Differential regulation of estrogen receptors α and β by 4-(E)-{(4-hydroxyphenylimino)-methylbenzene,1,2-diol}, a novel resveratrol analog

Abstract

Breast cancer is the second leading cause of death among women in the United States. Estrogens have been implicated as major risk factors in the development of breast neoplasms. Recent epidemiologic studies have suggested a protective role of phytoestrogens in prevention of breast and other cancers. Resveratrol, a naturally occurring phytoestrogen found notably in red grapes, berries and peanuts, has been shown to possess potent anti-cancer properties. However, the poor efficacy of resveratrol has prevented its use in a clinical setting. In order to improve the efficacy of resveratrol, we have synthesized a small combinatorial library of azaresveratrol analogs and tested them for their ability to inhibit the growth of breast cancer cell lines. We have recently shown that one of the synthesized analogs, 4-(E)-{(4-hydroxyphenylimino)-methylbenzene, 1, 2-diol} (HPIMBD), has better anti-cancer properties than resveratrol. The objective of this study was to investigate the differential regulation of estrogen receptors (ERs) α and β as a potential mechanism of inhibition of breast cancer by HPIMBD. Estrogen receptors α and β have been shown to have opposing roles in cellular proliferation. Estrogen receptor α mediates the proliferative responses of estrogens while ERβ plays an anti-proliferative and pro-apoptotic role. We demonstrate that HPIMBD significantly induces the expression of ERβ and inhibits the expression of ERα. HPIMBD also inhibits the protein expression levels of oncogene c-Myc, and cell cycle protein cyclin D1, genes downstream to ERα and important regulators of cell cycle, and cellular proliferation. HPIMBD significantly induces protein expression levels of tumor suppressors p53 and p21 in MCF-7 cells. Additionally, HPIMBD inhibits c-Myc in an ERβ-dependent fashion in MCF-10A and ERβ1-transfected MDA-MB-231 cells, suggesting regulation of ERs as an important upstream mechanism of this novel compound. Molecular docking studies confirm higher affinity for binding of HPIMBD in the ERβ cavity. Thus, HPIMBD, a novel azaresveratrol analog may inhibit the proliferation of breast cancer cells by differentially modulating the expressions of ERs α and β.

4-(E)-{(p-tolylimino)-methylbenzene-1,2-diol}, 1 a novel resveratrol analog, differentially regulates estrogen receptors α and β in breast cancer cells

“…Estrogen receptor β1-transfected MDA-MB-231 and empty vector-transfected MDA-MB-231 were a gift from Dr. Leigh C. Murphy (University of Manitoba, Canada). All breast epithelial and breast cancer cell lines were cultured as reported recently [30]. Cells were treated with 50 μM doses of Res or TIMBD for up to 72 hours.…”

“…Cells were seeded in 96-well plates for MTT assays and treated with 50 μM Res or TIMBD for 72 hours [30]. After treatments, cells were incubated with MTT reagent at 37°C.…”

“…Formazan crystals were dissolved in DMSO and plates were read in Benchmark Plus microplate spectrophotometer (Biorad, Hercules, CA). Percentage of viable cells were calculated as described recently [30]. …”

“…Cells were plated in 96-well plates and treated with different doses of Res or TIMBD as described recently [30]. Post-treatments, medium was collected and absorbance was measured using the Benchmark Plus microplate spectrophotometer (Biorad, Hercules, CA).…”

“…4-(E)-{(p-tolylimino)-methylbenzene-1,2-diol} (TIMBD) showed better potency than any of the other analogs in inhibiting the proliferation of breast cancer cell lines [29]. We have recently shown the effect of 4-(E)-{(4-hydroxyphenylimino)-methylbenzene, 1,2-diol} (HPIMBD) in regulating ERs in breast cancer cells [30]. In the present study, we investigated the effect of TIMBD on the regulation of ERα and β.…”