Brusatol-induced HL-60 cell differentiation involves NF-κB activation

Brusatol, a quassinoid isolated from the fruit of Brucea javant. ca, induces cell differentiation with various leukemic cell lines in the concentration range of 5-25 ng/ml. To investigate its mechanism of action, cultured HL-60 cells were treated with brusatol (25 ng/ml) for various periods of time and qualitatively analyzed for differential gene expression using a cDNA macroarray. As suggested by these preliminary data, we investigated the effect of brusatol on the Rel/nuclear factor-kappa B (NF-kappaB) family members and their role in HL-60 cell differentiation. When cells were treated with brusatol (25 ng/ml), p100/p52, p105/p50 and p65 mRNA were found to be up-regulated with a maximum after 8 h. As determined by electrophoretic mobility shift assays (EMSA), NF-kappaB was activated, involving p50 and p65. Moreover, immunoblots showed a decrease of IkappaBalpha and generation of the phosphorylated form of IkappaBalpha in whole cell lysates. Cell differentiation induced by brusatol was inhibited by SN50, a NF-kappaB translocation inhibitor. These results strongly suggest that brusatol induces activation of NF-kappaB and the activation and translocation of NF-kappaB into the nucleus is responsible for promoting HL-60 cell differentiation. (C) 2003 Elsevier Ireland Ltd. All rights reserved.