Strong Activation of p53 by Actinomycin D and Nutlin-3a Overcomes Resistance of Cancer Cells to the Pro-apoptotic Activity of FAS Ligand

The p53 protein activates pro-apoptotic gene FAS, which encodes death receptor for the FAS ligand (FASLG). Cancer cells are resistant to apoptosis triggered by FASLG. We found that actinomycin D and nutlin-3a (ActD+Nut3a) synergized in activation of p53 and in the induction of pro-apoptotic genes; however, the apoptotic cells were infrequent. We hypothesized that this drug combination sensitizes cancer cells to the pro-apoptotic activity of FASLG. We exposed various cancer cell lines to ActD+Nut3a for 45h and next we treated cells with recombinant FASLG. We observed apoptosis by flow cytometry and by activation status of caspase-3, -8, -9, and -10. The cell viability was determined by the MTS assay and cell staining on culture plates. Actinomycin D and nutlin-3a strongly synergized in sensitizing cells to apoptosis triggered by FASLG. This combination killed more than 99% of cells within 5h. The cell death was accompanied by a strong activation of all examined caspases. In engineered p53-deficient cells this pro-apoptotic effect was completely lost. Therefore, the combination of ActD+Nut3a activates p53 in a way, which overcomes the resistance of cancer cells to apoptosis triggered by FASLG.