Fig. 5

Lethality mediated by nutlin-3 + THZ1 required for effective p53. A MCF-7 cell was firstly given PFT β for 6 h, then treated with 50 M THZ1and 20uM nutlin-3. CCK8 assay was used to measure the 24 h or 48 h cell viability after combination was added. B Colony formation ability measurement of MCF-7 cell after a 6 h PFT β pretreatment, then added with 50 nM THZ1 + 20uM nutlin-3. C Quantitation of colony formation ability in B. D MCF-7 cell was firstly transfected with WT p53 plasmids or plasmids of control. After a 6 h incubation with PFT β, 50 M THZ1 was added into the medium of MCF-7 cell with WT p53 plasmids. CCK8 assay was used to measure the 24 h or 48 h cell viability after THZ1 was added. E Colony formation ability measurement of MCF-7 cell with WT p53 plasmids or plasmids of control, then treated with 50 nM THZ1. F Quantitation of colony formation ability in E. G CCK8 assay was used to measure the cell viability of SK-BR-3, MDA-MB-231, HS578T, HCC1937 cells under 50 M THZ1and 20uM nutlin-3 treatment for 24 h or 48 h respectively. H Western blot was meant to detect p53 and b-actin protein levels in SK-BR-3 cell which was treated with 50 nM THZ1 for 48 h after WT p53 plasmids transfection. I Quantity of P53 in H. J CCK8 assay was used to measure the cell viability of SK-BR-3 cell under 50 M THZ1 treatment for 24 h or 48 h respectively. K Western blot was meant to detect p53 and b-actin protein levels in MDA-MB-231 cell which was treated with 50 nM THZ1 for 48 h after WT p53 plasmids transfection. L Quantity of P53 in K. M CCK8 assay was used to measure the cell viability of MDA-MB-231 cell under 50 M THZ1 treatment for 24 h or 48 h respectively. N, O, P, Q Western blot was conducted to detect p53 protein levels in MCF-7 cell which was treated with increasing concentrations of THZ1 or nutlin-3 for 48 h. R, S, T, U, V, W, X, Y Western blot was conducted to detect p53 protein levels in nucleus, cytoplasm, cytosol or mitochondria of MCF-7 cell which was treated with 50 nM THZ1or 20uM nutlin-3 or combination for 48 h. Statistically significant (P < 0.05) are *, (P < 0.01) are **