The best cell inoculum density for the determination of doxorubicin sensitivity was preliminarily defined by the sulforhodamine B (SRB) assay17. Briefly, DHSF-BR16 cells were seeded onto 96-well microplates (Sarstedt) at a density of 1.0, 1.5, 2.0, 3.0 × 10cells/well. After 24 h of incubation at 37 °C in a humidified atmosphere with 5% CO2, the culture medium was replaced with fresh medium and doxorubicin was added at final drug concentrations varying from 3 × 10–8 to 1 × 10–4 M. After 72 h of incubation, the assay was terminated by the addition of cold TCA and the cells were stained according to the procedure reported in Coronnello et al.17 to determine the IC50 values. After choosing the optimal cell density, the sensitivity of DHSF-BR16 and MCF-7 cells (same cell density/well) to epirubicin, docetaxel and paclitaxel was evaluated at pharmacological concentrations ranging from 1 nM to 10 µM for epirubicin and from 0.003 nM to 0.1 µM for taxanes. The pharmacological sensitivity of the study cell line was compared with that of the MCF-7 reference line and was indicated by the resistance index (RI) obtained from the ratio between the IC50 values of the two cell lines for each individual drug. Respect to MCF-7 line, RI values higher than 1 indicate resistance of DHSF-BR16 cells to the tested drug and values lower than 1 indicate drug sensitivity. Cytotoxicity data are represented as mean ± standard error (SE).

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