Abstract
Agonists of mouse STING (TMEM173) shrink and even cure solid tumors by activating innate immunity; human STING (hSTING) agonists are needed to test this therapeutic hypothesis in humans. The endogenous STING agonist is 2'3'-cGAMP, a second messenger that signals the presence of cytosolic double-stranded DNA. We report activity-guided partial purification and identification of ecto-nucleotide pyrophosphatase/phosphodiesterase (ENPP1) to be the dominant 2'3'-cGAMP hydrolyzing activity in cultured cells. The hydrolysis activity of ENPP1 was confirmed using recombinant protein and was depleted in tissue extracts and plasma from Enpp1(-/-) mice. We synthesized a hydrolysis-resistant bisphosphothioate analog of 2'3'-cGAMP (2'3'-cG(s)A(s)MP) that has similar affinity for hSTING in vitro and is ten times more potent at inducing IFN-β secretion from human THP1 monocytes. Studies in mouse Enpp1(-/-) lung fibroblasts indicate that resistance to hydrolysis contributes substantially to its higher potency. 2'3'-cG(s)A(s)MP is therefore improved over natural 2'3'-cGAMP as a model agonist and has potential as a vaccine adjuvant and cancer therapeutic.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antineoplastic Agents / chemical synthesis
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Antineoplastic Agents / pharmacology*
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Cell Line, Tumor
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Gene Expression Regulation, Neoplastic*
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Humans
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Hydrolysis
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Interferon-beta
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Membrane Proteins / agonists*
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Membrane Proteins / deficiency
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Membrane Proteins / genetics
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Membrane Proteins / metabolism
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Mice
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Monocytes / drug effects
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Monocytes / metabolism
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Monocytes / pathology
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Nucleotides, Cyclic / chemistry
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Nucleotides, Cyclic / metabolism*
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Nucleotides, Cyclic / pharmacology
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Organothiophosphorus Compounds / chemical synthesis
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Organothiophosphorus Compounds / pharmacology*
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Phosphoric Diester Hydrolases / genetics
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Phosphoric Diester Hydrolases / metabolism
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Pyrophosphatases / antagonists & inhibitors*
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Pyrophosphatases / genetics
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Pyrophosphatases / metabolism
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Second Messenger Systems / genetics
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Signal Transduction
Substances
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Antineoplastic Agents
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Membrane Proteins
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Nucleotides, Cyclic
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Organothiophosphorus Compounds
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Recombinant Proteins
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STING1 protein, human
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Sting1 protein, mouse
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cyclic guanosine monophosphate-adenosine monophosphate
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Interferon-beta
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Phosphoric Diester Hydrolases
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ectonucleotide pyrophosphatase phosphodiesterase 1
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Pyrophosphatases