Inhibition of cell-cycle progression in human promyelocytic leukemia HL-60 cells by MCS-C2, novel cyclin-dependent kinase inhibitor

Abstract

To elucidate the action mechanism of MCS-C2, a novel analogue of toyocamycin and sangivamycin, its effect on the expression of cell cycle-related proteins in the human myelocytic leukemia cell line HL-60 was examined using Western blotting and a flow cytometric analysis. MCS-C2, a selective inhibitor of cyclin-dependent kinases, was found to inhibit cell growth in a time- and dose-dependent manner, and inhibits cell cycle progression by inducing the arrest at G1 and G2/M phases, in HL-60 cells. The flow cytometric analysis revealed an appreciable arrest of cells in the G2/M phase of the cell cycle after treatment with MCS-C2. The HL-60 cell population increased gradually from 13% at 0 h, to 28% at 12 h in the G2/M phase, after exposure to 2 muM MCS-C2. Furthermore, Western blot analysis demonstrated that MCS-C2 induced the cell cycle arrest at G1 phase through the inhibition of pRb phosphorylation. Hypophosphorylated pRb accumulated after treatment with 5 muM MCS-C2 for 12 h, whereas. the level of hyperphosphorylated pRb was reduced. Thus. treatment of the cell with MCS-C2 Suppressed the hyperphosphorylated form of pRb with a commensurate increase in the hypophosphorylated form.