Fig. 5

Synergistic effects of combining trastuzumab with Mcl-1-targeting approaches in trastuzumab-sensitive Her2-positive BC cells. a Genetic depletion of Mcl-1 or Her2 induces cell death in Her2-positive BC cells, but not benign breast cells (MCF-10A). BC and MCF-10A cells were transfected with siMCL1 (filled bars) or siHER2 (open bars) for 30 hours and then exposed to hypoxia for 2 days. Cell survival was determined by AlamarBlue^® assay. Data represent mean ± SD for triplicate samples. Results shown are representative of three independent experiments. b Synergistic effects of combining trastuzumab with siMCL1 in Her2-positive BC cells. BT-474 cells (5.5 × 10³ cells) were seeded per well in agar-coated 96-well plates prior to siMCL1 transfection. Spheroid formation was assessed in siMCL1-treated and/or trastuzumab-treated and control cells using an inverted fluorescence light microscope. Photographs (10× magnification) of spheroid formation are representative of each group and three independent experiments (left). Spheroid volumes were calculated as described in Materials and methods. Data represent mean ± SD (right). c Synergistic growth inhibition of trastuzumab and EU-5346 in trastuzumab-sensitive Her2-positive BC cells under hypoxic conditions. BT-474 cells were pretreated with trastuzumab or dimethyl sulfoxide (DMSO) for 24 hours under normoxic conditions followed by EU-5346 treatment or DMSO for 72 hours under hypoxic conditions. [³H]-thymidine was added during the last 8 hours. Data represent mean ± SD for triplicate samples. Results shown are representative of three independent experiments. Trastuzumab (white bars), EU-5346 (gray bars), and drug combination (black bars). Combination index (CI) was calculated using the Combosyn software as described in Materials and methods. a, b *p <0.05 and **p <0.001 by Student’s t test