Molecular and pharmacodynamic properties of estrogenic extracts from the traditional Chinese medicinal herb, Epimedium

Abstract

The Chinese medicinal herb, Epimedium, used traditionally for bone health exerts estrogenic activity (EA) in vitro. A genetically characterized Epimedium brevicornum (EB) extract induced biphasic responses in the mRNA and protein expression of the estrogen-regulated progesterone receptor gene in breast cancer (MCF-7) cells. These changes were mirrored changes in estrogenic receptor (ERα) content. In male Sprague–Dawley rats, administration of the estrogenic prodrug, estradiol valerate increased area-under-curve of serum effects for ERα (AUC difference: 18,900EA(ERα) min; 95% CI: 0–37,800; p = 0.05) and breast cancer cell (MCF-7) growth (AUC difference: 30,200EA(MCF-7) min; 95% CI: 24,200–36,200; p < 0.001), compared to placebo. Oral administration of Epimedium brevicornum increased ERα activity (1320EA(ERα) min, p < 0.01). Our data indicate that estrogen-responsive bioassays can measure the pharmacokinetic/pharmacodynamics of estrogenic activity in serum. Epimedium brevicornum extract increases estrogenic activity in serum and human studies are required to evaluate whether Epimedium extracts have utility for estrogen replacement therapy.

Introduction

Epimedium (Family Berberidaceau), a popular traditional Chinese medicinal plant, is colloquially known as Ying Yang Huo or Horny Goat weed (Yap and Yong, 2004). Dried leaves of several Epimedium species including Epimedium sagittatum (Sieb. Et Zucc), Epimedium koreanum Nakai, Epimedium pubescens Maxim., Epimedium wushanense T.S. Ying and Epimedium brevicornum Maxim (EB), alone or in complex formulations with other herbs, may be macerated in wine for oral consumption (Anon., 2000). Such extracts have, amongst other indications, been used traditionally to reinforce the Kidney Yang, for treatment of “coldness”, for male impotence, to improve female health, to strengthen bones and tendons, and to relieve pain in backbone and knees (Li, 1991).

The sudden drop in estrogen levels during menopause is associated with severe withdrawal symptoms, atrophy of connective tissues, poor bone health and osteoporosis. Estrogen replacement therapy (ERT) is an effective treatment modality. The targets for ERT are the estrogen receptors (ERα and ERβ), members of the 48 member steroid/nuclear receptor superfamily of transcription regulators (O’Malley, 2005). Like other nuclear receptors, ERα and ERβ have N-terminal transactivation domains, DNA-binding domains, and C-terminal ligand-binding domains. Ligands enter a pocket in the ligand-binding domain, activating the receptor and causing it to bind hormone-responsive elements in promoter regions, recruit coregulatory molecules, and modulate transcriptional activity of ER-regulated genes. Induction of the progesterone receptor (PR) is one of the best characterized and most physiologically important targets of estrogen action in breast cancer cells (Petz et al., 2004), and PR expression predicts survival in breast cancer patient (Oh et al., 2006). Unfortunately, estrogens in current use have unacceptable adverse effects. A large clinical trial recently demonstrated that a commonly used conjugated equine estrogen/progesterone combination for ERT was associated with increased hazard ratios for breast cancer, coronary heart disease and pulmonary embolism (Rossouw et al., 2002). The estrogen-only arm of this study indicated increased risks for coronary disease and thromboembolism (Anderson et al., 2004). The search for selective estrogen receptor modulators with favorable effects on menopausal symptoms and bone health, but without increasing breast cancer and cardiovascular risk, is an area of intense pharmaceutical activity (Riggs and Hartmann, 2003, Wallace et al., 2006). Nevertheless these adverse reports have led to unprecedented numbers of women abruptly stopping ERT medications containing synthetic sex steroids (Mclntosh and Blalock, 2005). Many have elected to use traditional herbal remedies without evidence for their efficacy (Beck et al., 2005).

Preliminary studies indicate that Epimedium, in particular extracts of Epimedium Brevicornum (EB), exert estrogenic effects in vitro (Yap et al., 2005). Extracts from Epimedium have been reported to exert osteoblastic activity in vitro (Meng et al., 2005) and its administration can prevent osteoporosis in rats (Zhang et al., 2006) and in post-menopausal women (Zhang et al., 2007), suggesting the possibility that this traditional herb may have utility for ERT. However unlike single-entity drugs, medicines of botanical origin contain multiple bioactive constituents. Classical pharmacokinetic studies of estrogenic compounds are dependent on measurements of single compounds or a very limited number of metabolites. Measurement of single compounds may not reflect in vivo bioactivity as ER-active compounds may exhibit additive, synergistic or antagonistic interactions. Recently, ER-driven cell-based bioassays have been shown to have good correlations with in vivo animal models of estrogenic activity (EA) (Sonneveld et al., 2006). There is therefore a possibility that ER-responsive cell-based bioassays may be utilized as biomarkers to measure summated EA in serum. Such an assay would measure the summated effects of all compounds, known and unknown, acting on the receptor and may therefore predict in vivo bioavailability.

The aims of this study are firstly, to clarify the molecular mechanisms whereby Epimedium brevicornum regulates PR expression in breast cancer cells. Secondly, we seek to evaluate the hypothesis that ERα-responsive bioassays, including ERα-driven breast cancer cell proliferation, can measure estrogenicity in serum of animal models at close time points following administration of a complex botanical like Epimedium brevicornum extract.