were washed, disintegrated, and extracted with CH3CH2OH (EtOH): H2O (100:0; 80:20; 60:40; 40:60; 20:80; 0:100) for 3 h. The crude extracts obtained were then subjected into silica gel chromatography and these elutes were evaporated to dry with a rotary evaporator.

Recombinant yeast cells

The Saccharomyces cerevisiae ER+LYS 8127 were gifted from Dr. Donald P. McDonnell

Estrogenic activity of the ethanol extracts of licorice root in recombinant yeast assays

A two-plasmid system consisting of human estrogen receptor (hER)-expression plasmid containing estrogen response element (ERE) was employed to study the estrogenic property of licorice root extracts. The reporter gene, β-galactosidase was expressed depending on the ligand-dependent transactivation. Treatment with increasing concentrations of the solvent, ethanol with licorice root induced a concentration-dependent increase in β-galactosidase activity in the yeast (Fig. 1). The activity of 80%

Discussion

This study investigated the antiproliferative activity of the ethanolic extract of G. uralensis licorice root in MCF-7 human breast cancer cells. We have chosen hormone-dependent cell line, MCF-7, because this cell line is ER-positive. In previous study, CHCl3, EtOAc, C6H14 and CH3OH–H2O (70:30) extracts of G. uralensis licorice root was found to exert anti-proliferative activity in this cell line [14].

Although the estrogenic activity of the licorice root of the genius G. glabra has been the

Acknowledgements

This study was supported by technology Development Program for Agriculture and Forestry, Ministry of Agriculture and Forestry (200001-03-3-CG000, 203004-03-HD110), Republic of Korea. And Research Institute for Veterinary Science, Seoul National University also supported this work. Professor Aruoma acknowledges the ‘Brain Pool Award’ from the Korean Ministry of Science and Technology (2004-2005) and its hosting by the Seoul National University, College of Veterinary Medicine.

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