Background: Daunorubicin is used clinically in the treatment of myeloma, acute lymphatic and myelocytic leukaemia. The toxic lesions caused by daunorubicin induce various modes of cell death, including apoptosis. Apoptosis is highly regulated programmed cell death that can be initiated mainly via two pathways, through death receptors (extrinsic) or involvement of the mitochondria (intrinsic). Induction of apoptosis via these pathways has been alluded following treatment with daunorubicin, but never compared in acute lymphoblastic leukaemia over a time course.
Methods: This study investigated the mechanisms of daunorubicin induced apoptosis in the treatment of CCRF-CEM, MOLT-4 (acute T-lymphoblastic leukaemia) and SUP-B15 (acute B-lymphoblastic leukaemia) cells. Cells were treated with daunorubicin for 4 h, and then placed in recovery medium (without daunorubicin) for 4 h, 12 h and 24 h. Apoptotic response was analysing using annexin-V expression, caspase activity, mitochondrial membrane potential change and an array to detect 43 apoptotic proteins.
Results: Daunorubicin induced apoptosis in all leukemic cell lines, but with different levels and duration of response. Both apoptosis levels and caspase activity increased after four hours recovery then declined in CCRF-CEM and MOLT-4 cells. However, SUP-B15 cells displayed initially comparable levels but remained elevated over the 24 h assessment period. Changes in mitochondrial membrane potential occurred in both MOLT-4 and CCRF-CEM cells but not in SUP-B15 cells. Expression of apoptotic proteins, including Bcl-2, Bax, caspase 3 and FADD, indicated that daunorubicin potentially induced both extrinsic and intrinsic apoptosis in both CCRF-CEM and MOLT-4 cells, but only extrinsic apoptosis in SUP-B15 cells.
Conclusions: This study describes variations in sensitivities and timing of apoptotic responses in different leukaemia cell lines. These differences could be attributed to the lack of functional p53 in coordinating the cells response following cytotoxic treatment with daunorubicin, which appears to delay apoptosis and utilises alternative signalling mechanisms that need to be further explored.
Keywords: Apoptosis; Caspases; Daunorubicin; Leukaemia; Mitochondrial membrane potential.