Combination of IAP antagonist and IFNγ activates novel caspase-10- and RIPK1-dependent cell death pathways

Abstract

Peptido-mimetic inhibitor of apoptosis protein (IAP) antagonists (Smac mimetics (SMs)) can kill tumour cells by depleting endogenous IAPs and thereby inducing tumour necrosis factor (TNF) production. We found that interferon-γ (IFNγ) synergises with SMs to kill cancer cells independently of TNF − and other cell death receptor signalling pathways. Surprisingly, CRISPR/Cas9 HT29 cells doubly deficient for caspase-8 and the necroptotic pathway mediators RIPK3 or MLKL were still sensitive to IFNγ/SMinduced killing. Triple CRISPR/Cas9-knockout HT29 cells lacking caspase-10 in addition to caspase-8 and RIPK3 or MLKL were resistant to IFNγ/SM killing. Caspase-8 and RIPK1 deficiency was, however, sufficient to protect cells from IFNγ/SM-induced cell death, implying a role for RIPK1 in the activation of caspase-10. These data show that RIPK1 and caspase-10 mediate cell death in HT29 cells when caspase-8-mediated apoptosis and necroptosis are blocked and help to clarify how SMs operate as chemotherapeutic agents.

Caspase‐8: regulating life and death

“…RIPK1 deubiquitylation can be achieved through the inhibition of cIAP1/2, for instance following SMAC release upon MOMP or by pharmacological SMAC mimetics. The absence of caspase-8 and RIPK3 suffices to block death in mouse cells even in the presence of MLKL following IFNγ and SMAC mimetic treatment (59). However, human caspase-8 and RIPK3 double-deficient HT29 cells die following IFNγ and SMAC mimetic treatment (59).…”

“…The absence of caspase-8 and RIPK3 suffices to block death in mouse cells even in the presence of MLKL following IFNγ and SMAC mimetic treatment (59). However, human caspase-8 and RIPK3 double-deficient HT29 cells die following IFNγ and SMAC mimetic treatment (59). Death was attributed to IFNγ-induced gene expression, RIPK1 and caspase-10 in human HT29 cells, but independent of TNFR1/2, Fas, TRAILR, TWEAKR, IFNAR and ATG5, a protein essential for autophagy.…”

“…59 However, human caspase-8 and RIPK3 double-deficient HT29 cells die following IFNγ and SMAC mimetic treatment. 59 Death was attributed to IFNγ-induced gene expression, RIPK1 and caspase-10 in human HT29 cells, but independent of TNFR1/2, Fas, TRAILR, TWEAKR, IFNAR, and ATG5, a protein essential for autophagy. These data hint at the existence of a yet unknown intrinsic signal that mediates death through RIPK1.…”

PD-L1 and IAPs co-operate to protect tumors from cytotoxic lymphocyte-derived TNF

“…6,7,11 However, TNF or other cytokines, supplied by immune cells are likely to contribute to SM efficacy, especially in tumors that fail to produce TNF upon IAP loss. [12][13][14][15][16][17] In support of this concept, SMs are more efficient in vivo when combined with inflammatory adjuvants, which promote immune cell-derived TNF secretion. 15,18 Similarly, IAP antagonism promotes antitumor immunity in myeloma, independently from cell death induction.…”

RIPK1 and RIPK3 - emerging targets in cancer?

“…Other co-agents promoting activation of necroptosis by SMAC mimetics may be proteasome inhibitors, such as bortezomib, or glucocorticoids, which have been recently shown to promote necroptosis in several non-Hodgkin lymphoma and acute lymphoblastic leukemia cell lines expressing low levels of caspase-8 (88, 89). Pro-death activity of SMAC mimetics can also be greatly enhanced by the combination with IFNγ, importantly rendering cell death independent of autocrine TNFα signaling (90,91). In the absence of caspase inhibition, SMAC mimetics and IFNγ induce RIPK1 dependent apoptosis, which is meditated by both caspase-8 and caspase-10.…”