Establishing Tamoxifen-Resistant Breast Cancer Cell Lines

Human ER-positive breast cancer cell lines MCF7 and T47D (ATCC), and HEK293T (ATCC) cells were cultured in DMEM (Hyclone) with 10% fetal bovine serum (Gibco) and 1% penicillin–streptomycin (Hyclone) at 37 °C in a humidified 5% CO₂ atmosphere. Tamoxifen-resistant (TamR) MCF7 and T47D cells were generated as described previously [63 (link)]. In brief, cells were cultured in the presence of increasing concentrations of 4-hydroxytamoxifen (Sigma-Aldrich) starting at 0.5 μM, and finally gradually increased up to 5 μM 4-hydroxytamoxifen when the growth of the cells could not be inhibited in this concentration. In parallel, parental MCF7 and T47D cells were cultured under identical conditions without tamoxifen. SP600125, etomoxir and puromycin was obtained from MedChem Express.