Abstract
Store-operated calcium (SOC) channels are vital for activation of the immune cells, and mutations in the channel result in severe combined immunodeficiency in human patients. In lymphocytes, SOC entry is mediated by the Orai1 channel, which is activated by direct binding of STIM1. Here we describe an alternative approach for identifying inhibitors of SOC entry using minimal functional domains of STIM1 and Orai1 to screen a small-molecule microarray. This screen identified AnCoA4, which inhibits SOC entry at submicromolar concentrations and blocks T cell activation in vitro and in vivo. Biophysical studies revealed that AnCoA4 binds to the C terminus of Orai1, directly inhibiting calcium influx through the channel and also reducing binding of STIM1. AnCoA4, unlike other reported SOC inhibitors, is a molecule with a known binding site and mechanism of action. These studies also provide proof of principle for an approach to ion channel drug discovery.
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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Benzodioxoles / chemistry
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Benzodioxoles / pharmacology
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Benzodioxoles / therapeutic use*
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Chromones / chemistry
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Chromones / pharmacology
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Chromones / therapeutic use*
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Disease Models, Animal
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Drosophila
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Drosophila Proteins / antagonists & inhibitors
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Drosophila Proteins / genetics
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Drosophila Proteins / metabolism*
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Fura-2 / chemistry
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Gene Expression / drug effects
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HEK293 Cells
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Humans
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Hypersensitivity, Delayed / drug therapy
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Hypersensitivity, Delayed / metabolism
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Hypersensitivity, Delayed / pathology
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Immunosuppressive Agents / chemistry
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Immunosuppressive Agents / pharmacology
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Immunosuppressive Agents / therapeutic use
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Lymphocyte Activation / drug effects
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Membrane Proteins / antagonists & inhibitors
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Membrane Proteins / chemistry
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Mice
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Mice, Inbred BALB C
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Molecular Docking Simulation
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Neoplasm Proteins / chemistry
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Neoplasm Proteins / genetics
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Neoplasm Proteins / metabolism
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ORAI1 Protein
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Protein Binding
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Protein Structure, Tertiary
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Recombinant Proteins / biosynthesis
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Small Molecule Libraries / chemistry*
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Small Molecule Libraries / metabolism
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Small Molecule Libraries / pharmacology
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Stromal Interaction Molecule 1
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T-Lymphocytes / drug effects
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T-Lymphocytes / immunology
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T-Lymphocytes / metabolism
Substances
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AnCoA4 compound
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Benzodioxoles
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Chromones
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Drosophila Proteins
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Immunosuppressive Agents
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Membrane Proteins
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Neoplasm Proteins
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ORAI1 Protein
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Recombinant Proteins
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STIM1 protein, human
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Small Molecule Libraries
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Stromal Interaction Molecule 1
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olf186-F protein, Drosophila
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Fura-2