Synthesis and characterization of trans-4-(4-chlorophenyl)pyrrolidine-3-carboxamides of piperazinecyclohexanes as ligands for the melanocortin-4 receptor

Article abstract of DOI:10.1016/j.bmcl.2007.10.032

A series of trans-N-alkyl-4-(4-chlorophenyl)pyrrolidine-3-carboxamides of piperazinecyclohexanemethylamines was synthesized and characterized for binding and function at the melanocortin-4 receptor (MC4R), and several potent benzylamine derivatives were i

Full text of DOI:10.1016/j.bmcl.2007.10.032

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 17 (2007) 6825–6831
Synthesis and characterization of trans-4-(4-chlorophenyl)
pyrrolidine-3-carboxamides of piperazinecyclohexanes
as ligands for the melanocortin-4 receptor
Caroline W. Chen,a Joe A. Tran,a Beth A. Fleck,b Fabio C. Tucci,a,
Wanlong Jianga 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
Received 25 July 2007; revised 5 October 2007; accepted 8 October 2007
Available online 17 October 2007
Abstract—A series of trans-N-alkyl-4-(4-chlorophenyl)pyrrolidine-3-carboxamides of piperazinecyclohexanemethylamines was syn-
thesized and characterized for binding and function at the melanocortin-4 receptor (MC4R), and several potent benzylamine deriv-
atives were identified. Compound 18v was found to bind MC4R with potent affinity (Ki = 0.5 nM) and high selectivity over the other
melanocortin subtypes and behaved as a functional antagonist (IC50 = 48 nM).
Ó 2007 Elsevier Ltd. All rights reserved.
The melanocortin-4 receptor (MC4R) is a member of
the G-protein-coupled receptor superfamily and plays
an important role in biological functions, including
regulation of feeding behavior.1 MC4R agonists have
been extensively studied in an effort to discover small
molecules for the treatment of obesity,2 and several
small molecule MC4R agonists from different chemical
classes have been reported.3 MC4R antagonists, on
the other hand, have been shown to reverse lean body
mass loss and increase food intake in animal models
of cachexia, suggesting potential in cancer cachexia
treatment.4,5
ent than many reported small molecule MC4R agonists,
such as 1a, in which a Tic-(4-Cl)Phe dipeptide is re-
quired as an ‘address element’.2 We have recently shown
that by combining the trans-4-arylpyrrolidine-3-car-
bonyl moiety of 1b with the piperazinebenzylamine
N,N-dimethylaminopropionamide of the antagonist 2,
potent MC4R agonists have been identified from the
resulting compounds. For example, 3 (Ki = 4.7 nM) pos-
sesses an EC50 value of 16 nM with intrinsic activity (IA)
of 102% relative to the endogenous ligand a-MSH.8 We
have also previously reported that the piperazinecyclo-
hexanes with the Tic-(4-Cl)Phe dipeptide are MC4 ago-
nists and can be converted into functional antagonists
by replacing the dipeptide. Thus, 4 is a potent agonist
(Ki = 16 nM, EC50 = 33 nM, IA = 96%),9 while the b-
Ala-(2,4-Cl)Phe analog 5a is a functional antagonist
(Ki = 8.8 nM, IC50 = 260 nM).10 Like the amide 5a, the
benzylamine 5b (Ki = 18 nM) also possesses potent bind-
ing affinity.
Pyrrolidine derivatives were first reported by Ujjainwal-
la as potent and selective MC4R agonists.6 An example
of this chemical class is compound 1b, which has a bind-
ing IC50 of 14 nM and a functional EC50 of 2 nM
(Fig. 1). This functional activity is similar to that re-
ported for THIQ 1a.7 One advantage of compound 1b
is its pyrrolidine structure. This structure is much differ-
Because of the success in converting antagonist 2 into ago-
nist 3, we were interested in the pharmacological proper-
ties of compounds derived from the combination of the
left-side of compound 4 with the pyrrolidine moiety of
1b. Here we report the synthesis and characterization
of trans-4-(4-chlorophenyl)pyrrolidine-3-carboxamides
of 1-(1-piperazine)cyclohexanemethylamine derivatives
7–22 as MC4R ligands.
Keywords: Synthesis; Pyrrolidine; Melanocortin-4 receptor; Agonist;
Antagonist; Structure–activity relationship.
*
Corresponding author. Tel.: +1 858 617 7600; fax: +1 858 617
7967; e-mail: cchen@neurocrine.com
Present address: Department of Medicinal Chemistry, Tanabe
Research Laboratories, U.S.A., Inc., 4540 Towne Centre Ct., San
Diego, CA 92121, USA.
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.10.032

6826
C. W. Chen et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6825–6831
F
Cl
Cl
F
N
O
N
NH
N
N
N
N
N
NH
NH
O
N
O
O
O
O
2: Ki = 2.8 nM
IC50 = 35 nM
N
1b
HN
1a
Cl
Cl
Cl
Cl
N
N
N
N
N
O
NH
NH
N
NH
NH
N
R
NH
O
O
O
O
NH2
O
O
HN
5a: R = PhCH2CO
Ki = 8.8 nM, IC50 = 260 nM
5b: R = PhCH2
N
3: Ki = 4.7 nM
EC50 = 16 nM
IA = 102%
4: Ki = 16 nM,
EC50 = 33 nM, IA = 96%
Ki = 18 nM
Figure 1. Chemical structures of some MC4R agonists and antagonists related to this study.
The core structure of 1-trifluoroacetamidomethyl-1-(1-
piperazine)cyclohexane 26 was synthesized as shown in
Scheme 1. A Strecker reaction of cyclohexanone 23 with
1-benzylpiperazine afforded the intermediate 24, which
was reduced with lithium aluminumhydride, followed
by a treatment with trifluoroacetic anhydride to provide
the protected diamine 25. Debenzylation of 25 catalyzed
by palladium gave the desired amine 26 in a good overall
yield.
tion of 6a with 2-vinylpyridine in the presence of acetic
acid in ethanol gave the 2-pyridinylethylamine 9r.
The key intermediate 6b was synthesized from N-
(methoxylmethyl)-N-(trimethylsilylmethyl)benzylamine
27b using a procedure similar to that for 6a. Reductive
alkylations of 6b with benzaldehyde and 4-pyridylcar-
boxaldehyde afforded the corresponding secondary
amines 10a and 10b, respectively. Debenzylation of
10a or 10b was accomplished using 1-chloroethyl chlo-
roformate to provide the secondary amine 11a or 11b.
Reductive alkylations of 11a with several ketones gave
the tertiary amines 12 after purification. Reactions of
11a–b with acyl chlorides in the presence of triethyl-
amine resulted in the amides 13 and 15, respectively.
Alternatively, coupling reactions of 11a with N-Boc-
amino acids, followed by TFA-treatment, gave the
amides 14. The urea 16 was obtained by a reaction
of 11b with ethylisocyanate, and the sulfonamide 17
was synthesized from methylsulfonyl chloride and 11b
(Scheme 3).
The trans-N-isopropyl-4-(4-chlorophenyl)pyrrolidine-3-
carboxylic acid 28a was synthesized from the cyclization
of N-(methoxylmethyl)-N-(trimethylsilylmethyl)isopro-
pylamine 27a with methyl trans-4-chlorocinnamate med-
iated by trifluoroacetic acid, followed by hydrolysis with
lithium hydroxide. Compound 28a was coupled with the
amine 26 to provide the amide 29a, which was deprotec-
ted under basic conditions to give the primary amine 6a
(Scheme 2).
Coupling reactions of 6a with various carboxylic acids
afforded the amides 7. Similarly, reactions of 6a with
several sulfonyl chlorides gave the sulfonamides 8.
Alternatively, reductive alkylations of 6a with a variety
of aldehydes provided the secondary amines 9. A reac-
Reductive alkylations of 6a with various substituted
benzaldehydes gave the corresponding benzylamines
18. The secondary amines 9p and 18j were further con-
a
b,c
d
N
N
NC
N
F3C
NH
F3C
NH
O
N
NH
N
Ph
Ph
O
O
24
23
25
26
Scheme 1. Reagents and conditions: (a) 1-Benzylpiperazine/KCN/Na2S2O5/H2O/rt, 18 h, 81%; (b) LiAlH4/Et2O/rt, 16 h, 93%; (c) (CF3CO)2O/TEA/
DCM/rt, 2 h, 100%; (d) Pd–C/HCO2NH4/MeOH, ꢀ100%.

C. W. Chen et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6825–6831
6827
Cl
Cl
c
a,b
O
N
Si
N
F3C
NH
N
HO
N
N
O
27a
O
O
29a
28a
Cl
Cl
g
d
N
N
NH
NH2
R3
N
N
N
N
O
6a
O
9a-q
e
h
f
Cl
Cl
N
Cl
N
NH
N
N
N
R1
NH
R2
NH
N
N
S
O
N
N
O
O
O
N
O
O
9r
7
8
Scheme 2. Reagents and conditions: (a) trans-4-ClPhCH@CHCOOMe/TFA/DCM/rt, 6 h, 80%; (b) LiOH/THF/H2O/rt, 3 h, 99%; (c) 26/HBTU/
DIEA/DMF/rt, 16 h, 85%; (d) K2CO3/MeOH/H2O/65 °C, 16 h, 92%; (e) R1COOH/EDC/HOBt/TEA/DCM/rt, 16 h; (f) R2SO2Cl/TEA/rt, 2 h; (g)
aldehyde/NaBH(OAc)3/DCE/rt, 16 h; (h) 2-PyCH@CH2/AcOH/EtOH/90 °C, 4 h, 30%.
Cl
Cl
d
a,b,c
O
N
Si
N
N
F3C
NH
NH2
N
N
N
N
O
O
O
27b
29b
6b
Cl
Cl
Cl
e
N
N
N
f
g
NH
NH
NH
Cl
N
N
N
NH
N
N
R4
O
O
O
A
A
11a: A = H
11b: A = N
10a: A = CH
10b: A = N
12a-d
k
h or i
j
Cl
Cl
N
N
N
NH
NH
NH
H
N
N
N
N
R5
N
O
N
N
S
O
O
O
O
O
O
13a-e,14a-d: A = CH
15a-b: A = N
16
17
A
N
N
Scheme 3. Reagents and conditions: (a) TFA/DCM/rt, 16 h, 57%; (b) LiOH/THF/H2O/rt, 3 h, quantitative; (c) 26/HBTU/DIEA/DMF/rt, 16 h, 9 4%;
(d) K2CO3/MeOH/H2O/65 °C, 16 h, 76%; (e) ArCHO/NaBH(OAc)3/DCE/rt, 16 h, ꢀ40%; (f) MeCH(Cl)OCOCl/DCE/reflux, 4 h, then MeOH/reflux,
2 h; (g) carbonyl compound/NaBH(OAc)3/DCE/rt, 16 h; (h) R5COCl/TEA/DCM/rt, 2 h; (i) N-Boc-amino acid/EDC/HOBt/DIEA/DCM/rt, 16 h,
then TFA/DCM/rt, 1 h; (j) EtNCO/TEA/DCM/rt, 1 h; (k) MeSO2Cl/TEA/DCM/rt, 2 h.

6828
C. W. Chen et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6825–6831
verted to the tertiary amines 19 and 20 and acetamides
21 and 22 as shown in Scheme 4.
results, in combination with those from the amides 7,
suggest that a basic amine as well as an aromatic ring
on the side chain is important for high affinity binding.
In the dipeptide series 5, the basicity of the amine is
unnecessary since both the amide 5a (Ki = 8.8 nM) and
the amine 5b (Ki = 18 nM) possess good binding affinity.
While its binding affinity was similar to that of the Tic-
(4-Cl)Phe dipeptide agonist 4 (Ki = 16 nM), 9c had poor
potency and low efficacy in the functional agonist assay
(EC50 = 1100 nM, IA = 31%). These results clearly indi-
cate that the current series is much different from the
dipeptides such as 4.
The synthesized compounds were tested in a competi-
tion binding assay using HEK293 cells expressing the
human melanocortin-4 receptor and [125I]-NDP-MSH
as the radiolabeled ligand.11 Compounds with potent
binding affinities were also tested in a functional agonist
assay measuring the stimulation of cAMP release. Se-
lected compounds were studied in a functional antago-
nist assay to determine the inhibition of a-MSH-
stimulated cAMP release. Results are shown in Tables
1–4.
The furan and thiophene derivatives 9d–g exhibited
binding affinities similar to 9c, while the electron-defi-
cient aromatic thiazole and imidazole analogs 9h–k were
less potent. The 1,5-dimethylpyrazolemethylamine 9l
(Ki = 11 nM) exhibited the best binding affinity in this
group. The pyrimidine 9m had a Ki value of 220 nM,
while the pyridine analogs 9n–p were more potent than
9m, especially the 4-pyridine 9p (Ki = 27 nM), which
was equal to 9c in binding affinity. In comparison to
9c, the indole derivative 9q and the pyridylethylamine
9r were significantly less potent.
The phenylacetamide 7a (R1 = PhCH2, Ki = 510 nM,
Table 1) only showed moderate binding affinity and
was 30-fold less potent than the Tic-(4-Cl)Phe dipeptide
4 (Ki = 16 nM). All the arylacetamides 7 (16 com-
pounds) showed similar binding affinity (Ki = 260–
820 nM, data not shown) and were only slightly more
potent than the parent primary amine 6a
(Ki = 1200 nM). These results are very different than
the SAR in the dipeptide series 5 where an amide side
chain increases the binding affinity as much as 30-fold.8,9
We also tested several sulfonamides, since this func-
tional group has been used in other small molecule
MC4R agonists.12,13 Like amides 7, the sulfonamides 8
(6 compounds) displayed moderate binding affinity
(Ki = 330–550 nM). For example, the benzyl- and phe-
nylsulfonamides 8a and 8b displayed Ki values of 340
and 330 nM, respectively.
While 9c was a weak partial agonist, several of its ana-
logs exhibited partial agonism with moderate potency
(Table 1). The most potent compound in this group
was 9l which had an EC50 of 50 nM with an intrinsic
activity of 43% of a-MSH. The 4-pyridine 9p displayed
an EC50 of 780 nM with very low efficacy (IA = 27%)
in the functional agonist assay. Additionally, 9c behaved
as a functional antagonist, dose-dependently inhibiting
a-MSH-stimulated cAMP release with an IC50 of
410 nM.
A series of secondary amines 9a–r were then examined
(Table 3). While the two alkylamines 9a and 9b only
exhibited 4-fold improvement in binding affinity over
the primary amine 6a, the benzylamine 9c displayed a
Ki value of 26 nM, which was over 10-fold better than
the cyclohexanemethyl analog 9b (Ki = 300 nM). These
The role of the N-group on pyrrolidine was also studied
in this series of compounds, and the results are summa-
rized in Table 2. For the benzylamines 10a and 12–14,
Cl
Cl
a
N
N
NH2
NH
X
N
N
N
N
O
O
6a
18a-v
Cl
Cl
Cl
c
b
N
N
N
NH
N
O
N
R6
N
N
N
N
N
N
O
O
O
18j: A = COMe
9p: A = N
21: A = COMe
22: A = N
19a-d: A = COMe
20a-c: A = N
A
A
A
Scheme 4. Reagents and conditions: (a) ArCHO/MeOH/rt, 2 h, then NaBH4; (b) aldehyde/NaBH(OAc)3/DCE/rt, 16 h; (c) MeCOCl/TEA/DCM/rt,
2 h.

C. W. Chen et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6825–6831
Table 1. SAR of secondary amines 9a–r at MC4R Table 2. SAR of the N-group of pyrrolidines 10–17 at MC4R
6829
Cl
Cl
N
N
N
N
R'
NH
N
NH
N
R3
9a-r
O
O
10a, 12-14: A = CH
10b, 11b, 15-17: A = N
A
Compound R1
Ki (nM) EC50 (nM)a
6a
7a
8a
8b
9a
9b
9c
9d
9e
9f
H
1200
510
PhCH2CO
PhCH2SO2
PhSO2
Compound
R0
Ki (nM)
EC50 (nM)
340
330
310
300
26
(40%)
(20%)
10a
9c
PhCH2
i-Pr
140
26
(1%)
1100 (31%)
(7%)
MeOCH2CH(Me)
CyclohexaneCH2
PhCH2
12a
12b
12c
12d
13a
13b
13c
13d
13e
14a
14b
14c
14d
10b
11b
15a
15b
16
CH(Me)CH2OMe
Cyclopentyl
Cyclohexyl
4-Tetrahydropyran
MeCO
100
17
5.5a
29
1800 (33%)
1500 (16%)
1000 (70%)
(13%)
1100 (31%)
570 (32%)
480 (53%)
590 (58%)
840 (31%)
740 (72%)
110 (49%)
230 (69%)
(31%)
2-FuranCH2
3-FuranCH2
38
75
190
210
190
210
180
100
430
110
670
200
2200
3000
3100
4600
3400
2-ThiopheneCH2
3-ThiopheneCH2
2-ThiazoleCH2
4-ImidazoleCH2
2-ImidazoleCH2
37
22
EtCO
nPrCO
i-PrCO
(19%)
(8%)
9g
9h
9i
130
220
560
390
(47%)
(26%)
CyclopentaneCO
MeNHCH2CO
MeCH(NH2)CO
NH2CH2CH2CO
i-PrCH(NH2)CO
PhCH2
9j
(8%)
(3%)
9k
9l
1-Me-2-imidazoleCH2
1,5-(Me)2-4-pyrazoleCH2 11
50 (43%)
(9%)
(0%)
9m
9n
9o
9p
9q
9r
5-PyrimidineCH2
2-PyCH2
3-PyCH2
220
76
220 (66%)
1200 (34%)
390 (72%)
780 (27%)
35
H
MeCO
4-PyCH2
1-Me-2-indoleCH2
2-PyCH2CH2
27b
1200
690
EtCO
EtNHCO
MeSO2
a Data are average of two or more independent measurements; intrinsic
activity is indicated in parentheses.
b Compound 9p dose-dependently inhibited a-MSH-stimulated cAMP
release with an IC50 value of 410 nM.
17
a Compound 12c dose-dependently inhibited a-MSH-stimulated
cAMP release with an IC50 value of 130 nM.
the N-benzyl intermediate 10a (Ki = 140 nM) was 5-fold
less potent in binding affinity than the N-isopropyl ana-
log 9c. Incorporating a methoxyl group into the isopro-
pane in 9c reduced its affinity by 4-fold (12a,
Ki = 100 nM). While the cyclopentane 12b was not much
different from 9c, the cyclohexane 12c (Ki = 5.5 nM) was
significantly more potent in binding affinity than 9c.
Functionally, 12c behaved as an antagonist with an
IC50 of 130 nM. The amides 13a–e and 14a–d had lower
binding affinity than the isopropyl 9c. Similarly, urea 16
and sulfonamide 17 exhibited poor binding affinity (Ta-
ble 2). These results suggest that the basic nitrogen and a
small lipophilic group such as isopropyl are important
for high binding affinity of these compounds.
bulky group is unfavored at this position. The elec-
tron-withdrawing fluorine (18f) and electron-donating
methoxy group (18g) alone at the 3-position of the phe-
nyl ring had no effect on the potency compared to 9c,
while substitution at the 4-position of 9c altered the
binding affinity (compounds 18h–q). For example, the
4-methoxy compound 18j (3.6 nM) displayed a Ki value
7-fold more potent than 9c, while the 4-trifluoromethyl
analog 18k (130 nM) was 5-fold less potent. The 4-iso-
propoxy 18q (Ki = 2.1 nM) was over 10-fold more po-
tent than 9c. These results indicate a hydrogen-bond
donor at this position increases binding affinity. For
the disubstituted phenyl derivatives 18r–v, the 2-fluoro-
4-methoxy 18t (Ki = 1.8 nM) improved the affinity over
6-fold from the 2-fluoro 18a, while the 3-fluoro-4-meth-
oxy analog 18v (Ki = 0.5 nM) exhibited the best binding
affinity among these compounds.
Because the benzyl side chain (9c) on the core structure
6a provided great improvement in binding affinity, a
more detailed survey was conducted using various benz-
aldehydes via reductive alkylations, and the results are
depicted in Table 3. For the 2-substituted phenyl deriv-
atives 18a–e, the small fluorine (18a, Ki = 12 nM)
slightly increased potency compared to the unsubstitut-
ed parent 9c, while the affinity of the chloro-derivative
18b remained unchanged. The other analogs (18c–e) dis-
played less potent affinity compared to 9c, suggesting a
Functionally, none of these substituted benzylamines
18a–v tested in the agonist assay showed high potency
or high efficacy. For example, compound 18a exhibited
an EC50 of 580 nM with an IA of 48%, indicative of a
moderately active partial agonist. Compound 18v had
an IA value of only 10% and functioned as a
potent antagonist with an IC50 value of 48 nM. The

6830
C. W. Chen et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6825–6831
Table 3. SAR of substituted benzylamines 18a–v at MC4R
Table 4. SAR of tertiary amines 19–20 and amides 21–22 at MC4R
Cl
Cl
N
N
N
N
NH
X
N
N
N
R6
O
O
19, 21: A = COMe
20, 22: A = N
18a-v
A
Compound
R6
Ki (nM)
Compound
X
H
Ki (nM)
EC50 (nM)a
18j
19a
19b
19c
19d
21
H
Me
3.6
110
9c
26
12
1100 (31%)
580 (48%)
1300 (75%)
1000 (48%)
18a
18b
18c
18d
18e
18f
18g
18h
18i
2-F
2-Cl
Et
i-Bu
190
25
62
1800
2800
1300
27
2-MeO
2-CF3
4-MeOC6H4CH2
140
110
24
MeCO
H
Me
2-CF3O
3-F
9p
(23%)
20a
20b
20c
22
750
3-MeO
4-F
4-Cl
30
44
1200 (39%)
6600 (61%)
4200 (29%)
(17%)
Et
i-Bu
MeCO
1300
2700
1500
90
18j
4-MeO
4-CF3
3.6
130
52
18k
18l
(16%)
4-NO2
4-AcNH
4-Me2N
4-COOMe
4-MeS
4-i-PrO
2,4-F
18m
18n
18o
18p
18q
18r
18s
18t
18u
18v
29
65
190 (27%)
1100 (16%)
(21%)
Compared to the dipeptide 1a, the pyrrolidine 1b is a
more compact molecule with fewer peptide features yet
still possesses potent MC4R agonist activity. We have
successfully converted the benzylamine antagonists such
as 2 into potent MC4R agonists such as 3 by incorporat-
ing the N-alkylpyrrolidine moiety. Compound 4 is a
functional agonist, but replacing its Tic-(4-Cl)Phe
with 1-isopropyl-4-(chlorophenyl)pyrrolidine-3-carbonyl
moiety results in a compound (7a) with low binding
affinity. The basic amine on the left-side was required
for the current series to have high binding affinity, con-
trary to the SAR in the b-Ala-(2,4-Cl)Phe dipeptides 5.
In addition, functional antagonists were identified.
These results demonstrate that the current series of com-
pounds has unique SAR. Previous studies indicated that
the basic nitrogen of the Tic-group might interact with
the Asp-126 residue of the human MC4 receptor,14
which is believed to be important for receptor activa-
tion.15 The requirement of the basic amine at the left-
side of current compounds might suggest that it inter-
acts with this residue, resulting in a binding mode that
is different from other series such as 1b and 3.
32
27
2.1
11
(6%)
1300 (27%)
(29%)
2,4-MeO
2-F,4-MeO
3,4-OCH2O
3-F,4-MeO
16
1.8
8.3
0.5b
1600 (26%)
(7%)
170 (10%)
a Data are average of two or more independent measurements; intrinsic
activity is indicated in parentheses.
b Compound 18v dose-dependently inhibited a-MSH-stimulated
cAMP release with an IC50 value of 48 nM.
discrepancy between the binding affinity and functional
activity of 18v is probably due todifferent assay conditions.
Compounds 18j and 9p were further derivatized to the
tertiary amines 19 and 20, respectively. A simple methyl-
ation of 18j reduced its affinity by 30-fold (19a,
Ki = 110 nM), while larger alkyl groups (19b–d) dis-
played further reduced affinity, suggesting the secondary
amine is optimal. Acetylation of 18j resulted in com-
pound 21 (Ki = 1300 nM) with poor binding affinity,
indicating the importance of the basic nitrogen of 18.
Similar results were also obtained for the pyridine ana-
logs of 9p (Table 4).
In conclusion, a series of trans-N-alkyl-4-(4-chloro-
phenyl)pyrrolidine-3-carboxamides of piperazinecyclo-
hexanemethylamines were synthesized and characterized
at the melanocortin-4 receptor. While the trans-4-aryl-
pyrrolidine-carbonyl group has been successfully used
in other templates to replace the Tic-(4-Cl)Phe dipeptide
for small molecule agonists such as 1b and 3, no potent
agonists were discovered in this study. It was found that
the basicity of the cyclohexanemethylamine was
essential for potent binding affinity, and several potent
benzylamine derivatives were identified with single digit
Ki values. Compound 18v was found to bind MC4R
with potent affinity (Ki = 0.5 nM) and high selectivity
Compound 18v was tested for affinity at the other mela-
nocortin receptor subtypes and was found to be highly
selective for MC4R. Thus, 18v bound to the MC1R,
MC3R, and MC5R with Ki values of 3,900, 760, and
540 nM, respectively, demonstrating 1000-fold selectiv-
ity at the melanocortin-4 receptor. Compound 18v also
bound to the mouse MC4R with a Ki of 0.6 nM, indicat-
ing no species difference for this compound.

C. W. Chen et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6825–6831
6831
9. Pontillo, J.; Tran, J. A.; White, N. S.; Arellano, M.; Fleck,
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over the other melanocortin subtypes and behaved as a
functional antagonist (IC50 = 48 nM).
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