Fig. 6
Effect of combined treatment with AMD3100 and docetaxel on acquired trastuzumab-resistant breast tumor growth. A, C BTRT or SKRT cells were grown in 3D Matrigel culture and treated with AMD3100 (5 µM), docetaxel (10 nM), or the combination. SDF-1α (4 ng/ml) was added at the same time. Photographs were taken on day 9. B, D Quantitative analysis of total acini area was performed using AlphaView SA, and the data were analyzed using one-way ANOVA. The data are reported as mean ± SD of triplicates, representing two independent experiments (*P < 0.0001 compared with vehicle). E BTRT cells grown in 3D Matrigel were treated with serial doses of AMD3100 (AMD) and/or serial doses of docetaxel. SDF-1α (4 ng/ml) was added at the same time. The combined effect of AMD3100 and docetaxel on acini growth was analyzed using CalcuSyn Dose Effect Analyzer. F HR6 cells, derived from BT474 cells and exhibiting in vivo acquired trastuzumab resistance, were implanted into the mammary fat pad of female athymic nude mice. When the tumor size reached 100 mm, the mice were randomized to treatment with vehicle, trastuzumab, AMD3100, docetaxel, or different combinations. Tumor volume was calculated using the formula V = lw2/2. Data were analyzed using two-way ANOVA (*P < 0.0001 compared with vehicle, #P < 0.0001 compared with AMD3100 alone, & P < 0.0001 compared with docetaxel alone). G BTRT cells were treated with docetaxel (5 nM) followed by BrdU pulse. Immunofluorescent staining for CXCR4, BrdU, and 7-AAD was performed and followed by flow cytometry analysis (“Materials and methods” section). The correlation between CXCR4 and 7-AAD or BrdU was analyzed using Pearson r coefficients. H BTRT cells were treated with docetaxel or vehicle. Dynamic expression of CXCR4 with cell cycle progression in BTRT cells was detected by flow cytometry