2-Chloro-2'-deoxyadenosine-induced apoptosis in T leukemia cells is mediated via a caspase-3-dependent mitochondrial feedback amplification loop

Abstract

Abstract. 2-Chloro-2'-deoxyadenosine (CdA; cladribine) is a chemotherapeutic agent used in the treatment of certain leukemias. However, the signalling events that govern CdAmediated cytotoxicity in leukemia cells remain unclear. We show here that CdA treatment caused Jurkat human T leukemia cells to die via apoptosis in a dose-and time-dependent fashion. Bcl-2 overexpression protected Jurkat T leukemia cells from CdA-induced apoptosis and loss of mitochondrial transmembrane potential (ΔΨ m ). Furthermore, mitochondria that were isolated from Jurkat T leukemia cells and then exposed to CdA showed a loss of ΔΨ m , indicating that CdA directly compromised outer mitochondrial membrane integrity. CdA treatment of Jurkat T leukemia cells resulted in the activation of caspase-3, -8, and -9, while inhibition of these caspases prevented the CdA-induced loss of ΔΨ m , as well as DNA fragmentation. In addition, caspase-3 inhibition prevented caspase-8 activation while caspase-8 inhibition prevented caspase-9 activation. Death receptor signalling was not involved in CdA-induced apoptosis since cytotoxicity was not affected by FADD-deficiency or antibody neutralization of either Fas ligand or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Taken together, these data suggested that CdA-induced apoptosis in Jurkat T leukemia cells was mediated via a caspase-3-dependent mitochondrial feedback amplification loop. CdA treatment also increased p38 mitogen-activated protein (MAPK) and extracellular signal-regulated kinase 1 and 2 (ERK1/2) phosphorylation in Jurkat T leukemia cells. Although ERK1/2 inhibition did not affect CdA-mediated cytotoxicity, inhibition of p38 MAPK had an enhancing effect, which suggested a cytoprotective function for p38 MAPK. Agents that inhibit p38 MAPK might therefore increase the effectiveness of CdA-based chemotherapy.