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Current
Drug Metabolism
ISSN: 1389-2002
Current Drug Metabolism
Volume 8, Number 7, October 2007
Contents
Prediction of Intestinal First-Pass Metabolism
Guest Editor: Aleksandra Galetin
Editorial: Intestinal
First-Pass Metabolism – Bridging the Gap Between In
Vitro and In Vivo Pp. 643
A. Galetin
Modeling Gastrointestinal Drug Absorption Requires
More In Vivo Biopharmaceutical Data: Experience from
In Vivo Dissolution and Permeability Studies in Humans
Pp. 645-657
H. Lennernäs
[Abstract]
In Vitro Methods to Study Intestinal Drug
Metabolism Pp. 658-675
E.G. van de Kerkhof, I.A.M. de Graaf and G.M.M. Groothuis
[Abstract]
Prediction of Intestinal First-Pass Drug Metabolism
Pp. 676-684
J. Yang, M. Jamei, K.R. Yeo, G.T. Tucker and A. Rostami
Hodjegan
[Abstract]
Maximal Inhibition of Intestinal First-Pass Metabolism
as a Pragmatic Indicator of Intestinal Contribution to the
Drug-Drug Interactions for CYP3A4 Cleared Drugs Pp.
685-693
A. Galetin, L.K. Hinton, H. Burt, R.S. Obach and J.B.
Houston
[Abstract]
The Role of the Intestine in Drug Metabolism and Pharmacokinetics:
An Industry Perspective Pp. 694-699
M.B. Fisher and G. Labissiere
[Abstract]
General Articles
Small Interfering RNA in Drug Metabolism and Transport
Pp. 700-708
A.-M. Yu
[Abstract]
Inhibition of Ataxia Telangiectasia-p53-E2F-1 Pathway
in Neurons as a Target for the Prevention of Neuronal Apoptosis
Pp. 709-715
A. Camins, E. Verdaguer, J. Folch, C. Beas-Zarate, A.M.
Canudas and M. Pallàs
[Abstract]
Brain Metabolism of Ethanol and Alcoholism: An Update
Pp. 716-727
L. Hipólito, M.J. Sánchez, A. Polache and
L. Granero
[Abstract]
Investigation of Xenobiotics Metabolism, Genotoxicity,
and Carcinogenicity Using Cyp2e1-/- Mice
Pp. 728-749
B.I. Ghanayem and U. Hoffler
[Abstract]
Abstracts
[Back to top]
Editorial: Intestinal First-Pass Metabolism
– Bridging the Gap Between In Vitro and In
Vivo
A. Galetin
The ability to successfully predict pharmacokinetic properties
plays a crucial role in the selection of candidate drugs and
significantly reduces the number of potential failures in
drug development. Current drug candidates typically show a
very high affinity for the target receptors; however, the
drawback is that many new lead compounds represent large,
lipophilic molecules with low solubility, dissolution and/or
permeability and consequently show poor absorption properties.
In vitro-in vivo prediction and application of in
silico methods for clearance and drug-drug interaction
prediction from hepatic cytochrome P450 data have been widely
accepted by both pharmaceutical companies and academia, and
meet certain regulatory requirements [1]. However, the application
of these approaches to extrahepatic tissues, including the
intestine, has proved challenging and less definitive. Estimates
of intestinal clearance are not routinely incorporated into
in vitro-in vivo strategies and this may partially
explain the clearance under-prediction trend often observed
[2].
The aim of this special issue of Current Drug Metabolism is
to provide a broad perspective, both from academia and industry,
on intestinal absorption and the impact of intestinal first-pass
metabolism on both clearance and drug-drug interaction prediction.
The issue also considers our in silico abilities
to bridge the gap between the increasing amount of intestinal
in vitro data and the importance of intestinal first-pass
metabolism in vivo. Overviews of the biopharmaceutical
and pharmacokinetic variables and their impact on the prediction
of the drug absorption from either chemical structure and/or
in vitro data is presented, focusing in particular
on the estimation and application of intestinal permeability
[3,4]. The role of uptake and efflux transporters and their
interplay with the metabolic enzymes is becoming an increasingly
important issue for drug candidates. Recently, drug elimination
mechanisms (e.g., via metabolism, or coupled with uptake/efflux
transporters or renal clearance) have been suggested as criteria
to extend the Biopharmaceutical Classification System beyond
the use of drug permeability/solubility characteristics to
the prediction of drug disposition [5]. The limitations of
the cellular systems and potential over-estimation of the
significance of efflux transporters on the intestinal absorption
are also discussed [3].
ROLE OF THE INTESTINE IN THE CLEARANCE PREDICTION
Over recent years a number of studies have assessed the catalytic
activity of intestinal metabolic enzymes in comparison to
the liver, focusing predominantly on CYP3A4/CYP3A5 as well
as a range of glucuronidating (UGT) enzymes. Although intuitively
one would expect the activity of enzymes in both organs to
be comparable, a number of in vitro studies have
reported differential importance of the intestine relative
to the liver. In addition, inter-individual variability in
P450/UGT expression in both liver and intestine and their
relative abundance in the corresponding organs have generally
not being taken into account. A recent study by Galetin and
Houston [6] has shown comparable intestinal and hepatic catalytic
activity (when expressed per pmol of P450 enzyme) of the major
P450 enzymes. Therefore, it would be reasonable to expect
that once normalized for the relative abundance of the enzyme
investigated, hepatic or recombinant data would be equally
useful for the prediction of intestinal clearance once an
appropriate mechanistic model is applied. Although this approach
is applicable for P450 enzymes [4], certain specific issues
need to be addressed in case of phase II enzymes. For example,
the relative UGT expression levels in vivo are not
clearly defined. A recent study by Cao et al. [7]
indicated a 3-fold higher expression of UGTs relative to CYP3A4
in human duodenum. However, as the expression of intestinal
metabolic enzymes, and transporters, follows certain gradient
patterns along the intestine and within the villi, this will
not necessarily reflect the UGT:P450 abundance ratio along
the whole length of the gut. In addition, the identification
of appropriate extrahepatic scaling factors represents a major
challenge. Only recently, has a consensus on such numbers
been reached for the scaling of hepatic data [8], and there
is no such comprehensive data available for the analysis of
the intestinal microsomal recovery.
Therefore, the choice of an appropriate in vitro
system for adequate assessment of intestinal metabolism is
an important consideration. This is of particular relevance
for compounds that are mutual substrates for efflux transporters
and metabolic enzymes, as the transporter-enzyme interplay
has been indicated as a contributing factor to the low and
variable intestinal availability (FG).
Intestinal in vitro systems of differing complexity
and from different animal species and their utility to study
phase I and II metabolism/transport are summarized here in
the review by van de Kerkhof et al. [9]. The advantages
and limitations of subcellular fractions, intestinal cell
lines and intact cell systems are considered for an array
of scenarios. A common concern for both metabolism and transporter
in vitro systems is the inconsistency and variability
of in vitro clearance and, in particular permeability,
data, which confounds straightforward in vitro-in vivo
extrapolation. In addition, the expression levels of both
enzymes and transporters vary between cell models and in most
cases do not correspond to the human intestine, which may
under- or over-estimate the contribution of either protein
of interest [3, 9].
MECHANISTIC MODEL TO PREDICT THE EXTENT OF INTESTINAL
EXTRACTION
Modelling of intestinal first-pass requires careful attention
to the physiological complexities unique to the intestine;
for example, enterocytic rather than organ blood flow needs
to be incorporated in addition to the cellular heterogeneity
and spatial arrangement for the metabolic enzymes and efflux
transporters in the intestine, which is distinct to the liver
[5]. In contrast to certain complex physiologically-based
models previously proposed [10], Yang et al. discuss
a ‘minimal’ Qgut
model in the current issue [4]. The model proposed overcomes
the inadequacy of the well-stirred approach (adopted from
the corresponding liver model) by combining passive permeability,
active transport, enterocytic blood flow and intrinsic activity
of intestinal enzymes and their relative abundance. In order
to incorporate a transporter component into the Qgut
model, in vitro data obtained in different transporter
cell lines are converted into a more physiological parameter,
i.e., effective intestinal permeability (Peff).
This conversion is based on the correlation of human in
vivo values [3] and in vitro apparent permeability
(Papp) data generated in
various transporter systems (e.g., MDCK-MDR1 or Caco-2), covering
a wide range (200-fold). Data obtained from various studies
and under inconsistent experimental conditions are used to
generate this correlation and this may significantly impact
on the accuracy of the FG
estimation for the compounds investigated. The current use
of the Qgut model assumes
that the effective free fraction in the gut equals 1; an accurate
estimate of this parameter is however difficult. This may
represent an issue of concern, in particular due to the sensitivity
of the model and the intestinal extraction predicted on this
parameter [4].
ROLE OF THE INTESTINE IN THE DRUG-DRUG INTERACTION
PREDICTION
There is an increasing concern within pharmaceutical
companies about the impact of inhibition of intestinal P450
enzymes [2], considering the high levels of a potential inhibitor
that are achieved during the absorption phase. However, only
in recent years has this issue been addressed in the prediction
of both reversible and time-dependent inhibition interactions.
The main question is – which IC50
or KI values constitute a
safe margin in terms of interaction at the level of the gut
wall? And in the case of time-dependent inhibition, would
either a fast inactivating (high kinact)
or a potent inhibitor (low KI)
represent the greater concern? A relative ratio of the inhibitor
intestinal concentration to its potency (Ig/Ki)
>100, in case of reversible inhibition, would indicate
a high interaction potential. Similarly, in the case of time-dependent
inhibitors, Ki<1µM
and kinact>0.01 would
indicate an interaction potential at inhibitor concentrations
>1 µM. However, in addition to the inhibitor properties,
FG of a victim drug is an
important determinant of the interaction magnitude. A minimal
50% intestinal extraction is indicated as an appropriate cut
off value for a potential interaction, irrespective of the
potency of the inhibitor and its inhibition mechanism. The
advantages and limitations of a ‘worst case scenario’
i.e., assuming maximal intestinal inhibition, on the magnitude
of an intestinal drug-drug interaction are addressed in this
issue [11]. Validation of the prediction success of a drug’s
intestinal extraction from in vitro data relies on
the accuracy of the in vivo estimates. Current special
issue discusses the limitations of both indirect method (from
oral and i.v. administration data) and estimates based on
the grapefruit juice interaction studies [4, 11]. If one considers
grapefruit juice as a potent and selective inhibitor of the
intestinal CYP3A4 (no effect on hepatic counterpart), this
approach can therefore represent an attractive alternative
to the indirect method. However, grapefruit juice shows only
weak inhibition towards the P-glycoprotein and has recently
been linked with an inhibition of uptake transporters (e.g.
OATP1B2). One would therefore argue a potentially limited
use of this approach to assess intestinal availability for
drugs whose disposition is dependent on both efflux/uptake
transporters and metabolic enzymes. An additional question
is whether the FG estimates
obtained via different approaches in vivo (anhepatic
patients, i.v./oral or from grapefruit juice interactions)
are interchangeable, as it has been observed in case of midazolam.
The articles presented in this special issue illustrate our
increasing confidence in bridging the gap between in vitro
data and in vivo absorption and intestinal first-pass
metabolism. Accurate prediction of the intestinal availability
directly affects not only the estimate of clearance of a compound,
but also the drug-drug interaction potential. Existing physiologically
based pharmacokinetic models have the potential to assess
the role of metabolic enzymes, transporters and their interplay
on the clearance by a particular organ. However, in order
to progress further it is essential to improve the quality
and standardize experimental conditions for in vitro
data generation, as well as improve the knowledgeable integration
of the data from different cell systems.
ACKNOWLDGEMENTS
Financial support for AG academic position in the School of
Pharmacy and Pharmaceutical Sciences, University of Manchester
is kindly provided by Pfizer, Sandwich, United Kingdom.
REFERENCES
[1] Rostami-Hodhegan, A. and Tucker, G.T. (2007) Nat.
Rev. Drug Discov., 6, 140-148.
[2] Fisher, M.B. and Labissiere, G. (2007) Curr. Drug
Metab., 8, 694-699.
[3] Lennernäs, H. (2007) Curr. Drug Metab.,
8, 645-657.
[4] Yang, J.; Jamei, M.; Rowland Yeo, K.; Tucker, G.T. and
Rostami-Hodjegan, A. (2007) Curr. Drug Metab., 8,
676-684.
[5] Wu, C-Y. and Benet, L.Z. (2005) Pharm. Res.,
22, 11-23.
[6] Galetin, A. and Houston, J.B. (2006) J. Pharmacol.
Exp. Ther., 318, 1220-1229.
[7] Cao, X.; Gibbs, S.T.; Fang, L.; Miller, H.A.; Landowski,
C.P.; Shin, H.-C.; Lennernas, H.; Zhong, Y.; Amidon, G.L.;
Yu, L.X. and Sun, D. (2006) Pharm. Res., 23(8),
1675-1686.
[8] Barter, Z.E.; Bayliss, M.K.; Beaune, P.H.; Boobis, A.R.;
Carlile, D.J.; Edwards, R.J.; Houston, J.B.; Lake, B.G.; Lipscomb,
J.C.; Pelkonen, O.R.; Tucker, G.T. and Rostami-Hodjegan, A.
(2007) Curr. Drug Metab., 8, 33-45.
[9] van de Kerkhof, E.G.; de Graaf, I.A.M. and Groothuis,
G.M.M. (2007) Curr. Drug Metab., 8,
658-675.
[10] Pang, K.S. (2005) Drug Metab. Dispos., 31,
1507-1519.
[11] Galetin, A.; Hinton, L.; Burt, H.; Obach, R.S. and Houston,
J.B. (2007) Curr. Drug Metab., 8,
685-693.
[Back to top]
Modeling Gastrointestinal Drug Absorption Requires
More In Vivo Biopharmaceutical Data: Experience from
In Vivo Dissolution and Permeability Studies in Humans
H. Lennernäs
The majority (84%) of the 50 most-sold pharmaceutical products
in the US and European markets are given orally. The dominating
role of this route in drug therapy is a consequence of it
being safe, efficient and easily accessible with minimal discomfort
to the patient in comparison with other routes of drug administration.
A successful drug discovery and development of oral pharmaceutical
products require an in-depth understanding of multiple biochemical
and physiological processes that determine the dissolution
rate, intestinal permeability, gastrointestinal transit, first-pass
extraction and systemic exposure-time profiles of drugs. It
is crucial to realize that these basic biopharmaceutic and
pharmacokinetic properties are crucial to focus on to allow
successful drug development. Identification of the rate-limiting
step(s) in order to overcome these barriers and understanding
of the sources of variability are important in the selection
of suitable candidate molecules in drug development.
Several reports based on in vitro investigations
in various cell models have suggested that carrier-mediated
intestinal efflux may be a major reason for incomplete absorption
and variable bioavailability of drugs, as well being a site
for drug-drug and specific food-drug interactions. However,
many drugs which were initially suggested to undergo significant
efflux in vitro were later shown to be completely
absorbed in vivo. This apparent discrepancy between
in vitro and in vivo results may be due
to several factors that will be discussed in this review.
Novel data on solubility and dissolution in human gastrointestinal
derived fluids will be reviewed. The effect of food intake
on solubility and dissolution rate of a range of drugs including
felodipine, danazol, griseofulvin, cyclosporine, probucol
and ubiquinone in simulated and real intestinal fluids is
discussed.
The biopharmaceutic and physicochemical data discussed here
can potentially be used as a benchmark set for validation
of new experimental techniques or in silico models
in future. Factors such as structural diversity, commercial
availability, price and a suitable analytical technique for
quantification were considered in the selection of a specific
drug set. Using the compiled data set lipophilicity as determined
by reverse phase HPLC and permeability across Caco-2 cell
monolayers were determined; means to overcome the experimental
difficulties due to the diversity of the data are also discussed.
[Back to top]
In Vitro Methods to Study Intestinal Drug
Metabolism
E.G. van de Kerkhof, I.A.M. de Graaf and G.M.M. Groothuis
Although the liver has long been thought to play the major
role in drug metabolism, also the metabolic capacity of the
intestine is more and more recognized. In vivo studies
eventually pointed out not only the significance of first-pass
metabolism by the intestinal wall for the bioavailability
of several compounds, but also the relevance of transporters
in this process. Only a few methods are available to study
drug metabolism in vivo or in situ and with
most of these methods it remains difficult to discriminate
between the contribution of liver and extrahepatic tissues.
To study intestinal drug metabolism in vitro, apart
from subcellular fractions, several intact cell systems are
nowadays available.
This review discusses the available intestinal in vitro
methods to study drug metabolism. The advantages and limitations
of intact cell systems (isolated intestinal perfusion, everted
sac, Ussing chamber preparations, biopsies, precision-cut
slices, primary cells), subcellular fractions (S9 fractions,
microsomes) and intestinal cell lines (caco-2, LS180 cells
amongst others) are discussed. Their applicability to different
species and to study phase I and II metabolism/transport and
drug-drug interactions are summarized. Furthermore, causes
of variation within and between methods are discussed and
metabolic rates obtained with different methods are compared.
Whereas subcellular fractions and cell lines are efficient
methods to study mechanistic aspects of drug metabolism at
the enzyme level, the isolated intestinal perfusion, everted
sac and Ussing chamber appear particularly useful for studying
drug metabolism of rapidly metabolised drugs and interactions
with transporters. Biopsies, precision-cut slices and primary
cells seem all appropriate to study induction and metabolism
of slowly metabolised drugs.
[Back to top]
Prediction of Intestinal First-Pass Drug Metabolism
J. Yang, M. Jamei, K.R. Yeo, G.T. Tucker and A. Rostami
Hodjegan
Despite a lower content of many drug metabolising enzymes
in the intestinal epithelium compared to the liver (e.g.
intestinal CYP3A abundance in the intestine is 1% that of
the liver), intestinal metabolic extraction may be similar
to or exceed hepatic extraction. Modelling of events on first-pass
through the intestine requires attention to the complex interplay
between passive permeability, active transport, binding, relevant
blood flows and the intrinsic activity and capacity of enzyme
systems. We have compared the predictive accuracy of the “well-stirred”
gut model with that of the “QGut”
model. The former overpredicts the fraction escaping first-pass
gut metabolism; the latter improves the predictions by accounting
for interplay between permeability and metabolism.
[Back to top]
Maximal Inhibition of Intestinal First-Pass Metabolism
as a Pragmatic Indicator of Intestinal Contribution to the
Drug-Drug Interactions for CYP3A4 Cleared Drugs
A. Galetin, L.K. Hinton, H. Burt, R.S. Obach and J.B.
Houston
For certain CYP3A4 substrates intestinal first-pass metabolism
makes a substantial contribution to low oral bioavailability
and extent of drug-drug interactions (DDI). In order to include
the contribution of enzyme inhibition in the gut wall in the
assessment of DDI potential, the ratio of the intestinal wall
availability in the presence and absence of an inhibitor (FG’
and FG, respectively) has
been incorporated into a prediction equation based on hepatic
enzyme interactions. This approach has been applied for both
reversible and irreversible DDIs, involving 36 different inhibitors
and 11 CYP3A4 substrates. The aim was to investigate the use
of maximal (complete) inhibition of intestinal CYP3A4 (FG’=1)
as a pragmatic measure of the intestinal enzyme interaction
and to compare this approach with observed in vivo values
(where available) and predicted FG
ratios from an intestinal model. The latter was obtained from
the decrease in the intestinal intrinsic clearance in the
presence of an inhibitor, using an estimated inhibitor concentration
in the intestinal wall during absorption phase (IG)
and an in vitro obtained Ki.
In addition, the impact of variability in the enterocytic
blood flow on the estimated IG
and subsequently the model predicted FG
ratio was investigated. The maximal FG
ratios for the 11 CYP3A4 substrates investigated ranged from
1.06-7.14 for alprazolam and tacrolimus, respectively. In
91% of the studies investigated the model predicted FG
ratio was within 40% of the maximal value. Maximal FG
ratio is proposed as an initial indicator of the magnitude
of intestinal enzyme interaction; the implications for drug
elimination involving substrates cleared either by metabolism
or by a combination of metabolism and efflux transporters
are discussed.
[Back to top]
The Role of the Intestine in Drug Metabolism and Pharmacokinetics:
An Industry Perspective
M.B. Fisher and G. Labissiere
Over the past decade, our knowledge concerning the importance
of intestinal metabolism in the disposition of xenobiotics
has significantly improved. Compounds such as midazolam can
be extensively extracted in the intestine upon first-pass
metabolism after oral dosing. Conversely, the intestine plays
a less important, albeit less characterized role in systemic
metabolism. This manuscript is meant to review the published
examples of pharmaceutical industry research on the intestinal
metabolism of xenobiotics, including the various in vitro
and in vivo models used. While it is clear that some
examples exist of published characterization of the role of
intestinal metabolism in drug disposition from the pharmaceutical
industry, the majority of industry literature ignores its
contribution. The opportunity exists to increase the examination
of intestinal metabolism of drugs and drug candidates in industry.
[Back to top]
Small Interfering RNA in Drug Metabolism and Transport
A.-M. Yu
RNA interference (RNAi) is a powerful technique that utilizes
RNA molecules to specifically knock down the expression of
targeted gene at posttranscriptional level. These small interfering
RNAs (siRNAs) not only have broad application to basic biomedical
research but may be developed as therapeutic agents. Drug-metabolizing
enzymes (DMEs) and drug transporters (DTs) are molecular determinants
of pharmacokinetic property of a drug. Transcriptional gene
expression of DMEs and DTs is controlled by xenobiotic-sensing
nuclear receptors (NRs). Because of complexity in studying
the function of individual DMEs, DTs and NRs, siRNAs can be
an excellent addition to chemical inhibitors and inhibitory
antibodies in delineating their specific roles in drug metabolism
and transport, gene regulation, and drug-drug interactions.
RNAi may be employed to modulate DT expression to overcome
multidrug resistance. Recent studies using RNAi to silence
gene expression of specific DME, DT and NR, and the impact
on drug metabolism and transport are discussed in this review.
Concerns remain about the efficiency, specificity, and off-target
effects when interpreting data obtained from RNAi studies.
Furthermore, potential role for endogenous siRNAs, microRNA
(miRNA) molecules, in controlling the posttranscriptional
gene regulation of DMEs, DTs and NRs is discussed.
[Back to top]
Inhibition of Ataxia Telangiectasia-p53-E2F-1 Pathway
in Neurons as a Target for the Prevention of Neuronal Apoptosis
A. Camins, E. Verdaguer, J. Folch, C. Beas-Zarate, A.M.
Canudas and M. Pallàs
Over the last few decades, understanding of the mechanisms
involved in the process of neuronal cell death has grown.
Recent findings have established that DNA damage, and specifically
ataxia telangiectasia mutated protein (ATM), is key to the
cascade of regulation of neuronal apoptosis. Another characteristic
common to all neurodegenerative diseases is oxidative stress.
Likewise, a common feature in the brain of patients with neurodegenerative
diseases such as Alzheimer’s and Parkinson’s diseases
and other neurological disorders is the expression of proteins
involved in cell-cycle control. In the process of re-entry
in the cell cycle, an additional component, transcription
factor E2F-1, also involved in the regulation of apoptosis,
is expressed. Finally, in this complex puzzle, mitochondrial
activation with the release of proteins and the activation
of cystein proteases, specifically caspase-3, is prominent
in the last step of neuronal apoptosis. This review focuses
on the role of ATM activation and its re-entry into the cell
cycle in neurons as a potential target for the prevention
of neuronal apoptosis. We suggest the mechanisms by which
ATM and E2F-1 orchestrate the apoptotic process. Among them,
p53 could be a common point on this apoptotic route. Finally,
we put forward drugs that are now being studied experimentally,
such as p53 inhibitors, ATM inhibitors and cyclin-dependent
kinase (CDKs) inhibitors, for the treatment of neurodegenerative
diseases.
[Back to top]
Brain Metabolism of Ethanol and Alcoholism: An Update
L. Hipólito, M.J. Sánchez, A. Polache and
L. Granero
It has long been suggested that some of the neuropharmacological,
neurochemical and behavioural effects of ethanol are mediated
by its first metabolite, acetaldehyde. In spite of the well
documented psychoactivity of acetaldehyde, the precise role
of this compound in alcohol abuse remains a matter of intense
debate among scientists devoted to the study of alcoholism.
Very frequently, the main drawback has been related to the
presence of adequate levels of acetaldehyde or its derivatives
inside the brain after ethanol ingestion. Since penetration
into the central nervous system from blood of peripherically
derived acetaldehyde is very low due to the high aldehyde
dehydrogenase activity at the blood-brain barrier, several
authors called into question the acetaldehyde implication
in the toxicity and neurobehavioral effects of ethanol. The
confirmation in several laboratories of the existence of enzymatic
mechanisms of ethanol oxidation in the brain has revitalized
the old theories supporting the acetaldehyde contribution
to alcohol abuse and alcoholism. In this paper, we review
current data on the brain metabolism of ethanol. We focused
on the description of the enzymatic mechanisms involved in
this metabolic process, reviewing the constitutive expression,
catalytic activity and inhibition and inducibility of the
enzymes involved in brain ethanol metabolism. We also analyze
old and recent data on their regional distribution and cellular
localization in the central nervous system, with special reference
to the mesocorticolimbic system, a dopaminergic brain pathway
that plays an important role in drug and ethanol reinforcement.
[Back to top]
Investigation of Xenobiotics Metabolism, Genotoxicity,
and Carcinogenicity Using Cyp2e1-/- Mice
B.I. Ghanayem and U. Hoffler
Cytochromes P450 (CYPs) comprise a number of enzyme subfamilies
responsible for the oxidative metabolism of a wide range of
therapeutic agents, environmental toxicants, mutagens, and
carcinogens. In particular, cytochrome P450 2E1 (CYP2E1) is
implicated in the oxidative bioactivation of a variety of
small hydrophobic chemicals including a number of epoxide-forming
drugs and environmentally important toxicants including urethane,
acrylamide, acrylonitrile, benzene, vinyl chloride, styrene,
1-bromopropane, tri-chloroethylene, dichloroethylene, acetaminophen,
and butadiene. Until recently, chemical modulators (inducers
and inhibitors) were used in order to characterize the enzymatic
basis of xenobiotic metabolism and the relationships between
CYP-mediated bioactivation and chemical-induced toxicity/carcinogenicity.
With the advent of genetically engineered knockout mice, the
ability to evaluate the roles of specific CYPs in the metabolism
of xenobiotics has become more attainable. The main focus
of the current review is to present studies that characterized
the enzymatic, metabolic, and molecular mechanisms of toxicity,
genotoxicity, and carcinogenicity of various xenobiotics using
Cyp2e1-/- mice. Data presented in this review demonstrated
that the most comprehensive studies using Cyp2e1-/-
mice, encompassing the entire paradigm of metabolism to toxicity,
genotoxicity, and carcinogenicity were possible when a substrate
was primarily metabolized via CYP2E1 (e.g. urethane and acrylamide).
In contrast, when multiple CYP enzymes were prevalent in the
oxidation of a particular substrate (e.g.: trichloroethylene,
methacrylonitrile, crotononitrile), investigating the relationships
between oxidative metabolism and biological activity became
more complicated and required the use of chemical modulators.
In conclusion, the current review showed that Cyp2e1-/-
mice are a valuable animal model for the investigation of
the metabolic and molecular basis of toxicity, genotoxicity,
and carcinogenicity of xenobiotics.
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