¹Drug Resistance Group, Centre for Cancer Research and Cell Biology, Queen's University Belfast, Belfast, UK

Correspondence: Dr DB Longley, Centre for Cancer Research and Cell Biology, Queen's University Belfast, University Floor, Belfast City Hospital, Lisburn Road, Belfast BT9 7AB, UK. E-mail: d.longley@qub.ac.uk

¹These authors contributed to this work equally.

Received 10 May 2005; Revised 9 August 2005; Accepted 17 August 2005; Published online 17 October 2005.

Abstract

c-FLIP inhibits caspase 8 activation and apoptosis mediated by death receptors such as Fas and DR5. We studied the effect of c-FLIP on the apoptotic response to chemotherapies used in colorectal cancer (CRC) (5-fluorouracil, oxaliplatin and irinotecan). Simultaneous downregulation of both c-FLIP splice forms c-FLIP_L and c-FLIP_S with siRNA synergistically enhanced chemotherapy-induced apoptosis in p53 wild-type (HCT116p53^+/+, RKO), null (HCT116p53^-/-) and mutant (H630) CRC cell lines. Furthermore, overexpression of c-FLIP_L, but not c-FLIP_S, potently inhibited apoptosis induced by chemotherapy in HCT116p53^+/+ cells, suggesting that c-FLIP_L was the more important splice form in mediating chemoresistance. In support of this, siRNA specifically targeted against c-FLIP_L synergistically enhanced chemotherapy-induced apoptosis in a manner similar to the siRNA targeted against both splice forms. Inhibition of caspase 8 blocked the enhanced apoptosis induced by c-FLIP-targeted (FT) siRNA and chemotherapy. Furthermore, we found that downregulating cell surface DR5, but not Fas, also inhibited apoptosis induced by FT siRNA and chemotherapy. Interestingly, these effects were not dependent on activation of DR5 by its ligand TRAIL. These results indicate that c-FLIP inhibits TRAIL-independent, DR5- and caspase 8-dependent apoptosis in response to chemotherapy in CRC cells. Moreover, targeting c-FLIP in combination with existing chemotherapies may have therapeutic potential for the treatment of CRC.