Conformationally constrained butyrophenones with affinity for dopamine (D1, D2, D4) and serotonin (5-HT(2A), 5-HT(2B), 5-HT(2C)) receptors: Synthesis of aminomethylbenzo[b]furanones and their evaluation as antipsychotics

Article abstract of DOI:10.1021/jm0009890

A series of novel conformationally restricted butyrophenones (6-aminomethyl-4,5,6,7-tetrahydrobenzo[b]furan-4-ones bearing 4-(6-fluorobenzisoxazolyl)piperidine, 4-(p-fluorobenzoyl)piperidine, 4-(o-methoxyphenyl)piperazine, 4-(2-pyridyl)piperazine, 4-(2-py

Full text of DOI:10.1021/jm0009890

4
678
J . Med. Chem. 2000, 43, 4678-4693
Con for m a tion a lly Con str a in ed Bu tyr op h en on es w ith Affin ity for Dop a m in e (D
, D ) a n d Ser oton in (5-HT2A, 5-HT2B, 5-HT2C) Recep tor s: Syn th esis of
Am in om eth ylben zo[b]fu r a n on es a n d Th eir Eva lu a tion a s An tip sych otics§
1
,
D
2
4
,
†
†
†
‡
‡
Enrique Ravi n˜ a,* Isabel Casariego, Christian F. Masaguer, J os e´ A. Fontenla, Gisela Y. Montenegro,
‡
‡
‡
‡
‡
|
Mar ´ı a E. Rivas, M. Isabel Loza, Maria J . Enguix, Maria Villazon, M. Isabel Cadavid, and Gian C. Demontis
Departamento de Qu ı´ mica Org a´ nica, Laboratorio de Qu ı´ mica Farmac e´ utica, and Departamento de Farmacolog ı´ a, Facultad de
Farmacia, Universidad de Santiago, E-15706 Santiago de Compostela, Spain, and Dipartimento di Psichiatria, Neurobiologia,
Farmacologia e Biotecnologia, Laboratorio di Neurobiologia, Universita di Pisa, Via Bonanno Pisano 6, I-56126 Pisa, Italy
Received May 16, 2000
A series of novel conformationally restricted butyrophenones (6-aminomethyl-4,5,6,7-tetrahydro-
benzo[b]furan-4-ones bearing 4-(6-fluorobenzisoxazolyl)piperidine, 4-(p-fluorobenzoyl)piperidine,
4
-(o-methoxyphenyl)piperazine, 4-(2-pyridyl)piperazine, 4-(2-pyrimidinyl)piperazine, or linear
butyro(or valero)phenone fragments) were prepared and evaluated as antipsychotic agents by
in vitro assays for affinity for dopamine receptors (D , D , D ) and serotonin receptors (5-HT2A
-HT2B, 5-HT2C), by neurochemical studies, and by in vivo assays for antipsychotic potential
1
2
4
,
5
and the risk of inducing extrapyramidal side effects. Potency and selectivity depended mainly
on the amine fragment connected to the cyclohexanone structure. Compounds 20b, with a
benzoylpiperidine moiety, and 20c, with a benzisoxazolyl fragment, were selective for 5-HT2A
receptors. The in vitro and in vivo pharmacological profiles of N-[(4-oxo-4,5,6,7-tetrahydrobenzo-
[b]furan-6-yl)methyl]-4-(p-fluorobenzoyl)piperidine (20b, QF1003B) and N-[(4-oxo-4,5,6,7-tetra-
hydrobenzo[b]furan-6-yl)methyl]-4-(6-fluorobenzisoxazol-3-yl)piperidine (20c, QF1004B) suggest
that they may be effective as antipsychotic (neuroleptic) drugs.
In tr od u ction
pharmacological deconnection of serotoninergic projec-
tions from the midline nuclei. Hence Meltzer et al.
1
6
17,18
Schizophrenia is a complex disorder affecting ap-
proximately 1% of the population. Classical neurolep-
tics used in its treatment, such as haloperidol (1, Chart
suggested that the profiles of clozapine and other
atypical antipsychotics, such as risperidone (3), olanz-
apine (4), and quetiapine (5), depend largely on their
relative affinities for D2 and 5-HT2A receptors.19 They
proposed that the ratio pKi 5-HT2A/D2 (between pKi for
1
1
), are poorly effective against its negative symptoms
(apathy, motor retardation, flat affectivity, poor speech,
etc.). Furthermore, the use of these classical antipsy-
chotics is associated with severe side effects, notably
acute extrapyramidal symptoms (EPS), that appear to
be an unavoidable consequence of their mechanism of
action. Indeed, both their induction of EPS and their
antipsychotic efficacy originate from blockade of dop-
amine (DA) receptors: striatal receptors in the case of
5
-HT2A and pKi for D2) may be used to discriminate
2
atypical antipsychotics, having a ratio > 1.12, from
classical antipsychotics, which have a ratio < 1.09. Note
that this latter finding raises the question of whether
there is a D2 affinity threshold, below which there is no
antipsychotic activity even if the pKi 5-HT2A/D2 ratio is
3
-5
high: clozapine has only moderate affinity for D , and
EPS induction
and mesolimbic receptors in the case
2
of antipsychotic activity.6
-8
the 5-HT2A-selective serotonin antagonist MDL 100907
(6), which has no specific affinity for DA receptors, has
Clozapine (2) was the first of a new group of “atypical”
shown antipsychotic potential in experiments with
neurochemical, electrophysiological, and behavioral
or nonclassical antipsychotics that cause no EPS and
are effective against negative symptoms.9-12 Since
2
0-22
models.
Nevertheless, the “5-HT2A/D2 concept” has
clozapine blocks not only DA receptors but also subtype
contributed to the development not only of atypical
antipsychotics with a close chemical similarity to cloza-
pine, such as olanzapine (4), quetiapine (5), and zotepine
5
-HT2A serotonin receptors, it is thought that its atypical
activity profile may be due to its effects on interaction
between the serotonin and DA systems;
specifically, it is believed that blockade of 5-HT2A
receptors in the limbic and cortical areas favors efficacy
against negative symptoms and that their blockade in
the striatal area reduces EPS, in both cases due to
1
3-15
more
(9), but also of others of an apparently different chemical
nature, such as risperidone (3), ocaperidone (7), and
sertindole (8).
As well as to 5-HT2A, atypical antipsychotics also bind
to other subtype 2 serotonin receptors (5-HT2B and
§
23
This is the 22nd paper in the series “Synthesis and CNS Activity
5-HT2C), and they are implicated not only in schizo-
of Conformationally Restricted Butyrophenones”; for preceding papers,
see ref 47.
phrenia but also in anxiety, depression, anorexia ner-
vosa, migraine, and cardiovascular disorders, including
*
Correspondence and reprints: Prof. Enrique Ravi n˜ a. Phone: +34-
9
81-594628. Fax: +34-981-594912. E-mail: qofara@usc.es.
24
hypertension. The subtypes of this family (5-HT2A,
†
Departamento de Qu ´ı mica Org a´ nica, Universidad de Santiago.
Departamento de Farmacolog ´ı a, Universidad de Santiago.
Universita di Pisa.
‡
5-HT2B, and 5-HT2C) are defined on the basis of primary
structure, secondary messengers, and operational pro-
|
1
0.1021/jm0009890 CCC: $19.00 © 2000 American Chemical Society
Published on Web 11/10/2000

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4679
Ch a r t 1
file24,25 and have different distributions in the body:
Ch a r t 2
5
-HT2A is found both in the brain and in peripheral
2
6
27
tissues, 5-HT2B is mainly peripheral, and 5-HT2C has
only been found in the central nervous system (CNS)
(
mainly in the choroid plexus, limbic structures, and
basal ganglia, a distribution different from that of
2
8
5
-HT2A). It is now thought that some pathologies or
pharmacological effects previously attributed to dys-
function or action at 5-HT2A may in fact be due to
dysfunction or action at other subtype 2 receptors. In
particular, it has been suggested that 5-HT2C plays a
2
9,30
role in anxiety, depression, and affective disorders,
and more recently the noninduction of EPS by atypical
antipsychotics has also been attributed to blockade of
3
1
5
-HT2C. Unfortunately, elucidation of these possibili-
ties has been hindered by the lack of any 5-HT2-blocking
compounds with high specificity for one or another
subtype, especially as regards differentiation of 5-HT2B
and 5-HT2C.
Despite the uncertainty about its mechanism of
action, clozapine is still the most widely used atypical
antipsychotic, and no other currently available agent
appears to have an equal or superior spectrum of
efficiency. However, its therapeutic use is severely
limited by its being associated with increased risk of
An alternative approach to the development of im-
proved antipsychotics originated from the discovery and
characterization of new types of DA receptors. Specifi-
cally, it has been reported that schizophrenia is associ-
37
agranulocytosis and seizures; likewise several cases
of myocarditis and cardiomyopathy associated with
3
2
ated with high levels of D4 receptor, leading to the
hypothesis that it is this receptor rather than D2 that
should be blocked by an antipsychotic.33 Furthermore,
the affinities of clozapine and some other atypical
antipsychotics for D4 are several times their affinities
for D2, raising the possibility that their superiority to
classical antipsychotics is due to more precise targeting
of the relevant DA receptor, rather than or in addition
to their action at the 5-HT2A receptor. However, com-
pounds such as L 74587034 (10, Chart 2) or CP 293019
38
clozapine treatment have been recently reported.
Accordingly, development of new antipsychotic agents
remains a great challenge today.
In previous papers we have described the synthesis
and atypical antipsychotic activities of 3-aminomethyl-
tetralones (13, Chart 3) and 2-aminoethylbenzocyclo-
39-42
alkanones (14),
which are conformationally restrict-
ed butyrophenone analogues of haloperidol; 5-amino-
ethyl- and 6-aminomethyl-4,5,6,7-tetrahydroindol-4-
35
43-45
ones (15, 16),
the former of which are butyro-
(
11), which block D4 almost exclusively, have proved
phenone homologues of the antipsychotic molindone;
ineffective in clinical trials (despite excellent perfor-
mance in preclinical trials, including experiments on live
animal models);36 the D2 antagonist remoxipride (12),
which has little affinity for D4, behaves like the least
EPS-inducing classical antipsychotics, and Meltzer et
and 2-aminomethyl-1,2,3,9-tetrahydro-4H-carbazol-4-
4
6,47
ones (17).
In all these series, the conformation of the
butyrophenone butyl chain is restricted by its partial
incorporation in a cycloalkanone ring fused to a benzene
ring or a heterocycle. We have also recently described
the synthesis, pharmacology, 3D QSARs, and molecular
modeling of some aminoalkylthienocycloalkanones (18,
1
7,18
al.
tinguished from 12 classical antipsychotics by pKi
-HT2A/D2 ratios but not by pKi 5-HT2A/D4 or pKi D2/D4
found that 13 atypical antipsychotics were dis-
4
8
5
19).
ratios. In short, more work must be done to clarify the
role of D4 in cerebral activity.
In continuation of this work, we have now synthesized
and pharmacologically evaluated some 6-aminomethyl-

4
680 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Ravi n˜ a et al.
Ch a r t 3
furoyl)propionic acid, formation of the Mannich base
corresponding to the relevant -NRR, and final cycliza-
tion. Route C involves condensation of â-(2-furoyl)-
propionic acid with formaldehyde, reduction of the
resulting â-(2-furoyl)-γ-butyrolactone, opening of the
lactone ring to obtain γ-bromo-â-furfurylbutyric acid,
nucleophilic substitution of the bromine for the relevant
-
NRR moiety, and final cyclization. In the route shown
in Scheme 2, the furan ring is constructed on a substi-
tuted cyclohexanone obtained by Birch reduction of 3,5-
dimethoxybenzoic acid, subsequent reduction of the
carboxyl group with a hydride, and acid hydrolysis of
the resulting 1,4-dihydro-3,5-dimethoxybenzoic alcohol;
condensation of the cyclohexanone with chloroacetal-
dehyde then affords a 6-substituted benzofuranone that
can be transformed into amino ketones 20 by tosylation
and nucleophilic substitution.
Route A was ruled out chiefly because of the long
reaction times that in our experience have been needed
for protection of the carbonyl in similar compounds, and
routes B and C because of the difficulty in obtaining
the common intermediate â-(2-furoyl)propionic acid.
This latter cannot be prepared by straightforward
succinoylation of furan or by reaction of diethyl mal-
onate with 2-bromoacetylfuran because these proce-
dures afford intractable resins. Although it can be
obtained by hydrolysis of the corresponding nitrile or
ester, which according to the literature (and in ac-
cordance with Scheme 1) is afforded by addition of
acrylonitrile or methyl acrylate to furfural in the pres-
5
3,54
4
3
,5,6,7-tetrahydrobenzo[b]furan-4-ones (20; see Scheme
and Table 1). One of these compounds (20b) has two
ence of sodium or potassium cyanide
or thiazolium
5
5
salts, the cyanide method has been reported to have
moderate yields at best, and our attempts at using
3-benzyl-5-(2-hydroxyethyl)-4-methylthiazolium as per
Stetter and Kuhlmann have provided very low yields
of the target compound, the major product being furoin.
5
4
butyrophenone pharmacophores: the semirigid amino-
alkyl furanone moiety and a 4-(p-fluorobenzoyl)piperi-
dine fragment. The latter has been described as an
antipsychotic pharmacophore with potency similar to
that of linear butyrophenones,49 and a SAR study of
ketanserin analogues has suggested that the benzoyl
carbonyl (present in 20b,h ,i) may be important for
5
4
We accordingly decided to pursue the route shown in
Scheme 2, which has the further advantage that 1,4-
dihydro-3,5-dimethoxybenzoic alcohol (21) is also a key
intermediate in the preparation of a series of heterocy-
clic butyrophenones based on indole and carbazole.
Alcohol 21 was hydrolyzed with 1 N HCl in tetrahydro-
furan, the resulting cyclohexanone (22) was condensed
with chloroacetaldehyde in the presence of NaHCO3,
and subsequent acid dehydration afforded the key
intermediate 6-hydroxymethyl-4-oxo-4,5,6,7-tetrahy-
drobenzo[b]furan (23) in 60% yield from 21 (Scheme 3).
Compound 23 was tosylated in 60% yield with p-
toluenesulfonyl chloride in anhydrous pyridine, and
reaction of the tosylate 24 with the amines of Table 1
in N-methyl-2-pyrrolidone (NMP) afforded 70-85%
yields of compounds 20a [HNRR ) morpholine], 20b
5
6
5
0
anchorage or orientation at the 5-HT2A receptor.
4
5
46,47
Compound 20c has the 4-(6-fluorobenzisoxazol-3-yl)-
piperidine moiety of risperidone; 1,2-benzisoxazole is
bioisosteric to benzoyl,51 and 3-(4-piperazinyl)-6-hy-
droxybenzisoxazoles have also recently been reported
to have potential as atypical antipsychotics. The new
compounds were all characterized pharmacologically by
determination of their affinities for D2, 5-HT2A, and
5
2
5
-HT2C (in radioligand binding studies) and for 5-HT2B
(in functional studies); the most active were further
characterized regarding their affinities for D1 and D4
and their effects on the behavior of experimental
animals and on the levels of DA and its major metabo-
lites in striatal tissue.
[
4
HNRR ) 4-(p-fluorobenzoyl)piperidine], 20c [HNNR )
-(6-fluorobenzisoxazol-3-yl)piperidine], 20d [HNNR )
Ch em istr y
N-phenylpiperazine], 20e [HNNR ) N-(o-methoxyphe-
nyl)piperazine], 20f [HNNR ) N-(2-pyridyl)piperazine],
20g [HNNR ) N-(2-pyrimidinyl)piperazine], and 25
[HNNR ) N-tert-butoxycarbonylpiperazine] (Scheme 3).
Cleavage of the tert-butoxycarbonyl group of 25 with
trifluoroacetic acid in anhydrous dichloromethane then
afforded piperazine 26 in 75% yield, and alkylation of
26 with 4-chloro-1,1-ethylenedioxy-1-(p-fluorophenyl)-
butane in methyl isobutyl ketone containing potassium
carbonate and catalytic amounts of potassium iodide,
For preparation of the 6-aminomethyl-4-oxo-4,5,6,7-
tetrahydrobenzo[b]furans 20a -i we initially considered
the routes shown in Schemes 1 and 2. Those shown in
Scheme 1 all start from furfural: Route A involves
Stobbe condensation with diethyl succinate, reduction
of the double bond, cyclization, formation of an amide
with the relevant -NRR moiety (Table 1), protection of
the ketone carbonyl, and reduction of the amide carbo-
nyl. Route B consists of conversion of furfural to â-(2-

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4681
Sch em e 1
Sch em e 2
Sch em e 3a
its affinity for 5-HT2C and was inactive for 5-HT2B (pA2
4). In addition, the affinities of 20i for 5-HT2A and
-HT2B were higher than for 5-HT2C.
0c was the new compound with greatest affinity for
D1, D2, 5-HT2A, and 5-HT2B (in this last case, jointly with
0b), while the greatest affinity for 5-HT2C was shown
by the pyridylpiperazine 20f. Affinity for D1, D2, and
-HT2A was reduced by replacing the benzisoxazolyl
<
5
2
2
5
moiety of 20c with the fluorobenzoyl group of 20b and
for D2 and 5-HT2A was reduced further by interpolating
a flexible carbon chain between the nitrogenated ring
and the benzoyl group as in 20h , which also showed
slightly reduced affinity for 5-HT2B. The affinities for
a
Reagents: (i) HCl; (ii) ClCH2CHO, NaHCO3; (iii) Ts-Cl, Py;
5
-HT2A, D2, and in most cases 5-HT2B fell further still
(
iv) HNRR.
upon lengthening the benzoyl-bearing chain, as in 20i,
or on replacing the benzoylalkyl moiety with pyridyl,
pyrimidinyl, or o-methoxyphenyl groups, as in 20e-g,
respectively (in particular, 20g was inactive at 5-HT2A,
pKi < 4). It appears that affinity for the D2 subtype
correlates with the presence of a piperidine moiety
(compounds 20b,c), while introduction of a piperazine
followed by acid hydrolysis of the ketal, afforded 20h
in 50% yield from 26. Similarly, alkylation of 26 with
5
-bromo-1-(p-fluorophenyl)-1-pentanone gave, via the
same procedure, valerophenone 20i, again in 50% yield.
Resu lts a n d Discu ssion
(
compounds 20d -i) is associated with loss of affinity.
The affinity of 20c for D2 (pKi ) 8.02) was similar to
The results of the in vitro experiments are sum-
marized in Table 2 and illustrated in Figures 1 and 2.
The salient finding is that, regarding their interactions
with serotonin receptors, both 20c,d were selective for
that of risperidone (8.09), greater than that of clozapine
(6.58), and less than that of haloperidol (8.48); see
Figure 2. Its affinity for D4 (pKi ) 7.68) was similar to
that of clozapine (7.42), slightly smaller than that of
haloperidol (8.20), and slightly greater than that of 20b,
5
-HT2A: the affinity of 20c for 5-HT2A was >23 times
its affinity for 5-HT2B and >1000 times its affinity for
-HT2C and 20d had an affinity for 5-HT2A >150 times
5

4
682 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Ravi n˜ a et al.
Sch em e 4a
a
Reagents: (i) TFA; (ii) (1) Na2CO3, KI, (2) HCl; (iii) Na2CO3, KI.
(
7.04) (affinity for D4 was only assayed for the new
blockers, R1-blockers, and reserpine (by induction of DA
depletion), and is therefore by itself indicative only of
general CNS depression. However, haloperidol (2 mg/
kg), clozapine (5 mg/kg), risperidone (5.0 mg/kg), 20b
(4 mg/kg), and 20c (0.5 mg/kg) also caused statistically
significant reductions in amphetamine-induced hyper-
motility (Figure 5), speed, and rearing (data not shown).
Amphetamine-induced hypermotility is attributed to the
activation of D2 receptors in the nucleus accumbens by
DA released due to amphetamine-induced reversal of
compounds with greatest affinity for D2 and 5-HT2A).
Neither 20c nor 20b were as selective as haloperidol
for D2 versus D1.
None of the new compounds had a pKi 5-HT2A/D2 ratio
1
4,17
greater than the Meltzer et al.
threshold for atypical
antipsychotic activity.1 It is also noteworthy that, like
haloperidol, they all had low affinity for 5-HT2C, for
which the atypical antipsychotics clozapine and risperi-
done have high affinity. This result is in keeping with
recent suggestions that it is 5-HT2C blockade rather
than 5-HT2A blockade that can prevent the EPS induced
,12
5
9,60
the DA re-uptake system.
However, effects on
behavior are dose-dependent: while doses in the range
3
1
by haloperidol (see Introduction).
4.5-5 mg/kg induced maximum increase in hypermo-
6
1
Experiments with rats or mice were carried out to
evaluate the new compounds with greatest affinity for
DA receptors (20b,c) for possible antipsychotic activity
and induction of EPS. Antipsychotic potential was
assayed in terms of reduction of spontaneous motor
activity, d-amphetamine-induced hypermotility, and
apomorphine-induced climbing behavior. Spontaneous
activity was reduced, as expected,5 by reference drugs
haloperidol (2 mg/kg), clozapine (5 mg/kg), and risperi-
done (0.16-5.0 mg/kg) (Figure 3A) and also, dose-
dependently, by both 20b,c (ED50 ) 1.21 and 0.48 mg/
kg, respectively; see Figure 4A,B) which like risperidone
generally exerted maximum effect within 20-30 min
of administration (Figure 3B,C). The reference drugs
and new compounds 20b,c also slowed movement and
inhibited rearing (data not shown).
The behavior of 20b,c in the spontaneous activity
tests correlates with their affinities for D1, D2, and D4
but without further evidence is not necessarily attribut-
able to action at these receptors; although D1 and D2
are known to be involved in the rapidity of initiation of
movement and speed,58 a reduction in spontaneous
motor activity can be caused by many substances that
are not DA antagonists including, for example, H1-
tility, higher doses cause a predominance of stereo-
typed activity by inducing the release of DA in other
6
2
pathways. Results obtained with the d-amphetamine
dosage used in this work (5 mg/kg) is a more specific
sign of in vivo D2-blocking activity, and although the
hypermotility induced by low amphetamine doses (0.5
mg/kg) can be inhibited by both D2 antagonist and
7
63,64
5-HT2A antagonist,
which appear to prevent the
6
5-68
5-HT2A-mediated activation of DA synthesis,
5-HT2A
blockers are ineffective against amphetamine dosages
6
4
> 2 mg/kg.
Further corroboration of the in vivo DA antagonism
of 20b ,c was provided by experiments in which they
inhibited the effects of the direct D1 and D2 agonist
apomorphine, which at low dosages causes hypomotility
and yawning (due to activation of autoreceptors6 ) and
at the high dosages used in this work (2 mg/kg) induces
climbing by simultaneous activation of postsynaptic D1
9,70
7
1-80
and D2 receptors.
In keeping with the in vitro
binding results on affinity for D2 (Table 2), 20b,c
inhibited apomorphine-induced climbing more effi-
ciently than clozapine but less efficiently than haloperi-
dol or risperidone, and 20c was more potent than 20b
(see Figure 6 and Table 3). However, the involvement

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4683
Ta ble 1. 6-Aminomethyl-4,5,6,7-tetrahydrobenzo[b]furan-4-ones
a
Recrystallized from i-PrOH. b Recrystallized from AcOEt.
Ta ble 2. In Vitro Assays
pKi
pKi ratio
pA2
compd
5-HT2A
5-HT2C
D1
D2
D4
5-HT2A/D2
5-HT2B
6
2
2
2
2
2
2
2
2
0b (QF 1003B)
0c (QF 1004B)
0d (QF 1007B)
0e (QF 1006B)
0f (QF 1008B)
0g (QF 1010B)
0h (QF 1005B)
0i (QF 1011B)
7.29 ( 0.20
7.97 ( 0.03
7.66 ( 0.05
5.64 ( 0.11
5.08 ( 0.10
<4
6.02 ( 0.03
5.90 ( 0.04
7.28 ( 0.03
7.56 ( 0.07
9.51 ( 0.03
33.3% (10- M)
4.74 ( 0.15
5.47 ( 0.08
4.89 ( 0.04
6.64 ( 0.02
<4
6.71 ( 0.11
7.54 ( 0.06
7.02 ( 0.14
8.02 ( 0.11
<5
7.04 ( 0.03
7.68 ( 0.19
1.03
0.99
6.71 ( 0.57
6.61 ( 0.38
<4
6.19 ( 0.20
5.46 ( 0.03
5.74 ( 0.24
6.36 ( 0.12
6.23 ( 0.08
<5
<5
5.14 ( 0.25
6.39 ( 0.12
<5
-
6
3.7% (10 M)
0.94
-
6
44.3% (10 M)
5.34 ( 0.03
8.30 ( 0.02
7.38 ( 0.03
haloperidol
clozapine
risperidone
6.70 ( 0.08
6.77 ( 0.13
8.48 ( 0.12
6.58 ( 0.05
8.09 ( 0.29
8.20 ( 0.03
7.42 ( 0.07
0.93
1.19
1.20
6.90 ( 0.2
of factor(s) other than D1 and D2 blockade is suggested
by the finding that risperidone was more potent than
haloperidol despite having less affinity for both D1 and
D2, and furthermore, risperidone was 4 times more
potent than 20c despite having quite similar affinity
for D2 and less affinity for D1.81 The putative additional
factor is unlikely to be 5-HT2A blockade, because al-
though risperidone has at least 30 times more affinity
for 5-HT2A than any of the other drugs used, the
selective 5-HT2A blocker MDL 100907 has no effect on

4
684 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Ravi n˜ a et al.
3
F igu r e 1. (A) Inhibition by 20c (b) and risperidone (O) of [ H]ketanserin binding by rat frontal cortex membrane preparations.
(
B) Inhibition by 20c of serotonin-induced contrations of rat stomach fundus (curves from a single representative of 3-4 replicate
3
experiments; vertical bars indicate SEM). (C) Inhibition by 20c (b) and risperidone (O) of [ H]mesulergine binding by bovine
choroid plexus membrane preparations. Graphs A and C show triplicate points from a single experiment; a total of two replicate
experiments were performed in each case.
ing, 0.34 mg/kg, was lower than the dose assayed in
amphetamine-induced hypermotility (0.5 mg/kg). Ad-
ditionally, clozapine, with affinity for 5-HT2A similar to
the new compounds, was less potent that 20c.
If it is accepted that apomorphine-induced climbing
is mediated chiefly by activation of mesolimbic DA
8
3,84
receptors
and that blocking these receptors has
antipsychotic effects because it blocks the mesolimbic
dopaminergic system, which is functionally altered in
schizophrenics, then the above results suggest that
2
0b,c may have antipsychotic activity. To evaluate the
risk of any such activity being accompanied by EPS, we
assayed their ability to induce catalepsy in mice, cata-
lepsy being attributed, like EPS, to blockade of striatal
F igu r e 2. Inhibition by reference drugs and new compound
3
2
2
0c of [ H]spiperone binding by striatal D .
8
5-87
DA receptors.
Neither 20b nor 20c induced cata-
apomorphine-induced climbing at dosages at which it
inhibits amphetamine-induced hypermotility,82 whereas
the ED50 value of 20c for apomorphine-induced climb-
lepsy at dosages lower than 2 mg/kg, but when higher
dosages were assayed they exhibited ED50 values of
respectively 3.44 mg/kg (95% CL 2.78-4.25) and 3.84

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4685
cantly affected by the reference drugs haloperidol and
risperidone or the new compound 20c. Clozapine, at the
highest dosage used, increased the DA level. Metabo-
lites, DOPAC and HVA, were significantly increased by
all four compounds, risperidone especially (see Figure
8
). These changes are thought to be due to the blockade
of DA receptors giving rise to a compensatory increase
in DA turnover that is reflected in the raised levels of
8
8-91
the metabolites.
Although 20c fails to meet the Meltzer et al. criterion
for atypical antipsychotic activity (pKi 5-HT2A/D2 >
1
.12), it nevertheless has a relative high ED50 catalepsy/
ED50 climbing ratio. The cause may be its poor selectiv-
ity for D2 versus D1 (see above), since cataleptogenic
potency may be favored by a high D2 selectivity like that
9
2
of haloperidol. Likewise, the similarity between the
D2:D1 selectivities of 20b,c may explain their having
very similar ED50 values for induction of catalepsy.
Since neither 20b nor 20c have high affinities for
5
-HT2C or high pKi 5-HT2A/D2 ratios, their being less
cataleptogenic than haloperidol is thus attributable to
a combination of lower affinity and lower selectivity for
D2, while their being more cataleptogenic than clozapine
or risperidone may be attributed to their low affinity
for 5-HT2C.
In conclusion, the new compounds 20b,c have moder-
ate-to-high affinity for 5-HT2A, D1, D2, and D4 (20c
especially), moderate affinity for 5-HT2B, and low affin-
ity for 5-HT2C. In behavioral experiments with rats and
mice they caused effects compatible with blockade of DA
receptors in the limbic region (reduction of d-amphet-
amine-induced hypermotility and apomorphine-induced
climbing) and the striatum (induction of catalepsy), and
the striatal activity of 20c was confirmed by determi-
nation of its effect on striatal levels of DA and its major
metabolites. These results are in keeping with their pKi
5
-HT2A/D2 ratios, which are in the range of that for
typical or classical antipsychotics. However, their induc-
tion of catalepsy less potently than haloperidol, which
is also in keeping with their pKi 5-HT2A/D2 ratios (1.03
and 0.99) and with their relatively poor selectivity for
D2 versus D1, suggests relatively low capacity to induce
EPS. The high selectivity of 20b,c for 5-HT2A and
F igu r e 3. Time dependence of the effects of various drug
dosages on the spontaneous activity of mice (means ( SEM):
5
-HT2B versus 5-HT2C and of 20d for 5-HT2A and 5-HT2C
(
A) risperidone, p < 0.01 with respect to vehicle at all dosages
and times except after 10 min with 0.63 and 0.16 mg/kg, for
which the difference was not significant; (B) 20b, p < 0.01 with
respect to vehicle with 10, 5, 3.5, and 2.5 mg/kg (except after
versus 5-HT2B may make these compounds useful for
researching the physiological and pharmacological roles
of these receptors.
4
0 min with 2.5 mg/kg, not significant), not significant with 1
mg/kg except after 50 min (p < 0.01) and 60 min (p < 0.05),
not significant with 0.5 mg/kg except after 20 and 50 min (p
Exp er im en ta l Section
Ch em istr y. 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
<
0.05 in both cases), not significant with 0.25 mg/kg; (C) 20c,
n.s. indicates no statistical significance with respect to vehicle,
p < 0.05, **p < 0.01.
*
-
1
1
13
bands are given, in cm
. H and C NMR spectra were
mg/kg (95% CL 2.78-5.30) (Figure 7); these values are
respectively 3.6 and 11.3 times the corresponding ED50
for inhibition of apomorphine-induced climbing (Table
recorded with a Bruker WM AMX apparatus (300 MHz);
chemical shifts are given in parts per million (δ) downfield
from tetramethylsilane, and J values are given in hertz (Hz).
Mass spectra were recorded on Kratos MS-50 or Varian Mat-
711 mass spectrometer in fast atom bombardment (FAB) mode
(with 2-hydroxyethyl disulfide as matrix) or by electron impact
3
). The ED50 catalepsy/ED50 climbing ratio for haloperi-
dol was 4.8, and the atypical antipsychotics used as
reference drugs, clozapine and risperidone, induced no
catalepsy even at dosages 20 times their ED50 values
for climbing inhibition.
The action of 20c in the striatum was confirmed by
determination of the concentrations of DA and of its two
major metabolites, DOPAC and HVA, in striatal tissue
(EI). Optical rotations at the sodium D-line were determined
in MeOH solutions of the indicated concentrations using a
Perkin-Elmer 241 polarimeter. Flash column chromatography
was performed using Kieselgel 60 (60-200 mesh; E. Merck
AG, Darmstadt, Germany). Reactions were monitored by thin-
layer chromatography (TLC) on Merck 60 GF254 chromatogram
sheets using iodine vapor and/or UV light for detection; unless
2
h after administration. DA levels were not signifi-

4
686 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Ravi n˜ a et al.
F igu r e 4. Total distance travelled in 1 h by mice treated with 20b (A) or 20c (B) and corresponding percent reduction in
spontaneous locomotor activity. The data shown are means ( SEM. Significant differences with respect to controls are indicated
by *p < 0.05 or **p < 0.01.
F igu r e 5. Inhibition of d-amphetamine-induced hypermotility
in mice by haloperidol, clozapine, risperidone, 20b, and 20c:
distances travelled in 1 h (means ( Sem) and percent reduc-
tions with respect to controls. All differences with respect to
F igu r e 6. Dose-effect curves for inhibition of apomorphine-
induced climbing by reference drugs and selected new com-
pounds.
controls are statistically significant (p < 0.01).
otherwise stated the purified compounds each showed a single
spot. Elemental combustion analyses were performed on a
Perkin-Elmer 240B apparatus at the Microanalysis Service of
the University of Santiago de Compostela; unless otherwise
stated all reported values are within (0.4% of the theoretical
compositions. Solvents were purified as per Vogel93 by distil-
lation over the drying agent under an argon atmosphere and
were used immediately. The drying agents used were Na/
benzophenone for THF, ether and toluene; P
2 5 2 2
O for CH Cl ;
K
2
CO for acetone and ethyl acetate; KOH for pyridine and
3

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4687
Ta ble 3. In Vivo Assays
ED50 (mg/kg)
rel potency (ED50 haloperidol/ED50 drug)
compd
climbing (95% CL)
catalepsy
climbing
catalepsy
ED50 catalepsy/ED50 climbing
2
2
0b (QF 1003B)
0c (QF 1004B)
0.95 (0.80-1.14)
0.34 (0.18-0.61)
0.14 (0.13-0.15)
2.21 (2.03-2.41)
0.09 (0.08-0.09)
3.44
3.84
0.67
0.15
0.41
1.00
0.06
1.56
0.19
0.17
1.00
3.62
11.29
4.79
haloperidol
clozapine
risperidone
>50.0
>2.0
with 1:1 AcOEt/hexane as eluant gave 1.84 g of a white solid
identified as the tosylate 24: yield 60%, mp 104-105 °C (i-
1
PrOH). IR: 2957, 1679, 1593. H NMR (CDCl
3
): δ 2.27-3.24
); 2.69-
); 3.99-
); 7.32 (d,
Ph), 7.78 (d,
): δ 22.1 (-CH ); 26.4
-OTs); 106.8 (C ); 121.4
Ph); 132.9 (C Ph); 143.7 (C );
Ph); 165.5 (C7a); 192.06 (-CO-). MS (FAB, m/z): 321
MH ). Anal. (C16 S) C, H, S.
-Mor ph olin ylm eth yl-4,5,6,7-tetr ah ydr oben zo[b]fu r an -
-on e (20a ). A solution of tosylate 24 (0.6 g, 1.87 mmol) and
(m, 1H, 1H
5
); 2.47 (s, 3H, -CH
.78 (m, 2H, 1H , H ); 3.01 (dd, 1H, J ) 15.6, 3.7, 1H
.10 (m, 2H, -CH -OTs); 6.62 (d, 1H, J ) 2 Hz, H
H, J ) 1.9, H ); 7.35 (d, 2H, J ) 8.22, H , H
H, J ) 8.3, H , H
Ph). 13C NMR (CDCl
); 35.5 (C ); 40.4 (C ); 72.1 (-CH
3a); 128.3 (C Ph); 130.4 (C
45.6 (C
3
); 2.50-2.52 (m, 1H, 1H
5
2
4
1
2
7
6
7
2
2
3
3
5
2
6
3
3
(
(
C
C
7
6
5
2
3
3
2
1
2
1
(
4
+
16 5
H O
6
4
morpholine (0.33 g, 3.75 mmol) was stirred in NMP (15 mL)
for 16 h at 85 °C. The solvent was removed in vacuo, and the
residue was dissolved in CH
2
Cl
2
. This solution was washed
twice with water, dried (Na SO
2
4
), and the solvent was removed
in vacuo, affording an orange solid that upon column chroma-
tography with AcOEt as eluant gave 20a (0.33 g, 75%) as a
white crystalline solid of mp 133-134 °C (i-PrOH). IR: 2958,
1
1
673. H NMR (CDCl
3
): δ 2.21-2.30 (m, 1H, 1H
, -N(CH -CH
HCH-N<); 3.03-3.08 (m, 1H, -HCH-N<); 3.69 (t, 4H, J ) 4.6,
N(CH -CH O); 6.65 (d, 1H, J ) 1.7, H ); 7.30 (d, 1H, J )
). C NMR (CDCl ): δ 28.4 (C ); 33.3 (C ); 42.9 (C );
4.4 (-N(CH -CH O); 63.6 (-CH -N<); 67.4 (-N(CH -CH O);
06.8 (C ); 121.4 (C3a); 143.3 (C ); 166.9 (C7a); 194.1 (-CO-). MS
) C, H, N. Hydro-
5
); 2.36-2.51
F igu r e 7. Results of catalepsy test, showing the time for
which initial posture was maintained by mice treated with
vehicle or new compounds (means ( SEM). Significant differ-
ences with respect to controls are indicated by *p < 0.05, **p
(
-
-
1
5
1
m, 6H, 1H
5
, 1H
7
2
2
)
2
O); 2.52-2.68 (m, 3H, H6, 1H
7
,
2
2
)
2
2
1
3
.7, H
3
3
7
6
5
<
0.01, or ***p < 0.001.
2
2
)
2
2
2
2 2
)
3
2
TEA; and CaSO /4 Å molecular sieves for DMF. Hydrochlo-
4
+
(
FAB, m/z): 236 (MH ). Anal. (C13
H17NO
3
rides were prepared by dropwise addition, with cooling, of a
saturated solution of HCl in anhydrous ether to a solution of
the amine in anhydrous ether or absolute ethanol/ether.
chloride: mp 138-139 °C (AcOEt).
Compounds 20b-g and 25 were prepared similarly, with
the following results.
6
-Hyd r oxym et h yl-4,5,6,7-t et r a h yd r oben zo[b]fu r a n -4-
5
6
6-[4-(p -F lu or ob en zoyl)p ip er id in -1-ylm et h yl]-4,5,6,7-
on e (23). A solution of alcohol 21 (1 g, 6.84 mmol) in a
mixture of THF (30 mL) and 1 N HCl (6 mL) was stirred at
room temperature for 2 h and then concentrated in vacuo,
giving 0.9 g of a yellow oil that was identified as alcohol 22
and was used without further purification in the next step, as
follows. A solution of 22 (0.9 g, 6.33 mmol) in 6 mL of water
was added at a rate of 0.4 mL/min, with stirring, to a mixture
tetr a h yd r oben zo[b]fu r a n -4-on e (20b): yield 75%, mp 113-
1
1
14 °C (i-PrOH). IR: 2949, 2768, 1671, 1595. H NMR
(
5
1
CDCl
H, 1H
3
): δ 1.81-1.83 (m, 4H, -N(CH
, -CH -N<, -N(HCH-CH CH-); 2.51-2.99 (m, 5H, 1H
, -N(HCH-CH CH-); 3.04-3.21 (m, 2H, H , -N(HCH-
CH-); 6.4 (d, 1H, J ) 2.02, H ); 7.13 (t, 2H, J ) 6.6, o-F-
); 7.96 (dd, 2H, J ) 5.3, 2, m-F-Ph).
): δ 28.4 (C ); 29.1 (-N(CH -CH CH-); 29.2
CH-); 33.9 (C ); 43.1 (C ); 43.9 (-N(CH -CH CH-
; 53.4 (-N(CH -CH CH-); 54.6 (-N(CH -CH CH-); 63.4
-CH -N<); 106.8 (C ); 116.2 (d, 2C, J ) 21.8, o-F-Ph); 121.4
3a); 131.2 (d, 2C, J ) 9.3, m-F-Ph); 132.8 (p-F-Ph); 143.3 (C );
67.2 (C7a); 166.0 (d, 1C, J ) 250.7, C-F); 194.2 (-CO-); 201.4
2 2 2
-CH ) C-); 2.05-2.46 (m,
7
2
2
)
2
5
,
H
6
, 1H
7
2
)
2
5
CH
2
)
2
2
Ph); 7.3 (d, 1H, J ) 1.9, H
3
3
of 50% chloroacetaldehyde (1.01 mL), NaHCO (0.63 g, 7.53
1
3
C NMR (CDCl
-N(CH -CH
3
7
2
2 2
)
2
mmol) and water (5 mL) maintained at 0-5 °C. Stirring was
continued at room temperature for 12 h, AcOEt (5 mL) was
added, followed by enough 10% HCl to bring the mixture to
pH 1, and after stirring for a further 1 h the organic layer
was separated, washed with 10% K CO , dried (Na SO ) and
2 3 2 4
concentrated to dryness, affording 0.73 g of an oil that upon
column chromatography with 4:1 AcOEt/hexane as eluant gave
(
)
(
(
2
2
)
2
6
5
2 2
)
2
2
)
2
2
2 2
)
2
3
C
2
1
+
3
(-CO-). MS (FAB, m/z): 356 (MH ). Anal. (C21H22FNO ) C, H,
N. Hydrochloride: mp 244-245 °C (AcOEt).
6-[4-(6-F lu or oben zisoxa zol-3-yl)p ip er id in -1-ylm eth yl]-
4,5,6,7-tetr a h yd r oben zo[b]fu r a n -4-on e (20c): yield 85%,
0
.61 g of an oil identified as the hydroxy ketone 23: yield 60%.
1
IR: 3394, 2935, 1668, 1598. H NMR (CDCl
m, 1H, 1H , 1H
); 3.01 (dd, J ) 17.2, 4.8, H
-OH); 6.63 (d, 1H, J ) 1.9, H ); 7.31 (d, 1H, J ) 1.8, H
): δ 26.6 (C ); 38.4 (C ); 40.9 (C ); 65.8 (-CH
); 121.2 (C3a); 143.6 (C ); 167.3 (C7a); 194.8
-CO-). MS (FAB, m/z): 167 (MH ). Anal. (C ) C, H.
-(p-Tolu en esu lfon yl)oxym eth yl-4,5,6,7-tetr ah ydr oben -
3
): δ 2.37-2.43
); 2.75 (dd, 1H, J )
); 3.66-3.70 (m, 2H,
).
(
1
CH
5
); 2.50-2.60 (m, 2H, 1H
7.2, 9.4, 1H
5
7
1
7
6
mp 131-132 °C (i-PrOH). IR: 2927, 2739, 1665, 1610. H NMR
2
2
3
(CDCl
2.40-2.59 (m, 1H, 1H
CH CH-); 3.04-3.51 (m, 4H, 1H
(d, 1H, J C-F ) 1.9, H ); 7.11 (dt, 1H, J
3
): δ 2.03-2.38 (m, 8H, -CH
2
-N(CH
); 2.61-2.99 (m, 3H, 1H
, -N(HCH-CH
) 8.8, 2.5, H ); 7.23-
2
-CH
2
)
2
CH-, 1H
, -N(HCH-
CH-); 6.67
7 6
, H );
1
3
C NMR (CDCl
OH); 106.7 (C
3
7
6
5
2
-
7
5
3
2
2
)
2
5
2 2
)
+
(
9 10 3
H O
2
C-F
5′
6
3
7.26 (m, 1H, 1H7′); 7.33 (d, 1H, J ) 1.9, H ); 7.69 (dd, 1H, J C-F
1
3
zo[b]fu r a n -4-on e (24). p-Toluenesulfonyl chloride (2.3 g, 12
mmol) was added to a solution of alcohol 23 (1.6 g, 9.64 mmol)
in dry pyridine (30 mL) at 0 °C, and the mixture was stirred
at this temperature for 24 h. After addition of water (20 mL)
) 8.7, 5.1, H4′). C NMR (CDCl
3
): δ 28.5 (C
CH-); 33.9 (C
); 53.9 (-N(CH -CH
CH-); 63.5 (-CH
7
), 30.9 (-N(CH
); 34.9 (-N(CH
) CH-); 54.7 (-N(CH
2 2
2
2
2
-
-
-
CH
CH
CH
2
2
2
)
2
)
2
)
2
CH-); 31.4 (-N(CH
CH-); 43.0 (C
2 2 2
-CH )
6
5
2
2
-N<); 97.8 (d, J C-F ) 26.8, C7′); 106.8
and extraction with CH
water, dried over Na SO
ing 2 g of a brown dark oil that upon column chromatography
2
Cl
2
, the organic layer was washed with
(C
(d, J C-F ) 11.1, C4′); 143.3 (C
6′); 164.3 (d, J ) 13.8, C7a′); 167.2 (C7a); 194.3 (-CO-). MS (FAB,
3
); 112.7 (d, J C-F ) 25.3, C5′); 117.7 (C3a′); 121.5 (C3a); 122.9
2
4
and concentrated to dryness, afford-
2
); 161.5 (C3′); 164.5 (d, J ) 250.7,
C

4
688 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Ravi n˜ a et al.
F igu r e 8. Effects of reference drugs and 20c on striatal DA, DOPAC, and HVA levels (means ( SEM): (A) DA, only for clozapine
(
20 mg/kg) is the difference with respect to vehicle statistically significant (p < 0.01); (B) DOPAC, p < 0.01 with respect to vehicle
for all drugs; (C) HVA, p < 0.01 with respect to vehicle for all drugs.
+
136 °C (i-PrOH). IR: 2934, 2811, 1656, 1591. 1H NMR
(CDCl ): δ 2.27-2.33 (m, 1H, 1H ); 2.43-2.47 (m, 2H, 1H
1H ); 2.58-2.70 (m, 8H, 1H , 1H , -CH -N(CH -CH N-); 3.85
(s, 4H, -N(CH -CH N-); 3.85 (s, 3H, -OCH ); 6.66 (d, 1H, J )
2.01, H ); 6.91-6.99 (m, 4H, Ph); 7.32 (d, 1H, J ) 1.9, H
NMR (CDCl ): δ 28.5 (C ); 33.7 (C ); 43.0 (C ); 51.1 (-N(CH
CH N-); 54.2 (N(CH -CH N-); 55.7 (-OCH ); 63.3 (-CH -N<);
106.8 (C ); 111.6 (C6′); 118.6 (C3′); 121.4 (C5′); 121.5 (C3a); 123.3
(C4′); 141.7 (C1′); 143.2 (C ); 152.7 (C2′); 167.2 (C7a); 194.2 (-CO-
). MS (FAB, m/z): 341 (MH ). Anal. (C20
m/z): 369 (MH ). Anal. (C21
H
21FN
2
O
3
) C, H, N. Hydrochlo-
ride: mp 271-272 °C (AcOEt). Anal. (C21
H
21FN
2
O
3
‚HCl) C,
3
5
5
,
H, N.
6
7
7
6
2
2
2 2
)
-(4-P h en ylpiper azin -1-ylm eth yl)-4,5,6,7-tetr ah ydr oben -
2
2
)
2
3
1
3
zo[b]fu r a n -4-on e (20d ): yield 75%, mp 121-122 °C (i-PrOH).
2
3
).
C
2
-
1
IR: 3852, 1673. H NMR (CDCl
3
): δ 2.24-2.62 (m, 9H, 2H
-N<); 2.68 (dd, 1H, J ) 15, 3.6,
); 3.08-3.15 (m, 1H, 1H ); 3.19 (t, 4H, J ) 5, -N(CH
N-); 6.67 (d, 1H, J ) 2, H ); 6.83-6.94 (m, 3H, o-Ph,
p-Ph); 7.23-7.29 (m, 2H, m-Ph); 7.3 (d, 1H, J ) 2, H
NMR (CDCl ): δ 28.5 (C ); 33.7 (C ); 43.0 (C ); 49.6 (-N(CH
CH N-); 53.9 (-N(CH -CH N-); 63.2 (-CH -N<); 106.8 (C
16.8 (C4′); 120.1 (C3′); 121.5 (C3a); 129.5 (C2′); 143.3 (C ); 151.6
7a); 167.1 (C1′); 194.2 (-CO-). MS (FAB, m/z): 311 (MH ).
Anal. (C19 ): C, H, N. Hydrochloride: mp 198-199 °C
AcOEt). Anal. (C19 ‚2HCl‚H O) C, H, N.
-[4-(o-Met h oxyp h en yl)p ip er a zin -1-ylm et h yl]-4,5,6,7-
tetr a h yd r oben zo[b]fu r a n -4-on e (20e): yield 70%, mp 135-
5
,
3
7
6
5
1
1
H
H
7
, -N(CH
2
-CH
2
)
2
N-, -CH
2
2
)
2
2
2
)
2
3
2
7
6
2
-
3
CH
2
)
2
2
2
1
3
+
3
).
C
24 2 3
H N O ) C, H, N.
3
7
6
5
2
-
24 2 3
Hydrochloride: mp 201-203 °C (AcOEt). Anal. (C20H N O ‚
2HCl) C, H, N.
6-[4-(2-P yr id yl)p ip er a zin -1-ylm et h yl]-4,5,6,7-t et r a h y-
2
)
2
2
2
)
2
2
3
);
1
2
+
(
C
d r oben zo[b]fu r a n -4-on e (20f): yield 80%, mp 108-109 °C
H
22
N
2
O
2
(i-PrOH). IR: 2826, 1677, 1592. 1H NMR (CDCl
3
): δ 2.24-
N-); 2.67 (dd, 1H, J
), 3.51 (t, 4H, J ) 5.1,
N-); 6.61 (d, 1H, J ) 2.2, H ); 6.63-6.66 (m, 2H,
(
H
22
N
2
O
2
2
2.64 (m, 9H, 2H
) 13.4, 4.5, 1H
-N(CH -CH
5
, 1H
7 2 2 2 2
, -CH -N(CH -CH )
6
7
); 3.04-3.15 (m, 1H, H
6
2
2
)
2
2

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4689
121.4 (C3a); 129.9 (p-F-Ph); 130.8 (d, 2C, J C-F ) 9.3, o-F-Ph);
H
H
3
, H5′); 7.32 (d, 1H, J ) 2.0, H
4′); 8.17 (dd, 1H, J ) 4.8, 1.2, H6′). C NMR (CDCl
); 33.6 (C ); 42.9 (C ); 45.6 (-N(CH -CH N-); 53.7 (-N(CH
CH N-); 63.3 (-CH -N<); 106.8 (C ); 107.4 (C3′); 113.7 (C5′);
21.5 (C3a); 137.8 (C4′); 143.3 (C ); 148.34 (C6′); 159.9 (C2′); 167.1
7a); 194.1 (-CO-). MS (FAB, m/z): 312 (MH ). Anal.
3
); 7.46 (dd, 1H, J ) 8.5, 5.1,
13
3
): δ 28.5
143.2 (C
); 198.9 (-CO-Ph). Anal. (C23
ride: mp 202-204 °C (AcOEt).
-{4-[5-(4-Fluorophenyl)-5-oxopentyl]piperazin-1-ylmeth-
2
); 165.9 (d, J C-F ) 251, C-F); 167.1 (C7a); 194.2 (-CO-
(C
7
6
5
2
2
)
2
2
-
2 3
H27FN O
) C, H, N. Hydrochlo-
2
)
2
2
3
1
2
6
+
(
(
C
yl}-4,5,6,7-tetr a h yd r oben zo[b]fu r a n -4-on e (20i). A solution
C
18
H
21
N
3
O
2
) C, H, N. Hydrochloride: mp 128-130 °C (AcOEt).
94
of 5-bromo-1-(p-fluorophenyl)-1-pentanone (0.35 g, 1.37 mmol)
6
-[4-(2-P yr im id in yl)p ip er a zin -1-ylm et h yl]-4,5,6,7-t et -
in methyl isobutyl ketone (15 mL) was added under argon,
r a h yd r oben zo[b]fu r a n -4-on e (20g): yield 70%, mp 112-113
with stirring, to a mixture of piperazine 26 (0.32 g, 1.37 mmol),
anhydrous Na CO (0.43 g) and KI (0.1 g) in 15 mL of the same
2 3
solvent, and the mixture was refluxed for 24 h. The precipitate
was filtered out, and removal of the solvent from the filtrate
under reduced pressure left 0.56 g of a dark brown oil that
1
°
C (i-PrOH). IR: 2826, 1677, 1592. H NMR (CDCl
3
): δ 2.23-
N-); 3.04-3.14 (m,
2 2
) N-); 6.46 (t, 1H, J
2
1
)
8
3
CH
1
)
.69 (m, 10H, 2H
H, H ), 3.80 (t, 4H, J ) 5.1, -N(CH
4.7, H5′); 6.65 (d, 1H, J ) 2.0, H ); 7.32 (d, 1H, J ) 2.0, H
.28 (d, 2H, J ) 4.7, H6′, H4′). C NMR (CDCl ): δ 28.5 (C
3.7 (C ); 43.0 (C ); 44.1 (-N(CH -CH
N-); 63.3 (-CH -N<); 106.8 (C ); 110.2 (C5′); 121.5 (C3a);
5
, 2H
7
, -CH
2
-N(CH
2
-CH
2
)
2
6
2
-CH
2
3
);
);
1
3
3
7
2 2 3
upon column chromatography with 19:1 CH Cl /CH OH as
6
5
2
2
)
2
N-); 53.8 (-N(CH
2
-
eluent afforded amino ketone 20i (0.28 g, 50%) as a white
1
2
)
2
2
3
solid: mp 92-94 °C (i-PrOH). IR: 2810, 1682, 1669. H NMR
43.3 (C
2
); 158.1 (C4′, C6′); 162.0 (C2′); 167.0 (C7a); 194.1 (-CO-
(CDCl ): δ 1.56-1.63 (m, 2H, -CH -CH -CH -CO-); 1.70-1.80
3
2
2
2
+
. MS (FAB, m/z): 313 (MH ). Anal. (C17
H
20
N
4
O
2
) C, H, N.
(m, 2H, -CH -CH -CH -CO-); 2.20-2.67 (m, 16H, H , 1H , 2H ,
2
2
2
5
6
7
Hydrochloride: mp 241-243 °C (AcOEt). Anal. (C17
HCl) C, H, N.
-[4-(ter t-Bu toxycar bon yl)piper azin -1-ylm eth yl]-4,5,6,7-
tetr a h yd r oben zo[b]fu r a n -4-on e (25): yield 70%, mp 86-
H
20
N
4
O
2
‚
-CH -N(CH -CH ) N-CH -); 2.96 (t, J ) 7.2, 2H, -CH -CO-Ph);
2
2
2
2
2
2
2
3.01-3.08 (m, 1H, 1H ); 6.65 (d, J ) 2.0, 1H, H ); 7.12 (t, J )
5
2
6
8.6, 2H, H
3
Ph); 7.32 (d, 1H, H
3
); 7.98 (dd, J ) 8.9, 5.4, 2H, H
2
Ph). 13C NMR (CDCl
3
2 2 2
): δ 22.7 (-CH -CH -CH -CO-); 26.8
8
1
1
3
7
7 °C (i-PrOH). IR: 2918, 1693, 1681. 1H NMR (CDCl
.45 (s, 9H, 3x -CH ); 2.30-2.38 (m, 7H, -N(CH -CH N-, 2H
); 2.40-2.67 (m, 3H, -CH N<, 1H ); 3.03-3.08 (m, 1H, H
.40 (t, 4H, J ) 5,.N(CH -CH N-); 6.65 (d, 1H, J ) 2, H
). C NMR (CDCl ): δ 28.4 (C
); 42.9 (C ); 44.2 (-N(CH -CH N-); 53.7
N-); 63.2 (-CH N<); 80.1 (-C(CH ); 106.8 (C );
21.5 (C3a); 143.3 (C ); 155.1 (-COO-); 166.9 (C7a); 194.1 (-CO-
. MS (FAB, m/z): 335 (MH ). Anal. (C18
-P ip er a zin -1-ylm eth yl-4,5,6,7-tetr a h yd r oben zo[b]fu -
r a n -4-on e (26). Trifluoroacetic acid (2.87 mL, 37.4 mmol) was
added to a solution of amine 25 (1.25 g, 3.74 mmol) in dry CH
Cl (10 mL), the mixture was stirred at room temperature for
0 min, and the solvent was then removed under reduced
pressure. The residue was dissolved in CH Cl , and this
solution was washed with 10% NaHCO
(2 × 20 mL). The
organic phase was dried over Na SO , and the solvent was
3
): δ
(-CH -CH -CH -CO-); 28.5 (C ); 33.7 (C ); 38.6 (-CH -CO-Ph);
2
2
2
7
6
2
)
2
5
,
43.0 (C
5
); 53.6 (2C, -N(CH
2
-CH
2
)
2
N-); 53.8 (2C,-N(CH
2
-CH
2
)
2
N-
3
2
2
H
7
2
7
6
2
);
);
); 58.6 (RRN-CH
C-F ) 21.8, o-F-Ph); 121.5 (C3a); 131.0 (d, 2C, J C-F ) 9.4, m-F-
Ph); 133.8 (d, 2C, J C-F ) 2.9, p-F-Ph); 143.2 (C ); 165.9 (d, J C-F
) 251, C-F); 167.1 (C7a); 194.2 (-CO-); 198.9 (-CO-Ph). Anal.
) C, H, N. Hydrochloride: mp 235-240 °C
(AcOEt). Anal. (C24 ‚HCl‚H O) C, H, N.
2 2 3
-); 63.2 (-CH -NRR); 106.8 (C ); 116.0 (d, 2C,
)
2
J
2
2
1
3
.31 (d, 1H, J ) 2, H
); 33.6 (C
-CH
3
3
7
); 28.8
2
(
(
3x -CH
-N(CH
3
6
5
2
2 2
)
)
2
2
3
)
3
3
2 3
(C24H29FN O
2
2
1
)
2
H
29FN
2
O
3
2
+
H
26
N
2
O
4
) C, H, N.
P h a r m a cologica l Test Meth od s. D1 a n d D2 Bin d in g
Assa ys. Male Sprague-Dawley rats were killed by decapita-
tion and their brains were rapidly removed and dissected on
an ice-cold plate. Striatal membrane preparations were ob-
tained by homogenization (Polytron homogenizer, setting 6,
10 s) in 50 mM Tris-HCl (pH 7.7 at 25 °C; about 100 µL/mg of
tissue) containing 5 mM EDTA; the homogenates were cen-
trifuged (49000g for 15 min at 4 °C; Sorvall RC-26 plus),
resuspended in 50 mM Tris-HCl buffer (pH 7.4 at 25 °C) and
centrifuged again (same conditions), and the final pellets were
stored at -80 °C pending use. All drugs were stored in 1 mM
solutions at -20 °C and diluted to the required concentrations
on ice immediately before binding assays. J ust before binding
assays, the striatal membrane pellets were resuspended (1.25
6
2
-
2
2
2
2
3
2
4
removed in vacuo, affording 26 (0.65 g, 75%) as a white
crystalline solid: mp 128-130 °C (i-PrOH). IR: 3342, 2938,
1
1
681, 1596. H NMR (CDCl
3
): δ 2.25-2.53 (m, 10H, 2H
7
, 2H
5
,
-CH
2
-N(CH
.93 (m, 1H, H
.25 (s, 1H, -NH). 13C NMR (CDCl
2.9 (C ); 43.8 (-N(CH -CH NH); 54.0 (-N(CH
3.7 (-CH -N<); 106.8 (C ); 121.4 (C3a); 142.3 (C ); 165.7 (C7a);
98 2 (-CO-). MS (FAB, m/z): 235 (MH ). Anal. (C13
2
-CH
); 6.4 (d, 1H, J ) 2, H
): δ 28.4 (C
2
)
2
NH); 2.75 (t, 4H, J ) 4.9, -N(CH
2
-CH
2
)
2
NH);
2
8
4
6
1
6
2
); 7.3 (d, 1H, J ) 2, H
); 33.3 (C
-CH
3
6
);
);
mg original wet weight/750 µL for D
for D ) in 50 mM Tris-HCl buffer (pH 7.4 at 25 °C) containing
120 mM NaCl, 5 mM KCl, 2 mM CaCl and 1 mM MgCl . For
2
assays, 1.00 mg/750 µL
3
7
5
2
2
)
2
2
2
)
2
NH);
1
2
3
2
2
2
+
H
18
N
2
O
2
)
D
2
binding assays, 750-µL aliquots of striatal membrane
C, H, N. Hydrochloride: mp 213-214 °C (AcOEt).
preparation were added to ice-cold tubes containing (a) 100
µL of [ H]spiperone, (b) 50 µL of ketanserin (final concentration
3
6
-{4-[4-(4-Flu or oph en yl)-4-oxobu tyl]piper azin -1-ylm eth -
5
0 nM) to block 5-HT2A receptors, and (c) either 100 µL of
yl}-4,5,6,7-tetr a h yd r oben zo[b]fu r a n -4-on e (20h ). A solu-
tion of 4-chloro-1,1-ethylenedioxy-1-(4-fluorophenyl)butane (0.55
g, 2.23 mmol) in methyl isobutyl ketone (15 mL) was added
under argon, with stirring, to a solution of the amine 26 (0.5
buffer (for total binding assay) or 100 µL of sulpiride (final
concentration 10 µM) to allow quantification of unspecific
3
binding of [ H]spiperone or 100 µL of the compounds to be
1
tested. For D binding assays, the same procedure was followed
g, 2.13 mmol), anhydrous Na
the same solvent (45 mL), and the mixture was refluxed for
6 h. The precipitate was filtered out, the solvent was removed
2 3
CO (0.65 g) and KI (0.17 g) in
3
3
except that [ H]spiperone was replaced by [ H]SCH23390,
ketanserin by buffer, and sulpiride by nonradiolabeled
SCH23390 (final concentration 1 µM) to allow quantification
3
from the filtrate under reduced pressure, and a solution of the
resulting residue in 10% HCl was refluxed for 1 h (100 °C).
Once cool, this solution was brought to basic pH with 10%
NaOH and extracted with AcOEt (3 × 50 mL). The organic
3
of nonspecific binding by [ H]SCH23390. The final assay
volume was thus 1 mL in all cases. All assays were performed
in duplicate. Incubations (15 min at 37 °C) were stopped by
rapid vacuum filtration through GF-52 glass fiber filters
2 4
phase was dried over Na SO , and removal of the solvent under
(Schleicher and Schuell) in a Brandel M-30 cell harvester. The
reduced pressure left 0.88 g of a dark brown oil that upon
column chromatography with 1:1 AcOEt/hexane as eluant
afforded 20h (0.35 g, 42%) as a white solid: mp 112-114 °C
filters were rinsed three times with 3 mL of ice-cold 50 mM
Tris-HCl buffer (pH 7.4), and radioactivity was determined by
liquid scintillation counting in a Beckman LS-6000LL ap-
paratus.
1
(
1
-
CH
(
i-PrOH). IR: 2810, 1682, 1669. H NMR (CDCl
3
): δ 1.85-
, 2H
, -N(CH
); 7.29-7.34
Ph); 7.84-7.89 (m, 2H, H
): δ 22.6 (-CH -CH -CH -CO-); 28.4
-CO-Ph); 46.2 (C ); 46.5 (2C, -N(CH
N-); 52.4 (2C,-N(CH -CH N-); 57.6 (RRN-CH -); 63.2
-NRR); 106.7 (C ); 115.8 (d, 2C, J C-F ) 21.8, m-F-Ph);
.92 (m, 2H, -CH
2
-CH
N-CH
-CO-Ph, H
); 7.56-7.63 (m, 2H, H
3
2
-CH
-); 2.83-2.98 (m, 8H, 1H
); 6.96-7.02 (m, 1H, H
2
-CO-); 2.34-2.52 (m, 11H, H
5
7
,
CH
2
-N(CH
2
-CH
2
)
2
2
6
2
-
D
4
Bin d in g Assa ys. Radioreceptor binding studies with
3
2
)
2
N-, -CH
2
5
2
[ H]YM-09-151-2 (emonapride, a D -like receptor antagonist,
2
m, 1H, 1H
3
3
2
81-87 Ci/mmol) were performed in membrane preparations
from bovine retina. Retinae were homogenized in a saline
1
del Ph). C NMR (CDCl
); 36.2 (C ); 38.6 (-CH
CH
-CH
3
2
2
2
(C
7
6
2
5
2
-
solution (1:20 w/v) containing in mM: Tris, 50; EDTA, 1; CaCl
1.5; MgCl , 4; KCl, 5; NaCl, 120; pH 7.4. The homogenate was
centrifuged at 48000g for 20 min at 4 °C, resuspended 1:20,
2
,
2
)
2
2
2
)
2
2
2
(
2
3

4
690 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Ravi n˜ a et al.
recentrifuged and resuspended at a final dilution of about 4
mg original wet tissue/mL of saline (1:250 w/v) for use in the
binding assay. For competition experiments, membranes were
of pA
2
(-log concentration of antagonist required to maintain
a constant response when the agonist concentration is doubled).
Beh a vior a l Assa ys. Mice weighing 25 ( 5 g and rats
weighing 250 ( 50 g were housed in groups of 12 (mice) or 4
(rats) under regulated conditions (light/dark cycle between 8.00
and 20.00 h at 21 ( 2 °C) in standard Makrolon cages (215 ×
465 × 145 mm). The animals received standard laboratory
chow and tap water ad libitum until the beginning of the
experiments.
Locom otor Activity. The locomotor activity of animals in
square arenas (50 × 50 × 30 cm) under a video camera
suspended from the ceiling was recorded over a 1-h period
between 10.00 and 14.00 h and analyzed, using a computerized
animal observation system (EthoVision V. 1.90, Noldus Infor-
mation Technology, Wageningen, The Netherlands) located in
a separated room. The variables considered were distance
travelled (in cm), rearing (reduction of vertical projection area
by 15% or more), and velocity.
3
incubated at 30 °C in the presence of 0.1 nM [ H]YM-09-151-
3
2
for 60 min. For [ H]YM-09-151-2 binding, duplicate test tubes
contained in a final volume of 1.5 mL: 0.5 mL membranes,
3
0
.15 tracer, and drug. The affinity of [ H]YM-09-151-2 was
measured by saturation experiments in similar conditions and
was 0.4 nM. Nonspecific binding was assessed by 2 mM DA.
At the end of the incubation period, the radioactivity bound
to the receptor was separated from the free ligand by rapid
vacuum filtration through glass fiber filters (GF/C) in a
Brandel 30-well filtration apparatus. Filters were counted by
liquid scintillation spectroscopy in a Packard LS-1600 ap-
paratus and converted from cpm to dpm.
5
-HT2A Bin d in g Assa ys. Male 200-250-g Sprague-Daw-
ley rats were asphyxiated with CO and decapitated. The
frontal cortex, containing 5-HT2A receptors,9
free on ice, frozen on dry ice and stored at -70 °C until use
generally less than 1 week later). All membrane preparation
2
5,96
was dissected
Am p h eta m in e-In d u ced Hyp er m otility. Animals were
treated with 5 mg/kg dextroamphetamine sulfate 30 min after
pretreatment with vehicle, haloperidol, clozapine, risperidone,
20b or 20c, and each animal was then immediately placed in
its experimental arena.
(
procedures were carried out at 4 °C. The tissue was thawed
on ice and homogenized with 10 volumes of 0.32 M sucrose in
a Polytron homogenizer (Brinkmann Instruments, Westbury,
NY). The homogenate was centrifuged twice at 4 °C (900g for
Ap om or p h in e-In d u ced Clim bin g. The method of Protais
et al.1 was used, with a minor modification. Vehicle, halo-
peridol, clozapine, risperidone, 20c (0.25-0.5 mg/kg) or 20b
(0.6-4 mg/kg) was administered to mice (6 for drug and
dosage, 12 for vehicle) that 30 min later were treated subcu-
taneously with 2 mg/kg apomorphine. The mice were then
placed in individual cylindrical wire cages (diameter, 12 cm;
height, 14 cm), and over the next 30 min their climbing activity
was recorded every 10 min using the following scale: 0, four
paws on the floor; 1, one or two paws against the cage wall; 2,
three or four paws used to cling to the grid wall. The scores
recorded 20 and 30 min post-apomorphine were added for use
in further calculations.
01
1
0 min followed by 40000g for 30 min), the supernatant was
discarded, and the pellet was resuspended in Tris HCl-buffer
pH 8.07) in a Teflon/glass homogenizer (10 strokes by hand).
(
This suspension was incubated at 37 °C for 15 min to remove
endogenous 5-HT and centrifuged for 30 min at 40000g, and
the final pellet was resuspended in Tris-HCl buffer of pH 8.07
2
containing 4 mM CaCl and 0.1% ascorbic acid. Competition
3
at [ H]ketanserin binding sites was assayed in triplicate in
assay mixtures consisting of 750 µL of membrane homogenate,
3
5
0 µL of [ H]ketanserin, 50 µL of either buffer or the compound
under test, 50 µL of masking ligand solution (1 µM methyser-
gide) as required, and buffer to a final volume of 1 mL.
Mixtures were incubated for 30 min at 37 °C. The assay was
terminated by rapid filtration through Whatman GF/C filter
strips (presoaked in 3% polyethylenimine) in a Brandel cell
harvester (Gaithersburg, MD) followed by washing with ice-
cold Tris-HCl buffer (pH 6.6) to remove unbound radioligand.
The radioactivity retained on filters was determined by liquid
scintillation counting in a beta counter (Beckman, LS-6000LL).
Ca ta lep sy. Thirty minutes after administration of vehicle,
haloperidol, clozapine, risperidone, 20c (0.5-8 mg/kg) or 20b
(0.5-6 mg/kg), mice (n ) 6-12) were placed with their
forepaws on one horizontal wire and their hindpaws on another
6
cm away and 2 cm lower. The time during which the mouse
maintained this position was recorded; maintenance for more
than 30 s was considered to indicate catalepsy.
Neu r och em ica l Assa ys. Sprague-Dawley rats were killed
by decapitation 120 min after injection of vehicle, reference
drugs haloperidol (0.5 mg/kg), clozapine (20 mg/kg), risperi-
done (10 mg/kg) or 20c (10 mg/kg). Within 1 min of decapita-
tion the brain was removed and the striatum dissected free
5
-HT2C Bin d in g Assa ys. Bovine choroid plexus containing
9
7
5
-HT2C receptors was treated as described in the 5-HT2A
binding assay. A suspension of the resulting pellet in the same
buffer was stored on ice while not being manipulated. Com-
petition at [ H]mesulergine binding sites was determined by
a protocol analogous to that described above for the 5-HT2A
binding assay, using a final [ H]mesulergine concentration of
nM and 1 mM mianserine as 5-HT2A masking ligand. The
mixtures were incubated for 1 h at room temperature. Mem-
branes were harvested on Whatman GF/B filters.
3
1
02
on an ice-cold plate as described by Glowinski and Iverssen.
Samples of striatal tissue were weighed in 1.5-mL conical test
3
tubes and 20 µL/mg of PCA solution (0.1 M perchloric acid
2
-5
and 4 × 10 M sodium bisulfite) was then added. The mixture
was homogenized (Ultra-turrax T8, position 5, 10 s), and the
homogenate was centrifuged twice for 15 min at 13 000 rpm
F u n ction a l Exp er im en ts. Male Sprague-Dawley rats
250-300 g) were asphyxiated with CO and decapitated. The
2
stomach was dissected free from the abdomen and immersed
in modified Krebs solution of the following composition (mM):
NaCl, 119; KCl, 4.7; MgSO
(Heraeus Biofugue-13). The supernatant was filtered (Millipore
(
HV 0.45 µm) and stored at -80 °C prior to HPLC, which was
performed using 20-µL samples, a Tracer RP-18 column (15
×
0.4 cm, 5 µm Spherisorb), a Waters pump 616 pump with a
4
‚7H
2
O, 1.2; CaCl
2
‚2H
2 2
O, 2.5; KH -
6
00S controller, a Waters 717 plus autosampler refrigerated
PO , 1.18; NaHCO , 25; glucose, 11. Strips of stomach fundus
4
3
at 4 °C and a Coulochem 5100 A detector with a 5021
conditioning cell and a 5011 analytical cell. The mobile phase
9
8
were prepared by Vane’s method and mounted in organ baths
containing 10 mL of the same Krebs solution as above,
maintained at 37 °C and aerated with carbogen (95% O
(
an 88:12 mixture of citrate buffer (pH 4.0) and methanol
containing 0.83 mM sodium octanesulfonate) was filtered
0.22-µm pore size) and degassed with ultrasound before use.
2
, 5%
CO ). Before addition of drugs, the tissue strips were equili-
2
(
brated for 1 h under a 1-g load. Isometric contractions were
recorded during cumulative addition of serotonin using a Grass
FTO3C transducer and a Grass 7D polygraph.
The flow rate was 1 mL/min and ambient temperature was
maintained. HPLC operations were controlled and the detec-
tion data processed using Waters Millenium software (V 2.10).
3
3
Concentration-response curves for serotonin were con-
Dr u gs a n d Ch em ica ls. [ H]Spiperone (104 Ci/mmol), [ H]-
9
9
3
structed as per Van Rossum. In the initial control runs, stable
contractions were achieved over the concentration range 0.01
nM-10 µM. Following the initial control run, each tissue strip
was run alternately with and without antagonist. Between
runs, the tissues were washed and allowed to rest for 60 min.
SCH23390 (81 Ci/mmol) and [ H]mesulergine (76 Ci/mmol)
were obtained from Amersham International (England), [ H]-
3
ketanserin (60.08 Ci/mmol) from DuPont NEN (Boston, MA),
3
[ H]YM-09-151-2 from New England Nuclear (Boston, MA),
unlabeled (R)-(+)-SCH23390‚HCl, mianserine and methyser-
gide from Research Biochemicals Inc., and unlabeled spiper-
1
00
Antagonist potency was measured as per Mackay in terms

Conformationally Constrained Butyrophenones
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4691
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the Xunta de Galicia Grants XUGA20308B94,
XUGA20308B96, and XUGA20319B97. M.E.R. was
aided by a predoctoral grant from Xunta de Galicia and
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