Figure 5. Bosutinib isomer is a potent sensitizer of gemcitabine in pancreatic cancer xenografts. (A) A patient derived pancreatic cell line, EGF-1, was treated with gemcitabine (100 nM) for 24h before the addition of UCN-01 (100 nM) or Bos-I (1 μM) for a further 9 h. Cells were then fixed and immunostained with α-tubulin (red) and DAPI (blue). Quantification of the mitotic index from a minimum of 100 cells is provided. (B) EGF-1 cell viability was assessed after treatment with increasing concentrations of gemcitabine for 24 h followed by Bos-I (1 μM) for a further 48 h before addition of MTS reagent. (C) Xenograft studies using EGF-1 cells to test combinatorial treatment were performed. Tumors were allowed to grow to 100mm³ before being randomized and treated with either vehicle, gemcitabine alone (50 mg/kg), Bos-I alone (100 mg/ml) or a combination of the two. The tumor growth index was used to determine efficacy of drug treatments. *P = 0.013 compared with vehicle treated animals on day 17. **P = 0.006 compared with gemcitabine-treated animals at day 17. ***P < 0.001 compared with Bos-I-treated animals at day 17. ^†P < 0.001 compared with either Bos-I or gemcitabine treated animals. Data are presented as the mean ± SEM (D) The average γH2AX intensity per tumor cell was determined via IHC staining after 1 round of treatments. A minimum of 100 cells were quantified and the data are presented as average ± SEM *P = 0.01, **P = 0.0001 compared with vehicle treated animals. (E) As for (D), except the average γH2AX intensity per mitotic tumor cell is shown ± SEM. A minimum of 25 mitotic cells were quantified. *P = 0.0015.