Design, synthesis and pharmacological evaluation of new series of naphthalenic analogues as melatoninergic (MT1/MT2) and serotoninergic 5-HT2C dual ligands (I)

Article abstract of DOI:10.1016/j.ejmech.2012.01.027

As part of our ongoing interest in developing new melatoninergic ligands bearing the same pharmacological profile as agomelatine, we focused our attention on this compound as a lead. Several chemical modifications have been performed on positions C-3 and 8 of the naphthalene ring determined as primary targets for the agomelatine metabolism. Herein we report the modulation of the positions C-3 and 7 in addition of the amide side chain because of this later prominent role in the affinity profile of such ligands. Synthesized compounds were then biologically evaluated at human cloned melatoninergic and serotoninergic receptors and showed different binding affinity and intrinsic activity profiles. Compounds bearing fluoroacetamide group (compounds 4 and 5) showed a high melatoninergic binding affinity particularly towards MT ₁ receptor subtype. Thus, the fluoroacetamide 4 exhibited a good melatoninergic (MT₁/MT₂) binding affinity (70 pM) higher than the lead. Moreover, other compounds (10a, 10e, 16, 17 and 18) issued from these modulations behaved as MT₁ and MT₂ agonists and exhibited a sub-nanomolar binding affinity towards these receptors. However, only compounds 10e, 17 and 18 showed a sub-nanomolar binding affinity at 5-HT_2C higher than the agomelatine.

Full text of DOI:10.1016/j.ejmech.2012.01.027

European Journal of Medicinal Chemistry 49 (2012) 310e323
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Design, synthesis and pharmacological evaluation of new series of naphthalenic
analogues as melatoninergic (MT1/MT2) and serotoninergic 5-HT2C dual ligands (I)
,
b
,
a
,
,
Mohamed Ettaoussi a , Ahmed Sabaouni b, Marouan Rami a b, Jean A. Boutin d, Philippe Delagrange c,
*
Pierre Renard c, Michael Spedding c, Daniel-Henri Caignard c, Pascal Berthelot a b, Saïd Yous a
,
,b
a Univ Lille Nord de France, F-59000 Lille, France
b UDSL, EA GRIIOT, UFR Pharmacie, 3 rue du Professeur Laguesse, 59000 Lille, France
c Département des Sciences Expérimentales, Institut de Recherches Servier, 92150 Suresnes, France
d Institut de Recherches Servier, 78290 Croissy-sur-Seine, France
a r t i c l e i n f o
a b s t r a c t
Article history:
As part of our ongoing interest in developing new melatoninergic ligands bearing the same pharma-
cological profile as agomelatine, we focused our attention on this compound as a lead. Several chemical
modifications have been performed on positions C-3 and 8 of the naphthalene ring determined as
primary targets for the agomelatine metabolism. Herein we report the modulation of the positions C-3
and 7 in addition of the amide side chain because of this later prominent role in the affinity profile of
such ligands. Synthesized compounds were then biologically evaluated at human cloned melatoninergic
and serotoninergic receptors and showed different binding affinity and intrinsic activity profiles.
Compounds bearing fluoroacetamide group (compounds 4 and 5) showed a high melatoninergic binding
affinity particularly towards MT1 receptor subtype. Thus, the fluoroacetamide 4 exhibited a good mel-
atoninergic (MT1/MT2) binding affinity (70 pM) higher than the lead. Moreover, other compounds
(10a, 10e, 16, 17 and 18) issued from these modulations behaved as MT1 and MT2 agonists and exhibited
a sub-nanomolar binding affinity towards these receptors. However, only compounds 10e, 17 and 18
showed a sub-nanomolar binding affinity at 5-HT2C higher than the agomelatine.
Received 22 November 2011
Received in revised form
10 January 2012
Accepted 12 January 2012
Available online 20 January 2012
Keywords:
Agomelatine
Agonist
Antagonist
Depression
5-HT2C
MT1
MT2
Ó 2012 Elsevier Masson SAS. All rights reserved.
1. Introduction
Search for new antidepressants has led to the discovery and
development of the agomelatine (Valdoxan), the first melatoni-
According to the US National Institute of Mental Health, anxiety
and depression are the most prevalent mental illnesses and consti-
tute a serious public health problem [1]. Depression is a major
contributor to the global disease burden but since the discovery of
their mechanisms, its treatments have been revolutionized.
Currently, monoaminergic compounds such as selective serotonin
reuptake inhibitors (SSRI), serotonin and noradrenaline reuptake
inhibitors (SNRI), and noradrenaline reuptake inhibitors (NRI) are
the most used antidepressants [2]. However, incomplete remission
(only 50%), action delay, side effects and poor tolerability [3] chal-
lenges remain until now and require the discovery of new generation
of medications with new mechanisms of action and low side effects.
nergic antidepressant (Chart 1). In February 2009, the agomelatine,
a novel antidepressant with an innovative pharmacological profile,
had received a marketing authorization by the European Medicines
Agency for the treatment of major depressive disorders [4]. It acts as
an agonist of the melatoninergic (MT1/MT2) receptors and antago-
nist of the serotoninergic 5-HT2C receptor [4e6]. Its antidepressant
properties have been demonstrated in animal models and clinical
studies, and showed an effect superior to placebo in the treatment of
depressed patients [7e9]. At a dose of 25 mg once daily, Valdoxan
has similar efficacy to paroxetine and venlafaxine [10e12].
The duality agonist MT1/MT2 and antagonist 5-HT2C suggests
that its antidepressant activity may be due to this combination or to
each of these properties. In addition, it might be involved in the
modulation of circadian rhythms, changes in sleepewake cycle
[12,13] and particularly effective for the treatment of seasonal
affective disorders, bipolar depression or other mood disorders
[14]. Although further studies are necessary in order to better
understand the various functions exerted by this antidepressant
and its mechanisms of action.
Abbreviations: MLT, melatonin; MT1, melatonin receptor subtype 1; MT2,
melatonin receptor subtype 2; 5-HT2C, serotonin receptor subtype 2C; [35S]GTP
gS,
[
35S]guanosine-50-O-(3-thio-triphosphate).
* Corresponding author. UDSL, EA GRIIOT, UFR Pharmacie, 3 rue du Professeur
Laguesse, 59000 Lille, France. Tel.: þ33 3 2096 4255; fax: þ33 3 2096 4913.
E-mail address: m.ettaoussi@yahoo.fr (M. Ettaoussi).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.01.027

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
311
6. The nucleophilic substitution of bromine atom of compound 2
was performed by treatment with N-methylbenzylamine and led to
compound 7 which was then debenzylated using hydrogen and
palladium charcoal into 8. Finally, acylated compounds 9aei were
prepared by treatment of the compound 1 by the appropriate acyl
chlorides under Friedel-Craft conditions [17]. Selective reduction of
the ketone groups by triethylsilane in trifluoroacetic acid [18] led to
the derivatives 10aei.
NHCOCH3
NHCOCH3
CH3
O
CH3
O
N
H
Melatonin
Agomelatine (1)
The results of our studies on bromination of compound 1 are
depicted in Scheme 2. Thus, its treatment at room temperature
with bromine in glacial acetic acid or with bromosuccinimide (NBS)
in carbon tetrachloride at reflux led exclusively to the formation of
the 8-brominated compound 11. Meanwhile, heat of this later at
70 ꢀC in glacial acetic acid furnished the 3-bromoderivative 12 by
migration of the bromine from C-8 position to C-3. Besides, the
compound 12 was exclusively obtained by heating the agomelatine
at 70 ꢀC in glacial acetic acid for 3 h in the presence of bromine.
The synthetic pathway for compounds 13e21 is outlined in
Scheme 3. First, the preparation of compounds 13 and 14 was
achieved by treatment of 3-bromoderivative 12 with respectively
morpholine and 4-methoxybenzylamine in the presence of palla-
dium dibenzylideneacetone and xantphos [19]. The catalytic
hydrogenolysis of the obtained compound 14 afforded then
compound 15 which was transformed into 3-fluoroagomelatine 16
in one-pot. Indeed, diazotization of compound 15 was accom-
plished by reaction with sodium nitrite in an acid medium followed
by nucleophilic substitution of the diazonium group over fluo-
roboric acid at reflux in xylene [20]. Otherwise, the 3-
bromoderivative 12 was converted to vinyl 17 and allyl 18 under
Stille conditions in the presence of tributylvinyltin or allyl-
tributyltin and tetrakis (triphenylphosphine) palladium. The alde-
hyde 19 was obtained in a good yield (72%) from the vinyl 17
according to the oxidative cleavage of Lemieux-Johnson by treat-
ment with osmium oxide in the presence of sodium periodate [21].
Therefore, condensation of the aldehyde 19 with hydroxylamine
Chart 1. Chemical structures of melatonin and agomelatine.
In our continuing efforts to develop new melatoninergic ligands
particularly bearing the same pharmacological profile as agome-
latine (MT1/MT2 agonist and 5-HT2C antagonist), we have intro-
duced several chemical modifications on positions known as
primary targets for the agomelatine metabolism, namely positions
C-3 and 8 of the naphthalene ring. Herein we report modulations
performed in positions C-3 and 7 of the cyclic core and the amide
side chain because of this later prominent role as a major factor for
the affinity profile of such ligands.
2. Results and discussion
2.1. Chemistry
In Scheme 1 is shown the synthetic route applied for the
preparation of compounds 1e10. First, the starting material 2-(7-
methoxy-naphthalen-1-yl)ethylamine hydrochloride (A) was
prepared as previously described [15] and N-acylated according to
a variant of the SchotteneBaumann reaction [16] to lead to deriv-
atives 1e3. Fluorinated derivatives 4 and 5 were prepared by
reaction of the free amine of compound A with respectively
ethyl fluoroacetate or methyl difluoroacetate in the 2,2,2-
trifluoroethanol. Otherwise, selective reduction of the ester func-
tion of compound 3 by lithium aluminum hydride led to compound
NHCOCH2(N)CH3Bz
CH3
NHCOCH2NHCH3,HCl
CH3
O
O
e
7
8
d
2, R1 = CH2Br
NHCOR1
NH2,HCl
b
a
CH3
O
CH3
O
8
5
1
7
9
2
a or b
6
3
1, R1 = CH3
10
NHCOCH3
NHCOCH3
4
f
1, R1 = CH3
A
CH3
O
2, R1 = CH2Br
3, R1 = CH2CO2Et
4, R1 = CH2F
CH3
O
g
c
5, R1 = CHF2
6, R1 = (CH2)2OH
COR2
CH2R2
9 a-i
10 a-i
R2
R2
a
b
c
d
e
CH3
C2H5
CH(CH3)2
CH2CH(CH3)2
c-C3H5
f
c-C4H7
c-C5H9
c-C6H11
C6H5
g
H
i
Scheme 1. Synthesis of compounds 9aei and 10aeia aReagents: (a) K2CO3, AcOEteH2O, R1COCl or BrCH2COBr for 2; (b) i) K2CO3, ii) ethyl fluoroacetate or methyl difluoroacetate for
compounds 4 and 5, 2,2,2-trifluoroethanol, reflux; (c) LiAlH4, THF, 0 ꢀC; (d) N-methylbenzylamine, NEt3, acetone, reflux; (e) i) H2, Pd/C, MeOH, r.t., ii) HCl(g), Et2O); (f) AlCl3, DCM/
DMF, 0 ꢀC, R2COCl; (g) Et3SiH, TFA, r.t.

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M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
NHCOCH3
NHCOCH3
NHCOCH3
a or b
To modulate the C-7 position of the agomelatine, we investi-
gated the introduction of amino and fluoro groups (Scheme 5). The
naphthol 27, obtained by demethylation of the compound 1 by
boron tribromide, was first activated as triflate [27] followed by
a nucleophilic substitution of the triflate group with p-methox-
ybenzylamine in the presence of palladium dibenzyledeneacetone
and xantphos [19] giving the amine 29 after debenzylation. The
fluorinated compound 30 was then readily prepared from the ob-
tained amine 29 according to the synthetic route previously
described for compound 16 (Scheme 3). Furthermore, the methyl
moiety of the methoxy group was replaced by fluorinated chains;
the naphthol 27 was O-alkylated in acetone at reflux by treatment
with the corresponding alkyl halide leading to compounds 31e33.
The hydroxyl group of compound 31 was then substituted over
a fluorine atom by reaction with DAST in dichloromethane to
furnish compound 34.
CH3 Br
O
CH3
O
CH3
O
c
Br
11
12
1
d
Scheme 2. Preparation of intermediate 12a aReagents: (a) Br2, AcOH, rt; (b) NBS, CCl4,
70 ꢀC; (c) AcOH, 70 ꢀC; (d) Br2, AcOH, 70 ꢀC.
hydrochloride led to the oxime 20 which was then reduced into the
corresponding amine 21 by catalytic hydrogenation over Raney
nickel in 46% yield (Scheme 3).
In Scheme 4 we report the synthesis of compound 23 in two
steps sequence; (1) the acid 22 was synthesized in a one-pot
procedure from the corresponding ketone 9a. The reaction took
place with sodium hypochlorite and potassium iodide in a mixture
of 1,4-dioxane/water at room temperature [22]. The acid 22 was
first esterified with methanol by treatment with thionyl chloride
then the ester function reduced using lithium aluminum hydride in
dry tetrahydrofuran [23], (2) Treatment of the hydroxyl group with
diethylaminosulfur trifluoride (DAST) at 0 ꢀC [24,25] led to the
fluoromethyl 23 in 50% yield. Otherwise, reaction of compound 9a
with hydroxylamine hydrochloride afforded the corresponding
oxime 24 in only one isomer determined as the (E)-isomer
according to the procedure previously described as Beckmann
transposition [26]. Therefore, the hydrogenation of this later
compound over Raney nickel in methanol yielded the corre-
sponding amine hydrochloride 25 after acidic treatment. Besides,
the acetamide 26 was obtained by treatment of the oxime 24 with
phosphorus pentachloride in dichloromethane.
Finally, the syntheses of 3-methoxy-naphthalene derivatives 39
and 40 have been accomplished as described in Scheme 6. The key
intermediate
B was synthesized from
a-amino-naphthalene
according to the procedure previously described by Newman [28].
Moreover and to introduce the ethylamine side chain, our proce-
dure involved reaction of the brominated intermediate B with
freshly prepared tributyl cyanomethyltin [29] under palladium-
catalyzed Stille reaction conditions [30]. The obtained nitrile 35
was then reduced into the amine 37 using LiAlH4eAlCl3 reagent.
Subsequently, we envisaged the preparation of its homologue via
Heck cross-coupling reaction [31]. Treatment of the bromoder-
ivative B with acrylonitrile in the presence of palladium acetate,
triphenylphosphine and triethylamine [32] to give the 3-(3-
methoxy-naphthalen-1-yl) acrylonitrile (36) in
a quantitative
yield as a mixture E/Z (3/1). This mixture was then reduced quan-
titatively in two steps: first by treatment with sodium borohydride
in methanol followed by reduction of the nitrile function with
NHCOCH3
NHCOCH3
CH3
O
CH3
O
c
15, R3 = NH2,HCl
R3
F
NHCOCH3
a
16
13, R3 = morpholine
14
15
, R3 = 4-methoxybenzylamine
, R3 = NH2,HCl
CH3
O
b
Br
NHCOCH3
NHCOCH3
12
NHCOCH3
d
CH3
O
CH3
O
CH3
O
f
e
H
17, R3 = vinyl
R3
CHO
N
20
17
19
, R3 = vinyl
, R3 = allyl
OH
18
g
NHCOCH3
CH3
O
CH2NH2,HCl
21
Scheme 3. Preparation of compounds 13e21a aReagents: (a) Pd2(dba)3, xantphos, morpholine or 4-methoxybenzylamine, Cs2CO3, dioxane, reflux; (b) H2, Pd/C, EtOH; (c) i) NaNO2,
HCl(aq), 0 ꢀC, ii) HBF4, xylene, reflux; (d) tributylvinyltin or allyltributyltin, Pd(PPh3)4, toluene, 100 ꢀC; (e) OsO4, NaIO4, THF-H2O, r.t.; (f) HONH2,HCl, pyridine, MeOH, reflux; (g) i) H2,
Ra-Ni, NH3(g), MeOH, ii) HCl(g), Et2O.

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
313
NHCOCH3
NHCOCH3
CH3
O
CH3
O
b
OH
NHCOCH3
CH2F
a
c
23
22
O
CH3
O
CH3
O
NHCOCH3
NHCOCH3
9a
CH3
O
CH3
O
d
CH3
CH3
25
24
N
NH2,HCl
OH
e
NHCOCH3
CH3
O
O
N
CH3
H
26
Scheme 4. Synthesis of compounds 23e26a aReagents: (a) KI, NaOCl, 1,4-dioxane/water, r.t., (b) i) SOCl2, MeOH, 0 ꢀC, ii) LiAlH4, THF, 0 ꢀC, iii) DAST, DCM, 0 ꢀC; (c) HONH2,HCl,
pyridine, MeOH, reflux; (d) i) H2, Ra-Ni, MeOH-NH3(g), r.t., ii) HCl(g), Et2O; (e) PCl5, DCM, r.t.
LiAlH4 to lead to the amine 38. The amines 37 and 38 were then
converted into acetamides 40 and 41 by N-acylation.
receptors stably transfected in Human Embryonic Kidney (HEK
293) cells or Chinese Hamster Ovarian (CHO) cells, using 2-[125I]
iodomelatonin as radioligand [33]. At each receptor, binding affin-
ities were verified for more than fifty selective and non-selective
molecules, either using the transfected HEK 293 or the CHO cell
lines. The correlations between affinities in HEK 293 and CHO cells
are highly significant (r ¼ 0.98), for both MT1 and MT2 receptors
[33]. This high correlation allowed the comparison of the affinities
on the receptors expressed in two different cell types and to draw
2.2. Biological results
The chemical structures, melatoninergic (MT1/MT2) and sero-
toninergic (5-HT2C
)
binding affinities of the synthesized
compounds are reported in Tables 1e4. These compounds were
evaluated for their binding affinity at human MT1 and MT2
NHCOCH3
NHCOCH3
NHCOCH3
CH3
O
HO
TfO
b
a
c
1
27
28
NHCOCH3
e
H2N
NHCOCH3
R
1
( )n
O
29
NHCOCH3
d
31-34
Compds
31
32
33
34
n
3
2
1
3
R1
OH
F
CF3
F
F
f
30
a
Scheme 5. Synthesis of compounds 29e34a Reagents: (a) BBr3, DCM, 0 ꢀC; (b) Tf2O, NEt3,DCM, ꢁ20 ꢀC; (c) i) Pd2(dba)3, xantphos, 4-methoxy-benzylamine, Cs2CO3, DMF, 100W,
microwave, ii) H2, Pd/C, MeOH; (d) i) NaNO2, HCl(aq), HBF4, 0 ꢀC ii) xylene, reflux; (e) K2CO3, acetone, reflux, TBAB, Br(CH2)3OH for 31 or X(CH2)nR1 for 32 and 33; (f) DAST, DCM, 0 ꢀC.

314
M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
N
c
NHCOCH3
NH2,HCl
O
( )n
35
( )n
Br
a
CH3
CN
e
O
O
O
39 , n = 1
40 , n = 2
37 , n = 1
38 , n = 2
B
CH3
CH3
CH3
b
d
O
36
CH3
a
Scheme 6. Synthesis of compounds 39 and 40a Reagents: (a) Bu3SnCH2CN, PdCl2(PPh3)2, toluene, reflux; (b) acrylonitrile, Pd(OAc)2, PPh3, NEt3, 100 ꢀC; (c) i) LiAlH4eAlCl3, 0 ꢀC,
Et2O, ii) HCl(g), Et2O; (d) i) NaBH4, MeOH, reflux, ii) LiAlH4eAlCl3, 0 ꢀC, Et2O, iii) HCl(g), Et2O; (e) K2CO3, AcOEteH2O, 0 ꢀC, CH3COCl.
structureeactivity relationships. Serotoninergic 5-HT2C binding
affinity was performed using Chinese Hamster Ovarian (CHO) cell
lines stably expressing the human 5-HT2C receptors.
groups showed a high melatoninergic binding affinity particularly
towards MT1 receptor subtype. The fluoroacetamide 4 showed
good MT1 and MT2 binding affinities (70 pM) higher than the lead
while the difluoroacetamide 5 behaved as a MT1-selective agonist
(143-fold). Otherwise, introduction of N-methylamine function
(compound 8) caused a fall of the MT1 binding affinity leading to
a weak MT2-selectivity factor of 25.
In search of new dual ligands we focused our attention on the
agomelatine as a lead. To better understand the role of the amide
nature, the methoxy group and naphthalene C-3 position on the
melatoninergic (MT1/MT2) and/or serotoninergic (5-HT2C) binding
affinities, we synthesized and biologically evaluated at human
cloned melatoninergic receptors subtypes a novel series of N-[2-(7-
methoxy-naphthalen-1-yl)ethyl]amide derivatives. These modula-
tions provided compounds with different binding and intrinsic
activity profiles (Tables 1e4). Compounds bearing fluoroacetamide
In the aim of understanding the role of the naphthalene ring C-3
position in the modulation of both melatoninergic and serotonin-
ergic binding affinities, several chemical modifications of this
position have been explored (Tables 1e3). Mainly, introduction of
an ethyl group (compound 10a) improved the MT1 and MT2 binding
affinities (MT1: Ki ¼ 0.06 nM; MT2: Ki ¼ 0.04 nM). Within this series,
the extension of the ethyl group to propyl (10b) [34], isobutyl or
isoamyl (compounds 10c and 10d) led to a decrease in the mela-
toninergic binding affinities particularly for MT1 leading to a MT2-
selectivity which is in agreement with findings observed previously
[35e37]. However, the bulky compound 10d possessed a higher
affinity and selectivity for MT2 (MT2: Ki ¼ 0.01 nM, S ¼ 7200-fold)
than agomelatine and remains one of the most potent MT2-selec-
tive compounds described to date in the naphthalenic series.
In further modulations of the C-3 position, we introduced
methylcycloalkyl groups. Thus, introduction of a methylcyclopropyl
(compound 10e) exhibited an interesting melatoninergic binding
affinity (MT1: Ki ¼ 3.9 nM; MT2: Ki ¼ 0.06 nM) [34]. It leads to a 39-
fold decrease of the MT1 binding affinity but improves the MT2
binding affinity of about 2-fold. Moreover, the methylcyclobutyl 10f
Table 1
Melatoninergic MT1 and MT2 binding affinities of compounds 1e26.
NHCOR1
O
R2
Compounds R1
R2
Ki (nM)
Ki (nM)
MT1
MT2
1
4
5
6
CH3
H
H
H
H
H
C2H5
C3H7
CH2CH(CH3)2
(CH2)2CH(CH3)2
CH2ceC3H5
CH2ceC4H7
CH2ceC5H9
CH2ceC6H5
CH2ceC6H11
Morpholine
NH2
0.1 ꢂ 0.01
0.12 ꢂ 0.02
CH2F
CHF2
(CH2)2OH
CH2NHCH3,HCl
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
0.07 ꢂ 0.01 0.07
0.03 ꢂ 0.0
4.5 ꢂ 0.4
4.3 ꢂ 1.0
Table 2
0.59 ꢂ 0.04
8.7 ꢂ 0.33
MT1 and MT2 GTPgS(þ) binding affinities of compounds 1e40.
8
218.0 ꢂ 14.0
0.06 ꢂ 0.02
0.39 ꢂ 0.03
65.0 ꢂ 4.0
72.0 ꢂ 11.0
3.9 ꢂ 1.8
10a
10b
10c
10d
10e
10f
10g
10h
10i
13
15
16
17
18
23
24
25
26
0.04 ꢂ 0.02
0.28 ꢂ 0.01
1.34 ꢂ 0.04
0.01 ꢂ 0.31
Compound
MT1
MT2
EC50 (nM)
Emax (%)
EC50 (nM)
Emax (%)
1
4
5
6
1.29 ꢂ 0.25
1.79 ꢂ 0.41
0.06 ꢂ 0.017
8.01 ꢂ 3.95
789.0 ꢂ 16.5
200.0 ꢂ 50.0
272.0 ꢂ 127.0
27.8 ꢂ 0.0
103.0 ꢂ 0.4
82.0 ꢂ 8.0
109.0 ꢂ 4.0
115.0 ꢂ 5.0
67.0 ꢂ 2.0
93.0 ꢂ 6.0
49.0 ꢂ 3.0
81.0 ꢂ 0.0
80.0 ꢂ 5.0
102.0 ꢂ 11.0
110.0 ꢂ 6.0
101.0 ꢂ 10.0
41.0 ꢂ 1.0
81.0 ꢂ 13.0
63.0 ꢂ 3.0
2.72 ꢂ 0.3
103.0 ꢂ 6.0
111.0 ꢂ 14.0
98.0 ꢂ 5.0
126.0 ꢂ 18.0
101.0 ꢂ 10.0
94.0 ꢂ 6.0
68.0 ꢂ 3.0
89.0 ꢂ 0.0
125.0 ꢂ 9.0
103 ꢂ 4.0
0.06
0.25 ꢂ 0.10
0.04 ꢂ 0.01
1.96 ꢂ 0.20
6.43 ꢂ 0.85
4.50 ꢂ 0.70
0.31 ꢂ 0.12
0.36 ꢂ 0.0
119.0 ꢂ 7.0
4.2 ꢂ 0.5
0.48 ꢂ 0.03
4.2 ꢂ 0.46
4.35
0.32
8
50.0 ꢂ 1.0
9.9 ꢂ 0.42
88.3 ꢂ 3.5
1.1 ꢂ 0.2
10c
10d
10e
10f
10g
17
23
24
39
40
4170
2.3 ꢂ 0.6
F
0.10 ꢂ 0.01
0.15 ꢂ 0.03
0.17 ꢂ 0.05
6.1 ꢂ 0.1
0.07 ꢂ 0.01
0.13 ꢂ 0.05
0.18 ꢂ 0.03
0.39 ꢂ 0.04
3.5 ꢂ 0.10
64.0 ꢂ 1.0
185.0 ꢂ 10.0
65.6 ꢂ 2.0
1.72 ꢂ 0.05
9.36 ꢂ 3.03
0.05 ꢂ 0.06
2.12 ꢂ 0.40
4.51 ꢂ 1.12
1.14 ꢂ 0.34
4.93 ꢂ 0.65
CH]CH2
CH2CH]CH2
CH2F
(E)C(CH3)NOH
(CH)CH3NH2,HCl
NHCOCH3
CH3
CH3
CH3
CH3
0.96 ꢂ 0.04
14.2 ꢂ 11.4
227.0 ꢂ 77.0
21.1 ꢂ 4.4
105.0 ꢂ 6.0
55.0 ꢂ 13.0
85.0 ꢂ 11.0
99.0 ꢂ 12.0
93.0 ꢂ 9.0
71.0 ꢂ 4.0
45.0 ꢂ 2.0
CH3
1560 ꢂ 88.0 124.0 ꢂ 12.0
31.8 ꢂ 1.2

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
315
Table 3
39 and 40 showed lower binding constants at MT1 and MT2 subtype
receptors respectively, than the parent ligand.
MT1 and MT2 binding affinities of compounds 30e34 and 39e40.
At last, all the synthesized compounds were evaluated at human
NHCOCH3
cloned serotoninergic2C (5-HT2C
)
receptors subtypes. Only
compounds 10e, 17 and 18 showed an interesting pharmacological
profile (Table 4). The methylcyclopropyl 10e (MT1: Ki ¼ 3.9 nM;
MT2: Ki ¼ 0.06 nM; 5-HT2C: Ki ¼ 391.0 nM) showed about 2-fold
increase of the 5-HT2C binding affinity than the lead. The vinyl 18
exhibited 1.4-fold less binding affinity at this receptor, whereas the
( )n
R3
R2
allyl 18 (MT1: Ki ¼ 0.17 nM; MT2: Ki ¼ 0.18 nM; 5-HT2C
:
Ki ¼ 144.0 nM) showed the most interesting serotoninergic binding
affinity 5-times more potent than the agomelatine. Intrinsic activity
of these three selected compounds (10e, 17 and 18) is under study.
Compound
n
R2
R3
Ki (nM)
MT1
Ki (nM)
MT2
1
1
1
1
1
1
1
2
H
H
H
H
H
OCH3
OCH3
OCH3
F
O(CH2)2F
OCH2CF3
O(CH2)3F
H
0.1 ꢂ 0.01
7.74 ꢂ 1.60
1.71 ꢂ 0.45
5.58 ꢂ 0.0
5.24 ꢂ 1.70
1.73 ꢂ 0.04
4.0 ꢂ 0.3
0.12 ꢂ 0.02
0.56 ꢂ 0.05
0.98 ꢂ 0.12
8.70 ꢂ 1.31
1.05 ꢂ 0.19
1.19 ꢂ 0.02
3.8 ꢂ 0.09
30
32
33
34
39
40
3. Conclusions
A series of new naphthalenic compounds has been synthesized
using agomelatine as a lead and their affinity and functional activity
were evaluated in vitro at human melatoninergic (MT1/MT2) and
serotoninergic (5-HT2C) receptors stably transfected in Human
Embryonic Kidney (HEK 293) cells or Chinese Hamster Ovarian
(CHO) cells. To better understand the role of the acetamide group of
the agomelatine in the melatoninergic and/or the serotoninergic 5-
HT2C binding affinity, its replacement by a fluoro or difluor-
oacetamide (compounds 4, 5) led to a good melatoninergic binding
affinity. Moreover, compound 5 showed a MT1-selectivity of about
143-fold. Otherwise, the C-3 position of naphthalene core modu-
lation provided compounds with different binding and intrinsic
activity profiles. Thus, the introduction of a bulky group (compound
10d) led to the most MT2-selective (S ¼ 7200-fold) compound
described nowadays in the naphthalenic series. Introduction of
a methylcyclopropyl (compound 10e) or an allyl (compound 18)
provided the most interesting compounds of these series. Biological
evaluation of all the synthesized compounds at human serotonin-
ergic (5-HT2C) receptors showed that only compounds 10e, 17 and
18 exhibited a good serotoninergic binding affinity while their
intrinsic activity is under study.
H
(MT1: Ki ¼ 119.0 nM; MT2: Ki ¼ 0.48 nM) showed a MT2-selectivity
of about 248-fold. Otherwise, the introduction in this position of an
embedded group such as morpholine (compound 13) decreased
notably the binding affinity at MT1 receptor leading to the
appearance of a weak MT2-selectivity (47-fold). Furthermore, the
introduction in the C-3 position of vinyl (compound 17, MT1:
Ki ¼ 0.15 nM; MT2: Ki ¼ 0.13 nM) and allyl (compound 18, MT1:
Ki ¼ 0.17 nM; MT2: Ki ¼ 0.18 nM) groups didn’t have a strong
influence on the melatoninergic binding affinities. By contrast, our
next modulation was the insertion of hydrophilic substituents. In
this case, oxime, amine and alcohol functions were grafted but their
binding affinities were low than the agomelatine. Finally, intro-
duction of a fluorine atom (compound 16) exhibited a good mela-
toninergic binding affinity (MT1: Ki ¼ 0.10 nM; MT2: Ki ¼ 0.07 nM)
than the parent compound. Meanwhile, the introduction of a fluo-
romethyl (compound 23) led to the decrease of both MT1 (61-fold)
and MT2 (3-fold) binding affinities.
To broaden the structureeactivity relationships concerning the
effect of the methoxy group on both melatoninergic activity and
affinity [38,39], first we replaced this group by a fluorine atom
(compound 30) then by fluoroethers (compounds 32e34). These
modulations led to a decrease of the melatoninergic binding
affinity (Table 3). We have also synthesized the C-3 methoxy group
analogues of the agomelatine (Table 3). The resultant compounds
4. Experimental section
4.1. Chemistry
All reagents and dry solvents were purchased from
SigmaeAldrich, Alfa Aesar or Acros organics and were used without
further purification except as noted below. Dichloromethane was
distilled from phosphorus pentoxide and Tetrahydrofuran was
purified by distillation from sodium and benzophenone. Dimethyl
formamide was dried by distillation from anhydrous calcium
sulfate and was stored under argon. TLC was conducted on Merck
precoated silica gel 60 F-254. Melting points were determined on
a Büchi SMP-20 capillary apparatus and are uncorrected. IR spectra
were recorded on a PerkineElmer 297 or a Vector 22 Bruker
spectrometer. 1H and 13C NMR spectra were recorded on an AC 300
Bruker spectrometer. Chemical shifts are reported in ppm units
(parts per million) relative to (CH3)4Si, and J values are reported in
hertz (Hz). Purity of all final products was determined by analytical
HPLC to be ꢃ95%. HPLC purity of compounds was measured with
Table 4
Melatoninergic (MT1/MT2) and serotoninergic (5-HT2C) binding affinities of
compounds 1, 10e, 17 and 18.
NHCOCH3
O
R2
a
reverse phase HPLC (Lichrospher 100 RP-18 column,
m). Mass spectra were performed on
4.6 mm ꢄ 150 mm, 12
m
Compound
R2
Ki (nM) MT1
Ki (nM) MT2
Ki (nM) 5-HT2C
a Finnigan MAT SSQ 710 Advantage spectrometer and were recor-
ded in the APCI positive mode. Elemental analyses for tested
compounds were performed by CNRS Laboratories (Vernaison,
France). Obtained results were within ꢂ0.4% of the theoretical
values.
1
H
0.1 ꢂ 0.01
3.9 ꢂ 1.8
0.12 ꢂ 0.02
0.06
708 ꢂ 45.0
391.0 ꢂ 4.0
1000
144.0 ꢂ 43.0
10e
17
18
CH2c-C3H5
CH]CH2
CH2CH]CH2
0.15 ꢂ 0.03
0.17 ꢂ 0.05
0.13 ꢂ 0.05
0.18 ꢂ 0.03

316
M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
4.1.1. Ethyl N-[2-(7-methoxy-naphthalen-1-yl)ethyl]malonate (3)
To a suspension of 2-(7-methoxy-naphthalen-1-yl)ethylamine
hydrochloride (compound A) (5.0 g, 21.0 mmol) in 100 mL of
a mixture of water-ethyl acetate (1/1), potassium carbonate (8.7 g,
63.0 mmol) was added and the reaction mixture was cooled at 0 ꢀC
in an ice-bath. Ethyl malonyl chloride (3.58 mL, 27.8 mmol) was
added and the mixture stirred for 1 h at room temperature. The
organic phase was then washed respectively with an aqueous
solution of HCl (1 M) and water, dried over MgSO4, filtered and
evaporated under reduced pressure to give 6.09 g (92% yield) of the
desired product after recrystallization from isopropyl ether as
a white solid; m.p. 90e92 ꢀC; 1H NMR (300 MHz, CDCl3): dH 1.29 (t,
J ¼ 7.2, 3H, CH3), 3.20e3.40 (m, 4H, CH2-a,c), 3.70 (m, 2H, CH2-b),
4.00 (s, 3H, OCH3), 4.18 (quintet, J ¼ 7.2, 2H, CH2-d), 7.18 (dd, J ¼ 9.0,
J ¼ 2.4, 1H, H-6), 7.25e7.35 (m, 3H, H-2,3 and NH), 7.50 (d, J ¼ 2.4,
1H, H-8), 7.70 (d, J ¼ 7.6, 1H, H-4), 7.76 (d, J ¼ 9.0, 1H, H-5); IR (neat,
cmꢁ1): 3230 (NH), 1738 (C]O), 1641 (C]O); LC/MS m/z 316.4
(M þ 1); Anal. calcd for C18H21NO4: C 68.65%, H 6.71%, N 4.44%;
Found: C 68.66%, H 6.68%, N 4.41%.
simultaneously with 0.7 mL of water, 0.7 mL of sodium hydroxide
(15%) and 2.1 mL of water. The solid formed was filtered and the
filtrate washed with water, dried over MgSO4 and evaporated under
reduced pressure. Recrystallization from acetonitrile gave 0.30 g of
the desired product as a white solid (12% yield); m.p. 84e86 ꢀC; 1H
NMR (300 MHz, CDCl3): dH 1.60 (signal exchangeable in D2O, 1H,
OH), 2.40 (t, J ¼ 5.4, 2H, CH2-c), 3.30 (t, J ¼ 7.0, 2H, CH2-a), 3.70 (m,
2H, CH2-b), 3.90 (t, J ¼ 5.4, 2H, CH2-d), 4.05 (s, 3H, OCH3), 5.85 (br s,
1H, NH), 7.18 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.27e7.32 (m, 2H, H-2,3),
7.47 (d, J ¼ 2.4, 1H, H-8), 7.70 (dd, J ¼ 6.3, J ¼ 2.9, 1H, H-4), 7.79 (d,
J ¼ 9.0, 1H, H-5); 13C NMR (CDCl3): dC 22.8 C-a, 35.4 Cec, 39.4 C-b,
55.6 OCH3, 60.44 C-d,102.9 C-8,118.3 C-6,123.5 C-5,127.2 C-2,127.5
C-4, 129.4 C-10, 130.4 C-3, 132.9 C-1, 133.0 C-9, 158.0 C-7, 172.1 CO;
IR (neat, cmꢁ1): 3444, 3332, 1655; LC/MS m/z 274.3 (M þ 1); Anal.
calcd for C16H19NO3: C 70.31%, H 7.01%, N 5.12%. Found: C 70.28%, H
7.00%, N 5.09%.
4.1.4. 2-(Benzyl-methylamino)-N-[2-(-7-methoxy-naphthalen-1-
yl)ethyl]acetamide (7)
To a solution of compound 2 (3.2 g, 10.0 mmol) in 50 mL of
acetone was added successively N-methyl benzylamine (1.2 g,
10.0 mmol) and triethylamine (2.8 mL, 20.0 mmol), the reaction
mixture was refluxed for 1 h. The acetone was then evaporated
under reduced pressure and the residue taken up with water and
extracted with ethyl ether. The organic layer was washed with
water, dried over MgSO4 and evaporated under reduced pressure to
lead to 2.86 g (79% yield) of the desired product as a white solid
after recrystallization from cyclohexane; m.p. 88e90 ꢀC; 1H NMR
(300 MHz, DMSO-d6): dH 2.12 (s, 3H, NCH3), 2.95 (s, 2H, CH2-d), 3.17
(t, J ¼ 8.0, 2H, CH2-a), 3.46 (m, 2H, CH2-b), 3.52 (s, 2H, CH2-c), 3.94
(s, 3H, OCH3), 7.14e7.86 (m, 11H, H-aromatic), 8.00 (br s, 1H, NH); IR
(neat, cmꢁ1): 3342 (NH), 1655 (C]O); LC/MS m/z 363.5 (M þ 1).
4.1.2. General protocol for the preparation of compounds 4, 5
To a solution of free base of compound A (5.0 mmol) in 10 mL of
ethanol trifluoride, was added ethyl 2-fluoroacetate or methyl 2,2-
difluoroacetate (15.0 mmol), the reaction mixture was refluxed for
12 h. After concentration under reduced pressure, the obtained
residue was poured into 40 mL of water, acidified with an aqueous
solution of HCl (3 M) and extracted twice with 50 mL of ethyl
acetate. The organic phase was washed with water, dried over
MgSO4 and evaporated under reduced pressure to lead to the
desired product after recrystallization.
4.1.2.1. 2-Fluoro-N-[2-(7-methoxy-naphthalen-1-yl)ethyl]acetamide
(4). Recrystallized from cyclohexane as a white solid (0.65 g, 50%
yield); m.p. 90e92 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 3.19 (t,
J ¼ 8.3, 2H, CH2-a), 3.45 (m, 2H, CH2-b), 3.94 (s, 3H, OCH3), 4.85 (d,
J ¼ 47.0, 2H, CH2-c), 7.18 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.22e7.35
(m, 2H, H-2,3), 7.60 (d, J ¼ 2.4, 1H, H-8), 7.70 (d, J ¼ 7.6, 1H,
H-4), 7.83 (d, J ¼ 9.0, 1H, H-5), 8.49 (br s, 1H, NH); 13C NMR
(DMSO-d6): dC 33.4 C-a, 39.5 C-b, 55.7 OCH3, 79.4 and 81.8
(1J ¼ 181.0) Cec, 103.0 C-8, 118.5 C-6, 123.6 C-3, 127.1 C-2, 127.5 C-
4, 129.3 C-10, 130.6 C-5, 133.3 C-9, 134.3 C-1, 158.0 C-7, 168.0 CO; IR
(neat, cmꢁ1): 3354 (NH), 1685 (C]O); LC/MS m/z 262.3 (M þ 1);
Anal. calcd for C15H16FNO2: C 68.97%, H 6.17%, N 5.36%; Found: C
69.01%, H 6.21%, N 5.32%.
4.1.5. N-[2-(7-Methoxy-naphthalen-1-yl)ethyl]-2-(methylamino)
acetamide hydrochloride (8)
To a solution of compound 7 (3.08 g, 8.5 mmol) in 50 mL of
methanol was added 0.3 g of palladium on charcoal. The reaction
mixture was hydrogenated at atmospheric pressure for 12 h. After
filtration, the mixture was evaporated under reduced pressure to
lead to the desired product after treatment with HCl gas. Recrys-
tallized from acetonitrile as a white solid (2.1 g, 80% yield); m.p.
194e196 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 2.54 (d, J ¼ 8.5, 3H,
NCH3), 3.17 (t, J ¼ 8.0, 2H, CH2-a), 3.46 (m, 2H, CH2-b), 3.69 (s, 2H,
CH2-c), 3.94 (s, 3H, OCH3), 7.18 (dd, J ¼ 9.0, J ¼ 2.4,1H, H-6), 7.27 (dd,
J ¼ 6.0, J ¼ 8.0, 1H, H-3), 7.34 (d, J ¼ 6.0, 1H, H-2), 7.54 (d, J ¼ 2.4, 1H,
H-8), 7.72 (d, J ¼ 8.0,1H, H-4), 7.85 (d, J ¼ 9.0,1H, H-5), 8.88 (br s,1H,
NH), 9.02 (br s, 2H, NHþ2 Clꢁ); 13C NMR (DMSO-d6): dC 33.0 CH3,
33.2 C-a, 39.8 C-b, 49.4 Cec, 55.8 OCH3, 103.0 C-8, 118.4 C-6, 123.6
C-3, 127.1 C-2, 127.6 C-4, 129.3 C-10, 130.7 C-5, 133.2 C-9, 134.1 C-1,
158.0 C-7, 166.0 CO; IR (neat, cmꢁ1): 3238 (NH), 3000-2500
(NHþ2 Clꢁ), 1686 (C]O); LC/MS m/z 309.8 (M þ 1); Anal. calcd for
C16H21ClN2O2: C 62.23%, H 6.85%, O 10.36%; Found: C 61.99%, H
6.88%, O 10.40%.
4.1.2.2. 2,2-Difluoro-N-[2-(7-methoxy-naphthalen-1-yl)ethyl]acetamide
(5). Recrystallized from cyclohexane as a white solid (0.95 g, 68%
yield); m.p. 77e79 ꢀC; 1H NMR (300 MHz, CDCl3): dH 3.30
(t, J ¼ 6.7, 2H, CH2-a), 3.75 (m, 2H, CH2-b), 4.00 (s, 3H, OCH3), 5.90
(t, J ¼ 54.2, 1H, H-c), 6.40 (br s, 1H, NH), 7.18 (dd, J ¼ 9.0, J ¼ 2.4,
1H, H-6), 7.25e7.35 (m, 2H, H-2,3), 7.44 (d, J ¼ 2.4, 1H, H-8),
7.70 (m, 1H, H-4), 7.80 (d, J ¼ 9.0, 1H, H-5); 13C NMR (CDCl3): dC
23.8 C-a, 39.7 C-b, 55.5 OCH3, 102.1 C-8, 108.5 and 111.8
(1J ¼ 253.0) Cec, 118.5 C-6, 123.2 C-3, 127.3 C-2, 127.5 C-4, 129.4 C-
10, 130.4 C-5, 132.5 C-9, 133.0 C-1, 158.2 C-7, 163.2 CO; IR (neat,
cmꢁ1): 3278, 1675; LC/MS m/z 280.3 (M þ 1); Anal. calcd for
C15H15F2NO2: C 64.51%, H 5.41%, N 5.02%; Found: C 64.48%, H
5.39%, N 4.99%.
4.1.6. General procedure for preparation of compounds 9aei
To a solution of compound 1 (20.5 mmol) in 60 mL of a mixture
of dichloromethane-DMF at 0 ꢀC, was added the aluminum chloride
(61.6 mmol) and the mixture was allowed to stir for 5 min while
keeping the temperature at 0 ꢀC. The desired acyl chloride
(26.7 mmol) was added dropwise cautiously and the mixture stir-
red for 1 h at room temperature. The reaction mixture was then
hydrolyzed with ice-water (100 mL) and the two phases were
separated. The organic phase was washed with water, dried
(MgSO4), filtered and evaporated under reduced pressure to give
the desired product.
4.1.3. 3-Hydroxy-N-[2-(7-methoxy-naphthalen-1-yl)ethyl]
propanamide (6)
Compound 3 (2.93 g, 9.3 mmol) was dissolved in dry tetrahy-
drofuran (75 mL) then cooled at 0 ꢀC, the aluminum lithium
hydride (0.70 g, 18.6 mmol) was added dropwise to the reaction
mixture. After 1 h stirring, the reaction mixture was hydrolyzed

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
317
4.1.6.1. N-[2-(3-Acetyl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(9a). Recrystallized from toluene as a white solid (4.68 g, 80%
yield); m.p. 152e153 ꢀC; 1H NMR (300 MHz, CDCl3): dH 1.83 (s, 3H,
CH3-c), 2.65 (s, 3H, CH3), 3.16 (m, 2H, CH2-a), 3.36 (m, 2H, CH2-b),
3.99 (s, 3H, OCH3), 7.26 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.68 (d, J ¼ 2.0,
1H, H-8), 7.80 (s, 1H, H-2), 8.03 (d, J ¼ 9.0, 1H, H-5), 8.15 (br s, 1H,
NH), 8.47 (d, J ¼ 2.0, 1H, H-4); IR (neat, cmꢁ1): 3297 (NH), 1662 (C]
O), 1639 (C]O); LC/MS m/z 286.3 (M þ 1).
6), 7.68 (d, J ¼ 2.4, 1H, H-8), 7.81 (d, J ¼ 2.0, 1H, H-2), 8.07 (d, J ¼ 9.0,
1H, H-5), 8.12 (br s, 1H, NH), 8.50 (d, J ¼ 2.0, 1H, H-4); IR (neat,
cmꢁ1): 3383 (NH), 1672 (C]O), 1657 (C]O); LC/MS m/z 340.4
(M þ 1).
4.1.6.8. N-[2-(3-Cyclohexanecarbonyl-7-methoxy-naphthalen-1-yl)
ethyl]acetamide (9h). Recrystallized from acetonitrile as a white
solid (5.80 g, 80% yield); m.p. 130e132 ꢀC; 1H NMR (300 MHz,
DMSO-d6): dH 1.35e1.43 (m, 6H, H-30,40,50), 1.70e1.77 (m, 4H, H-
20,60), 1.80 (s, 3H, CH3), 3.16 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.53
(m, 1H, H-10), 3.97 (s, 3H, OCH3), 7.27 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6),
7.68 (d, J ¼ 2.4, 1H, H-8), 7.78 (d, J ¼ 2.0, 1H, H-2), 8.07 (d, J ¼ 9.0, 1H,
H-5), 8.12 (br s, 1H, NH), 8.50 (d, J ¼ 2.0, 1H, H-4); IR (neat, cmꢁ1):
3380 (NH), 1672 (C]O), 1655 (C]O); LC/MS m/z 354.5 (M þ 1).
4.1.6.2. N-[2-(7-Methoxy-3-propanoyl-naphthalen-1-yl)ethyl]acetamide
(9b). Compound recrystallized from toluene as a white solid
(4.17g, 68% yield); m.p. 139e141 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 1.10 (t, J ¼ 7.2, 3H, CH3), 1.80 (s, 3H, CH3), 3.05e3.20 (m, 4H, CH2-
a,c), 3.35 (m, 2H, CH3-b), 4.00 (s, 3H, OCH3), 7.27 (dd, J ¼ 9.0, J ¼ 2.4,
1H, H-6), 7.68 (d, J ¼ 2.0, 1H, H-8), 7.80 (s, 1H, H-2), 8.07 (d, J ¼ 9.0,
1H, H-5), 8.12 (br s, 1H, NH), 8.50 (d, J ¼ 2.0, 1H, H-4); IR (neat,
cmꢁ1): 3397 (NH), 1685 (C]O), 1671 (C]O); LC/MS m/z 300.4
(M þ 1).
4.1.6.9. N-[2-(3-Benzoyl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(9i). Recrystallized from toluene as a white solid (5.7 g, 80% yield);
m.p. 142e144 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.85 (s, 3H,
CH3), 3.20 (t, J ¼ 8.0, 2H, CH2-a), 3.40 (m, 2H, CH2-b), 4.00 (s, 3H,
OCH3), 7.27 (dd, J ¼ 9.0, J ¼ 2.0, 1H, H-6), 7.53e7.84 (m, 7H, H-
2,8,phenyl), 8.03 (d, J ¼ 9.0, 1H, H-5), 8.10e8.20 (m, 2H, H-4 and
NH); IR (neat, cmꢁ1): 3363 (NH), 1671 (C]O), 1642 (C]O); LC/MS
m/z 348.4 (M þ 1).
4.1.6.3. N-[2-(3-Isobutyryl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(9c). Compound recrystallized from acetonitrile as a white solid
(3.34 g, 52% yield); m.p. 114e116 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 1.15 (d, J ¼ 6.6, 6H, CH3), 1.80 (s, 3H, CH3), 3.15 (m, 2H, CH2-a),
3.32 (m, 2H, CH2-b), 3.80 (m, 1H, CH), 3.96 (s, 3H, OCH3), 7.27 (dd,
J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.68 (d, J ¼ 2.0, 1H, H-8), 7.80 (d, J ¼ 2.4, 1H,
H-2), 8.07 (d, J ¼ 9.0, 1H, H-5), 8.12 (br s, 1H, NH), 8.50 (d, J ¼ 2.0,1H,
H-4); IR (neat, cmꢁ1): 3398 (NH), 1683 (C]O), 1664 (C]O); LC/MS
m/z 314.3 (M þ 1).
4.1.7. General procedure for compounds 10aei synthesis
To a solution of the acylated compound 9aei (6.4 mmol) in
20 mL of trifluoroacetic acid was added dropwise triethylsilane
(15.6 mmol). The mixture was allowed to stir for 12 h at room
temperature then evaporated under reduced pressure and the
precipitate formed in petroleum ether was recrystallized to lead to
the desired product.
4.1.6.4. N-[2-(3-Isovaleryl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(9d). Recrystallized from acetonitrile as a white solid (4.56 g, 68%
yield); m.p. 106e108 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 0.95 (d,
J ¼ 6.6, 6H, CH3), 1.80 (s, 3H, CH3), 2.20 (m, 1H, CH), 2.96 (d, J ¼ 6.6,
2H, CH2), 3.15 (m, 2H, CH2-a), 3.32 (m, 2H, CH2-b), 3.96 (s, 3H,
OCH3), 7.27 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.68 (d, J ¼ 2.0, 1H, H-8),
7.80 (d, J ¼ 2.4, 1H, H-2), 8.07 (d, J ¼ 9.0, 1H, H-5), 8.12 (br s, 1H, NH),
8.50 (d, J ¼ 2.0, 1H, H-4); IR (neat, cmꢁ1): 3387 (NH), 1663 (C]O),
1636 (C]O); LC/MS m/z 328.2 (M þ 1).
4.1.7.1. N-[2-(3-Ethyl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(10a). Recrystallized from cyclohexane as a white solid (0.95 g, 55%
yield); m.p. 106e108 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.24 (t,
J ¼ 7.6, 3H, CH3-b0), 1.83 (s, 3H, CH3), 2.72 (quintet, J ¼ 7.6, 2H, CH2-
a0), 3.10 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.90 (s, 3H, OCH3), 7.12
(dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.19 (d, J ¼ 2.0, 1H, H-8), 7.48 (d, J ¼ 2.0,
1H, H-2), 7.56 (d, J ¼ 2.4,1H, H-5), 7.75 (d, J ¼ 9.0,1H, H-4), 8.12 (br s,
1H, NH); 13C NMR (DMSO-d6): dC 22.5 C-b0, 23.1 CH3, 33.5 C-b,
45.1 C-a,a0, 55.5 OCH3, 102.8 C-8, 118.6 C-7, 126.0 C-6, 129.2 C-4,
129.7 C-10,130.2 C-5, 131.8 C-2, 134.3 C-9, 136.0 C-1, 157.7 C-3, 170.0
CO; IR (neat, cmꢁ1): 3272 (NH), 1644 (C]O); LC/MS m/z 272.3
(M þ 1); Anal. calcd for C17H21NO2: C 75.25%, H 7.80%, N 5.16%;
Found: C 75.28%, H 7.78%, N 5.13%.
4.1.6.5. N-[2-(3-cyclopropanecarbonyl-7-methoxy-naphthalen-1-yl)
ethyl]acetamide (9e). Recrystallized from toluene as a white solid
(4.14 g, 65% yield); m.p. 153e155 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 1.05 (m, 4H, H-20,30), 1.80 (s, 3H, CH3), 3.05 (quintet, 1H, H-10),
3.15 (m, 2H, CH2-a), 3.35 (m, 2H, CH2-b), 4.00 (s, 3H, OCH3), 7.27 (dd,
J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.68 (d, J ¼ 2.4, 1H, H-8), 7.84 (d, J ¼ 2.0, 1H,
H-2), 8.07 (d, J ¼ 9.0,1H, H-5), 8.12 (br s,1H, NH), 8.64 (d, J ¼ 2.0,1H,
H-4); IR (neat, cmꢁ1): 3375 (NH), 1670 (C]O), 1646 (C]O); LC/MS
m/z 312.2 (M þ 1).
4.1.7.2. N-[2-(7-methoxy-3-propyl-naphthalen-1-yl)ethyl]acetamide
(10b). Recrystallized from cyclohexane as a white solid (80% yield);
m.p. 84e86 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 0.90 (t, J ¼ 7.6, 3H,
CH2-c0), 1.65 (m, 2H, CH2-b0), 1.83 (s, 3H, CH3), 2.65 (t, J ¼ 7.6, 2H,
CH2-a0), 3.10 (t, J ¼ 8.4, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.95 (s, 3H,
OCH3), 7.12 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.19 (d, J ¼ 0.9, 1H, H-2),
7.48 (d, J ¼ 0.9, 1H, H-4), 7.56 (d, J ¼ 2.4, 1H, H-8), 7.75 (d, J ¼ 9.0, 1H,
H-5), 8.12 (br s, 1H, NH); 13C NMR (DMSO-d6): dC dC 22.5 Cec0, 23.5
CH3, 33.3 C-b, 33.8 C-b0, 39.8 C-a,a0, 55.5 OCH3, 103.1 C-8, 118.3 C-7,
125.5 C-6,128.9 C-4,129.5 C-10,130.1 C-5,131.7 C-2,134.6 C-9,137.6
C-1, 157.4 C-3, 169.9 CO; IR (neat, cmꢁ1): 3255 (NH), 1630 (C]O);
LC/MS m/z 286.4 (M þ 1); Anal. calcd for C18H23NO2: C 75.76%, H
8.12%, N 4.91%; Found: C 75.75%, H 8.10%, N 4.89%.
4.1.6.6. N-[2-(3-Cyclobutanecarbonyl-7-methoxy-naphthalen-1-yl)
ethyl]acetamide (9f). Recrystallized from toluene as a white solid
(5.33 g, 80% yield); m.p. 142e144 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 1.80 (s, 3H, CH3), 1.82 (m, 1H, H-30 ), 2.05 (m, 1H, H-30
a b), 2.28 (m,
4H, H-20,40), 3.16 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.97 (s, 3H,
OCH3), 4.23 (m, 1H, H-10), 7.27 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.68 (d,
J ¼ 2.0, 1H, H-8), 7.76 (d, J ¼ 2.4, 1H, H-2), 8.07 (d, J ¼ 9.0, 1H, H-5),
8.12 (br s, 1H, NH), 8.36 (d, J ¼ 2.0, 1H, H-4); IR (neat, cmꢁ1): 3378
(NH), 1673 (C]O), 1655 (C]O); LC/MS m/z 326.1 (M þ 1).
4.1.6.7. N-[2-(3-Cyclopentanecarbonyl-7-methoxy-naphthalen-1-yl)
ethyl]acetamide (9g). Recrystallized from acetonitrile as a white
solid (3.48 g, 50% yield); m.p. 134e136 ꢀC; 1H NMR (300 MHz,
DMSO-d6): dH 1.64 (m, 4H, H-30,40), 1.77 (m, 2H, H-20), 1.80 (s, 3H,
CH3), 1.94 (m, 2H, H-50), 3.16 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-a),
3.94 (m, 1H, H-10), 3.97 (s, 3H, OCH3), 7.27 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-
4.1.7.3. N-[2-(3-Isobutyl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(10c). Recrystallized from cyclohexane as a white solid (0.96 g, 50%
yield); m.p. 94e96 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 0.90 (d,
J ¼ 9.0, 6H, 2CH3-c0),1.83 (s, 3H, CH3),1.97 (m,1H, H-b0), 2.53 (m, 2H,
CH2-a0), 3.10 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.95 (s, 3H, OCH3),

318
M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
7.12 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.14 (d, J ¼ 2.0, 1H, H-2), 7.43 (d,
J ¼ 2.4, 1H, H-8), 7.56 (d, J ¼ 9.0, 1H, H-5), 7.75 (m, 1H, H-4), 8.10 (br
s, 1H, NH); 13C NMR (DMSO-d6): dC 22.7 Cec0, 23.0 CH3, 30.0 C-b0,
33.5 C-b, 45.1 C-a,a0, 55.6 OCH3, 102.9 C-8,118.4 C-7,126.2 C-6,128.9
C-4, 129.4 C-10, 130.0 C-5, 131.8 C-2, 134.3 C-9, 136.2 C-1, 157.4 C-3,
169.9 CO; IR (neat, cmꢁ1): 3253 (NH), 1636 C]O); LC/MS m/z 300.4
(M þ 1); Anal. calcd for C19H25NO2: C 76.22%, H 8.42%, N 4.68%;
Found: C 76.24%, H 8.46%, N 4.70%.
C21H27NO2: C 77.50%, H 8.36%, N 4.30%; Found: C 77.45%, H 8.39%,
N 4.25%.
4.1.7.8. N-[2-(3-Cyclohexylmethyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (10h). Recrystallized from acetonitrile as a white solid
(1.41 g, 65% yield); m.p. 106e108 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 0.92 (m, 2H, H-40), 1.14 (m, 2H, H-30), 1.58e1.64 (m, 6H, H-20,50
and 60), 1.83 (s, 3H, CH3), 2.53 (m, 2H, CH2-a0), 3.10 (m, 2H, CH2-a),
3.33 (m, 2H, CH2-b), 3.37 (m, 1H, H-10), 3.91 (s, 3H, OCH3), 7.12 (dd,
J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.15 (d, J ¼ 2.0, 1H, H-2), 7.43 (d, J ¼ 2.4, 1H,
H-8), 7.53 (d, J ¼ 2.0, 1H, H-4), 7.75 (d, J ¼ 9.0, 1H, H-5), 8.05 (br s,
1H, NH); 13C NMR (DMSO-d6): dC 23.2 CH3, 26.3 C-30 and 50, 26.5
C-40, 33.1 C-20 and 60, 33.5 C-b, 39.7 C-a and a0, 43.7 C-50, 55.8 OCH3,
103.1 C-8, 118.4 C-7, 126.3 C-6, 129.4 C-4, 129.6 C-10, 130.1 C-5, 131.7
C-2, 134.3 C-9, 135.9 C-1, 157.5 C-3, 169.9 CO; IR (neat, cmꢁ1): 3255
(NH), 1638 (C]O); LC/MS m/z 340.5 (M þ 1); Anal. calcd for
C22H29NO2: C 77.84%, H 8.61%, N 4.13%; Found: C 77.84%, H 8.59%,
N 4.07%.
4.1.7.4. N-{2-[7-Methoxy-3-(3-methylbutyl)naphthalen-1-yl]ethyl}
acetamide (10d). Recrystallized from cyclohexane as a white solid
(0.68 g, 34% yield); m.p. 72e74 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 0.92 (d, J ¼ 6.0, 6H, 2CH3-d0), 1.53 (m, 2H, CH2-b0), 1.58 (m, 1H,
H-c0), 1.83 (s, 3H, CH3), 2.67 (m, 2H, CH2-a0), 3.10 (m, 2H, CH2-a),
3.33 (m, 2H, CH2-b), 3.95 (s, 3H, OCH3), 7.12 (dd, J ¼ 9.0, J ¼ 2.4, 1H,
H-6), 7.18 (d, J ¼ 2.0, 1H, H-2), 7.49 (d, J ¼ 2.4, 1H, H-8), 7.56 (d,
J ¼ 9.0, 1H, H-4), 7.75 (d, J ¼ 8.9, 1H, H-5), 8.10 (br s, 1H, NH); 13C
NMR (DMSO-d6): dC 22.7 C-d0, 23.1 CH3, 27.7 Cec0, 33.4 C-b,
33.7 C-b0, 40.8 C-a,a0, 55.7 OCH3, 103.1 C-8, 118.5 C-7, 125.3 C-6,
128.9 C-4, 129.5 C-10, 130.0 C-5, 131.7 C-2, 134.6 C-9, 137.6 C-1, 157.4
C-3, 169.9 CO; IR (neat, cmꢁ1): 3254 (NH), 1637 C]O); LC/MS m/z
314.4 (M þ 1); Anal. calcd for C20H27NO2: C 76.64%, H 8.68%, N
4.47%; Found: C 76.67%, H 8.70%, N 4.47%.
4.1.7.9. N-[2-(3-Benzyl-7-methoxy-naphthalen-1-yl)ethyl]acetamide
(10i). Recrystallized from isopropyl ether as a white solid (0.81 g,
38% yield); m.p. 93e95 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.80 (s,
3H, CH3), 3.10 (t, J ¼ 8.2, 2H, CH2-a), 3.30 (m, 2H, CH2-b), 3.90 (s, 3H,
OCH3), 4.05 (s, 2H, CH2-a0), 7.12 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-8),
7.16e7.32 (m, 6H, H-8 and aromatic), 7.51e7.56 (m, 2H, H-2 and 4),
7.75 (d, J ¼ 9.0, 1H, H-5), 8.06 (br s, 1H, NH); 13C NMR (DMSO-d6): dC
23.1 CH3, 33.5 C-a, 41.0 C-a0, 41.5 C-b, 55.7 OCH3, 103.1 C-8, 118.6 C-
6, 126.0 C-2, 126.4 C-4, 128.9 C-30, 129.1 C-40, 129.2 C-20, 129.4 C-10,
130.1 C-5, 131.8 C-9, 134.8 C-1, 136.3 C-3, 141.8 C-10, 157.6 C-7, 169.9
CO; IR (neat, cmꢁ1): 3251 (NH), 1632 (C]O); LC/MS m/z 334.4
(M þ 1); Anal. calcd for C22H23NO2: C 79.25%, H 6.95%, N 4.20%;
Found: C 79.24%, H 6.91%, N 4.21%.
4.1.7.5. N-[2-(3-Cyclopropylmethyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (10e). Recrystallized from petroleum ether as a white
solid (1.14 g, 60% yield); m.p. 80e81 ꢀC; 1H NMR (300 MHz,
DMSO-d6): dH 0.90e1.40 (m, 5H, H-10,20 and 30), 1.83 (s, 3H, CH3),
3.00e3.70 (m, 6H, CH2-a,b and a0), 3.90 (s, 3H, OCH3), 7.10e7.30 (m,
2H, H-6 and 8), 7.40e7.60 (m, 2H, H-2 and 4), 7.80 (d, J ¼ 9.0, 1H, H-
5), 8.12 (br s, 1H, NH); 13C NMR (DMSO-d6): dC 23.0 CH3, 27.3 C-20
and 30, 33.7 C-10, 37.1 C-a and a0, 42.5 C-b, 55.6 OCH3,103.1 C-8,118.2
C-7, 125.4 C-6, 129.0 C-4, 129.5 C-10, 130.1 C-5, 131.9 C-2, 134.4 C-9,
135.8 C-1, 157.4 C-3, 170.0 CO; IR (neat, cmꢁ1): 3259 (NH), 1633 (C]
O); LC/MS m/z 298.3 (M þ 1); Anal. calcd for C19H23NO2: C 76.74%, H
7.80%, N 4.71%; Found: C 76.78%, H 7.81%, N 4.70%.
4.1.8. N-[2-(3-Bromo-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (12)
To a solution of compound 1 (7.0 g, 28.8 mmol) in 150 mL of
acetic acid at 70 ꢀC, was added dropwise cautiously a solution of
bromine (1.8 mL, 34.8 mmol) in 20 mL of acetic acid. The reaction
mixture was allowed to stir for 6 h. After cooling, the reaction
mixture was poured into an ice-water mixture and allowed to stir
for 30 min. The solid formed was filtered, washed with ethyl ether
and recrystallized from toluene to lead to the desired product
(6.77 g, 73% yield); White solid, m.p. 106e108 ꢀC; 1H NMR
(300 MHz, DMSO-d6): dH 1.80 (s, 3H, CH3), 3.12 (t, J ¼ 8.3, 2H, CH2-
a), 3.32 (m, 2H, CH2-b), 3.96 (s, 3H, OCH3), 7.18 (dd, J ¼ 9.0, J ¼ 2.4,
1H, H-6), 7.46 (d, J ¼ 2.0, 1H, H-2), 7.63 (d, J ¼ 2.4, 1H, H-8), 7.83 (d,
J ¼ 9.0, 1H, H-5), 8.00 (d, J ¼ 2.0, 1H, H-4), 8.14 (br s, 1H, NH); IR
(neat, cmꢁ1): 3300 (NH), 1633 (C]O); LC/MS m/z 323.2 (M þ 1).
4.1.7.6. N-[2-(3-Cyclobutylmethyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (10f). Recrystallized from cyclohexane as a white sol-
id (1.33 g, 67% yield); m.p. 108e110 ꢀC; 1H NMR (300 MHz, DMSO-
d6): dH 1.73e1.78 (m, 4H, H-20 and 40), 1.83 (s, 3H, CH3), 1.97 (m, 2H,
H-30), 2.61 (m, 1H, H-10), 2.75 (m, 2H, CH2-a0), 3.10 (m, 2H, CH2-a),
3.30 (m, 2H, CH2-b), 3.91 (s, 3H, OCH3), 7.10 (dd, J ¼ 9.0, J ¼ 2.4, 1H,
H-6), 7.15 (d, J ¼ 2.0, 1H, H-2), 7.43 (d, J ¼ 2.4, 1H, H-8), 7.56 (d,
J ¼ 2.0, 1H, H-4), 7.75 (d, J ¼ 9.0, 1H, H-5), 8.07 (br s, 1H, NH); 13C
NMR (DMSO-d6): dC 18.2 C-30, 23.1 CH3, 28.0 C-20 and 40, 33.5 C-10,
37.1 C-a and a0, 42.6 C-b, 55.6 OCH3, 103.0 C-8, 118.3 C-7, 125.4 C-6,
129.1 C-4, 129.5 C-10, 130.1 C-5, 131.9 C-2, 134.4 C-9, 135.8 C-1, 157.4
C-3, 170.0 CO; IR (neat, cmꢁ1): 3253 (NH), 1630 (C]O); LC/MS m/z
312.4 (M þ 1); Anal. calcd for C20H25NO2: C 77.14%, H 8.09%, N
4.50%; Found: C 77.10%, H 8.08%, N 4.47%.
4.1.9. N-[2-(7-Methoxy-3-morpholino-4-yl-naphthalen-1-yl)ethyl]
acetamide (13)
To a solution of 3-bromoagomelatine 12 (3.1 mmol) in 6.0 mL of
dioxane under an inert atmosphere, were added cesium carbonate
(1.40 g, 4.3 mmol), palladium dibenzylydeneacetone (0.006 mmol),
xantphos (0.007 mmol) and morpholine (3.7 mmol). The reaction
mixture was refluxed for 24 h, cooled and hydrolyzed with 100 mL
of water then acidified with an aqueous solution of hydrochloride
acid (1M). After washing with ethyl acetate, the aqueous phase was
alkalinized with an aqueous solution of potassium carbonate 10%
then extracted with ethyl acetate. The organic layer was dried over
MgSO4 and evaporated under reduced pressure to give the desired
product as a white solid after recrystallization from toluene (0.13 g,
12% yield); m.p. 152e154 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.84
(s, 3H, CH3), 3.17 (t, J ¼ 7.6, 2H, CH2-a), 3.36 (m, 2H, CH2-b), 3.54 (m,
4H, H-20and 60), 3.94 (s, 3H, OCH3), 4.04 (m, 4H, H-30 and 50), 4.94
4.1.7.7. N-[2-(3-Cyclopentylmethyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (10g). Recrystallized from acetonitrile as
a white
solid (1.47 g, 71% yield); m.p. 104e106 ꢀC; 1H NMR (300 MHz,
DMSO-d6): dH 1.20 (m, 2H, H-30), 1.55 (m, 2H, H-40), 1.60e1.64 (m,
4H, H-20 and 50), 1.83 (s, 3H, CH3), 2.14 (m, 1H, H-10), 2.67 (m, 2H,
CH2-a0), 3.10 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.91 (s, 3H, OCH3),
7.14 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.18 (d, J ¼ 2.0, 1H, H-2), 7.46 (d,
J ¼ 2.4, 1H, H-8), 7.54 (d, J ¼ 2.0, 1H, H-4), 7.73 (d, J ¼ 9.0, 1H, H-5),
8.07 (br s, 1H, NH); 13C NMR (DMSO-d6): dC 23.1 CH3, 25.0 C-30 and
40, 32.5 C-20 and 50, 33.5 C-b, 41.7 C-10, 41.9 C-a and a0, 55.7 OCH3,
103.1 C-8, 118.4 C-7, 125.8 C-6, 129.0 C-4,129.5 C-10,129.8 C-5, 131.7
C-2, 134.5 C-9, 136.9 C-1, 157.4 C-3, 169.6 CO; IR (neat, cmꢁ1): 3252
(NH), 1635 (C]O); LC/MS m/z 326.5 (M þ 1); Anal. calcd for

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
319
(br s, 2H, NHþ2 Clꢁ), 7.24 (d, J ¼ 9.0, 1H, H-6), 7.55 (s, 1H, H-4),
7.60e7.70 (m, 2H, H-2 and 8), 7.83 (d, J ¼ 9.0,1H, H-5), 8.22 (br s,1H,
NH); IR (neat, cmꢁ1): 3432-2427 (NH, NHþ2 Clꢁ), 1659 (C]O); LC/MS
m/z 365.6 (M þ 1); Anal. calcd for C19H24N2O3: C 69.49%, H 7.37%, N
8.50%; Found: C 69.50%, H 7.41%, N 8.51%.
262.3 (M þ 1); Anal. calcd for C15H16FNO2: C 68.95%, H 6.17%, N
5.36%; Found: C 68.99%, H 6.20%, N 5.35%.
4.1.13. N-[2-(7-Methoxy-3-vinyl-naphthalen-1-yl)ethyl]
acetamide (17)
To a solution of compound 12 (3.0 mmol) in 20 mL of dry toluene
4.1.10. N-{2-[7-Methoxy-3-(4-methoxy-benzylamino)-naphthalen-
1-yl]ethyl}acetamide (14)
under
an
inert
atmosphere
was
added
tetrakis(-
triphenylphosphine) palladium (0) (0.2 mmol). The mixture was
allowed to stir and heated at 100 ꢀC for 3 h after adding tribu-
tylvinyltin (4.5 mmol). After cooling, the reaction mixture was
poured into an aqueous solution of potassium fluoride (10%) and
extracted with ethyl acetate. The organic phase was washed with
water, dried over MgSO4 and evaporated under reduced pressure to
give the desired product as a white solid after recrystallization from
acetonitrile (0.40 g, 50% yield); m.p.109e110 ꢀC; 1H NMR (300 MHz,
DMSO-d6): dH 1.80 (s, 3H, CH3), 3.12 (m, 2H, CH2-a), 3.33 (m, 2H,
This compound was prepared from 12 according to the proce-
dure described for compound 13 by using p-methoxybenzylamine.
Recrystallized from toluene as a white solid (0.63 g, 54% yield); m.p.
142e144 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.80 (s, 3H, CH3),
2.98 (t, J ¼ 8.9, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.71 (s, 3H, OCH3),
3.84 (s, 3H, OCH3), 4.23 (s, 2H, CH2-a0), 6.14 (br s, 1H, NH amine),
6.50e7.50 (m, 9H, H aromatic), 8.10 (br s, 1H, NH amide); IR (neat,
cmꢁ1): 3399 (NH), 3249 (NH), 1632 (C]O); LC/MS m/z 379.5
(M þ 1).
CH2-b), 3.95 (s, 3H, OCH3), 5.27 (d, J ¼ 11.1, 1H, H-b0
a), 5.89 (d,
J ¼ 17.4, 1H, H-b0
b
), 6.60 (dd, J ¼ 17.4, J ¼ 11.1, 1H, H-a0), 7.16 (dd,
4.1.11. N-[2-(3-Amino-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (15)
J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.53 (d, J ¼ 2.0, 1H, H-2), 7.60 (d, J ¼ 2.4, 1H,
H-8), 7.70 (s,1H, H-4), 7.80 (d, J ¼ 9.0,1H, H-5), 8.10 (br s,1H, NH); IR
(neat, cmꢁ1): 3265 (NH), 1633 (C]O); LC/MS m/z 270.3 (M þ 1);
Anal. calcd for C17H19NO2: C 75.81%, H 7.11%, N 5.20%; Found: C
75.80%, H 7.08%, N 5.19%.
To a solution of compound 14 (0.9 g, 2.2 mmol) in 5 mL of
a mixture of ethanol-hydrochloride acid solution 0.5 M, was added
dropwise palladium hydroxide (0.076 g, 0.5 mmol). The reaction
mixture was placed under hydrogen flow and stirred at room
temperature for 12 h. After filtration of the catalyst, the filtrate
was alkalinized with an aqueous solution of sodium hydroxide
0.5 M then extracted with dichloromethane. The organic phase
was washed with water, dried over MgSO4 and evaporated under
reduced pressure to give the desired product after recrystallization
from toluene-cyclohexane (1/1) as a white solid (0.41 g, 73%
yield); m.p. 174e176 ꢀC; 1H NMR (300 MHz, CDCl3): dH 1.97 (s, 3H,
CH3), 3.19 (t, J ¼ 7.3, 2H, CH2-a), 3.63 (m, 2H, CH2-a), 3.72 (br s, 2H,
NH2), 3.95 (s, 3H, OCH3), 5.60 (s, 1H, NH), 6.83 (d, J ¼ 2.3, 1H, H-2),
6.91 (d, J ¼ 2.3, 1H, H-4), 7.10 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.33 (d,
J ¼ 2.4, 1H, H-8), 7.55 (d, J ¼ 9.0, 1H, H-5); 13C NMR (DMSO-d6): dC
23.1 CH3, 33.6 C-a, 40.1 C-b, 55.5 OCH3, 103.4 C-8, 106.2 C-6, 118.2
C-2, 119.9 C-4, 126.1 C-10, 127.8 C-5, 131.2 C-9, 135.2 C-1, 144.7 C-3,
154.7 C-7, 169.9 CO; IR (neat, cmꢁ1): 3421 and 3341 (NH2), 3265
(NH), 1636 (C]O); LC/MS m/z 259.3 (M þ 1); Anal. calcd for
C15H18N2O2: C 69.74%, H 7.02%, N 10.81%; Found: C 69.69%, H
7.00%, N 10.79%.
4.1.14. N-[2-(3-Allyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (18)
This compound was prepared from 12 by reaction with tribu-
tylallyltin according to the procedure described for compound 17.
Recrystallized from acetonitrile as a white solid (0.47 g, 56% yield);
m.p. 130e132 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.80 (s, 3H,
CH3), 3.12 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.50 (d, 2H, J ¼ 6.7,
CH2-a0), 3.95 (s, 3H, OCH3), 5.29 (d, 1H, J ¼ 11.0, H-c0
a), 5.89 (d, 1H,
J ¼ 17.4, H-c0
b
), 6.60 (dd, 1H, J ¼ 11.0, J ¼ 17.4, H-b0), 7.15 (dd, 1H,
J ¼ 9.0, J ¼ 2.4, H-6), 7.53 (d, 1H, J ¼ 2.0, H-2), 7.60 (d, 1H, J ¼ 2.4, H-
8), 7.70 (s, 1H, H-4), 7.80 (d, 1H, J ¼ 9.0, H-5), 8.10 (br s, 1H, NH); IR
(neat, cmꢁ1): 3265 (NH), 1633 (C]O); LC/MS m/z 284.3 (M þ 1);
Anal. calcd for C18H21NO2: C 76.30%, H 7.47%, N 4.94%; Found: C
76.35%, H 7.45%, N 4.98%.
4.1.15. N-[2-(3-Formyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (19)
To a solution of compound 17 (3.6 g, 13.4 mmol) and osmium
oxide (0.04 g, 0.16 mmol) in 70 mL of a mixture of tetrahydrofuran-
water (1/1), was added dropwise sodium periodate (5.7 mL,
26.9 mmol). The reaction mixture was stirred at room temperature
for 2 h, poured into a solution of hydrogenocarbonate 5% and the
mineral solid formed was filtered. The aqueous phase was then
extracted twice with ethyl ether and the organic phase was washed
with water, dried over MgSO4 and evaporated under reduced
pressure to give the desired product after recrystallization from
toluene; White solid (2.62 g, 72% yield); m.p. 146e148 ꢀC; 1H NMR
(300 MHz, CDCl3): dH 2.00 (s, 3H, CH3), 3.30 (t, J ¼ 7.0, 2H, CH2-a),
3.60 (m, 2H, CH2-b), 4.05 (s, 3H, OCH3), 5.75 (br s, 1H, NH), 7.25 (dd,
J ¼ 9.0, J ¼ 2.3, 1H, H-6), 7.65 (d, J ¼ 2.3, 1H, H-8), 7.78 (d, J ¼ 1.2, 1H,
H-2), 7.91 (d, J ¼ 9.0, 1H, H-5), 8.16 (s, 1H, H-4), 10.10 (s, 1H, CHO); IR
(neat, cmꢁ1): 3331 (NH),1693 (C]O),1649 (C]O); LC/MS m/z 272.3
(M þ 1).
4.1.12. N-[2-(3-Fluoro-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (16)
Compound 15 (2.3 g, 8.9 mmol) was dissolved in 30 mL of an
aqueous solution of HCl (0.4 M) and cooled at 0 ꢀC then a solution of
sodium nitrite (0.6 g, 8.9 mmol) in 5 mL of water was added. Next,
a solution of fluoroboric acid (48%) (5.4 mL, 37.1 mmol) was added
to the mixture, and the precipitate formed was filtered then taken
up with xylene and refluxed. After cooling, the mixture was
hydrolyzed and extracted with dichloromethane. The organic
phase was washed with water, dried (MgSO4) and evaporated
under reduced pressure to give the desired product after purifica-
tion on a silica gel column eluted by a mixture of acetone-
cyclohexane (4/6). Recrystallized from cyclohexane as a white
solid (0.23 g, 10% yield); m.p. 133e134 ꢀC; 1H NMR (300 MHz,
CDCl3): dH 2.06 (s, 3H, CH3), 3.27 (t, J ¼ 7.5, 2H, CH2-a), 3.65 (m, 2H,
CH2-b), 4.05 (s, 3H, OCH3), 5.65 (br s, 1H, NH), 7.11 (dd, J ¼ 9.9,
J ¼ 2.6, 1H, H-2), 7.21 (dd, J ¼ 8.8, J ¼ 2.2, 1H, H-6), 7.31 (dd, J ¼ 9.9,
J ¼ 2.6, 1H, H-4), 7.49 (d, J ¼ 2.4, 1H, H-8), 7.71 (d, J ¼ 8.8, 1H, H-5);
13C NMR (DMSO-d6): dC 23.1 CH3, 33.1 C-a, 39.9 C-b, 55.8 OCH3,
103.5 C-8, 109.1 and 109.9 (2J ¼ 20.0) C-2, 116.9 and 117.2 (2J ¼ 25.0)
C-4, 119.8 C-6, 129.7 and 129.8 (3J ¼ 9.0) C-10, 129.9 C-5, 130.0 C-9,
138.1 and 138.2 (3J ¼ 9.0) C-1, 156.9 and 157.5 (1J ¼ 39.0) C-3, 160.1
C-7, 174.0 CO; IR (neat, cmꢁ1): 3243 (NH), 1640 (C]O); LC/MS m/z
4.1.16. N-{2-[3-Hydroxyiminomethyl)-7-methoxy-naphthalen-1-
yl)]ethyl}acetamide (20)
To a solution of compound 19 (4.2 mmol) in methanol (250 mL)
was added hydroxylamine hydrochloride (16.8 mmol) and pyridine
(21.1 mmol). The reaction mixture was refluxed for 3 h, cooled and
hydrolyzed with water then the precipitate formed was filtered and
washed leading to the desired product as a white solid after

320
M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
recrystallization from ethanol (0.90 g, 75% yield); m.p. 98e100 ꢀC;
1H NMR (300 MHz, DMSO-d6): dH 1.82 (s, 3H, CH3), 3.13 (m, 2H,
CH2-a), 3.33 (m, 2H, CH2-b), 3.94 (s, 3H, OCH3), 7.18 (dd, J ¼ 9.0,
J ¼ 2.3, 1H, H-6), 7.65e7.75 (m, 2H, H-2 and 8), 7.80e7.95 (m, 2H, H-
4 and 5), 8.11 (br s, 1H, NH), 8.20 (s, 1H, H-a0), 11.15 (br s, 1H, OH); IR
(neat, cmꢁ1): 3436 (OH), 3279 (NH), 1645 (C]O); LC/MS m/z 287.3
(M þ 1).
desired product in 53% yield (1.34 g). To a solution of the obtained
alcohol (7.0 mmol) in 25 mL of dry dichloromethane at 0 ꢀC, was
added dropwise diethylaminosulfur trifluoride (DAST) (13.9 mmol)
and the mixture was allowed to warm to room temperature, poured
into water (100 mL) and acidified with an aqueous solution of
hydrochloride acid (1 M). The mixture was then extracted with
dichloromethane and the organic phase was washed with water,
dried over MgSO4 and evaporated under reduced pressure to give
the desired product as a white solid after recrystallization from
acetonitrile (0.96 g, 50% yield); m.p. 118e120 ꢀC; 1H NMR
(300 MHz, DMSO-d6): dH 1.83 (s, 3H, CH3), 3.14 (t, J ¼ 6.7, 2H, CH2-a),
3.34 (m, 2H, CH2-b), 3.96 (s, 3H, OCH3), 5.50 (d,, J ¼ 48.2, 2H, CH2-
a0), 7.21 (dd, J ¼ 9.0, J ¼ 2.4,1H, H-6), 7.39 (s,1H, H-2), 7.65 (d, J ¼ 2.4,
1H, H-8), 7.78 (s, 1H, H-4), 7.87 (d, J ¼ 9.0, 1H, H-5), 8.12 (br s, 1H,
NH); IR (neat, cmꢁ1): 3254 (NH), 1635 (C]O); LC/MS m/z 276.3
(M þ 1); Anal. calcd for C16H18FNO2: C 69.80%, H 6.59%, N 5.09%;
Found: C 69.79%, H 6.66%, N 5.08%.
4.1.17. N-[2-(3-Aminomethyl-7-methoxynaphthalen-1-yl)]ethyl}
acetamide hydrochloride (21)
Compound 20 (2.8 mmol) was dissolved in 75 mL of methanol
and Raney nickel (0.1 g) was added. The mixture was stirred and
saturated with ammonia then placed under hydrogen pressure
(20 bars) for 72 h. After the catalyst filtration, solvent was evapo-
rated and the obtained residue was taken up with ethyl ether then
HCl gas was bubbled in. The precipitate formed was filtered and
recrystallized from acetonitrile; Yellow solid (0.4 g, 46% yield); m.p.
134e136 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.85 (s, 3H, CH3), 3.10
(m, 2H, CH2-a), 3.35 (m, 2H, CH2-b), 3.95 (s, 3H, OCH3), 4.10 (m, 2H,
CH2-a0), 7.20 (dd, J ¼ 9.0, J ¼ 2.3, 1H, H-6), 7.47 (d, J ¼ 1.5, 1H, H-2),
7.60 (d, J ¼ 2.3,1H, H-8), 7.70e7.90 (m, 2H, H-4 and 5), 8.25 (br s,1H,
NH), 8.45 (br s, 3H, NH3þClꢁ); IR (neat, cmꢁ1): 3420-2700 (NH,
NHþ3 Clꢁ), 1630 (C]O); LC/MS m/z 309.6 (M þ 1).
4.1.20. N-{2-[3-(1E-Hydroxyiminoethyl)-7-methoxy-naphthalen-1-
yl]ethyl}acetamide (24)
To a solution of the ketone 9a (1.3 g, 4.2 mmol) in 25 mL of
methanol were added respectively hydroxylamine hydrochloride
(1.17 g, 16.9 mmol) and pyridine (1.7 mL, 21.1 mmol). The reaction
mixture was refluxed for 3 h and allowed to cool then poured into
water (50 mL) and the solid formed was filtered, washed with
water and recrystallized from ethanol 95ꢀ to give the desired
product as a white solid (1.03 g, 82% yield); m.p. 143e145 ꢀC; 1H
NMR (300 MHz, DMSO-d6): dH 1.80 (s, 3H, CH3), 2.23 (s, 3H, CH2-b0),
3.12 (t, J ¼ 8.3, 2H, CH2-a), 3.30 (m, 2H, CH2-b), 3.93 (s, 3H, OCH3),
7.18 (dd, J ¼ 9.0, J ¼ 2.4,1H, H-6), 7.60 (d, J ¼ 2.4,1H, H-8), 7.73 (s,1H,
H-2), 7.89 (d, J ¼ 9.0, 1H, H-5), 7.93 (s, 1H, H-4), 8.12 (br s, 1H, NH),
11.18 (s, 1H, OH); 13C NMR (DMSO-d6): dC 11.7 C-b0, 23.1 CH3, 33.6 C-
a, 40.0 C-b, 55.8 OCH3, 103.3 C-8, 118.7 C-6, 124.4 C-2, 124.6 C-4,
129.0 C-10, 131.2 C-5, 132.0 C-9, 133.5 C-1, 134.6 C-3, 153.2 Cb0, 158.4
C-7, 170.0 CO; IR (neat, cmꢁ1): 3462 (NH), 3272 (OH), 1633 (C]O);
LC/MS m/z 301.4 (M þ 1); Anal. calcd for C17H20N2O3: C 67.98%, H
6.71%, N 9.33%; Found: C 68.00%, H 6.70%, N 9.38%.
4.1.18. 2-{4-[2-(Acetylamino)ethyl]-6-methoxy}naphthoic acid (22)
To a solution of compound 9a (1.0 g, 3.5 mmol) in 56 mL of
a mixture 1,4-dioxane-water (1/1), was added potassium iodide
(1.3 g, 15.4 mmol) and a solution of sodium hypochlorite 9.6%
(18.0 mL, 14.0 mmol). The reaction mixture was allowed to stir at
room temperature then filtered. The filtrate was treated with an
aqueous solution of sodium thiosulfate, acidified with an aqueous
solution of hydrochloride acid (3 M) and extracted with ethyl
acetate. The organic phase was washed with water, dried over
MgSO4 and evaporated under reduced pressure to give the desired
product after recrystallization from 1,4-dioxane; Beige solid (0.70 g,
70% yield); m.p. 248e250 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.80
(s, 3H, CH3), 3.12 (m, 2H, CH2-a), 3.30 (m, 2H, CH2-b), 4.00 (s, 3H,
OCH3), 7.25 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.67 (d, J ¼ 2.4, 1H, H-8),
7.79 (d, J ¼ 2.0, 1H, H-2), 8.00 (d, J ¼ 9.0, 1H, H-5), 8.12 (br s, 1H, NH),
8.40 (d, J ¼ 2.0, 1H, H-4), 12.80 (br s, 1H, OH); IR (neat, cmꢁ1): 3363
(NH), 3200-2500 (OH), 1693 (C]O); LC/MS m/z 288.3 (M þ 1).
4.1.21. N-{2-[3-(1-Aminoethyl)-7-methoxy-naphthalen-1-yl]ethyl}
acetamide hydrochloride (25)
Compound 24 (0.85 g, 2.8 mmol) was dissolved in 50 mL of
methanol and 0.1 g of Raney nickel was added. The reaction
mixture was placed in a high pressure tube under 20 bars hydrogen
pressure, and allowed to stir at room temperature for 3 days. After
filtration and evaporation under reduced pressure, the residue
obtained was dissolved in ethyl ether and treated with HCl gas to
give the desired product as a white solid (0.31 g, 34% yield) after
recrystallization from ethanol-ethyl ether (1/1); m.p. 239e241 ꢀC;
1H NMR (300 MHz, DMSO-d6): dH 1.60 (d, J ¼ 6.7, 3H, H-b0), 1.86
(s, 3H, CH3), 3.12 (t, J ¼ 6.4, 2H, CH2-a), 3.63 (m, 2H, CH2-b), 3.93 (s,
3H, OCH3), 4.49 (m, 1H, H-a0), 7.22 (dd, J ¼ 9.0, J ¼ 2.0, 1H, H-6), 7.52
(s, 1H, H-2), 7.62 (s, 1H, H-4), 7.79e7.86 (m, 2H, H-5 and 8), 8.27 (br
s, 1H, NH), 8.61 (br s, 3H, NHþ3 Clꢁ); IR (neat, cmꢁ1): 3254 (NH),
3200-2500 (NHþ3 Clꢁ) 1632 (C]O); LC/MS m/z 322.7 (M þ 1).
4.1.19. N-[2-(3-Fluoromethyl-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (23)
This compound was synthesized via two steps sequence: (1)
treatment of compound 22 with thionyl chloride in methanol to
afford the corresponding ester (methyl 2-{4-[2-(acetylamino)
ethyl]-6-methoxy-naphthoate), (2) followed by reduction of the
ester function to hydroxyl by lithium aluminum hydride. Conse-
quently, to a solution of the acid 22 (4.2 g, 14.6 mmol) in methanol
(100 mL) at 0 ꢀC, was added dropwise cautiously thionyl chloride
(1.6 mL, 22.0 mmol) and the mixture was allowed to stir at room
temperature for 1 h. After concentration under reduced pressure,
the obtained residue was poured into water (100 mL) and extracted
with ethyl ether (2 ꢄ 50 mL). The organic layers were washed, dried
over MgSO4 and evaporated under reduced pressure to give the
desired product after recrystallization from acetonitrile (2.72 g, 62%
yield). This compound (2.8 g, 9.3 mmol) was then dissolved in dry
tetrahydrofuran (75 mL) and cooled in an ice-bath. Aluminum
lithium hydride (0.7 g, 18.6 mmol) was added dropwise. The reac-
tion mixture was allowed to stir for 2 h then hydrolyzed simulta-
neously with 0.7 mL of water, 0.7 mL of a solution of sodium
hydroxide (15%) and 2.1 mL of water. The solid formed was filtered
and the filtrate washed, dried over MgSO4 and evaporated under
reduced pressure. The recrystallization from acetonitrile gave the
4.1.22. N-[2-(3-Acetylamino-7-methoxy-naphthalen-1-yl)ethyl]
acetamide (26)
To a solution of compound 24 (1.5 g, 5.0 mmol) in dry
dichloromethane (25 mL) was added dropwise cautiously phos-
phorus pentachloride (2.1 g, 10.0 mmol). The reaction mixture was
allowed to stir at room temperature for 6 h, then poured into water
and extracted with dichloromethane. The organic phase was
washed with water, dried over MgSO4 and evaporated under
reduced pressure to give the desired product as a yellow solid
(1.07 g, 71% yield); m.p. 182e184 ꢀC; 1H NMR (300 MHz, DMSO-d6):

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
321
dH 1.80 (s, 3H, CH3), 2.07 (s, 3H, CH3), 3.12 (t, J ¼ 8.3, 2H, CH2-a), 3.34
(m, 2H, CH2-b), 3.93 (s, 3H, OCH3), 7.12 (dd, J ¼ 9.0, J ¼ 2.4, 1H, H-6),
7.38 (d, J ¼ 2.0, 1H, H-2), 7.52 (d, J ¼ 2.4, 1H, H-8), 7.72 (d, J ¼ 2.0, 1H,
H-5), 8.07e8.15 (m, 2H, H-4 and NH), 10.00 (br s, 1H, NH); 13C NMR
(DMSO-d6): dC 23.1 CH3, 24.5 CH3, 33.5 C-a, 40.0 C-b, 55.7 OCH3,
103.1 C-8, 114.8 C-6, 118.9 C-2, 121.3 C-4, 129.6 C-10, 129.9 C-5, 130.0
C-9, 135.0 C-1, 135.2 C-3, 157.0 C-7, 168.8 CO, 169.9 CO; IR (neat,
cmꢁ1): 3297 (NH), 1643 (C]O); LC/MS m/z 301.4 (M þ 1); Anal.
calcd for C17H23ClN2O2: C 63.24%, H 7.19%, N 9.91%; Found: C
63.23%, H 7.24%, N 10.06%.
C-6, 124.7 C-3, 127.2 C-4, 127.6 C-2, 130.9 C-10, 131.2 and 131.3
(3J ¼ 9.0) C-5, 134.5 and 134.6 C-1, 136.0 C-9, 159.3 and 162.6
(1J ¼ 248.0) C-7, 170.2 CO; IR (neat, cmꢁ1): 3244 (NH), 1633 (C]O);
LC/MS m/z 232.3 (M þ 1); Anal. calcd for C14H14FNO: C 72.70%, H
6.10%, N 6.06%; Found: C 72.68%, H 6.11%, N 6.10%.
4.1.27. General procedure for the synthesis of compounds 31e33
To a solution of N-[2-(7-hydroxy-naphthalen-1-yl)ethyl]acet-
amide (4.4 mmol) in dry acetone (50 mL) were added potassium
carbonate (13.1 mmol) and tetrabutylammonium bromide
(0.4 mmol). The reaction mixture was refluxed for 30 min, then 3-
bromo-1-propanol, 1-bromo-2-fluoroethane (6.6 mmol) or 1,1,1-
trifluoro-2-iodoethane (6.6 mmol) was added and the mixture
was refluxed for 5 h. After concentration of the solvent the residue
was taken up with 100 mL of water and extracted with ethyl
acetate. The organic phase was washed with water, dried and
evaporated to lead to desired product.
4.1.23. N-[2-(7-Hydroxy-naphthalen-1-yl)ethyl]acetamide (27)
Compound 1 (8.5 g, 35.0 mmol) was dissolved in dichloro-
methane (250 mL) at
0
ꢀC and boron tribromide (6.7 mL,
70.0 mmol) was added dropwise cautiously. The reaction mixture
was allowed to stir at 0 ꢀC for 2 h then hydrolyzed and the two
phases were separated. The organic layer was washed, dried and
evaporated under reduced pressure to give the desired product
after recrystallization from toluene as a white solid (6.42 g, 80%
yield); m.p. 123e125 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.80 (s,
3H, CH3), 3.04 (t, J ¼ 7.4, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 7.10 (dd,
J ¼ 9.0, J ¼ 2.1, 1H, H-6), 7.15e7.25 (m, 2H, H-2 and 3), 7.32 (d, J ¼ 2.1,
1H, H-8), 7.65 (d, J ¼ 7.4,1H, H-4), 7.76 (d, J ¼ 9.0,1H, H-5), 8.02 (br s,
1H, NH), 9.75 (br s, 1H, OH); IR (neat, cmꢁ1): 3345, 3102, 1639; LC/
MS m/z 230.3 (M þ 1).
4.1.27.1. N-{2-[7-(3-Hydroxypropoxy)-naphthalen-1-yl]ethyl}acetamide
(31). Recrystallized from acetonitrile as a white solid (0.86 g,
68% yield); m.p. 143e145 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.83
(s, 3H, CH3), 1.96 (q, 2H, CH2-b0), 3.12 (t, J ¼ 8.4, 2H, CH2-a), 3.33 (m,
2H, CH2-b), 3.63 (m, 2H, CH2-c0), 4.24 (t, J ¼ 6.1, 2H, CH2-a0), 4.62 (t,
J ¼ 5.2, 1H, OH), 7.14e7.34 (m, 3H, H-2,3 and 6), 7.56 (d, J ¼ 2.0, 1H,
H-8), 7.71 (d, J ¼ 4.1, 1H, H-4), 7.83 (d, J ¼ 9.1, 1H, H-5), 8.11 (br s, 1H,
NH); IR (neat, cmꢁ1): 3298 (OH), 3240 (NH), 1637 (C]O); LC/MS m/
z 288.3 (M þ 1).
4.1.24. N-[2-(7-Trifluoromethansulfonyl-naphthalen-1-yl)ethyl]
acetamide (28)
To a solution of N-[2-(7-hydroxy-naphthalen-1-yl)ethyl]acet-
amide (7.5 g, 33.0 mmol) in 50 mL of dichloromethane was added
triethylamine (13.8 mL, 100.0 mmol). The mixture was cooled
at ꢁ20 ꢀC then trifluoromethanesulfonic anhydride acid (6.0 mL,
35.0 mmol) was added dropwise cautiously. The reaction mixture
was allowed to warm to room temperature for 3 h then hydrolyzed
and the organic phase was washed twice with an aqueous solution
of sodium hydroxide 1 M then with water, dried (MgSO4) and
evaporated under reduced pressure to give the desired product
after purification on a silica gel column eluted with a mixture
acetone-cyclohexane (3/7). Recrystallized from cyclohexane as
a white solid (7.75 g, 65% yield); m.p. 81e83 ꢀC; 1H NMR (300 MHz,
DMSO-d6): dH 1.81 (s, 3H, CH3), 3.18 (t, J ¼ 7.7, 2H, CH2-a), 3.33 (m,
2H, CH2-b), 7.40e7.60 (m, 3H, H-2,3 and 6), 7.93 (dd, J ¼ 8.1, J ¼ 2.1,
1H, H-4), 8.06 (br s, 1H, NH), 8.15 (d, J ¼ 8.1, 1H, H-5), 8.31 (d, J ¼ 2.1,
1H, H-8); IR (neat, cmꢁ1): 3300 (NH), 1620 (C]O), 1130 (SO2); LC/
MS m/z 362.3 (M þ 1).
4.1.27.2. N-{2-[7-(2-Fluoro-ethoxy)-naphthalen-1-yl]ethyl}acetamide
(32). Recrystallized from isopropyl ether as a white solid (0.91 g,
75% yield); m.p. 60e62 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.84 (s,
3H, CH3), 3.12 (t, J ¼ 6.2, 2H, CH2-a), 3.34 (m, 2H, CH2-b), 4.45 (td,
J ¼ 30.4, J ¼ 7.3, 2H, CH2-a0), 4.95 (td, J ¼ 47.9, J ¼ 7.3, 2H, CH2-b0),
7.18e7.40 (m, 3H, H-2,3 and 6), 7.64 (s, 1H, H-8), 7.74 (d, J ¼ 7.6, 1H,
H-4), 7.87 (d, J ¼ 9.0, 1H, H-5), 8.15 (br s, 1H, NH); 13C NMR (DMSO-
d6): dC 23.1 CH3, 33.5 C-a, 40.1 C-b, 67.5 and 67.7 (2J ¼ 19.0) C-a0, 81.5
and 83.7 (1J ¼ 167.0) C-b0, 104.1 C-8, 118.5 C-6, 123.8 C-3, 127.0 C-2,
127.5 C-4,129.4 C-10,130.7 C-5,133.3 C-9,134.7 C-1, 156.7 C-7,170.0
CO; IR (neat, cmꢁ1): 3311 (NH), 1648 (C]O); LC/MS m/z 276.3
(M þ 1); Anal. calcd for C16H18FNO2: C 69.80%, H 6.59%, N 5.09%;
Found: C 69.74%, H 6.59%, N 5.17%.
4.1.27.3. N-{2-[7-(2,2,2-Trifluoroethoxy)-naphthalen-1-yl]ethyl}acet-
amide (33). Recrystallized from acetonitrile as a white solid (0.13 g,
10% yield); m.p. 79e81 ꢀC; 1H NMR (300 MHz, CDCl3): dH 1.87 (s,
3H, CH3), 3.27 (t, J ¼ 6.0, 2H, CH2-a), 3.61 (m, 2H, CH2-b), 4.66 (q,
J ¼ 8.3, 2H, CH2-a0), 5.64 (br s, 1H, NH), 7.24 (dd, J ¼ 9.0, J ¼ 2.4, 1H,
H-6), 7.30e7.35 (m, 2H, H-2 and 4), 7.67 (d, J ¼ 2.4, 1H, H-8), 7.73 (t,
J ¼ 4.5, 1H, H-3), 7.84 (d, J ¼ 9.0, 1H, H-5); 13C NMR (DMSO-d6): dC
23.1 CH3, 33.4 C-a, 39.5 C-b, 64.2 C-a0, 64.8 CF3, 105.1 C-8, 118.0 C-6,
124.4 C-3,127.0 C-2,127.7 C-4,129.9 C-10,131.0 C-5, 133.0 C-9,135.0
C-1, 155.4 C-7, 170.0 CO; IR (neat, cmꢁ1): 3288 (NH), 1635 (C]O);
LC/MS m/z 312.3 (M þ 1); Anal. calcd for C16H16F3NO2: C 61.73%, H
5.18%, N 4.50%; Found: C 61.76%, H 5.14%, N 4.52%.
4.1.25. N-[2-(7-Amino-naphthalen-1-yl)ethyl]acetamide (29)
This compound was prepared from N-(2-{7-[(4-Methoxybenzyl)
amino]-naphthalen-1-yl}ethyl)acetamide according to the proce-
dure described for compound 15; Brown oil (1.14 g, 59% yield); 1H
NMR (300 MHz, CDCl3): dH 1.95 (s, 3H, CH3), 3.15 (t, J ¼ 7.0, 2H, CH2-
a), 3.55e3.65 (m, 4H, CH2-b and NH2), 5.60 (br s, 1H, NH), 6.95 (dd,
J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.17e7.25 (m, 3H, H-2,3 and 8), 7.61 (d,
J ¼ 8.0, 1H, H-4), 7.68 (d, J ¼ 9.0, 1H, H-5); IR (neat, cmꢁ1): 3408,
3340 (NH and NH2), 1651 (C]O); LC/MS m/z 229.3 (M þ 1).
4.1.28. N-{2-[7-(3-Fluoropropoxy)-naphthalen-1-yl]ethyl}
acetamide (34)
4.1.26. N-[2-(7-Fluoro-naphthalen-1-yl)ethyl]acetamide (30)
This compound was synthesized from 29 according to the
procedure described for compound 16. Recrystallized from cyclo-
hexane as a white solid (0.27 g, 13% yield); m.p. 136e138 ꢀC; 1H
NMR (300 MHz, CDCl3): dH 1.95 (s, 3H, CH3), 3.25 (t, J ¼ 7.6, 2H, CH2-
a), 3.65 (q, J ¼ 7.2, 2H, CH2-b), 5.50 (br s, 1H, NH), 7.29 (m, 1H, H-6),
7.33e7.43 (m, 2H, H-2 and 3), 7.67e7.78 (m, 2H, H-4 and 8), 7.87
(dd, J ¼ 9.1, J ¼ 5.9, 1H, H-5); 13C NMR (CDCl3): dC 23.4 CH3, 32.9 C-a,
40.1 C-b, 107.3 and 107.5 (2J ¼ 21.0) C-8, 115.9 and 116.3 (2J ¼ 24.0)
Prepared from compound 31 according to the procedure
described above for compound 23. Recrystallized from cyclohexane
as a white solid (0.30 g, 15% yield); m.p. 87e89 ꢀC; 1H NMR
(300 MHz, CDCl3): dH 1.95 (s, 3H, CH3), 3.25 (t, J ¼ 7.0, 2H, CH2-a),
3.30 (m, 2H, CH2-b0), 3.45 (m, 2H, CH2-b), 4.30 (t, J ¼ 6.0, 2H, CH2-
a0), 4.75 (dd, J ¼ 47.1, J ¼ 5.9, 2H, CH2-c0), 5.60 (br s, 1H, NH), 7.18 (dd,
J ¼ 9.0, J ¼ 2.4, 1H, H-6), 7.26e7.33 (m, 2H, H-2 and 3), 7.47 (d,
J ¼ 2.4, 1H, H-8), 7.70 (dd, J ¼ 6.0, J ¼ 3.3, 1H, H-4), 7.79 (d, J ¼ 9.0,

322
M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
1H, H-5); 13C NMR (CDCl3): dC 23.4 CH3, 30.3 and 30.5 (2J ¼ 20.2) C-
b0, 33.1 C-a, 40.1 C-b, 63.6 and 63.7 (3J ¼ 5.7) C-a0, 79.8 and 82.0
(1J ¼ 163.9) Cec0,103.3 C-8,118.5 C-6,123.2 C-3,127.1 C-2, 127.2 C-4,
129.4 C-10, 130.3 C-5, 133.2 C-9, 133.7 C-1, 157.1 C-7, 170.2 CO; IR
(neat, cmꢁ1): 3280 (NH), 1633 (C]O); LC/MS m/z 290.4 (M þ 1);
Anal. calcd for C17H20FNO2: C 70.57%, H 6.97%, N 4.84%; Found: C
70.52%, H 7.00%, N 4.86%.
temperature and stirred for 48 h, concentrated under reduced
pressure. The obtained residue was dissolved in water (10 mL) and
acidified with an aqueous solution of HCl (1 M). The mixture was
then extracted with ethyl acetate and the organic phase was
washed with saturated NaHCO3 and water, dried and evaporated.
The obtained product was then treated with aluminum lithium
hydride as described above for compound 37 to furnish the desired
product after recrystallization from toluene as a white solid (2.89 g,
81% yield); m.p. 198e200 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.97
(m, 2H, CH2-b), 2.86 (m, 2H, CH2-a), 3.08 (m, 2H, CH2-c), 3.86 (s, 3H,
OCH3), 7.06 (d, J ¼ 2.0, 1H, H-2), 7.19 (d, J ¼ 2.0, 1H, H-4), 7.34e7.50
(m, 2H, H-6 and 7), 7.81 (d, J ¼ 8.1, 1H, H-5), 8.01 (d, J ¼ 8.4, 1H, H-8),
8.15 (br s, 3H, NHþ3 Clꢁ); IR (neat, cmꢁ1): 2959e2744 (NHþ3 Clꢁ); LC/
MS m/z 251.6 (M þ 1).
4.1.29. 2-(3-Methoxy-naphthalen-1-yl)acetonitrile (35)
To a solution of compound B (3.5 g, 14.5 mmol) in 40 mL of
anhydrous toluene under an inert atmosphere, was added drop-
wise palladium dichloro-di(triphenylphosphine) (0.5 g, 0.7 mmol)
and tributyl cyanomethyltin (5.5 g, 16.5 mmol) freshly prepared in
20 mL of anhydrous toluene. The reaction mixture was refluxed for
24 h, cooled and hydrolyzed with saturated solution of potassium
fluoride then filtered on celite. The filtrate was extracted with ethyl
acetate and the organic phase was washed with water, dried and
evaporated to give the desired product after purification on a silica
gel column eluted by a mixture of ethyl acetate-cyclohexane (1/9);
Brown solid (2.66 g, 93% yield); m.p. 81e83 ꢀC; 1H NMR (300 MHz,
CDCl3): dH 3.94 (s, 3H, OCH3), 4.08 (s, 2H, CH2-a), 7.14 (d, J ¼ 2.4, 1H,
H-2), 7.30 (d, J ¼ 2.4, 1H, H-4), 7.43e7.55 (m, 2H, H-6 and 7),
7.75e7.82 (m, 2H, H-5 and 8); IR (neat, cmꢁ1): 2251 (CN); LC/MS m/
z 198.2 (M þ 1).
4.1.33. N-[2-(3-Methoxy-naphthalen-1-yl)ethyl]acetamide (39)
Synthesized from 37 according to the procedure described for
compound 3 and recrystallized from toluene as a white solid
(4.03 g, 79% yield); m.p. 151e153 ꢀC; 1H NMR (300 MHz, DMSO-d6):
dH 1.78 (s, 3H, CH3), 3.11 (m, 2H, CH2-a), 3.33 (m, 2H, CH2-b), 3.86 (s,
3H, OCH3), 7.01 (d, J ¼ 2.4, 1H, H-2), 7.19 (d, J ¼ 2.4, 1H, H-4),
7.35e7.47 (m, 2H, H-6 and 7), 7.81 (d, J ¼ 8.1, 1H, H-5), 8.03 (m, 2H,
H-8 and NH); 13C NMR (DMSO-d6): dC 23.2 CH3, 30.6 C-a, 38.8 C-b,
55.6 OCH3, 104.9 C-4, 119.0 C-2, 124.0 C-7, 124.1 C-6, 126.5 C-5, 127.5
C-9, 128.1 C-8, 135.5 C-10, 140.2 C-1, 157.2 C-3, 169.6 CO; IR (neat,
cmꢁ1): 3246 (NH), 1638 (C]O); LC/MS m/z 244.3 (M þ 1); Anal.
calcd for C15H17NO2: C 74.05%, H 7.04%, N 5.76%; Found: C 74.02%, H
7.00%, N 5.80%.
4.1.30. (Z/E) 3-(3-methoxy-naphthalen-1-yl)acrylonitrile (36)
To
a solution of 1-bromo-3-methoxy-naphthalene (4.0 g,
16.9 mmol) in 150 mL of dry DMF was added dropwise acrylonitrile
(1.7 mL, 25.3 mmol), palladium acetate (0.2 g, 0.8 mmol), triphe-
nylphosphine (0.4 g, 1.7 mmol) and triethylamine (3.5 mL,
25.3 mmol). The reaction mixture was allowed to stir and heated at
100 ꢀC for 5 h. After cooling, the mixture was taken up with satu-
rated solution of NaHCO3 and extracted with ethyl acetate. The
organic phase was washed with water, dried over MgSO4 and
evaporated under reduced pressure to give the desired product
after purification on silica gel column eluted by a mixture of ethyl
acetate-cyclohexane (1/9); Yellow solid (3.46 g, 98% yield); m.p.
72e74 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 3.93 (s, 3H, OCH3), 6.58
(d, J ¼ 16.5, 1H, CH2-a), 7.41e7.57 (m, 3H, H-4,6 and 7), 7.60 (d,
J ¼ 2.4, 1H, H-2), 7.88 (d, J ¼ 8.1, 1H, H-5), 8.22 (d, J ¼ 9.1, 1H, H-8),
8.48 (d, J ¼ 16.5, 1H, H-b); IR (neat, cmꢁ1): 2216 (CN); LC/MS m/z
210.2 (M þ 1).
4.1.34. N-[3-(3-Methoxy-naphthalen-1-yl)propyl]acetamide (40)
Prepared from 38 as for compound
3 above, and then
recrystallized from toluene as a white solid (4.05 g, 75% yield);
m.p. 106e108 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 1.75 (m, 2H,
CH2-b), 1.81 (s, 3H, CH3), 2.95 (m, 2H, CH2-a), 3.11 (m, 2H, CH2-c),
3.83 (s, 3H, OCH3), 7.01 (d, J ¼ 2.4, 1H, H-2), 7.14 (d, J ¼ 2.4, 1H,
H-4), 7.31e7.47 (m, 2H, H-6 and 7), 7.79 (d, J ¼ 8.1, 1H, H-5), 7.92
(d, J ¼ 8.4, 1H, H-8), 7.98 (br s, 1H, NH); 13C NMR (DMSO-d6): dC
23.2 CH3, 30.0 C-b, 30.6 C-a, 38.8 Cec, 55.6 OCH3, 104.9 C-4, 119.0
C-2, 124.0 C-7, 124.1 C-6, 126.5 C-5, 127.5 C-9, 128.1 C-8, 135.5 C-
10, 140.2 C-1, 157.2 C-3, 169.6 CO; IR (neat, cmꢁ1): 3313 (NH),
1642 (C]O); LC/MS m/z 258.3 (M
C16H19NO2: C 74.68%, H 7.44%, N 5.44%; Found: C 74.70%, H 7.50%,
N 5.39%.
þ
1); Anal. calcd for
4.1.31. N-[2-(3-Methoxy-naphthalen-1-yl)ethyl]amine
hydrochloride (37)
To a suspension of aluminum and lithium hydride (1.1 g,
30.4 mmol) at 0 ꢀC in dry ethyl ether (100 mL), was added dropwise
a solution of aluminum chloride (4.0 g, 30.4 mmol) in dry ethyl
ether (50 mL). After 10 min under stirring, a solution of compound
35 (3.0 g, 15.2 mmol) in dry ethyl ether (50 mL) was added. The
reaction mixture was stirred for additional 1 h then hydrolyzed
with a sodium hydroxide solution (4.2 g, 106.4 mmol). After
filtration, the filtrate was washed with water, dried (MgSO4) and
evaporated. The obtained oil was then treated with HCl gas and
recrystallized from acetonitrile; White solid (2.20 g, 61% yield);
m.p. 245e247 ꢀC; 1H NMR (300 MHz, DMSO-d6): dH 3.08 (m, 2H,
CH2-a), 3.31 (m, 2H, CH2-b), 3.86 (s, 3H, OCH3), 7.09 (d, J ¼ 1.5, 1H,
H-2), 7.25 (m, 1H, H-4), 7.38e7.51 (m, 2H, H-6 and 7), 7.84 (d, J ¼ 8.1,
1H, H-5), 8.03 (d, J ¼ 8.1, 1H, H-8); IR (neat, cmꢁ1): 3003-2940
(NHþ3 Clꢁ); LC/MS m/z 237.6 (M þ 1).
4.2. Pharmacology
4.2.1. Reagents and chemicals
2-[125I]Iodomelatonin (2200 Ci/mmol) was purchased from NEN
(Boston, MA). Other drugs and chemicals were purchased from
SigmaeAldrich (Saint Quentin, France).
4.2.2. Cell culture
HEK (provided by A.D. Strosberg, Paris, France) and CHO cell
lines stably expressing the human melatonin MT1 or MT2 receptors
were grown in DMEM medium supplemented with 10% foetal calf
serum, 2 mM glutamine, 100 IU/mL penicillin and 100
mg/mL
streptomycin. Grown at confluence at 37 ꢀC (95% O2/5% CO2), they
were harvested in PBS containing EDTA 2 mM and centrifuged at
1000ꢄ g for 5 min (4 ꢀC). The resulting pellet was suspended in Tris
5 mM (pH 7.5), containing EDTA 2 mM and homogenized using
a Kinematica polytron. The homogenate was then centrifuged
(95,000ꢄ g, 30 min, 4 ꢀC) and the resulting pellet suspended in
75 mM Tris (pH 7.5), 12.5 mM MgCl2 and 2 mM EDTA. Aliquots of
membrane preparations were stored at ꢁ80 ꢀC until use.
4.1.32. 3-(3-Methoxy-naphthalen-1-yl)propylamine hydrochloride (38)
To a solution of compound 36 (14.3 mmol) in methanol (100 mL)
at 0 ꢀC was added dropwise cautiously sodium borohydride
(290.7 mmol). The reaction mixture was allowed to warm to room

M. Ettaoussi et al. / European Journal of Medicinal Chemistry 49 (2012) 310e323
323
4.2.3. Binding assays
Acknowledgments
2-[125I]Iodomelatonin binding assay conditions were essentially
as previously described [33]. Briefly, binding was initiated by
addition of membrane preparations from stable transfected HEK or
CHO cells diluted in binding buffer (50 mM TriseHCl buffer, pH 7.4,
containing 5 mM MgCl2) to 2-[125I]iodomelatonin (25 or 200 pM for
MT1 and MT2 receptors, respectively, expressed in HEK cells or 20
pM for MT1 and MT2 receptors expressed in CHO cells) and the
tested drug. Non-specific binding was defined in the presence of
The 300 MHz NMR facilities were funded by the Région
Nord-Pas de Calais (France), the Ministère de la Jeunesse, de
l’Education Nationale et de la Recherche (MJENR) and the Fonds
Européens de Développement Régional (FEDER).
References
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which defined the non-specific binding. Data from the
doseeresponse curves (seven concentrations in duplicate) were
analyzed by using the program PRISM (Graph Pad Software Inc.,
San Diego, CA) to yield EC50 (effective concentration 50%) and Emax
(maximal effect) for agonists. Antagonist potencies are expressed
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gS
binding in the presence of a fixed concentration of melatonin
([Ago]) and EC50 ago is the EC50 of the molecule when tested
alone. Imax (maximal inhibitory effect) was expressed as
a percentage of that observed with melatonin at 3 nM for MT2
receptor.
Serotonin 5-HT2C binding assay was determined according to
reported tests [40]. First incubation of 200
mL solution from
membrane CHO cell lines, stably expressing the human 5-HT2C
receptors, for 60 min at 37 ꢀC in binding buffer (50 mM TriseHCl
buffer, pH 7.4, containing 10 mM MgCl2 and 0.1% BSA) containing
the radioligand [3H]-mesulergine (1 nM). Non-specific binding was
defined in the presence of 10 mM mianserine. Doseeresponse curves
are obtained by displacement of the radioligand. Reaction
was stopped by rapid filtration through GF/B filters presoaked in
0.1% (v/v) polyethylenimine. Filters were washed three times
with 1 mL of ice-cold 50 mM TriseHCl buffer, pH 7.4. Residual
radioactivity was revealed by addition of Microscint 20 and
measured by using TopCount calculator (Packard). IC50 was deter-
mined from doseeresponse curves and results are expressed as
Ki ¼ IC50/1 þ ([L]/KD), where [L] is the concentration of radioligand
used in the assay and KD, the dissociation constant of the radio-
ligand characterising the membrane preparation.
[38] M. Mor, S. Rivara, C. Silva, F. Bordi, P.V. Plazzi, G. Spadoni, G. Diamantini,
C. Balsamini, G. Tarzia, F. Fraschini, V. Lucini, R. Nonno, B.M. Stankov, J. Med.
Chem. 41 (1998) 3831e3844.
[39] P.J. Garratt, R. Jones, S.J. Rowe, D. Sugden, Bioorg. Med. Chem. Lett. 4 (1994)
1555e1558.
[40] R.S. Westphal, E. Sanders-Bush, Mol. Pharmacol. 46 (1994) 937e942.