Smac mimetic triggers necroptosis in pancreatic carcinoma cells when caspase activation is blocked

Evasion of apoptosis represents a key mechanism of treatment resistance of pancreatic cancer (PC) and contributes to the poor prognosis of this cancer type. Here, we report that induction of necroptosis is an alternative strategy to trigger programmed cell death in apoptosis-resistant PC cells. We show that the second mitochondrial activator of caspases (Smac) mimetic BV6 that antagonizes inhibitor of apoptosis (IAP) proteins induces necroptosis in PC cells in which apoptosis is blocked by the caspase inhibitor zVAD.fmk. Intriguingly, BV6 switches autocrine/paracrine production of tumor necrosis factor (TNF)α by PC cells into a death signal and also acts in concert with exogenously supplied TNFα to trigger necroptosis, when caspase activation is simultaneously blocked. BV6 stimulates TNFα production and formation of the receptor-interacting protein (RIP)1/RIP3-containing necrosome complex in PC cells. Knockdown of TNF receptor 1 (TNFR1) protects PC cells from BV6- or BV6/TNFα-mediated cell death, demonstrating that TNFα autocrine/paracrine signaling by PC cells contributes to BV6-induced necroptosis. Importantly, genetic silencing of receptor interacting protein kinase 3 (RIPK3) or mixed lineage kinase domain-like protein (MLKL) significantly rescues PC cells from BV6- or BV6/TNFα-induced cell death. Similarly, pharmacological inhibition of RIP1, RIP3 or MLKL significantly reduces BV6- or BV6/TNFα-stimulated cell death. By demonstrating that Smac mimetics can bypass resistance to apoptosis by triggering necroptosis as an alternative form of programmed cell death, our findings have important implications for the design of new treatment concepts for PC.