Androgen Receptor Regulation by Physiological Concentrations of the Isoflavonoid Genistein in Androgen-Dependent LNCaP Cells Is Mediated by Estrogen Receptor β
Introduction
Prostate cancer is the most common invasive tumor and, after lung, the leading cause of cancer deaths in males [1]. Despite the high incidence, little is known about its etiology. Accepted risk factors are age, race ethnicity and geographical dependence [2].
In the Asian population the incidence of this cancer is low compared with that in the USA and Western countries, with the highest incidence and mortality rates in African Americans [3]. Because of high soy intake, Asian subjects have 5–100 times higher serum levels of isoflavonoids, a group of phytoestrogens, than have subjects living in Western countries [4]. Adlercreutz and coworkers found a mean concentration of 276 nM of the isoflavonoid genistein in Asian men on regular soy diet [4].
It has been suggested that consumption of phytoestrogens may contribute to the low rate of prostate cancer in countries such as China and Japan [5], [6] and these substances are currently under investigation for their use in prostate cancer treatment and chemoprevention [7]. Indeed, these non-steroidal, diphenolic phytoestrogens found in many plants (especially soy products) have been shown in animal and in limited clinical investigations to be protective from osteoporosis [8], [9], cardiovascular disease [10] and also hormone-dependent cancers, namely cancers of the breast, uterus and prostate [11], [12].
In cell cultures genistein inhibited the growth of prostate cancer cells, possibly due to its ability to inhibit tyrosine kinases [1]. It was shown that genistein also inhibits prostate 5α-reductase enzyme in vitro [13], but this effect was not seen in an animal model [12]. Similarly, controversial data were obtained with respect to the effect on testosterone serum levels [12], [13], [14]. Other possible beneficial mechanisms that could be involved are inhibition of aromatase in peripheral tissues [15] and inhibition of 17β-hydroxysteroid oxidoreductase [16].
Epidemiologic, in vitro and laboratory animal studies provide evidence for the hypothesis that genistein has anticarcinogenic properties, but the molecular basis for its function in the prostate is still unclear. At submicromolar concentrations found in people on soy diet genistein binds to and activates the estrogen receptor (ER) with a preference for subtype β (ER-β) [17]. Higher concentrations in the upper micromolar range inhibit tyrosine kinase activities of growth factors and growth factor receptors, thus inhibiting cell proliferation by inhibiting growth factor signaling [18].
Exposure to a genistein rich diet may alter steroid receptor expression and activity resulting in a modified response to hormonal signals. We investigated these effects in prostate cancer cells with a focus on AR which mediates the action of androgens, one important risk factor for the development of prostate cancer. Their contribution is underscored by studies showing that this tumor rarely occurs in eunuchs or in men with a deficiency in 5α-reductase [19]. Our results suggest that genistein is down-regulating androgen receptor expression and this down-regulation is mediated through ER-β.
Section snippets
Culture of prostate cancer cell lines
LNCaP, PC-3 and DU-145 were obtained from the American Type Culture Collection (Rockville, MD, USA). PC-3 and DU-145 cells were grown in RPMI 1640 (HyClone, Logan, UT, USA) and LNCaP cells in MCDB 131 medium (Sigma-Aldrich-Chemie, Steinheim, Germany). Media were supplemented with 10% fetal calf serum (FCS; Biological Industries, Kibbutz Beit Haemek, Israel), 100 units/ml of penicillin and 0.1 mg/ml of streptomycin (Biological Industries) and the cells were cultured at 37 °C in a humidified
AR is down-regulated by genistein at the mRNA and the protein level
Treatment of androgen-sensitive LNCaP prostate cancer cells with physiological concentrations of genistein up to 1 μM resulted in a dose-dependent decrease of AR protein level within 24 hours starting at a concentration of 100 nM (Fig. 1). At this concentration genistein binds to the estrogen receptor and does not inhibit growth factor receptor tyrosine kinases [15]. To analyze whether genistein influences the AR mRNA, LNCaP cells were exposed to genistein for 24 hours and total RNA was isolated
Discussion
Prostatic carcinoma is one of the most frequently diagnosed cancers and one of the leading causes of cancer death in males and the opportunities for its prevention and therapy are limited. As agents used in the treatment of prostate cancer are of limited efficacy and often overshadowed by side effects, scientists are looking for prevention strategies and alternatives or additions to currently available therapies. Based on the observation that the concentration of isoflavones is much higher in
Acknowledgements
This project was supported by the European Community FP5 program (Project EUROESTROGENES, QLTR-2000-565).
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