Abstract
We have recently described the development of a series of small-molecule inhibitors of human tumour necrosis factor (TNF) that stabilise an open, asymmetric, signalling-deficient form of the soluble TNF trimer. Here, we describe the generation, characterisation, and utility of a monoclonal antibody that selectively binds with high affinity to the asymmetric TNF trimer-small molecule complex. The antibody helps to define the molecular dynamics of the apo TNF trimer, reveals the mode of action and specificity of the small molecule inhibitors, acts as a chaperone in solving the human TNF-TNFR1 complex crystal structure, and facilitates the measurement of small molecule target occupancy in complex biological samples. We believe this work defines a role for monoclonal antibodies as tools to facilitate the discovery and development of small-molecule inhibitors of protein-protein interactions.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Antibodies, Monoclonal / metabolism*
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Antibodies, Monoclonal / pharmacology
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Cells, Cultured
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Crystallography, X-Ray
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Epitopes / chemistry
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Epitopes / metabolism
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HEK293 Cells
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Humans
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Models, Molecular
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Multiprotein Complexes / chemistry
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Multiprotein Complexes / metabolism*
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Protein Binding / drug effects
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Protein Conformation / drug effects
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Receptors, Tumor Necrosis Factor, Type I / chemistry
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Receptors, Tumor Necrosis Factor, Type I / metabolism*
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Signal Transduction / drug effects
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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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Tumor Necrosis Factor-alpha / chemistry
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Tumor Necrosis Factor-alpha / metabolism*
Substances
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Antibodies, Monoclonal
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Epitopes
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Multiprotein Complexes
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Receptors, Tumor Necrosis Factor, Type I
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Small Molecule Libraries
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TNF protein, human
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Tumor Necrosis Factor-alpha