Synthesis and characterization of pyrrolidine derivatives as potent agonists of the human melanocortin-4 receptor

Article abstract of DOI:10.1016/j.bmcl.2007.09.079

A series of trans-4-phenylpyrrolidine-3-carboxamides were synthesized and characterized as potent ligands of the human melanocortin-4 receptor. Interestingly, a pair of diastereoisomers 20f-1 and 20f-2 displayed potent functional agonist and antagonist activity, respectively. Thus, the 3S,4R-compound 20f-1 possessed a K_i of 11 nM and an EC₅₀ of 24 nM, while its 3R,4S-isomer 20f-2 exhibited a K_i of 8.6 and an IC₅₀ of 65 nM. Both compounds were highly selective over other melanocortin receptor subtypes. The MC4R agonist 20f-1 also demonstrated efficacy in diet-induced obese rats.

Full text of DOI:10.1016/j.bmcl.2007.09.079

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 17 (2007) 6546–6552
Synthesis and characterization of pyrrolidine derivatives as
potent agonists of the human melanocortin-4 receptor
Wanlong Jiang,a Joe A. Tran,a Fabio C. Tucci,a, Beth A. Fleck,b Sam R. Hoare,b
Stacy Markison,c Jenny Wen,d Caroline W. Chen,a Dragan Marinkovic,a
Melissa Arellano,a Alan C. Fosterc and Chen Chena,*
aDepartment of Medicinal Chemistry, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA
bDepartment of Pharmacology, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA
cDepartment of Neuroscience, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA
dDepartment of Preclinical Development, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA
Received 9 August 2007; revised 20 September 2007; accepted 24 September 2007
Available online 22 October 2007
Abstract—A series of trans-4-phenylpyrrolidine-3-carboxamides were synthesized and characterized as potent ligands of the human
melanocortin-4 receptor. Interestingly, a pair of diastereoisomers 20f-1 and 20f-2 displayed potent functional agonist and antagonist
activity, respectively. Thus, the 3S,4R-compound 20f-1 possessed a Ki of 11 nM and an EC50 of 24 nM, while its 3R,4S-isomer 20f-2
exhibited a Ki of 8.6 and an IC50 of 65 nM. Both compounds were highly selective over other melanocortin receptor subtypes. The
MC4R agonist 20f-1 also demonstrated efficacy in diet-induced obese rats.
ꢀ 2007 Elsevier Ltd. All rights reserved.
The melanocortin-4 receptor (MC4R) is a member of
the G-protein-coupled receptor (GPCR) superfamily,
and plays an important role in regulating feeding
behavior and other biological functions.1 Therefore,
MC4R agonists have been extensively studied in efforts
to discover small molecules for the treatment of obes-
ity.2 Several MC4R agonists from different chemical
classes have been reported.3 Recently, a series of pyrr-
olidines exemplified by 1 (Fig. 1) has been characterized
as potent and selective MC4R agonists.4 In our efforts
to find orally active small molecule MC4R antagonists,
we have discovered a series of piperazinebenzylamines
attaching a 3-phenylpropionyl group (2) as potent and
selective MC4R antagonists.5 Introducing a small group
next to the carbonyl moiety increases the binding affin-
ity of 2a (Ki = 74 nM). Thus the R-methyl derivative 2b
displayed a Ki of 26 nM, while the pyrrolidinone 2c
(Ki = 4.5 nM) had over 15-fold improvement over 2a,
demonstrating a role of a small group at this site. To
further reduce flexibility of the molecule, we synthesized
a trans-pyrrolidine 3 (Ki = 610 nM), which exhibited
only moderate binding affinity. We then embarked upon
an SAR study around this core structure, and here we
report the characterization of this series of compounds
as potent agonists and antagonists of the human MC4
receptor.
A set of close analogs of 3 were synthesized as shown in
Scheme 1. Coupling reactions of the trans-N-benzylpyrr-
olidinecarboxylic acids 5 with the benzylamine 46 affor-
ded the amides 6. Selective deprotection of 6b using HCl
in methanol gave the Boc-derivative 8. Treatment of 6
with trifluoroacetic acid selectively removed the Boc-
group to afford the key intermediates 7. Acylation of
7b provided, after a HCl/MeOH treatment, the amides
9a–c, while treatment of 7b with methanesulfonyl chlo-
ride in the presence of triethylamine gave the sulfon-
amide 10. Reductive alkylations of 7 with carbonyl
compounds afforded the tertiary amines 3, 11a–c, and
12, after removing the sulfinyl protecting group. The
aniline 11d was obtained by the reaction of 7b with
bromobenzene under palladium-catalyzed conditions,7
followed by HCl in methanol at room temperature for
1 h. For the compounds with a Boc-protected amine
Keywords: Synthesis; Pyrrolidine; Melanocortin-4 receptor; Agonist;
Antagonist; Potency; Efficacy.
*
Corresponding author. Tel.: +1 858 617 7634; fax: +1 858 617
7967; e-mail: cchen@neurocrine.com
Present address: Department of Medicinal Chemistry, Tanabe
Research Laboratories, Inc., 4540 Towne Centre Ct., San Diego,
CA 92121, USA.
0960-894X/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.09.079

W. Jiang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6546–6552
6547
F3C
F3C
N
N
R
N
N
N
N
O
N
NH2
N
Cl
F
NH2
O
O
O
Cl
O
2a: R = H, Ki = 74 nM
2b: R = Me, Ki = 26 nM
2c: R = 2-oxopyrrolidin-1-yl, Ki = 4.5 nM
Cl
F
1
Cl
3 (trans): Ki = 610 nM
Figure 1. Chemical structures of pyrrolidine agonists and piperazinebenzylamine antagonists of the MC4 receptor.
F3C
O
O
O
F3C
O
X
N
N
N
N
a
+
N
NH
NH
NH
HO
S
O
S
O
O
O
X
5a: X = 2,4-Cl
5b: X = 4-Cl
5c: X = 4-MeO
6
4
b
c
R1
N
F3C
F3C
O
F3C
O
O
d,c
NH
N
N
N
N
N
N
N
NH
S
NH2
NH2
O
O
O
O
X
7
8
9
Cl
Cl
g,c
e,c
f,c
F3C
F3C
F3C
O
R2
N
S
N
O
N
N
N
N
N
N
N
NH2
NH2
NH2
O
O
O
3: X = ,2,4-Cl;
11: X = 4-Cl
12: X = 4-MeO
X
Cl
Cl
10
11d
Scheme 1. (a) EDC/HOBt/NaHCO3/DMF/CH2Cl2/rt, 16 h, 87% for 6b; (b) TFA/CH2Cl2/rt, 1 h, quantitative; (c) HCl/MeOH/rt, 1 h; (d) R1COCl/
Et3N/THF/rt, 8 h, >30%, or BocNHCH2COOH/EDC/HOBt/NaHCO3/CH2Cl2/rt, 10 h, then TFA/CH2Cl2/rt, 1 h, 54% for 9c; (e) MeSO2Cl/Et3N/
THF/rt, 1 h; (f) carbonyl compound/NaBH(OAc)3/HOAc/CH2Cl2/rt, 8 h, 17% for 11a; (g) C6H5Br/Pd(OAc)2/( )BINAP/Cs2CO3/dioxane/100 ꢁC,
24 h, 7.4%.
such as 9c and 11c, a TFA treatment was required
before purification using an HPLC instrument.8
pyrrolidinecarboxamides 15, which were separated by
chromatography into the two diastereoisomers 15-1
and 15-2 (Scheme 2).9 The absolute stereochemistry of
15–2 was resolved by X-ray crystal structure determina-
tion (Fig. 2a). The crystal structure of S-benzyloxazoli-
none 16-1 was also obtained using a similar process
from (4S)-benzyl-2-oxazolidinone to confirm the stereo-
chemistry of 15-1 (Fig. 2b).10 Debenzylation of 15-1 or
To obtain the two single stereoisomers of pyrrolidine 5b,
we first coupled (4R)-benzyl-2-oxazolidinone (R-13)
with 4-chlorocinnamic acid to give the oxazoline
14, which was cyclized with N-(methoxymethyl)-N-
(trimethylsilylmethyl)benzylamine to provide the

6548
W. Jiang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6546–6552
O
O
Cl
b
O
O
N
N
O
O
NH
N
a
S
R
R
S
O
+
O
N
N
O
R
R
O
O
14
R-13
Cl
Cl
15-2
15-1
(X-ray structure)
c
c
O
O
O
O
O
O
O
O
N
N
O
N
S
R
O
N
S
HO
HO
O
Cl
16-1 (X-ray structure)
Cl
Cl
5b-1
5b-2
Scheme 2. (a) i—4-chlorocinnamic acid/Me3CCOCl/Et3N/THF/ꢀ20 ꢁC, 2 h; ii—LiCl/ꢀ20 ꢁC, 16 h, 93%; (b) i—Me3SiCH2N(Bn)(CH2OMe)/TFA/
toluene/0 ꢁC, 16 h; ii—Chromatography separation, 40% for 15-1 and 46% for 15-2; (c) i—ACE-Cl/proton sponge/ClCH2CH2Cl/0 ꢁC to reflux, 1 h,
then MeOH/reflux, 0.5 h; ii—(Boc)2O/NaHCO3/H2O/dioxane/rt, 16 h, 97%; iii—H2O2/LiOH//H2O/0 ꢁC, 2 h, 96% for 5b-1.
radiolabeled ligand. Compounds with good binding
affinity were also tested in a whole cell functional ago-
nist assay measuring accumulation of cAMP with com-
petitive ELISA as previously reported.11
In the binding assay, the secondary amine
7
(Ki = 82 nM) was more potent than its N-Boc analog 8
(Ki = 390 nM). In comparison, the acetamide 9a had
better binding affinity than 8, while the benzoyl analog
9b slightly improved potency, and the glycine derivative
9c and the methanesulfonamide 10 had similar Ki values
to 7. Therefore, except for 8, there was no clear SAR at
this site of the molecule. In the functional assay, only the
acetamide 9a (EC50 = 1,900 nM, IA = 72%) had compa-
rable intrinsic activity (IA) to a-MSH, which was used
as a standard, but with much lower potency. The other
amides, as well as the parent amine 7, had low intrinsic
activity (IA < 35%). Introducing an N-isopropyl group
to 7 not only improved its binding affinity but also
significantly increased its intrinsic activity (11a, Ki =
26 nM, EC50 = 1,100 nM, IA = 94%). The N-benzyl
compound 11b was less potent in both binding and func-
tional assays than 11a, while the N-phenyl analog 11d
had very low intrinsic activity, suggesting that the basic
nitrogen of the pyrrolidine plays a role in receptor
activation. However, an additional amine in 11c did
not increase efficacy. Finally, the 4-methoxy compound
12 (Ki = 67 nM) was less potent than its 4-chloro analog
11a, which was the most efficacious among the com-
pounds in Table 1.
Figure 2. (a) (Left) and (b) (right). X-ray crystal structures of 15-2 and
16-1.
15-2 was achieved with ACE-Cl, and the resultant amine
was protected with a Boc-group, which was hydrolyzed
using H2O2/LiOH to give the single isomer 5b-1 or 5b-2.
Coupling reactions of 5b with a-isopropylbenzylamines
174 gave the amides 18. Acylations of 18a and 18b with
various carboxylic acids provided the amides 19 and 21,
respectively, after a HCl/MeOH treatment. Reductive
alkylations of 18a and 18b with a variety of aldehydes
and ketones provided the tertiary amines 20 and 22,
respectively, after deprotections. Compounds 20f-1 and
20f-2 were synthesized using a similar procedure from
5b-1 and 5b-2, respectively. The piperidine intermediate
23 was selectively deprotected with trifluoroacetic acid,
followed by reductive alkylations to give 22i–k after an
HCl/MeOH treatment at room temperature (Scheme 3).
These initial results prompted us to expand the SAR
study, which is summarized in Table 2. The acetamides
19a and 21a had moderate binding affinity which was
slightly less potent than 9a, although 19a had lower effi-
cacy than 9a. Other amides 19b–g and 21c–g did not
All of the final compounds were tested in a binding as-
say using membranes from HEK293 cells expressing
human MC4 receptor and [125I]-NDP-MSH as the

W. Jiang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6546–6552
6549
H
N
O
O
R1
Y
Y
N
Y
N
N
a,b
c,d
N
e
NH
N
O
N
NH
S
O
NH
S
O
Cl
NH2
Cl
17a: Y = 4-Me
17b: Y = 6-F
18a,b
19,21
e,d
R3
N
Boc
N
R2
b,e,d
N
N
N
Y
N
N
N
N
N
N
O
O
O
NH2
NH
S
O
NH2
Cl
Cl
Cl
20,22
23
20i-k
Scheme 3. (a) 5b/EDC/HOBt/NaHCO3/DMF/CH2Cl2/rt, 16 h, 88% for 18a; (b) TFA/CH2Cl2/rt, 1 h, 98%; (c) R1COOH/EDC/HOBt/NaHCO3/
DMF/CH2Cl2/rt, 16 h; (d) HCl/MeOH/rt, 1 h; (e) carbonyl compound/NaBH(OAc)3/CH2Cl2/rt, 1–16 h.
Table 1. SAR of pyrrolidine derivatives 7–12 at MC4Ra
compounds 20d (Ki = 6.6 nM) and 22h (Ki = 9.3 nM)
exhibited good binding affinity, however, their intrinsic
activity was low (21% and 49%, respectively). An
F3C
R
N-cyclopentyl analog 22g (Ki = 26 nM, EC50 = 400 nM,
IA = 80%) had high efficacy but moderate potency in the
cAMP assay. A series of N-piperidin-4-yl derivatives
20h–k possessed high binding affinities (Ki 6 20 nM),
especially 20k (Ki = 5.4 nM) with an N-isobutylpiperidi-
nyl group, but these compounds exhibited negligible
intrinsic activity.
N
N
N
NH2
O
7-12
X
Compound
X
R
H
Ki (nM)
EC50 (nM)b
While the N-tetrahydropyran-4-yl derivative 20f
(Ki = 12 nM) as a mixture of trans-isomers exhibited
high binding affinity and moderate intrinsic activity
(IA = 36%), the two single stereoisomers 20f-1 and
20f-2 (Fig. 3), prepared separately from the pyrrolidin-
ecarboxylic acids 5b-1 and 5b-2, displayed quite inter-
esting results. While these two stereoisomers
possessed similar binding affinity (Ki of 11 and
8.6 nM, respectively, for 20f-1 and 20f-2), 20f-1 was a
potent and full agonist with an EC50 of 24 nM, and
20f-2 had very low intrinsic activity (18% cAMP stim-
ulation up to 10 lM concentrations). Instead, in a
functional antagonist assay, 20f-2 dose-dependently
inhibited a-MSH-stimulation of cAMP production
with an IC50 value of 65 nM (Fig. 3), demonstrating
that 20f-2 was a functional antagonist.12 In compari-
son, both single isomers (22i-1 and 22i-2) of the
trans-N-benzylpyrrolidine 22i had moderate binding
affinity and low intrinsic activity, suggesting the N-sub-
stituent of the pyrrolidine plays an important role in
receptor binding as well as activation.
7
Cl
Cl
82
390
66
(34%)
(32%)
8
Boc
9a
9b
9c
Cl
Cl
MeCO
PhCO
COCH2NH2
1900 (72%)
(3%)
(12%)
30
96
Cl
Cl
10
11a
11b
11c
11d
12
MeSO2
i-Pr
95
26
(34%)
Cl
Cl
1100 (94%)
7300 (52%)
(37%)
Bn
CH2CH2NH2
120
180
260
67
Cl
Cl
Ph
i-Pr
(24%)
450 (67%)
MeO
a Average of two or more independent measurements.
b Intrinsic activity (100% of the endogenous ligand a-MSH) is indi-
cated in parentheses.
have significant improvement on binding affinity except
the cyclopropylcarboxamide 21b (Ki = 31 nM) which
was about 7-fold better than 21a. The N-isopropyl
amines 20a (Ki = 12 nM, EC50 = 105 nM, IA = 99%)
and 22a (Ki = 21 nM, EC50 = 270 nM, IA = 86%) pos-
sessed good binding affinity and moderate agonist po-
tency with full efficacy. Similar to 11b, the N-benzyl
analogs 20g and 22i had low intrinsic activity. Among
the tertiary amines 20b–g and 22b–i, the N-cyclohexyl
Compounds 20f-1 and 20f-2 were also found to be
highly selective at the MC4 receptor over the other
receptor subtypes (Table 3). This potent and selective

6550
W. Jiang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6546–6552
Table 2. SAR of 1-substituent of pyrrolidines 19–22 at hMC4Ra
R'
N
Y
6
4
N
1
N
O
NH2
Cl
19,20: Y = 4-Me
21,22: Y = 6-F
Compound
Y
R0
Ki (nM)
EC50 (nM)b
19a
19b
19c
19d
19e
19f
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
6-F
MeCO
EtCO
110
59
(35%)
(31%)
(14%)
(22%)
(0)
n-PrCO
cBuCO
Ph(CH2)2CO
68
78
75
Ph(CH2)3CO
Ph(CH2)4CO
100
65
(0)
(0)
19g
21a
21b
21c
21d
21e
21f
MeCO
cPrCO
220
31
6-F
6-F
n-PrCO
i-PrCO
260
250
540
350
160
12
6-F
6-F
t-BuCO
cHxCO
6-F
6-F
21g
20a
20b
20c
20d
20f
PhCO
i-Pr
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
4-Me
6-F
105 (99%)
(36%)
(27%)
MeOCH2CH(Me)
(MeOCH2)2CH
64
200
6.6
12
cHx
4-THP
(21%)
(36%)
20f-1
20f-2
20g
20h
20i
S,S,R
S,R,S
11
24 (105%)
(18%)c
(52%)
8.6
36
Bn
t-Boc
15
18
(0)
(3%)
1-Ethyl-4-piperidinyl
1-Isopropyl-4-piperidinyl
20j
18
(1%)
(1%)d
270 (86%)
20k
22a
22b
22c
22d
22e
22f
1-Isobutyl-4-piperidinyl
i-Pr
Me
5.4
21
6-F
6-F
81
HOCH2CH2
CF3CH2CH2
i-Bu
580
205
43
6-F
6-F
1400 (52%)
6-F
6-F
cBu
cPn
32
26
22g
22h
22i
400 (80%)
(49%)
(19%)
6-F
6-F
cHx
Bn
9.3
94
22i-1
22i-2
6-F
6-F
S,S,R
S,R,S
340
98
(22%)
(3%)
a Data are average of two or more independent measurements.
b Intrinsic activity is indicated in parentheses.
c Dose-dependent inhibition of a-MSH-stimulated cAMP release with an IC50 of 65 nM (Fig. 3, right).
d IC50 = 1800 nM.
MC4R agonist 20f-1 was further characterized for its
pharmacokinetic properties in rats. After an intravenous
injection at 2.5 mg/kg, 20f-1 displayed a very high plas-
ma clearance of 118 mL/min kg, which was associated
with its extremely high tissue distribution (Vd = 270 L/
kg), resulting in a long half life of 27 h in this species.
The high volume of distribution could be the result of
its dibasic structure. However, the brain distribution
of this compound was low. Thus the whole brain con-
centrations were only 5 and 32 ng/g at 1 and 4 h post-
dosing, respectively, resulting in a brain to plasma
ratio of 0.7–1.3 during this time frame. 20f-1 had a logD
value of 1.8 measured by a shake–flake method, and the
low brain penetration most likely is associated with its
transporter activity. In an in vitro Caco-2 assay, 20f-1
displayed a Papp of 0.8 · 10ꢀ6cm/s from apical (a) to
basolateral (b) direction, which resulted in a (b to a/a
to b) ratio of 25, demonstrating strong P-glycoprotein
activity.
After oral administration of 20f-1 at a 10 mg/kg dose,
the Cmax was only 16 ng/mL, which appeared at about
the 3 h time point, and the AUC was 154 ng/mL h.
The oral bioavailability of 20f-1 was moderate (22%).

W. Jiang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6546–6552
6551
O
O
N
N
N
N
N
N
NH2
NH2
O
O
20f-1 (S,S,R)
Cl
20f-2 (R,S,S)
Cl
100
110
α-MSH
20f-1
20f-2
100
90
80
70
60
50
40
30
20
10
0
90
80
70
60
50
40
30
20
10
0
-10
-9
-8
-7
-6
-5
-10
-9
-8
-7
-6
-5
log [ligand] (M)
log [20f-2] (M)
Figure 3. Chemical structures of 20f-1 and 20f-2 with absolute stereochemistry and functional agonist activity of 20f-1 and 20f-2 (left) and functional
antagonist activity of 20f-2 (right).
positive control fenfluramine significantly decreased
cumulative food intake at all time points tested. By the
24 h time point all doses of 20f-1 significantly and
dose-dependently reduced feeding (p < 0.05; Fig. 4).13
Table 3. Binding affinity (Ki, nM) of 20f-1 and 20f-2 at the human
melanocortin receptorsa
Compound
MC1
MC3
MC4
MC5
20f-1
20f-2
3350
(33%)b
1600
1400
11
240
380
8.6
In conclusion, a series of 3-phenylpyrrolidine-3-carbox-
amides were synthesized and studied as MC4 receptor
ligands. While the initial 2,4-dichlorophenylpyrrolidine
3 displayed lower binding affinity than the correspond-
ing phenylpropionyl analogs 2, potent functional
antagonists such as 20f-2 were identified. More inter-
estingly, its diastereoisomer 20f-1 possessed potent ago-
nist activity. Thus, 20f-1 had an EC50 of 24 nM in a
cAMP assay and was highly selective over other mela-
nocortin receptor subtypes in binding affinity. This
compound also demonstrated efficacy in an acute
DIO model.
a Data are average of two and more independent measurements.
b Percentage inhibition at 10 lM concentration.
The low oral plasma exposure is due to its high tissue
distribution coupled with its low oral bioavailability.
This MC4R agonist 20f-1 was further characterized
in vivo to examine the effects on deprivation-induced
food intake in diet-induced obese (DIO) rats (n = 8 per
group). Both the 30 mg/kg dose (ip) of 20f-1 and the
Vehicle
3 mg/kg
10 mg/kg
30 mg/kg
Fen (5 mg/kg)
35
30
9
8
7
6
5
4
3
2
1
25
*
20
*
*
15
10
5
*
*
*
*
*
*
*
*
*
24 hrs
1 hr
2 hrs
4 hrs
6 hrs
Figure 4. Effect of 20f-1 (3, 10, 30 mg/kg, ip) on deprivation-induced food intake in diet-induced obese rats. Cumulative food intake was significantly
decreased at all time points for fenfluramine and the 30 mg/kg dose of 20f-1 and for all treatments at the 24 h time point (*p < 0.05). Values are
means SEM.

6552
W. Jiang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6546–6552
Crystallographic Data Centre, 12, Union Road, Cam-
bridge CB2 1EZ, UK.
References and notes
11. Nickolls, S. A.; Cismowski, M. I.; Wang, X.; Wolff, M.;
Conlon, P. J.; Maki, R. A. J. Pharmacol. Exp. Ther. 2003,
304, 1217.
1. Cone, R. D. Nat. Neurosci 2005, 8, 571.
2. Yang, Y. K. Obes. Rev. 2003, 4, 239.
3. For a recent review, see: Chen, C. Prog. Med. Chem. 2007,
45, 111.
4. Ujjainwalla, F. 230th ACS National Meeting, Washington,
DC, USA. 2005, MEDI 275.
5. Jiang, W.; Tucci, F. C.; Chen, C. W.; Arellano, M.; Tran,
J. A.; White, N. S.; Marinkovic, D.; Pontillo, J.; Fleck, B.
A.; Wen, J.; Saunders, J.; Madan, A.; Foster, A. C.; Chen,
C. Bioorg. Med. Chem. Lett. 2006, 16, 4674.
12. Functional efficacy of 20f-1 (agonist) and 20f-2 (antago-
nist): Accumulation of cAMP production in CHO cells
transfected with hMC4R was measured and analyzed using
an ELISA. Data are from representative experiments
performed 2–8 times with similar results. (Fig. 3, left): 20f-
1, 20f-2, and a-MSH stimulation of cAMP production,
where 100% cAMP is defined as the Emax of a-MSH
stimulated cAMP production. (Fig. 3, right): 20f-2 antag-
onism of a-MSH stimulated cAMP production, where
100% cAMP is defined as stimulation of cAMP production
by 10 nM a-MSH (ꢁEC50) in the absence of antagonist.
13. Male CD rats (Charles River) were fed a high fat diet from
weaning until 12 weeks of age. Food was removed 24 h prior
to testing. On the test day rats were injected ip with vehicle
(sterile water), 5 mg/kg fenfluramine (Sigma), or 3, 10, or
30 mg/kg of 20f-1. Rats were then placed into individual test
cages with water available. Fifteen minutes after dosing,
pre-measured test diet was presented to the rats and food
intake was measured 1, 2, 4, 6, and 24 h later.
6. Jiang, W.; Chen, C.; Marinkovic, D.; Tran, J. A.; Chen, C.
W.; Arellano, L. M.; White, N. S.; Tucci, F. C. J. Org.
Chem. 2005, 70, 8924.
7. Wolfe, J. P.; Buchwald, S. L. J. Org. Chem. 2000, 65, 1144.
8. For experimental detail, see: WO 2005/040109.
9. (a) Karlsson, S.; Han, F.; Hogberg, H.-E.; Caldirola, P.
Tetrahedron: Asymmetry 1999, 10, 2605; (b) Karlsson, S.;
Hogberg, H.-E. Tetrahedron: Asymmetry 2001, 12, 1975.
10. CCDC 661517 (for 15-2) and CCDC 661518 (for 16-1)
contain the supplementary crystallographic data for this
paper. These data can be obtained via the CDCC website
(www.ccdc.cam.ac.uk), or by contacting The Cambridge