s 1999 Elsevier Science B.V. All rights reserved.Variability in Human Drug Response 29G.T. Tucker, Editor

Enzyme Inhibition and Induction - PK/PD Impact

Ulrich Klotza and Kari T. Kivistob

aDr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany

^Department of Clinical Pharmacology, University of Hèlsinki, Finland

ABSTRACT

Disposition and action of many drugs depend on then- (extra-) hepatic metabolismwhich is accomplished by several cytochrome P450 (CYP) enzymes and/or conjugatingactivities. The CYP3a subfamily is the most abundant in the liver and in the smallintestine.At these metabolic sites high clearance drugs sustain presystemic elimination.Ingredients of grapefruit juice have no influence on the systemic clearance of various drugswhereas intestinal drug metabolism may be decreased and consequently oral bioavailabilityincreases. Midazolam represents a probe of CYP3A4. Its oral availability is higher duringtreatment with fluconazole, itraconazole, ketoconazole, diltiazem, verapamil, erythromycinor grapefruit juice ingestión. On the other side midazolam metabolism is induced byrifampicin. Thus, switching from inhibition to induction causes an about 400 fold changein the AUC of oral midazolam which has a significant impact on the sedative action of thisbenzodiazepine. Using verapamil as a model drug it was shown that rifampicin will induce,not only hepatic but also intestinal drug metabolism. During oral coadministration of bothdrugs, steady state concentrations of verapamil decreased more than 30 fold and theresponse was nearly abolished. These examples show that inhibition or induction of drugmetabolizing enzymes will have an impact on PK and both contribute to variability in drugresponse.

Key words: inhibition, induction, drug metabolism, pharmacokinetics, pharmacodynamics

Correspondence: Prof. Dr. Ulrich Klotz, Dr. Margarete Fischer-Bosch-Institut fur Klinische Pharmacologie,Auerbachstrasse 112, D-70376 Stuttgart, Germany, Tel: +49-711-81-01-37-02, Fax: 449-711-85-92-95, Email:ulrich.klotz@ikp-stuttgart.de

Funding acknowledgement: This work was supported by the Robert Bosch Foundation Stuttgart and HelsinkiUniversity Central Hospital Research Fund.

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INTRODUCTION

Most drugs undergo biotransformation before excretion by renal, biliary or other routes.The liver, which contains a number of drug metabolising systems, is the major site ofmetabolism. Among those enzymes the cytochrome (CYP) P450 superfamily enzymesrepresent the most important class [1,2]. More recently it has been revealed that theseenzymes are also present at extrahepatic sites, especially in the small intestine [3],

During the past numerous in vitro and in vivo studies have been performed indicatingthat the activity of the various CYP can be modified, i.e. drugs, environmental factors orfood constituents can inhibit or induce these enzymes. Impairment or acceleration of drugmetabolism will lead to pharmacokinetic (PK) changes which may have alsopharmacodynamic (PD) consequences [4,5].

This article will concentrate on in vivo studies in man where interactions betweendifferent compounds have been investigated. In these interaction studies often probe drugshave been used to characterize indirectly the in vitro activity of certain CYP [6,7]. Thepharmacokinetic modifications due to inhibiting or inducing agents can be best visualizedby changes in the plasma level-time profiles (AUC measurements) or by the calculation ofsystemic (CL) and oral (CL0) clearance of the affected drugs [8].

We will present some recent examples from literature and our own work to illustratethe clinical impact of the observed PK- and/or PD-changes, rather than provide acomprehensive overview of the involved problems.

GRAPEFRUIT JUICE AS INHIBITOR

The inhibitory potential of grapefruit juice was serendipitously discovered during aninteraction study with ethanol (EtOH) and Ca-channel blockers to mask the typical flavorof EtOH [9]. Since then, with an increasing number of drugs, such as felodipine,nifedipine, nitrendipine, nisoldipine, terfenadine, cyclosporine and midazolam, elevatedplasma concentrations following oral administration have been observed when grapefruitjuice was concomitantly ingested. Most studies focused on drugs that are CYP3Asubstrates, since this subfamily is particularly inhibited, especially in the intestinalepithelium. Consequently, the affected drugs showed a substantial increase in their relativeoral bioavailability (F) ranging from a mean value of about 40% to over 250% [10, 11]. Inthe case of terfenadine [12,13] and midazolam [14] these PK-changes were associated withmore pronounced PD-effects. Recently, it was suggested that 6',7'-dihydroxybergamottin isthe putative inhibiting agent in the grapefruit juice [15]. The unpredictability of themagnitude of the individual increase in F would suggest to reduce the oral dosage of theaffected CYP3A-substrates.

EtOH AS INDUCER

EtOH is consumed worldwide in tremendous amounts and it is an effective inducer ofhepatic drug metabolism, especially involving pathways mediated by CYP2E1. Thereforeindividuals consuming chronically EtOH have an accelerated metabolism of CYP2E1

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substrates, such as chlorzoxazone (CZX), paracetamol, halothane, enflurane,methoxyflurane, sevoflurane and many organic solvents [16]. The metabolic ratio of CZX,a probe of CYP2E1, increased significantly from 0.43 in non-induced subjects to 1.2 inEtOH-induced patients and it normalized within 1 week after withdrawal of EtOH (17].

The hepatotoxic potential of paracetamol resides in its reactive metabolite N-acetyl-p-benzo-chinonimine which is formed by CYP2E1 [18]. In two recent reviews, 67 and 94cases of paracetamol-induced hepatotoxicity associated with regular intake of EtOH werepresented indicating the risk of combined use of both agents [19,20], Likewise, the hepato-and nephrotoxic potential of fluorinated volatile anaesthetics depends on the CYP2E1-mediated formation of reactive metabolites and inorganic fluoride. Therefore it isconceivable that induction of CYP2E1 by EtOH may increase the toxicity of these drugs[16].

INHIBITION OF DRUG METABOLISM BY CIMETIDINE

Due to its imidazole ring structure cimetidine binds to the heme part of several CYP[21]. Therefore it is not surprising that this H2-receptor antagonist inhibits the metabolismof many drugs (Table 1) which are substrates of different CYP. CL of these substances isdecreased on average by 20 to 50% by cimetidine [22,23]. With some drugs (e.g.diazepam, theophylline, carbamazepine, phenytoin, lidocaine, warfarin), the increasedplasma concentrations were associated with a more pronounced PD response [22-24].Therefore it appears prudent to reduce the dose of these drugs or to use ranitidine whichdoes not inhibit drug metabolism.

Table 1.Drugs, whose hepatic elimination is impaired by cimetidine

acenocoumarolealprazolamamitriptylineamoxicillinantipyrinebromazepambupivacainecaffeinecarbamazepinecarbocysteinechlordiazepoxidechlormethiazolechloroquinechlorpromazineclobazam

cyclosporinedapsonedebrisoquinedesipraminedesmethyldiazepamdiazepamdigitoxindiltiazemdoxepineencainideenoxacinestradiolethanolfelodipineflecainide

5-fluorouracilflurazepamflurbiprofenglibenclamideglipizideimipramineindomethacinlabetalollidocainemebendazolemetforminemethadonemetoprololmetronidazolemidazolam

moricizinenicardipinenicotinenifedipinenimodipinenitrazepamnitrendipinenortriptylineparoxettaepentoxifyllinepethidinephenytoinpindololpiroxicamprednisone

propranololquinidinequininesparteinesulindactacrinetheobrominetheophyllinetolbutamidetriamterentriazolamvalproic acidverapamilwarfarin

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RIFAMPICIN AND VERAPAMIL

Drug metabolism in the liver and gut wall can be induced by rifampicin and especiallysubstrates of CYP3A4 (e.g. cyclosporine, midazolam) are cleared much faster duringcoadministration of rifampicin and consequently F of these drugs is dramatically reduced[25,26].

In other studies verapamil was chosen as a model drug, because several CYP(CYP3A4, CYP1A2, CYP2C) are involved in its metabolism and stereoselective aspectscan be investigated with this racemic drug. In young [27] and elderly [28] subjects it wasfound that 12 days' co-treatment with rifampicin (600 mg once daily) and verapamil (120mg twice daily) resulted in a more than 30-fold increase (p < 0.001) in the oral CL of thecardioactive S-verapamil, whereas systemic CL following an iv-dose of 10 mg wasincreased only about 1.3-fold. F of both enantiomers was dropped almost to zero andconsequently the effect of oral verapamil on atrioventricular conduction was nearlyabolished (p < 0.01). Induction by rifampicin caused a considerable reduction ofstereoselectivity after both iv- and po-administration. As rifampicin altered the PK and PDof verapamil in both populations to a much greater extent after po-dosing compared withiv-administration, it was concluded that - independent of the age - prehepatic (gut wall)metabolism of verapamil was preferentially induced by rifampicin [27,28].

INHIBITION AND INDUCTION OF THE METABOLISM OF MIDAZOLAM

Midazolam represents a well-established probe for CYP3A4 [6,7] and its PK/PD-properties and relationships have been described thoroughly [29]. There is a markedinterindividual variation in both CYP3A4 activity in the liver and pharmacokinetics ofCYP3A4 substrates. Furthermore, CYP3A4 is readily inhibited or induced by a number ofdrugs, resulting in many clinically significant drug interactions. The erythromycin breathtest is currently the best validated measure of CYP3A4 activity in the liver in vivo. Theresults of this test have been shown to correlate with the clearance of intravenouslyadministered midazolam [30], and the clearance of intravenous midazolam seems toprovide a good estimate of the activity of CYP3A4 in the liver. It should be noted thatassays of CYP3A4 activity in the liver are not suitable for accurate assessment of intestinalCYP3A4 activity. This is an important issue because many CYP3A4 substrates undergosignificant first-pass metabolism not only in the liver but also in the gut. The first-passmetabolism of CYP3A4 substrates, i.e. the combined CYP3A4 activity in the liver and gut,can, however, be predicted by administering the probe orally.

After oral administration, midazolam undergoes extensive first-pass metabolism in theintestinal wall and the liver and, therefore, its concentrations in plasma are markedlyaltered by inhibition and induction of CYP3A4. The effects of various drugs on the AUCof oral midazolam are illustrated in Figure 1 [31-36]. In these studies, a single dose ofmidazolam was given orally after a 2- to 6-day pretreatment with the inhibitor. Most ofthese interactions are clinically significant, that is, the sedative effect of midazolam isgreater and lasts longer during co-administration of a potent CYP3A4 inhibitor. This holdsalso true for triazolam, the pharmacokinetics of which are also very susceptible toinhibition (and induction) of CYP3A4.

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Plasma concentrations of numerous drugs are reduced to a clinically significant extentby co-administration of rifampicin. In a recent study, in which a single dose (15 mg) ofmidazolam was given orally after a 5-day pretreatment with rifampicin (600 mg oncedaily) or placebo, rifampicin decreased the AUC of midazolam by 96% compared withplacebo [37]. As a result, the pharmacodynamic effects of midazolam, measured by severalpsychomotor tests, were practically abolished. It is therefore reasonable to assume thatorally administered midazolam (and triazolam) are ineffective as hypnotic agents duringconcomitant treatment with rifampicin and other potent CYP3A4 inducers such asphenytoin and carbamazepine.

AUC of oral midazolam

o_aCDüCD

CooCD.C

Keto Itra Eryt Dilt Clar Vera Roxi Azit Plac

Figure 1: The effect of various drugs on the AUC of oral midazolam compared withplacebo. Keto, ketoconazole; Itra, itraconazole; Eryt, erythromycin; Dilt, diltiazem; Clar,clarithromycin; Vera, verapamil; Roxi, roxithromycin; Azit, azithromycin; Plac, placebo.N.S., not significantly different from placebo. Data from references [31-36].

To summarise, midazolam is a sensitive probe of CYP3A4 activity in vivo, since itspharmacokinetics are very susceptible to inhibition and induction of CYP3A4. By usingorally administered midazolam (or triazolam) as a probe, the potential of drugs to affectCYP3A4 activity in vivo can be assessed. For example, the pharmacokinetics of midazolamwere unaffected by azithromycin [35], strongly suggesting that azithromycin does not causeclinically significant interactions with CYP3A4 substrates.

PROTON PUMP INHIBITORS (PPI) AS INHIBITORS OR INDUCERS

Omeprazole can induce (CYP1A subfamily) and inhibit (CYP2C19) drug metabolism[38,39]. It is controversial whether this is of clinical significance and whether other PPI,such as pantoprazole or lansoprazole share the same interaction potential [22,40]. Therefore

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we recently tested in a randomized, placebo-controlled, 4-way cross-over study in 20healthy, non-smoking and drug-free volunteers (extensive metabolizers of CYP2C19,H. pylori negative) whether 10 days' treatment with omeprazole (40mg/day), pantoprazole(SOmg/day) or lansoprazole (60mg/day) affected the steady state disposition of theophylline(350 mg bid given as a slow release preparation), a common probe of CYP1A2. During all4 parts of the study plasma concentration time profiles following the last theophylline dosewere nearly identical and the 90% C.I. for the ratios for bioequivalence criteria (e.g. AUC;peak; mean and trough Css; % PTF) fluctuated around unity. Therefore it appears veryunlikely that the 3 PPI, tested at a sufficiently high dosage, have a significant effect on theCYP1A2 activity [41].

CONCLUSIONS

According to the examples given drug metabolizing enzymes can be more or lessspecifically inhibited or induced by a variety of compounds ingested in form of foodconstituents or drugs (Table 2). The affected (pre-)systemic elimination will influence thedisposition (PK) of many therapeutic agents and consequently their action (PD) can beaffected as well. The interactions may necessitate dosage modifications, and sometimesselection of alternative drugs with a lower potential for drug interactions should beconsidered. In summary, enzyme inhibition and induction can significantly contribute tovariability in human drug response.

Table 2.Typical inhibitors and inducers of some CYP-enzymes

Enzyme

CYP1A1

CYP1A2

CYP2A6

CYP2C8/9/18

CYP2C19

CYP2D6

CYP2E1

CYP3A(4)

Inhibitory referencecompounds

cc-naphtoflavone

furafylline, fluvoxamine

methoxsalen, pilocarpine

sulfaphenazole

fluconazole, omeprazole

quinidine

diethyldithiocarbamate

ketoconazole,itraconazole

Inducers

methylcholanthrene

smoking, omeprazole

phenobarbital

phenobarbital, rifampicin

phenobarbital, rifampicin

not inducible

ethanol, INH

rifampicin, dexamethasone,anticonvulsants

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ACKNOWLEDGEMENT: The valuable secretarial help of Mrs. H. Kohler and Mrs. G. Wilder is highlyappreciated.

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Discussion: Enzyme inhibition of polymorphic drug metabolism

E.M. Sellers:Perhaps we need to give a little bit more attention to how we characterise variations in

activity. We tend to use ratios of parent drug to metabolite, which from a psychometric pointof view are very unsatisfactory. We should probably be focusing much more on fractionalclearances and absolute recoveries by particular pathways, because these give a much moreaccurate estimate of what is the real activity of the enzyme.

U. Klotz:We discovered in this interaction study that there were quite dramatic differences by

comparing the ratio technique with using area under the curves. But it is much more tediousto measure a whole time profile during the dosing intervals than just a single measurement.For routine use the ratio technique is much more convenient, but it is much more variable.

P. Rolan:The extent of the variability of the caffeine metabolic ratio is surprising, because that test

has been suggested as a screening tool and if its variability is that large, it means that youcould fail to find an effect just due to the variability. I wonder whether part of the problemwas again selecting the ratio, because there are at least five metabolic caffeine ratios.

U. Klotz:We used the ratio (MR) which, according also to Geoffrey Tucker and other experts, is

independent of the renal excretion [MR = (AFMU+1X+1U)/1,7U]

G.T. Tucker:The issue of what index of enzyme activity you use is really quite important. From a

theoretical point of view certainly I would defend the metabolic ratio over urine metaboliterecovery as an index, because the latter depends not just on the pathway you are interestedin but on parallel pathways also. Theoretically, the urinary drug to metabolite ratio for asimple system is a function of the partial clearance by the enzyme you are interested in, in thedenominator, and in the numerator you have the renal clearance of the parent drug. Thus, renalclearance is the potential source of noise in using these ratios as markers of enzyme activity.The kinetic bases of the indices that we use should be considered more carefully.

N.L. Benowitz:In the verapamil studies when you look at inhibition, is there a general relationship between

hepatic CYP3A4 activity and intestinal activity, or are these regulated separately?

U. Klotz:For a long time it was thought that CYP3A4 in the intestinal wall and in the liver were using

the same substrates. The same inhibitors and the same inducers could affect both sites, andthere might be some kind of co-regulation. But more recently, some evidence has appearedshowing that both enzymes are probably regulated in a slightly different way.

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P. du Souich:I would like to reiterate again that it is very difficult to interpret the effect of a drug on

theophylline kinetics, because at least four enzymes are contributing to its clearance. Whenyou modify one of these enzymes you do not know exactly how the alternative pathways willrespond. We do not know if they will remain exactly the same, and follow predictablefirst-order kinetics or not. Another practical point I would like to make from the examples youhave shown, should we think about developing drugs for alternative routes of administration,for example, sublingual, transcutaneous or even inhaled drugs, to try to avoid these effects ofintestinal and liver variability?

U. Klotz:This could be one alternative to get round intestinal metabolism, although comparative

studies with different routes of administration are required.

A.J.J. Wood:We need to be careful just extrapolating from studies done with one route to another route

and assuming that the effects will be exactly the same. I suspect that the effects are notidentical.

J. Urquhart:If you are going to use other routes, you need more potent drugs and there are not very

many of them.

U. Klotz:Bioavailability then becomes an issue, because to develop a new formulation you have to

complete new studies in terms of efficacy and toxicity. Then, it is probably much easier toperform 2 or 3 interaction studies, instead.