Discovery and SAR of 4-amino-2-biarylbutylurea MCH 1 receptor antagonists through solid-phase parallel synthesis

Article abstract of DOI:10.1016/j.bmcl.2005.05.039

-4-Amino-2-arylbutylbenzamides such as 1 were identified as micromolar MCH 1 receptor (MCH1R) antagonists via screening using a scintillation proximity assay based on [¹²⁵I]-MCH binding to recombinant, human MCH1R. Subsequent lead optimization

Full text of DOI:10.1016/j.bmcl.2005.05.039

Bioorganic & Medicinal Chemistry Letters 15 (2005) 3691–3695
Discovery and SAR of 4-amino-2-biarylbutylurea MCH 1
receptor antagonists through solid-phase parallel synthesis
Tao Guo,* Rachael C. Hunter, Huizhong Gu, Laura L. Rokosz,
Tara M. Stauffer and Doug W. Hobbs
Pharmacopeia Drug Discovery, Inc., P.O. Box 5350, Princeton, NJ 08543-5350, USA
Received 1 March 2005; revised 25 April 2005; accepted 11 May 2005
Available online 13 June 2005
Abstract—-4-Amino-2-arylbutylbenzamides such as 1 were identified as micromolar MCH 1 receptor (MCH1R) antagonists via
screening using a scintillation proximity assay based on [125I]-MCH binding to recombinant, human MCH1R. Subsequent lead
optimization efforts using solid-phase parallel synthesis resulted in the defined structure–activity relationships and the identification
of 4-amino-2-biarylbutylureas, such as 11g, as potent single digit nanomolar MCH1R antagonists.
Ó 2005 Elsevier Ltd. All rights reserved.
Melanin-concentrating hormone (MCH) is a cyclic
19-amino-acid neuropeptide expressed broadly through-
out the brain, with the highest concentration located in
the lateral hypothalamus and zona incerta regions.1a It
has been demonstrated that MCH plays an important
role in the central regulation of food intake and energy
homeostasis through the following studies: central
administration of MCH in mice stimulates food intake
while fasting results in an increase in MCH expression;1b
MCH knockout mice are hypophagic and leaner than
wild-type mice but otherwise healthy;1c and transgenic
mice over-expressing MCH are susceptible to obesity
and insulin resistance.1d
However, the physiological function of MCH2R has not
been established.
Based on the evidence implicating MCH1R in the con-
trol of feeding behavior and energy expenditure, the
development of potent and selective MCH1R antago-
nists as new therapeutics for the treatment of obesity
has become an attractive field of research. A number of
structurally diverse and small molecule MCH1R antago-
nists have emerged over the past few years.5,6 Some of
these antagonists have been further demonstrated to
show in vivo efficacy in several animal models of body
weight regulation and feeding behavior.6a,6b,6m,6n In this
article, we describe our efforts in the discovery of novel 4-
amino-2-biarylbutylureas as highly potent MCH1R
antagonists by using solid-phase parallel synthesis.
MCH binds and activates two distinct receptors in the
brain, MCH1R and MCH2R, both of which belong to
the rhodopsin superfamily of G-protein-coupled recep-
tors (GPCRs), but which have low (38%) sequence
homology to one another.2,3 MCH1R is present in all
mammals whereas MCH2R is found only in ferrets,
dogs, rhesus monkeys, and humans but not in rodents
and lagomorphs. More recently, it has been demonstrat-
ed that MCH1R knockout mice are lean, hyperphagic
but hyperactive and resistant to diet-induced obesity,
strongly implicating MCH1R in playing a critical role
in the regulation of food intake and energy homeostasis.4
Initial screening for MCH1R antagonists, using a
scintillation proximity assay (SPA) that is based on
Keywords: MCH; MCH1R antagonists; Obesity; Solid-phase parallel
synthesis.
*
Corresponding author. Tel: +1 609 452 3746; fax: +1 609 452 3699;
e-mail: tguo@pharmacop.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.05.039

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T. Guo et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3691–3695
Scheme 1. Solid-phase synthesis of parallel optimization libraries 8 and 11. Reagents and conditions: (a) LDA, allyliodide, THF, ꢁ78 °C; (b)
LiAlH4, H2SO4, THF; (c) Na(OAc)3BH, ClCH2CH2Cl; (d) R2COCl (or R2NCO, R2OCOCl, R2SO2Cl), pyridine, CH2Cl2; (e) OsO4, NMMO,
0
acetone, H2O; (f) NaIO4, acetone, H2O; (g) R3R3 NH, Na(OAc)3BH, ClCH2CH2Cl; (h) TFA, CH2Cl2; (i) Ar1B(OH)2, Pd(PPh3)4, K2CO3, DMF,
70 °C.
[
125I]-MCH binding to membranes prepared from CHO
generated secondary amine 5. Treatment of 5 with vari-
ous reagents such as acid chlorides, sulfonyl chlorides,
isocyanates, and chloroformates produced 6 as amides,
sulfonamides, ureas, and carbamates, respectively. Fi-
nally, olefin 6 was treated with OsO4 and NMMO fol-
lowed by NaIO4 to generate aldehyde 7, which was
reductively alkylated with various primary and second-
ary amines to provide the aryl library 8 after TFA-med-
iated cleavage from the resin. When X = Br or I,
treatment of 6 with various arylboronic acids under
standard Suzuki coupling conditions provided biaryl
olefin 9. Subsequently, olefin 9 was treated with OsO4
and NMMO followed by NaIO4 to generate aldehyde
10, which was reductively alkylated with various pri-
mary and secondary amines to provide the biaryl library
11 after TFA-mediated cleavage from the resin.
cells that express human MCH1R, resulted in the iden-
tification of a series of 4-amino-2-(3,4-dichlorophenyl)-
butylbenzamides such as 1 (MCH1R, Ki = 4.1 lM) with
potencies in the micromolar range.
To explore the SAR around compound 1 and to opti-
mize its potency, parallel libraries 8 and 11 (Scheme 1)
were designed to allow systematic variations of the three
regions of compound 1: variation of the 3,4-dichlor-
ophenyl group in the center with various aryl and biaryl
groups; variation of the 3,5-dichlorobenzamide substitu-
ent on the left-hand side with diverse amides, sulfona-
mides, ureas, and carbamates; and variation of the
4-piperidylpiperidyl unit on the right-hand side with
various secondary and tertiary amines.
Scheme 1 shows the synthesis of libraries 8 and 11. First,
the requisite 2-aryl-pent-4-enylamine scaffold 3 was pre-
pared via treatment of various commercially available
arylacetonitriles 2 with LDA and allyliodide followed
by reduction using LiAlH4. Overall, 15 scaffolds were
prepared including, for example, 2-(3,4-dichloro-
phenyl)-pent-4-eneamine and 2-(4-iodophenyl)-pent-4-
eneamine. Next, reductive alkylation of resin-bound
aldehyde linker 4 (which was readily prepared via
coupling of the acid labile linker 4-(4-formyl-3-meth-
oxy-phenoxy)-butyric acid with the amine resin Argo-
Gel-NH2 using DIC and HOBt) with scaffold 3
Overall, more than 500 compounds were prepared. Each
compound (ꢀ5 mg) was purified by HPLC prior to
biological evaluation.7
Table 1 highlights the SAR from library 8. First, there is
a dramatic enhancement in potency (>10-fold) when the
3,5-dichlorobenzamide group in 1 is replaced with vari-
ous dihalo substituted phenylureas. As can be seen, 3,5-
dichlorophenylurea (8a), 3,4-dichlorophenylurea (8b),
3,4-difluorophenylurea (8c), and 3-chloro-4-fluoro-phe-
nylurea (8d) display MCH1R Kis ranging from 100 to
300 nM. However, the unsubstituted phenylurea (8e)

T. Guo et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3691–3695
Table 1. Key SAR from parallel library 8
3693
donating groups. For example, 3-methoxyphenylurea
(8f) and 3-toluylurea (8g) are 7-fold and 4-fold less po-
tent, respectively, than the 3-chlorophenylurea (8h).
Also, meta-substituents are preferred over para-substitu-
ents, and ortho substitution is detrimental. For example,
in the mono-CF3-substituted series, the potency rank is
as follows: meta (8i) > para (8j) > ortho (8k). However,
the 3,5-di-CF3-substituted analog 8 is 6-fold less active
than 8i. Finally, aromatic carbamates and sulfonamides
did not show any potency enhancement as compared to
amide 1 (data not shown).
Compound
R2
MCH1R, Ki (nM)7
8a
8b
8c
8d
8e
8f
3,5-Cl2C6H3
3,4-Cl2C6H3
3,4-F2C6H3
3-Cl-4-FC6H3
C6H5
135
300
Table 2 summarizes the SAR from parallel library 11.
Only para-substituted biaryl analogs from library 11
are listed in the table since meta- or ortho-linked biraryl
analogs were all >10-fold less active than the corre-
sponding para-biaryl compounds (data not shown). As
is evident from Table 2, installing an aryl group at the
para-position can further enhance the potency, in many
cases yielding potent low nanomolar compounds. Com-
pounds 11a–j highlight the SAR at the Ar position in
this series. Substitution at the 3-position of the distal
phenyl ring with a chloro group increased the potency
by 2-fold (11b, Ki = 26 nM) relative to the unsubstituted
compound (11a, Ki = 50 nM). Substitution at the
2-position (11c) or the 4-position (11d) both resulted
in a decrease in potency relative to 11a. Similar to the
3-chloro compound, the 3-fluoro analog 11e, and the
240
100
18,500
11,000
6800
1641
205
3-MeOC6H4
3-MeC6H4
3-ClC6H4
8g
8h
8i
3-CF3C6H4
4-CF3C6H4
2-CF3C6H4
3,5-(CF3)2C6H3
8j
1080
22,500
1223
8k
8l
has a 4-fold drop in potency (Ki = 18,500 nM) relative to
1. Second, phenylureas appended with some electron
withdrawing groups are more potent than with electron
Table 2. Key SAR from parallel library 11
0
Compound
Ar
R2
NR3R3
MCH1R Ki (nM)7
11a
11b
11c
C6H5
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
3,5-Cl2C6H3
C6H5
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
cyclo-Pentylamino
Methylamino
50
26
3-ClC6H4
2-ClC6H4
4-ClC6H4
3-FC6H4
130
280
37
11d
11e
11f
3-CF3OC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
4-NCC6H4
3-pyridyl
20
11g
(+)-11g
(ꢁ)-11g
11h
11i
3.0
3.0
4.0
46
24
11j
4-pyridyl
122
2.6
4.7
4.3
5.6
5.4
2.4
1.1
9.2
1.2
7.4
0.98
0.84
0.88
11k
11l
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
3-NCC6H4
Dimethylamino
Ethylamino
11m
11n
11o
11p
11q
11r
Isopropylamino
Piperidin-1-yl
Dimethylamino
Dimethylamino
Dimethylamino
Dimethylamino
Dimethylamino
Dimethylamino
Dimethylamino
Dimethylamino
3-ClC6H4
4-ClC6H4
11s
11t
3-FC6H4
4-FC6H4
11u
11v
3-Cl-4-FC6H3
3,4-F2C6H3
3,5-F2C6H3
11w

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T. Guo et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3691–3695
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11a. To our delight, when a cyano group is introduced
at the 3-position (11g, Ki = 3.0 nM), a dramatic 17-fold
increase in potency relative to that of 11a was achieved.
However, the 4-cyano analog (11h) showed no improve-
ment in potency as compared to 11a. Furthermore, the
3-pyridyl compound 11i is 2-fold more potent than the
benzene analog 11a, while the 4-pyridyl compound 11j
is 2-fold less potent than 11a.
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position. A wide variety of acyclic and cyclic amines
are explored—methylamino, dimethylamino, ethyl
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Compounds 11p–w were made to probe the SAR at the
R2 position. As is shown, the potency for this series of
compounds is relatively insensitive to changes in the
R2 aryl group. Even the simple phenyl urea 11p exhib-
ited potency (Ki = 2.4 nM) as good as the substituted
phenyl ureas (11q–w). However, in the mono-substituted
cases, the 4-chloro analog 11q and 4-fluoro analog 11s
exhibited 6- to 8-fold decreases in potency relative to
the corresponding 3-chloro (11r) and 3-fluoro (11t)
compounds. Interestingly, the 3,4-disubstituted analogs
11u–w displayed slightly greater potency than the
mono-substituted ones (11p–r).
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It should be noted that all the compounds tested are race-
mic mixtures except that in the case of 11g, its two enan-
tiomers, (+)- and (ꢁ)-11g, resolved via chiral HPLC
(Chiracel AD column), were also evaluated. Interestingly,
both enantiomers showed almost identical activity in the
MCH1R binding assay: (+)-11g Ki = 3.0 nM; and (ꢁ)-
11g Ki = 4.0 nM. The two enantiomers were further tested
in aCa2+ mobilization FLIPRÒ assay8 and both displayed
identical and potent functional antagonism of MCH1R:
(+)-11g Kb = 1.0 nM; and (ꢁ)-11g Kb = 1.0 nM.
In summary, starting from micromolar 4-amino-2-aryl-
butylbenzamide MCH1R receptor antagonists identified
from screening, solid-phase parallel synthesis of optimi-
zation libraries resulted in the discovery of 4-amino-2-
biarylbutylureas, such as 11g, as potent single digit
nanomolar MCH1R antagonists along with defined
SAR.
Acknowledgment
We thank Dr. Brian E. Hawes of Schering-Plough Re-
search Institute for obtaining the Kb data for (+)- and
(ꢁ)-11g.
References and notes
1. (a) Bittencourt, J. C.; Presse, F.; Arias, C.; Peto, C.;
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3695
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competition binding model for IC50 determination using the
program GraphPad Prism (GraphPad Software, Inc., San
Diego, CA) and Ki values were calculated using the Cheng–
Prusoff equation. All Kis represent the average of two or
more determinations. The standard deviations were no
greater than 30% from the mean; (b) Cheng, Y.; Prusoff, W.
H. Biochem. Pharmacol. 1973, 22, 3099.
7. (a) MCH1R Kis were determined as follows: membranes
prepared from CHO cells that express human MCH1R
(0.1 mg/mL final) were incubated with wheatgerm-aggluti-
nin (WGA) coated SPA beads (1 mg/mL final, Amersham
Biosciences, Piscataway, NJ) in assay buffer (25 mM
HEPES, 10 mM NaCl, 10 mM MgCl2, 5 mM MnCl2,
0.1% BSA, pH 7.4) for 5 min on ice and subsequently
washed two times in assay buffer. The test compound (5 ll)
prepared in DMSO, DMSO (vehicle, 5%) or MCH (2.5 lM
final—used to quantify the non-specific signal) was mixed
with 45 ll assay buffer in 96-well assay plates (Corning
#3604). The bead/membrane mixture (100 ll) was added to
the compounds followed by 50 ll of [125I]-MCH (0.5 nM
final, Perkin-Elmer, Boston, MA). The assay plates were
shaken for 5 min on a plate shaker and then incubated for
2 h. Binding of [125I]-MCH to the bead/membrane mixture
was detected using a Microbeta TriluxTM scintillation
counter (Perkin-Elmer). The data were fit to a one-site
8. Calcium mobilization assay: MCH1R/CHO cells were
incubated with 4 lM Fluoro-3-AM/0.04% pluronic acid in
assay buffer (HanksÕ BSS, 0.1% BSA, 20 mM HEPES,
2.5 mM probencid) for 1 h at 37 °C. The cells were
centrifuged for 7 min at 1000g and the pellet was resus-
pended in assay buffer to a density of 500 cells/ll. The cells
(40 ll) were plated at 20,000 cells per well into 96-well BD
Biocoat poly-D-lysine black/clear plates, centrifuged for
3 min at 1000g and incubated at room temperature for
30 min. Compounds or 2% DMSO (vehicle) were added to
the cells and incubated for 10–15 min at room temperature.
The cells were stimulated with 100 nM MCH and the
calcium response was monitored for 1 min per well in a
FLIPRÒ system from Molecular Devices (Sunnyvale, CA).
Kb values were determined as described for Ki in Ref. 7a.
All KbS represent the average of two or more determina-
tions, and the standard deviations were no greater than 30%
from the mean.