Fig. 5

Chemo-residual tumor cells exhibit higher DNA repair capability and increased expression/phosphorylation of DNA-dependent protein kinase (DNA-PK _CS ). a Left panel: SUM159 chemo-residual cells and parental (untreated) cells were re-challenged with doxorubicin (Dox) (1 μg/ml) for 3 h. Fresh medium was added after chemotherapy removal. DNA damage at sequential time points after chemotherapy treatment was analyzed by neutral comet assay. Representative images are shown at each time point. Cells scored as comet tail-positive are indicated with red arrows in the 72-h (72h) time frame. Right panel: the percentage of cells with comet tails at the indicated time points was quantified with a fluorescence microscope. Error bars represent SD, n = 3 fields. Significance of data points at 24h, 48h and 72 h was determined relative to data reported at 0h for the indicated cell population (*p <0.05, **p <0.01, ***p <0.001, two-tailed Student’s t test). Cells scored as comet tail-positive are indicated with red arrows in the 72h time frame. b. Left panel: BT549 chemo-residual cells and parental (untreated) cells were challenged with Dox (0.5 μg/ml) for 4 h. DNA damage was assessed at the indicated times using the neutral comet assay as in a. Right panel: the percentage of cells with comet tails at the indicated time points was quantified as in a. Cells scored as comet tail-positive are indicated with red arrows in the 72h time frame. c Left and middle panel: SUM159 cells (left) and BT549 cells (middle) were treated with Dox as described in Fig. 1a. Nuclear protein from parental and chemo-residual cells was extracted. Equivalent amounts were immunoblotted with phospho (Ser 2056)-DNA-PK_CS, DNA-PK_CS or Lamin A antibody. Right panel: protein bands from three independent trials (SUM159 cells treated with Dox as described in Fig. 1a) were quantified, and the relative ratio of DNA-PK_CS to loading control is shown. Error bars represent SD, n = 3, **p <0.01, two-tailed Student’s t test