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Nat Chem Biol. 2014 Aug;10(8):656-63. doi: 10.1038/nchembio.1578. Epub 2014 Jul 6.

A high-throughput, multiplexed assay for superfamily-wide profiling of enzyme activity.

Author information

1
The Eli and Edythe L. Broad Institute, Cambridge, Massachusetts, USA.
2
Department of Biochemistry, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, Massachusetts, USA.
3
Arisaph Pharmaceuticals, Boston, Massachusetts, USA.
4
Laboratory of Clinical Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
5
1] Department of Biochemistry, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, Massachusetts, USA. [2] Arisaph Pharmaceuticals, Boston, Massachusetts, USA.
6
1] The Eli and Edythe L. Broad Institute, Cambridge, Massachusetts, USA. [2] Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA. [3] Harvard Medical School, Boston, Massachusetts, USA. [4] Howard Hughes Medical Institute, Chevy Chase, Maryland, USA.

Abstract

The selectivity of an enzyme inhibitor is a key determinant of its usefulness as a tool compound or its safety as a drug. Yet selectivity is never assessed comprehensively in the early stages of the drug discovery process, and only rarely in the later stages, because technical limitations prohibit doing otherwise. Here, we report EnPlex, an efficient, high-throughput method for simultaneously assessing inhibitor potency and specificity, and pilot its application to 96 serine hydrolases. EnPlex analysis of widely used serine hydrolase inhibitors revealed numerous previously unrecognized off-target interactions, some of which may help to explain previously confounding adverse effects. In addition, EnPlex screening of a hydrolase-directed library of boronic acid- and nitrile-containing compounds provided structure-activity relationships in both potency and selectivity dimensions from which lead candidates could be more effectively prioritized. Follow-up of a series of dipeptidyl peptidase 4 inhibitors showed that EnPlex indeed predicted efficacy and safety in animal models. These results demonstrate the feasibility and value of high-throughput, superfamily-wide selectivity profiling and suggest that such profiling can be incorporated into the earliest stages of drug discovery.

Comment in

PMID:
24997602
PMCID:
PMC5953424
DOI:
10.1038/nchembio.1578
[Indexed for MEDLINE]
Free PMC Article

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