Review Article | Published:

Mechanistic enzymology in drug discovery: a fresh perspective

Nature Reviews Drug Discovery volume 17, pages 115132 (2018) | Download Citation

  • An Erratum to this article was published on 15 December 2017

This article has been updated

Abstract

Given the therapeutic and commercial success of small-molecule enzyme inhibitors, as exemplified by kinase inhibitors in oncology, a major focus of current drug-discovery and development efforts is on enzyme targets. Understanding the course of an enzyme-catalysed reaction can help to conceptualize different types of inhibitor and to inform the design of screens to identify desired mechanisms. Exploiting this information allows the thorough evaluation of diverse compounds, providing the knowledge required to efficiently optimize leads towards differentiated candidate drugs. This review highlights the rationale for conducting high-quality mechanistic enzymology studies and considers the added value in combining such studies with orthogonal biophysical methods.

Key points

  • The application of detailed mechanistic enzyme kinetics is vital for the characterization of enzyme targets; furthermore, it is crucial to aid in the design and prosecution of assays to effectively profile compound properties and to gain insight into the mechanism of action required for drug efficacy.

  • Enzyme assays are used extensively in hit identification and hit validation and for detailed characterization of the compound mechanism to guide lead optimization.

  • It is important to understand the limitation of IC50 values and to more deeply probe the relationship between the molecular structures of hits and leads and their kinetics of binding, inhibition and mechanism of action.

  • Combining information from enzyme kinetic studies with that derived from biophysical methods can be advantageous in assessing protein quality, generating suitable assays to identify a range of desired mechanisms and to complement detailed mechanistic characterization.

  • The efficient use of these methods enables the identification, prioritization and progression of truly differentiated compound series that have an enhanced probability of success in translation through to the clinic as a consequence of detailed understanding of their mechanism.

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Change history

  • 15 December 2017

    In the affiliations of this article, the address provided for Discovery Sciences, IMED Biotech Unit, AstraZeneca was incorrect and should be Building 310, Cambridge Science Park, Milton Road, Cambridge CB4 0WG, UK. The article has been corrected in the print and online version.

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Acknowledgements

The authors would like to thank S. Thrall for critical input into Fig. 1.

Author information

Affiliations

  1. Discovery Sciences, IMED Biotech Unit, AstraZeneca, Building 310, Cambridge Science Park, Milton Road, Cambridge, CB4 0WG, UK.

    • Geoffrey A. Holdgate
    •  & Rachel L. Grimley
  2. Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas 77843, USA.

    • Thomas D. Meek

Authors

  1. Search for Geoffrey A. Holdgate in:

  2. Search for Thomas D. Meek in:

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Competing interests

G.H. and R.G. are current employees of AstraZeneca. T.M. is a former employee of GSK.

Corresponding author

Correspondence to Geoffrey A. Holdgate.

Glossary

Mechanism

A process by which a reaction takes place, fully determined when all the intermediates, complexes and conformational states of an enzyme are characterized and the rate constants associated with the conversion between them are quantified. The term is often used in inhibition studies to distinguish between different modes of inhibition.

Half-maximal inhibitory concentration

(IC50). The inhibitor concentration that gives a 50% decrease in rate under the specific assay conditions employed.

Random-order mechanism

A reaction mechanism in which either of the two substrates may bind to the enzyme first to form a binary complex, followed by the other to form a ternary complex.

Non-competitive inhibitors

Inhibitors that bind with equal affinity both before and after the varied substrate.

Uncompetitive inhibitors

Inhibitors that bind only after the substrate.

Slow-binding inhibition

An inhibition that occurs slowly on the timescale of the assay as the enzyme–inhibitor complex concentration increases to its steady-state level.

Competitive inhibition

A type of inhibition where the inhibitor binds only before the varied substrate (see also non-competitive and uncompetitive inhibition).

Reversible inhibition

An inhibition that can be reversed on the timescale of the assay by competition or dilution. Reversible inhibitors are not precluded from forming covalent bonds with the enzyme.

Irreversible inhibition

An inhibition that cannot be reversed on the timescale of the assay. Truly irreversible inhibitors never dissociate from the enzyme and are characterized by an inactivation rate constant, not a dissociation constant. Irreversible inhibitors do not necessarily have to be covalently bound.

Tight-binding inhibition

A type of inhibition occurring under conditions when the concentration of inhibitor required to cause inhibition is similar to the enzyme concentration, leading to depletion of the free inhibitor concentration. This results in breakdown of the usual assumptions leading to simple inhibition kinetics and requires a more complex rate equation.

Mixed inhibition

A type of inhibition in which the inhibitor binds both before and after the varied substrate.

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https://doi.org/10.1038/nrd.2017.219