Activation of programmed cell death (apoptosis) by cisplatin, other anticancer drugs, toxins and hyperthermia.

Cell death induced by cisplatin was studied in Chinese hamster ovary cell lines, one proficient and the other deficient (100-fold sensitive) in DNA excision repair. Previous experiments demonstrated that cells progressed to and arrested in the G2 phase of the cell cycle before dying. DNA double-strand breaks were detected following G2 arrest and prior to loss of membrane integrity. These DNA breaks have been studied in more detail. DNA fragments were observed consisting of multimers of approximately 180 base pairs. These fragments are consistent with internucleosomal cleavage of chromatin by an endonuclease. At LC90 concentrations, DNA digestion began 48 hr cisplatin treatment followed by loss of membrane integrity and cell shrinkage 24 hr later. High concentrations of cisplatin (170 logs of kill) induced DNA digestion 12 hr after drug treatment but loss of membrane integrity occurred 12 hr later. Both cell death and DNA fragmentation were inhibited by cycloheximide, suggesting the requirement for new protein synthesis. Cells incubated with many other agents demonstrated the same characteristic pattern of DNA degradation. At 90% lethal conditions, DNA digestion was induced within 30 min by hyperthermia, 18 hr by methotrexate, and 48-72 hr by all other agents tested. DNA digestion always preceded loss of membrane integrity and cell shrinkage. These observations are consistent with cell death occurring by the process of apoptosis, or prorammed cell death, and demonstrate the importance of DNA digestion as an early and presumably essential step in cell death. The results suggest that, irrespective of the primary site of action of a drug, cell death by most pharmacologic agents is mediated by activation of the signal transduction pathway for apoptosis. The results also suggest two signal pathways for apoptosis, one directly associated with drug action and a second that requires cell cycle-related events.