Inhibition of cyclin-dependent kinase 1 (CDK1) by indirubin derivatives inhuman tumour cells

The bisindole indirubin has been described, more than 30 years ago, as bein g clinically active in the treatment of human chronic myelocytic leukaemia. However, the underlying mechanism of action has remained unclear. We have reported previously that indirubin and its analogues are potent and selecti ve inhibitors of cyclin-dependent kinases (CDK). In this study, we investig ated the influence of indirubin and derivatives on CDK1/cyclin B kinase in human tumour cells at concentrations known to induce growth inhibition. Cel ls of the mammary carcinoma cell line MCF-7, synchronized by serum deprivat ion. after serum repletion stay arrested in the G(1)/G(0) phase of the cell cycle in the presence of 2 muM indirubin-3'-monoxime. At higher drug conce ntrations (greater than or equal to 5 muM) an increase of the cell populati on in the G(2)/M phase is additionally observed. Cells synchronized in G(2) /M phase by nocodazole remain arrested in the G(2)/M phase after release, i n the presence of indirubin-3'-monoxime (greater than or equal to5 muM). Af ter 24 h treatment with 10 muM indirubin-3'-monoxime a sub-G(2) peak appear s. indicative for the onset of apoptotic cell death. Treatment of MCF-7 cel ls with growth inhibitory concentrations of indirubin-3'-monoxime induces d ose-dependent inhibition of the CDK1 activity in the cell. After 24 h treat ment. a strong decrease of the CDK1 protein level along with a reduction of cyclin B in complex with CDK1 is observed. Taken together, the results of this study strongly suggest that inhibition of CDK activity in human tumour cells is a major mechanism by which indirubin derivatives exert their pote nt antitumour efficacy. (C) 2001 Cancer Research Campaign.