Synthesis, binding affinity, and molecular docking analysis of new benzofuranone derivatives as potential antipsychotics

Article abstract of DOI:10.1021/jm800602w

The complex etiology of schizophrenia has prompted researchers to develop clozapine-related multitarget strategies to combat its symptoms. Here we describe a series of new 6-aminomethylbenzofuranones in an effort to find new chemical structures with balanced affinities for 5-HT₂ and dopamine receptors. Through biological and computational studies of 5-HT_2A and D₂ receptors, we identified the receptor serine residues S3.36 and S5.46 as the molecular keys to explaining the differences in affinity and selectivity between these new compounds for this group of receptors. Specifically, the ability of these compounds to establish one or two H-bonds with these key residues appears to explain their difference in affinity. In addition, we describe compound 2 (QF1004B) as a tool to elucidate the role of 5-HT_2C receptors in mediating antipsychotic effects and metabolic adverse events. The compound 16a (QF1018B) showed moderate to high affinities for D₂ and 5-HT_2A receptors, and a 5-HT _2A/D₂ ratio was predictive of an atypical antipsychotic profile.

Full text of DOI:10.1021/jm800602w

J. Med. Chem. 2008, 51, 6085–6094
6085
Synthesis, Binding Affinity, and Molecular Docking Analysis of New Benzofuranone Derivatives
as Potential Antipsychotics
Reyes Aranda,† Karen Villalba,† Enrique Ravin˜a,† Christian F. Masaguer,*,† Jose´ Brea,‡ Filipe Areias,‡ Eduardo Dom´ınguez,‡
Jana Selent,§ Laura Lo´pez,§ Ferran Sanz,§ Manuel Pastor,§ and Mar´ıa I. Loza‡
Departamento de Qu´ımica Orga´nica, Laboratorio de Qu´ımica Farmace´utica, and Departamento de Farmacolog´ıa, Facultad de Farmacia,
UniVersidad de Santiago de Compostela, E-15782 Santiago de Compostela, Spain, and Research Unit on Biomedical Informatics (GRIB),
IMIM, UniVersitat Pompeu Fabra, Dr. Aiguader 88, E-08003 Barcelona, Spain
ReceiVed May 21, 2008
The complex etiology of schizophrenia has prompted researchers to develop clozapine-related multitarget
strategies to combat its symptoms. Here we describe a series of new 6-aminomethylbenzofuranones in an
effort to find new chemical structures with balanced affinities for 5-HT2 and dopamine receptors. Through
biological and computational studies of 5-HT2A and D2 receptors, we identified the receptor serine residues
S3.36 and S5.46 as the molecular keys to explaining the differences in affinity and selectivity between
these new compounds for this group of receptors. Specifically, the ability of these compounds to establish
one or two H-bonds with these key residues appears to explain their difference in affinity. In addition, we
describe compound 2 (QF1004B) as a tool to elucidate the role of 5-HT2C receptors in mediating antipsychotic
effects and metabolic adverse events. The compound 16a (QF1018B) showed moderate to high affinities
for D2 and 5-HT2A receptors, and a 5-HT2A/D2 ratio was predictive of an atypical antipsychotic profile.
Introduction
pathology.11,12 Since the 1980s,13-15 a drug’s relative affinity
for the D2 and 5-HT2A receptors has been used as a good
predictive marker of an atypical antipsychotic profile, as is the
case with clozapine, risperidone, and olanzapine. In fact, the
ratio of a compound’s pKi for 5-HT2A and for D2, known as
the Meltzer ratio, is used to discriminate atypical antipsychotics
(ratio >1.12) from classical antipsychotics (<1.09).16
Experimental and clinical studies seem to confirm the major
role of the 5-HT2A receptor in the atypical profile of antipsycho-
tics.17-19 Additionally, many of the atypical antipsychotic agents
block not only 5-HT2A but also other serotonin receptors, and
the set of affinities for the 5-HT2A, 5-HT2C, 5-HT6, D2, and D3
receptors is considered to be predictive of an optimal antipsy-
chotic profile.20 Although there is no consensus on the involve-
ment of the 5-HT2C receptor in the therapeutic profile21 or in
the adverse metabolical events,11 this receptor remains a
potential target in the treatment of psychotic illnesses.22,23
Schizophrenia is a severe disorder that affects around 24
million people worldwide, and it typically begins in late
adolescence or early adulthood. It is characterized by profound
disruptions in thinking, and it affects language, perception, and
a sense of self. It often includes psychotic experiences such as
hearing voices or delusions. It can impair functioning through
the loss of the acquired capability of earning one’s own
livelihood or through the disruption of studies.1 Classical
(typical) neuroleptics such as haloperidol (Figure 1) are currently
used to treat this disease, but their use is associated with severe
mechanism-related side effects including the induction of acute
extrapyramidal symptoms (EPS)a.2 Also, these compounds are
ineffective against the negative symptoms of schizophrenia. A
common feature of the clinical effects of classical antipsychotics
is their ability to block dopamine D2 receptors in the brain.3-5
However, it has been reported that the blockage of the dopamine
receptor in the striatum is closely associated with EPS.6,7
Four decades after its introduction into the clinic, clozapine
remains the prototype for atypical antipsychotic drugs. Its
reintroduction for use in cases of treatment-resistant schizo-
phrenia gave rise to a new group of atypical or nonclassical
antipsychotics that have no EPS at therapeutic doses and are
also effective against schizophrenia’s negative symptoms.8-10
These drugs exhibit potent antagonism at multiple receptor
subtypes including serotonin and dopamine receptors, which
suggests the involvement of the serotoninergic system in this
Despite clozapine’s efficacy, it is associated with an increased
risk of agranulocytosis,24 which strongly limits its therapeutic
use. However, none of the other currently available drugs appear
to have clozapine’s spectrum of efficacy. Therefore, the
discovery of novel antipsychotic agents that are chemically
different, more effective, and free of side effects remains a
challenging research goal.
As a part of our ongoing work in developing multitarget
ligands for the potential use as treatments for schizophrenia,
we have studied the modulation of the butyrophenone system
with the aim of synthesizing a single molecule that can
antagonize receptors of both the 5-HT2 and the D2 families.25-27
So far, we have reported the synthesis, pharmacology, and 3D-
QSAR analysis of a number of aminoalkyl-1-benzofuranones
as potential antipsychotics.28 This study introduced the ami-
nobutyrophenones 1-4 (substructure in bold), which show
different pharmacological profiles (Table 1), and 6-aminom-
ethylbenzofuranones 1 and 2, which exhibit a classical antip-
sychotic profile, whereas 5-aminoethylbenzofuranones 3 and 4
may display an atypical antipsychotic profile.
* To whom correspondence should be addressed. Tel: +34 981 563 100.
Fax: +34 981 594 912. E-mail: qojcfm@usc.es.
†
Departamento de Qu´ımica Orga´nica, Universidad de Santiago de
Compostela.
‡
Departamento de Farmacolog´ıa. Universidad de Santiago de Compostela.
Universitat Pompeu Fabra.
§
a Abbreviations: EPS, extrapyramidal symptoms; 3D-QSAR, three-
dimensional quantitative structure-activity relationship; PF, Pseudomonas
fluorescens; THF, tetrahydrofuran; TFAA, trifluoroacetic anhydride; TFA,
trifluoroacetic acid; DCC, dicyclohexylcarbodiimide; HOBt, 1-hydroxy-
benzotriazole; FB, 4-fluorobenzoyl; BI, 6-fluorobenzisoxazolyl; SAR,
structure-activity relationship; GPCR, G protein-coupled receptor.
10.1021/jm800602w CCC: $40.75
2008 American Chemical Society
Published on Web 09/11/2008

6086 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19
Aranda et al.
route from this precursor to the hydroxymethylbenzofuranone
5 is depicted in Scheme 3. The carboxylic acid (S)-8 was
reduced to the alcohol (S)-9 by the use of a borane dimethyl
sulfide complex in THF at 0 °C. Subsequent acid-catalyzed
cyclization yielded the lactone (S)-10, which was treated with
HBr in acetic acid. The resulting bromoacid was cyclized to
produce the bromobenzofuranone (+)-7 with the S absolute
configuration.
New compounds that bear different substituents on the furane
ring were prepared as illustrated in Schemes 3 and 4. Different
strategies of furannulation (Scheme 4) were applied to our
previously described synthon 12, which was generated in situ
from bis-enolether 11 by acid hydrolysis32 to obtain benzofura-
none systems with different substituents.
Therefore, the condensation of diketone 12 with ethyl
2-bromopyruvate in methanolic KOH produced (6-hydroxym-
ethyl-4-oxo-4,5,6,7-tetrahydrobenzofuran-3-yl)carboxylic acid
(13a) as a yellow solid in 40% yield. This was esterified with
diazomethane to yield 13d quantitatively.
Figure 1. Structures of a typical (haloperidol) and some atypical
antipsychotics.
Compounds 2 (QF1004B) and 4 (QF0703B) are closely
structurally related and have affinities for the 5-HT2A and D2
receptors, but their Meltzer ratios vary from 1.24 in the case of
compound 4, which is characteristic of an atypical antipsychotic,
to 1.06 in the case of 2, which is characteristic of a typical
antipsychotic. These two compounds have been characterized
as interesting pharmacological tools for the study of how a
compound’s antipsychotic profile is influenced by different
selectivities for the 5-HT2 receptor subtypes (the potency of
compound 4 at the 5-HT2C receptor is approximately 10 times
(1 log unit) higher than that of compound 2), by different
5-HT2A/D2 ratios (Meltzer ratios), and by different conformation-
dependent functional behaviors at the 5-HT2A and 5-HT2C
receptors.29
To explore the therapeutic potential of the aminoalkylben-
zofuranone cores of 1-4 further, we describe in this article the
synthesis and binding affinity of a novel series of 5-HT2/D2
ligands. To investigate the mode of receptor binding of these
new compounds and to gain an understanding of the structural
and conformational features that underlie their pharmacological
properties, we docked some compounds into homology models
of the dopamine D2 and serotonin 5-HT2A receptors. The detailed
analysis of the obtained complexes provides an excellent
explanation of some of the largest differences in their binding
affinities and thus validates the potential use of such complexes
in the design of novel compounds.
Chemistry. The synthesis of the optically pure aminobuty-
rophenones (+)- and (-)-1 and (+)- and (-)-2 was ac-
complished by a lipase-catalyzed kinetic resolution of the
intermediate 6-hydroxymethyl-4,5,6,7-tetrahydrobenzofuran-4-
one ((()-5)30 that used lipase from Pseudomonas fluorescens
(PF) that was adsorbed on Celite in benzene (Scheme 1). This
yielded the hydroxyketone (-)-5 (98% ee) and the acetate (+)-6
(97% ee). Subsequent ester hydrolysis of (+)-6 with LiOH
produced the corresponding hydroxymethylbenzofuranone (+)-5
in 78% yield. The bromination of the enantiomerically pure
alcohols (Scheme 2), followed by the nucleophilic displacement
of the bromine atom with 4-(p-fluorobenzoyl)piperidine, af-
forded (-)-1 and (+)-1, whereas the analogous reaction with
4-(6-fluorobenzisoxazol-3-yl)piperidine gave (-)-2 and (+)-2.
The hydrochloride salts of these compounds prove to be suitable
for use in binding assays.
The manganese(III) acetate-mediated addition33,34 of diketone
12 to phenylacetylene in glacial acetic acid afforded the desired
phenylbenzofuranone 13b in 30% yield. Likewise, by following
the method described by Stetter and Lauterbach,35 we cyclo-
condensed 12 with ethyl 2-chloroacetoacetate in the presence
of KOH to produce the ester 13c in 40% yield.(Scheme 5).
The tosylation of alcohols 13a-c with p-toluenesulfonyl
chloride in anhydrous pyridine afforded the corresponding
sulfonates 14a-c in 50-65% yield (Scheme 4). The tosylates
underwent nucleophilic substitution with 4-(p-fluorobenzoyl)pi-
peridine or 4-(6-fluorobenzisoxazol-3-yl)piperidine to give the
desired amines 15a-c and 16a-c, respectively. Amide 18 was
prepared in 45% yield from piperazine derivative 1730 by acid-
amine coupling with DCC carboxylate activation in the presence
of HOBt (Scheme 6).
Results and Discussion
In some related compounds that were previously published
by our group, we observed that the differences in the binding
affinities of the R and S enantiomers are not large but they are
certainly not negligible.36 Therefore, our first objective in the
present study was to investigate whether the receptor affinities
of these compounds correlate with the absolute stereochemistry.
For this purpose, we prepared the benzofuranone derivative 2
as single enantiomers and determined their binding affinities
for the D2, 5-HT2A, and 5-HT2C receptors. The affinities of the
two enantiomers for cloned human D2, 5-HT2A, and 5-HT2C
receptors were evaluated by in vitro binding assays, and the
data are summarized in Table 2.
As shown in Table 2, the individual enantiomers and their
racemic mixture show nearly identical affinities for the 5-HT2A,
5-HT2C, and D2 receptors. Consequently, it is not possible to
correlate the absolute configuration with the potency in any of
the receptors that were studied here.
The structures of the complexes that were obtained in the
molecular modeling studies for the enantiomers of compound
2 fully support this finding. As expected, the piperidinylmethyl
substituent always adopts an equatorial position, whereas the
hydrogen that is attached to the chiral center adopts a pseudo-
axial orientation in which it points toward different sides of
the binding site depending on the configuration. These structural
differences do not significantly affect the binding behavior, and
as shown in Figure 2, both enantiomers can adopt nearly
identical orientations within the receptor and thus establish the
same polar interactions.
The exact stereochemistry of these enantiomers could not be
determined by crystallographic analysis. The absolute config-
uration was therefore assigned on the basis of the known
configuration of the optically pure precursor (S)-4-(tert-butoxy)-
2-(furan-2-ylmethyl)-4-oxobutanoic acid (8).31 The synthetic

Analysis of New Benzofuranone DeriVatiVes
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19 6087
Table 1. Binding Profile of Compounds 1-4 for Dopamine D2 and Serotonins 5-HT2A and 5-HT2C Human Receptorsa
compd
pKi D2
pKi 5-HT2A
pKi 5-HT2C
pA2 5-HT2A
MRb
1 (QF1003B)
2 (QF1004B)
3 (QF0702B)
4 (QF0703B)
6.82 ( 0.17
7.79 ( 0.18
6.14 ( 0.31
7.34 ( 0.13
7.02 ( 0.14
8.26 ( 0.12
7.11 ( 0.20
9.14 ( 0.11
5.81 ( 0.21
6.47 ( 0.26
6.47 ( 0.18
7.46 ( 0.30
7.28 ( 0.07
7.95 ( 0.07
7.58 ( 0.20
9.25 ( 0.05
1.03
1.06
1.16
1.24
a Data are expressed as mean ( SEM of three experiments. b MR ) Meltzer ratio (pKi 5-HT2A/pKi D2).
Scheme 1a
Scheme 4a
a Reagents and conditions: (i) vinyl acetate, lipase from PF, benzene, rt,
70 h and (ii) LiOH, DME, reflux, 17 h, 78%.
Scheme 2a
a Reagents and conditions: (i) CBr4, PPh3, CH2Cl2, reflux, 28 h, 95%
and (ii) HNRR, NMP, 85 °C, 16 h, 62-70%.
a Reagents and conditions: (i) HCl, THF, rt,
4
h, 96%; (ii)
BrCH2COCOOC2H5, KOH, MeOH, 12 h, 55%; (iii) (CH3COO)3Mn ·2H2O,
PhC′CH, AcOH, from 80 to 45 °C, 2 h, 30%; (iv) 2-chloroacetoacetate,
KOH, MeOH, H2O, rt, 36 h, 40%; and (v) CH2N2, MeOH, Et2O, rt, 12 h,
97%.
Scheme 3a
Scheme 5a
a Reagents and conditions: (i) (CH3)2S·BH3, THF, from 0 °C to rt, 2 h,
86%; (ii) p-TsOH, benzene, reflux, 3 h, 76%; and (iii) (i) HBr, AcOH, 80
°C, 6 h and (ii) TFAA, TFA, rt, 12 h, 10% overall.
Because the stereochemistry of the chiral center does not seem
to determine the affinities of these compounds for the target
receptors, we focused our studies on the benzofuranone core.
Therefore, several new compounds (15a-c and 16a-c) were
prepared with substituents on the furan ring. The binding
affinities of these new aminoalkylbenzofuranone derivatives at
the dopamine D2 and serotonin 5-HT2A and 5-HT2C receptors
are summarized in Table 3.
Table 3 shows that the binding affinities of all of the
derivatives that bear the p-fluorobenzoyl (FB) moiety (15a-c)
are consistently lower than those of the compounds with the
6-fluorobenzisoxazolyl (BI) fragment (16a-c). For the 5-HT2A
receptor, the difference in the affinities of the two series is of
a Reagents and conditions: (i) TsCl, Py, from 0 °C to rt, 24 h, 45-63%;
(ii) 4-(4-fluorobenzoyl)piperidine, CH3CN, reflux, 22 h, 25-45%; and (iii)
4-(6-fluorobenzisoxazol-3-yl)piperidine, CH3CN, reflux, 24 h, 30-84%.
approximately 1 order of magnitude, and the effect is even more
remarkable for the D2 receptor, where the FB compounds show
no affinity. The compounds can be ranked according to their
binding affinities as follows: BI-5-HT2A > BI-D2 and FB-5-
HT2A > FB-D2.
A close analysis of the obtained complexes of our modeling
studies suggests an interesting explanation of their binding-

6088 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19
Aranda et al.
Scheme 6a
This raises the question of why the same affinities are not
observed for compounds 1 and 2. Both are unsubstituted at the
furan ring and are free of the bulky groups that are present for
15a-c and 16a-c. Consequently, they are free to bind the
receptors in an alternative orientation that interchanges the
position of the molecule’s ends at the binding site; in this
alternative orientation, the interactions with the 5-HT2A and D2
receptors are weaker. These results were confirmed by docking
simulations, which yielded docking solutions in both orientations
for only the unsubstituted compounds. The presence of two
alternative binding modes for this series of compounds has been
previously reported,36,41 and it is consistent with both molecular
modeling and experimental results.
The 4-(p-fluorobenzoyl)piperidine fragment has been de-
scribed as an antipsychotic pharmacophore with a potency that
is similar to that of linear butyrophenones,42 and a SAR study
of ketanserin analogs has suggested that the carbonyl of the
benzoyl moiety (present in 1, 3, and 15a-c) may be important
for the anchoraging or the orientation in the 5-HT2A receptor’s
binding site.43 To determine whether this carbonyl’s identity
as a ketone group is essential for binding, we synthesized an
amide analog. The resultant benzoylpiperazine derivative 18
(Table 4) bound to the 5-HT2A receptor with 10-fold lower
affinity than that for analog 1. Furthermore, the substitution of
the piperidine ring with a pirerazine ring in these compounds
drastically reduces the binding affinity for both dopamine D2
and serotonin 5-HT2C receptors.
a Reagents and conditions: 4-fluorobenzoic acid, DCC, HOBt, CH2Cl2,
from 0 °C to rt, 45%.
Table 2. Binding Profile of the Individual Enantiomers and the Racemic
Mixture of 2 for the Dopamine D2 and Serotonins 5-HT2A and 5-HT2C
Receptorsa
compd
pKi D2
pKi 5-HT2A
pKi 5-HT2C
(+)-2
(-)-2
(()-2
7.38 ( 0.15
7.80 ( 0.10
7.79 ( 0.18
8.60 ( 0.22
8.29 ( 0.15
8.26 ( 0.12
6.61 ( 0.15
6.43 ( 0.22
6.47 ( 0.26
a Data are expressed as mean ( SEM of two experiments.
Again, the experimental results may be rationalized on the
basis of the structures of the complexes that were obtained by
modeling studies. In compound 18, the carbonyl oxygen is part
of the planar amide group that orients this oxygen in a way
that prevents it from interacting with the aforementioned S3.36
residue. Figure 4 shows a superposition of compounds 1 and
18 bound to the 5-HT2A receptor. The superposition reveals the
different abilities of the two compounds to interact with S3.36.
Figure 2. Superimposition of the docking solutions for the R and S
enantiomers of compound 2 over the binding site of the 5-HT2A receptor.
Conclusions
affinity rankings. As shown in Figure 3, the main polar
interactions between the ligand and the receptor at this part of
the binding pocket are established between the H-bond acceptor
atoms of the FB or BI moieties and the H-bond donor side chains
of the amino acid residues at positions 3.36 and 5.46. The BI
moiety contains two H-bond acceptors, and it can therefore
establish two H-bonds with the 5-HT2A receptor (S3.36 and
S5.46). This proposed forked interaction of the BI moiety that
is simutaneously mediated by the nitrogen and oxygen heteroa-
toms is supported by experimental structural data.37-39 In the
D2 receptor, however, the serine at position 3.36 is replaced by
a cysteine, and the ligand can establish only one strong H-bond.
The importance of these residues in the binding of antipsychotics
has been shown in a previous recent study.40 The compounds
of the FB series have only one H-bond acceptor group that is
oriented toward residue 3.36, which limits them to establishing
a single H-bond with the 5-HT2A receptor and no strong H-bonds
with the D2 receptor. In addition, the entropic cost of fixing the
more flexible FB moiety will probably contribute to the further
reduction of the binding affinities of all of the compounds in
this series. Overall, these differences in hydrogen bonding can
explain the previously mentioned ranking of binding affinity:
BI-5-HT2A (two H-bonds) > BI-D2, and FB-5-HT2A (one
H-bond) > FB-D2 (no H-bonds). It should be emphasized that
the quality of our structural models allows only a qualitative
interpretation of the observed differences and not a complete
quantitative description of individual binding affinities.
Butyrophenone analogs with benzofuranone cores were
prepared and examined here for their affinities and selectivities
as 5-HT2A/D2 dual ligands. The SAR study focused on the
chirality and substitutions of the piperidine ring and the
benzofuranone core. The use of almost enantiopure compounds
indicated that the chirality of the compounds in this series does
not influence their affinity or the selectivity for the studied
receptors. This finding is supported by both experimental and
molecular docking studies. For example, the docking studies
showed that both enantiomers can establish the same polar
interactions in the receptor binding site. Compound 16a shows
the greatest potential for antipsychotic activity. It binds with
moderate-to-high affinities for the D2 and 5-HT2A receptors with
an appropriate affinity ratio at these two receptors for atypicality;
further work is needed to assess its effect in animal models of
schizophrenia. Because they show a range of affinity for this
receptor, these compounds stand out as useful pharmacological
tools in the elucidation of the role of the 5-HT2C receptor in
antipsychotic efficacy and metabolic side effects.
Apart from the usefulness of the new compounds, the
modeling reported here provides a rationalization for the
extreme differences in the binding affinities observed in some
compounds with respect to the 5-HT2A and D2 receptors.
These differences seem to be related to the presence of either
the 4-benzoylpiperidine (15 series) or the 4-benzisoxazolylpi-
peridine (16 series) moieties. The differences can be ex-
plained in terms of their abilities to establish key polar

Analysis of New Benzofuranone DeriVatiVes
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19 6089
Table 3. Binding Profile of the Aminoalkylbenzofuranone Derivatives 15a-c, 16a-c (Scheme 2), 1, and 2 and Reference Compounds at the Dopamine
D2 and Serotonin 5-HT2A and 5-HT2C Receptorsa
compd
R1
R2
pKi D2
pKi 5-HT2A
pKi 5-HT2C
MRb
15a (QF1014B)
15b (QF1064B)
15c (QF1034B)
16a (QF1018B)
16b (QF1068B)
16c (QF1038B)
1 (QF1003B)
2 (QF1004B)
haloperidol
H
Ph
CO2Et
H
Ph
CO2Et
H
H
CO2Me
H
<5
7.59 ( 0.23
7.76 ( 0.27
7.66 ( 0.14
8.81 ( 0.22
8.16 ( 0.18
8.08 ( 0.50
7.02 ( 0.14
8.26 ( 0.12
6.78 ( 0.25
8.04 ( 0.31
<5
<5
<5
Me
CO2Me
H
Me
H
<5
<5
7.07 ( 0.08
7.92 ( 0.15
6.67 ( 0.14
6.82 ( 0.17
7.79 ( 0.18
9.22 ( 0.12
6.65 ( 0.17
<5
1.25
1.03
1.21
1.03
1.06
0.73
1.21
5.48
<5
5.81 ( 0.21
6.47 ( 0.26
5.14 ( 0.18
7.98 ( 0.11
H
clozapine
a Data are expressed as mean ( SEM of two experiments. b MR ) Meltzer ratio (pKi 5-HT2A/pKi D2).
Figure 3. Structures of the D2 and 5-HT2A receptors complexed with compounds bearing p-fluorobenzoyl (FB) and 6-fluorobenzisoxazolyl (BI)
moieties. FB compounds cannot establish strong H-bonds with the residues at positions 3.36 and 5.46 of the D2 receptor, and they can establish
only one H-bond with the same residues of the 5-HT2A receptor. In contrast, the BI compounds can establish one H-bond and two H-bonds with
the same receptors, respectively. See the text for details.
Table 4. Effects the Substitution of the Benzoylpiperidine Moiety with
a Benzoylpiperazine (1 vs 18) on Receptor Binding Affinitya
for the study of binding sites on these and other receptors. This
research may lead to the design of new chemical agents that
show an optimal affinity profile and that avoid the adverse side
effects that are associated with the dibenzodiazepine structure
of clozapine.
compd
pKi D2
pKi 5-HT2A
pKi 5-HT2C
5.81 ( 0.21
<5
1 (QF1003B)
18 (QF1012B)
6.82 ( 0.17
7.02 ( 0.14
5.88 ( 0.46
<5
a Data are expressed as mean ( SEM of two experiments.
Experimental Section
interactions with the residues at positions 3.36 and 5.46 in
both receptors. This hypothesis deserves further experimental
confirmation by synthesizing and testing derivatives with
alternative H-bond patterns that confirm the importance of
these residues in the overall ligand selectivity. The finding
presented in this work may prove useful in the future design
of selective compounds for either the previously mentioned
receptors or the other receptors involved in the treatment of
schizophrenia. The fact that our modeling studies concur with
experimental work in explaining binding selectivity as well
as other minor findings (such as the low activity of compound
18) argues for the accuracy and usefulness of our homology
modeling approach.
Chemistry. Melting points were determined with a Kofler hot
stage instrument or a Gallenkamp capillary melting point apparatus
and are uncorrected. Infrared spectra were recorded with a Perkin-
Elmer 1600 FTIR spectrophotometer; the main bands are given in
cm-1 1H NMR spectra were recorded on a Bruker WM AMX
.
(300 MHz) spectrometer; chemical shifts were recorded in parts
per million (δ) downfield from tetramethylsilane (TMS). Mass
spectra were performed on a Kratos MS-50 or a Varian Mat-711
mass spectrometer by chemical ionization (CI) or by electron impact
(EI) methods. We performed flash column chromatography by using
Kieselgel 60 (60-200 mesh, E. Merck AG, Darmstadt, Germany).
We monitored the reactions by thin layer chromatography (TLC)
on Merck 60 GF254 chromatogram sheets using iodine vapor, UV
light, or both for detection. Unless otherwise stated, each purified
compound showed a single spot. HPLC analysis was performed
on a Waters HPLC system that consisted of a vacuum degasser, a
binary pump, a column compartment, and a UV detector (254 and
330 nm). Chiral HPLC was carried out at room temperature on a
Chiralcel OD-H column (Daicel Chemical Industries, Japan).
On the basis of our present study, our aminomethylbenzo-
furanones have provided new insights into the binding mode
of ligands to the D2 and 5-HT2A receptors and have thereby
contributed to this very active research area. Work is underway
to develop these compounds further into pharmacological tools

6090 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19
Aranda et al.
and water, the organic phase was dried (Na2SO4), and the solvent
was distilled off. Purification of the residue by column chroma-
tography with 1:2 AcOEt/hexane as the eluent gave the title
compound (210 mg, 95%) as a white solid. mp 59-60 °C. IR: 3139,
1
1675, 1450, 1121. H NMR (CDCl3, δ): 2.50 (dd, 1H, J ) 16.3,
11.5 Hz, H5), 2.61-2.76 (m, 2H, H5, H6), 2.85 (dd, 1H, J ) 16.7,
10.0 Hz, H7), 3.14 (dd, 1H, J ) 16.8, 5.1 Hz, H7), 6.68 (d, 1H, J
) 2.0 Hz, H3), 7.35 (d, 1H, J ) 2.0 Hz, H2). CIMS m/z: 229 (MH+),
231 [(MH + 2)+]. [R]2D5 ) +43.5 (c 2.0, CHCl3). 97% ee. Chiralcel
OD-H; hexane/2-propanol, 93:7; flow, 0.4 mL/min; λ ) 254 nm;
tR ) 74.5 min.
(R)-6-(Bromomethyl)-4,5,6,7-tetrahydro-1-benzofuran-4-one, (R)-
7. The title compound was prepared according to the above
procedure, starting from the alcohol (R)-5. [R]2D5 ) -44.6 (c 0.7,
CHCl3). 99% ee. Chiralcel OD-H; hexane/2-propanol, 93:7; flow,
0.4 mL/min; λ ) 254 nm; tR ) 70.5 min.
General Procedure for the Preparation of 6-(Aminomethyl)-
4,5,6,7-tetrahidro-1-benzofuran-4-ones (R)- and (S)-1-2. To a
solution of (R)- or (S)-bromobenzofuranone 7 (100 mg, 0.44 mmol)
and the appropriate amine (0.90 mmol) was stirred in N-methyl-
2-pyrrolidone (5 mL) for 16 h at 85 °C. The solvent was removed
in vacuo, and the residue was dissolved in CH2Cl2. This solution
was washed twice with water and was dried (Na2SO4), and the
solvent was removed in vacuo, which afforded an orange solid that
upon column chromatography with AcOEt as the eluent gave the
final amine as a white crystalline solid. The spectroscopic data were
identical to those of the racemic compounds.30
Figure 4. Modeled complexes of receptor 5-HT2A with compounds 1
(carbons in green) and 18 (carbons in pink). The planar amide group
present in 18 prevents any H-bonding between the amide carbonilic
oxygen and S3.36, such as the H-bond shown here for 1.
Elemental combustion analyses were performed by the Microanaly-
sis Service of the University of Santiago de Compostela on a Perkin-
Elmer 240B apparatus. Unless otherwise stated, all reported values
were within (0.4% of the theoretical compositions. Solvents were
purified by distillation over an appropriate drying agent under an
argon atmosphere and were immediately used. Diazomethane
solution was prepared according to Vogel.44
(R)-6-{[4-(p-Fluorobenzoyl)piperidin-1-yl]methyl}-4,5,6,7-tet-
rahydro-1-benzofuran-4-one, (R)-1. The title compound was pre-
pared in 70% yield according to the general procedure using 4-(p-
fluorobenzoyl)piperidine as the amine. [R]2D5 ) -37.9 (c 0.68,
AcOEt). Hydrochloride: Anal. Calcd (C21H22FNO3 ·HCl·1.5H2O):
C, H, N.
(S)-6-{[4-(p-Fluorobenzoyl)piperidin-1-yl]methyl}-4,5,6,7-tet-
rahydro-1-benzofuran-4-one, (S)-1. The title compound was pre-
pared in 65% yield according to the general procedure using 4-(p-
fluorobenzoyl)piperidine as the amine. [R]2D5 ) +33.4 (c 0.40,
AcOEt). Hydrochloride: Anal. Calcd (C21H22FNO3 ·HCl·0.6H2O):
C, H, N.
(R)-6-{[4-(6-Fluorobenzisoxazol-3-yl)piperidin-1-yl]meth-
yl}-4,5,6,7-tetrahydro-1-benzofuran-4-one, (R)-2. The title com-
pound was prepared in 62% yield according to the general procedure
using 4-(6-fluorobenzisoxazol-3-yl)piperidine as the amine. [R]D25
) -31.6 (c 1.65, AcOEt). Hydrochloride: Anal. Calcd (C21H21FN2-
O3 ·HCl·0.2H2O): C, H, N.
(S)-6-{[4-(6-Fluorobenzisoxazol-3-yl)piperidin-1-yl]methyl}-4,5,6,7-
tetrahydro-1-benzofuran-4-one, (S)-2. The title compound was
prepared in 65% yield according to the general procedure using
4-(6-fluorobenzisoxazol-3-yl)piperidine as the amine. [R]2D5 ) +29.6
(c 0.46, AcOEt). Hydrochloride: Anal. Calcd (C21H21FN2O3 ·2H-
Cl·0.1H2O): C, H, N.
tert-Butyl-(S)-4-(furan-2-yl)-3-(hydroxymethyl)butanoate, (S)-
9. To a solution of hemiester (S)-8 (102 mg, 0.4 mmol) in anhydrous
THF (5 mL) cooled in an ice bath was added borane dimethyl
sulfide complex (0.058 mL, 0.61 mmol) dropwise. The reaction
mixture was stirred at 0 °C for 15 min and then at room temperature
for 2 h. After the mixture was cooling again in an ice bath, methanol
was added dropwise until effervescence ceased, and then the mixture
was stirred at room temperature for 1 h. The solvent was then
evaporated under vacuum, and the crude oil was purified by column
chromatography with 8:1 AcOEt/hexane as the eluent to give the
title compound (84 mg, 86%) as a yellowish oil. IR: 3424, 2976,
1724, 1151. 1H NMR (CDCl3, δ): 1.44 (s, 9H, C(CH3)3), 2.27-2.71
(m, 3H, H2, H3), 2.63-2.73 (m, 2H, furan-CH2), 3.53 (dd, 1H, J )
11.0, 5.4 Hz, -HCH-OH), 3.61 (dd, 1H, J ) 11.1, 4.7 Hz,
-HCH-OH), 6.04 (d, 1H, J ) 3.1 Hz, H3 furan), 6.28 (dd, 1H, J
) 3.0, 1.9 Hz, H4 furan), 7.30 (d, 1H, J ) 1.5 Hz, H5 furan). CIMS
m/z: 241 (MH+). [R]D25 ) +1.32 (c 2.8; CH2Cl2).
Enzymatic Resolution of (()-6-(Hydroxymethyl)-4,5,6,7-tet-
rahydro-1-benzofuran-4-one ((()-5). To a solution of the hy-
droxymethylbenzofuranone (()-530 (226 mg, 1.35 mmol) in
benzene (12 mL) were added vinyl acetate (82 µL, 0.84 mmol)
and PF lipase on Celite (136 mg). The mixture was stirred for 70 h
at room temperature, was filtered through Celite, and was concen-
trated in vacuo. The residue was purified by column chromatog-
raphy with 1:3 AcOEt/hexane as the eluent to give (R)-3-
hydroxymethylbenzofuranone ((-)-5) (68 mg, 30%) and the (S)-
acetate (+)-6 (62 mg, 22%) as a yellowish oil.
(R)-6-(Hydroxymethyl)-4,5,6,7-tetrahydro-1-benzofuran-4-
one, (-)-5. The spectroscopic data were identical to those of the
racemic compound.30 [R]D20 ) -45.0 (c 2.5, AcOEt). 99% ee.
Chiralcel OD-H; hexane/2-propanol, 93:7; flow, 0.4 mL/min; λ )
254 nm; tR ) 99.6 min.
(S)-(4-Oxo-4,5,6,7-tetrahydro-1-benzofuran-6-yl)methyl Acetate,
(+)-6. IR: 1733, 1683, 1246, 1039. 1H NMR (δ): 2.08 (s, 3H, CH3),
2.34-2.43 (m, 1H, H5), 2.58-2.78 (m, 3H, 1H5, H6, 1H7),
2.98-3.08 (m, 1H, 1H7), 4.09-4.17 (m, 2H, CH2-OAc), 6.67 (d,
1H, J ) 1.9 Hz, H3), 7.34 (d, 1H, J ) 1.8 Hz, H2). EIMS m/z: 208
(M+). [R]2D5 ) +41.7 (c 2.6, AcOEt). 99% ee. Chiralcel OD-H;
hexane/2-propanol, 93:7; flow, 0.4 mL/min; λ ) 254 nm; tR ) 78.1
min.
(S)-6-(Hydroxymethyl)-4,5,6,7-tetrahydro-1-benzofuran-4-one (+)-
5. To a solution of acetate (S)-(+)-6 (188 mg, 0.90 mmol) in
dimethoxyethane (10 mL) was added 1 N aqueous LiOH (2.26 mL,
2.26 mmol). The mixture was refluxed for 13 h, 1 N aqueous LiOH
(1.36 mL, 1.36 mmol) was added, and the mixture was heated for
4 h. After cooling to room temperature, the reaction mixture was
diluted with Et2O (25 mL) and acidified with 1 N HCl. The organic
phase was separated, and the aqueous layer was extracted with ether.
The combined organic extracts were dried (Na2SO4) and concen-
trated in vacuo to give a residue that was purified by column
chromatography with 4:1 AcOEt/hexane as the eluent. The title
compound (117 mg, 78%) was yielded as a white solid. The
spectroscopic data were identical to those of the racemic com-
pound.30 [R]2D5 ) +45.1 (c 2.4; AcOEt). 99% ee. Chiralcel OD-H;
hexane/2-propanol, 93:7; flow, 0.4 mL/min; λ ) 254 nm; tR ) 94.1
min.
(S)-6-(Bromomethyl)-4,5,6,7-tetrahydro-1-benzofuran-4-one, (S)-
7. A stirred solution of alcohol (S)-5 (160 mg, 0.96 mmol), carbon
tetrabromide (400 g, 1.2 mmol), and triphenyl phosphine (380 mg,
1.44 mmol) in CH2Cl2 (10 mL) was refluxed under argon for 28 h.
After cooling, the reaction mixture was washed with 5% NaHCO3
(S)-4-[(2-Furyl)methyl]-2,3,4,5-tetrahydrofuran-2-one, (S)-10. A
solution of the hydroxyester (S)-9 (1.0 g, 4.16 mmol) and p-TsOH
(catalytic) in benzene (25 mL) was refluxed under argon for 3 h.

Analysis of New Benzofuranone DeriVatiVes
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19 6091
After cooling to room temperature, the solution was washed with
10% NaHCO3 and water. Aqueous phases were extracted with
AcOEt, and the combined organic extracts were dried (Na2SO4)
and concentrated in vacuo. The residue was purified by column
chromatography with 5% MeOH/CH2Cl2 as the eluent to give the
title compound (0.53 g, 76%) as a yellowish oil. IR: 1775, 1170,
1008. 1H NMR (CDCl3, δ): 2.31 (dd, 1H, J ) 17.6, 6.8 Hz, 1H3),
2.65 (dd, 1H, J ) 17.6, 8.2 Hz, 1H3), 2.79 (d, 2H, J ) 7.3 Hz,
furyl-CH2-), 2.84-2.96 (m, 1H, H4), 4.07 (dd, 1H, J ) 9.2, 6.0
Hz, 1H5), 4.40 (dd, 1H, J ) 9.2, 7.1 Hz, 1H5), 6.06 (d, 1H, J ) 3.3
Hz, H3 furan), 6.29 (dd, 1H, J ) 3.2, 1.9 Hz, H4 furan), 7.30 (d,
1H, J ) 1.7 Hz, H5 furan). EIMS m/z: 166 (M+). [R]D24) -5.76 (c
1.15; CH2Cl2).
(S)-6-(Bromomethyl)-4,5,6,7-tetrahydro-1-benzofuran-4-one, (S)-
7. To a solution of lactone (S)-10 (0.59 g, 3.05 mmol) in glacial
acetic acid (7.5 mL) cooled in an ice bath was added a 33% solution
of HBr in acetic acid (1 mL, 4.57 mmol). The mixture was heated
to 80 °C and was stirred at this temperature for 6 h. After cooling
to room temperature, the mixture was poured onto crushed ice and
extracted with CH2Cl2. The organic extracts were washed with
water, were dried (Na2SO4), and were concentrated under reduced
pressure to afford crude (S)-3-(bromomethyl)-4-(2-furyl)butanoic
acid (0.77 g) as a yellowish oil, which was used without further
purification. IR: 2924, 1710, 1430, 1250. IEMS m/z: 246 (M+),
248 [(M + 2)+]. A mixture of this compound (380 mg, 1.53 mmol),
trifluoroacetic anhydride (8.3 mL), and trifluoroacetic acid (1.6 mL)
was stirred under an argon atmosphere overnight at rt. The brown
mixture was then poured onto crushed ice and extracted with
CH2Cl2. The organic extracts were washed successively with ice-
cold 5% sodium hydroxide solution and brine and were then dried
(Na2SO4) and concentrated. The crude material obtained was
purified by column chromatography with 1:2 EtOAc/hexane as the
eluent to give the title compound (35 mg, 10%) as a beige solid.
Spectroscopic data were identical to those of the aforementioned
compound. [R]2D5) +38 (c 0.27, CHCl3). 85% ee. Chiralcel OD-
H; hexane/2-propanol, 93:7; flow, 0.4 mL/min; λ ) 254 nm; tR)
75.6 min.
to dryness. The crude product was purified by column chromatog-
raphy with 1:1 AcOEt/hexane as the eluent to afford the title
compound (403 mg, 30%) as a light-brown solid. mp 131-132 °C
(toluene). IR: 3362, 1650. 1H NMR (CDCl3,δ): 2.38-2.48 (m, 1H,
1H5), 2.59-2.65 (m, 2H, H6, 1H5), 2.87 (dd, 1H, J ) 17.2, 9.4 Hz,
1H7), 3.02 (dd, 1H, J ) 17.3, 4.8 Hz, 1H7), 3.72-3.74 (m, 2H,
-CH2OH), 6.88 (s, 1H, H3), 7.30 (t, 1H, J ) 7.3 Hz, H4 Ph), 7.40
(dd, 2H, J ) 8.0, 6.9 Hz, H3+H5 Ph), 7.65 (d, 2H, J ) 8.0 Hz,
H2+H6 Ph). EIMS m/z): 242 (M+). Anal. Calcd (C15H14O3 ·
0.15H2O): C, H.
Ethyl 6-(Hydroxymethyl)-3-methyl-4-oxo-4,5,6,7-tetrahydro-1-
benzofuran-2-carboxylate (13c). A solution of 12 (0.60 g, 4.2 mmol)
and ethyl 2-chloroacetoacetate (0.64 mL, 4.64 mmol) in MeOH
(2.3 mL) was added with stirring to a mixture of KOH (0.39 g, 6.9
mmol) and water (14 mL). Stirring was continued at room
temperature for 36 h, and H2O (9 mL) was added, followed by
enough 10% HCl to bring the mixture to pH 3. After being stirred
for 1 h, the mixture was extracted with ether and the organic layer
was washed with 5% NaHCO3, was dried (Na2SO4), and was
concentrated to dryness, affording 0.26 g of an oil that, upon column
chromatography with 1:1 AcOEt/hexane as the eluent, gave 0.16 g
of a white solid that was identified as the ester 13c. From the basic
aqueous phase, more ester 13c (0.23 g) crystallized and was
collected by filtration in 40% yield. mp 81-83 °C. IR: 3380, 1685,
1
1600. H NMR (CDCl3, δ): 1.39 (t, 3H, J ) 7.1 Hz, -CH2CH3),
2.41-2.57 (m, 3H, H6 + H5), 2.55 (s, 3H, -CH3), 2.82 (dd, 1H, J
) 17.5, 9.3 Hz, 1H7), 3.06 (dd, 1H, J ) 17.9, 5.3 Hz, 1H7),
3.60-3.80 (m, 2H, C H2OH), 4.38 (q, 2H, J ) 7.1 Hz, -CH2CH3).
EIMS m/z: 252 (M+). Anal. Calcd (C13H16O5): C, H.
Methyl 6-(Hydroxymethyl)-4-oxo-4,5,6,7-tetrahydro-1-benzo-
furan-3-carboxylate (13d). To a solution of the acid 13a (0.50 g,
2.38 mmol) in anhydrous methanol (20 mL), an excess amount of
a diazomethane solution was added, followed by agitation at room
temperature overnight. The solution was then concentrated to
dryness to afford the title compound (0.52 g, 97%) as a yellowish
oil that crystallized upon standing. mp 104-106 °C. IR: 3392, 1734,
1684, 1549. 1H NMR (CDCl3, δ): 2.36-2.60 (m, 3H, H5, H6), 2.78
(dd, 1H, J ) 17.2, 8.8 Hz, 1H7), 3.02 (dd, 1H, J ) 17.2, 4.5 Hz,
1H7), 3.59-3.76 (m, 2H, CH2-OH), 3.80 (s, 3H, -CH3), 7.86 (s,
1H, H2). CIMS m/z: 225 (MH+). Anal. Calcd (C11H12O5): C, H.
[3-(Methoxycarbonyl)-4-oxo-4,5,6,7-tetrahydro-1-benzofuran-6-
yl]methyl p-Toluenesulfonate (14a). p-Toluenesulfonyl chloride
(200 mg, 1.03 mmol) was added to a solution of hydroxyester 13d
(180 mg, 0.80 mmol) in dry pyridine (2 mL) at 0 °C, and the
mixture was stirred at this temperature for 24 h. After the addition
of water (5 mL) and the extraction with CH2Cl2, the organic layer
was washed with water, was dried (Na2SO4), and was concentrated
to dryness, affording 315 mg of a clear solid that was crystallized
in 2-propanol to give the title compound (136 mg, 45%) as a white
solid. mp 124-126 °C. IR: 1741, 1678, 1591, 1356, 1173. 1H NMR
(CDCl3, δ): 2.32-2.54 (m, 2H, H5), 2.46 (s, 3H, CH3-Ph),
2.65-2.84 (m, 2H, H6, H7), 3.05 (dd, 1H, J ) 16.7, 4.8 Hz, 1H7),
3.85 (s, 3H, COOCH3), 3.97-4.10 (m, 2H, CH2OH), 7.36 (d, 2H,
J ) 8.0 Hz, H3 + H5 Ts), 7.78 (d, 2H, J ) 7.7 Hz, H2 + H6 Ts),
7.89 (s, 1H, H2). CIMS m/z: 379 (MH+). Anal. Calcd (C18H18O7-
S·0.25C3H8O): C, H, S.
6-(Hydroxymethyl)-4-oxo-4,5,6,7-tetrahydro-1-benzofuran-3-
carboxylic Acid (13a). A solution of alcohol 1132 (2.0 g, 11.7 mmol)
in a mixture of THF (50 mL) and 1 N HCl (10 mL) was stirred at
room temperature for 4 h and was then concentrated in vacuo,
giving 1.6 g of a yellow oil that was identified as the hydroxy-
diketone 12 and was used without further purification in the next
step, as follows. A solution of 12 (834 mg, 5.86 mmol) in methanol
(2 mL) was added with stirring to a mixture of KOH (328 mg,
5.86 mmol) in methanol (3 mL) that was maintained at 0-5 °C.
The mixture was basified until it reached pH 11 by the addition of
a 5 M solution of KOH in methanol, and then ethyl 2-bromopiruvate
(0.77 mL, 6.15 mmol) was added at a rate of 0.4 mL/min. Stirring
was continued at room temperature for 12 h, 60% NaOH aqueous
solution (0.60 mL) was added, and the mixture was left to stand at
room temperature for 15 h. After it was acidified by concentrated
HCl, the mixture was left to crystallize at +4 °C in a refrigerator
for 2 days. The precipitated solid was collected by filtration to afford
the title compound (683 mg, 55%) as a yellowish solid. mp
1
164-165 °C. IR: 3465, 1701, 1616, 1547. H NMR (CD3OD, δ):
2.64-2.80 (m, 3H, H5, H6), 2.92 (dd, 1H, J ) 17.4, 10.3 Hz, 1H7),
3.05 (dd, 1H, J ) 17.6, 4.7 Hz, 1H7), 3.68-3.71 (m, 2H, CH2-OH),
8.30 (s, 1H, H2). EIMS m/z: 210 (M+). Anal. Calcd (C10H10O5):
C, H.
(4-Oxo-2-phenyl-4,5,6,7-tetrahydro-1-benzofuran-6-yl)methyl p-
Toluenesulfonate (14b). We prepared the title compound in 63%
yield by using the same procedure that was described for 14a by
substituting 13d for 13b. mp 160-161 °C (2-propanol). IR: 1667,
1358, 1173. 1H NMR (CDCl3, δ): 2.31-2.56 (m, 2H, H5), 2.44 (s,
3H, -CH3), 2.70-2.77 (m, 1H, H6), 2.84 (dd, 1H, J ) 17.1, 9.7
Hz, 1H7), 3.00 (dd, 1H, J ) 16.7, 4.5 Hz, 1H7), 4.01-4.14 (m,
2H, -CH2OTs), 6.84 (s, 1H, H3), 7.28-7.43 (m, 5H, H3 + H5 Ts,
and H3 + H4 + H5 Ph), 7.62-7.65 (m, 2H, H2 + H6 Ph), 7.80 (d,
2H, J ) 8.2 Hz, H2 + H6 Ts). EIMS m/z: 396 (M+). Anal. Calcd
(C22H20O5S·0.25H2O·0.25C3H8O): C, H, S.
6-(Hydroxymethyl)-2-phenyl-4,5,6,7-tetrahydro-1-benzofuran-
4-one (13b). A suspension of manganese(III) acetate dihydrate (3.0
g, 11.5 mmol) in glacial acetic acid (45 mL) was heated to 80 °C
under argon until it dissolved. The solution was cooled to 45 °C,
a solution of compound 12 (823 mg, 5.78 mmol) and phenylacety-
lene (0.76 mL, 6.94 mmol) in glacial acetic acid (12 mL) was added
dropwise, and the mixture was stirred for 2 h. After the mixture
was cooled to room temperature, the solvent was removed under
reduced pressure, and the residue was dissolved in water (6 mL),
neutralized with NaHCO3 saturated solution, and extracted with
AcOEt. The organic phase was dried (Na2SO4) and concentrated
[2-(Ethoxycarbonyl)-3-methyl-4-oxo-4,5,6,7-tetrahydro-1-benzo-
furan-6-yl]methyl p-Toluenesulfonate (14c). We prepared the title
compound in 52% yield by using the same procedure that was
described for 14a by substituting 13d for 13c. mp 112-115 °C

6092 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19
Aranda et al.
1
(2-propanol). IR: 1705, 1677, 1372, 1177. H NMR (CDCl3, δ):
6-{[4-(6-Fluorobenzisoxazol-3-yl)piperidin-1-yl]methyl}-2-phen-
yl-4,5,6,7-dihydro-1-benzofuran-4-one (16b). We prepared the title
compound by using the same procedure that was described for 16a
by substituting 14a for 14b. Purification by column chromatography
with 1:2 AcOEt/hexane as the eluent afforded the title compound
(30%) as a light-brown solid. mp 162-164 °C. IR: 2931, 1673,
1611, 1440, 1119. 1H NMR (CDCl3, δ): 2.06-2.49 (m, 9H,
-CH2-N(HCH-CH2)2CH-, 1H5), 2.67-2.78 (m, 3H, 1H5, H6,
1H7), 2.96-3.20 (m, 4H, -N(HCH-CH2)2CH- + 1H7), 6.88 (s,
1H, H3), 7.06 (dt, 1H, J ) 8.8, 2.1 Hz, H5 benzisox), 7.22-7.37
(m, 2H, H7 benzisox, and H4 Ph), 7.41 (dd, 2H, J ) 6.9, 1.4 Hz,
H3 + H5 Ph), 7.64-7.72 (m, 3H, H4 benzisox, and H2 + H6 Ph).
CIMSm/z:445(MH+).Hydrochloride:Anal.Calcd(C27H25FN2O3 ·H-
Cl·0.8H2O): C, H, N.
Ethyl 6-{[4-(6-Fluorobenzisoxazol-3-yl)piperidin-1-yl]methyl}-
3-methyl-4-oxo-4,5,6,7-tetrahydro-1-benzofuran-2-carboxylate (16c).
We prepared the title compound by using the same procedure that
was described for 16a by substituting 14a for 14c. Purification by
column chromatography with 1:4 AcOEt/hexane as the eluent
afforded the title compound (84%) as a colorless oil. IR: 3524,
1711, 1680, 1612, 1553. 1H NMR (CDCl3, δ): 1.39 (t, 3H, J ) 7.1
Hz, -CH2CH3), 2.04-2.18 (m, 4H, -N(HCH-CH2)2CH-),
2.28-2.36 (m, 4H, -CH2-N(HCH-CH2)2CH-), 2.49-2.55 (m,
2H, H5), 2.55 (s, 3H, -CH3), 2.65-2.78 (m, 2H, H6 + 1H7),
3.02-3.20 (m, 4H, -N(HCH-CH2)2CH- + 1H7), 4.38 (q, 2H, J
) 7.0 Hz, -CH2CH3), 7.08 (dt, 1H, J ) 8.8, 2.1 Hz, H5 benzisox),
7.23-7.26 (m, 1H, H7 benzisox), 7.71 (dd, 1H, J ) 8.4, 5.1 Hz,
H4 benzisox). IEMS m/z: 454 (M+). Hydrochloride: mp 245-247
°C. Anal. Calcd (C25H27FN2O5 ·HCl·2/3H2O): C, H, N.
6-{[4-(4-Fluorobenzoyl)piperazin-1-yl]methyl}-4,5,6,7-tetrahydro-
1-benzofuran-4-one (18). A solution of piperazine 1730 (96 mg,
0.41 mmol), 1-hydroxy benzotriazole (HOBt) (55 mg, 0.41 mmol),
and 4-fluorobenzoic acid (57 mg, 0.41 mmol) in anhydrous CH2Cl2
(5 mL) was stirred under argon at room temperature for 15 min
and was then cooled to 0 °C. At this temperature, dicyclohexyl-
carbodiimide (DCC) (84 mg, 0.41 mmol) was added, and the
reaction mixture was kept at 0-5 °C for 1 h, was then allowed to
reach room temperature, and was left for 48 h. The precipitated
dicyclohexylurea was filtered off, and the filtrate was washed several
times with 5% NaHCO3 and water, was dried (Na2SO4), and was
condensed to dryness. We purified the oily residue by flash
chromatography by using 1:4 AcOEt/hexane as the eluent to give
the title compound (65 mg, 45%) as a white crystalline solid. mp
129-130 °C (2-propanol). IR: 2927, 1677, 1628, 1433.1H NMR
(CDCl3, δ): 2.28 (dd, 1H, J ) 16.0, 10.1 Hz, H5), 2.35-2.69 (m,
9H, 1H5, H6, 1H7, -CH2-N(CH2-CH2)2N,), 3.07 (dd, 1H, J )
16.0, 3.6 Hz, H7), 3.36-3.65 (m, 4H, N(CH2-CH2)2NCO), 6.66
(d, 1H, J ) 2.0 Hz, H3), 7.08 (t, 2H, J ) 8.6 Hz, H3+ H5 Ph), 7.33
(d, 1H, J ) 2.0 Hz, H2), 7.32 (dd, 2H, J ) 8.7, 5.4 Hz, H2 + H6
Ph).EIMSm/z:356(M+).Hydrochloride:Anal.Calcd(C20H21FN2O3·HCl·2/
3H2O): C, H, N.
1.38 (t, 3H, J ) 7.0 Hz, -CH2CH3), 2.30-2.55 (m, 2H, H5), 2.46
(s, 3H, H3C-Ph), 2.51 (s, 3H, -CH3), 2.65-2.80 (m, 2H, 1H7 +
H6), 3.02 (dd, 1H, J ) 16.7, 3.8 Hz, 1H7), 3.98-4.10 (m, 2H,
-CH2-OTs), 4.37 (q, 2H, J ) 7.1 Hz, -CH2CH3), 7.36 (d, 2H, J
) 7.9 Hz, H3 + H5 Ph), 7.78 (d, 2H, J ) 8.2 Hz, H2 + H6 Ph).
EIMS m/z: 361 (M+). Anal. Calcd (C20H22O7S·0.1H2O): C, H, S.
Methyl 6-{[4-(4-Fluorobenzoyl)piperidin-1-yl]methyl}-4-oxo-
4,5,6,7-tetrahydro-1-benzofuran-3-carboxylate (15a). A mixture of
tosylate 14a (120 mg, 0.3 mmol) and 4-(4-fluorobenzoyl)piperidine
(132 mg, 0.64 mmol) in acetonitrile (3 mL) was stirred under reflux
for 22 h. After cooling to room temperature, the solvent was
removed under reduced pressure, and the residue was dissolved in
CH2Cl2, was washed with water, was dried (Na2SO4), and was
concentrated to dryness. The crude product was purified by column
chromatography with ethanol as the eluent to afford the title
compound (58 mg, 45%) as a white solid. mp 155-157 °C (2-
1
propanol). IR: 1680, 1616. H NMR (CDCl3, δ): 1.81-1.84 (m,
4H, -N(HCH-CH2)2CH-), 2.04-2.55 (m, 5H, -CH2-N-
(HCH-CH2)2CH-, 1H5), 2.62-2.70 (m, 3H, 1H5, H6, 1H7),
2.85-3.08 (m, 2H, -N(HCH-CH2)2CH-), 3.02-3.22 (m, 2H,
1H7, -N(HCH-CH2)2CH-), 3.86 (s, 3H, -CH3), 7.10-7.16 (m,
2H, o-F-Ph), 7.89 (s, 1H, H2), 7.93-7.98 (m, 2H, m-F-Ph). CIMS
m/z: 414 (MH+). Anal. Calcd (C23H24FNO5): C, H, N.
6-{[4-(4-Fluorobenzoyl)piperidin-1-yl]methyl}-2-phenyl-4,5,6,7-
tetrahydro-1-benzofuran-4-one (15b). We prepared the title com-
pound by using the same procedure that was described for 15a by
substituting 14a for 14b. Purification by column chromatography
with 1:2 AcOEt/hexane as the eluent afforded the title compound
(37%) as a white solid. mp 181-182 °C (2-propanol). IR: 1741,
1668, 1517. 1H NMR (CDCl3, δ): 1.84-2.04 (m, 4H,-N-
(HCH-CH2)2CH-), 2.29-2.50 (m, 5H, 1H5 and -CH2-N-
(HCH-CH2)2CH-), 2.63-2.76 (m, 3H, 1H7 + H6 + 1H5),
2.85-2.88 (m, 2H, -N(HCH-CH2)2CH-), 3.04-3.20 (m, 2H,
1H7, -N(HCH-CH2)2CH-), 6.87 (s, 1H, H3), 7.11-7.16 (m, 2H,
o-F-Ph), 7.30-7.33 (m, 1H, H4 Ph), 7.39 (dd, 2H, J ) 16.0, 8.2
Hz, H3 Ph), 7.65 (d, 2H, J ) 6.5 Hz, H2 Ph), 7.94-7.99 (m, 2H,
m-F-Ph). EIMS m/z: 431 (M+). Anal. Calcd (C27H26FNO3 ·3/4H2O):
C, H, N.
Ethyl 6-{[4-(4-Fluorobenzoyl)piperidin-1-yl]methyl}-3-methyl-
4-oxo-4,5,6,7-tetrahydro-1-benzofuran-2-carboxylate (15c). We pre-
pared the title compound by using the same procedure that was
described for 15a by substituting 14a for 14c. Purification by
column chromatography with 1:2 AcOEt/hexane as the eluent
afforded the title compound (25%) as a white solid. mp 130-132
1
°C. IR: 1715, 1694, 1673. H NMR (CDCl3, δ): 1.38 (t, 3H, J )
7.11 Hz, -CH2CH3), 1.80-1.84 (m, 4H, -N(HCH-CH2)2CH-),
2.04-2.39 (m, 6H, H5, -CH2-N(HCH-CH2)2CH-), 2.54 (s, 3H,
-CH3), 2.61-2.71 (m, 2H, H6 + 1H7), 2.85-2.90 (m, 2H,
-N(HCH-CH2)2CH-), 3.06-3.08 (m, 2H, 1H7 + -N(HCH--
CH2)2CH-), 4.37 (q, 2H, J ) 7.0 Hz, -CH2CH3), 7.13 (t, 2H, J
) 6.5 Hz, o-F-Ph), 7.93-7.98 (m, 2H, m-F-Ph). IEMS m/z: 441
(M+). Anal. Calcd: (C25H28FNO5 ·0.25C3H8O): C, H, N.
Pharmacology. The affinities of the new compounds for cloned
human D2, 5-HT2A, and 5-HT2C receptors were evaluated by in vitro
binding assays that used the radioligands [3H]spiperone, [3H]ket-
anserin, and [3H]mesulergine, respectively, according to previously
described procedures.28 Ki values expressed as pKi were calculated
according to the Cheng-Prusoff equation.45
Residue Numbering. For residues belonging to helix regions of
the G-protein-coupled receptors (GPCRs), the generalized number-
ing scheme that was proposed by Ballesteros and Weinstein46 was
used.
GPCR Modeling. The human sequences of the 5-HT2A and D2
receptors were retrieved from the Swiss-Prot database47 and were
aligned with the crystal structure of the human ꢀ2 adrenergic
G-protein-coupled receptor (PDB entry 2RH1)48,49 by the use of
ClustalX software50,51 that used the PAM250 matrix and penalties
of 10 and 0.05, respectively, for gap opening and gap elongation.
The alignment was then manually refined to ensure a perfect
alignment of the highly conserved residues of the GPCR super-
family according to Baldwin et al.52 The conserved disulfide bond
between residue C3.25 at the beginning of TM3 and the cysteine
Methyl 6-{[4-(6-Fluorobenzisoxazol-3-yl)piperidin-1-yl]methyl}-
4-oxo-4,5,6,7-tetrahydro-1-benzofuran-3-carboxylate (16a). A solu-
tion of tosylate 14a (95 mg, 0.25 mmol) and 4-(6-fluorobenzisoxazol-
3-yl)piperidine (108 mg, 0.49 mmol) was refluxed in acetonitrile
(3 mL) for 24 h. The precipitate was filtered off, the solvent was
removed in vacuo, and the residue was dissolved in CH2Cl2. This
solution was washed twice with water and was dried (Na2SO4),
and the solvent was removed in vacuo, affording 149 mg of a brown
solid that, upon column chromatography with 2:1 AcOEt/hexane
as the eluent, gave the title compound (86 mg, 80%) as a light-
brown crystalline solid. mp 140-142 °C (2-propanol). IR: 1744,
1
1683, 1614, 1546. H NMR (CDCl3, δ): 2.04-2.44 (m, 9H, 1H5,
-CH2-N(HCH-CH2)2CH-), 2.64-2.75 (m, 3H, 1H5, H6, 1H7),
2.92-3.05 (m, 4H, 1H7, -N(HCH-CH2)2CH-), 3.86 (s, 3H,
-CH3), 7.06 (dt, 1H, J ) 8.8, 2.1 Hz, H5 benzisox), 7.22-7.23
(m, 1H, H7 benzisox), 7.68 (dd, 1H, J ) 8.7, 5.1 Hz, H4 benzisox),
7.90 (s, 1H, H2). EIMS m/z: 426 (M+). Anal. Calcd (C23H23FN2O5 ·1/
6H2O): C, H, N.

Analysis of New Benzofuranone DeriVatiVes
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 19 6093
(8) Filton, A.; Heel, R. C. Clozapine. A review of its pharmacological
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747.
in the middle of extracellular loop 2 (a feature common to many
GPCR receptors) was also created and was kept as a constraint in
the geometric optimization.
(9) Schwarz, J. T.; Brotman, A. W. A clinical guide to antipsychotic drugs.
We then built 3D models by using the MODELER suite of
programs,53 which yielded 15 candidate models for each receptor
final structure. The best structures were selected and were
optimized with the molecular modeling program MOE (molec-
ular operating environment; Chemical Computing Group), which
applied the Amber99 force field.54 PROCHECK software55 was
used to assess the stereochemical quality of the minimized
structures, and this resulted in good-quality parameters with an
excellent distribution of ψ and ꢁ angles in the Ramachandran
plot; more than 90% of the residues were in the most favored
regions. Furthermore, the resulting models must reproduce the
correct orientation of the side chains for the amino acids that
are strongly conserved in the GPCR superfamily,56-59 in
particular, F6.51, F6.52, and W6.48, which some authors60 have
implicated in the activation process. In the data that were recently
published for 2RH1,48,49 the cocrystallized partial inverse agonist
carazolol interacts with F6.51 and F6.52, which form an extended
aromatic network that surrounds W6.48. As a result, W6.48
adopts the rotamer that is associated with the inactive state.On
the basis of these findings, the conformation of these residues
in the present study was set to the inactive state, which is
probably more appropriate for modeling the docking of antago-
nists and more consistent with the inactive state of the main
template structure (2RH1).
Docking Simulation. We analyzed the binding modes of the
compounds in this study for the 5-HT2A and D2 receptors by using
docking simulations in the GOLD3.1.1 program.61 We docked the
ligands in the active site of 5-HT2A/D2 by defining a region of 15
Å that was centered on the CG of D3.32, a residue that is conserved
in all aminergic receptors and is known to be important in the
interaction with the ligand.62,63 The best docking solution, according
to the scoring function of GOLD and mutagenesis data, was
subjected to energy minimization by the use of MOE. The complex
was further refined in a molecular dynamics simulation that lasted
200 ps (force field MMF94x, 300 K, time step of 2 fs); subse-
quently, the energy was minimized by the application of gradient
minimization until the rms gradient was lower than 0.001 kcal/
mol·Å.
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Acknowledgment. This work was supported by grants from
the CICYT (Spain, SAF2005-08025-C03) and from the Xunta
de Galicia (Spain, PIDIT06PXIB203173PR). K.V. and J.S. were
supported, respectively, by a predoctoral fellowship from the
Diputacio´n de A Corun˜a and by grants from the Instituto de
Salud Carlos III (Red HERACLES RD06/0009 and COM-
BIOMED). J.B. received financial support from the Programa
Isabel Barreto (Xunta de Galicia, Spain).
Supporting Information Available: Elemental analysis data of
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