CPSA Digest 2002

Proceedings -Wednesday, October 9, 2002

WeOD1

Metabolism Dependent Drug-Drug Interactions

Background
Enzyme inhibition represents a common strategy in the development of drug candidates. Potency (in terms of inhibition) and selectivity are important factors that influence the likelihood of drug-drug interactions. Traditionally, enzyme inhibition is thought of as reversible, when a drug or its metabolite enters the active site of an enzyme, interacts for a period of time, and leaves the enzyme unchanged. In contrast, mechanism-based inactivation is an irreversible process that has been demonstrated for a variety of enzymes and inhibitors [e.g., tienlinic acid, erythromycin, gestodene, delavirdine and mibefradil (Posicor®)]. The focus of this presentation is on cytochrome P450 (CYP450) metabolism dependent inhibition, also known as mechanism-based inhibition or suicide inhibition. Here, a drug binds to a particular active site within CYP450 and forms a reactive intermediate that destroys the enzyme.

Premise
Mechanism-based inhibition is an irreversible inhibition where a covalent bond is formed between a metabolite and the active site of the enzyme, destroying the enzyme’s activity. This inactivation is seen as two types—true irreversible and quasi-irreversible.

Below are listed some characteristics that distinguish reversible from irreversible CYP450 inhibition:

• Inhibition effect extended beyond the elimination of the drug
• Inactivation of enzyme (i.e., the inhibitor becomes covalently attached to the enzyme)
• Effect tends to accumulate after each dosing
• Inhibition effect is generally greater than predicted based on 'reversible' IC50 or Ki values
• Most compounds will have non-linear pharmacokinetics
• Rare cases of hepatotoxicity associated with covalently bound adducts
• Difficult to predict inhibitory effect in patients

The kinetic analysis of metabolism-dependent inhibition utilizes two parameters:

K_inact - the maximal rate of enzyme inactivation
K_i - the concentration of the inhibitor that gives 50% maximal inhibition

The Dixon and Kitz-Wilson methods represent two of the most commonly used methods for the estimation of K_i. Both of these methods require the use of several assumptions and data linearization. The parameter lambda (combining K_inact, K_i and inhibitor concentration) has been developed to provide improved interpretation of kinetic data. Lambda is the inactivation rate constant which can be compared to known irreversible inhibitors with clinically significant drug interactions (reference: Mayhew, Hall and Jones).

The utility of lambda can be seen in the numbers below which compare values to known inhibitor compounds:

A reversible P450 inhibition screen can be used that measures the inhibitory concentration (IC50) at different time points; if there is true irreversible inhibition, IC50 does not change but if it is reversible then IC50 changes with time.

Some metabolism dependent inhibition experiments include:

1. Precincubation—dilution in liver microsomes, in which the inhibitor is exposed to microsomes and cofactors before the addition of substrate
2. Dialysis of microsomal incubations
3. Spectral determination of MI complex formation
4. Partition ratio

Metabolism dependent CYP450 inhibition is a liability that should be evaluated during preclinical development along with other drug characteristics. Irreversible CYP inhibition is undesirable and should be seriously evaluated in conjunction with dose, therapeutic
area, enzyme kinetics and competition from other drugs on the market. In vitro models are predictive with varying degrees of confidence. Human clinical studies are the true test of drug interactions, especially metabolism dependent inhibition.

References and/or Links
Bradley S. Mayhew, David R. Jones, and Stephen D. Hall, "An In Vitro Model for Predicting In Vivo Inhibition of Cytochrome P450 3A4 by Metabolic Intermediate Complex Formation." Drug Metabolism Disposition 28 (2000) 1031-1037.

T. Maurer and H.L. Fung, "Comparison of Methods for Analyzing Kinetic Data From Mechanism-Based Enzyme Inactivation: Application to Nitric Oxide Synthase," AAPS PharmSci 2000; 2 (1) article 8 can be viewed at: www.aapspharmsci.org/scientificjournals/pharmsci/journal/8.html

Richard B. Silverman web pages at Northwestern University
www.chem.northwestern.edu/~agman/silverman.html

Books by R.B. Silverman:
The Organic Chemistry of Enzyme-Catalyzed Reactions, Academic Press, San Diego (2000).
Mechanism-Based Enzyme Inactivation: Chemistry and Enzymology, Vols. I and II, CRC Press, Boca Raton, Florida (1988).
The Organic Chemistry of Drug Design and Drug Action, Academic Press, San Diego (1992).

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