t47d  (ATCC)

Journal: iScience

Article Title: Distinct 2-phenylimidazo[1,2- a ]pyridine derivatives that inhibit breast cancer cell proliferation identified as AHR ligands

doi: 10.1016/j.isci.2026.115936

Figure Lengend Snippet: Attenuation of estrogen receptor action by the imidazopyridines (see also ). (A and B) Four-way plots show differentially expressed genes (DEGs) in MCF-7 cells (A) and T47D cells (B) treated with X15695 under estrogen-deprived condition ( X15695 vs. vehicle) (X axis) or E 2 -supplemented condition ( X15695 + E 2 vs. E 2 ) (Y axis). Genes with |Log 2 (Fold Change)| ≥ 1 and adj. p -value ≤0.05 were considered significant. Upregulated and downregulated DEGs are highlighted in red and blue, respectively; non-significant genes are shown in gray. Select representative ER target genes (blue) and AHR target genes (red) are explicitly labeled. In the 4-way plot, each gene has a coordinate (X,Y), the X value shows the log 2 FC of X15695 vs. vehicle, while Y value refers to the log 2 FC of X15695_E 2 vs. E 2. (C and D) Results of reporter gene assay on the action of the indicated imidazopyridines on the transactivation function of the ERα. Cells were hormone starved for 24 h and treated with the indicated compounds (10 nM–10 μM) in the presence of 10 nM (C) or 0.1 or 10 nM (D) E 2 for 24 h. The results are the mean value ± SEM of independent biological replicates (n = 2–4). (E) Quantitative RT-PCR to detect the effect of the imidazopyridine derivatives on the expression of the indicated ER target genes in MCF-7 cells in the presence and absence of E 2 . Cells were serum starved for 72 h and treated with the indicated compounds (1 μM) in the presence and absence of 10 nM E 2 for 24 h. The data represent the mean ± SEM. Statistical significance was assessed by multiple t test ( n = 3; ∗ p < 0.05; ∗∗< 0.01; ∗∗∗< 0.001; ns is not significant).

Article Snippet: T47D , ATCC , RRID: CVCL_0553.

Techniques: Labeling, Reporter Gene Assay, Quantitative RT-PCR, Expressing

Journal: iScience

Article Title: Distinct 2-phenylimidazo[1,2- a ]pyridine derivatives that inhibit breast cancer cell proliferation identified as AHR ligands

doi: 10.1016/j.isci.2026.115936

Figure Lengend Snippet: The aryl hydrocarbon receptor is the direct molecular target of antiproliferative imidazopyridines (A and B) Nano-DSF analysis of the interaction between Hsp90-XAP2-AHR complexes and the indicated imidazopyridines. DMSO and Indirubin (indi) were used as negative and positive controls. The results are representative of independent biological replicates ( n = 3). The results in (A) show the first derivative plot for X15695 , X19724, and X19728. In (B) are the fluorescence ratio plots (F350/F330) as a function of temperature that show the raw protein unfolding transition. (C and D) Results of molecular docking calculations. The binding modes of six imidazopyridine derivatives ( X15695 , X19724, X19728, X19712, X19718, and X19167) and (D) three additional imidazopyridines (X19720, X15696 , and X20046) within the PAS-B binding domain of AHR shown in a three-dimensional representation: ligands and the main interacting residues are shown as sticks, protein as white cartoons; π- π stacking interactions, as identified by the ligand interaction diagram (Maestro, Schrödinger), are represented as green dashed lines; the interacting residues and the corresponding centroid-to-centroid distances are labeled. (E) Results of reporter gene assay on the action of the indicated imidazopyridines on the transactivation function of the AHR. HAHLH cells were serum starved for 24 h and treated with the indicated concentrations of the compounds for 8 h. Results were expressed as % of the maximal luciferase activity obtained with dioxin 10 nM. The results are the mean value ± SEM of independent biological replicates (n = 2–4). (F and G) Quantitative RT-PCR to detect the effect of the indicated imidazopyridine derivatives on the expression of three AHR target genes in MCF-7 and T47D cells. Cells were serum starved for 72 h and treated with the indicated compounds (1 μM) for 16 h. The data represent the mean ± SEM. Statistical significance was assessed by multiple t test (n = 3–12; ∗ p < 0.05; ∗∗< 0.01; ∗∗∗< 0.001; ns is not significant).

Article Snippet: T47D , ATCC , RRID: CVCL_0553.

Techniques: Fluorescence, Binding Assay, Labeling, Reporter Gene Assay, Luciferase, Activity Assay, Quantitative RT-PCR, Expressing

Journal: iScience

Article Title: Distinct 2-phenylimidazo[1,2- a ]pyridine derivatives that inhibit breast cancer cell proliferation identified as AHR ligands

doi: 10.1016/j.isci.2026.115936

Figure Lengend Snippet: The antiproliferative effect of imidazopyridines is strictly dependent on the AHR (see also ). (A) Effect of the AHR antagonist CH223191 on the inhibition of clonal expansion by the X compounds. Quantification of the action of the AHR antagonist CH223191 on the inhibition of the clonal expansion of MCF-7 cells by the indicated X compounds. Cells were treated with 10 nM of the X compounds in the absence and presence of 1 μM CH223191 for 14 days. The values are the means ± SEM. Statistical significance was assessed by multiple t test. ( n = 4. ∗∗∗ p < 0.001; ns is not significant). (B and C) Proliferation of empty vector-transfected MCF-7 and T47D cells, as well as MCF-7 AHR KO clones #2 and #17, and T47D AHR KO clones #7 and #30. Cells were cultured in complete media and counted on days 0 and 5. The starting number of cells was 1 ×10 4 per well. Data are the averages of three independent experiments ± SEM, normalized to day 0. Statistical significance was assessed by two-way ANOVA (∗∗∗∗ p ≤ 0.0001). (D) Representative images of the clonal expansion of empty vector-transfected MCF-7 cells or AHR KO MCF-7 cell clones #2 and #17 after treatment with the indicated concentrations of X compounds. (E) Quantification of the concentration-dependent action of the indicated compounds on the clonal expansion of empty vector-transfected MCF-7 cells and AHR KO MCF-7 clones #2 and #17. The results are shown as bar charts, and they represent the means ± SEM. Statistical significance was assessed by one-way ANOVA (n = 3–4. ∗∗∗ p < 0.001; ∗∗∗∗ <0.0001; ns is not significant). (F) Quantification of the concentration-dependent action of the indicated compounds on the clonal expansion of empty vector-transfected T47D cells and AHR KO T47D clones #7 and #30. The results are shown as bar charts, and they represent the means ± SEM. Statistical significance was assessed by one-way ANOVA (n = 3–4. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; and ∗∗∗∗ p < 0.0001; ns is not significant).

Article Snippet: T47D , ATCC , RRID: CVCL_0553.

Techniques: Inhibition, Plasmid Preparation, Transfection, Clone Assay, Cell Culture, Concentration Assay

Journal: microPublication Biology

Article Title: Minoxidil May be a Partial Agonist of Estrogen Receptor Alpha

doi: 10.17912/micropub.biology.002173

Figure Lengend Snippet: A) Line-stick structures of 17β-estradiol (E 2 ), minoxidil (MX), and minoxidil intermediate (MXI). T47D and MCF-7 cell proliferation after 4 days of treatment with B) EtOH, E 2 , or MX; C) EtOH, E 2 , or MXI; D) EtOH, E 2 , 100 pM E 2 and 4-hydroxytamoxifen (4-OHT), or 1-20 nM MX and 1 µM 4-OHT. E) Tryptophan emission spectrum of 200 nM full-length ERα with E 2 or MX. F) Representative images of E 2 (left) and MX (right) docked to the ERα ligand-binding domain (PDB ID: 1ERE). Key residues are labeled with some interaction distances shown. G) Average docking energies of E 2 (n=53) and MX (n=66) bound to ERα. Data in B-D and G is average ± standard deviation; with at least n=3 biological replicates for B-D. All statistical analyses are Student’s t-test where n.s.=not significant; *p<0.05, **p<0.01, ***p<0.001, and ****p<0.0001.

Article Snippet: Cell Maintenance T47D cells were courtesy of David J. Shapiro and MCF-7 cells were from ATCC.

Techniques: Binding Assay, Ligand Binding Assay, Labeling, Standard Deviation

Journal: Journal of the Endocrine Society

Article Title: Estrogenic activity in tampon products

doi: 10.1210/jendso/bvag094

Figure Lengend Snippet: Estradiol dose-response and recovery of spiked estradiol from tampon extracts. (A) T47D-luc cells were treated with an 11-point estradiol dilution series starting at 30nM, followed by 10nM, 3.3nM, 1.11nM, 370pM, 123pM, 41pM, 14pM, 4.6pM, 1.5pM, and 0.5pM). For each concentration, 2 μL of steroid dilution was added to 198 μL assay medium, and cells were incubated for 24 hours. Estrogen-responsive luciferase activity was quantified using BrightGlo substrate. Luminescence values represent the mean of 4 technical replicates, and a sigmoidal dose-response curve with EC 50 determination was generated using GraphPad v10. (B) Recovery of estradiol from tampon brands A-D following spiking and extraction. One gram of each tampon from Brands A, B, C, and D were spiked with 2nM estradiol, soaked in a 1:1 water:ethanol solution for 4 hours, extracted, evaporated to dryness, and reconstituted in methanol. Extracts were diluted 1:100 in cell culture medium and then serially diluted 1:2 to generate 4 assay points. T47D-luc cells were incubated with a further 1:10 dilution of each tampon extract for 24 hours and luciferase activity measured using BrightGlo. Each point represents mean ± SEM of 4 technical replicates corrected for the blank control sample (1:1 ethanol:water solution SPE-extracted as per samples). The data for the spiked tampons are plotted alongside estradiol standards run in parallel.

Article Snippet: Estrogen Receptor Luciferase Reporter T47D Stable Cell Line cells (Signosis, Santa Clara, CA, USA: Cat # SL-0002, T47D-luc)) were grown to 90% confluence in a 550 mL cell culture flask with RPMI 164 medium supplemented with 10% (v/v) Hyclone fetal bovine serum (GE Lifesciences) and Geneticin (75 μg mL −1 , G148, Life Technologies).

Techniques: Concentration Assay, Incubation, Luciferase, Activity Assay, Generated, Extraction, Cell Culture, Control

Journal: Journal of the Endocrine Society

Article Title: Estrogenic activity in tampon products

doi: 10.1210/jendso/bvag094

Figure Lengend Snippet: Estrogenic activity detected in extracts from 8 commercially available tampon brands (A-H). T47D-luc cells were exposed to methanolic extracts prepared from 1 g of core tampon material (string removed) from 8 tampon brands sold in New Zealand. Each 1 g sample was soaked in a 1:1 ethanol:water solution for 4 hours, sonicated, filtered, extracted, and then evaporated to dryness, and reconstituted in methanol. The final extract was diluted into assay medium at 1:1000 (1), followed by serial dilutions to generate 1:2000 (2), 1:4000 (3), and 1:8000 (4) exposure concentrations. Cells were incubated with each dilution for 24 hours, and estrogen-responsive luciferase activity was quantified using a luminescence plate reader. Data are presented as the mean ± SEM of 2 independently extracted tampons per brand, each tested as 2 portions of 1 g each, and each dilution tested with 4 technical replicates.

Article Snippet: Estrogen Receptor Luciferase Reporter T47D Stable Cell Line cells (Signosis, Santa Clara, CA, USA: Cat # SL-0002, T47D-luc)) were grown to 90% confluence in a 550 mL cell culture flask with RPMI 164 medium supplemented with 10% (v/v) Hyclone fetal bovine serum (GE Lifesciences) and Geneticin (75 μg mL −1 , G148, Life Technologies).

Techniques: Activity Assay, Sonication, Incubation, Luciferase, Microplate Reader Luminescence Measurement

Journal: Journal of the Endocrine Society

Article Title: Estrogenic activity in tampon products

doi: 10.1210/jendso/bvag094

Figure Lengend Snippet: Independent batches of Tampon Brand D show estrogenic activity. T47D-luc cells were treated with methanolic extracts generated from 1 g of core tampon material (string removed) from 3 independent batches of Brand A and Brand D. For each batch, 3 separate tampons were extracted independently. Each 1 g sample was soaked in 1:1 ethanol:water solution for 4 hours, sonicated, filtered, extracted, and evaporated to dryness, and reconstituted in methanol. Reconstituted extracts were diluted into assay medium at 1:1000 (1), followed by serial 1:2 dilutions to produce 1:2000 (2), 1:4000 (3), and 1:8000 (4) exposure concentrations. Cells were incubated for 24 hours, and estrogen-responsive luciferase activity was quantified by using a luminescence plate reader. Data are shown as mean ± SEM of 3 independently extracted tampons per batch, tested as 2 portions of 1 g each with each dilution tested as 4 technical replicates.

Article Snippet: Estrogen Receptor Luciferase Reporter T47D Stable Cell Line cells (Signosis, Santa Clara, CA, USA: Cat # SL-0002, T47D-luc)) were grown to 90% confluence in a 550 mL cell culture flask with RPMI 164 medium supplemented with 10% (v/v) Hyclone fetal bovine serum (GE Lifesciences) and Geneticin (75 μg mL −1 , G148, Life Technologies).

Techniques: Activity Assay, Generated, Sonication, Incubation, Luciferase, Microplate Reader Luminescence Measurement

Journal: Journal of the Endocrine Society

Article Title: Estrogenic activity in tampon products

doi: 10.1210/jendso/bvag094

Figure Lengend Snippet: Estrogenic activity of internationally sourced tampon brands. T47D-luc cells were treated with methanolic extracts prepared from intact tampons (string removed) from a panel of internationally sourced tampon brands (Brands I-R). For each brand, 2 or 3 independent tampons were extracted separately on independent days. Each tampon was soaked in a 1:1 ethanol:water solution for 4 hours, sonicated, filtered, extracted, evaporated to dryness, and reconstituted in methanol. Reconstituted extracts were diluted into assay medium at 1:1000, and cells were incubated for 24 hours before estrogen-responsive luciferase activity was quantified using a luminescence plate reader. Data are shown as mean ± SEM of 2 or 3 independently extracted tampons per brand with each dilution tested as 4 technical replicates.

Article Snippet: Estrogen Receptor Luciferase Reporter T47D Stable Cell Line cells (Signosis, Santa Clara, CA, USA: Cat # SL-0002, T47D-luc)) were grown to 90% confluence in a 550 mL cell culture flask with RPMI 164 medium supplemented with 10% (v/v) Hyclone fetal bovine serum (GE Lifesciences) and Geneticin (75 μg mL −1 , G148, Life Technologies).

Techniques: Activity Assay, Sonication, Incubation, Luciferase, Microplate Reader Luminescence Measurement

Journal: Journal of the Endocrine Society

Article Title: Estrogenic activity in tampon products

doi: 10.1210/jendso/bvag094

Figure Lengend Snippet: Effect of sonication on estrogenic activity of tampon extracts. T47D-luc cells were exposed to methanolic extracts prepared from 2 independent tampons per brand (Brands A and D, string removed), comparing extraction with and without sonication. Each tampon was soaked in a 1:1 ethanol:water solution for 4 hours, either sonicated or left nonsonicated, then filtered, extracted, evaporated to dryness, and reconstituted in methanol. Extracts were diluted into assay medium at 1:1000 (1), followed by serial 1:2 dilutions to generate 1:2000 (2) and 1:4000 (3) exposure concentrations. Cells were incubated for 24 hours, and estrogen-responsive luciferase activity was quantified. Data represent mean ± SEM of t2wo independently extracted tampons per condition with each dilution tested as 4 technical replicates.

Article Snippet: Estrogen Receptor Luciferase Reporter T47D Stable Cell Line cells (Signosis, Santa Clara, CA, USA: Cat # SL-0002, T47D-luc)) were grown to 90% confluence in a 550 mL cell culture flask with RPMI 164 medium supplemented with 10% (v/v) Hyclone fetal bovine serum (GE Lifesciences) and Geneticin (75 μg mL −1 , G148, Life Technologies).

Techniques: Sonication, Activity Assay, Extraction, Incubation, Luciferase