Mechanism-based inhibition: deriving K(I) and k(inact) directly from time-dependent IC(50) values

J Biomol Screen. 2009 Sep;14(8):913-23. doi: 10.1177/1087057109336751. Epub 2009 Aug 12.


The potential of enzyme inhibition of a drug is frequently quantified in terms of IC(50) values. Although this is a suitable quantity for reversible inhibitors, concerns arise when dealing with irreversible or mechanism-based inhibitors (MBIs). IC(50) values of MBIs are time dependent, causing serious problems when aiming at ranking different compounds with respect to their inhibitory potential. As a consequence, most studies and ranking schemes related to MBIs rely on the inhibition constant (K(I)) and the rate of enzyme inactivation (k(inact)) rather than on IC(50) values. In this article, the authors derive a novel relation between potentially time-dependent IC(50) values and K(I), k(inact) parameters for different types of inhibition. This allows for direct estimation of K(I) and k(inact) values from time-dependent IC(50) values, even without the need of additional preincubation experiments. The application of this approach is illustrated using a fluorimetric assay to access the drug-drug interaction potential associated with new chemical entities. The approach can easily be implemented using standard software tools (e.g., XLfit) and may also be suitable for applications where mechanism-based inhibition is a desired mode of action (e.g., at particular pharmacological drug targets).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding, Competitive
  • Cytochrome P-450 CYP1A2 / metabolism
  • Cytochrome P-450 CYP1A2 Inhibitors
  • Cytochrome P-450 CYP3A / metabolism
  • Cytochrome P-450 CYP3A Inhibitors
  • Drug Antagonism*
  • Enzyme Activation / drug effects*
  • Enzyme Inhibitors / pharmacokinetics*
  • Humans
  • Inhibitory Concentration 50*
  • Models, Chemical
  • Models, Theoretical
  • Time Factors


  • Cytochrome P-450 CYP1A2 Inhibitors
  • Cytochrome P-450 CYP3A Inhibitors
  • Enzyme Inhibitors
  • CYP1A2 protein, human
  • Cytochrome P-450 CYP1A2
  • Cytochrome P-450 CYP3A
  • CYP3A4 protein, human