Cancer Letters

Cancer Letters

Volume 276, Issue 1, 8 April 2009, Pages 68-73
Cancer Letters

The inhibitory effect of diarylpropionitrile, a selective agonist of estrogen receptor beta, on the growth of MC38 colon cancer line

https://doi.org/10.1016/j.canlet.2008.10.050Get rights and content

Abstract

The protective role of estrogens in the colon carcinogenesis has been suggested for many years and attributed mainly to estrogen receptor beta (ERβ). However, the direct effect of estrogens and their action through ERβ on the growth of colon cancer have been rarely studied.

The aim of this study was to examine the effect of various concentrations (10−4–10−12 M) of diarylpropionitrile (DPN) – a selective agonist of ERβ – on the growth of murine MC38 colon cancer line. Moreover, the aim of this paper was the immunohistochemical assessment of estrogen and progesterone receptor expression in human colon tissues and in MC38 cells (only ERβ).

We found that DPN induced a growth inhibition of MC38 cancer (50–94% of control group) at the highest (10−4 M) and two lowest concentrations (10−11 and 10−12 M). Furthermore, we detected a nuclear-cytoplasmic expression of ERβ in human normal and neoplastic colon tissues and in the studied MC38 cancer cells.

The inhibitory effect of DPN on the growth of MC38 colon cancer line suggests a possibility of using a selective estrogen receptor agonist in the treatment of colon cancer.

Introduction

In the developed countries colorectal cancer belongs to the major health problems because of high incidence rate, mortality [1] and unsatisfactory efficacy of standard chemotherapy [2]. Therefore, researches with new antineoplastic substances including cytostatic drugs as well as biomodulators are still being conducted.

The role of female sex hormones, especially estrogens, in colon carcinogenesis has been discussed for many years. This involvement was implied by some epidemiological data such as sex- and age-specific occurrence of colorectal cancer [3], [4] and the protective influence of increasing parity [5]. Furthermore, many observational studies [6] and the randomized trial WHI (Women’s Health Initiative) [7] confirmed that the application of a hormonal replacement therapy in menopausal women decreased the risk of colon cancer. The protective effect of estrogens in colorectal carcinogenesis is attributed mainly to estrogen receptor beta (ERβ), which was shown to be the prevailing subtype of ER in human colon and its expression was decreased in colon cancer [8], [9], [10], [11]. ERα was reported to be minimally expressed in both – normal and neoplastic − colon cells [8], [9]. The predominant role of ERβ in colon physiology and carcinogenesis is also suggested by some experimental studies. The experiments with βERKO mice (β Estrogen Receptor Knockout) revealed the significance of ERβ in the growth, organization and maintenance of the normal colonic crypt-villus architecture [12]. In addition, the over-expression of ERβ in engineered HCT8 human colon cancer line inhibited cell proliferation and increased cell adhesion in a ligand-independent manner [13].

Despite many facts supporting an association of female sex hormones with colorectal carcinogenesis, the studies examining the direct effect of estrogens on the colon cancer growth have been rarely performed and gave often conflicting results [14], [15]. To our knowledge, the effect of selective agonist of ERβ on the growth of colon cancer has not been studied at all, although recently a potential use of these agonists in colon cancer treatment has been suggested [16]. Diarylpropionitrile (DPN) belongs to such compounds. It is a quite newly synthesized substance, which acts as an agonist on both the ER subtypes, but has a 70-fold higher relative binding affinity and 170-fold higher relative potency in transcription assays with ERβ than with ERα [17]. DPN and other specific agonists or antagonists of ER subtypes are used in studies examining the role of ERα and ERβ in immunology [18], central nervous system [19] and other organs [20].

Thus, the aim of this paper was to examine the direct effect of DPN on the growth of murine MC38 colon cancer in vitro assessed by two colorimetric methods reflecting changes in proliferation and apoptosis. Moreover, the aim of this study was the immunohistochemical assessment of the expression of estrogen and progesterone receptor (ER and PR) in human colon tissues (normal and pathological) and in the studied cell line (only ERβ).

Section snippets

Cell line

Murine MC38 colon cancer cells were used in the study. The cells were routinely grown in a humidified incubator at 37 °C with 5% CO2 in RPMI 1640 medium (Sigma), supplemented with: 25 nM Hepes buffer (Sigma), 4 mM l-glutamine (Sigma), 100 U/ml penicillin and 100 μg/ml streptomycin solution (Sigma), 2 g/l sodium bicarbonate (Sigma) and 5% fetal calf serum (FCS, Biochrom). The cells were passaged every 5–7 days with 0.05% trypsin/0.02% EDTA (trypsin–EDTA, Sigma), and the medium was changed every 3–4

Influence of DPN on MC38 growth

DPN inhibited the growth of MC38 cancer with a moderate potency (50–94% of control group). Its inhibitory effect was shown in Mosmann method in 72 h culture for the highest (10−4 M; 77% of control group) and two lowest concentrations (10−12 and 10−11 M; 92 and 94% of control group) and in 24 h culture for the extreme concentrations (10−4 and 10−12 M; 84 and 75% of control group) (Fig. 1). The cancer growth inhibition induced by DPN at those concentrations was also confirmed in the method based on

Discussion

In this study, we have shown for the first time that a selective ERβ agonist, DPN, inhibited the growth of murine MC38 colon cancer line, which expressed ERβ. Moreover, we have also confirmed the recent data of some other authors [8], [9], [10], [11] reporting the presence of ERβ in the human colon tissues, both normal and neoplastic, and the decrease of its expression in colon tumors. However, it should be underlined that the results of studies examining the expression of sex steroid receptors

Conclusions

Summing up, in this paper we have shown the inhibitory effect of DPN on the growth of the murine MC38 colon cancer line possessing ERβ and the expression of this subtype of estrogen receptor in human colon cancers. These data suggest a possibility of using selective ERβ agonists in the treatment of colon cancer, however, further studies are necessary.

Conflicts of interest statement

All authors have not got any financial and personal relationships with other people or organisations that could inappropriately influence (bias) their work.

Acknowledgement

This study was supported by grant of Medical University of Lodz No 503-1084-3.

References (30)

  • D. Schrag

    The price tag on progress-chemotherapy for colorectal cancer

    N. Engl. J. Med.

    (2004)
  • R.K. Peters et al.

    Reproductive factors and colon cancers

    Br. J. Cancer

    (1990)
  • S. Singh et al.

    Sex differences in the incidence of colorectal cancer: an exploration of oestrogen and progesterone receptors

    Gut

    (1993)
  • J.E. Rossouw et al.

    Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results from the women’s health initiative randomized controlled trial

    JAMA

    (2002)
  • E.F. Foley et al.

    Selective loss of estrogen receptor β in malignant human colon

    Cancer Res.

    (2000)
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