Figure 3.

Blockade of CHK1 phosphorylation by VE‐821 (ATR inhibitor) and radio‐potentiation. A. Effect of VE‐821 on breast cancer cell growth. For clonogenic assays, data are mean ± standard deviation from three separate experiments in MCF7 (B) and MDA‐MB‐231 (C). D. LD50 values (dose of IR where 50% of cells no longer survive) were calculated from each experiment and the potentiation factor at 50% cell kill (PF50) was calculated (see Supplementary Table S3 for further details). E. Exponentially growing MCF7 or MDA‐MB‐231 were seeded into 10 cm tissue culture dishes and allowed to adhere for 24 h. Where indicated, cells were pre‐treated with 1 μM VE‐821 for 1 h before IR (2 Grays) or mock treatment. After 24 h, media from cells was collected, the cells washed and harvested and fixed and frozen in ice cold methanol overnight. Cells were stained with 200 μg/ml PI and 200 μg/ml RNAase A in PBS and samples run using a FACScalibur counting a minimum of 20000 events. Single cell populations were then gated into phases of the cell cycle by DNA content. The resulting FACS profiles analysed using Cyflogic software (mean ± S.D. of three individual experiments) is show here.