LY303511 amplifies TRAIL-induced apoptosis in tumor cells by enhancing DR5 oligomerization, DISC assembly, and mitochondrial permeabilization

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 cFLIP S 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. Tumor necrosis factor-related apoptosis inducing ligand (TRAIL) has been shown to induce apoptosis preferentially in tumor cells. 1,2 The selective responsiveness of TRAIL is attributed to the increased surface expression of TRAIL receptors (DR4 and DR5) on tumor cells. [3][4][5][6] In contrast, nontransformed cells express decoy receptors that provide a mechanism for evading apoptotic signaling initiated by TRAIL, and hence its tumor selectivity. 7,8 Unfortunately, as with most chemotherapeutic compounds, TRAIL-responsive tumors acquire a resistant phenotype which renders TRAIL therapy ineffective. 9,10 This has stimulated an enormous interest in identifying small molecule compounds that, when used in combination with TRAIL could sensitize tumor cells to TRAILinduced apoptosis. The desired consequence would be the need for a much lower therapeutic dose of TRAIL and at the same time an increase in the efficacy of the sensitizing drug. To that end, various groups have demonstrated that a variety of compounds and proteins (either upon silencing or upregulation) sensitize several classes of tumor cells to TRAIL-induced apoptosis. 11,12 LY303511 (LY30) is an inactive analog of LY294002 (LY29), a widely used inhibitor of the phosphatidylinositide-3-kinase (PI3K)/Akt survival pathway. 13 Previously, LY30 has been purported to have no effect on cells in contrast to its active counterpart, LY29. However, recent studies from our laboratory and other groups have demonstrated that LY30 does have a...

LY303511 Enhances TRAIL Sensitivity of SHEP-1 Neuroblastoma Cells via Hydrogen Peroxide–Mediated Mitogen-Activated Protein Kinase Activation and Up-regulation of Death Receptors

“…We linked the apoptosis sensitizing effect of these compounds to an increase in intracellular hydrogen peroxide (H 2 O 2 ) production and amplification of the mitochondrial death pathway. More recently, we showed DR5 oligomerization and amplification of TRAIL signaling in ovarian cancer cells upon exposure to LY30 (19). Stimulated by these findings, we investigated the effect of preincubation of SHEP-1 cells with LY30 on TRAIL-induced apoptosis and questioned the specific role of intracellular H 2 O 2 on apoptosis execution.…”

Multivalent DR5 Peptides Activate the TRAIL Death Pathway and Exert Tumoricidal Activity

“…It is well established that death receptor oligomerization is important for DISC function/efficacy, and structural studies provide novel insight into this process (32). Indeed, it has been shown that strategies that increase receptor oligomerization amplify TRAIL-induced apoptosis (33,34). Therefore, we used previously described sequences (30) to systematically study the effect of peptide dimerization and trimerization on DR5 binding and selective DR5-mediated death induction.…”

Interferon-α sensitizes human gastric cancer cells to TRAIL-induced apoptosis via activation of the c-CBL-dependent MAPK/ERK pathway

“…[6][7][8] Deregulated expression of apoptotic-related signaling molecules accounts for the resistance of various tumor cells to TRAIL-induced apoptosis. 9,10 In addition, studies have implicated multiple signaling pathways were involved in TRAIL-induced apoptosis. 11,12 For example, a recent study showed that c-Jun NH 2 -terminal kinase (JNK) and extracellular-regulated protein kinase (ERK) are two critical players involved in the hydrogen peroxide-mediated increase in TRAIL sensitization of tumor cells upon exposure to LY303511.…”