ª 2009 John Wiley & Sons A/S
Chem Biol Drug Des 2010; 75: 3–17
Review Article
Structure-based Drug Metabolism Predictions for
Drug Design
Hao Sun* and Dennis O. Scott
Department of Pharmacokinetics, Dynamics, and Drug Metabolism,
Pfizer Inc., Groton, CT 06340, USA
*Corresponding author: Hao Sun hao.sun@pfizer.com
Significant progress has been made in structure-
based drug design by pharmaceutical companies
at different stages of drug discovery such as iden-
tifying new hits, enhancing molecule binding affin-
ity in hit-to-lead, and reducing toxicities in lead
optimization. Drug metabolism is a major consid-
eration for modifying drug clearance and also a
primary source for drug metabolite-induced toxic-
ity. With major cytochrome P450 structures identi-
fied and characterized recently, structure-based
drug metabolism prediction becomes increasingly
attractive. In silico methods based on molecular
and quantum mechanics such as docking, molecu-
lar dynamics and ab initio chemical reactivity cal-
culations bring us closer to understand drug
metabolism and predict drug–drug interactions. In
this study, we review important progress in drug
metabolism and common in silico techniques
adopted to predict drug regioselectivity, stereose-
lectivity, reactive metabolites, induction, inhibi-
tion and mechanism-based inactivation, as well as
their implementation in hit-to-lead drug discovery.
Key words: cytochrome P450, drug design, drug metabolism,
structure-based
The goal of drug discovery is to find best medicines to prevent,
treat and cure diseases quickly and efficiently. To fulfill this goal,
computational tools have helped medicinal chemists modify and
optimize molecules to potent drug candidates, have led biologists
and pharmacologists to explore new disease genes and novel drug
targets, and have been also guiding drug metabolism scientists to
achieve better pharmacokinetic profiles and avoid drug toxicities.
These in silico approaches have been widely applied to predict drug
absorption, distribution, metabolism, excretion and toxicity (ADMET).
The optimization of chemical space in early discovery stage using
these in silico tools will shorten the total drug discovery cycle time
and at the same time enhance the late-stage drug survival rate.
Lipinski's 'Rule-of-Five' is a rule of thumb to predict small-molecule
Editor’s invited manuscript to celebrate the 4th Anniversary of Chemical Biology & Drug Design.
doi: 10.1111/j.1747-0285.2009.00899.x
oral drug bioavailability, which initiated the era of data mining and
computer-aided drug design in pharmaceutical industry (1). Another
virtual 'prediction' example is the 'structure alert chart' summarized
by medicinal chemists for drug design purpose. To develop compre-
hensive drug metabolism prediction methods will offer a powerful
means to identify and analyze potential pharmacokinetic and toxico-
logical problems.
Metabolism is a major consideration for modifying drug clearance
and a primary source for drug metabolite-induced drug toxicity. An
appropriate pharmacokinetic profile such as a reasonable half-life is
mainly controlled by drug metabolism, which should be adapted to
the desired purpose and is extremely important for drug develop-
ment. Drug-metabolizing enzymes catalyze phase I (such as hydroly-
sis, reduction and oxidation) and phase II (such as glucuronidation,
sulfation, acetylation, methylation and glutathione conjugation) reac-
tions (2). The oxidation reactions in phase I often cause toxicities
and ⁄ or drug–drug interactions. Cytochrome P450s are major enzymes
to catalyze the oxidative bioactivation, and other enzymes are flavin
monooxygenase, aromatase, monoamine oxidase, and alcohol ⁄ alde-
hyde dehydrogenase. To elucidate the underlying oxidation mecha-
nisms will help improving pharmacokinetic and drug safety profiles.
To date, 57 human P450 isoforms are known (3). They catalyze the
metabolic activation of xenobiotics ⁄ drugs, sterols, fatty acids, eico-
sanoids and vitamins. Most of them are expressed in human livers
including CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 3A4
and 3A5, and others are in human lungs such as CYP1A1, 1B1, 2A6,
2B6, 2E1, 2F1, 2J2, 2S1, 3A5 and 4B1 or in minor sites including kid-
ney, brain, small intestine, peripheral blood cells, platelets, neutroph-
ils, and seminal vesicles (3). CYP1A2, 2C9, 2C19, 2D6, 2E1, and
3A4 ⁄ 5 are major ones generally used for in vitro drug metabolism
profiling studies in current discovery settings. They contribute to a
combined metabolism of over 90% of all drugs in the market, and
their individual contribution to the total hepatic P450 expression is
approximate 15%, 20%, 5%, 5%, 10%, and 30%, respectively (3).
Over the past 30 years, researchers have attempted to pinpoint the
nature and mechanism of P450-catalyzed drug metabolism by com-
paring substrate selectivity, identifying reaction intermediates, char-
acterizing products, and most recently studying enzyme structures in
a molecular level. The most relevant and accepted catalysis mecha-
nism includes several major steps. In the catalytic cycle, it starts
with the binding of the substrate to the active site of P450s, then
reduction of the ferric heme iron to its ferrous state by co-enzyme
3
Sun and Scott
and reductase, then binding of molecular oxygen followed by a sec-
ond one-electron transfer and two-step protonation to form the
iron-oxo intermediate (compound I), and then electron transfer for
catalysis (3). It is difficult to simulate this whole metabolic process
by computers accurately, but with the availability of human P450
structures gradually, structure-based modeling has been developed
to facilitate drug design.
In silico drug metabolism prediction methods were ligand-based such
as building pharmacophore and QSAR quantitative structure–activity
relationship (QSAR) modeling before structure-based drug design
emerges (4). QSAR modeling still plays a big role in pharmaceutical
industry because of the significant growth of high-throughput screen-
ing data. In QSAR, neural networks, classification methods such as
recursive partitioning, decision trees, and support vector machines
are constructed with topological, structural and electronic descrip-
tors. ADMET properties are thus predicted on the basis of these de-
scriptors such as lipophilicity and solubility. Usually QSAR predictions
work particularly well with structurally similar compounds whereas
molecules from other regions of the chemical space can cause out-
liers. Furthermore, QSAR is limited by the quality of the available
experimental data. QSAR can sometimes provide hints about the
active site if according descriptors appear in the regression equation
such as steric constraints and lipophilicity. Pharmacophore models
are constructed to overlay structures of all ligands also to simulate
the spatial and chemical properties of the binding site (5). Pharmaco-
phore models, structure-based docking and molecular dynamics are
all mechanism-based approaches. Recently, the prediction power has
been improved in building 3D-QSAR pharmacophore models with
spatial atomic descriptors in consideration. These descriptors include
molecular interaction fields, electronic properties, and shapes of
active sites. The purpose of in silico ADMET prediction is that even-
tually we should be able to predict drug absorption based on the
combination of lipophilicity, solubility, permeability, metabolism, and
transporter activity; to predict drug distribution based on the volume
of distribution, plasma-protein binding and blood–brain barrier pene-
tration; and to predict clearance based on renal clearance, hepatic
metabolism, biliary excretion, and gut stability.
Structure-based methods have been an integral part of drug design.
These mechanistic approaches are encroaching on drug metabolism
studies such as to predict the binding modes of substrates, conforma-
tion change of enzymes, catalysis, and their consequences on physio-
logical system. For example, docking method and local binding energy
calculation were used to evaluate the relationship between the
metabolism and carcinogenicity (6). Indeed, structure-based modeling
by combining molecular and quantum mechanics enables us to predict
substrate affinity, lability and metabolic pathways. In practice, mass
spectrometry, nuclear magnetic resonance spectroscopy, proteomics,
metabolomics, and other advanced bioanalytical techniques have
been improved significantly, which can help us validate these in silico
predictions quickly. There are still challenges using these tools such
as the difficulties in calculating free energies in the thermodynamic
cycles, enthalpy and entropy differences and solvation issues (7), but
they already saved us time and recourses discovering new drugs fas-
ter and more efficiently, and thus provide us more opportunities in
drug discovery. We believe, with breakthroughs, in silico methods will
play an increasingly important role in drug discovery.
4 Chem Biol Drug Des 2010; 75: 3–17
To simplify the complexity of drug metabolism prediction, a hybrid
method of combining molecular and quantum mechanics has been
proposed, which is designed to predict the substrate binding and
rate-limiting steps of catalysis with cytochrome P450s. The primary
goal of this review is to present these computational approaches
on drug metabolism prediction. We first review drug-induced toxici-
ties and mechanisms, then specific prediction examples for regiose-
lectivity, stereoselectivity, inhibition, induction, and mechanism-
based inhibition, as well as the application in hit-to-lead drug
discovery.
Drug Metabolite-induced Toxicities
The attrition rate of drug candidates as a result of toxicities remains
high. Toxicity is still a major safety concern for drug withdrawal
(such as drugs suprofen, terfenadine, rofecoxib, mibefradil, troglitaz-
one, nefazodone, cisapride, remoxipride, tolcapone, and tienilic acid),
the 'black box warning' (such as the most recent GlaxoSmithKline's
anti-diabetic drug rosiglitazone), and the discontinuation of clinical
trials (such as Pfizer's hypercholesterolemia drug torcetrapib with-
drawal from Phase III) (7,8). An analysis of the first-in-human regis-
tration for ten big pharma companies demonstrated only a 10%
total success rate leading to the final FDA approval (8). The failure
rate becomes even higher when all drug candidates in preclinical
research are included in the statistics. In this regard, it becomes
important to define strategies for drug safety assessment. The tradi-
tional drug safety testing approaches include in vivo animal models
and in vitro cell-based assays, and most recently in silico assess-
ment is also introduced. QSAR-based expert systems are mainly
used in early drug discovery to predict toxicological endpoints includ-
ing carcinogenicity, teratogenicity, mutagenicity, immunotoxicity, neu-
rotoxicity, developmental toxicity, respiratory sensitization, and skin
irritation (9), but the quality of experimental datasets (10) is still a
big challenge to develop a predictive model.
Drug metabolism continues to be a prominent contributor in drug
safety (11). To understand the molecular and cellular mechanisms
responsible for drug- and metabolite-induced toxicity will be critical
to reduce the attrition rate. In-target and off-target toxicities are
two different mechanisms (12), in detail, the toxicity can be catego-
rized into 4 types (13): type-1 is related to pharmacological path-
ways either on primary target or non-primary target; type-2 refers
to the idiosyncratic toxicities such as drug-induced allergies; type-3
is due to chemical activation such as drug metabolism and the
metabolites adducting with macromolecules; and type-4 is the
chronic toxicity such as carcinogenesis and teratogenesis. Mitochon-
drial dysfunction is also a mechanism. It is very possible that the
withdrawal anti-diabetic drug troglitazone and cholesterol lowering
drug cerivastatin caused mitochondrial dysfunction (14). Long-term
mechanisms may come from the impaired mitochondrial replication
and protein synthesis, and short-term from the inactivation of the
electron transport complexes in the inner membrane of mitochon-
dria through the covalent binding with reactive metabolites (15).
Although in silico predictions of drug metabolism seem unlikely to
replace the well-established in vitro and in vivo methods in the
near future, these computational tools help early drug design

greatly. One potential toxicological factor the can be eliminated in
early stage is the mechanism-based toxicity. Xenobiotics may be
directly toxic such as nicotine, whereas the toxicity of drugs is lar-
gely due to their metabolites, either highly reactive electrophiles or
free radicals. Various mechanisms were proposed such as reactive
metabolite formation and mechanism-based inactivation. Primarily,
metabolic activation process depends on physicochemical properties
of compounds, the interactions between compounds and active site
amino acid residues of drug-metabolizing enzymes, and catalysis. In
addition, transporters such as P-glycoprotein or organic ion-trans-
porting polypeptides can transport drug metabolites to the off-site
target for toxicity. Moreover, drug toxicity can be idiosyncratic or
caused by genetic diversity. Polymorphisms in P450, N-acetyltrans-
ferase and transporter genes impact individual responses to drugs
because of the population genetic difference of these enzymes. The
ultimate objective of drug metabolism prediction is to evaluate the
toxicity, but currently it focuses on the selection and optimization of
drug candidates for the advancement of drug discovery pipelines,
also with the goal of reducing recourses and recreating more reli-
able candidates (11).
One of the key roles of preclinical drug metabolism assessment is
to identify potential liabilities in new chemical series as early as
possible. For example, the identification of reactive metabolites
offers promise for decreasing the high rate of attrition, because
reactive metabolites may bind covalently to the locus of their for-
mation or at a distant site to inactivate target proteins, change the
biochemistry, signal transduction, or even trigger immunological
response. Various forms of reactive metabolites are identified and
used for the in vitro reactive metabolite screening. These moieties
include quinones, quinone methides, quinone imines, imine met-
hides, epoxides, anilines and derivatives, furans, benzylamines, thi-
ophenes, glitazones, thioureas, alkenes, and alkynes (16). However,
some reactive metabolites still escape from the detection even with
the most advanced bioanalytical instruments, and also there is a
gap between reactive metabolite formation and its toxicological
consequences (15). A solution especially in design is to incorporate
in silico tools as reviewed here.
Several P450-related pharmacological processes can cause drug–
drug interactions including induction, inhibition, and mechanism-
based inactivation. Induction refers to a process that causes the
increase of P450 enzyme expression, an adaptive response to xeno-
biotics or certain drugs by human body, which is mediated by
nuclear hormone receptors such as constitutive androstane receptor
(CAR), pregnane X-receptor (PXR), peroxisome proliferator activated
receptor (PPAR) and glucocorticoid receptor (GR). Induction-caused
drug–drug interactions can reduce the efficacy and ⁄ or increase the
toxicity of co-administrated drugs (17). Inhibition occurs during sub-
strate or molecular oxygen binding and mechanistically, P450 inhibi-
tors are categorized as reversible or irreversible ones that are
measured by analyzing the inhibition of typical substrates of P450s
by inhibitors such as using Michaelis–Menten kinetics to determine
a reversible inhibition (18). Mechanism-based inactivation is also
referred as 'suicide' or 'time-dependent' inactivation. It is a process
that reactive intermediates inactivate the enzyme without leaving
the active site of P450s (19). Mechanism-based inactivators can
either covalently bind to the amino acid residues of apoprotein
Chem Biol Drug Des 2010; 75: 3–17
Structure-based Drug Metabolism Predictions
or destruct the prosthetic heme group of P450s. Some
known mechanism-based inactivators are acetylenes (such as
2-ethynylnaphatalene, 9-ethynylphenathrene, 7-ethynylcoumarin and
5-phenyl-1-pentyne), organosulfur compounds (such as disulfiram,
cimetidine, tienilic acid, ticlopidine, and thiazolidinediones), arylam-
ines (such as 1-aminobenzotriazole), cyclic tertiary amines (such as
phencyclidine), and furanocoumarins (such as bergamottin, 8-gera-
nyloxypsoralen, and 8-methoxypsoralen). Others include tamoxifen
and raloxifene, both of which are the selective estrogen receptor
modulators, glabridin (an isoflavan) and anticancer drug N,N ¢,N ¢¢-
triethylenethiophosphoramide (19). In addition, the pneumotoxin
3-methylindole was found to be bioactivated by CYP2F1 in human
lung to form electrophilic methylene imine reactive intermediates
that cause mechanism-based inactivation. Studies showed that 3-
methylindole structural analogues are also potential mechanism-
based inactivators (Figure 1). For example, zafirlukast (a leukotriene
receptor antagonist), MK-0524 (a prostaglandin D2 receptor antago-
nist), and SPD-304 (a tumor necrosis factor-a inhibitor) were found
to form methylene imine reactive intermediates, and the mecha-
nism-based inactivation of CYP3A4 was observed in zafirlukast,
SPD-304 and tadalafil.
Drug Metabolism Prediction: insights
from Docking, Molecular Dynamics and
Quantum Chemical Calculations
The success of structure-based drug design is well known, by which
approximately 50 compounds have been introduced to the clinical
trials and ⁄ or approved until 2004 (20). To date, many more drug
candidates have been discovered with the help of advanced compu-
tational techniques for the reason that in silico tools have been
improved considerably and more widely used by pharmaceutical
companies in the last several years. For example, using AutoDock
combined with relaxed complex method has led to the approval of
the first HIV integrase inhibitor raltegravir (Isentress) by Merck in
2007 (21). Furthermore, the advantage of structure-based drug
metabolism prediction not only predicts the site of metabolism
(metabolic liability) but also elucidates important molecular interac-
tions between substrates and active site residues of drug-metabo-
lizing enzymes, which produce invaluable information for chemical
design and redesign to enhance pharmacokinetic profiles and
reduce toxicities.
To perform structure-based drug metabolism prediction, experimen-
tal determined (such as using X-ray crystallography) structures of
P450s or other drug-metabolizing enzymes are required. If no corre-
sponding X-ray crystal structures are available, appropriate homol-
ogy models are required, but need to be carefully evaluated during
the construction process based on enzyme kinetics data and ⁄ or
site-directed mutagenesis results. The first mammalian P450 struc-
ture, rabbit microsomal CYP2C5 was determined in 2003 by Dr. Eric
Johnson's group in the Scripps Research Institute (22) and subse-
quently, the crystal structures of major human forms become avail-
able including CYP1A2, 2A6, 2A13, 2C8, 2C9, 2D6, 2E1, and 3A4
(Table 1). These structures yielded invaluable molecular information
of P450 enzymes, and also revealed the coming of fast crystalliza-
tion of drug-metabolizing enzymes.
5
Sun and Scott

O
O HN S
P450 O
O
H
N N O N
H H
3-methylindole 3-methyleneindolenine O
N

Zafirlukast (Accolate)

O O
N N
N
F O

O N
OH O
N
N
O S O N
H
Cl F F O Figure 1: 3-Methylindole and
O its structural analogues are poten-
F
MK-0524 tial mechanism-based inactivators
SPD-304 Tadalafil (Cialis) of cytochrome P450s (86-91).

Table 1: Available structures of human cytochrome P450s for public access, as determined by X-ray crystallography (37,77–85)

CYPs PDB CODE Resolution () Notes

CYP1A2 2HI4 1.95 Bound with Alpha-Naphthoflavone

CYP2A6 1Z10, 1Z11 1.90, 2.05 Bound with Coumarin and Methoxsalen
2FDU, 2FDV, 2FDW, 2FDY 1.65 – 2.05 Bound with N,N-Dimethyl(5-(pyridin-3-yl)furan-2-yl)methanamine,
N-Methyl(5-(pyridin-3-yl)furan-2-yl)methanamine, (5-(Pyridin-3- yl)furan-2-yl)methanamine,
and Adrithiol
2PG6, 2PG7, 2PG5 1.95 – 2.80 2A6 Mutants L240C ⁄ N297Q, N297Q ⁄ I300V and N297Q
CYP2A13 2P85 2.35 Bound with Indole
CYP2C8 1PQ2 2.70 No Substrate Bound
2NNI, 2NNJ, 2NNH 2.28 – 2.80 Bound with Montelukast, Felodipine, and 9-Cis-Retinoic Acid
CYP2C9 1OG2, 1OG5 2.55, 2.60 Bound with Warfarin
1R9O 2.00 Bound with Flurbiprofen
CYP2D6 2F9Q 3.00 No Substrate Bound
CYP2E1 3E4E, 3E6I 2.60, 2.20 Bound with 4-Methylpyrazole and Indazole
CYP3A4 1TQN 2.05 No Substrate Bound
1W0E, 1W0F, 1W0G 2.65 – 2.80 No Substrate Bound, and Bound with Metyrapone and Progesterone
2J0D, 2V0M 2.75, 3.80 Bound with Erythromycin and Ketoconazole

Modeling drug metabolism requires predicting substrate binding to
the active sites and adapting computational approaches to account
for the catalysis. There is no single fully integrated structure-based
computational tool that is able to simulate all the possible pro-
cesses of drug metabolism. However, a combined computational
strategy was designed to predict the regioselectivity of macrolide
immunosuppressants sirolimus and everolimus: their hydroxylation
and O-dealkylation reactions catalyzed by CYP3A4 (23). In address-
ing the molecular mechanisms of sirolimus and everolimus bioacti-
vation, the proposed prediction method consisted of three separate
steps. First, the substrates were docked into the active site of
CYP3A4 with energetically favored binding poses selected. Second,
the chosen docking poses were placed into AMBER force field for
molecular dynamic simulation with the substrate binding orientation
and the distance to the heme iron calculated, respectively. Third,
the hydrogen atom abstraction of selected fragments of both sub-
strates was calculated by quantum chemistry method (Figure 2).
This prediction strategy has been gradually accepted and adopted
by other computational chemists and drug metabolism scientists to
predict drug metabolism.
One of the goals of drug metabolism prediction is to find the low-
est-energy binding structure complex containing drug-metabolizing
enzyme and substrate, which in many cases may account for one of
the most possible orientations for catalysis. Other goals include the
prediction of the rates of metabolism based on drug molecule
activation mechanisms. The success of prediction requires the
development of an accurate energy potential function in the above-
mentioned sequential calculations. Simplification of the process is
invariably necessary in considering the computing time; however, a
single one-step modeling is not adequate from a mechanistic view,
but sometimes still produces relative or comparable energy informa-
tion that is sufficient for drug design purpose especially for the
compounds within the same chemical series. For example, docking
was successfully used to predict CYP2C9 metabolic activation of
the COX-2 inhibitor celecoxib and its 13 analogues (26) Figure 3

6 Chem Biol Drug Des 2010; 75: 3–17

 Structure-based Drug Metabolism Predictions

Figure 2: Energy change from

substrate binding to product forma-
tion in cytochrome P450-catalyzed
drug metabolism. A combination of
docking, molecular dynamics and
quantum chemical calculation is
proposed for in silico prediction of
drug metabolism.

demonstrated the predicted two binding pose clusters of midazolam
with docking method applied alone, and Figure 4 demonstrated the
quantum chemistry-based calculation for the activation energy of
testosterone carbon hydrogens indicating the 6-b position is the
most likely CYP bioactivation place.
Docking method which involves the prediction of substrate confor-
mation and orientation within the active site of enzymes (27) is
composed of a tandem prediction process: posing and ranking. Pos-
ing searches energetically favored binding orientations of substrates
that is often determined by a certain computational algorithm. On
the contrary, ranking scores various poses derived from posing with
different interaction energy calculation functions such as electro-
static, hydrogen bonding, van der Waals, entropy, and explicit solva-
tion (24). Searching protocols may be systematic, stochastic or a
simulation. A systematic searching usually incrementally grows the
substrate within the active site of CYPs, in other words, rigid cores
of substrates are docked in first and then the flexible parts, based
on libraries of pre-generated conformations. They are less challeng-
ing comparing to stochastic methods, which apply random changes
of substrates and then evaluate a predefined probability function.
Monte-Carlo and genetic algorithms are classic stochastic algo-
rithms. Simulation method like molecular dynamics accommodates
ligands in local minima of the energy surface that will be discussed
later.
A scoring function is required to rank the favored ligand conforma-
tions, such as using force field-, empirical-, or knowledge-based
methodologies. Fundamentally, these scoring functions make various
assumptions; therefore, the evaluation process is simplified and can
only roughly simulate the biological process. Researchers have tried
to eliminate assumptions and combine as many as energy parame-
ters to make it more reliable. In force field-based scoring functions,
energetic terms are derived from classic mechanics such as
AMBER95 or CHARMm22, and the atomic interactions between pro-
tein and ligands are simulated based on known factors controlling
the molecular recognition. Both enzyme–substrate interaction energy
and substrate internal energy are calculated, but the internal pro-

A B

Figure 3: Two major energeti-

cally favored binding clusters of
midazolam in the active site of
CYP3A4 from molecular docking
prediction (A) 1¢-hydroxylation Helix l Helix I
position (B) 4-hydoxylation position.
CYP3A4 is shown in a cartoon
format (transparency gray), and
midazolam (yellow) and heme (pink)
are shown with colored sticks: Heme Heme
nitrogen, blue; oxygen, red;
chlorine, green; fluorine, cyan.

Chem Biol Drug Des 2010; 75: 3–17 7

Sun and Scott
A B
Figure 4: Dynamic analysis of active site residues of CYP3A4 crystal structures (A) active site of CYP3A4 (B) overlapped selected active
site residues from various crystal structures with different substrates bound.
tein energy are generally excluded. For example, the van der Waals
energy is given by a Lennard–Jones potential function and electro-
static energy is calculated using Coulombic formulation with a dis-
tance-dependent dielectric function. Empirical scoring functions
depend on the molecular datasets used for regression analysis and
fitting that are obtained from experimentally determined binding
energies. It is usually simpler than force field-based scoring func-
tion but does not provide mechanistic information for the purpose
of molecular modification. Knowledge-based scoring is designed to
reproduce experimental structures rather than binding energies. The
advantage is its computational simplicity, but the function is only
based on limited sets of protein–ligand complex structures and also
limited by the atom-types being used to derive the corresponding
potential of mean force.
Our perception of the dynamic change of P450 active site grows
with the expanding X-ray crystal structures, and also from site-
directed mutagenesis and molecular dynamic analysis. Molecular
dynamics simulate the flexibility of CYP active site residues in a
time scale, generally in the order of nanoseconds. Depending on
the size of the binding pocket, multiple binding poses are possible
to give rise to different metabolic products. This might help explain
why regioselectivity or stereoselectivity often occurs for P450 sub-
strates. With molecular dynamics combined, docking methods can
also be enhanced. Although our understanding of P450-catalyzed
drug metabolism and our ability to model the process are expand-
ing, the prediction of active site residue dynamic change in a time
scale remains a challenge. The stability of the substrate-P450 com-
plex is likely the result of attractive van de Waals interactions, sol-
vation free energy, hydrogen bonds as well as hydrophobic and
polar interactions. In light of these interactions, molecular dynamic
approaches are often applied to identify specific residues that direct
8 Chem Biol Drug Des 2010; 75: 3–17
substrate binding. Ito et al. simulated the motion of active site resi-
dues of CYP2D6 in the process of propranolol binding, and found a
significant movement of F-G loop and H-I loop, with several resi-
dues identified as key sites such as Phe120, Glu216, Asp301,
Phe483, Phe219, and Glu222 (25). Their theoretical analysis was
indeed correlated with site-directed mutagenesis results. In another
study, molecular dynamic analysis was placed in-line with neural
network modeling to predict the binding of 82 compounds to
CYP2D6, which demonstrated well-defined predictive behavior (28).
Another molecular dynamic analysis with spectroscopic study
showed the large-scale structural transition of CYP2B4 when the
substrate 4-(4-chlorophenyl)imidazole binds. Interestingly, Muralidha-
ra and coworkers proposed a theory of 'open from' and 'closed
form' of P450 enzymes (29), which states that the structural varia-
tions are mainly located in the B' helix and F-G region.
A continuous time-course structural determination would be an
ideal method to visualize the dynamic change of P450 active site
residues, but still, not possible in the current experimental settings.
However, the co-crystallization with different substrate bound has
created a small database illustrating the dynamic change of P450
enzymes especially the dynamic orientations of side chains. Over-
lapping several CYP3A4 structures, we analyzed the dynamic
change of active site residues. Primary interests have centered on
different substrate binding, which showed some residues are extre-
mely flexible such as the obvious movement of Arg212, Ser312,
Phe304, and Glu308 (Figure 5). The interplay between these resi-
dues makes it difficult, if not impossible, to fully grasp the protein
dynamics when individual substrate binds. But the information is
important for us to determine docking parameters such as to set
part of protein flexible and the rest in rigid to increase the predic-
tion accuracy and reduce computing time.
 Structure-based Drug Metabolism Predictions

OH ation and epoxidation reaction pathway ratios and they

(i) concluded both electrostatic and hydrogen bonding interactions
(j) are important for the difference, especially for propene, the
(h) penta-radical and ⁄ or tri-radical determine the mechanisms of
(k) (g)
hydrogenation and epoxidation. For compound I, the Fe–S bond
(m) length changes when the substrate binds, which can be calculated
(n) (l) by QM ⁄ MM methods, for example, it was found the Fe–S bond
(e) (f) of CYP3A4 compound I tends to elongate (34). Moreover, the
(o) parameters calculated by quantum mechanics were tested as
(d)
theoretical descriptors to build QSAR models such as molecular
orbital derived descriptors for flavone substrates binding to CYP1A
(c) (35).
O
(a) (b)
Water molecules often play an essential role in drug–enzyme
Position of H ab initio Position of H ab initio interactions: the displacement of water molecules from binding
Abstration (kcal/mol) Abstration (kcal/mol) site is one principal source of binding free energy (24). Water
molecules were found in the binding pockets of drug-metabolizing
a 116.23 i 107.33
enzymes, which can make favorable interactions with either
b 83.54 j 101.98
c 102.12 k 98.01
ligands or protein active site residues. Therefore, active-site water
d 95.52 l 94.62 molecules are often included in computational methods as one of
e 95.05 m 106.31 the strategies to predict drug metabolism. But current efforts in
f 102.90 n 102.49 the simulation of these water molecules mainly focus on the
g 103.05 o 95.85 molecular dynamics of the 'on' and 'off' states to unravel their
h 96.48 significant roles in substrate binding as well as their effects on
docking and virtual screening accuracy. In some docking protocols
Figure 5: Hydrogen atom abstraction of testosterone calculated water molecules are allowed to rotate around their three principal
by density functional theory. The DFT energy shows 6b position axes. With this consideration, a study on the prediction of 65
hydrogen is the most energetically favored for abstraction. CYP2D6 substrates binding and their sites of metabolism demon-
strated that water molecules are located in the energetically
favored locations. Applying this strategy, the accuracy of docking
Quantum chemical calculation is a major tool for predicting P450 algorithms was significantly enhanced (36). Thus, an improved
catalysis. From the calculated energy barrier value, we can tell drug metabolism prediction strategy could plausibly include water
the absolute or relative oxidation potential in xenobiotic bioactiva- molecules to the settings of simulation. Not all water molecules
tion. The ab initio calculations using Hartree–Fock or density func- are resolved in crystal structures of cytochrome P450s, but
tional theory are often used to calculate the reactivity of researchers have identified some of these structural water mole-
substrates. For aliphatic hydroxylation and N,O-dealkylation reac- cules recently (Figure 6). For example, an ordered water (wat733)
tions, the hydrogen atom abstraction to form a radical is the rate- was found in the active site of CYP1A2, which is located in an
limiting step for catalysis. However, the mechanism of aromatic appropriate distance range to form hydrogen bonds with the car-
hydroxylation is still unclear. The arene epoxides are generally bonyl group of the inhibitor a-naphthoflavone (2.92 ) and also
accepted intermediates for the next-step hydrogen migration from the carbonyl group of Gly316 (2.84 ) of helix I (83). Several
hydroxylation site to the adjacent carbon ('NIH shift') (3). Except water molecules were identified to be important factors for
the activation energy, the orientation of the aromatic ring is also CYP2C9 substrate binding: one (wat600) is located between the
important, which is orientated for either 'side-on' addition (ring substrate flurbiprofen and the heme iron at a distance of <3.0 
vertical to the heme porphyrin) or 'face-on' addition (ring parallel that accounts for the hydrogen bond with the backbone carbonyl
to the heme porphyrin). To ensure the active site environment group of Ala297, one (wat819) is located at the distortion region
parameters included to determine the orientation, heme-oxygen of helix I, and another one (wat842) is between flurbiprofen and
species and other interacting active site residues are needed to the turn region of the substrate recognition site-6 (85). In CYP3A4,
be incorporated into quantum chemical calculation. The hybrid a cluster of water molecules was also observed above the heme
quantum mechanical ⁄ molecular mechanical (QM ⁄ MM) method is porphyrin (37), which can be replaced totally or partially that
ideal to do the calculation (30). For example, a hybrid QM ⁄ MM depends on the size of substrates.
method was used to simulate the benzene hydroxylation by
CYP2C9 by Bathelt et al. and the results suggested a roughly Regioselectivity
equal 'side-on' and 'face-on' pathways, which posed a theoreti- Many drug metabolism prediction efforts have focused on the
cal challenge because early opinions differed because the 'face- prediction of P450 regioselectivity or metabolic pathways (38,39).
on' addition was considered as the dominant mechanism for The derived information is important for modifying metabolic lia-
p-stacking interactions (31). Quantum chemical calculations also bility in hit-to-lead. Camitro's metabolism models were widely
shed light on the regioselectivity studies of cyclohexene and pro- used in early years to predict enzyme-substrate binding affinities
pene. Cohen et al. (32) and Hirao et al. (33) studied the hydroxyl- and metabolic sites using empirical and quantum chemical

Chem Biol Drug Des 2010; 75: 3–17 9

Sun and Scott
A B
wat733
wat842
ANF G316 FBP
Helix I
Heme Heme
Figure 6: Essential water molecules in the active sites of cytochrome P450s. (A) CYP1A2 and inhibitor a-naphthoflavone (ANF) (B) CYP2C9
and substrate flurbiprofen FBP, and (C) CYP3A4. Water molecules are depicted as red spheres.
approaches (40). By definition, the regioselectivity of P450-cata-
lyzed reactions is the preference of one or several substrate
sites of metabolism over others. Zhang et al. investigated the
metabolism of the C3¢ substituents of a series of taxane ana-
logues by CYP2C8 and 3A4 (41) using docking approaches, and
found the binding of molecules were re-oriented from in favor of
3¢-p-hydroxylation to 4¢¢-hydroxylation reaction when replacing the
C3¢ aromatic ring (paclitaxel) with an aliphatic chain (cephalo-
mannine). In addition, several combined models were used to
predict regioselectivity. In one model, docking was combined with
an empirical activation energy model as well as reaction
enthalpy and ionization potential descriptors to predict the sub-
strate reactivity (42). The authors evaluated 72 well-known
CYP3A4 substrates and produced a 76% success rate. In another
model, docking and ligand-based in silico methods were applied
to predict metabolism sites of 70 known CYP2C9 substrates suc-
cessfully (43). These studies also indicated that the regioselectivi-
ty is probably controlled by one or several active site residues.
For example, Lafite et al. investigated the hydroxylation of terfen-
adine derivatives by CYP2J2 (44) and found the presence of
bulky residues Ile127, Ala311, Ile375, Ile376, and Val380 above
the heme iron to form a hydrophobic core, and a hydrogen bond
formed between the keto groups of terfenadine derivatives and
Arg117. These results demonstrated that the control of substrate
accessibility can direct the regioselectivity, although improved cal-
culations are needed to further test the hypothesis. In another
study, the bioactivation of tobacco procarcinogen 4-(methylnitrosa-
mino)-1-(3-pyridyl)-1-butanone (NNK) by CYP2A6 and 2A13 was
compared by docking approaches. It was found that both Met365
and Ile366 of CYP2A13 provided more space for NNK positioning
in the active site for a-methyl hydroxylation and a-methylene
hydroxylation reactions but not in CYP2A6 (84), which explains
the regioselectivity difference of these two enzymes. The regiose-
lectivity was also observed using enzyme mutants. Keizers et al.
found that the O-demethylation of 3,4-methylenedioxy-N-alkylam-
phetamines was dominant in wild type CYP2D6 but N-demethyla-
tion and N-hydroxylation reactions occurred instead in its F120A
mutants (45).
10
C
Helix I R212
wat819
A297 Helix I
wat600
Heme
Stereoselectivity
Structure-based drug metabolism prediction is also a good option to
elucidate the metabolic differences in enantiomers. Indeed, many
drugs are synthesized for one enantiomer only to avoid the stere-
oselectivity by drug-metabolizing enzymes (Figure 7). The difference
of metabolic profiles of phenylahistin enantiomers was evaluated
using the energy calculation of docked enantiomers by molecular
mechanics and Poisson–Boltzmann surface area approaches. It dem-
onstrated that (-)-phenylahistin is 1.5–8 times less metabolized than
(+)-phenylahistin by CYP3A4 (46). Docking of (+)-phenylahistin in the
active site of CYP3A4 showed that both isoprenyl group and nitro-
gen atom at the imidazolyl ring can interact with the heme iron for
the hydroxylation reaction; however, for (-)-phenylahistin, the predic-
tion results differed dramatically (isoprenyl group is far from the
heme and the lone pair of the imidazolyl nitrogen does not point
toward the heme iron), because they possess very similar physico-
chemical properties. The difference of enantiomer metabolites from
non-steric compounds can also cause enantioselectivity. Bikadi and
Hazai studied CYP2C-catalyzed O-demethylation of methoxychlor
(47), and demonstrated that both CYP2C9 and 2C19 favored the for-
mation of S-mono-OH-methoxychlor, no enantioselectivity found for
CYP2C3, but CYP2C5 favored the formation of R-mono-OH-methoxy-
chlor. The chiral preference was further studied by molecular
dynamics, and the authors found it was caused by the variance and
flexibility of active site residues. In addition, deoxypodophyllotoxin
was found to be stereoselectively bioactivated by CYP3A4 to epip-
odophyllotoxin but not to podophyllotoxin (48). The docking and
Monte-Carlo-based refinement revealed that the epipodophyllotoxin
formation is in a more appropriate orientation, or specifically, the
hydroxylation is mediated by the hydrogen bonding between de-
oxypodophyllotoxin and active site residues Arg212, Glu374 and
Arg105.
Reactive metabolites
The mechanism of reactive metabolite formation by CYPs has
been extensively studied computationally, mainly by quantum
chemical calculations. With different substituents added to lead
Chem Biol Drug Des 2010; 75: 3–17
 Structure-based Drug Metabolism Predictions

O O

NH N NH N
NH NH
HN HN

O O

(–)-phenylahistin (+)-phenylahistin

H CCl3 H CCl3 H CCl3

(R) (S)

MeO OMe HO OMe MeO OH

Methoxychlor R-mono-OH-methoxychlor S-mono-OH-methoxychlor

H H HO H H OH
O O O
O O O
O O O

MeO OMe MeO OMe MeO OMe

OMe OMe OMe

Deoxypodophyllotoxin Podophyllotoxin Epipodophyllotoxin

Figure 7: Enantioselectivity and stereoselectivity of several known compounds.

compounds, the relative free energy change for activation is calcu-
lated to guide lead optimization. Docking and molecular dynamics
are often combined with quantum chemical calculations to
increase the prediction power. Many successful applications have
been documented. For example, the mechanism of phenacetin bio-
activation to form reactive metabolite quinone imine was studied
with ab initio energy and spin distribution calculation, to compare
the hydrogen atom abstraction at the a-methylene carbon atom
and electron abstraction from the nitrogen atom at the acetylami-
no side (49). A similar study on the metabolic activation of acet-
aminophen to quinone imine by Loew and Goldblum predicted the
hydrogen atom abstraction from the phenol group is more proba-
ble than from the amide nitrogen (50), which was also confirmed
by in vitro studies (51). In another study, the bioactivation of car-
bamazepine by CYP3A4 showed that the reactive metabolite10,11-
epoxide formation is more probable than the hydroxylation of the
aromatic ring calculated by density functional theory (52). With
these mechanistic results, we can design new compounds with
reduced oxidation potential and thus eliminate the reactive metab-
olite formation. For example, the bioactivation energy of the anti-
psychotic drug remoxipride to from a hydroquinone metabolite and
further to an electrophilic quinonone was calculated by hybrid
density functional theory (53). The authors then redesigned
remoxipride by adding amide and methoxy groups. In addition, the
ab initio calculation was used to compare the oxidation potential
of nefazodone and buspirone to form reactive intermediate
quinone imines (54). It demonstrated buspirone is much more diffi-
cult to undergo two-electron oxidation than nefazodone, and thus
less reactive metabolite is expected. From these drug design
examples, we know by calculating the oxidation potential of reac-
tive metabolite formation, we can modify molecules to enhance
drug metabolic profiles or reduce toxicity issues.
Inhibition
The mechanism of inhibition was studied by docking and molecular
dynamic approaches. Quinidine is a potent competitive inhibitor of
CYP2D6 but not a substrate. McLaughlin et al. investigated the
binding modes of quinidine together with site-directed mutagenesis
experiments (55). They found in CYP2D6's F120A and E216F
mutants, quinidine indeed became a substrate, indicating the impor-
tance of these two residues in controlling substrate ⁄ inhibitor
switching. Both substrates and inhibitors bind to enzymes in ener-
getically favored manner, but for inhibitors they are not in the
appropriate orientation for catalysis or bind to no-catalysis sites
such as the ligand entrance region of P450s. Structurally diverse
compounds were evaluated by several research groups for their
inhibition of CYP3A4 using docking methods and also considering
ligand conformation, orientation, stereoisomer, and protonation
state. The results demonstrated that docking approach is indeed a
good option to screen a library for potential inhibition and drug-
drug interactions (56,57). In addition, co-operativity properties that
may determine the inhibition showed the homotropic and ⁄ or hetero-
tropic co-operativity of 1-alkoxy-4-nitrobenzenes and 1,4-phenylene
diisocyanide in CYP1A2 (58). These calculations were used to
design new compounds with modified inhibition. For example, sub-
stitution of terfenadine benzylic alcohol group to ketone demon-
strated 10-fold inhibition increase with CYP2J2 (59).

Chem Biol Drug Des 2010; 75: 3–17 11

Sun and Scott
Figure 8: Structure of pregnane X receptor. The monomers are
in yellow and green, ligands in hot pink, and steroid receptor coac-
tivator 1 (SRC-1) in red on the back.
Induction
CYP3A4 is induced by the activation of PXR, and its structure has
been determined recently (60). Docking showed the addition of
polar groups to one end of the activator attenuated the activation
because the new molecules destabilize the hydrophobic interactions
in the binding pocket (Figure 8) (61). With more structures of key
induction regulators determined, we expect to predict the induction
using the similar in silico tools above.
Mechanism-based inactivation
Heme porphyrin ring and some active site residues are primary tar-
gets for the binding of reactive metabolites that can consequently
cause the mechanism-based inactivation before their release from
P450 active site (19). Recent structural modeling and proteomic
studies have shed light on possible inactivation mechanisms for
either heme porphyrin or apoprotein residue alkylation with drug
molecules. Raloxifene is a mechanism-based inhibitor of CYP3A4,
which is bioactivated to either diquinone methide or epoxide reac-
tive metabolites. Several research teams have identified these
adduction sites such as Cys239 and Tyr75 for quenching reactive
intermediates (62–64), in particular, it was postulated that the
nucleophilic OH group of Tyr75 or the sulfur group of Cys239 can
adduct with reactive metabolite directly. In addition, CYP3A4 and
CYP3A5 were compared to elucidate the mechanism of raloxifene
bioactivation. It was found that the mechanism-based inactivation
was only present in CYP3A4 but not in CYP3A5 because Cys239 is
replaced by a serine residue in CYP3A5 (64). Kang et al. showed
that the formation of reactive 10,11-epoxide, 2,3-arene oxide and
iminoquinone metabolites of carbamazepine adducted with Cys239
for mechanism-based inactivation (65). The cysteine-specific modify-
ing reagent iodoacetamide was also applied to test the importance
of the cysteine residue, and the results showed that iodoacetamide
can prevent the mechanism-based inactivation (63), and thus indi-
cated Cys239 may be one of the cysteine to quench the reactive
metabolite. Moreover, the apoprotein alkylation by reactive metabo-
lites also depends on the structures of substrates. Docking showed
the furan ring of bergamottin is bioactivated to an epoxide that
inactivates CYP3A5 and CYP2B6. The furan site was observed as
the primary site for the exposure to the oxyferryl center of heme
(66). Ser360 was also found to be the adduction site by the reac-
tive metabolites of 17-a-ethynylestrodiol (67).
12
Application in hit-to-lead Drug Discovery
It is important to maintain both quality and quantity of early lead
series to discover a new therapeutic agent in a reasonable time
frame (68). In hit-to-lead drug discovery, high-throughput screening
requires a suitable assay to be developed; as attractive alternatives
fragment-based and docking methods are especially useful for
focused screening. The application of better hit-to-lead approaches
can help us achieve the goal of fast and efficient drug discovery.
Structure-based drug metabolism prediction has the potential to
offer new drug design approaches based on metabolic profiles for
hit-to-lead discovery, which should facilitate our endeavor on
expanding chemical space and searching high-quality drug candi-
dates. Drug metabolism prediction using docking, molecular dynamic
and quantum chemical methods provided a powerful insight into
the application of these in silico tools in drug discovery. We have
found great success in predicting drug metabolism using the similar
methods in our drug discovery projects. Indeed, we need a cost-
effective and high-efficient hit-to-lead process to expand the chemi-
cal space and find the best lead compounds with minimum
resources, so the in silico drug metabolism approaches described in
this review can be a great benefit for the optimization of chemical
space with the purpose of exploring best-in-class medicines
(Figure 9).
Substantial attention has been given to use docking-related molecu-
lar mechanical approaches to make molecules more potent. The
modification such as adding or changing functional groups primarily
focuses on the increase of ligand–protein interactions, which gener-
ally lowers binding energies or increases the potency. For examples,
a docking and scoring method was applied to explore potent b-sec-
retase inhibitors (69). More importantly, to develop a safer com-
pound means more chance to move a drug candidate into the
market. For example, the cyclooxygenase-2 inhibitor celecoxib is
less potent than rofecoxib, but much safer according to meta-analy-
sis postmarket studies, which was the main reason it is still in the
market but the rest of same-family drugs such as rofecoxib got
withdrawal for increased cardiovascular toxicity. As discussed
above, the current trend of hit-to-lead and lead optimization is the
early evaluation of ADMET properties such as the optimization of
clearance profiles and elimination of P450 inhibition (70). The trade-
off for improved ADMET properties is sometimes decrease the
potency, and vice versa. Therefore, to find the overlap region of AD-
MET and potency properties early trade-off is essential for reducing
chemical liabilities and accelerating hit-to-lead process. Lead com-
pounds can then be optimized based on the collected pharmacologi-
cal and ADMET properties. It was noted that the receptor binding
sites are difficult to define for many drug targets (71), but for P450
enzymes they are highly conserved in a fixed region. Considering
this, the success rate for drug metabolism modeling can be higher
than target protein simulation. Overall, these in silico approaches
are helpful for the identification of tighter or specific binders with
better ADMET properties (72).
The strategy of blocking the metabolic labile sites of compounds
was driven by attaching a metabolic stable group such as a fluorine
atom or a trifluoromethyl group. For example, the metabolic activa-
tion of a series of celecoxib structural analogues showed that
Chem Biol Drug Des 2010; 75: 3–17

Figure 9: Schematic represen-
tation of hit-to-lead and lead opti-
mization approaches to defining
chemical space based on both drug
receptor space and ADME space.
replacing the methyl group of 4-methylphenyl moiety with a trifluo-
romethyl group totally blocked the methyl hydroxylation by CYP2C9
(but COX-2 inhibition rate remains the same) (73), which indicate
that new compounds have improved metabolic properties without
losing the potency and selectivity. However, the merit of adding a
trifluoromethyl or fluorine group is unpredictable because new met-
abolic lability can be introduced by switching the metabolic path-
ways. Therefore, the blind practice of including them for every drug
design without considering substrate-protein interactions can block
our vision during the exploration of wider chemical space. Once the
structures of receptors or P450 enzymes are available, we can use
structure-based computational tools to expand the chemical space
and also achieve better pharmacokinetic profiles for safer drugs.
Therefore, a promising strategy to reduce attrition rate and
shorten discovery cycle is to harness these in silico tools to refine
ADMET properties in the early stage. Indeed, many recent drug
discovery efforts have focused on drug metabolism prediction
early. In the discovery of chemokine receptor CCR5 antagonists,
the original hits contained imidazopyridine moieties that are com-
mon CYP2D6 inhibitors (74). The authors redesigned the candi-
dates after modeling the binding of selected compounds in the
active site of CYP2D6, which demonstrated that the pyridine nitro-
gen atom is the closest atom to the heme iron. Based on the
interaction information, imidazopyridine moieties were replaced by
the carbon analogues benzimidazoles. Consequently, the drug-drug
interaction was avoided early, and thus a high quality lead was
discovered. The development of metabotropic glutamate receptor
antagonists tells a similar story. The pyrazolo[3,4-d]pyrimidin-4-one
series were found to be original hits as the potent and selective
antagonists but showed high clearance. The authors then
improved the ADMET properties by adding electron-withdrawing
groups to the molecule (75). In addition, rational approaches to
eliminating reactive metabolite formation are worthy of consider-
Chem Biol Drug Des 2010; 75: 3–17
Structure-based Drug Metabolism Predictions
ation. The trifluoromethylpyrimidine series were found to be pro-
line-rich tyrosine kinase-2 inhibitors but also positive in reactive
metabolite screening. Clearly, to remove the potential toxicities
was required for further development. The modification of the
carbonyl group of the 5-aminooxindole moiety to a stronger elec-
tron-withdrawing sulfone group was found to eliminate reactive
metabolite formation, which was predicted correctly by density
functional theory calculations (92). These successful hit-to-lead
efforts in drug metabolic profile improvement suggest that the
combination use of in silico tools at both receptor space and drug
metabolism space will be likely to turn the serendipity of drug
discovery into powerful drug design strategies.
Several limitations of these tools should be noticed. Crystal struc-
ture resolution, accuracy of the force field, docking algorithms and
quantum chemical calculation all determine the prediction power.
The quality of crystal structures are different because the flexible
regions of some proteins are difficult to be determined. A higher-
quality structure will generally mean higher prediction power. For
trend analysis purpose, the high-throughput docking methods have
been developed for fast and economic discovery process (71). To
avoid these limitations various strategies have been applied.
Scientists at Merck found that the virtual screening success rate
was enhanced by the combination of two docking methods (93). In
addition, free-energy perturbation calculation in the context of
Monte-Carlo statistical mechanic simulation was used concurrently
to optimize docked compounds. For example, modification of core
heterocyclic moieties of 2-anilinyl-5-benzyloxadiazole with free-
energy calculation led to potent non-nucleoside inhibitor of human
immunodeficiency virus reverse transcriptase (76). Moreover, de
novo ligand design such as fragment-based design and 'scaffold
hopping' are attractive approaches that ultimately allow more hit
series explored, and thus produce better chance of successful hit-
to-lead drug discovery.
13
Sun and Scott
In summary, recent drug metabolism prediction efforts reviewed
here highlighted the importance of computational methods for drug
design and hit-to-lead drug discovery. Given the rapid accumulation
of new structures of drug-metabolizing enzymes, it is increasingly
clear that computational tools to predicting drug metabolism will be
crucial in faster and more efficient drug discovery. To conclude, con-
siderable challenges remain for in silico drug metabolism prediction
using structure-based methods, but the opportunities for in silico
drug metabolism prediction to become a mainstream component of
drug design are now apparent, and the accurate prediction now
appears to be an achievable goal in the near future.
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