Ikarugamycin induces DNA damage, intracellular calcium increase, p38 MAP kinase activation and apoptosis in HL-60 human promyelocytic leukemia cells

Abstract

Ikarugamycin (IKA) is an antibiotic with strong antiprotozoal and cytotoxic activity. The purpose of our work was to provide insight into the mechanism of action characterizing the cytotoxic effect of IKA in HL-60 leukemia cells in order to evaluate its potential as an antineoplastic agent. Cell viability was reduced in response to IKA (IC 50 of 221.3 nM), while the amount of HL-60 cells with a subdiploid DNA content increased significantly after 24 h. Apoptotic cell death was confirmed by the cleavage of caspase-9, -8 and -3 using immunoblotting. Single cell gel electrophoresis pointed to an early genotoxic effect. Monitoring of intracellular calcium ([Ca 2+] i) levels by flow cytometric analysis of Fluo-3-AM fluorescence indicated an increase in cytosolic calcium that correlated with the cleavage of caspases. In addition, IKA triggered the activation of p38 MAP kinase which was partly dependent on elevated [Ca 2+] i concentrations and contributed to caspase activation. The data demonstrate that IKA induced apoptosis in HL-60 cells through genotoxicity and caspase activation which was in part correlated to an increase in intracellular calcium levels and activation of p38 MAP kinase.