EMT-Related Gene Expression in MCF-7 Cells Treated with Monocarbonyl Analogs of Curcumin C66 and B2BrBC Following EMT Induction – RT-qPCR Array Dataset

Description

Monocarbonyl analogs of curcumin (MACs), C66 [(2E,6E)-2,6-bis[(2-trifluoromethyl)benzylidene]cyclohexanone] and B2BrBC [(2E,6E)-2,6-bis(2-bromobenzylidene)cyclohexanone], were synthesized at the Institute of Chemistry, Ss. Cyril and Methodius University in Skopje, North Macedonia, following the previously published method (DOI: 10.17632/vdgz7pk6vh.1). MCF-7 human breast cancer cells (ATCC, Manassas, VA, USA; Cat. # HTB-22) were cultured in DMEM/F12 (50:50) medium (Corning Life Sciences, Corning, NY, USA; Cat. # 10-092-CM) supplemented with 10% FBS (Avantor, Radnor, PA, USA; Cat. # 89510-186) and antibiotic/antimycotic mixture (Corning Life Sciences, Corning, NY, USA; Cat. # 30-004-Cl) at 37 °C in 5% CO2 with 95% atmospheric air. For experiments, cells were seeded into 6-well plates (0.3 x 106 cells per well) in complete tissue culture media, supplemented with StemXVivo epithelial-to-mesenchymal transition (EMT)‑inducing media supplement (Bio‑Techne, Minneapolis, MN, USA; Cat. # CCM017), containing TGFβ1, Wnt5a, and antibodies against E‑cadherin, sFRP1, and Dkk‑1. After 48 h of incubation, the medium was replaced with pre‑warmed complete medium, and fresh EMT supplement was added along with either vehicle (DMSO), C66, or B2BrBC (100 µM each) for an additional 72 h. MCF-7 cells cultured in medium without a StemXVivo EMT Inducing Media Supplement, but with vehicle alone, served as the control. Total RNA was isolated using TRIzol reagent (Invitrogen, Thermo Fisher Scientific, Waltham, MA, USA; Cat. # 15596026) and quantified with a NanoDrop One spectrophotometer (Thermo Fisher Scientific, Wilmington, DE, USA). RNA samples were normalized to 1 µg total RNA and reverse transcribed into cDNA using qScript cDNA SuperMix (Quantabio, Beverly, MA, USA; Cat. # 95048) on a SimpliAmp Thermal Cycler (Applied Biosystems, Thermo Fisher Scientific, Carlsbad, CA, USA). qPCR was carried out using the Human Epithelial to Mesenchymal Transition PCR Array kit (QIAGEN, Hilden, Germany; Cat. # 330231 PAHS‑090ZA) with PerfeCTa SYBR Green FastMix (Quantabio, Beverly, MA, USA; Cat. # 95072) on a QuantStudio 3 Real‑Time PCR System (Applied Biosystems, Thermo Fisher Scientific, Carlsbad, CA, USA) under the following conditions: 2 min at 50 °C, 10 min at 95 °C, followed by 40 cycles of 15 s at 95 °C and 1 min at 60 °C, and a final dissociation stage of 15 s at 95 °C, 1 min at 60 °C, and 15 s at 95 °C. Cycle threshold (Ct) values were determined using QuantStudio Design & Analysis Software (Applied Biosystems, Thermo Fisher Scientific).

Institutions

• Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

  NY, Hempstead
• Feinstein Institute for Medical Research

  NY, Manhasset
• Northwell Health

  NY, Glen Cove
• Saints Cyril and Methodius University in Skopje Faculty of Natural Sciences and Mathematics

  Skopje, Skopje

Categories

Funders

• Gerald J. and Dorothy R. Friedman New York Foundation for Medical Research

Related Links

Licence