Assessment of Enzyme Inhibition: A Review with Examples from the Development of Monoamine Oxidase and Cholinesterase Inhibitory Drugs

Abstract

The actions of many drugs involve enzyme inhibition. This is exemplified by the inhibitors of monoamine oxidases (MAO) and the cholinsterases (ChE) that have been used for several pharmacological purposes. This review describes key principles and approaches for the reliable determination of enzyme activities and inhibition as well as some of the methods that are in current use for such studies with these two enzymes. Their applicability and potential pitfalls arising from their inappropriate use are discussed. Since inhibitor potency is frequently assessed in terms of the quantity necessary to give 50% inhibition (the IC50 value), the relationships between this and the mode of inhibition is also considered, in terms of the misleading information that it may provide. Incorporation of more than one functionality into the same molecule to give a multi-target-directed ligands (MTDLs) requires careful assessment to ensure that the specific target effects are not significantly altered and that the kinetic behavior remains as favourable with the MTDL as it does with the individual components. Such factors will be considered in terms of recently developed MTDLs that combine MAO and ChE inhibitory functions.

Untargeted metabolomic profiling of three Crataegus species (hawthorn) and their in vitro biological activities

“…Despite the increase in the use of drugs for receptors to modulate signals from outside the cell, 47% of all current approved drugs inhibit enzyme targets to treat or delay the onset of these major health problems. 41 However, several of these drugsfor instance, ⊍-glucosidase and ⊍-amylase inhibitors used to treat diabetes mellitus, and cholinesterase and tyrosinase inhibitors used for managing Alzheimer's disease and hyperpigmentation respectivelyhave been found to pose side effects such as gastrointestinal disturbance and liver damage. There is, therefore, an urgent need to search for alternative inhibitors from natural sources with minimal or no side effects.…”

Carprofen: a theoretical mechanistic study to investigate the impact of hydrophobic interactions of alkyl groups on modulation of COX-1/2 binding selectivity

“…The relationship between the half maximal inhibitory concentration (IC 50 ) and inhibition constant (K i ) was described in several reports [28,29]. Both IC 50 and K i are used to express the relative potency of an inhibitor.…”

Kinetics, mechanism, and inhibition of monoamine oxidase

“…Thus, the relationship between IC50 and Ki depends on the substrate concentration used and on the type of the reversible inhibition (Ramsay and Tipton 2017). The accurate assessment of reversible inhibitors depends on reliable quantitative assays for determination of initial IC50 values, for which the substrate concentration used must be defined.…”

“…If the IC50 is determined by adding enzyme to substrate and inhibitor at the same time, the parameter obtained will be the Ki for reversible binding. If the inhibitor is pre-incubated with MAO for 30 minutes before adding substrate, then the IC50 obtained will come from a mix of inactivated and reversibly inhibited MAO and cannot be used to compare different classes of compounds with different inactivation rates (Ramsay and Tipton 2017). Proper information can be obtained from progress curves where the oxidation of substrate is measured in the presence of the irreversible inhibitor (McDonald and Tipton 2012).…”