Similarly, silencing of survivin expression in MDA-MB-231 (p53 mut) and PC-3 (p53 null) cells activates caspase-3 (Fig. 6), a hallmark of apoptosis. Epigenetics inhibitor These
studies provide direct evidence for the involvement of survivin expression in bortezomib resistance. Figure 5 Effects of silencing of survivin expression on bortezomib sensitivity in HCT116p53-/- cells. The highly survivin expressing HCT116p53-/- cells at 50% confluence were transfected with survivin mRNA-specific siRNAs or with control siRNAs. After 16 hours post transfection, cells were treated with and without bortezomib for 48 hours. A part of the transfected cells were then collected for western blots to determine survivin expression (A), a part of the transfected cells was used to determine cell growth inhibition by MTT assay (B), and the other part of the transfected cells was used to determine cell death/DNA fragmentation by cell death ELISA assay (C). Data shown in B and C are the mean ± SD derived from three independent determinations. Note: Results from cells without transfection were similar to cells transfected with control siRNA/shRNA (not shown). The expression of survivin in HCT116p53-/- cells was set at 10 and relative survivin expression levels are shown after normalized to
actin. Figure 6 Effects of silencing Entospletinib of survivin expression on bortezomib sensitivity in other cancer cell with mutant p53. Cell treatment condition is the same as in Figure 5. Cells were then collected for western blots to determine survivin expression and/or caspase-3
activation. Rho A, MDA-MB-231 breast cancer cells are with mutant p53. B, PC-3 prostate cancer cells are with p53 null. Cancer cell sensitivity to bortezomib treatment is dependent on p53 Alpelisib status but not cancer cell types Previous studies indicated that modulation of survivin expression by bortezomib, and cancer cell sensitivity to bortezomib-induced apoptosis are cell type-dependent . Based on the data provided above, we hypothesized that the different sensitivity to bortezomib for cancer cells is due to p53 status-associated differential survivin expression, and induction by bortezomib, rather than cancer cell type. Here, we tested four pairs of cancer cell lines with different p53 status from lung cancer (EKVX with mutant p53 versus A549 with wild type p53), breast cancer (MDA-MB-231 with mutant p53 versus MCF-7 with wild type p53), prostate cancer (PC-3 with null p53 versus LNCaP with wild type p53) and myeloma (RPMI-8226 with mutant p53 versus Kms11 with wild type p53). Consistent with our early data and our rationale, bortezomib-mediated inhibition of cell growth is significantly better in cancer cell lines with wild type p53 in comparison to those cell lines with a p53 null or p53 mutant status (Fig. 7), which is consistent with the relative expression level of survivin in these cells (Fig. 3A and 3B). Figure 7 p53 status but not cancer cell type is a critical indicator for bortezomib sensitivity.