Isoflavonoid Profiling and Estrogen-Like Activity of Four Genista Species from the Greek Flora

As part of our ongoing research on phytoestrogens, we investigated the phytochemical profile and estrogen-like activities of eight extracts from the aerial parts of four Genista species of Greek flora using estrogen-responsive cell lines. Ethyl acetate and methanolic extracts of G. acanthoclada, G. depressa,G. hassertiana, and G. millii were obtained with accelerated solvent extraction and their phytochemical profiles were compared using ultra-high-performance liquid chromatography-high-resolution mass spectrometry (uHPLC-HRMS). Fourteen isoflavonoids, previously isolated from G. halacsyi, were used as reference standards for their identification in the extracts. Thirteen isoflavonoids were detected in both extracts of G. acanthoclada and G. hassertiana, while fewer and far fewer were detected in G. millii and G. depressa, respectively. The ethyl acetate extracts of G. hassertiana and G. acanthoclada displayed 2.45- and 1.79-fold higher, respectively, estrogen-like agonist activity in Ishikawa cells compared to MCF-7 cells at pharmacologically relevant concentrations. Both these extracts, but not that of G. depressa, contained mono- and di-O-β-d-glucosides of genistein as well as the aglycone, all three of which are known to display full estrogen-like activity at lower-than-micromolar concentrations. The possibility of using preparations rich in G. hassertiana and/or G. acanthoclada extracts as a potentially safer substitute for low-dose vaginal estrogen for menopausal symptoms is discussed

Biological evaluation of isoflavonoids from Genista halacsyi using estrogen-target cells: Activities of glucosides compared to aglycones.

The purpose of this study was to evaluate the response of estrogen target cells to a series of isoflavone glucosides and aglycones from Genista halacsyi Heldr. The methanolic extract of aerial parts of this plant was processed using fast centrifugal partition chromatography, resulting in isolation of four archetypal isoflavones (genistein, daidzein, isoprunetin, 8-C-β-D-glucopyranosyl-genistein) and ten derivatives thereof. 7-O-β-D-glucopyranosyl-genistein and 7,4΄-di-O-β-D-glucopyranosyl-genistein were among the most abundant constituents of the isolate. All fourteen, except genistein, displayed low binding affinity for estrogen receptors (ER). Models of binding to ERα could account for the low binding affinity of monoglucosides. Genistein and its glucosides displayed full efficacy in inducing alkaline phosphatase (AlkP) in Ishikawa cells, proliferation of MCF-7 cells and ER-dependent gene expression in reporter cells at low concentrations (around 0.3 μM). ICI182,780 fully antagonized these effects. The AlkP-inducing efficacy of the fourteen isoflavonoids was more strongly correlated with their transcriptional efficacy through ERα. O-monoglucosides displayed higher area under the dose-response curve (AUC) of AlkP response relative to the AUC of ERα-transcriptional response compared to the respective aglycones. In addition, 7-O-β-D-glucopyranosyl-genistein and 7,4΄-di-O-β-D-glucopyranosyl-genistein displayed estradiol-like efficacy in promoting differentiation of MC3T3-E1 cells to osteoblasts, while genistein was not convincingly effective in this respect. Moreover, 7,4΄-di-O-β-D-glucopyranosyl-genistein suppressed lipopolysaccharide-induced tumor necrosis factor mRNA expression in RAW 264.7 cells, while 7-O-β-D-glucopyranosyl-genistein was not convincingly effective and genistein was ineffective. However, genistein and its O-glucosides were ineffective in inhibiting differentiation of RAW 264.7 cells to osteoclasts and in protecting glutamate-challenged HT22 hippocampal neurons from oxidative stress-induced cell death. These findings suggest that 7-O-β-D-glucopyranosyl-genistein and 7,4΄-di-O-β-D-glucopyranosyl-genistein display higher estrogen-like and/or anti-inflammatory activity compared to the aglycone. The possibility of using preparations rich in O-β-D-glucopyranosides of genistein to substitute for low-dose estrogen in formulations for menopausal symptoms is discussed

New hydroxystilbenoid derivatives endowed with neuroprotective activity and devoid of interference with estrogen and aryl hydrocarbon receptor-mediated transcription

We have synthesized a series of new (E) stilbenoid derivatives containing hydroxy groups at ring positions identical or similar to those of trans-resveratrol and bearing one or two bulky electron donating groups ortho to 4′-OH and we have evaluated their neuroprotective activity using glutamate-challenged HT22 hippocampal neurons to model oxidative stress-induced neuronal cell death. The most active derivatives, 5-{(E)-2-[3,5-bis(1-ethylpropyl)-4-hydroxyphenyl]ethenyl}-1,3-benzenediol (2), 5-[(E)-2-(3,5-di-tert-butyl-4-hydroxyphenylethenyl)]-1,3-benzenediol (4) and 5-{(1E,3E)-4-[3,5-bis(1-ethylpropyl)-4-hydroxyphenyl]-1,3-butadienyl}-1,3-benzenediol (6), had EC50 values of 30, 45 and 12 nM, respectively, and were ca. 100 to 400-fold more potent than resveratrol. Derivatives 2, 4 and 6 lacked cytotoxic activity against HT22 cells and estrogen receptor agonist or antagonist activity in estrogen response element-dependent gene expression and in estrogen-dependent proliferation of MCF-7 human breast cancer cells. In addition, they were incapable of interfering with aryl hydrocarbon receptor-mediated xenobiotic response element-dependent gene expression. Derivatives 2, 4 and 6 might assist in the development of lead candidates against oxidative stress-driven neurodegenerative diseases that will not increase endocrine cancer risk nor affect drug activation and detoxification mechanisms

Engineered chimeric enzymes as tools for drug discovery: generating reliable bacterial screens for the detection, discovery, and assessment of estrogen receptor modulators

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