Synthesis and biological activity of new melatonin receptor ligands

Article abstract of DOI:10.1211/146080800128735638

To discover analogues of melatonin with a longer half-life, novel non- indole analogues of the compound, in which the amide group of the side-chain has been reversed, have been prepared and evaluated in binding assays to determine their activity on melato

Full text of DOI:10.1211/146080800128735638

Pharm. Pharmacol. Commun. 2000, 6: 49±60
Received December 9, 1999
Accepted December 22, 1999
# 2000 Pharm. Pharmacol. Commun.
Synthesis and Biological Activity of New Melatonin
Receptor Ligands
I. CHARTON, A. MAMAI, C. BENNEJEAN*, P. RENARD*, P. DELAGRANGE{, P. J. MORGAN{,
H. E. HOWELL{, M. E. GOURDEL-MARTIN, M. C. VIAUD AND G. GUILLAUMET
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Institut de Chimie Organique et Analytique, Associe au CNRS, Universite d'Orleans, B.P. 6759, 45067
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Orleans Cedex 2, *ADIR, 1, rue Carle Hebert, 92415 Courbevoie Cedex, {IRI Servier, 6, Place des
Pleiades, 92415 Courbevoie Cedex, France and {Rowett Research Institute, Aberdeen, Scotland, UK
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Abstract
To discover analogues of melatonin with a longer half-life, novel non-indole analogues of
the compound, in which the amide group of the side-chain has been reversed, have been
prepared and evaluated in binding assays to determine their activity on melatonin
receptors.
The two most active compounds were those with the N-methylbutyramide side-chain.
Butyramide and pentanoylamide side-chains resulted in similar af®nities, irrespective of
the skeleton tested whereas a propionamide side-chain led to loss of af®nity. The biological
activity of the molecules was more in¯uenced by the length of the side-chain than by the
nature of the skeleton, which had little effect.
The results obtained show the relative importance of the length of the side-chain and of
the nature of the skeleton in both the binding to and the activity on the melatonin receptor
of the retroamide series.
Since the hormone melatonin was isolated and
identi®ed as N-acetyl-5-methoxytryptamine (Ler-
ner et al 1958) interest in it has been steadily
growing. In vertebrates this hormone has its pri-
mary sites of production in the pineal gland and in
the photoreceptor cells of the retina, where it is
synthesized from 5-hydroxytryptamine (5-HT) via
a two-step biochemical pathway (Weichmann
1986; Reiter 1991). Its activity, mediated through
high-af®nity G protein-coupled receptors, is prin-
cipally related to regulation of photoperiodic
responses (Hagan & Oakley 1995), entrainment of
mammalian circadian rhythms (Armstrong 1989),
induction of sleep in man (Dollins et al 1994), and
retinal physiology (Cahill & Besharse 1995).
Recently, the antitumour properties of melatonin,
its implication in the responsiveness of the immune
system (Maestroni 1993), and its free-radical scaven-
ger properties have also been described (Reiter et al
1995).
short biological half-life (15±30 min) of melatonin,
because of its rapid catabolism to 6-hydroxy-
melatonin and N-acetylkynurenamines, currently
limits its use as a therapeutic agent (Claustrat et al
1989; Di et al 1997). For this reason there is con-
siderable interest in discovering new molecules
capable of mimicking or antagonizing responses to
melatonin. Such compounds constitute important
tools in the elucidation of the physiological roles of
melatonin. Several indole analogues (Spadoni et al
1993, 1997, 1998; Garratt et al 1994, 1995; Henin
et al 1997; Tarzia et al 1997; Davies et al 1998) of
melatonin have been found to act as ligands and
many papers have reported the synthesis of several
non-indole bioisosteres (Yous et al 1992; Copinga
et al 1993; Depreux et al 1994; Sugden 1994;
Langlois et al 1995; Garratt et al 1996; Jansen et al
1996; Leclerc et al 1996; Li et al 1997; Four-
maintraux et al 1998; Kloubert et al 1998).
We recently synthesized a series of benzopyran
and benzodioxin analogues of melatonin (Viaud
et al 1998; Mamai et al 1999) and evaluated their
competitive inhibition of 2-[125I]melatonin binding
in ovine pars tuberalis membrane preparation.
In our continuing study of heterocyclic com-
pounds with potential biological activity (Savelon
Despite its potential involvement in the regula-
tion of many physiological processes, the very
Correspondence: G. Guillaumet, Institut de Chimie Organi-
Â
que et Analytique, Associe au CNRS, Universite d'Orleans,
B.P. 6759, 45067 Orleans Cedex 2, France.
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50
I. CHARTON ET AL
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et al 1998; Boye et al 1999), we present here initial
results on the design of new non-indole melatonin-
like compounds in which the amide group has a
carbon-nitrogen sequence (Leclerc et al 1998), in
contrast with that of melatonin in which the order is
opposite (nitrogen-carbon). The aim of this study
was to evaluate for three ®xed skeletonsÐ1,4-
benzodioxin, 2,3-dihydro-1,4-benzodioxin and 2,3-
dihydro-1,4-benzoxathiinÐthe importance of the
length of the retroamide side-chain in the inter-
action of the molecule when binding to, and acting
on, the melatonin receptor.
3-(2,3-Dihydro-1,4-benzodioxin-5-yl)propionic acid
(10a). Sodium hydroxide solution (10%, 3Á4 mL,
8Á46 mmol) was added to a solution of ethyl 3-(2,3-
dihydro-1,4-benzodioxin-5-yl)propionate 9a (1Á00 g,
4Á23 mmol) in ethanol (15 mL) under argon. The
reaction mixture was heated under re¯ux for 2 h,
then extracted with ethyl acetate. The aqueous layer
was acidi®ed with 3 M HCl which caused precipita-
tion of acid 10a. The white solid was collected by
®ltration and dried under high vacuum over P2O5 to
give a white solid (91%), mp 73ꢀC. 1H NMR
(300 MHz, DMSO-d6) d 2Á44 (t, 2H, J ˆ 7Á8,
CH2), 2Á73 (t, 2H, J ˆ 7Á8, CH2), 4Á19±4Á24 (m,
4H, H2 and H3), 6Á65±6Á73 (m, 3H, H6, H7 and H8),
12Á1 (br s,1H, CO2H). IR (KBr) n 3300±2700,
1712, 1282, 1101 cmÀ1. Calculated for C11H12O4:
C, 63Á44; H, 5Á82; found: C, 63Á29; H, 5Á80.
Compounds 1±8 (Figure 1) were synthesized and
evaluated in binding assays; the results obtained
were compared with those from N-[3-(2,3-dihydro-
1,4-benzodioxin-5-yl)propyl]acetamide (A; Guil-
laumet et al 1998; Charton et al 1999).
5-(2,3-Dihydro-1,4-benzodioxin-5-yl)pentanoic acid
(10c). This compound was prepared as a white
solid, mp 56ꢀC, in 79% yield, from ethyl 5-
(2,3-dihydro-1,4-benzodioxin-5-yl)pentanoate 9c
Materials and Methods
1
according to the procedure described for 10a. H
Chemistry
Melting points were determined on a Ko¯er hot-
NMR (300 MHz, DMSO-d6) d 1Á50±1Á55 (m, 4H,
2 6 CH2), 2Á23±2Á29 (m, 2H, CH2), 2Á62±2Á70 (m,
2H, CH2), 4Á20±4Á25 (m, 4H, H2 and H3), 6Á67±
6Á72 (m, 3H, H6, H7 and H8), 12Á00 (br s,1H,
CO2H), IR (KBr) n 3300±2700, 1705, 1278,
1100 cmÀ1. Calculated for C13H16O4: C, 66Á08; H,
6Á84; found: C, 65Á82; H, 6Á78.
È
stage apparatus and are uncorrected. Proton and
carbon NMR were recorded on a Bruker AM 300
W. B. or DPX 250 spectrometer. Coupling con-
stants are in Hz and chemical shifts are d ppm
down®eld from tetramethylsilane, which was used
as an internal standard. IR spectra were obtained by
use of a Perkin-Elmer 297 spectrophotometer. Mass
spectra were recorded on a Nermag R 10-10 C
(70 eV) apparatus. Organic solvents were puri®ed
as necessary as described by Perrin et al (1986) or
purchased from Aldrich. Analytical thin-layer
chromatography was performed on precoated plates
(silica gel, 60F254) and spots were visualized with
UV light or with a solution of ammonium cerium
nitrate in ethanol. Column chromatography was
performed on silica gel 60 (Merck) j 70±400 mesh
for gravity columns and 230±400 mesh for ¯ash
columns. The solvents used for column chromato-
graphy were distilled before use; solvent mixtures
are reported as v=v ratios.
N-Methyl-[3-(2,3-dihydro-1,4-benzodioxin-5-yl)]-
propanamide (1). Hydroxybenzotriazole (379 mg,
2Á81 mmol) and 1-(3-dimethylaminopropyl)-3-ethyl-
carbodiimide hydrochloride (538 mg, 2Á81 mmol)
were added at 0ꢀC to a solution of acid 10a
(500 mg, 2Á55 mmol) in anhydrous N, N-dimethyl-
formamide (10 mL) under argon. The argon ¯ow
was then stopped and methylamine (10% in ben-
zene, 195 mL) was introduced. The temperature
was left to increase slowly to 20ꢀC, and stirring
was continued for 18 h. The reaction mixture was
concentrated in-vacuo, and diluted with dichloro-
methane. The organic layer was washed with
Figure 1. Structures of the retroamides with 1,4-benzodioxin, 2,3-dihydro-1,4-benzodioxin, and 2,3-dihydro-1,4-benzooxathiin
skeletons. A ˆ N-[3-(2,3-dihydro-1,4-benzodioxin-5-yl)propyl]acetamide (Guillaumet et al 1998; Charton et al 1999).

SYNTHESIS AND ACTIVITY OF NEW MELATONIN RECEPTOR LIGANDS
51
water. The combined organic extracts were dried
(MgSO4) and concentrated in-vacuo. Puri®cation
by column chromatography (chloroform±ethyl
acetate, 7 : 3) gave the retroamide 1 as a solid in
78% yield, mp 99ꢀC. 1H NMR (300 MHz,
CDCl3=D2O) d 2Á35 (t, 2H, J ˆ 8Á0, CH2), 2Á66
(s, 3H, CH3) 2Á81 (t, 2H, J ˆ 8Á0, CH2), 4Á13±4Á18
(m, 4H, H2 and H3), 6Á59±6Á65 (m, 3H, H6, H7 and
H8). IR (KBr) n 3326, 1646, 1274, 1192,
1102 cmÀ1. MS(CI =NH3) m=z 222 (M ‡ 1). Cal-
culated for C12H15NO3: C, 65Á13; H, 6Á85; N, 6Á33;
found: C, 65Á04; H, 6Á81; N, 6Á28.
product was hydrolysed and extracted with dichlor-
omethane. The combined organic extracts were
dried (MgSO4) and concentrated in-vacuo. Puri®-
cation by column chromatography (ethyl acetate±
petroleum ether, 3 : 7) gave the tosylate 14 (92%)
1
as a colourless oil. H NMR (300 MHz, CDCl3) d
1Á86 (q, 2H, J ˆ 7Á0, CH2), 2Á38 (s, 3H, CH3), 2Á54
(t, 2H, J ˆ 7Á0, CH2), 3Á98 (t, 2H, J ˆ 7Á0, CH2O),
4Á14±4Á17 (m, 4H, H2 and H3), 6Á47 (dd,1H,
J ˆ 7Á4, 2Á9, Harom), 6Á59±6Á66 (m, 2H, Harom),
7Á27 (m, 2H, Harom), 7Á72 (m, 2H, Harom). IR
(neat) n 1292, 1184, 1102 cmÀ1. Calculated for
C18H20O5S : C, 62Á04; H, 5Á80; S, 9Á20; found: C,
61Á82; H, 5Á85; S, 9Á12.
N-Methyl-[5-(2,3-dihydro-1,4-benzodioxin-5-yl)]-
pentanamide (3). This compound (mp 79ꢀC) was
prepared in 75% yield from acid 10c according to
the procedure described for 1. 1H NMR (250 MHz,
CDCl3=D2O) d 1Á56±1Á72 (m, 4H, CH2), 2Á19 (t,
2H, J ˆ 6Á7, CH2), 2Á58 (t, 2H, J ˆ 6Á7, CH2), 2Á78
(s, 3H, CH3), 4Á21±4Á27 (m, 4H, H2 and H3), 6Á61±
6Á78 (m, 3H, H6, H7 and H8). 13C NMR (62Á9 MHz,
CDCl3) d 25Á6 and 26Á3 (CH2 and CH3), 29Á3 and
29Á5 (CH2), 36Á7 (CH2), 64Á3 (C2 and C3), 115Á1
(C7), 120Á6 and 121Á9 (C6 and C8), 131Á10 (C5),
141Á5 (C9 and C10), 173Á7 (C ˆ O). IR (KBr) n
3290, 1636, 1288, 1079 cmÀ1. MS(CI =NH3) m=z
250 (M ‡ 1). Calculated for C14H19NO3: C, 67Á45;
H, 7Á68; N, 5Á62; found: C, 67Á50; H, 7Á70; N, 5Á41.
4-(2,3-Dihydro-1,4-benzodioxin-5-yl)butyronitrile
(16). Potassium cyanide (448 mg, 6Á89 mmol) was
added to a solution of tosylate 14 (2Á00 g,
5Á74 mmol) in N, N-dimethylformamide. The reac-
tion mixture was heated at 100ꢀC for 5Á5 h, and then
hydrolysed. The solvent was evaporated and the
aqueous layer extracted with dichloromethane. The
combined organic extracts were dried (MgSO4) and
concentrated in-vacuo. Puri®cation by column
chromatography (ethyl acetate-petroleum ether,
3 : 7) gave the cyano compound 16 (77%) as a
1
yellow oil. H NMR (300 MHz, CDCl3) d 1Á95 (q,
2H, J ˆ 7Á4, CH2), 2Á30 (t, 2H, J ˆ 7Á4, CH2), 2Á71
(t, 2H, J ˆ 7Á4, CH2), 4Á20±4Á26 (m, 4H, H2 and
H3), 6Á65±6Á75 (m, 3H, H6, H7 and H8). IR (neat) n
2250, 1292, 1092 cmÀ1. Calculated for C12H13NO2:
C, 70Á90; H, 6Á46; N, 6Á89; found: C, 70Á75; H, 6Á50;
N, 6Á81.
3-(2,3-Dihydro-1,4-benzodioxin-5-yl)propanol (12).
Lithium aluminium hydride (401 mg, 10Á6 mmol)
was added in portions to a solution of ethyl 3-(2,3-
dihydro-1,4-benzodioxin-5-yl)propionate 9a (2Á5 g,
10Á6 mmol) in anhydrous ether (20 mL) at 0ꢀC
under argon. The reaction mixture was stirred for
1 h at room temperature. The reaction was
quenched by adding water (320 mL), 15% NaOH
(320 mL) and then water (960 mL). After stirring for
1 h, the salts were removed by ®ltration and washed
with ether. The organic layer was dried (MgSO4)
and concentrated in-vacuo. The crude alcohol 12
was obtained as a colourless oil (97%), and used in
4-(2,3-Dihydro-1,4-benzodioxin-5-yl)butanoic acid
(10b). Sodium hydroxide solution (10%, 7Á9 mL,
19Á65 mmol) was added to a solution of cyano
compound 16 (800 mg, 3Á93 mmol) in ethanol
(10 mL). The reaction mixture was heated at 60ꢀC
for 16 h, and the ethanol was then evaporated.
Acidi®cation with 2 M HCl led to precipitation of
the acid as a white solid which was collected by
®ltration, and dried under high vacuum over P2O5
(74%, mp 54ꢀC). 1H NMR (300 MHz, DMSO-d6) d
2Á00 (q, 2H, J ˆ 7Á8, CH2), 2Á46 (t, 2H, J ˆ 7Á8,
CH2), 2Á78 (t, 2H, J ˆ 7Á8, CH2), 4Á45±4Á58 (m, 4H,
H2 and H3), 6Á90±7Á01 (m, 3H, H6, H7 and H8),
12Á10 (br s, 1H, CO2H). IR (KBr) n 3000±2500,
1730, 1300, 1110 cmÀ1. Calculated for C12H14O4:
C, 64Á84; H, 6Á36; found: C, 64Á77; H, 6Á34.
1
the next step without further puri®cation. H NMR
(300 MHz, CDCl3) d 1Á70 (br s,1H, OH), 1Á82 (q,
2H, J ˆ 7Á3, CH2), 2Á66 (t, 2H, J ˆ 7Á3, CH2), 3Á60
(m, 2H, CH2OH), 4Á21±4Á27 (m, 4H, H2 and H3),
6Á67±6Á77 (m, 3H, H6, H7 and H8). IR (neat) n
3690±3110, 1282, 1068 cmÀ1.
[3-(2,3-Dihydro-1,4-benzodioxin-5-yl)propyl]-(4-
methylphenyl)sulphonate (14). Tosyl chloride
(1Á57 g, 8Á24 mmol) was added to a solution of
alcohol 12 (800 mg, 4Á12 mmol) and triethylamine
(1Á15 mL, 8Á24 mmol) in dichloromethane (20 mL).
After stirring at room temperature for 12 h, the
N-Methyl-[4- (2,3- dihydro-1,4-benzodioxin-5-yl)]-
butanamide (2a). This compound (mp 79ꢀC) was
prepared in 77% yield from acid 10b (600 mg,
2Á70 mmol) according to the procedure described
1
for 1. H NMR (300 MHz, CDCl3=D2O) d 1Á86 (q,

52
I. CHARTON ET AL
2H, J ˆ 7Á4, CH2), 2Á13 (t, 2H, J ˆ 7Á4, CH2), 2Á55
(t, 2H, J ˆ 7Á4, CH2), 2Á73 (s, 3H, CH3), 4Á16±4Á20
(m, 4H, H2 and H3), 6Á60±6Á69 (m, 3H, H6, H7 and
H8). 13C NMR (62Á9 MHz, CDCl3) d 24Á4 and 25Á0
(CH2 and CH3), 27Á8 and 34Á8 (CH2), 62Á9 and 63Á0
(C2 and C3), 114Á0 (C7), 119Á3 and 120Á8 (C6 and
C8), 129Á0 (C5), 140Á2 and 142Á1 (C9 and C10),
172Á2 (C ˆ O). IR (KBr) n 3300, 1646, 1284, 1208,
1118 cmÀ1. MS(CI =NH3) m=z 236 (M ‡ 1). Cal-
culated for C13H17NO3: C, 66Á36; H, 7Á28; N, 5Á95;
found: C, 66Á31; H, 7Á21; N, 5Á95.
(dd,1H, J ˆ 7Á5, 2Á2, Harom), 6Á84 (dd,1H, J ˆ 7Á5,
2Á2, Harom), 6Á70 (m,1H, Harom). IR (neat) n 3640±
3130, 1280 cmÀ1. Calculated for C11H12O3: C,
68Á72; H, 6Á30; found: C, 68Á56; H, 6Á25.
[3-(1,4-Benzodioxin-5-yl)propyl](4-methylphenyl)-
sulphonate (15). This compound was prepared from
alcohol 13 in 90% yield according to the procedure
described for 14 (24 h). 1H NMR (300 MHz,
CDCl3) d 1Á86 (m, 2H, CH2), 2Á44 (s, 3H, CH3),
2Á47 (t, 2H, J ˆ 7Á2, CH2), 4Á01 (t, 2H, J ˆ 6Á2,
CH2), 5Á83 (d,1H, J ˆ 3Á7, ˆ CH), 5Á85 (d,1H,
J ˆ 3Á7, ˆ CH), 6Á44 (dd,1H, J ˆ 8Á0, 1Á7, Harom),
6Á51 (dd,1H, J ˆ 8Á0, 1Á7, Harom), 6Á65 (m,1H,
N-Propyl-4-(2,3-dihydro-1,4-benzodioxin-5-yl)but-
anamide (2b). A solution of acid 10b (260 mg,
1Á17 mmol) in N, N-dimethylformamide (1 mL)
was added to a solution of n-propylamine hydro-
chloride (134 mg, 1Á75 mmol) in N, N-dimethyl-
formamide (5 mL) at 0ꢀC. 1-(3-Dimethylaminopro-
pyl)-3-ethylcarbodiimide hydrochloride (EDCI;
247 mg, 1Á29 mmol) was then added. Stirring was
continued for 10 h, and the reaction mixture left to
warm to room temperature. After evaporation of N,
N-dimethylformamide, the residue was hydrolysed
and then extracted with ethyl acetate. The com-
bined organic extracts were dried (MgSO4), and
concentrated in-vacuo. Puri®cation by column
chromatography (ethyl acetate±petroleum ether,
2 : 8) gave the retroamide 2b (65%) as a white
solid, mp 57±58ꢀC. 1H NMR (250 MHz, CDCl3) d
0Á92 (t, 3H, J ˆ 7Á3, CH3), 1Á51 (sext, 2H, J ˆ 7Á3,
CH2), 1Á92 (q, 2H, J ˆ 7Á3, CH2), 2Á20 (t, 2H,
H ˆ 7Á3, CH2), 2Á62 (t, 2H, J ˆ 7Á3, CH2), 3Á17±
3Á25 (m, 2H, CH2N), 4Á22±4Á27 (m, 4H, H2 and
H3), 5Á47 (br s, 1H, NH), 6Á67±6Á76 (m, 3H, H6, H7
and H8). 13C NMR (62Á9 MHz, CDCl3) d 11Á3
(CH3), 22Á9, 25Á8, 29Á1 and 36Á3 (CH2), 41Á1
(CH2N), 64Á16 and 64Á22 (C2 and C3), 115Á2 (C7),
122Á1 and 120Á6 (C6 and C8), 130Á2 (C5), 141Á5 and
143Á4 (C9 and C10), 172Á8 (C ˆ O). IR (KBr) n
3300, 1641. Calculated for C15H21NO3: C, 68Á40;
H, 8Á05; N, 5Á32; found: C,68Á59; H, 8Á14; N, 5Á38.
H
arom), 7Á34 (d, 2H, J ˆ 8Á5, Harom), 7Á78 (d, 2H,
J ˆ 8Á5, Harom). IR (neat) n 1360, 1250, 1185 cmÀ1.
Calculated for C18H18O5: C, 62Á40; H, 5Á25; S,
9Á25; found: C, 62Á29; H, 5Á26; S, 9Á12.
4-(1,4-Benzodioxin-5-yl)butyronitrile (17). Potas-
sium cyanide (500 mg, 7Á70 mmol) was added to a
solution of tosylate 15 (1Á07 g, 3Á07 mmol) in N, N-
dimethylformamide (15 mL) under argon and the
reaction mixture was heated under re¯ux for 4 h.
The solvent was then evaporated and the residue
diluted with water. The aqueous layer was
extracted with dichloromethane. The combined
organic extracts were dried (MgSO4) and concen-
trated in-vacuo. Puri®cation by column chromato-
graphy (ethyl acetate±petroleum ether, 1 : 9) gave
1
the cyano compound 17 (85%) as an oil. H NMR
(300 MHz, CDCl3) d 1Á89 (m, 2H, CH2), 2Á32 (t,
2H, J ˆ 7Á0, CH2), 2Á60 (t, 2H, J ˆ 7Á5, CH2), 5Á86
(d,1H, J ˆ 3Á5, ˆ CH), 5Á90 (d,1H, J ˆ 3Á5, ˆ CH),
6Á49 (dd,1H, J ˆ 7Á7, 2Á0, Harom), 6Á66 (dd,1H,
J ˆ 7Á7, 2Á0, Harom), 6Á74 (m,1H, Harom). IR (neat)
n 2250, 1290 cmÀ1. Calculated for C12H11O2N: C,
71Á62; H, 5Á52; N, 6Á96; found: C, 71Á49; H, 5Á50;
N, 6Á95.
4-(1,4-Benzodioxin-5-yl)butanoic acid (18). A solu-
tion of cyano compound 17 (580 mg, 2Á90 mmol) in
10% sodium hydroxide solution (20 mL) was
heated under re¯ux for 20 h. The solvent was
removed and the residue was dissolved in 1 M
HCl (20 mL). The aqueous layer was extracted
with ethyl acetate and the combined organic
extracts were dried (MgSO4) and concentrated in-
vacuo. The acid 18 was obtained quantitatively as a
3-(1,4-Benzodioxin-5-yl)propanol (13). Lithium
aluminium hydride (233 mg, 6Á14 mmol) was
added to a solution of ethyl 3-(1,4-benzodioxin-5-
yl)propanoate 11 (900 mg, 4Á09 mmol) in dry ether
(30 mL). The reaction mixture was heated under
re¯ux for 30 min. After cooling and hydrolysis, the
resulting salts were removed by ®ltration. The
®ltrate was dried (MgSO4), and concentrated in-
vacuo. Puri®cation by column chromatography
(ethyl acetate±petroleum ether, 4 : 6) gave alcohol
1
white solid, mp 75±76ꢀC. H NMR (300 MHz,
CDCl3) d 1Á88 (q, 2H, J ˆ 7Á5, CH2), 2Á37 (t, 2H,
J ˆ 7Á5, CH2), 2Á51 (t, 2H, J ˆ 7Á5, CH2), 5Á86
(d,1H, J ˆ 3Á5, ˆ CH), 5Á89 (d,1H, J ˆ 3Á5,
ˆ CH), 6Á47 (dd,1H, J ˆ 7Á5, 1Á8, Harom), 6Á66
(dd,1H, J ˆ 7Á5, 1Á8, Harom), 6Á73 (m,1H, Harom),
10Á93 (br s,1H, CO2H). IR (KBr) n 3560±2560,
1
12 (95%) as an oil. H NMR (300 MHz, CDCl3) d
1Á51 (m,1H, OH), 1Á76 (m, 2H, CH2), 2Á50 (t, 2H,
J ˆ 7Á2, CH2), 3Á60 (m, 2H, CH2), 5Á83 (d,1H,
J ˆ 3Á5, ˆ CH), 5Á87 (d,1H, J ˆ 3Á5, ˆ CH), 6Á43

SYNTHESIS AND ACTIVITY OF NEW MELATONIN RECEPTOR LIGANDS
53
1705, 1290 cmÀ1. Calculated for C12H12O4: C,
65Á44; H, 5Á50; found: C, 65Á29; H, 5Á51.
added to a solution of acetate 20 (1Á20 g,
4Á75 mmol) in methanol (48 mL) and water
(12 mL). The reaction mixture was stirred for
2Á5 h at room temperature. The salts were removed
by ®ltration and the methanol evaporated. The
residual aqueous layer was extracted with ethyl
acetate. The combined organic extracts were dried
(MgSO4), and concentrated in-vacuo. Filtration
through silica gel (CH2Cl2) gave the alcohol 21
(94%) as a yellow oil. 1H NMR (300 MHz,
CDCl3=D2O) d 1Á69 (t, 2H, J ˆ 7Á0, CH2), 2Á67
(t,1H, J ˆ 7Á0, CH2), 3Á13 (t, 2H, J ˆ 4Á4, H3), 3Á67
(t, 2H, J ˆ 7Á0, CH2), 4Á37 (t, 2H, J ˆ 4Á4, H2), 6Á69±
6Á77 (m, 2H, H6 and H8), 6Á93 (m,1H, H7). IR (neat) n
3700±3100, 1300, 1242, 1076 cmÀ1. Calculated for
C11H14O2S : C, 62Á82; H, 6Á72; S, 15Á24; found: C,
62Á65; H, 6Á70; S, 15Á10.
N-Propyl-4-(1,4-benzodioxin-5-yl)butyramide (4b).
4-Dimethylaminopyridine (330 mg, 3Á54 mmol), and
then acid 18 (410 mg, 2Á36 mmol) were added at 0ꢀC
to a suspension of n-propylamine hydrochloride
(290 mg, 2Á60 mmol) in dry N, N-dimethylformamide
(7 mL). EDCI (500 mg, 2Á60 mmol) was then intro-
duced. The reaction mixture was stirred at room
temperature for 12 h under argon. After evaporation
of the solvent, the residue was diluted with water and
ethyl acetate (1 : 1). The aqueous layer was extracted
with ethyl acetate and the combined organic extracts
were dried (MgSO4) and concentrated in-vacuo.
Chromatography (ethyl acetate±petroleum ether,
1
7 : 3) gave the retroamide 4b in 88% yield. H
NMR (300 MHz, CDCl3) d 0Á90 (t, 3H, J ˆ 7Á3,
CH3), 1Á49 (m, 2H, CH2), 1Á89 (m, 2H, CH2), 2Á15
(t, 2H, J ˆ 7Á5, CH2), 2Á47 (t, 2H, J ˆ 7Á5, CH2), 3Á20
(t, 2H, J ˆ 7Á2, CH2), 5Á57 (br s,1H, NH), 5Á84 (d,1H,
J ˆ 3Á7, ˆ CH), 5Á87 (d,1H, J ˆ 3Á7, ˆ CH), 6Á44
(dd,1H, J ˆ 7Á5, 2Á0, Harom), 6Á64 (dd,1H, J ˆ 7Á5,
2Á0, Harom), 6Á72 (m,1H, Harom). IR (neat) n 3410±
3170, 1645, 1300 cmÀ1. MS(CI =NH3) m=z: 262
(M ‡ 1). Calculated for C15H19NO3: C, 68Á93; H,
7Á34; N, 5Á36; found: C, 68Á81; H, 7Á34; N, 5Á35.
3-(2,3-Dihydro-1,4-benzoxathiin-5-yl)propanal (22).
A
solution of dimethylsulphoxide (1Á15 mL,
15Á67 mmol) in dry dichloromethane (3Á7 mL) was
added to a solution of oxalyl chloride (680 mL,
7Á85 mmol) in dry dichloromethane (18Á8 mL) at
À50ꢀC under argon. The reaction mixture was
stirred for 2 min, and a solution of alcohol 21
(1Á50 g, 7Á13 mmol) in dichloromethane (7Á5 mL)
was then added via a cannula at À50ꢀC. After
stirring for 30 min, triethylamine (5Á0 mL,
35Á7 mmol) was introduced. The reaction mixture
was left to warm to room temperature, and stirring
was continued for 2 h. The reaction mixture was
diluted with dichloromethane. The organic layer
was washed with brine, dried (MgSO4) and con-
centrated in-vacuo. Puri®cation by column chroma-
tography (ethyl acetate±petroleum ether, 3 : 7) gave
aldehyde 22 (84%) as a yellow oil. 1H NMR
(300 MHz, CDCl3) d 2Á75 (t, 2H, J ˆ 7Á8, CH2),
2Á85 (t, 2H, J ˆ 7Á8, CH2), 3Á13 (t, 2H, J ˆ 4Á4, H3),
4Á38 (t, 2H, J ˆ 4Á4, H2), 6Á70±6Á78 (m, 2H, H6 and
H8), 6Á95 (m,1H, H7), 9Á85 (s,1H, CHO). IR (neat) n
2870, 1738, 1310, 1247, 1080 cmÀ1. Calculated for
C11H12O2S : C, 63Á43; H, 5Á82; S, 15Á39; found: C,
63Á35; H, 5Á81; S, 15Á24.
3-(2,3-Dihydro-1,4-benzoxathiin-5-yl)propyl acetate
(20). Mercuric acetate (1Á17 g, 3Á66 mmol) was
added to a solution of 5-(3-bromopropyl)-2,3-dihy-
dro-1,4-benzoxathiin 19 (1Á00 g, 3Á66 mmol) in gla-
cial acetic acid (20 mL). The reaction mixture was
heated under re¯ux for 4 h, and concentrated in-
vacuo. Ethyl acetate was added and the resulting
precipitate was removed by ®ltration. The ®ltrate
was washed with a saturated bicarbonate aqueous
solution. The combined organic extracts were
dried, and concentrated in-vacuo. Puri®cation by
column chromatography (ethyl acetate±petroleum
ether,
dichloromethane±cyclohexane (1 : 1) to remove
1 : 3),
and
then
trituration
in
solid impurity gave the acetate 20 (83%) as a
1
yellow oil. H NMR (300 MHz, CDCl3) d 1Á96 (t,
2H, J ˆ 7Á1, CH2), 2Á65 (t, 2H, J ˆ 7Á1, CH2), 3Á13
(t, 2H, J ˆ 4Á7, H3), 4Á11 (t, 2H, J ˆ 7Á1, CH2), 4Á38
(t, 2H, J ˆ 4Á7, H2), 6Á69±6Á76 (m, 2H, H6 and H8),
6Á93 (m,1H, H7). 13C NMR (62Á9 MHz, CDCl3), d
19Á2 (CH2), 23Á7 (C3), 26Á2 and 27Á6 (CH2), 62Á1
and 62Á9 (C2 and CH2) 114Á5 (Ar), 115Á2 (C10),
120Á1 (Ar), 123Á0 (C7), 136Á7 (C5), 150Á1 (C9),
169Á4 (C ˆ O). IR (neat) n 1738, 1305, 1247,
1044, 1087 cmÀ1.
3-(2,3-Dihydro-1,4-benzoxathiin-5-yl)propanoic
acid (23). A solution of 3-(2,3-dihydro-1,4-benzox-
athiin-5-yl)propanal 22 (190 mg, 0Á91 mmol) in
ethanol (4 mL) was added to a solution of silver
nitrate (310 mg, 1Á82 mmol) in distilled water
(3Á6 mL). The reaction mixture was stirred at
room temperature, and a solution of sodium hydro-
xide (146 mg, 3Á63 mmol) in distilled water
(7Á3 mL) was then added. The reaction mixture
was stirred for 30 min and ®ltered. The solid
residue was washed with ether and water. After
separation, the aqueous layer was acidi®ed. The
3-(2,3-Dihydro-1,4-benzoxathiin-5-yl)propanol (21).
Potassium carbonate (986 mg, 7Á13 mmol) was

54
I. CHARTON ET AL
resulting precipitate was ®ltered, washed with
water, and dried under high vacuum over P2O5.
Acid 23 was obtained as a white solid in 64% yield.
1H NMR (250 MHz, DMSO-d6) d 2Á40±2Á50 (m,
2H, CH2), 2Á70 (t, 2H, J ˆ 7Á2, CH2), 3Á15 (t, 2H,
J ˆ 4Á9, H3), 4Á28 (t, 2H, J ˆ 4Á9, H2), 6Á65 (m,1H,
(br s,1H, CO2H). 13C NMR (62Á9 MHz, DMSO-d6)
d 23Á8 and 26Á8 (CH2), 31Á9 (CH2), 63Á5 (C2), 115Á2
(C10), 116Á0 (Ar), 120Á4 (Ar), 123Á7 (C7), 136Á8
(C5), 150Á7 (C9), 172Á7 (C ˆ O).
according to the procedure described for 10b (heat
under re¯ux for 24 h). 1H NMR (DMSO- d6) d 1Á74 (t,
2H, J ˆ 7Á5, CH2), 2Á20 (t, 2H, J ˆ 7Á5, CH2), 2Á48 (t,
2H,J ˆ 7Á5,CH2),3Á13(t, 2H,J ˆ 4Á8,H3),4Á27(t,2H,
J ˆ 4Á8, H2), 6Á63 (d,1H, J ˆ 7Á1, Harom), 6Á71 (d,1H,
7Á1, Harom), 6Á89 (m,1H, H7), 12Á00 (br s,1H, CO2H).
13C NMR (300 MHz, CDCl3) d 23Á6 and 24Á3 (C3 and
CH2), 31Á3 and 32Á8 (CH2), 64Á0 (C2), 115Á5 (CH),
116Á4 (C10), 121Á2 (CH), 124Á1 (C7), 138Á2 (C5), 151Á2
(C9), 173Á7 (C ˆ O). IR (KBr) n 3300±2790, 1712,
1040 cmÀ1. Calculated for C12H14SO3: C, 60Á47; H,
5Á93; S, 13Á45; found: C, 60Á28; H, 5Á91; S, 13Á39.
H
arom), 6Á75 (m,1H, Harom), 6Á90 (m,1H, H7), 12Á16
N-Methyl-3-(2,3-dihydro-1,4-benzoxathiin-5-yl)-
propanamide (5). This compound was prepared in
95% yield from acid 23 according to the procedure
described for 4b. Puri®cation by column chromato-
graphy (chloroform±ethyl acetate, 7 : 3) gave the
N-Methyl-4-(2,3-dihydro-1,4-benzoxathiin-5-yl)but-
anamide(6a).Thiscompoundwaspreparedasawhite
solid, mp 94ꢀC, in 74% yield, from acid 25 and
methylamine hydrochloride according to the proce-
1
1
retroamide 5 as a white solid, mp 98±99ꢀC. H
dure described for the retroamide 4b. H NMR
NMR (250 MHz, CDCl3) d 2Á47 (m, 2H, CH2), 2Á79
(d, 3H, J ˆ 4Á9, CH3), 2Á93 (m, 2H, CH2), 3Á14 (m,
2H, H3), 4Á39 (m, 2H, H2), 5Á36 (br s,1H, NH), 6Á71
(dd,1H, J ˆ 8Á1, 1Á2, Harom), 6Á78 (br d,1H, J ˆ 7Á6,
(300 MHz, CDCl3=D2O) d 1Á51±1Á70 (m, 2H, CH2),
1Á96 (s, 3H, CH3), 2Á61 (t, 2H, J ˆ 7Á5, CH2), 2Á81 (t,
2H,J ˆ 7Á5,CH2),3Á13(t, 2H,J ˆ 4Á7,H3),4Á38(t,2H,
J ˆ 4Á7, H2), 6Á69±6Á76 (m, 2H, H6 and H8), 6Á93
(m,1H, H7). 13C NMR (62Á9 MHz, CDCl3) d 25Á5
(CH2), 26Á0 (CH2), 26Á7 (CH3), 32Á9 (CH2), 36Á4
(CH2), 65Á2 (C2), 116Á7, 122Á5 and 125Á1 (C6, C7,
C8), 117Á5 (C10), 139Á3 (C5), 152Á3 (C9), 173Á6
(C ˆ O). IR (KBr) n 3590±3270, 1722, 1274, 1116,
1075 cmÀ1. MS(CI, NH 3), m=z 252 (M ‡ 1). Calcu-
lated for C13H17NO2S : C, 62Á11; H, 6Á83; N, 5Á57; S,
12Á75; found: C, 61Á93; H, 6Á80; N, 5Á55; S, 12Á66.
H
arom), 6Á93 (m,1H, H7). Calculated for
C12H15NO2S : C, 6Á073; H, 6Á37; N, 5Á90; S,
13Á51; found: C, 60Á46; H, 6Á35; N, 5Á75; S, 13Á53.
4-(2,3-Dihydro-1,4-benzoxathiin-5-yl)butyronitrile
(24). Potassium cyanide (52 mg, 0Á80 mmol) was
added to a solution of 5-(3-bromopropyl)-2,3-dihy-
dro-1,4-benzoxathiin 19 (200 mg, 0Á73 mmol) in N,
N-dimethylformamide (5 mL) under argon at room
temperature. After stirring for 12 h, further potas-
sium cyanide (52 mg, 0Á80 mmol) was added, and
stirring was continued for 12 h. The reaction mix-
ture was the hydrolysed, the solvent evaporated,
and the aqueous layer extracted with dichloro-
methane. The combined organic extracts were
dried (MgSO4), and concentrated in-vacuo. Puri®-
cation by column chromatography (ethyl acetate±
petroleum ether, 3 : 7) gave the cyano compound
N-Propyl-4-(2,3-dihydro-1,4-benzoxathiin-5-yl)-bu-
tanamide (6b). This compound was prepared as a
white solid, mp 93ꢀC, in 96% yield, from acid 25 and
propylamine hydrochloride, according to the proce-
1
dure described for the retroamide 4b. H NMR
(300 MHz, CDCl3=D2O) d 0Á93 (t, 2H, J ˆ 7Á6,
CH3), 1Á54 (q, 2H, J ˆ 7Á6, CH2), 1Á98 (q, 2H, 7Á6,
CH2), 2Á23 (t, 2H, J ˆ 7Á6, CH2), 2Á62 (t, 2H, J ˆ 7Á6,
CH2), 3Á14 (t, 2H, J ˆ 4Á2, H3), 3Á22 (t, 2H, J ˆ 7Á6,
CH2), 4Á39 (t, 2H, J ˆ 4Á2, H2), 6Á69±6Á77 (m, 2H, H6
and H8), 6Á94 (m,1H, H7). IR (KBr) n 3590±3260,
1668, 1274, 1120, 1075 cmÀ1. MS(EI) m =z 279 (M).
Calculated for C15H21NO2S : C, 64Á48; H, 7Á58; N,
5Á01; S, 11Á48; found: C, 64Á79; H, 7Á59; N, 5Á08; S,
11Á39.
1
24 (100%) as a colourless oil. H NMR (250 MHz,
CDCl3) d 1Á99 (q, 2H, J ˆ 7Á2, CH2), 2Á36 (t, 2H,
J ˆ 7Á2, CH2), 2Á74 (t, 2H, J ˆ 7Á2, CH2) 3Á13 (t, 2H,
J ˆ 4Á6, H3), 4Á37 (t, 2H, H ˆ 4Á6, H2), 6Á71±6Á76
(m, 2H, H6 and H8), 6Á95 (m,1H, J ˆ 7Á9, H7). 13C
NMR (62Á9 MHz, CDCl3) d 16Á0 (CH2), 24Á1
(CH2), 25Á0 (C3), 31Á3 (CH2), 64Á0 (C2), 116Á2 (C6
or C8), 119Á0 (C10 or CN), 121Á7 (C6 or C8), 124Á3
(C7), 137Á5 (C5), 152Á1 (C9). IR (neat) n 2234, 1302,
1252, 1084, 1054. Calculated for C12H13NOS: C,
65Á71; H, 5Á99; N, 6Á39; S, 14Á62; found: C, 65Á56;
H, 5Á98; N, 6Á31; S, 14Á53.
Ethyl 5-(2,3-dihydro-1,4-benzoxathiin-5-yl)pent-2-
enoate (26). (Carbethoxymethylene)triphenylphos-
phorane (1Á83 g, 5Á28 mmol) was added to a solu-
tion of aldehyde 22 (500 mg, 2Á40 mmol) in dry
toluene. The reaction mixture was heated under
re¯ux for 4 h. Triphenylphosphine oxide was pre-
cipitated by adding hexane to the ice-cooled reac-
tion mixture, and removed by ®ltration. The ®ltrate
was concentrated in-vacuo. Puri®cation of the resi-
4-(2,3-Dihydro-1,4-benzoxathiin-5-yl)butanoic acid
(25).Thiscompoundwaspreparedasawhitesolid,mp
93ꢀC, in 71% yield, from the cyano compound 24

SYNTHESIS AND ACTIVITY OF NEW MELATONIN RECEPTOR LIGANDS
55
due by column chromatography (ethyl acetate±
petroleum ether, 1 : 9) gave ester 26 (92%) as a
colourless oil. H NMR (250 MHz, CDCl3) d 1Á28
white solid, mp 66ꢀC, in 93% yield, from acid 28
according to the procedure described for retroamide
4b. 1H NMR (250 MHz, CDCl3) d 1Á60±1Á83 (m, 4H,
CH2), 2Á23 (t, 2H, J ˆ 7Á8, CH2), 2Á60 (t, 2H, J ˆ 7Á8,
CH2), 2Á81(s,3H,CH3), 3Á15(t,2H,J ˆ 4Á6,H3), 4Á39(t,
2H, J ˆ 4Á6, H2), 6Á63±6Á80 (m, 2H, H6 and H8), 6Á95
(m,1H, H7). 13C NMR (62Á9 MHz, CDCl3) d 24Á9 and
25Á6(C3 and CH2), 25Á6(CH3), 28Á2(CH2), 32Á1(CH2),
35Á8(CH2),64Á1(C2),115Á4,121Á2and124Á0(C6,C7and
C8), 138Á8 (C5), 151Á2 (C9 and C10), 172Á8 (Cˆ O). IR
(KBr) n 3261±3590, 1720, 1276, 1113, 1075. MS(EI)
m=z265(M). CalculatedforC14H19NSO2:C,63Á36;H,
7Á22;N,5Á28;S,12Á08;found:C,63Á65;H,7Á19;N,5Á35;
S , 12Á40.
1
(t, 3H, J ˆ 6Á1, CH3), 2Á52 (q, 2H, J ˆ 5Á6, CH2),
2Á71 (t, 2H, J ˆ 5Á6, CH2), 3Á15 (t,1H, J ˆ 5Á5, H3),
4Á20 (q, 2H, J ˆ 6Á1, CH2), 4Á41 (t, 2H, J ˆ 5Á5, H2),
5Á86 (d,1H, J ˆ 11Á1, ˆ CH), 6Á70±6Á75 (m, 2H,
Harom and ˆ CH), 6Á90±7Á10 (m, 2H, Harom). 13C
NMR (62Á9 MHz, CDCl3) 14Á3 (CH3), 25Á6 (C3),
31Á7 (2CH2), 60Á2 (CH2), 64Á8 (C2), 116Á5 (Ar),
117Á0 (C10), 121Á8 (Ar and ˆ CH), 124Á9 (C7),
138Á2 (C5), 148Á0 ( ˆ CH), 152Á0 (C9), 166Á6
(C ˆ O). IR (neat) n 1719, 1305, 1247, 1193,
1081. Calculated for C15H18SO3: C, 64Á71; H,
6Á53; S, 11Á52; found: C, 64Á55; H, 6Á49; S, 11Á47.
N-Methyl-3-(4-dioxo-2,3-dihydro-1,4-benzoxathiin-
5-yl)propanamide(8). Sodiumdihydrogenphosphate
buffer solution (0Á16 M, 0Á2 mL, 0Á14 mmol) was
added to a solution of aldehyde 22 (150 mg,
0Á72 mmol). Potassium permanganate solution (1 M,
4Á30 mL, 4Á32 mmol) was then introduced. After stir-
ring for 40 min at room temperature, the reaction was
quenchedwithsaturatedsulphitesodiumsolutionand
manganese oxide was removed by ®ltration. The
®ltrate was washed with dichloromethane. Precipita-
tion of the acid 29 occurred after acidi®cation of the
aqueous layer. Acid 29 (53%) was collected by ®ltra-
tion,driedunderhighvacuumandusedinthenextstep
without further puri®cation. The retroamide 8 was
thenpreparedasawhitesolid,mp157ꢀC,in61%yield,
from acid 29 according to the procedure described for
compound1.1HNMR(300 MHz,CDCl3=D2O)d2Á57
(t, 2H, J ˆ 7Á3, CH2), 2Á78 (s, 3H, CH3), 3Á38 (t, 2H,
J ˆ 7Á3, CH2), 3Á58 (t, 2H, J ˆ 5Á1, H3), 4Á73 (t, 2H,
J ˆ 5Á1, H2), 6Á91 (d,1H, J ˆ 8Á0, Harom), 7Á01 (d,1H,
J ˆ 8Á0, Harom), 7Á35 (m,1H, H7). IR (KBr) n 3292,
1652, 1286, 1230, 1177, 1134, 1075 cmÀ1. MS(CI,
NH3) m=z 270 (M ‡ 1). Calculated for C12H15NO4S:
C,53Á52;H,5Á61;N,5Á20;S,11Á91;found:C,53Á56;H,
5Á57; N, 5Á27; S, 11Á91.
Ethyl 5-(2,3-dihydro-1,4-benzoxathiin-5-yl)pentan-
oate (27). Palladium on charcoal 10% (10% by
weight) was added to a solution of unsaturated ester
26 (350 mg, 1Á24 mmol) in ethanol (10 mL). The
compound was hydrogenated, by means of a Parr
apparatus, at 45 psig and room temperature for 12 h.
The reaction mixture was ®ltered, and the ®ltrate
concentrated in-vacuo. Puri®cation by column
chromatography (ethyl acetate±petroleum ether,
3 : 7) gave the saturated ester 27 (95%) as a yellow
1
oil. H NMR (250 MHz, CDCl3) d 1Á24 (t, 3H,
J ˆ 7Á1, CH3), 1Á59±1Á75 (m, 4H, CH2), 2Á33 (t, 2H,
J ˆ 6Á8, CH2), 2Á56 (t, 2H, J ˆ 6Á8, CH2), 3Á11 (t,
2H, J ˆ 4Á7, H3), 4Á11 (q, 2H, J ˆ 7Á1, CH2), 4Á36 (t,
2H, J ˆ 4Á7, H2), 6Á66±6Á74 (m, 2H, H6 and H8),
6Á91 (m,1H, H7). IR (neat) n 1732, 1305, 1247,
1081. Calculated for C15H20SO3: C 64Á25; H, 7Á20;
S , 11Á43; found: C, 64Á17; H, 7Á18; S, 11Á39.
5-(2,3-Dihydro-1,4-benzoxathiin-5-yl)pentanoic
acid (28). This compound was prepared as a white
solid, mp 92±93ꢀC, in 87% yield, from ester 27
1
according to the procedure described for 10a. H
NMR (250 MHz, DMSO-d6) 1Á30±1Á50 (m, 4H,
CH2), 2Á06±2Á21 (m, 2H, CH2), 2Á34±2Á48 (m,
2H, CH2), 3Á10 (t, 2H, J ˆ 4Á8, H3), 4Á22 (t, 2H,
J ˆ 4Á8, H2), 6Á56 (d,1H, J ˆ 7Á3, Harom), 6Á67 (d,1H,
J ˆ 7Á3, Harom), 6Á85 (m,1H, H7), 11Á95 (br s,1H,
CO2H). 13C NMR (62Á9 MHz, DMSO- d6) 24Á3 and
24Á7 (CH2 and C3), 28Á2 (CH2), 32Á2 (CH2), 33Á4
(CH2), 64Á4 (C2), 115Á7 (Ar), 116Á7 (C10), 121Á6
(Ar), 124Á5 (C7), 139Á1 (C5), 151Á5 (C9), 174Á3
(C ˆ O). IR (KBr) n 3550±2500, 1706, 1305,
1232, 1082. Calculated for C13H18SO3: C, 61Á87;
H, 6Á40; S, 12Á70; found: C, 61Á79; H, 6Á38; S,
12Á65.
Pharmacology
Melatonin receptor-binding assays. The af®nity of
the compounds was evaluated on ovine pars tuber-
alis membranes according to the method described
by Morgan et al (1989a). Different concentrations
of the new compounds were tested, in triplicate, for
competitive inhibition of binding of 50 pM 2-
[
125I]iodomelatonin to ovine pars tuberalis melato-
nin receptors under standard conditions (2 h, 37ꢀC).
Non-speci®c binding was assessed with 10 mM
melatonin and data were analysed by computerized
non-linear regression to determine af®nity values
N-Methyl-[5-(2,3-dihydro-1,4-benzoathiin-5-yl)]pen-
tanamide (7). This compound was prepared as a

56
I. CHARTON ET AL
(Ki, equilibrium dissociation constant). Experi-
ments were repeated three times. Results are
expressed as Àlog KiÆ s.d.
alcohols 12 and 13, respectively. Tosylation then
displacement of the resulting tosylates 14 and 15
with potassium cyanide gave the required nitriles
16 and 17. Hydrolysis of the nitriles 16 and 17 with
aqueous sodium hydroxide in ethanol provided,
after acidi®cation, the acids 10b and 18 which react
with appropriate amines to give the desired amides
2 and 4 (Figure 3).
Reaction of the bromide derivative 19 (Charton et
al 1999) with mercury(II) acetate in acetic acid
gave an ester that under basic conditions (K2CO3,
CH3OH, H2O) afforded the corresponding alcohol
21. Oxidation of this compound under Swern con-
ditions led to aldehyde 22 and treatment of this
product with silver nitrate in the presence of aque-
ous sodium hydroxide enabled access to acid 23.
The target compound 5 was prepared by reaction of
acid 23 with methylamine hydrochloride in the
presence of 4-dimethylaminopyridine (DMAP) and
EDCI in N, N-dimethylformamide (Figure 4).
The bromide derivative 19 was converted into the
nitrile 24 which was then hydrolysed with aqueous
sodium hydroxide to the acid 25. Treatment of 25
with methylamine in the presence of EDCI and
HOBt or with n-propylamine hydrochloride in the
presence of DMAP and EDCI, led to the desired
amides 6 (Figure 5).
Cyclic AMP studies. The functional activity of the
compounds was determined by means of a single-
dose cAMP inhibition bioassay, using cultured
ovine pars tuberalis cells as described in detail by
Morgan et al (1989b). Melatonin causes dose-
dependant inhibition of forskolin stimulated
cAMP production in these cells which can be
compared with the melatonin-blocking or mimick-
ing effects of a de®ned concentration of test com-
pound. From these comparisons an activity index is
calculated and provides a semi-quantitative mea-
sure of drug activity in this system. Drugs (10 mM)
were tested, both alone and in combination with
1 nM melatonin, on cells stimulated by 10 mM for-
skolin; inhibitory effects were compared with that
of forskolin or melatonin only. The forskolin=drug
vs forskolin=melatonin index (F=D) provides a
measure of the agonist-like activity of the drug
and the forskolin=drug=melatonin vs forsko-
lin=melatonin index (F=M=D) provides a measure
of antagonist activity (Table 1). All experiments
were conducted in triplicate for each treatment and
the experiments were repeated three times.
The aldehyde 22 enabled access to compound 27
by successive use of Wittig's reaction with (carb-
ethoxymethylene)triphenylphosphorane and cata-
lytic reduction, in the presence of palladium on
carbon, of the intermediate unsaturated ester 26
(E isomer only). Saponi®cation (10% NaOH,
C2H5OH) and amidi®cation (methylamine hydro-
chloride, DMAP, EDCI) gave the desired amide 7
(Figure 6).
The target sulphone 8 was synthesized by oxi-
dation of the aldehyde 22 with potassium perman-
ganate then treatment of the acid obtained, 29, with
methylamine, as before (Figure 7).
Results and Discussion
Chemistry
The synthesis of the novel melatonin analogues 1±
8 was achieved by the routes depicted in Figures
2±7. Saponi®cation of esters 9a and 9c (Guillaumet
et al 1998) with aqueous sodium hydroxide in
ethanol gave, after acidi®cation, the corresponding
acids 10a and 10c. Treatment of these compounds
with methylamine in the presence of EDCI and 1-
hydroxybenzotriazole (HOBt) in N, N-dimethyl-
formamide led to the desired amides 1 and 3
(Figure 2).
Pharmacology
The two best compounds of the series were (2a)
and (6a) with identical N-methylbutyramide side-
Reduction of 9a and 11 (Guillaumet et al 1998)
with lithium aluminium hydride in ether gave the
Figure 2. Synthesis of 2,3-dihydro-1,4-benzodioxin derivatives 1 and 3. Reagents and conditions: i. 10% NaOH in C2H5OH,
re¯ux, then HCl; ii. CH3NH2, HOBt, EDCI, DMF, room temp.

SYNTHESIS AND ACTIVITY OF NEW MELATONIN RECEPTOR LIGANDS
57
Figure 3. Synthesis of derivatives 2a, 2b and 4b. Reagents and conditions: i. LiAlH4, (C2H5)2O, room temp., then re¯ux; ii. TsCl,
(C2H5)3N, CH2Cl2, room temp.; iii. KCN, DMF, 100ꢀC; iv. 10% NaOH in C2H5OH, re¯ux, then 1 M HCl, room temp.; v. CH3NH2,
‡
HOBt, EDCI, DMF, room temp.; vi. n-PrNH3 ClÀ, EDCI, DMAP, DMF, room temp.
Figure 4. Synthesis of 2,3-dihydro-1,4-benzooxathiin analogue 5. Reagents and conditions: i. mercury(II) acetate, CH3COOH,
re¯ux; ii. K2CO3, CH3OH, H2O, room temp.; iii. (ClCO)2, DMSO, CH2Cl2, À50ꢀC, then (C2H5)3N, À50ꢀC; iv. AgNO3, aq. NaOH,
C2H5OH, room temp.; v. CH3NH3 ClÀ, DMAP, EDCI, DMF, room temp.
‡
chains (Table 1). They were better melatoninergic
ligands than A (Charton et al 1999).
dependent on side-chain length. Similar dis-
crepancies in binding and activation have been
reported (Leclerc et al 1998) for other retroamide
compounds.
For each skeleton tested, butyramide (2a, 2b, 4b,
6a, 6b) and pentanoylamide (3, 7) side-chains
resulted in similar af®nities. This was not true of
the propionamide side-chain (1), which led to loss
of af®nity. The worst af®nity was obtained with the
4,4-dioxo-2,3-dihydro-1,4-benzoxathiin skeleton
(compound 8).
The biological activity of the molecules was most
in¯uenced by variation of the length of the side-
chain. Indeed, irrespective of the type of skeleton,
we obtained either agonist compounds with a
butyramide side-chain (2a, 2b, 6a) or partial ago-
nist or antagonist compounds with a pentanoyla-
mide side-chain (3, 7).
In this retroamide series, propyl substitution of
the retroamide did not lead to the increase in af®-
nity usually observed in amide series (Copinga et al
1993; Depreux et al 1994; Garratt et al 1996;
Leclerc et al 1998; Teh & Sugden 1998). Accom-
modation of this alkyl group in the putative
hydrophobic binding pocket possibly differs
because the retroamide group induces a change of
conformation at the binding site and reduced
hydrophobic interaction compared with the usual
amide function.
If the ratio F=D (an estimate of agonist activity)
is compared for compounds in the 2,3-dihydro-1,4-
benzodioxin, 1,4-benzodioxin, and 2,3-dihydro-
1,4-benzoxathiin series with side-chains of the
Therefore, even though these compounds have
similar binding interactions with the receptor,
subsequent activation of biological effects is

58
I. CHARTON ET AL
Figure 5. Synthesis of retroamides 6a and 6b. Reagents and conditions: i. KCN, DMF, room temp.; ii. 10% NaOH in C2H5OH,
‡
re¯ux, then 1 M HCl, room temp.; iii. CH3NH2, HOBt, EDCI, DMF, room temp.; iv.n-PrNH3 ClÀ, EDCI, DMAP, DMF, room
temp.
Figure 6. Synthesis of pentanoylamide analogue 7. Reagents and conditions: i. (C6H5)3P ˆ CHCOOC2H5, toluene, re¯ux; ii. H2,
EDCI, DMF, room temp.
‡
Pd=C, C2H5OH, 45 psig, room temp.; iii. 10% NaOH in C2H5OH, re¯ux, then 1 M HCl, room temp.; iv. CH3NH3 ClÀ, DMAP,
Figure 7. Synthesis of sulphone derivative 8. Reagents and conditions: i. KMnO4, NaH2PO4, t-BuOH; ii. CH3NH2, HOBt, EDCI,
DMF.
same length it seems that a double bond (4b) or a
sulphur atom in the ortho position of the side-chain
(6b) lead to partial or total loss of agonist activity
(reduced value of F=D).
Despite the similar binding values, in the 2,3-
dihydro-1,4-benzodioxin series the butyramide 2b
(F=D ˆ 0Á90) and pentanoylamide 3 (F=D ˆ 0Á56)
are agonist and partial agonist, respectively,
whereas in the 2,3-dihydro-1,4-benzoxathiin series
the butyramide 6b (F=D ˆ 0Á47) and pentanoyla-
mide 7 (F=D ˆ 0Á24) are partial agonist and
antagonist, respectively, and in the 1,4-benzodioxin
series the butyramide 4b (F=D ˆ 0Á53) is a partial
agonist.
Only 2a, 2b and 6a have the same full agonist
activity, with F=D ˆ 0Á97, 0Á90 and 0Á86, respec-
tively. These three compounds have the main fea-
tures required if the activity of these retroamide
compounds is to be highÐoxygen in the position
meta to the side-chain and a butyramide side-chain.
In conclusion, the series of melatoninergic
ligands with a retroamide group in common con-
®rms the importance of the length of the side-chain
in both binding to and activity at the melatonin

SYNTHESIS AND ACTIVITY OF NEW MELATONIN RECEPTOR LIGANDS
59
Table 1. Pharmacological evaluation of melatonin analogues on ovine pars tuberalis membranes.
Binding Activity
Ovine pars tuberalis
Àlog KiÆ s.d. (n ˆ 3)
Agonist index
F=DÆ s.d. (n ˆ 3)
Antagonist index
F=M=DÆ s.d. (n ˆ 3)
Comment
Melatonin
1
2a
2b
3
4b
6a
6b
7
9Á52Æ 0Á10
5Á10Æ 0Á02
8Á08Æ 0Á13
7Á57Æ 0Á06
7Á58Æ 0Á04
7Á60Æ 0Á01
8Á23Æ 0Á07
7Á84Æ 0Á15
7Á71Æ 0Á12
< 4
1
±
±
No activity
0Á07Æ 0Á19
0Á97Æ 0Á26
0Á90Æ 0Á05
0Á56Æ 0Á09
0Á53Æ 0Á01
0Á86Æ 0Á10
0Á47Æ 0Á07
0Á24Æ 0Á05
0Á06Æ 0Á01
0Á85Æ 0Á21
0Á94Æ 0Á12
0Á86Æ 0Á34
0Á87Æ 0Á02
0Á48Æ 0Á12
0Á76Æ 0Á03
0Á87Æ 0Á13
0Á80Æ 0Á01
0Á35Æ 0Á24
0Á98Æ 0Á21
0Á94Æ 0Á25
Full agonist
Full agonist
Partial agonist
Partial agonist
Full agonist
Partial agonist
Antagonist
8
A
No activity
Full agonist
7Á11Æ 0Á01
Dollins, A. B., Zhadanova, I. V., Wurtman, R. J., Lynch, H. J.,
Deng, M. H. (1994) Effect of inducing nocturnal serum
melatonin concentrations in daytime on sleep, mood, body
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91: 1824±1828
receptor, and shows the in¯uence of the skeleton on
the activity of the compounds.
Ã
Fourmaintraux, E., Depreux, P., Lesieur, D., Guardiola-Lemaõ-
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Tetrahydronaphthalenic derivatives as new agonist and
antagonist ligands for melatonin receptors. Bioorg. Med.
Chem. 6: 9±13
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