Elsevier

Toxicology

Volume 270, Issues 2–3, 11 April 2010, Pages 59-65
Toxicology

Identification of selected therapeutic agents as inhibitors of carboxylesterase 1: Potential sources of metabolic drug interactions

https://doi.org/10.1016/j.tox.2010.01.009Get rights and content

Abstract

A series of studies were designed and carried out in order to explore the potential for the major human hepatic hydrolase, carboxylesterase 1 (hCES1), to serve as a target of metabolic inhibition by a variety of medications. The risk of adverse drug–drug interaction(s) is present when metabolic inhibitors are combined with known or suspected substrates of a given enzyme. In the present report the abundantly expressed hepatic enzyme, hCES1, was examined as a potential target of metabolic inhibition by a number of routinely prescribed medications. hCES1 has been seldom assessed in this regard despite its role in the metabolism and detoxification of many compounds. The psychostimulant methylphenidate (MPH) was chosen as an hCES1 selective substrate. In vitro studies were performed using previously developed cell lines which overexpress hCES1 with both p-nitrophenyl acetate and d-MPH serving as known substrates. Aripiprazole, perphenazine, thioridazine, and fluoxetine were determined to be the potent hCES1 inhibitors. A complementary animal study followed in vitro screening studies to further evaluate the inhibitory effect of aripiprazole on CES1 activity in FVB mice. The results suggest that the concurrent administration of racemic (i.e. dl-) MPH with aripiprazole significantly increased the plasma concentrations of both total MPH as well as the less active l-isomer. The ratio of d-MPH and l-MPH plasma concentrations was significantly decreased in the mice treated with aripiprazole compared to the control animals, indicating an overall decrease of CES1 catalytic activity in aripiprazole treated animals. Additionally, a quantitative structure–activity relationship based analysis identified a number of structural similarities of CES1 inhibitors. In conclusion, drug–drug interactions with MPH are likely mediated via CES1 inhibition as a result of concomitant drug therapies. CES1 inhibition represents an overlooked and little studied source of variability in MPH disposition, tolerability, and response.

Introduction

Medications inadvertently result in significant patient morbidity and mortality while contributing significantly to the overall costs of medical/psychiatric care. Adverse drug reactions (ADRs) are considered to be a significant source of iatrogenic illness, potential toxicity leading to hospitalization and/or the prolongation of hospitalizations, and ultimately, therapeutic failure. Many ADRs are inevitable due to their idiosyncratic nature and the inability to predict unintended yet extremely serious adverse consequences of drug therapy such as hypersensitivity reactions (e.g. Stevens-Johnson Syndrome). Such ADRs often occur when initiating pharmacotherapy in a patient that may represent their first exposure to the drug. However, in other instances, ADRs may be attributed to unanticipated drug–drug interactions (DDIs) as a result of combining various medications, an increasing practice throughout most every clinical specialty including pediatric psychiatry. DDIs may occur due to clinician unfamiliarity with drug combinations known to be problematic, the prescribing of medications by multiple clinicians unaware of other ongoing treatments by a second clinician, and the use of drug combinations which have simply never been assessed in a systematic fashion using more modern screening techniques required by the US Food and Drug Administration (FDA). This latter situation may be particularly problematic for drugs which were approved long ago and that remain in routine clinical use.

dl-threo-methylphenidate (MPH) is the most widely prescribed medication for the treatment of attention-deficit/hyperactivity disorder (ADHD) in children, and a molecule that has been in continuous clinical use for approximately one-half century. Racemic MPH is a 50:50 mixture of d-threo-(R,R)-MPH and l-threo-(S,S)-MPH isomers (Fig. 1).

Although MPH is often viewed as the drug of choice for patients with ADHD (Elia et al., 1999), and there are numerous case reports suggesting that this agent may serve as a metabolic inhibitor of drug metabolism, or may have its own metabolism inhibited by other agents, upon closer evaluation of these reports it is apparent that the overwhelming majority of these cases lack rigorous, or in many instances, the most rudimentary evidence or documentation of a DDI (Markowitz and Patrick, 2001). Additionally, almost no systematic studies assessing the potential for DDIs with MPH as a participant have been performed or published in humans. This lack of documented case reports or other studies confirming a pharmacokinetic DDI between MPH and concurrent medication(s) should not be interpreted as evidence that few or no DDIs of consequence occur. There are a number of explanations for the dearth of documentation in the biomedical literature with regard to DDIs and measurable, significant changes in blood concentrations of a widely utilized medication such as MPH. First, since there is no identified “therapeutic window” or threshold concentration associated with a superior response to MPH (Patrick and Markowitz, 1997), therapeutic drug monitoring (TDM) is of little value. Secondly, the accurate analytical measurement of MPH is generally accomplished via a mass spectrometry method which is both costly, and not routinely available through most hospital or other laboratories performing more routine TDM assays (e.g. lithium). Thus, there has been little motivation for clinicians to order an MPH concentration in a patient treated with the drug as part of any routine practice. However, had clinicians measured MPH concentrations more frequently over the years—no matter what the reason for doing so, there is a much greater likelihood that DDIs may have been discovered and subsequently documented over time. Finally, carboxylesterase 1 (CES1), the major hepatic enzyme responsible for the metabolism of MPH, has not generally been viewed or rigorously studied as a potential source of drug variability and DDIs, even in a medication almost exclusively metabolized through this pathway.

The primary metabolic fate of MPH is the initial rapid de-esterification to the major (and inactive) metabolite, ritalinic acid (RA) (Fig. 1) (Patrick et al., 1981, Patrick et al., 1987). This process appears to be exclusively mediated by the human CES1 (hCES1) enzyme (Sun et al., 2004) which is primarily localized in the liver. hCES1 represents one of two major hepatic isoenzymes hCES1 and hCES2, belonging to the classes CES1 and CES2, respectively, and are currently viewed as the most relevant to drug metabolism in humans. hCES1 and hCES2 are known to exhibit broad substrate specificities (Imai et al., 2006). Both are widely distributed throughout the body including the heart, lung, kidney, stomach, small intestine, colon, liver, testes, skin, and brain (Satoh et al., 2002, Fleming et al., 2005). However, the highest hydrolytic activity overall occurs in the liver with hCES1 expression and activity far exceeding hCES2, while the opposite situation exists in the lower gut (i.e. hCES2 predominates). Sun et al. (2004) demonstrated that hCES1 is highly enantioselective in that the catalytic efficiency of hCES1 is 6–7-fold higher for l-MPH relative to d-MPH—a finding in agreement with that of essentially every published enantioselective pharmacokinetic study conducted to date. In general, peripheral blood sampling finds the l-isomer attaining only 1–2% of the systemic concentration of d-MPH (Srinivas et al., 1993, Patrick et al., 2005). Not surprisingly, the plasma t1/2 of d-MPH is markedly longer than that of l-MPH (Patrick et al., 2005). Thus, it is generally accepted that pronounced stereoselective first-pass or presystemic metabolism via hepatic hCES1 occurs following the oral administration of the racemate. By comparison, MPH undergoes only minimal biotransformation mediated by other metabolic enzymes (e.g. CYP450s). Although significant interindividual variability in MPH pharmacokinetics has long been recognized, only recently have potential influences upon hCES1 activity been explored as sources of the variability in the disposition of MPH.

This report describes for the first time, the use of transfected cell lines over-expressing hCES1 as an in vitro tool to identify commonly used therapeutic agents as potential inhibitors of hCES1, and accordingly, impede MPH metabolism, or the metabolism of other therapeutic agents or possible detoxification from chemical exposures. Such a tool might be adopted by the pharmaceutical industry and others to screen various compounds of interest for their ability to inhibit this major hydrolytic pathway as it is not routinely assessed presently. Although preliminary, a confirmatory in vivo study was also described in a mouse model of one of the more potent inhibitors identified through in vitro screening. Finally, based upon the molecular structures of all of the compounds screened in vitro, a preliminary profile was developed describing the structural features of a molecule (i.e. pharmacophore) which may be more likely to produce hCES1 inhibition.

Section snippets

Choice of potential hCES1 inhibitors for assessment

These studies sought to screen a variety of clinically used agents for their potential to inhibit hCES1, and by extension, MPH metabolism. A novel approach was chosen which exploited the use of our previously generated human cell line which over-expresses hCES1 (Zhu et al., 2008). With regard to the selection of candidate drugs to screen as potential inhibitors of hCES1, given the dearth of information presently known regarding inhibition of hCES1, the rationale for the selection of medications

In vitro inhibition of hCES1 activity by selected agents

The PNPA incubation studies indicated that all tested compounds (with the exception of phenylephrine, clonidine, risperidone and its active and marketed metabolite, paliperidone) exhibited varying degrees of inhibitory effects on hCES1 mediated PNPA hydrolysis (Fig. 2). Among the agents found to be significant inhibitors as defined by the exhibition of IC50 values <15 μM were aripiprazole, perphenazine, thioridazine, fluoxetine were the most potent inhibitors with IC50 values of 5.7 μM, 13.9 μM,

Discussion

Drug interactions are recognized as one of the significant contributing factors which may lead to the occurrence of an ADR (Markowitz et al., 1999). Children in particular are believed to represent a high risk population in this regard (Ferranti et al., 2008). It is noteworthy that most clinical studies conducted per FDA guidelines assessing the safety and efficacy of a single drug entity intended for a pediatric population are carried out in less than 1000 individuals (Smith et al., 2008).

Conflict of interest

None declared.

References (37)

  • A.K. Christman et al.

    Atomoxetine, a novel treatment for attention-deficit-hyperactivity disorder

    Pharmacotherapy

    (2004)
  • J. Elia et al.

    Treatment of attention-deficit-hyperactivity disorder

    N. Engl. J. Med.

    (1999)
  • J. Ferranti et al.

    Reevaluating the safety profile of pediatrics: a comparison of computerized adverse drug event surveillance and voluntary reporting in the pediatric environment

    Pediatrics

    (2008)
  • G. Grunder et al.

    Brain and plasma pharmacokinetics of aripiprazole in patients with schizophrenia: an [18F]fallypride PET study

    Am. J. Psychiatry

    (2008)
  • L.B. Hansen et al.

    Plasma concentrations of perphenazine and its sulphoxide metabolite during continuous oral treatment

    Psychopharmacology (Berl)

    (1977)
  • L.B. Hansen et al.

    Plasma levels of perphenazine (Trilafon) related to development of extrapyramidal side effects

    Psychopharmacology (Berl)

    (1981)
  • T. Imai et al.

    Substrate specificity of carboxylesterase isozymes and their contribution to hydrolase activity in human liver and small intestine

    Drug Metab. Dispos.

    (2006)
  • K.M. Kirschbaum et al.

    Serum levels of aripiprazole and dehydroaripiprazole, clinical response and side effects

    World J. Biol. Psychiatry

    (2008)
  • Cited by (43)

    • The Pharmacokinetics and Pharmacogenomics of Psychostimulants

      2022, Child and Adolescent Psychiatric Clinics of North America
      Citation Excerpt :

      We do know that neither dl-MPH nor ritalinic acid themselves can inhibit CES1 activity in vitro. On the other hand, agents which may be used concurrently with MPH which have been screened by in vitro methods and not found to significantly inhibit CES1 include; the nonstimulant ADHD medications atomoxetine, clonidine, and guanfacine, the antipsychotics clozapine, olanzapine, risperidone, paliperidone, aripiprazole, ziprasidone, quetiapine and haloperidol, the TCA imipramine, the antibiotics amoxicillin, erythromycin, and ciprofloxacin, and antihistamine diphenhydramine and decongestant phenylephrine.70 The concurrent use of MPH with ethanol is associated with at least two metabolic phenomena.

    • Carboxylesterase inhibitors from clinically available medicines and their impact on drug metabolism

      2021, Chemico-Biological Interactions
      Citation Excerpt :

      Recently, Zhao et al. found that reserpine exhibited strong hCES2A inhibition activity, with an IC50 value of 0.94 μM [86]. Methylphenidate (MPH), the most commonly prescribed CNS stimulant drug for attention-deficit hyperactivity disorder (ADHD) in children, is also an hCES1A selective substrate [87]. Zhu et al. chose several older conventional antipsychotic drugs and characterized their inhibitory effects against hCES1A using p-nitrophenyl acetate (PNPA) or d-MPH as hCES1A substrate [87].

    • Rapid bioluminescence assay for monitoring rat CES1 activity and its alteration by traditional Chinese medicines

      2020, Journal of Pharmaceutical Analysis
      Citation Excerpt :

      Since NLMe had been demonstrated to be a selective hCES1 substrate, to validate the specificity of NLMe towards CES1 in rat plasma, potential inhibition assays of various CES inhibitors on NLMe hydrolysis were conducted in blank rat plasma. Assessment was done by comparing activity in the presence or absence of various reported CES inhibitors including BNPP (a general CESs inhibitor) [34], fluoxetine (a selective CES1 inhibitor) [35] and loperamide (a selective CES2 inhibitor) [36]. The study was conducted in a 96-well microplate in a total volume of 100 μL.

    View all citing articles on Scopus
    View full text