Background: The estrogen receptor alpha (ERα) is found predominately in the nucleus, both in hormone stimulated and untreated cells. Intracellular distribution of the ERα changes in the presence of agonists but the impact of different antiestrogens on the fate of ERα is a matter of debate.Results: A MCF-7 cell line stably expressing GFP-tagged human ERα (SK19 cell line) was created to examine the localization of ligand-bound GFP-ERα. We combined digitonin-based cell fractionation analyses with fluorescence and immuno-electron microscopy to determine the intracellular distribution of ligand-bound ERα and/or GFP-ERα.Using fluorescence- and electron microscopy we demonstrate that both endogenous ERα and GFP-ERα form numerous nuclear focal accumulations upon addition of agonist, 17β-estradiol (E2), and pure antagonists (selective estrogen regulator disruptor; SERD), ICI 182,780 or RU58,668, while in the presence of partial antagonists (selective estrogen regulator modulator; SERM), 4-hydroxytamoxifen (OHT) or RU39,411, diffuse nuclear staining persisted.Digitonin based cell fractionation analyses confirmed that endogenous ERα and GFP-ERα predominantly reside in the nuclear fraction. Overall ERα protein levels were reduced after estradiol treatment. In the presence of SERMs ERα was stabilized in the nuclear soluble fraction, while in the presence of SERDs protein levels decreased drastically and the remaining ERα was largely found in a nuclear insoluble fraction. mRNA levels ofESR1were reduced compared to untreated cells in the presence of all ligands tested, including E2. E2 and SERDs induced ERα degradation occurred in distinct nuclear foci composed of ERα and the proteasome providing a simple explanation for ERα sequestration in the nucleus.Conclusions: Our results indicate that chemical structure of ligands directly affect the nuclear fate and protein turnover of the estrogen receptor alpha independently of their impact on transcription. These findings provide a molecular basis for the selection of antiestrogen compounds issue from pharmacological studies aimed at improving treatment of breast cancer.

Auraptene is a prenyloxycoumarin from Citrus species with chemopreventive properties against colitis-related colon and breast cancers through a yet-undefined mechanism. To decipher its mechanism of action, we used a ligand-structure based approach. We established that auraptene fits with a pharmacophore involved in both the inhibition of acyl-CoA:cholesterol acyl transferase (ACAT) and the modulation of estrogen receptors (ERs). We confirmed experimentally that auraptene inhibits ACAT and binds to ERs in a concentration-dependent manner and that it inhibited ACAT in rat liver microsomes and in intact cancer cells of murine and human origins, with an IC₅₀ value in the micromolar range. Auraptene bound to ERs with affinities of 7.8 μM for ERα and 7.9 μM for ERβ, stabilized ERs, and modulated their transcriptional activity via an ER-dependent reporter gene and endogenous genes. We further established that these effects correlated well with the control of growth and invasiveness of tumor cells. Our data shed light on the molecular mechanism underlying the anticancer and chemopreventive effects of auraptene.

Obesity is a major public health problem in recent decades. The accumulation of excessive fat promotes inflammatory status. Meanwhile, herbal products are marketed for their weight-loss properties, such as Nigella sativa (N. Sativa) which has been used for centuries to treat rheumatoid arthritis, diabetes, and asthma; recently, the anti-obesity characteristics of N. sativa have also been indicated. However, the exact mechanisms and cellular-related pathways are still unclear. Thus, we will aim to assess the effects of oral N. sativa on the gene expression of inflammatory and adipogenesis-related factors, including TNF-α, PPAR-γ, and adiponectin as well as assessing their serum concentrations among obese and overweight individuals. Obese and overweight women aged 25–55 years with a body mass index (BMI) of 25–35 kg/m2 will be recruited from the Obesity Clinic in Shahid Sadoughi University of Medical Sciences and will be assessed for eligibility against inclusion criteria. They will be randomly assigned into two groups to receive either two capsules of N. sativa or two capsules of placebo per day for eight weeks (each capsule contains 1000 mg of N. sativa or placebo). There will be a four-week wash-out period and then participants will receive the reverse supplements for another eight weeks. Biochemical assessments and gene expressions (using real-time polymerase chain reaction) will be conducted at the beginning and at the end of every intervention period. The present study will investigate the probable cellular pathways for the anti-obesity effects of N. sativa in overweight/obese women. Iranian Registry of Clinical Trials, IRCT20180528039884N1 . Registered on 2nd of July, 2018.

Tamoxifen Resistance and CYP2D6 Copy Numbers in Breast Cancer Patients Tamoxifen;CYP2D6 gene;copy numbers;drug resistance; Background: Breast cancer accounts about one million from total annual ten million new diagnosed cases of neoplasia worldwide and is the main cause of death due to cancer in women. Tamoxifen is the most popular selective estrogen receptor modulator used in anti estrogen treatments. Tamoxifen must be converted into its metabolite endoxifen for biologic effects; this conversion process is catalysed by highly polymorphic cytochrome P450 2D6 (CYP2D6). This study surveyed copy number variation of the CYP2D6 gene and its possible correlation with Tamoxifen resistance in breast cancer patients. Methods: This case control study was performed on samples taken from 79 patients with breast cancer who used tamoxifen in Yazd and Tehran Cities, Iran. Real time reactions were conducted for 10 healthy samples using the comparative $C_t$ (Cycles threshold) method, each pair of genes being compared and samples with ratios around 1 were taken as control samples. Proliferation reactions were done by Real-Time PCR ABI Prism 7500. All registered data were transformed into SPSS 15 program and analyzed. Results: Efficiency of PCR for both CYP2D6 and ALB genes was 100%. From all 23 drug resistant patients 21.7% had one copy, 47.8% two copies and 30.4% had three copies. Also from all 56 drug sensitive patients, 26.8% had one copy, 51.8% two copies and 21.4% had three copies. The percentage of patients with one and two copies was similar between two groups but patients with three copies were more likely to belong to the drug resistant group more. Odd ratios for one and two copies were 0.759 and 0.853 respectively, indicating possible protective effects while that for three copies was 1.604. Conclusions: Based on our study there is no significant link between CYP2D6 gene copy numbers and tamoxifen resistance in women with breast cancer. But more studies considering other influencing factors appear warranted.

The TP53 is important in functions of cell cycle control, apoptosis, and maintenance of DNA integrity. Studies on the association between p53 codon 72 polymorphism and primary open-angle glaucoma (POAG) risk have yielded conflicting results. Published literature from PubMed and Web of Science databases was retrieved. All studies evaluating the association between p53 codon 72 polymorphisms and POAG were included. Pooled odds ratio (OR) and 95% confidence interval (CI) were calculated. Eleven separate studies including 2541 cases and 1844 controls were pooled in the meta-analysis. We did not detect a significant association between POAG risk and p53 codon 72 polymorphism overall population except allele genetic model (C vs. G: OR = 0.961, 95% CI = 0.961–0.820, P = 0.622). In the stratified analysis for Asians and Caucasians, there was an association between p53 codon 72 polymorphism and POAG. In the dominant model in the overall population and by ethnicity subgroups, the highest elevated POAG risk was presented. In summary, these results indicate that p53 codon 72 polymorphism is likely an important genetic factor contributing to susceptibility of POAG. However, more case–controls studies based on larger sample size and stratified by ethnicity are suggested to further clarify the relationship between p53 codon 72 polymorphism and POAG.