Cell-death by apoptosis following anticancer drug-treatment in-vitro.

The mechanism of cell death, apoptosis or necrosis, was determined morphologically and by DNA gel electrophoresis in 3 human leukaemic T-cell lines (CCRF-CEM.f2, CCRF-HSB and MOLT.4) after treatment with cytotoxic drugs. These include one hormone, dexamethasone (DXM); the DNA damaging agents, melphalan, cisplatin, bleomycin, mitomycin C and mithramycin; inhibitors of DNA synthesis, aphidicolin, cytosine arabinoside (Ara-C), methotrexate (MTX), 5-fluoro-2'-deoxyuridine (FUdR) and 5-fluorouracil (5-FU); and other metabolic inhibitors, bromo-2'-deoxy-2'-uridine (BUdR), actinomycin D, 5-azacytidine (5-AC), cycloheximide, vincristine, etoposide and adriamycin. When cell death was assessed morphologically apoptotic cell death was apparent in the three cell lines 48 hours after all drug treatments. However, a distinct pattern of DNA breakdown was observed for each cell line. A smear of DNA on agarose gels was seen for CCRF-CEM.f2 with 5-FU and mithramycin treatments whilst CCRF-HSB cells showed a similar DNA profile after 5-FU and MTX treatments. All drug treatments of MOLT.4 cells produced a necrotic pattern of DNA degradation. Cycloheximide, an inhibitor of protein synthesis reduced DNA fragmentation of CCRF-CEM.f2 cells treated with DXM, MTX and FUdR indicating that protein synthesis is required for cytotoxicity by apoptosis. However, the extent of DNA fragmentation caused by 5-FU was not significantly affected by cycloheximide. These results indicate that at least morphological and electrophoretic criteria should be used to avoid differing conclusions about modes of cell death.