Figure 5: Multinucleated BRCA2-deficient cells arising from failed chromatin bridge resolution after PARP inhibitor treatment are not viable.
(a,b) HeLa cells were transfected with siRNA targeting BRCA2 for 24 h and were then treated with olaparib (0.5 μM) or DMSO for 72 h. Cells were then incubated with Hoechst for 45 min at 37 °C, after which cells containing 2n, 4n or >4n were sorted as shown in a. Subsequently, cells were sorted at a density of 5,000 cells per well in six-wells plates. After 7 days, colony formation was quantified. The graph shows averages and s.d.'s from three replicates. P values were calculated using two-tailed Student’s t-test. (c) KB2P1.21 (Brca2−/−) cells were treated, sorted and stained as described for a and b. After 7 days, colony formation was quantified. The graph shows means with error bars indicating s.d. of three replicates. P values are calculated using two-tailed Student’s t-test. (d) H&E staining of a Brca2−/−;p53−/− mammary tumour derived from a tumour-bearing mouse, treated with vehicle or olaparib (50 mg kg−1) for 21 days i.p. daily. Arrowheads indicate multinucleated cells. Scale bars represent 100 μm. (e) Quantification of the percentage of multinucleated cells in tumours described in d. (f) H&E staining of a Brca1−/−;p53−/− tumour derived from a tumour-bearing mouse, treated with vehicle or olaparib (50 mg kg−1) for 21 days i.p. daily. Multinucleated cells and apoptotic cells are indicated. Scale bars represent 100 μm. (g) Quantification of the percentage of multinucleated cells in tumours described in f. P values were calculated using two-tailed Student’s t-test. Throughout the figure ‘NS’ indicates not significant.
