1. ¹Centro Andaluz de Biología Molecular y Medicina Regenerativa, Consejo Superior de Investigaciones Cientificas (CSIC), Avda Américo Vespucio s/n, 41092 Sevilla, Spain;
2. ²Department of Hematology, Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy

Correspondence: Abelardo López-Rivas, Tel: +34-95-446-7997; Fax: +34-95-446-1664 E-mail: abelardo.lopez@cabimer.es

Received 17 October 2007; Revised 3 December 2007; Accepted 11 December 2007.

Abstract

The tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL/APO2L) is a member of the TNF gene superfamily that induces apoptosis upon engagement of cognate death receptors. While TRAIL is relatively non-toxic to normal cells, it selectively induces apoptosis in many transformed cells. Nevertheless, breast tumor cells are particularly resistant to the effects of TRAIL. Here we report that, in combination with the cyclin-dependent kinase inhibitor roscovitine, exposure to TRAIL induced marked apoptosis in the majority of TRAIL-resistant breast cancer cell lines examined. Roscovitine facilitated TRAIL death-inducing signaling complex formation and the activation of caspase-8. The cFLIP_L and cFLIP_S FLICE-inhibitory proteins were significantly down-regulated following exposure to roscovitine and, indeed, the knockdown of cFLIP isoforms by siRNA sensitized breast tumor cells to TRAIL-induced apoptosis. In addition, we demonstrate that roscovitine strongly suppressed Mcl-1 expression and up-regulated E2F1 protein levels in breast tumor cells. Significantly, the silencing of Mcl-1 by siRNA sensitized breast tumor cells to TRAIL-induced apoptosis. Furthermore, the knockdown of E2F1 protein by siRNA reduced the sensitizing effect of roscovitine in TRAIL-induced apoptosis. In summary, our results reveal a pleitropic mechanism for the pro-apoptotic influence of roscovitine, highlighting its potential as an antitumor agent in breast cancer in combination with TRAIL.