Article

pubs.acs.org/molecularpharmaceutics

The Contribution of Atom Accessibility to Site of Metabolism Models

for Cytochromes P450
Patrik Rydberg,* Michal Rostkowski, David E. Gloriam, and Lars Olsen
Department of Drug Design and Pharmacology, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen, Denmark
*
S Supporting Information

ABSTRACT: Three different types of atom accessibility

descriptors are investigated in relation to site of metabolism
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predictions. To enable the integration of local accessibility we

have constructed 2DSASA, a method for the calculation of the
Downloaded via BITS PILANI PILANI CAMPUS on October 4, 2019 at 05:52:36 (UTC).

atomic solvent accessible surface area that is independent of

3D coordinates. The method was implemented in the
SMARTCyp site of metabolism prediction models and
improved the results by up to 4 percentage points for nine
cytochrome P450 isoforms. The final models are made
available at http://www.farma.ku.dk/smartcyp.
KEYWORDS: cytochromes P450, solvent accessible surface area, site of metabolism, circular fingerprints

■ INTRODUCTION
Understanding the pharmacokinetic characteristics of drug
candidates is crucial both in the early drug discovery and
subsequent development processes. The metabolism and
elimination of drugs gives a major contribution to their kinetic
profile and is heavily influenced by interactions with the
cytochrome P450 (CYP) enzyme family. This family of
ubiquitous enzymes is the major determinant of phase I
metabolism. The nine most prevalent isoforms in human are
1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4, among
which 1A2, 2C9, 2C19, 2D6, and 3A4 are considered to be the
most important for drug metabolism.1
The prediction of the site of metabolism for CYP mediated
drug metabolism has received a lot of interest in recent years,
and models have been based on many different techniques.2
While attempts to construct models that include the protein
structure in the modeling have been made,3−5 they so far seem
to offer very little (if any) improvement in prediction accuracy
compared to the best ligand-based models.6−8 This is probably
because the CYP enzymes are very flexible,9 making the
sampling of the full conformational space too computationally
expensive.
We have developed the 2D structure based SMARTCyp
methodology,10−12 which in contrast to other models is
completely independent of experimental data. To achieve
this, the reactivity toward oxidation by the heme group of the
CYPs of all atoms is assigned using fragment matching toward a
fragment library for which the reactivities have been
precomputed using density functional theory (DFT) transition
state calculations.
These reactivities in combination with a simple atom
accessibility descriptor (the relative span) lead to a model that
is quite accurate for most CYP isoforms.8 This is because
reactivity is the most important factor in determining the site of
metabolism, and accessibility is less important for most
isoforms. This is also why it works less well when the binding
orientation of a molecule within the active site is a more
important determinant than reactivity. This is the case for the
CYP isoforms 2C9 and 2D6. To take this into account, we
therefore recently constructed simple pharmacophore correc-
tions that simply take the distance between each atom and the
pharmacophoric element into account (carboxylic acid and its
bioisosteres in CYP 2C9 and protonated amines in CYP
2D6).6,13
In principle there are only two contributions that a ligand-
based model for CYP-mediated site of metabolism should
include: the reactivity and descriptors that mimic the binding
mode to the active site. Descriptors that mimic the binding
mode can be separated into two classes. First are those that
describe overall accessibility of an atom, which is determined by
the orientation of the molecule inside the active site. Examples
of such descriptors are distances to pharmacophoric elements,
bond counts to the center of the molecule, etc. Second are the
descriptors that describe the local accessibility of an atom. Such
descriptors estimate how likely an atom is to be accessible for a
reaction to take place, if the binding mode suggests that this
atom will be close to the heme group.
In SMARTCyp, all descriptors that mimic the binding mode
belong to the first class. In the standard model the relative span
is used to describe the orientation. The relative span is defined
Special Issue: Predictive DMPK: In Silico ADME Predictions in
Drug Discovery
Received: September 12, 2012
Revised: December 13, 2012
Accepted: January 7, 2013
Published: January 22, 2013

© 2013 American Chemical Society 1216 dx.doi.org/10.1021/mp3005116 | Mol. Pharmaceutics 2013, 10, 1216−1223
Molecular Pharmaceutics
as the relative distance to the center of the molecule (see Figure
1). Its maximum and minimum values within a molecule are
Figure 1. Description of the relative span and Span2End descriptors
used in the different SMARTCyp models. The example of calculated
accessibility concerns the atom in the red circle. The green bonds
represent the largest number of bonds between this atom and any
other atom. The black bold bonds represent the largest number of
bonds between any two atoms in the molecule. Reprinted with
permission from ref 6. Copyright 2012 American Chemical Society.
independent of molecule size. In the CYP 2C9 and CYP 2D6
models the Span2End descriptor is used together with a
pharmacophore descriptor. The Span2End descriptor is the
distance to the end of the molecule, defined as the maximum
number of bonds between any two atoms in the molecule
minus the maximum number of bonds between the atom of
interest and any other atom in the molecule (see Figure 1).
This descriptor has a minimum value of zero, but its maximum
value is dependent on the size of the molecule. Both span
descriptors give preference to sites of metabolism that require
binding modes with a vertical orientation compared to sites of
metabolism that require binding modes with a horizontal
orientation (described in Figure 2). Additionally, in the CYP
Figure 2. Vertical (left) and horizontal (right) binding modes
exemplified with testosterone. The bold horizontal lines represent
the heme group at the bottom of the active site. The span descriptors
give preference to sites of metabolism that are close to the heme iron
in a vertical binding mode, as shown by the relative span values in the
figure.
2C9 and CYP 2D6 models there is a pharmacophore-based
distance descriptor, which counts the number of bonds from
the atom of interest to the pharmacophoric element
(protonated amines in CYP 2D6 and negatively charged
groups in CYP 2C9). This descriptor penalizes binding modes
with the pharmacophoric element close to the heme group.
However, a descriptor for the local accessibility is missing in all
SMARTCyp models. The atomic solvent accessible surface area
(SASA, illustrated in Figure 3) is the most commonly used local
accessibility descriptor and has previously been shown to give a
significant contribution to several methods for prediction of
CYP mediated drug metabolism.5,7,14,15
The atomic SASA was first defined by Lee and Richards16 as
“The area on the surface of a sphere of radius R, on each point
of which the center of a solvent molecule can be placed in
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Figure 3. A 2-dimensional example of how atomic SASA is defined.
The gray, blue, green, and red filled circles are atoms, and their
corresponding atomic SASA are shown as dashed arcs in the same
colors. R is the sum of the atom and solvent probe radii. The distance
between each atom and its atomic SASA is the radius of the solvent
probe, which is shown as a black circle. The SASA of atom B is very
small as it is only accessible from one direction in this 2D
representation.
contact with this atom without penetrating any other atoms of
the molecule.” The sphere radius, R, is the sum of the atomic
van der Waals and solvent molecule radii.
The atomic SASA has hitherto required 3D structures,
typically in one of two different ways: either numerically and
accurately using the Shrake−Rupley dot density method17 or
analytically and to a rather high accuracy using approximate
methods.18−22
Here, we describe the first method for calculation of atomic
SASA from 2D structures and include it in site of metabolism
prediction models for cytochromes P450. The atomic SASA for
non-hydrogen atoms are computed based on circular finger-
prints, which previously have been used successfully to build
models for prediction of toxicity,23,24 atomic properties such as
pKa,25 and site of metabolism.26,27
■ METHODS AND DATA SETS
Cytochrome P450 Data Sets. Nine data sets with site of
metabolism data for CYP mediated metabolism were taken
from the recent work by Zaretzki et al.8 and comprise the
following numbers of compounds for each isoform: 1A2 (271),
2A6 (105), 2B6 (151), 2C8 (142), 2C9 (226), 2C19 (218),
2D6 (270), 2E1 (145), and 3A4 (475). The CYP 3A4 data set
was divided into a training set (50 compounds) and a test set
(425 compounds) by selecting a diverse subset using MACCS
fingerprints and Tanimoto similarities in the MOE software.28
Calculation of Site of Metabolism Prediction Accu-
racies. The models were validated by using the top 1, top 2,
and top 3 standard prediction rates by the algorithm developed
by Zaretzki et al.7 This algorithm is used to ascertain that
results from different software are comparable even when some
software gives multiple atoms the same rank.
Solvent Accessible Surface Area (SASA) Data Sets. The
subset of druglike compounds available for purchase (subset 23,
“drugs-now”, version update 2010-11-03) was downloaded in
smiles format from the ZINC database.29 MACCS fingerprints,
with 64 bit precision, were calculated for all molecules using the
Schrö dinger canvasFPGen utility 30 and applied by the
canvasDBCS utility to generate a diverse compound subset.
The Soergel metric31 and sphere exclusion method were
applied with an exclusion distance of 0.3 (similarity of 0.7). For
each of these compounds, the lowest energy ionization and
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Molecular Pharmaceutics Article

tautomeric state at pH 7 was calculated with Epik.32,33

Subsequently, the most likely bioactive conformation was
determined using Confgen.34,35 The compound set was split
into training and test sets containing 7,000 and 20,849
compounds, respectively. Finally, hydrogen atoms were deleted,
and the two molecule sets were both split into four atom
subsets (Table 1) having 1−4 neighboring atoms.

Table 1. Description of the Data Sets in Terms of Number of

Atoms and Atomic SASA
1a 2a 3a 4a all
Training Sets
no. of atoms 29,692 70,451 42,936 3,784 146,863
SASA range 7−97 0−62 0−35 0−12 0−97
(Å2)
av SASA (Å2) 48.8 25.7 5.4 0.4 23.8
Test Sets
no. of atoms 84,188 191,005 113,888 11,309 400,390 Figure 4. The atom type counts for the atomic circular fingerprint of
SASA range 0−96 0−59 0−37 0−11 0−103 the chlorine atom in 1-chloro-3-methoxybenzene.
(Å2)
av SASA (Å2) 49.6 25.9 5.3 0.3 24.3
a
Number of neighboring atoms.
for the atomic SASA. 7-fold cross validation was used to
compute q2 values, and the auto fit function was used to
determine if the PLS components were significant. The two
3D SASA Calculation. The atomic SASA was computed atom types for neutral (N.3) and charged (N.4) sp3 nitrogen
using the Shrake−Rupley dot density method17 with a probe atoms were merged into one variable since knowledge about
radius of 1.4 Å (representing water36) and 16,000 sphere points the charge of pyramidal nitrogen atoms is usually not included
(this quantity of sphere points generates a grid fine enough to in topological methods. When selecting which variables to
make numerical errors negligible). Atom radii were taken from include in the models, we always looked at one level of the
Mantina et al.37 and complemented with van der Waals radii circular fingerprints at a time, and kept or removed all variables
from Bondi.38,39 Atoms that are topologically symmetric can of the same type in that level. The final PLS models were
have different SASA when using 3D coordinates. However, for defined as those having the highest explained variation and
prediction of accessibility we are only interested in the largest goodness of prediction (r2 and q2, respectively) and the lowest
SASA if there are topologically symmetric atoms. To define the complexity (smallest number of variables).
atoms that are topologically symmetric, we have used the Variable Importance Determination: Since the SMART-
algorithm by Hu and Xu.40 When describing local accessibility Cyp model is applied to site of metabolism data, which, in
for the purpose of CYP-mediated site of metabolism prediction principle, describes atom ranks within molecules, a traditional
we are, in principle, interested in describing the accessibility of determination of the variable importance in regression models
the oxygen atom that is bound to the heme iron atom, which is is not applicable. Instead we performed a variable exclusion
the atom that participate directly in the oxidation reaction, and evaluation to estimate the contribution of the parameters to the
not the full heme molecule itself. We assume that the atomic models. An exclusion evaluation is performed in the following
SASA calculated with a standard water probe radius (1.4 Å) is way: For each variable, the reduction of prediction accuracy
similar to the accessibility of this oxygen atom. when this variable is excluded from the scoring function
Atomic Circular Fingerprint Generation. The basic circular (without refitting the scoring function) is measured. The sum
fingerprints were generated using the algorithm of Xing et al.25 of the reductions for all variables is defined as being the total
without grouping of atom types. In our data sets of neighboring model accuracy, and the contribution of each variable is then its
atoms, there were in total 25 different atom types as described reduction divided by the total model accuracy. Thus, this gives
in Table SI1 in the Supporting Information. To these an estimate of the relative contributions of each variable to the
fingerprints we added atom counts and counts of rings of model for the data set in question. Below, the evaluation was
performed using the top 2 prediction accuracy.

sizes 3−8 or larger for each level (a level describes atoms
located a specific number of bonds from the atom of interest),
resulting in 33 variables at each level. Since we are counting
atom type, rings, and total atom count, each atom always
contributes to at least two variables at each level (atom type
and atom count), and can potentially contribute to three
different variables (if it is part of a ring). Rings were defined
using the minimum cycle bases algorithm41 as implemented in
the CDK.42,43 For each atom we generated circular fingerprints
in six levels, where level 0 is the atom of interest and levels 1−5
encompass the atoms separated from this atom by 1−5 bonds,
respectively, as shown in Figure 4 for atom type counts.
Construction of Partial Least-Squares (PLS) Models.
SIMCA-P (version 11)44 was used to create the PLS models45
1218
■
RESULTS AND DISCUSSION
Introduction of the Solvent Accessible Surface Area
for Site of Metabolism Prediction. We wanted to
investigate if adding the atomic solvent accessible surface area
(SASA) to the SMARTCyp score would improve the
prediction accuracy. As a test case, a small subset of the CYP
3A4 data set (50 compounds) was used to determine the
contribution of atomic SASA, which was then validated by the
remainder of the CYP 3A4 data set (425 compounds). The
atomic SASA was computed from the 3D structures of these
475 compounds. The contribution of atomic SASA to the
SMARTCyp scoring function was optimized to achieve the best
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Molecular Pharmaceutics Article

top 1, top 2, and top 3 prediction accuracies for the training set.
The optimal parameter for the training set was found to be
0.04, resulting in the SMARTCyp score equation shown (eq 1).
score = reactivity − 8 × relative span
− 0.04 × atomic SASA
The score determines the likelihood for metabolism, with a
low score suggesting an atom is more likely to be metabolized
than an atom with a high score. The reactivity is a measure of
the transition state energy for the oxidation reaction
(determined by fragment matching against precomputed
transition state energies). The relative span is defined as
shown in Figure 1, and the constant 8 was determined to reflect
the standard deviation of the original reactivity rules in
SMARTCyp, and has not been changed in this work.10
The improvement in prediction accuracy was found to be 2−
4.5 percentage points for the training set and 1.2−3.0
percentage points for the test set (see Table 2), suggesting
Table 2. SMARTCyp Prediction Accuracy on CYP 3A4 Data
with and without 3D Atomic SASA Contribution
training set test set
no SASA SASA no SASA
top 1%a
66.0 68.0 64.2
top 2%a 77.5 80.0 75.7
top 3%a 83.5 88.0 81.2
a the number of neighbors and the size of the atomic SASA
Top n accuracy is the percentage of compounds for which a site of
metabolism is found among the top n atoms.
that atomic SASA can make a valuable contribution to
SMARTCyp models. An example of how rankings can change
when including atomic SASA is shown in Figure 5. A variable
Figure 5. An example of improved site of metabolism prediction when
including atomic SASA. The arrow represents the site of metabolism,
and the numbers represent the top two ranked atoms (black with
atomic SASA and gray without).
exclusion evaluation on the test set shows that the reactivity is
ten times more important than the two accessibility descriptors,
contributing 84% of the model (relative span and atomic SASA
contribute 9% and 7%, respectively).
Hence, we have shown that the addition of atomic SASA to
the SMARTCyp model adds a valuable contribution to the
model (∼7% of the prediction accuracy).
(1) How To Calculate the Atomic SASA without Using 3D
Structures. Since SMARTCyp is a purely 2D-based method,
we decided to investigate if it was possible to predict the atomic
SASA from 2D structure-based properties, avoiding the need to
generate 3D structures.
As starting point for the building of a predictive model for
atomic SASA, we compiled a diverse selection of 27,838
druglike molecules from the ZINC database.29 The most likely
tautomer and conformation were generated for each molecule
in this subset, and their atomic SASA were computed as
described above. This data set was split into training and test
sets, and atoms were divided into subsets according to the
number of neighboring atoms as described in Table 1.
Using circular fingerprints (described above) predictive
models were built from each training set using the partial
least-squares regression method (PLS).45 The final PLS models
are described in Table 3. Applied to the test sets, the models
have mean absolute errors (MAE) of 0.2−3.8 Å2, with
SASA coefficients of determination (r2) ranging from 0.57 to 0.85.
65.4
In the final model (2DSASA), in which the appropriate PLS
78.1
model is applied to each atom, the MAE is 2.8 Å2 and r2 is 0.96.
84.2
The MAEs of the four PLS models are inversely correlated to
distribution of the data sets (see Tables 1 and 3). This is
because data sets with small data ranges typically get smaller
absolute errors. As can be seen in Figure 7 and Figure S2 in the
Supporting Information, the atomic SASA of the test set
calculated by 2DSASA more often have large positive errors
than large negative ones. However, this is not reflected in the
average error, which is −0.06 for the test set. The errors for
specific atom types correlate with the occurrence of the atom
types in the different data sets (1−4 neighbors), and hence with
the PLS model applied. More details on the errors for specific
atom types are available in the Supporting Information.
Figure 6 describes the variables that are included in the final
PLS models. The atom type variables (T) contribute to levels
0−1 in all models, level 2 in the 1−3-neighbor models, and
level 3 in the 1-neighbor model. The ring count variables (O)
contribute to level 0 in all models except the 1-neighbor one
(an atom with one neighbor cannot be part of a ring). It also
contributes to level 1 in the 1- and 2-neighbor models, and level
2 in the 1-neighbor model. The variable for total number of
atoms (#) contributes to levels 2−3 in all models, and level 4 in
all models except that for 4 neighbors. It is clear that the fewer
atoms that are bound to an atom, the more atoms further away

Table 3. Statistics of the PLS Models in 2DSASA

variables components r2 q2 MAEa,b r2preda MAEpreda,b
c c
2DSASA 35−99 3−5 0.95 N/A 2.83 Å2 0.96 2.76 Å2
PLS models used in 2DSASA
1 neighbor 99 4 0.85 0.85 3.87 Å2 0.85 3.79 Å2
2 neighbors 79 5 0.81 0.81 3.30 Å2 0.82 3.22 Å2
3 neighbors 70 5 0.67 0.67 1.53 Å2 0.67 1.50 Å2
4 neighbors 35 3 0.51 0.49 0.24 Å2 0.57 0.20 Å2

a
Mean absolute error of the training (MAE) and test (MAEpred) sets. bThe predicted atomic SASA are set to zero if the PLS model gives a negative
value. c2DSASA uses the appropriate PLS model for each atom.

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Molecular Pharmaceutics
Figure 6. Atom of interest (filled black circle) with 1−4 neighboring
atoms, respectively. The PLS model for each atom of interest
incorporates variables for atom types (T), ring counts (O), and total
number of atoms (#).
Figure 7. Atomic SASA in Å2 computed by the reference Shrake−
Rupley dot density method and 2DSASA.
will contribute to the PLS model. This is most likely due to the
fact that atoms with many neighboring atoms have less
flexibility, and hence their accessible surface area depends
more on the closest neighbors.
To validate that the atomic SASA computed from our 2D
model gives an accuracy that is useful for the prediction of site
of metabolism, we applied it to the same test set used to
validate the calibration of the atomic SASA contribution to
SMARTCyp above. The results are compared to the results
using atomic SASA calculated from 3D structures in Table 4,
and show that there is no significant difference of using atomic
SASA computed from 2D and 3D structures. Therefore, while
it might be possible to build more accurate 2D predictive
models of atomic SASA using nonlinear models such as random
forests46 and support vector machines,47 there seems to be no
gain in going beyond a simple PLS model for the purpose of
application to site of metabolism modeling, as the difference
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Table 4. Prediction Accuracy for SMARTCyp with 2D and
3D Atomic SASA Contribution on the CYP 3A4 Test Set
3D SASA 2DSASA
top 1% 65.4 65.3
top 2% 78.1 77.9
top 3% 84.2 84.5
between using atomic SASA from our 2D model and from 3D
structures is minimal.
Application of 2D Based Atomic SASA to the
SMARTCyp Prediction Models and Validation to Nine
CYP Isoforms. Since the application of 2DSASA to the CYP
3A4 data set showed good results, we decided to apply this new
standard SMARTCyp model to the other eight isoform data
sets released by Zaretzki et al.8 The results are shown in Table
5 and show improvements in prediction accuracies for all
isoforms ranging from 0.0 to 3.6 percentage points for top 1,
1.5 to 3.6 percentage points for top 2, and 1.7 to 4.3 percentage
points for top 3. The average improvements are 1.4, 2.3, and
2.7 percentage points for top 1, top 2, and top 3 prediction
accuracies, respectively, indicating that the contribution of
atomic SASA is more important for compounds that are harder
to predict (have a lower top-ranked site of metabolism without
atomic SASA).
As seen in Table 5, the prediction accuracies for SMARTCyp
are the lowest for the 2C9 and 2D6 isoforms. This has been
noted earlier and was the major reason for our investigation
into pharmacophore corrections to the prediction of 2C9 and
2D6 substrates which were introduced in version 2 of
SMARTCyp.6,13 Applying the same atomic SASA corrections
to these models gives similar increases in prediction accuracies.
Since the 2C9 and 2D6 models have a different accessibility
descriptor than the standard model, this shows that the simple
correction derived from 50 CYP 3A4 substrates is consistent
across all isoforms and model types (see Table 6).
After the inclusion of 2DSASA in the standard model and the
pharmacophore models, it is obvious that the isoforms for
which SMARTCyp cannot perform as well are CYP 2C8 and
CYP 2C19. Since these are closely related to CYP 2C9, one
might expect that a model similar to the 2C9 model would give
better predictions for these isoforms than the standard model.
Indeed, Liu et al. have shown that a model similar to the
SMARTCyp 2C9 model13 performs better than the standard
model for a small set of CYP 2C19 substrates.48 To investigate
how their results extrapolate to the much larger data sets used
in this study, we applied the 2C9 model to the 2C8 and 2C19
data sets, with and without the pharmacophore correction
included (all with 2DSASA correction included). The results
are significantly better than for the standard model, and the
pharmacophore correction also gives a small positive
contribution to the prediction accuracy for the 2C8 and
2C19 data sets (see Table 7). While these isoforms are not
usually considered to metabolize as many carboxylic acid
containing substrates as 2C9, the results in Table 7 and an
investigation of the data sets show that, when they do, they
tend to oxidize them at the same positions as 2C9.
Thus, the new SMARTCyp version includes three models, all
depending solely on the 2D structure of the compounds to be
predicted:
1. a standard model including reactivity, relative span, and
2DSASA, that is applicable to CYP isoforms 1A2, 2A6,
2B6, 2E1, and 3A4
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Table 5. Prediction Accuracy for the Standard SMARTCyp Model with and without 2D Atomic SASA Contribution on Nine
CYP Isoforms and the Improvements with Atomic SASA
1A2 2A6 2B6 2C8 2C9 2C19 2D6 2E1 3A4
Standarda
top 1% 64.3 71.3 66.0 61.9 55.7 59.6 48.9 64.1 64.4
top 2% 78.5 84.1 74.6 73.8 69.6 74.0 59.8 81.0 75.9
top 3% 85.4 88.7 85.0 80.2 78.8 80.3 68.6 87.1 81.4
+SASAb
top 1% 64.9 72.4 66.2 65.5 58.8 60.1 49.3 64.1 65.4
top 2% 80.0 85.7 76.8 77.5 71.7 76.1 61.1 82.1 78.1
top 3% 88.4 90.5 86.8 84.5 81.4 83.0 70.7 89.0 85.0
a
Equation 1 without SASA. bEquation 1.

Table 6. Prediction Accuracy for the Pharmacophore

SMARTCyp Models with and without 2D Atomic SASA
Contribution on CYP 2C9 and CYP 2D6
SMARTCyp 2.3 +2DSASA diff
2C9
top 1% 70.1 71.0 1.0
top 2% 81.5 83.8 2.4
top 3% 88.0 90.5 2.5
2D6
top 1% 73.5 73.7 0.2
top 2% 82.0 83.0 0.9
top 3% 87.5 89.6 2.1
Figure 8. Variable importance for the final SMARTCyp models
Table 7. Prediction Accuracy for the 2C9 Pharmacophore applied to data sets for nine CYP isoforms. Standard model applied to
SMARTCyp Model with 2D Atomic SASA on CYP 2C8 and 1A2, 2A6, 2B6, 2E1, and 3A4; 2C model applied to 2C8, 2C9, and
CYP 2C19 2C19; 2D6 model applied to 2D6.
std 2C9 model 2C9 model − pharmacophore
2C8 2E1 have a preference for smaller substrates (the median
top 1% 65.5 68.0 63.7 molecular weights in these data sets are 206 and 200,
top 2% 77.5 82.7 81.3 respectively, compared to 271−326 for the other seven data
top 3% 84.5 90.5 89.8 sets), which is because they have the smallest binding sites
2C19 among these isoforms.49,50 However, we know that CYP 2E1
top 1% 60.1 69.7 67.9 also can metabolize various fatty acids,50 for which the relative
top 2% 76.1 85.8 85.3 span might be more important, but such molecules are a
top 3% 83.0 91.3 91.3 minority in the current data set.
The application of the 2C model to the three 2C enzymes
2. a CYP 2C model including reactivity, Span2End, distance also shows a relatively systematic contribution from the local
to carboxylic acid bioisostere, and 2DSASA accessibility (5−7%), whereas the contributions from the other
3. a CYP 2D6 model including reactivity, Span2End, descriptors vary more. The reactivity is less important than in
distance to positively charged amine, and 2DSASA the standard model, contributing 62−69%, and being most
To investigate the relative importance of the descriptors in important in CYP 2C8. The Span2End descriptor contributes
the final models, we performed the same variable exclusion 21−29%, being least important in CYP 2C8, and the
evaluation as done above for all the final models toward the pharmacophore descriptor contributes 1−7%, being most
nine data sets. A summary of the results is shown in Figure 8. important in CYP 2C9 as expected. The importance of the
The standard model was applied to data sets for isoforms 1A2, Span2End descriptor correlates inversely to the likelihood of
finding sites of metabolism that are located further away from
2A6, 2B6, 2E1, and 3A4. In this model, reactivity is highly
the end of the molecules. The average number of bonds from
important, contributing 86−97% of the model, and the two the end of the molecule to a site of metabolism (i.e., the average
accessibility descriptors contribute to the remaining 3−14%. Span2End descriptor value for sites of metabolism) is 1.74 in
The contribution of the relative span and 2DSASA varies from CYP 2C8, 1.46 in CYP 2C9, and 1.33 in CYP 2C19. The larger
0−10% and 3−7%, respectively, suggesting that, while the difference between CYP 2C8 and the other two enzymes most
contribution of the local accessibility developed in this work likely is due to the larger size of its active site,51 since a larger
(2DSASA) gives a relatively systematic contribution, the active site would make it easier to orient the substrates in a way
contribution of the relative span descriptor depends more on that makes more centrally located atoms in a substrate position
the data set. The relative span is least important for the CYP themselves close to the heme group. However, the small
2A6 and CYP 2E1 data sets (1.4 and 0%, respectively), which is difference between CYP 2C19 and CYP 2C9 is harder to
quite interesting since these isoforms have different substrate explain. It could possibly be explained by the difference in
preferences than the other isoforms. Both CYP 2A6 and CYP positions of Phe 114 and Phe 476 as shown in the recent work
1221 dx.doi.org/10.1021/mp3005116 | Mol. Pharmaceutics 2013, 10, 1216−1223
Molecular Pharmaceutics
by Reynald et al.,51 but it might as well be a result of the data
sets used in the current work.
The same evaluation performed for the CYP 2D6 model on
the CYP 2D6 data set shows that reactivity is even less
important than for the 2C family (55%), whereas the
pharmacophore is much more important (35%). This reflects
the substrate preference of CYP 2D6, which has a high
preference for protonated amines, and binds these specifically
with the amine group far away from the heme. The N-
dealkylation of protonated amines, which is initiated by a
hydrogen abstraction from the α-carbon atom, is one of the
CYP-mediated reactions with lowest energy barrier. Thus, to
make a reactivity-based model such as SMARTCyp predict
CYP 2D6 mediated metabolism correctly, a large contribution
from the pharmacophore descriptor is required.
Comparison of the Final Models to More Complex
Models. To compare our methods to others, we show the
prediction accuracies in relation to the recent work on the RS-
predictor methodology by Zaretzki et al.,8 in which models
were built using the MIRank algorithm52 on SMARTCyp
reactivities together with either 148 topological descriptors, or
the same descriptors plus 392 quantum chemical descriptors
computed with AM1. They also compared the results to two
methods implemented in StarDrop and Schrö dinger. A
comparison of the top 2 prediction accuracies is shown in
Figure 9. It shows that our simple 3−4 parameter methods
compare well to the much more complex RS-predictor based
models, as well as the StarDrop and Schrödinger software
packages.
Figure 9. Comparison of top 2 prediction accuracies for SMARTCyp
with 2DSASA included to other prediction methods.
■
CONCLUSIONS
In this work we created 2DSASA, a model for prediction of
atomic SASA from purely 2D structure information. 2DSASA
enables methods built from 2D structure data, e.g., toxicity
alerts and site of metabolism predictions, to take atomic
accessibility into account. We showed that, for a test set
consisting of 20,847 compounds, 2DSASA predict the atomic
SASA with an average absolute error of only 2.8 Å2.
We integrated 2DSASA in SMARTCyp and showed that for
a set of 425 CYP3A4 substrates it gave a model as accurate as
one built from atomic SASA computed from 3D structures. It
was also applied to data sets for eight other CYP isoforms (1A2,
2A6, 2B6, 2C8, 2C9, 2C19, 2D6, and 2E1) and was shown to
consistently improve the predictions. The final models were
1222
Article
compared to much more complex methods and showed better
or similar prediction accuracies for all isoforms.
■
*
ASSOCIATED CONTENT
S Supporting Information
ZINC IDs of all compounds in the data sets, PLS equations for
the 2DSASA models, SYBYL atom type descriptions of the
atom types included in the 2DSASA models, atom type
distributions in the 2DSASA data sets, and errors by atom type
in 2DSASA. This material is available free of charge via the
Internet at http://pubs.acs.org.
■
AUTHOR INFORMATION
Corresponding Author
*Phone: +45 35 33 66 50. Fax: +45 35 33 60 41. E-mail: pry@
sund.ku.dk.
Notes
The authors declare no competing financial interest.
■
ACKNOWLEDGMENTS
The work was supported by grants from the Alfred Benzon
Foundation, the Danish Council for Independent Research
(Medical Sciences), and Lhasa Limited. The authors wish to
thank Nina Jeliazkova for constructing the initial version of the
circular fingerprint java code.
■ ABBREVIATIONS USED
SASA, solvent accessible surface area; CDK, chemistry
development kit; CYP, cytochrome P450
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