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/SM-induced 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.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Apoptosis / drug effects*
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CRISPR-Cas Systems / genetics
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Caspase 10 / chemistry
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Caspase 10 / genetics
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Caspase 10 / metabolism*
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Caspase 8 / chemistry
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Caspase 8 / genetics
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Caspase 8 / metabolism
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Caspase Inhibitors / pharmacology
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Cell Line
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Cytokine TWEAK / pharmacology
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Drug Synergism
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HT29 Cells
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Humans
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Inhibitor of Apoptosis Proteins / antagonists & inhibitors
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Inhibitor of Apoptosis Proteins / metabolism*
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Interferon-gamma / genetics
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Interferon-gamma / metabolism
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Interferon-gamma / pharmacology*
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Mice
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Mice, Knockout
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Pentanoic Acids / pharmacology
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Protein Kinases / deficiency
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Protein Kinases / metabolism
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Receptor-Interacting Protein Serine-Threonine Kinases / deficiency
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Receptor-Interacting Protein Serine-Threonine Kinases / genetics
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Receptor-Interacting Protein Serine-Threonine Kinases / metabolism*
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Receptors, Tumor Necrosis Factor, Type I / deficiency
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Receptors, Tumor Necrosis Factor, Type I / genetics
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Receptors, Tumor Necrosis Factor, Type I / metabolism
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Recombinant Proteins / biosynthesis
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Recombinant Proteins / isolation & purification
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Recombinant Proteins / pharmacology
Substances
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3-(2-(2-tert-butylphenylaminooxalyl)aminopropionylamino)-4-oxo-5-(2,3,5,6-tetrafluorophenoxy)pentanoic acid
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Caspase Inhibitors
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Cytokine TWEAK
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Inhibitor of Apoptosis Proteins
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Pentanoic Acids
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Receptors, Tumor Necrosis Factor, Type I
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Recombinant Proteins
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Interferon-gamma
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MLKL protein, human
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Protein Kinases
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RIPK1 protein, human
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RIPK3 protein, human
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Receptor-Interacting Protein Serine-Threonine Kinases
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Caspase 10
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Caspase 8