Phytoestrogen Bakuchiol Exhibits In Vitro and In Vivo Anti-breast Cancer Effects by Inducing S Phase Arrest and Apoptosis

Abstract

Phytoestrogen has been proposed as an alternative to hormone replacement therapy, which has been demonstrated to promote a high risk of breast cancer. However, the effect of phytoestrogen on breast cancer development has not been fully understood. Bakuchiol is an active ingredient of a traditional Chinese herbal medicine Fructus Psoraleae, the dried ripe fruit of Psoralea corylifolia L. (Fabaceae). The in vitro and in vivo estrogenic activities and anti-breast cancer effects of bakuchiol have not been well-studied. We found that bakuchiol induced the GFP expression in transgenic medaka (Oryzias melastigma, Tg, Chg:GFP) dose-dependently (0–1 μg/ml), demonstrating its in vivo estrogenic activity. Low dose of bakuchiol (1 μg/ml) induced the cell proliferation and ERα expression in MCF-7 cells, which could be blocked by the anti-estrogen ICI 182780, suggesting the in vitro estrogenic activity of bakuchiol. Our data indicated that high doses of bakuchiol (>2 μg/ml) inhibited breast cancer cell growth, with a stronger anti-proliferative effect than resveratrol, a widely studied analog of bakuchiol. High doses of bakuchiol (4, 7, and 10 μg/ml) were used for the further in vitro anti-breast cancer studies. Bakuchiol induced ERβ expression and suppressed ERα expression in MCF-7 cells. It also induced S phase arrest in both MCF-7 and MDA-MB-231 cells, which could be rescued by caffeine. Knock-down of p21 also marginally rescued S phase arrest in MCF-7 cells. The S phase arrest was accompanied by the upregulation of ATM, P-Cdc2 (Tyr15), Myt1, P-Wee1 (Ser642), p21 and Cyclin B1, suggesting that blocking of Cdc2 activation may play an important role in bakuchiol-induced S phase arrest. Furthermore, bakuchiol induced cell apoptosis and disturbed mitochondrial membrane potential in MCF-7 cells. The bakuchiol-induced apoptosis was associated with increased expression of Caspase family and Bcl-2 family proteins, suggesting that bakuchiol may induce apoptosis via intrinsic apoptotic pathway. The in vivo anti-breast cancer effect of bakuchiol was further proved in zebrafish (Danio rerio, wild-type AB) xenografts. 0.5 μg/ml of bakuchiol significantly reduced the MCF-7 cell mass in zebrafish xenografts. Overall, these results suggested the potential of using bakuchiol in HRT and breast cancer treatment.

Genus Psoralea: A review of the traditional and modern uses, phytochemistry and pharmacology

“…Angelicin (2) n (Backhouse et al, 2001;Labbe et al, 1996;Li et al, 2016;Madrid et al, 2012Madrid et al, , 2013Madrid et al, , 2015aMadrid et al, , 2015b Bakuchiol (…”

An Insight into the Prophylactic Effects of the Leguminosae Family Plants against Oxidative Stress-Induced Pathophysiological Conditions

“…In recent studies, a phytoestrogen, bakuchiol, isolated from a Fabaceae family plant Psoralea corylifolia L. showed significant antiproliferative and anticancer effect against breast cancer [133]. It can effectively induce cell cycle arrest in S phase by blocking the activation of cyclin-dependent kinases (Cdc2) along with the upregulation of different proapoptotic signaling pathways.…”

Estrogenic Potentials of Traditional Chinese Medicine

“…Due to its easy handling and great sensitivity, the reporter-gene assay has been widely used to examine estrogenic activity and effects of TCM and their effective chemicals; for example, CALUX assay for Justicia pectoralis (Locklear et al, 2010); GFP-based assay for bakuchiol (Li et al, 2016); luciferase-based assay for Danggui Buxue Tang (Zhang et al, 2013a); MVLN cell assay for Si-Wu-Tang (Liu et al, 2012); YES assay for essential oils (Tabanca et al, 2004), Glycine soja (Agradi et al, 2006) and Butea superba (Cherdshewasart et al, 2010).…”

“…In a Xenopus assay, embryos/eggs were exposed to test chemicals, quercetin, or control solvent, and, after certain periods of time, frogs were subjected to morphological and histological examinations (Cong et al, 2006). The results of animal tests are often reevaluated by other assays to confirm the effects or to find the differences in species, principles or materials among the assays; for example, the rainbow trout assay was reevaluated by reverse-transcription polymerase chain-reaction (RT-PCR), DNA microarray assay and Western blotting (Tilton et al, 2006), rodent uterotrophic assay by ligand-binding assay (Owens and Ashby, 2002) and medaka assay by signaling pathway analyses using human breast cancer cells (Li et al, 2016).…”

“…For example, the angiogenic effects of calycosin were examined by evaluating the diameter of subintestinal vein vessels of zebrafish, which was re-evaluated by other assays, suggesting the involvement of ERs and crosstalk of ER-signaling with vascular endothelial growth factor receptor (VEGFR) and mitogen-activated protein kinase (MAPK) pathways (Tang et al, 2010). A transgenic medaka strain containing the gene for green fluorescence protein (GFP), which was transcriptionally regulated with the estrogen-responsive, liverspecific choriogenin H gene promoter, was developed and used to evaluate the effects of bakuchiol on the cell cycle and apoptosis (Li et al, 2016). Estrogenicity of 3,3 0 -diindolylmethane, a dietary phytochemical, was examined by the expression level of the Lee et al (2006) a Only representative references are shown, and the TCM or chemicals examined are shown in parentheses.…”