Intracellular quantitative detection of human thymidylate synthase engagement with an unconventional inhibitor using tetracysteine-diarsenical-probe technology

Demonstrating a candidate drug's interaction with its target protein in live cells is of pivotal relevance to the successful outcome of the drug discovery process. Although thymidylate synthase (hTS) is an important anticancer target protein, the efficacy of the few anti-hTS drugs currently use...

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Bibliographic Details
Published in:Scientific reports 2016-06, Vol.6 (1), p.27198-27198, Article 27198
Main Authors: Ponterini, Glauco, Martello, Andrea, Pavesi, Giorgia, Lauriola, Angela, Luciani, Rosaria, Santucci, Matteo, Pelà, Michela, Gozzi, Gaia, Pacifico, Salvatore, Guerrini, Remo, Marverti, Gaetano, Costi, Maria Paola, D'Arca, Domenico
Format: Article
Language:English
Subjects:
Amino Acid Motifs
Arsenic - chemistry
Arsenic - metabolism
Catalytic Domain
Cysteine - chemistry
Cysteine - metabolism
Enzyme Inhibitors - chemistry
Enzyme Inhibitors - pharmacology
HEK293 Cells
Humans
Models, Molecular
Protein Binding
Protein Conformation
Thymidylate Synthase - antagonists & inhibitors
Thymidylate Synthase - chemistry
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Summary:Demonstrating a candidate drug's interaction with its target protein in live cells is of pivotal relevance to the successful outcome of the drug discovery process. Although thymidylate synthase (hTS) is an important anticancer target protein, the efficacy of the few anti-hTS drugs currently used in clinical practice is limited by the development of resistance. Hence, there is an intense search for new, unconventional anti-hTS drugs; there are approximately 1600 ongoing clinical trials involving hTS-targeting drugs, both alone and in combination protocols. We recently discovered new, unconventional peptidic inhibitors of hTS that are active against cancer cells and do not result in the overexpression of hTS, which is a known molecular source of resistance. Here, we propose an adaptation of the recently proposed tetracysteine-arsenic-binding-motif technology to detect and quantitatively characterize the engagement of hTS with one such peptidic inhibitor in cell lysates. This new model can be developed into a test for high-throughput screening studies of intracellular target-protein/small-molecule binding.
ISSN:2045-2322
2045-2322
DOI:10.1038/srep27198