Abstract 3813: TRAIL and IAP inhibitor AZD5582 combination treatment induces profound tumor cell death, which is differentially dependent on the mitochondrial apoptotic pathway
Authors: Polanski Radoslaw , Vincent John , Polanska Urszula , Tang Eric 
Published: August 2015

Background

It has been suggested that activation of the death receptors may induce selective tumour cell lethality and therefore represent a safe and effective therapeutic approach. However, several recent clinical trials evaluating either recombinant TRAIL or TRAIL receptor agonistic antibodies have shown very limited responses despite favourable safety profiles. This lack of efficacy suggests that combination with other agents will be required to unleash the therapeutic potential of the death receptor pathway.

Through a high-throughput combination screen, we identified the combination of TRAIL and an IAP (Inhibitor of Apoptosis Proteins) inhibitor AZD5582 to be highly synergistic in inducing tumour cell death across a number of tumour cell line panels. Since the molecular mechanisms underlying the synergistic activity of combined IAP inhibition and TRAIL remain unclear, we investigated the modes of the death signal transduction and its execution in a panel of breast cancer cell lines.

Methods

Cell number and viability were assessed using the nuclear staining dye Sytox green. BID, BAX and BAK gene expression was modulated using siRNA, protein expression was evaluated by western blotting. Total caspase activity was inhibited with Q-VD-OPh.

Results

We observed a diverse sensitivity profile in response to the AZD5582/TRAIL combination in it's ability to enhance cell killing over the single agent across the breast cancer cell line panel. A smaller panel was selected to contain members either resistant or exquisitely sensitive to AZD5582/TRAIL. Analysis of cell death confirmed early induction of apoptosis following treatment in sensitive cell lines. Combination treatment induced both caspase-8 activation and a concomitant apoptotic cell death in sensitive cell lines but we observed neither event in resistant cell lines. Importantly, knockdown of BID demonstrated the requirement of this BH3-only protein in two out of four IAPi/TRAIL sensitive cell lines.

Conclusions

In this study we investigated the exceptionally synergistic effect of combining TRAIL and the IAP inhibitor AZD5582 in mediating enhanced tumour cell lethality. We have shown that the combination induces both caspase-8 activation and a concomitant apoptotic cell death in sensitive cell lines but not in resistant cell lines upon combination treatment. We also demonstrated that the BID-initiated mitochondrial apoptotic pathway plays an essential role in some cell lines but is dispensable in others. Thus our study provides new insights into the molecular events responsible for the death signal transduction and execution in response to the IAPi/TRAIL combination treatment.

Citation Format: Radoslaw Polanski, John Vincent, Urszula Polanska, Eric Tang. TRAIL and IAP inhibitor AZD5582 combination treatment induces profound tumor cell death, which is differentially dependent on the mitochondrial apoptotic pathway. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3813. doi:10.1158/1538-7445.AM2015-3813