Binding of serotonin and N1-benzenesulfonyltryptamine-related analogs at human 5-HT6 serotonin receptors: Receptor modeling studies

Article abstract of DOI:10.1021/jm070910s

A population of 100 graphics models of the human 5-HT₆ serotonin receptor was constructed based on the structure of bovine rhodopsin. The endogenous tryptamine-based agonist serotonin (5-HT; 1) and the benzenesulfonyl-containing tryptamine-deri

Full text of DOI:10.1021/jm070910s

J. Med. Chem. 2008, 51, 603–611
603
Binding of Serotonin and N1-Benzenesulfonyltryptamine-Related Analogs at Human 5-HT6
Serotonin Receptors: Receptor Modeling Studies
Małgorzata Dukat,† Philip D. Mosier,† Renata Kolanos,† Bryan L. Roth,‡ and Richard A. Glennon*,†
Department of Medicinal Chemistry, School of Pharmacy, Medical College of Virginia Campus, Virginia Commonwealth UniVersity, Richmond,
Virginia 23298-0540, and Departments of Pharmacology, Psychiatry, DiVision of Medicinal Chemistry and Natural Products, Neurosciences
Center and ComprehensiVe Cancer Center Schools of Medicine and Pharmacy, UniVersity of North Carolina,
Chapel Hill, North Carolina 27599
ReceiVed July 26, 2007
A population of 100 graphics models of the human 5-HT6 serotonin receptor was constructed based on the
structure of bovine rhodopsin. The endogenous tryptamine-based agonist serotonin (5-HT; 1) and the
benzenesulfonyl-containing tryptamine-derived 5-HT6 receptor antagonist MS-245 (4a) were automatically
docked with each of the 100 receptor models using a genetic algorithm approach. Similar studies were
conducted with the more selective 5-HT6 receptor agonist EMDT (5) and optical isomers of EMDT-related
analog 8, as well as with optical isomers of MS-245 (4a)-related and benzenesulfonyl-containing pyrrolidine
6 and aminotetralin 7. Although associated with the same general aromatic/hydrophobic binding cluster,
5-HT (1) and MS-245 (4a) were found to preferentially bind with distinct receptor conformations, and did
so with different binding orientations (i.e., poses). A 5-HT pose/model was found to be common to EMDT
(5) and its analogs, whereas that identified for MS-245 (4a) was found common to benzenesulfonyl-containing
compounds. Specific amino acid residues were identified that can participate in binding, and evaluation of
a sulfenamide analog of MS-245 indicates for the first time that the presence of the sulfonyl oxygen atoms
enhances receptor affinity. The results indicate that the presence or absence of an N1-benzenesulfonyl group
is a major determinant of the manner in which tryptamine-related agents bind at 5-HT6 serotonin receptors.
Chart 1. Structures of 5-HT (1), 5-HT6 antagonists Ro 04-6790
(2), SB-258510 (3a), SB-271046 (3b), MS-245 (4a) and its
des-methoxy analog 4b, and the agonist EMDT (5)
Introduction
Many actions of the neurotransmitter substance serotonin (5-
hydroxytryptamine; 5-HT, 1) are mediated by its interaction with
various populations (5-HT1-5-HT7) of 5-HT receptors.1 Apart
from ionotropic 5-HT3 receptors, other members of this receptor
family are class A rhodopsin-like G protein-coupled receptors
(GPCRs).1,2 Human 5-HT6 receptors, first cloned in 1996,3 are
positively coupled to an adenylate cyclase second messenger
system, and there is evidence that this receptor population is
involved in cognition, obesity, and certain neurological and
neuropsychiatric disorders, including schizophrenia and depres-
sion.4 This receptor population is also of interest because
clozapine and a number of other typical and atypical antipsy-
chotic agents bind at 5-HT6 receptors with high affinity.5 Only
within the last six or seven years have 5-HT6-selective agents
been identified, and among some of the first antagonists
described were Ro 04-6790a (2),6 SB-258510 (3a) and its
N-desmethyl analog SB-271046 (3b),7 MS-245a (4a),8,9 and the
agonist EMDT (5;8 Chart 1; reviewed4,10).
by their human sequence number followed, in brackets, by their
numbering using the Ballesteros-Weinstein convention)11 and
threonine (Thr196 [5.46]) moieties as being directly or indirectly
involved in the actions of serotonergic agonists and that their
individual mutation substantially reduces both the affinity
(radioligand binding) and the efficacy (stimulation of adenylate
cyclase action) of the nonselective agonists 5-HT (1) and (+)-
lysergic acid diethylamide (LSD).12,13
To date, there have been four attempts to provide insight into
how 5-HT6 receptor–ligands interact with 5-HT6 receptors using
graphical models.14–17 Bromidge14 was first to describe a
possible docking mode for a 5-HT6 antagonist (i.e., SB-258510;
3a) to a 5-HT6 receptor model and his epigrammatic description
implicated Phe277 [6.44] and Trp281 [6.48] as participating in
possible π-π stacking interactions (Table 1). Hirst et al.16
Prior to the availability of selective agents, site-directed
mutagenesis studies with rat 5-HT6 receptors identified con-
served aspartate (Asp106 [3.32]). (There is a high degree of
homology between rat and human 5-HT6 receptors, and their
amino acid sequence numbering is identical for the amino acids
described herein; for convenience, amino acids are numbered
* To whom correspondence should be addressed. Phone: (804) 828-8487.
Fax: (804) 828-7404. E-mail: glennon@vcu.edu.
†
Virginia Commonwealth University.
University of North Carolina.
‡
aAbbreviations: MS-245, N1-benzenesulfonyl-5-methoxy-N,N-dimeth-
yltryptamine; EMDT, 2-ethyl-5-methoxy-N,N-dimethyltryptamine; LSD,
lysergic acid diethylamide; SB-258510, 5-chloro-N-[4-methoxy-3-(4-me-
thylpiperazin-1-yl)phenyl]-3-methyl-2-benzothiophenesulfonamide; Ro 04-
6790, 4-amino-N-(2,6-bis-methylamino-pyrimidin-4-yl)benzenesulfonamide.
10.1021/jm070910s CCC: $40.75
2008 American Chemical Society
Published on Web 01/18/2008

604 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
Dukat et al.
Table 1. Proposed Modes of Interaction for Sulfonamide-Containing 5-HT6 Antagonists
Aryl′-SO2-N-aryl-X-amine
feature
Bromidge14
Pullagurla et al.17
Hirst et al.16
Lopez-Rodriguez et al.15
currenta
amine
aryl
Asp106 [3.32]
Trp281 [6.48]
Asp106 [3.32]
Phe284 [6.51]
Phe285 [6.52]
Asp106 [3.32]
Trp281[6.48]
Phe284 [6.51]
Phe302 [7.35]
Asp106 [3.32]
Phe198 [5.48]
Phe285 [6.52]
Asp106 [3.32]
Val107 [3.33]
Leu182 [e2]
Trp281 [6.48]
Phe284 [6.51]
Ser111 [3.37]
Thr196 [5.46]
Ala157 [4.56]
Ala192 [5.42]
Phe285 [6.52]
SO2
Leu182 [e2]
Gln291 [e3]
Phe302 [7.35]
Asn288 [6.55]
Gln216b
Phe188 [5.38]
Ser193 [5.43]
Asn288 [6.55]
Val107 [3.33]
Phe188 [5.38]
Ala192 [5.42]
Aryl′
Phe277 [6.44]
a Other residues within 5 Å of docked molecules, in 0.5 Å distance increments, are listed in the Supporting Information section. b Both the rat and human
receptors contain a glutamine moiety at position 218, but an arginine residue at position 216. Hence, it is uncertain which of the two amino acids was
actually identified in this study.
examined the docking of several antagonists including Ro 04-
6790 (2) and SB-258510 (3a) at mouse, rat, and human 5-HT6
receptor models to explain the reduced affinity of certain
antagonists (in particular, Ro 04-6790) for mouse receptors.
Implicated in possible π-π stacking interactions (rat model)
were Phe188 [5.38], Trp281 [6.48], Phe284 [6.51], and Phe302
[7.35]. Pullagurla et al.,17 using a human 5-HT6 receptor model,
examined several MS-245 (4a) analogs and identified Phe284
[6.51], Phe285 [6.52], and Phe302 [7.35] as being involved in
π-π interactions. More recently, Lopez-Rodriguez et al.15
studied the binding of a series of 5-HT6 antagonists (including
2, 3b, and 4) using a human 5-HT6 receptor model. An aromatic/
hydrophobic binding region was defined by Val107 [3.33],
Phe188 [5.38], Ala192 [5.42], Phe198 [5.48], and Phe285 [6.52].
Despite using different approaches, each of the four studies
identified certain common amino acids as possibly participating
in the interaction of antagonists with 5-HT6 receptors. That is,
the studies all began with recognition that a role for Asp106
[3.32] is key to binding and identified a common binding pocket
or “cluster” consisting of aromatic/hydrophobic residues in TM5
and TM6 (e.g., Trp281, Phe284, and Phe285). However,
orientation of the ligands within this region differed consider-
ably. For example, the SO2 oxygen atoms of the sulfonyl-
containing antagonists have been variously suggested to be
within hydrogen bonding distance to, for example, Asn288
[6.55], Ser193 [5.43], Leu182 (in the e2 loop), and Gln291 (at
the TM6-e3 junction) (see Table 1).14,15
nonselective agonist 5-HT (1) and of the more 5-HT6-selective
agonist EMDT (5) at human 5-HT6 receptors. We also examined
the docking of the tryptamine-derived antagonist MS-245 (4a)
and structurally related and stereochemically and conforma-
tionally defined benzenesulfonyl analog pyrrolidine 6 and
aminotetralin 7. In light of empirical evidence that N1-
unsubstituted and N1-benzenesulfonyltryptamines likely bind in
a dissimilar manner at 5-HT6 receptors,18,19 and with realization
that agonists and antagonists might bind optimally with different
receptor conformations (and that there might even be differences
in the binding of different agonists at a given receptor),20 we
utilized a docking technique that should emphasize and account
for such differences. Finally, we examined the binding of
stereochemically defined optical isomers of EMDT analog 8
and also determined whether or not the sulfonyl oxygen atom(s)
contributes to binding of benzenesulfonyltryptamines by ex-
amining sulfenamide 9 in comparison with its sulfonyl coun-
terpart 4b. As such, this investigation employed (a) 5-HT6
receptor–ligands bearing a single basic amine, (b) stereochemi-
cally or (somewhat) conformationally defined analogs of the
5-HT6 ligands MS-245 (4a) and EMDT (5), and (c) an
automated and unbiased docking procedure that might account
for differences in the binding of agonist versus antagonist
analogs at human 5-HT6 receptors.
Chemistry. The synthesis of certain compounds required for
this investigation has been previously reported by us, including
MS-245 (4a),8 4b,21 EMDT (5),8 R-(+)-6, and S-(-)-6.22 The
synthesis of compounds 7R and 7S employed the isomers of
11 (Scheme 1). Isomers 11 were prepared in five steps from
5-amino-2-naphthol (10) following a literature procedure23 that
involved protection of the amine, Birch reduction, and stereo-
selective synthesis of 11R and 11S from the resultant ketone.
Dimethylation of the individual optical isomers of 11 afforded
isomers 12, which were deprotected to isomers 13 and then
sulfonylated to isomers 7 by reaction with benzenesulfonyl
chloride (Scheme 1). Compounds 12 and 13 have been previ-
ously reported in the patent literature as their racemates;23
although optical isomers of 13 also were mentioned, synthesis
was not described nor were physicochemical properties re-
corded.23
Two investigations examined agonists,16,17 and only one of
these examined the possible binding modes of 5-HT at human
5-HT6 receptors.17 The two models implicated a role for Asp106
[3.32] and Thr196 [5.46] for interaction with the agonists 5-HT
and/or LSD.16,17
Using graphics models of the receptor, attempts have been
made to identify how 5-HT6 receptor antagonist ligands interact
with various receptor amino acid features. For the most part,
however, these studies involved antagonists with multiple basic
amine groups (with consequent and inherent uncertainty, and
assumptions, as to which amine specifically interacts with the
TM3 aspartate moiety Asp106), employed agents with substan-
tial conformational flexibility, and have assumed that the
sulfonyl moiety (common to nearly all 5-HT6 antagonists)
participates in binding. From what we can discern, only one of
the prior studies explicitly states that a nonmanual docking
method was utilized,17 and only a single study examined a
possible binding mode for 5-HT.17
Isomers 8 were prepared from 2-ethyl-5-methoxyindole24 in
a manner that paralleled the synthesis of 622 and related
compounds.25 Compound 9 was obtained by interaction of N,N-
dimethyltryptamine (DMT) with N-(phenylthio)succinimide.26
The present investigation was designed to specifically ex-
amine the binding of tryptamine-related analogs; the binding
of other 5-HT6 ligands will be examined in due course. In this
investigation we examined possible docking modes for the
Results and Discussion
Human 5-HT6 receptor affinities (Ki values) of certain
compounds employed in this investigation already have been

5-HT6 Receptor Models
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 605
Scheme 1a
a Reagents and conditions: (a) H2CdO (37%), NaCNBH3, HOAc,
MeOH/MeCN, rt, 3 h; (b) concd HCl, EtOH, ∆, 2 h; (c) PhSO2Cl, CH2Cl2,
0 °C, 3 h.
Figure 1. Overlap of the two selected receptor conformations
highlighting those side chains in the binding site that differ most
substantially between the two sites. The position of the receptors relative
to one another was determined by calculating the minimum rmsd of
the R carbon atoms of residues in the binding pocket. Side chains that
are colored by atom type belong to the receptor preferred by ligands
that do not contain a benzenesulfonyl group (green ribbon). Side chains
that are colored either red (rmsd g 2.0 Å) or yellow (1.0 Å g rmsd >
2.0 Å) belong to the receptor selected for the ligands that do contain
a benzenesulfonyl group (purple ribbon). The conformations of other
side chains in the binding site differed between the two receptors but
to a smaller degree. These have been omitted for clarity.
reported from our laboratories: MS-245 and its des-methoxy
counterpart (4a and 4b; Ki ) 2.1 and 4.1 nM, respectively),21
EMDT (5; Ki ) 16 nM),8 R-(+)-6 (Ki ) 0.3 nM), and S-(-)-6
(Ki ) 1.7 nM).22 Aminotetralin isomers 7R and 7S displayed
little enantioselectivity (Ki ) 49 ( 12 and 90 ( 20 nM,
respectively), whereas isomers of the pyrrolidine counterpart
of EMDT showed greater stereoselectivity of binding (8R Ki )
1.8 ( 0.2 nM; 8S Ki ) 220 ( 25 nM). For several structurally
related chiral pyrrolidines, we have previously found greater
enantioselectivity in the absence of the benzenesulfonyl group
(up to 70-fold) than in its presence (<6-fold).22 Sulfenamide 9
(Ki ) 90 ( 10 nM) displayed >20-fold reduced affinity relative
to its sulfonamide counterpart 4b (Ki ) 4.1 nM).
Model Construction and Docking Studies. Human 5-HT6
receptor models were constructed and various docking solutions
were explored for 5-HT (1), MS-245 (4a), EMDT (5), MS-245
analogs 6, aminotetralins 7, and EMDT-related optical isomers
8 such that both the receptor models and the ligands were
allowed to interact in a fairly flexible manner. Receptor
flexibility was explicitly addressed via the generation of a
population of 100 h5-HT6 receptor models whose conforma-
tional state varied among the members of the population.
Automated docking routines were then used to flexibly place
each ligand into each receptor model. The resulting receptor-
–ligand complexes were scored and subsequently ranked using
the ChemScore27,28 fitness function. Thus, each ligand was
allowed to select an optimal receptor conformation from the
population, reducing the bias that might occur when using a
single receptor model to dock different ligands. Two common
receptor conformations were identified: one for agents lacking
a benzenesulfonyl group (i.e., 5-HT, EMDT and analogs) and
another for agents possessing such a group (i.e., MS-245 and
related analogs). The top-scoring receptor conformation for each
nonbenzenesulfonyl analog was identical. The top-scoring
receptor conformation for benzenesulfonyl-containing analogs
also was the same, except that for MS-245 (4a), the second
highest top-scoring receptor conformation was selected. The
latter (ChemScore ) 39.32 relative to the top score of 41.20)
was selected because of its commonality with the receptor
conformation identified for the other benzenesulfonyl-containing
analogs. In the top-scoring MS-245 receptor conformation, the
hydrogen bonding component suffers at the expense of a greater
lipophilic component, reflecting a somewhat different pose for
the benzenesulfonyl moiety.
The primary differences in the two receptor models are
depicted in Figure 1. The largest difference in side chain
orientation occurs for Trp281 [6.48]. The phenyl portion of the
indole nucleus of Trp281 is oriented toward transmembrane
helix 5 (TM5) for the nonbenzenesulfonyl-containing com-
pounds, and toward transmembrane helix 3 (TM3) for benze-
nesulfonyl-containing compounds. In addition, residues com-
prising the second extracellular loop (e2) also exhibited a
significantly different position and orientation between the two
receptor models. This is not surprising because the overall
movement of the e2 loop is expected to be greater than that for
the transmembrane helical segments. A complete list of rmsd
differences for individual residues of the binding cavity may
be found in the Supporting Information section. The two top
receptor solutions are described in more detail in the following
sections as they relate to docked ligands.
5-HT and EMDT Analogs. With the amine function docked
within 2.7 Å of Asp106 [3.32], the indolic nucleus of 5-HT is
situated in a cluster of amino acids consisting of Leu182 [e2],
Trp281 [6.48], and Phe284 [6.51] (Figure 2). Previous models
for docked 5-HT17 and LSD16 suggest that the indolic NH might

606 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
Dukat et al.
Figure 3. Proposed binding mode of MS-245 (4a) in the h5-HT6
receptor, looking through the extracellular side of the helical bundle.
TM5, TM6, and TM7 are nearest to the viewer. The MS-245 ligand is
rendered as a ball-and-stick model with the carbon backbone in green,
and the side chains of residues whose heavy atoms fall within 4.0 Å of
the ligand’s heavy atoms are rendered as capped sticks.
Figure 2. Proposed binding mode of serotonin (5-HT; 1) in the h5-
HT6 receptor, looking through the extracellular side of the helical
bundle. TM5, TM6, and TM7 are nearest to the viewer. The backbone
ribbon trace is color-coded based on the assignment of secondary
structure using the Kabsch-Sander29 algorithm (red ) helix, blue )
sheet, violet ) turn, and yellow ) coil). 5-HT (1) is rendered as a
CPK-style space-filling model, and the side chains of residues whose
heavy atoms fall within 4.0 Å of the ligand’s heavy atoms are rendered
as capped sticks.
individual ChemScore components indicated that the hydrogen
bond component was higher for the R-isomer (2.62) than for
the S-isomer (2.40). This suggests that the ammonium group
of 8R is able to form a stronger ionic/hydrogen bond with
Asp106 than can the ammonium group of 8S. Other differences
in the ChemScore components occurred for the clash (8R )
3.84; 8S ) 3.30) and internal torsion (8R ) 4.12; 8S ) 5.77)
penalty terms. The clash penalty term, which is a measure of
undesirable van der Waals overlap between atoms of the ligand
and those of the receptor, was higher for 8R, whereas the internal
torsion penalty, which is a measure of the deviation from ideality
of the rotatable bonds (i.e., gauche vs eclipsed substituents),
was higher for 8S. Analyses of molecular dynamics simulation
data to explore more fully the significance of these terms as
they relate to the binding modes of the two isomers are ongoing.
hydrogen bond with Thr196 [5.46]. In the present model, the
indolic NH is directed toward Arg181 [e2], which is <4 Å
distant. This virtually turns the indolic nucleus “upside down”
relative to a prior binding motif described by us.17 Both Ser111
[3.37] and Thr196 [5.46] (Figure 2) are within hydrogen bond
distance of the 5-HT hydroxyl group (O-O distance 3.0–3.1
Å), indicating that the hydroxyl group can form a hydrogen
bond with either one or, simultaneously, with both amino acids.
This is consistent with the observation that removal of the
5-hydroxyl group of 5-HT results in several-fold decreased
affinity.30 These hydrogen bond interactions were stable during
a 100-ps dynamics run (see Experimental Section). Similar
results were obtained with EMDT (5) except that the methoxy
oxygen atom is a little closer (2.6 Å) to Ser111 [3.37] and
Thr196 [5.46].
Both optical isomers of EMDT analog 8 were also docked
in a similar manner. Of the 100 docking solutions, the top
solution (receptor conformation) for both isomers was the same
docking model identified for 5-HT (1) and EMDT (5). The
primary difference in docking modes for 8R and 8S is that the
N-methyl group of 8R was directed toward the extracellular
region (opening) of the binding cavity, whereas for 8S, the
N-methyl group faced the intracellular region (bottom) of the
binding cavity. The pose distinctions for the isomers of 8 are
reflected in their ChemScore values of 36.96 for 8R and 35.13
for 8S. The ChemScore is larger for the higher-affinity 8R,
which is consistent with the observed binding affinities for the
two isomers. While the scores are similar in magnitude, there
are some important but subtle differences in the components
that contribute to the total ChemScore values. Analysis of the
MS-245 and Analogs. A common binding model was
identified for MS-245 (4a), pyrrolidine 6R, and aminotetralin
7R (as illustrated in Figure 3 for MS-245). With the basic amine
of each ligand within 2.7 Å of Asp106, their aryl moieties are
situated in essentially the same hydrophobic pocket described
above for 5-HT (1); however, they are positioned differently
than 5-HT. For example, the benzenesulfonyl aryl groups (see
Aryl′ in Table 1) are within 3.5 to 4.0 Å of Ala157, Ala192,
and Phe285. Furthermore, the sulfonyl oxygen atoms are within
hydrogen bond distance (O-O distance: 2.8–3.3 Å) to both
Ser111 [3.37] and Thr196 [5.46], except for 6R, which is located
within 3.0 Å of Ser111 and 3.8 Å away from Thr196. The model
indicates that the sulfonyl oxygen atoms should be able to
participate in hydrogen bond formation with at least one, if not
both, of these amino acids. That the model is common to MS-
245 (4a) and stereochemically or conformationally defined
analogs 6R and 7R is telling and suggests that the conformation
of MS-245, shown in Figure 3, reflects a conformation optimal
for binding. Furthermore, as shown in Figure 3, the benzene-
sulfonyl group is nearly 90° out of the plane of the indole
nucleus; this is a conformation we had previously predicted

5-HT6 Receptor Models
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 607
would be optimal for binding on the basis of studies with
conformationally constrained benzenesulfonyl analogs.18
The current model, like prior models, shows that the sulfonyl
oxygen atoms of sulfonyl-containing antagonists are positioned
for a productive interaction with certain receptor-associated
features. This raises the obvious question: Are the sulfonyl
oxygen atoms actually required for binding? To address this
issue, we prepared and examined sulfenamide 9 (Ki ) 90 nM),
which was found to bind with 20-fold lower affinity than its
sulfonyl counterpart 4b (Ki ) 4.1 nM). This provides the first
evidence that at least one of the sulfonyl oxygen atoms of MS-
245-type analogs is required for enhanced affinity. Moreover,
that the sulfonyl oxygen atom(s), and not the methoxy group,
of MS-245 (4a; Ki ) 2.1 nM) interacts with Thr196 is consistent
with the finding that removal of the methoxy group has little
impact on affinity (i.e., 4b Ki ) 4.1 nM). Also, as shown in
Figure 3, there are no amino acid residues within about 7 Å
that could potentially hydrogen bond with this methoxy group.
Indeed, even an intact indole nucleus is not required for
tryptamines to bind at 5-HT6 receptors. This is evident from
the binding of phenylethylamine 15 (Ki ) 52 nM)31 and
aminotetralins 7. Compound 16 (Ki ) 15 nM),31 which lacks
the benzenoid ring and methoxy substituent of MS-245 (4a),
binds with only 4-fold lower affinity than 4b. This further
supports the notion that the methoxy substituent of MS-245 is
unlikely to interact with a specific receptor-associated feature.
The somewhat reduced affinity of 16 relative to 4 can be
explained by the fewer aromatic/hydrophobic contacts possible
with the abbreviated heterocyclic nucleus.
either display reduced or enhanced affinity for 5-HT6 receptors
(depending upon the nature of the N1-substituent), and have
previously attributed these affinity differences to different modes
of binding.9,18,19
The present model shows that the hydroxyl group, not the
N1-H, of 5-HT (1) is situated within hydrogen bonding distance
to Thr196 [5.46] (and Ser111 [3.37]). Interesting to note is that
there are two conserved hydroxyl-containing amino acids
(Ser204 [5.43] and Ser207 [5.46]) common to rhodopsin-like
G-protein coupled receptors that bind catecholamine agonists
(i.e., dopamine receptors, adrenergic receptors)33 and that it has
been shown on the basis of site-directed mutagenesis studies
that these two residues are important for binding and agonist
action and probably participate in hydrogen bond interactions
with the hydroxyl groups of catecholaminergic agonists.33–35
Other than for Asp106 and Thr196, few other single amino
acid mutations have been reported. Mutation of the rat Trp102
[3.28] (i.e., W102F mutant) had only a small effect on the
affinity of 5-HT (<2-fold) or on several nonselective antagonists
(<6-fold).12 Although Trp102 is not within 5 Å of docked 5-HT,
it is within 4.5-5.0 Å of EMDT (5) and the other agents
examined (see Supporting Information). Given its distance from
the docked molecules, Trp102 might not be expected to exert a
major influence on binding. In the rat, mutation of Ala154 [4.53]
to serine had no effect on the receptor affinity or agonist action
of 5-HT;12 Ala154 was not found within 5.0 Å of docked 5-HT
in the current model. Teitler and co-workers36 found that
mutation of Ser267 [6.34] to lysine enhances the affinity of 5-HT
at human 5-HT6 receptors. However, it is unlikely that Ser267
is directly involved in the binding of 5-HT because it is near
the intracellular margin of the receptor. Furthermore, it has been
demonstrated that the Ser267 mutation results in a constitutively
active receptor,36,37 and it is known that constitutively active
forms of 5-HT receptors characteristically display enhanced
affinity for 5-HT.38 Human 5-HT6 receptor mutation constructs
also have been examined in the intracellular loop regions.39 On
this basis, a particular residue (Lys265) has been shown
instrumental in the coupling of 5-HT6 receptors to GRs; however,
there is no reason to suspect any direct involvement in ligand
binding.
Taken together, new models are presented for the interaction
of 5-HT and several tryptamine-related ligands at 5-HT6
receptors. Docking solutions were identified for the interaction
of agonist 5-HT (1), EMDT (5), and EMDT analog 8, each at
100 different receptor conformations. In each case, their
preferred (based on ChemScore) docking solution was the same.
With this model, the basic amines interact with Asp106, the
5-hydroxy (or methoxy) group interacts with Thr196 and/or
Ser111, and the indolic nucleus is located in an aromatic/
hydrophobic pocket. Antagonist MS-245 (4a) binds differently;
MS-245 utilizes the same aromatic/hydrophobic pocket but
otherwise binds in an altogether different fashion than 5-HT.
Specifically, the sulfonyl oxygen atoms are situated within
hydrogen bond distance of Thr196 and Ser111, and the
benzenesulfonyl aryl group (Aryl′; Table 1) interacts with
Ala157, Ala192, and Phe285. (Note: other nearby amino acid
residues are either shown in Figure 3 or are listed in Supporting
Information.) Benzenesulfonyl compounds 6R and 7R were
found to dock in a similar manner to this common model.
Consistency with Site-Directed Mutagenesis Results. As
already mentioned, Asp106 and Thr196 have been shown to
be important for ligand binding at 5-HT6 receptors. Two prior
modeling studies with N1-unsubstituted tryptamine analogs (i.e.,
5-HT and LSD) have suggested that the indolic N1-H moiety
might be within hydrogen bonding distance to Thr196,16,17
a
finding not inconsistent with earlier site-directed mutagenesis
studies that showed the importance of this residue for agonist
binding and agonist efficacy. That is, on the basis of site-directed
mutagenesis studies showing that the affinity of N1-unsubstituted
LSD-related ergolines (i.e., LSD, ergotamine, lisuride) is
decreased and that the affinity of N1-methyl ergolines (i.e.,
methysergide and mesulergine, but not metergoline) is enhanced
when Thr196 is mutated to alanine, it was speculated that the
indolic NH of 5-HT (1) might participate in hydrogen bond
formation with Thr196.13 This is certainly a possibility; however,
it is not known how ergolines bind relative to 5-HT. In fact,
comparing the affinities of various ergolines at wild type and
several mutant 5-HT2A receptors, it has been suggested that there
are differences in binding modes even among the ergolines.32
Hence, there is no compelling or a priori reason to suspect, or
assume, that 5-HT (1) and LSD-related ergolines must bind in
a comparable manner to 5-HT receptors. Furthermore, we have
found that N1-substituted analogs of 5-HT-related tryptamines
In summary, two relatively similar, but distinct, 5-HT6
receptor binding models are described: one for the agonist 5-HT
(1) and another for the antagonist MS-245 (4a). Both ligands
utilize a common binding domain but are oriented quite
differently within this region. There is consistency with the

608 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
Dukat et al.
Table 2. Results of Microanalysis (Atlantic Microlab)
R-(+)-2-Ethyl-5-methoxy-3-[(1-methylpyrrolidin-2-yl)methyl]-
1H-indole Hydrochloride (8R). A solution of 14R (0.54 g, 1.38
mmol) in dry THF (7 mL) was added in a dropwise manner at 0
°C to a stirred suspension of LiAlH4 (0.24 g, 6.21 mmol) in dry
THF (7 mL). The resulting mixture was heated under a N2
atmosphere for 4 h. The reaction mixture was cooled, and sodium
sulfate decahydrate (7 g) was added very carefully, portionwise,
followed by H2O (0.24 mL) and EtOAc (7 mL). The resulting
mixture was allowed to stir at room temperature under a N2
atmosphere for 24 h. The reaction mixture was then filtered through
Celite, and solvent was evaporated under reduced pressure. The
residual oil was then chromatographed on a silica gel column
(Aldrich silica gel 60) using first EtOAc (to remove benzyl alcohol)
and then EtOAc/MeOH/NH4OH (9:1:2 drops) as eluent to give a
crude product (0.33 g, 88%) as an oil. The product was converted
to its hydrochloride salt in anhydrous MeOH by addition of HCl/
Et2O to give an oil that solidified under high vacuum. The solid
was recrystallized from CH2Cl2/Et2O to give the product (0.27 g,
64%) as off-white crystals: mp 157–159 °C; 1H NMR (DMSO-d6)
δ 1.25 (t, 3H, CH3), 1.68–1.92 (m, 4H, CH2), 2.72 (m, 2H, CH2),
2.87 (s, 3H, CH3), 2.92–3.06 (m, 2H, CH2), 3.27 (dd, 1H, CH2),
3.48–3.58 (m, 2H, CH2), 3.77 (s, 3H, CH3), 6.67 (d, 1H, ArH),
calculated/found
%C
%H
%N
7R
7S
8R
8S
9
57.51
57.87
57.51
57.45
66.11
66.29
66.11
65.83
62.16
61.94
6.44
6.21
6.44
6.10
8.16
8.13
8.16
8.12
5.74
5.76
7.45
7.46
7.45
7.36
9.07
9.18
9.07
8.88
7.25
7.26
binding of other tryptamine-related analogs at one or the other
of these two models, depending upon the presence or absence
of an N1-benzenesulfonyl group, and the models are also
consistent with structure-affinity findings and the results of site-
directed mutagenesis. We already have provided empirical
evidence that tryptamine analogs likely bind differently at 5-HT6
receptors, depending upon whether or not they possess an N1-
benzenesulfonyl group; thus, it is particularly gratifying that
two distinct modes of binding were identified by the automated
docking procedure employed herein. The models will now be
employed to examine the binding of other 5-HT6 receptor
agonist and antagonist ligands to define pharmacophore models
for 5-HT6 receptor agonists and antagonists.
7.00 (s, 1H, ArH), 7.16 (d, 1H, ArH), 10.75 (s, 1H, NH); [R]20
)
D
+19.6 (c 0.5, MeOH). Anal. Calcd for (C17H24N2O·HCl) C, H, N
(see Table 2).
S-(-)-2-Ethyl-5-methoxy-3-[(1-methylpyrrolidin-2-yl)methyl]-
1H-indole Hydrochloride (8S). Beginning with the acid chloride
of N-(benzyloxycarbonyl)-L-proline (i.e., 14S), the target compound
was prepared in 61% yield in the same manner as its R-(+)-
enantiomer 8R, and isolated as off-white crystals after recrystalli-
zation from CH2Cl2/Et2O: mp 156.5–158.5 °C; 1H NMR (DMSO-
d6) δ 1.24 (t, 3H, CH3), 1.68–1.94 (m, 4H, CH2), 2.73 (m, 2H,
CH2), 2.86 (s, 3H, CH3), 2.94–3.08 (m, 2H, CH2), 3.27 (dd, 1H,
CH2), 3.48–3.58 (m, 2H, CH2), 3.77 (s, 3H, CH3), 6.67 (d, 1H,
ArH), 7.00 (s, 1H, ArH), 7.16 (d, 1H, ArH), 10.75 (s, 1H, NH);
[R]D ) -24.6 (c 0.5, MeOH). Anal. Calcd for (C17H24N2O·HCl)
C, H, N (see Table 2).
1-Phenylthio-N,N-dimethyltryptamine Oxalate (9). A solution
of N-(phenylthio)succinimide26 (0.21 g, 1.0 mmol) in CH2Cl2 was
added to a vigorously stirred mixture of N,N-dimethyltryptamine
(0.09 g, 0.5 mmol), tetrabutylammonium hydrogen sulfate (0.02 g,
0.05 mmol), and 50% aqueous KOH (0.5 mL) in CH2Cl2 (2.5 mL)
over a 10-15 min period at room temperature. After half of the
succinimide was added, additional tetrabutylammonium hydrogen
sulfate (0.02 g, 0.05 mmol) was added to the reaction mixture. The
reaction mixture was washed with H2O (3 × 10 mL), the organic
portion was separated and dried (Na2SO4), and the solvent was
evaporated under reduced pressure to give an oily yellow residue.
Purification by chromatography on a silica gel column (Aldrich
silica gel 60) using CH2Cl2/MeOH (9:1) as eluent gave 9 (free base;
0.10 g, 68%) as a yellow oil: 1H NMR (CDCl3) δ 2.37 (s, 6H,
CH3), 2.67 (t, 2H, CH2), 2.96 (t, 2H, CH2), 6.92 (d, 2H, ArH),
7.06 (s, 1H, ArH), 7.13–7.31 (m, 5H, ArH), 7.58–7.65 (m, 2H,
ArH). A small sample of the free base was converted to the oxalate
salt and recrystallized from MeOH to give the product as white
crystals: mp 191–192 °C. Anal. Calcd for (C18H20N2S·C2H2O4) C,
H, N (see Table 2).
R-5-Acetamido-2-dimethylaminotetralin (12R). A solution of
aqueous H2CO (37% w/w, 0.79 mL) was added in a dropwise
manner to a cooled (ice bath) and stirred mixture of R-acetamido-
2-aminotetralin23 (11R; 0.51 g, 2.5 mmol), sodium cyanoborohy-
dride (0.35 g, 5.6 mmol), and glacial HOAc (0.14 mL, 2.5 mmol)
in a mixture of dry MeOH (30 mL) and MeCN (10 mL). The
mixture was allowed to warm to room temperature and stirring was
continued for 3 h before addition of saturated aqueous K2CO3 (25
mL). Organic solvent was removed under reduced pressure, and
the residue was diluted with H2O (25 mL) and extracted with EtOAc
(3 × 30 mL). The combined organic portion was washed with brine
and dried (Na2SO4). Solvent was removed under reduced pressure
to give the product (0.53 g, 91%) as an oily residue that was used
in the preparation of 13R: 1H NMR (CDCl3) δ 1.72 (m, 1H, CH2),
Experimental Section
Chemistry. Melting points were taken in glass capillary tubes
on a Thomas-Hoover melting point apparatus and are uncorrected.
1H NMR spectra were recorded with a Varian EM-390 spectro-
meter, and peak position are given in parts per million (δ) downfield
from tetramethylsilane as internal standard. Microanalyses were
performed by Atlantic Microlab (GA) for the indicated elements,
and the results are within 0.4% of calculated values. Reactions and
product mixtures were routinely monitored by thin-layer chroma-
tography (TLC) on silica gel precoated F254 Merck plates, and
chromatographic separations were performed on silica gel columns.
R-(+)-5-Benzenesulfonamido-2-(dimethylamino)tetralin Hy-
drochloride (7R). A solution of benzenesulfonyl chloride (0.08 g,
0.5 mmol) in dry CH2Cl2 (2 mL) was added in a dropwise manner
at 0 °C to a solution of 13R (0.09 g, 0.5 mmol) in dry CH2Cl2 (2
mL). The reaction mixture was allowed to stir at 0 °C for 3 h,
solvent was evaporated under reduced pressure, and the residue
was recrystallized from MeOH/Et2O. The residue was converted
to a salt by treatment of a dry methanolic solution with HCl-
saturated anhydrous Et2O. The precipitate was collected by filtration
and recrystallized from MeOH/Et2O to give 7R (0.05 g, 28%) as
light-pink crystals: mp 194–196 °C; [R]20 ) +69.0° (c 0.5,
D
MeOH); 1H NMR (DMSO-d6) δ 1.50 (m, 1H, CH2), 2.19 (m, 1H,
CH2), 2.47 (m, 1H, CH2), 2.74–3.00 (m, 8H, CH3, CH2), 3.14 (d,
1H, CH2), 3.36 (m, 1H, CH2), 6.73 (d, 1H, ArH), 6.99–7.09 (m,
2H, ArH), 7.55–7.60 (m, 2H, ArH), 7.64–7.69 (m, 3H, ArH), 9.64
(bs, 1H, NH), 10.66 (bs, 1H, NH+). Anal. Calcd for
(C18H22N2O2S·HCl·0.5H2O) C, H, N (see Table 2).
S-(-)-5-Benzenesulfonamido-2-(dimethylamino)tetralin Hy-
drochloride (7S). Compound 7S was prepared from 13S in 32%
yield in the same manner as 7R. The salt was obtained as light-
pink crystals after recrystallization from MeOH/Et2O: mp 195–197
°C; [R]20D ) -75.7° (c 0.5, MeOH); 1H NMR (DMSO-d6) δ 1.51
(m, 1H, CH2), 2.19 (m, 1H, CH2), 2.47 (m, 1H, CH2), 2.75–3.01
(m, 8H, CH3, CH2), 3.13 (d, 1H, CH2), 3.35 (m, 1H, CH2), 6.73
(d, 1H, ArH), 6.99–7.09 (m, 2H, ArH), 7.54–7.60 (m, 2H, ArH),
7.64–7.69 (m, 3H, ArH), 9.64 (bs, 1H, NH), 10.65 (bs, 1H, NH+).
Anal. Calcd for (C18H22N2O2S · HCl · 0.5H2O) C, H, N (see
Table 2).

5-HT6 Receptor Models
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 609
2.19–2.29 (m, 4H, CH2, CH3), 2.48–2.56 (m, 7H, CH2, CH3), 2.66
(m, 1H, CH2), 2.82–2.94 (m, 2H, CH2), 3.06 (d, 1H, CH2), 6.97
(d, 1H, ArH), 7.14–7.19 (m, 2H, ArH, NH), 7.54 (d, 1H, ArH).
S-5-Acetamido-2-N,N-dimethylaminotetralin (12S). The com-
pound was prepared in 87% yield from S-5-acetamido-2-aminote-
tralin (11S),23 in the same manner as 12R, and isolated as an oily
tion. Cells were then washed by centrifugation and resuspension
once in phosphate-buffered saline (pH ) 7.40; PBS) and then frozen
as tight pellets at -80 °C until use. Binding assays were performed
at room temperature for 90 min in binding buffer (50 mM Tris-Cl,
10 mM MgCl2, 0.1 mM EDTA, pH ) 7.40) with [3H]LSD (1 nM
final concentration) using 10 µM clozapine for nonspecific binding.
Various concentrations of unlabeled test agent were used for Ki
determinations, with Ki values calculated using the program
GraphPad Prism. Specific binding represented 80–90% of total
binding. Ki values represent a minimum of triplicate determinations.
Computational Methods. Molecular modeling studies were
conducted using AMD 64 Dual-Core Opteron-based HP xw9400
and Intel Dual-Core Xeon-based SGI VSS40 Linux workstations
utilizing SYBYL (version 7.3; Tripos Associates, Inc., St. Louis,
MO).
The amino acid sequence of the human 5-HT6 receptor (accession
code P50406) was retrieved from the SwissProt-ExPASY databank
and aligned to the sequence of bovine rhodopsin (P02699) based
on the Bissantz et al.40 multiple alignment profile of several diverse
type A human GPCRs: dopamine D3 (P35462), muscarinic acetyl-
choline m1 (P11229), vasopressin V1a (P37288), ꢀ2-adrenoceptor
(P07550), and δ-opioid (P41143). The ClustalX41,42 multiple
alignment program was used; in short, (i) the amino acid sequences
of the GPCR profile were aligned to the sequence of bovine
rhodopsin in two steps starting from the first transmembrane helix
(TM1) to the first five residues of the third intracellular loop (i3)
and then from the last five residues of the i3 loop to the end of the
seventh transmembrane helix (TM7) using the BLOSUM weight
matrices and with a gap opening penalty of 15.0, and (ii) the
alignment of extracellular loop e2 was manually adjusted to align
the conserved disulfide bridge in the structure of bovine rhodopsin
with the corresponding cysteines in all five aminergic receptors.
This alignment provided an unambiguous mapping of residues in
the TM regions containing no additions or deletions.
When the alignment described above was used, 3D models of
the h5-HT6 receptor were generated based on the 2.2 Å resolution
X-ray crystal structure43 of bovine rhodopsin (“A” chain of PDB
code 1U19). Using MODELER44 (version 9.1), a population of
100 models was generated whose members possessed varied side
chain and, to a lesser extent, backbone conformations. The N- and
C-termini were truncated prior to the first and after the seventh
TM helix, respectively. The e3 loop (which varies considerably
both in terms of length and sequence identity between bovine
rhodopsin and 5-HT6) was modeled as a simple poly Gly sequence
whose initial backbone coordinates were taken from rhodopsin.
Following the procedure of Xhaard et al.,47 the variation in the
side chain conformations of the MODELER-generated h5-HT6
receptors was maximized by mutating to alanine the residues falling
within 12.0 Å of the retinal ligand in the 1U19 MODELER
rhodopsin template. After adding hydrogen atoms to the one
hundred MODELER-generated h5-HT6 models, the receptors were
energy-minimized (Tripos Force Field, Powell minimization,
termination criterion: energy gradient e 0.05 kcal/(mol · Å),
Gasteiger-Hückel charges with a distance-dependent dielectric
constant ) 4.0, and a nonbonded cutoff of 8 Å) using a SYBYL
programming language script (written in-house).
1
residue: H NMR (CDCl3) δ 1.72 (m, 1H, CH2), 2.19–2.29 (m,
4H, CH2, CH3), 2.48–2.56 (m, 7H, CH2, CH3), 2.66 (m, 1H, CH2),
2.82–2.94 (m, 2H, CH2), 3.06 (d, 1H, CH2), 6.97 (d, 1H, ArH),
7.14–7.19 (m, 2H, ArH, NH), 7.54 (d, 1H, ArH).
R-5-Amino-2-dimethylaminotetralin (13R). Concentrated HCl
(20 mL) was added to a solution of R-5-acetamido-2-dimethylami-
notetralin (12R; 0.50 g, 2.2 mmol) in absolute EtOH (60 mL), and
the reaction mixture was heated at reflux for 2 h. The mixture was
cooled to 0 °C and basified with 40% NaOH to pH 12. The mixture
was extracted with CH2Cl2 (3 × 50 mL), the combined organic
portion was dried (Na2SO4), and the solvent was evaporated under
reduced pressure to give the product (0.35 g, 86%) as a semisolid
1
material: H NMR (CDCl3) δ 1.72 (m, 1H, CH2), 2.30 (m, 1H,
CH2), 2.41–2.56 (m, 7H, CH3, CH2), 2.68–2.86 (m, 3H, CH2), 2.97
(d, 1H, CH2), 3.60 (bs, 2H, NH2), 6.56 (d, 1H, ArH), 6.59 (d, 1H,
ArH), 6.99 (t, 1H, ArH). The compound was used without further
purification in the synthesis of 7R.
S-5-Amino-2-dimethylaminotetralin (13S). Beginning with
12S, compound 13S was prepared in 78% yield in the same manner
1
as 13R and isolated as a semisolid material: H NMR (CDCl3) δ
1.71 (m, 1H, CH2), 2.27 (m, 1H, CH2), 2.44–2.55 (m, 7H, CH3,
CH2), 2.63–2.84 (m, 3H, CH2), 2.96 (d, 1H, CH2), 3.60 (bs, 2H,
NH2), 6.55 (d, 1H, ArH), 6.59 (d, 1H, ArH), 6.98 (t, 1H, ArH).
R-3-[[N-(Benzyloxycarbonyl)pyrrolidin-2-yl]carbonyl]-2-
ethyl-5-methoxy-1H-indole (14R). Oxalyl chloride (0.19 mL, 2.21
mmol) was added to a stirred solution of N-(benzyloxycarbonyl)-
D-proline (0.35 g, 1.47 mmol) in dry CH2Cl2 (5 mL) with a trace
of DMF (1 drop). The resulting effervescent solution was allowed
to stir at room temperature under a N2 atmosphere for 3 h. Solvent
was evaporated under reduced pressure, dry hexane (3 mL) was
added, the resulting solution was again evaporated under reduced
pressure to afford N-(benzyloxycarbonyl)-D-proline acid chloride,
and the crude product was dissolved in dry benzene (3 mL).
Concomitantly, a solution of ethyl magnesium bromide (3.0 M
in Et2O, 1.07 mL, 3.09 mmol) was added in a dropwise manner to
a stirred solution of 2-ethyl-5-methoxyindole24 (0.52 g, 2.94 mmol)
in benzene (10 mL) at 0 °C under a N2 atmosphere. The resulting
mixture was allowed to stir at the same temperature for 15 min,
then the solution of N-(benzyloxycarbonyl)-D-proline acid chloride
in benzene was added in a dropwise manner with vigorous stirring.
The reaction mixture was allowed to stir at 0 °C under a N2
atmosphere for 1 h, a saturated aqueous solution of NaHCO3 (7
mL) and EtOAc (9 mL) were successively added, and the mixture
was allowed to stir at room temperature for an additional 1 h. The
organic layer was removed and the aqueous portion was extracted
with EtOAc (3 × 10 mL). The organic extracts were combined
and dried (MgSO4), and the solvent was evaporated under reduced
pressure. The residue was chromatographed on a silica gel column
(Aldrich silica gel 60) using CH2Cl2/MeOH (195:5) as eluent to
give a crude product (0.27 g, 47%) as a semisolid material. The
product was used without further characterization in the preparation
of 8R.
The 3D structures of the ligands were generated using the
SKETCH MOLECULE command within SYBYL. For chiral
compounds, the individual isomers were sketched as separate
structures. Basic amines were protonated. All ligand structures were
then energy-minimized using the same parameters as described
above for the receptor models.
S-3-[[N-(Benzyloxycarbonyl)pyrrolidin-2-yl]carbonyl]-2-
ethyl-5-methoxy-1H-indole (14S). Compound 14S was prepared
in 51% yield from N-(benzyloxycarbonyl)-L-proline in the same
manner as 14R and isolated as a yellow semisolid material. The
product was used without further characterization in the preparation
of 8S.
Radioligand Binding Assay. The h5-HT6 radioligand binding
assays were performed as previously described.3 In brief, h5-HT6
cDNA was transiently expressed in HEK-293 cells using Fugene6
according to the manufacturer’s recommendations. At 24 h after
transfection, the medium was replaced; 24 h later, medium
containing dialyzed serum (to remove 5-HT) was added. At 75 h
after transfection, cells were harvested by scraping and centrifuga-
The automated docking program GOLD 45,46 (Genetic Optimiza-
tion for Ligand Docking, version 3.1, CCDC, Cambridge, U.K.)
was employed to dock ligands into each of the one hundred h5-
HT6 receptor models. A 20 Å radius from the Asp106 [3.32] side
chain Cγ atom was used to define the receptor binding cavity. A
protein hydrogen bonding constraint was defined that included the
oxygen atoms of the Asp106 [3.32] side chain. Docking simulations
were not restricted to early termination and the ligands were allowed
to rotate around single bonds and nonaromatic rings were free to

610 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
Dukat et al.
(8) Glennon, R. A.; Lee, M.; Rangisetty, J. B.; Dukat, M.; Roth, B. L.;
Savage, J. E.; McBride, A.; Rauser, L.; Hufesien, L.; Lee, D. K. H.
2-Substituted tryptamines: Agents with selectivity for 5-HT6 serotonin
receptors. J. Med. Chem. 2000, 43, 1011–1018.
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J. E.; Roth, B. L.; Hufesein, S.; Lee, M.; Glennon, R. A. N1-
(Benzenesulfonyl)tryptamines as novel 5-HT6 antagonists. Bioorg.
Med. Chem. Lett. 2000, 10, 2295–2299.
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flip. A total of 10 genetic algorithm docking runs were performed
for each ligand structure. The ChemScore fitness function (an
estimation of the total free energy change that occurs on ligand
binding) was used to rank the docked ligand–receptor complexes.
Based on the resulting ChemScores, one of the 100 MODELER-
generated receptor models was selected for each ligand. Two such
receptors were consistently identified: one being common to ligands
lacking the sulfonamide group and another common to ligands
containing a sulfonamide moiety.
The PROTABLE facility within SYBYL and the PROCHECK
program were employed to identify potential, unusual, and sterically
unfavorable side chain geometries; these were iteratively corrected
as necessary.
All receptor/ligand complexes were subjected to short molecular
dynamics simulations using the DYNAMICS routine within
SYBYL to provide additional evidence that the GOLD-docked
poses were not artifactual (i.e., to assess the stability of the bound
pose of the ligands in the h5-HT6 receptor models). Molecular
dynamics simulations were carried out for 100 ps and snapshots
were taken every 25 fs. Otherwise, the default settings for the
DYNAMICS were employed. The energy setup was analogous to
that described for energy minimization. To maintain the integrity
of the ligand–receptor complexes, all residues except the ligand
and amino acids within an 8 Å radius of the ligand were maintained
as an aggregate; these atoms did not move during the course of the
simulation.
Acknowledgment. This work was supported, in part, by
NIMH grant MH-60599 (R.A.G.), the A. D. Williams Fund
(R.A.G.), the School of Pharmacy, Medical College of Virginia,
Virginia Commonwealth University, MH-57635 (B.L.R.), and
the NIMH Psychoactive Drug Screening Program (B.L.R.). R.K.
was supported by T32 grant DA 007027.
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A. J.; Riley, G.; Routledge, C.; Middlemiss, D. N.; Price, G. W.
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on the binding of N1-substituted tryptamines at h5-HT6 receptors.
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of G protein-coupled receptors. Endocr. ReV. 2000, 21, 90–113.
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N1-Benzenesulfonylgramine and N1-benzenesulfonylskatole: Novel
5-HT6 receptor ligand templates. Bioorg. Med. Chem. Lett. 2003, 13,
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Supporting Information Available: A table showing the root-
mean-square distances (RMSD) between the heavy atoms of
corresponding residues in the binding pocket of the two selected
receptor models [i.e., binding models for 5-HT (1) and MS-245
(4a), as shown in Figure 1; Table S1] and of amino acids within
various distances (in 0.5 Å distance increments from 3.0 to 5.0 Å)
of the docked ligands (Table S2). This material is available free of
charge via the Internet at http://pubs.acs.org.
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