Original Paper
Cell Death and Differentiation (2007) 14, 1813–1825. doi:10.1038/sj.cdd.4402177; published online 22 June 2007
LY303511 amplifies TRAIL-induced apoptosis in tumor cells by enhancing DR5 oligomerization, DISC assembly, and mitochondrial permeabilization
Edited by J Tschopp
T W Poh1,4, S Huang1,4, J L Hirpara1 and S Pervaiz1,2,3
- 1Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore
- 2NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore 117597, Singapore
- 3Singapore-MIT Alliance, National University of Singapore, Singapore 117597, Singapore
Correspondence: S Pervaiz, Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, 2 Medical Drive, Bldg MD9, Singapore 117597, Singapore. Tel: +65 9661 4732; Fax: +65 6778 8161; E-mail: phssp@nus.edu.sg
4These two authors contributed equally to this work
Received 23 November 2006; Revised 2 May 2007; Accepted 2 May 2007; Published online 22 June 2007.
Abstract
Certain classes of tumor cells respond favorably to TRAIL due to the presence of cell surface death receptors DR4 and DR5. Despite this preferential sensitivity, resistance to TRAIL remains a clinical problem and therefore the heightened interest in identifying compounds to revert tumor sensitivity to TRAIL. We recently demonstrated that the phosphatidylinositide-3-kinase (PI3K) inhibitor, LY294002, and its inactive analog LY303511, sensitized tumor cells to vincristine-induced apoptosis, independent of PI3K/Akt pathway. Intrigued by these findings, we investigated the effect of LY303511 on TRAIL-induced apoptosis in HeLa cells. Preincubation of cells with LY30 significantly amplified TRAIL signaling as evidenced by enhanced DNA fragmentation, caspases 2, 3, 8, and 9 activation, and reduction in the tumor colony formation. This increase in TRAIL sensitivity involved mitochondrial membrane permeabilization resulting in the egress of cytochrome c and second mitochondrial activator of caspase/direct IAP-binding protein with low PI, cleavage of X-linked inhibitor of apoptosis protein, and activation of caspase 9. We link this execution signal to the ability of LY30 to downregulate cFLIPS and oligomerize DR5, thus facilitating the signaling of the death initiating signaling complex. The subsequent exposure to TRAIL resulted in processing/activation of caspase 8 and cleavage of its substrate, the BH3 protein Bid. These data provide a novel mechanism of action of this small molecule with the potential for use in TRAIL-resistant tumors.
Keywords:
LY30, TRAIL, DR5, DISC, mitochondria, apoptosis
Abbreviations:
TRAIL, TNF-related apoptosis-inducing ligand; DR4, death receptor 4; DR5, death receptor 5; DISC, death-inducing signaling complex; FADD, Fas-associated death domain-containing protein; PI3K, phosphatidylinositide-3-kinase; Cyt c, cytochrome c; DiOC6, 3,3'-dihexyloxacarbocyanine iodide; MOMP, mitochondrial outer membrane permeablization; MPT, mitochondrial permeability transition; 
m, mitochondrial transmembrane potential; XIAP, X-linked inhibitor of apoptosis protein; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide; cFLIP, cellular FLICE-like inhibitory protein; CM-H2DCFDA, 5-(and-6)-chloromethyl-2',7'-dichlorofluorescin diacetate; DTSSP, 3,3'-dithiobis sulfosuccinimidylpropionate; ROS, reactive oxygen species; Smac/Diablo, second mitochondrial activator of caspase/direct IAP-binding protein with low pI
