Investigating the Adverse Effects of Novel Tumour-Specific IAP Antagonists

Inhibitor of apoptosis (IAP) proteins are negatively regulated by the mitochondrial pro-apoptotic protein, SMAC, through a tetrapeptide binding-motif. A class of drugs mimicking this motif have been developed as anti-cancer agents and have further highlighted IAP signalling in regulation of cell death. These SMAC mimetics (SMs) activate the non-canonical NF-κB pathway, induce TNFα secretion and selectively promote cell death in tumour cells; an effect potentiated in a TNFα-rich microenvironment. However, due to this mechanism of action, SMs have the potential to induce toxicity in non-tumour cells residing in this environment. The aim of this thesis was two-fold: to determine the effect of SMs in non-tumour derived cells both in vitro and in vivo. I determined a robust methodology in sensitive and resistant tumour cells with SMs as single agents. I found that MDA-MB-231 and MDA-MB-468 cells were sensitive and resistant to SMs as single agents, respectively. Furthermore, the pan caspase inhibitor, zVAD, actually inhibited TNFα secretion in combination with the bivalent SM, BV6. In hepatocyte-like and epithelial non-tumour cells in vitro, I found that SMs did not induce cell death, even when co-treated with TNFα. These data suggest that SMs under clinical development do not have significant toxicity in non-tumour derived cells suggesting a large therapeutic window in the clinic. As liver regeneration is also TNFα-dependent, SMs have the potential to exacerbate liver injury. To determine whether the SM, TL-32711, could induce NF- κB activation in vivo, I used mice expressing an NF-κB reporter gene (miceluc) allowing for quantification of NF-κB activity via in situ live imaging. Miceluc were dosed with TL-32711, either alone, or in combination with two hepatotoxins. Strikingly, TL-32711 alone induced significant liver-specific NF-κB activation and protected against liver injury. These data show that SMs could have real clinical relevance for use as a treatment to ameliorate hepatoxicity.