Discovery of potent, selective, and orally bioavailable 3H-spiro[isobenzofuran-1,4′-piperidine] based melanocortin subtype-4 receptor agonists

Article abstract of DOI:10.1016/j.bmcl.2010.06.068

Design, synthesis, and SAR of a series of 3H-spiro[isobenzofuran-1, 4′-piperidine] based compounds as potent, selective and orally bioavailable melanocortin subtype-4 receptor (MC4R) agonists are disclosed.

Full text of DOI:10.1016/j.bmcl.2010.06.068

Bioorganic & Medicinal Chemistry Letters 20 (2010) 4895–4900
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of potent, selective, and orally bioavailable
3H-spiro[isobenzofuran-1,40-piperidine] based melanocortin
subtype-4 receptor agonists
a
a
a
a
a
Liangqin Guo a, , Zhixiong Ye , Jian Liu , Shuwen He , Raman K. Bakshi , Iyassu K. Sebhat ,
Peter H. Dobbelaar a, Qingmei Hong a, Tianying Jian a, James P. Dellureficio a, Nancy N. Tsou a,
Richard G. Ball a, David H. Weinberg b, Tanya MacNeil b, Rui Tang b, Constantin Tamvakopoulos d,
Qianping Peng d, Howard Y. Chen c, Airu S. Chen c, William J. Martin b, D. Euan MacIntyre b,
Alison M. Strack b, Tung M. Fong c, Matthew J. Wyvratt a, Ravi P. Nargund a
*
a Department of Medicinal Chemistry, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065, USA
b Department of Pharmacology, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065, USA
c Department of Obesity Research, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065, USA
d Department of Drug Metabolism and Pharmacokinetics, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065, USA
a r t i c l e i n f o
a b s t r a c t
Design, synthesis, and SAR of a series of 3H-spiro[isobenzofuran-1,40-piperidine] based compounds as
potent, selective and orally bioavailable melanocortin subtype-4 receptor (MC4R) agonists are disclosed.
Ó 2010 Elsevier Ltd. All rights reserved.
Article history:
Received 11 May 2010
Revised 10 June 2010
Accepted 11 June 2010
Available online 17 June 2010
Keywords:
Melanocortin subtype-4 receptor
MC4R agonists
Obesity
Oral bioavailability
Food intake
Spiro isobenzofuran
Over the past 20 years, obesity has become one of the fastest
growing diseases in the world. The medical problems associated
with obesity include increased mortality, hypertension, diabetes,
heart disease, and other substantial health risks. The current pre-
scription drug market for anti-obesity agents is small compared
to overall disease prevalence, so the unmet demand for more effec-
tive anti-obesity agents has prompted many pharmaceutical com-
panies and research groups to conduct research in this area.
The melanocortin receptors are a family of seven-transmem-
brane G-protein-coupled receptors. All the five known subtype
melanocortin receptors are activated by their endogenous ligands
(the corticotropins and melanocortins) to mediate a variety of
physiological functions, such as feeding behavior, sexual function,
skin pigmentation, steroidogenesis, and exocrine gland secretion.
The melanocortin subtype-4 receptor (MC4R), highly expressed
in the hypothalamus and brainstem, has been clearly linked to
the regulation of appetite, body weight, and energy homeostasis.1
Both rodent and human genomic studies indicate that inactivation
of MC4R results in obesity. In the past decade, considerable efforts
have been devoted in drug discovery to develop potent and selec-
tive non-peptide MC4R agonists for the potential treatment of hu-
man obesity.2 Previously, we disclosed the discovery of t-butyl
pyrrolidine derived, potent and orally bioavailable MC4R modula-
tors by modifying the 2,4-difluorophenyl in 1.3 Herein, we report
our continued effort in this area, which has led to the discovery
of a series of potent and orally bioavailable MC4R agonists contain-
ing a 3H-spiro[isobenzofuran-1,40-piperidine] moiety (Fig. 1). The
lead compound 2 is a potent, selective, orally bioavailable, and
brain penetrable MC4R agonist with good efficacy in lowering food
intake and body weight in rodents.
Our earlier efforts in modifying the piperidine part-structure
to enhance MC4R potency and oral bioavailability revealed a
4,4-disubstituted piperidine bearing a gem -dimethyl acetamide
moiety, exemplified by compound 3 (Fig. 2a).4a Aiming to further
* Corresponding author.
E-mail address: liangqin_guo@merck.com (L. Guo).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.06.068

4896
L. Guo et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4895–4900
O
Boc
N
Br
Br
a, b
c
N
N
OH
F
Cl
Cl
Cl
F
O
O
5
6
N
d
N
F
F
Boc
N
Boc
N
Boc
N
O
Cl
NHAc
e,f, g, h
O
CN
O
O
Cl
Cl
CN
Cl
Cl
CN
OH
2
1
8B
8A
7
O
N
Figure 1. MC4R selective agonists.
RCO2H;
i, j, k
R
O
N
R
N
Cl
F
F
F
N
F
O
O
F
N
F
O
OH
OH
O
11
Boc
N
F
F
9
N
N
AcHN
F
F
N
O
OH
F
1
N
F
O
Cl
R1
R2
CN
N
O
F
O
F
O
R3
OH
OH
12
10
2: R = 4-THP
R1 = Cl,
Scheme 1. Reagents and conditions: (a) NIS, TFA, RT, 2 h, 50 °C, 1 h; (b)
Bu3Sn(CH@CH2), Pd(dppf)2Cl2,LiCl, DMF, rt, 48 h; (c) nBuLi, N-Boc-4-oxopiperidine,
THF–ether, ꢀ78 °C to rt; (d) MCPBA, DCM, rt-reflux, O/N; (e) TsCl, DIEA, DMAP, rt,
overnight; (f) KCN, KI, DMSO, 110 °C, O/N; (g) NaHMDS, MeI, ꢀ78 °C to rt; (h) chiral
HPLC resolution: ChiralCel OD column; mobile phase = 0.5% IPA/heptane; flow
rate = 9 ml/min; (i) 4 N HCl, dioxane, rt, 1 h; (j) RCO2H (9–12), DIEA, HOAt, HATU,
DCM; (k) for RCO2H = 12: (i) 4 N HCl, dioxane, rt, 1 h; (ii) DIEA, molecular sieve,
aldehyde or ketone, Na(OAc)3BH, DCM, rt.
NHAc
3
R2 = Me
R3 = CN
N
N
4
NHAc
NHAc
moiety is summarized in Scheme 1. Iodination of 4-bromo-2-
chloro-toluene, followed by a Stille coupling reaction, gave vinyl
intermediate 5. Trans metalation of 5 and addition to N-Boc-4-
oxopiperidine yielded tertiary alcohol 6, which, upon treatment
with MCPBA, furnished the spiro isobenzofuran alcohol 7 via
epoxide formation and subsequent intramolecular substitution.
Tosylation of 7, followed by substitution with cyanide and then
methylation afforded a racemic gem-dimethyl nitrile, which under-
went chiral HPLC resolution to give the separated enantiomers 8A
and 8B.
Figure 2a. Design of spiro isobenzofuran piperidine based MC4R selective agonists.
O
O
O
O
O
N
N
The absolute stereochemistry of the faster eluting isomer 8A
was determined as ‘S’ by single-crystal X-ray diffraction analysis
(Fig. 3).5 Following deprotection, 8A was coupled to pyrrolidine
acids 9–11 to generate MC4R agonists 2, 20 and 23. Coupling to
12, followed by removal of Boc and reductive amination, afforded
a further series of MC4R agonists variously substituted on the
pyrrolidine ring (Scheme 1).
Amides analogs were generated from the nitrile of 8A via
hydrolysis to the de-protected amino acid followed by a one-pot,
two-step coupling procedure (Scheme 2a). Reversed amides and
a gem-dimethyl acetamide moiety were generated from intermedi-
ate 13 via Curtis rearrangement to give an isocyanate intermediate
and subsequent reaction with 2-(trimethylsilyl)ethanol affording
the protected amine 14. De-protection and acetylation gave acet-
amide 15. Removal of Boc followed by coupling to 11 afforded 16
(Scheme 2b).
cLogP = 4.28
cLogP = 3.38
Figure 2b. Calculated Log P.
improve pharmacokinetic properties, this was coupled with piper-
idine design in 1, leading to the cyclized analog 4.4 Attempts to
lower the Log P of the molecules in hope of attenuating plasma
protein binding and ultimately improving in vivo efficacy inspired
us to design the oxygen tethered spiro indane or spiro isobenzofu-
ran piperidine moiety (Fig. 2b), which in turn yielded a series of
interesting MC-4 agonists with good MC4R in vitro potency and
in vivo efficacy.
The general synthetic procedure for synthesizing 3H-spiro[iso-
benzofuran-1,40-piperidine] containing structures bearing a nitrile

L. Guo et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4895–4900
4897
Figure 3. Structure of 8A by single-crystal X-ray diffraction.
O
which was converted to the nitrile 17 via Michael addition using
acrylonitrile followed by selective protection of the primary alco-
hol using tert-butyl chlorodiphenylsilane. Activation of the benzyl
alcohol using diethyl phosphoryl chloride followed by treatment
with LiHMDS effected the intramolecular SN2 reaction to construct
the pyrrolidine ring (18) with inversion of the benzyl stereo center
from ‘S’ to ‘R’. Refluxing 18 under basic conditions epimerized the
N
F
O
Boc
N
N
a,b,c
stereo center at the
a-position of the nitrile group to ‘S’ favoring
F
the more stable trans conformation, and hydrolyzed the nitrile to
the carboxylic acid. The TBPS protecting group was also removed
in this step to afford 10.
O
O
O
CN
Cl
Cl
NRR'
Compound 11 was synthesized from 126b via esterification,
deprotection, reductive amidation, and hydrolysis (Scheme 4).
All final compounds were evaluated in MCR binding and
functional assays for initial SAR studies.7 Selected compounds were
also evaluated in pharmacokinetic studies in rats. As shown in
Table 1, compound 20A, derived from intermediate 8A, is signifi-
cantly more potent than 20B, derived from intermediate 8B, indi-
cating that ‘S’ configuration in the 3-position of the spiro
isobenzofuran is preferred for MC4R binding affinity and functional
activity.
8A
30-36
Scheme 2a. Reagents and conditions: (a) concd HCl, reflux; (b) 11, DIEA, HOAt,
HATU, DMF; (c) RR0NH.
Compound 10 was synthesized using a modified literature pro-
cedure (Scheme 3).6a Substitution of the chloride in (1S)-2-chloro-
1-(2,4-diflurophenyl)ethanol with the amine of 2-amino-2-
methyl-1-propanol resulted in a hydroxylamine intermediate,

4898
L. Guo et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4895–4900
Table 1
Boc
N
Boc
N
Effects of chirality of spiro isobenzofuran on binding affinity and functional activity at
the human MC4R7
a, b
O
Cl
O
Cl
CO2H
N
Cl
CN
F
13
N
8A
*
O
O
c, d
CN
F
Boc
Boc
N
N
Compd
*
MC4R binding IC50 (nM)
MC4R cAMP EC50 (nM) [%Max]a
g, h
e, f
20A
20B
‘S’
‘R’
0.9
31.4
1.1 [122]
195.6 [127]
Si
16
O
O
H
N
a
Cl
Cl
Percentage of cAMP accumulation at 10 lM relative to a-MSH.
NHAc
O
O
15
14
Table 2
Scheme 2b. Reagents and conditions: (a) concd HCl, reflux; (b) (Boc)2O, NaOH,
dioxane–H2O; (c) DPPA, Et3N, toluene, reflux; (d) HO(CH2)2TMS, reflux; (e) TBAF,
THF, rt, O/N; (f) Ac2O, pyridine, CH2Cl2, rt, overnight; (g) HCl in dioxane, rt, 1 h; (h)
11, DIEA, HOAt, HATU, DCM.
Effects of N-substitutions of pyrrolidine part in MC-4 agonists on oral bioavailability,
MC4R activity and selectivity at the human melanocortin receptors7
Cl
R
N
CN
OH
O
N
OH
F
Cl
a, b, c
N
CN
F
OTBPS
O
F
F
F
17
F
d, e
Compd
R
MC4R binding IC50 (nM)
EC50 (nM)
[%Max]
F%a
OH
N
f, g, h
OTBPS
F
MC4R
MC1b
F
N
21
22
Boc
H
479.2
15.0
1006.0
[68%]
78.1
—
[3]
2100
[36]
ndb
nd
O
OH
NC
[99%]
F
18
10
1.1
115
20A
23
0.9
2.8
32
44
F
[122%]
[50%]
2.6
342.9
[77]
Scheme 3. Reagents and conditions: (a) 2-amino-2-methylpropan-1-ol, NaOH,
MeOH; (b) acrylonitrile, 80 °C, overnight; (c) DIEA, DMAP, Me3Si(Ph)2Cl; (d)
(EtO)2P(O)Cl; (e) LiHMDS; (f) NaOH, EtOH–H2O, reflux; (g) SOCl2, MeOH, reflux,
1 h; (h) NaOH, THF–H2O, rt
[112%]
OH
2.7
1697
[44]
2
2.0
64
nd
[106%]
O
O
95.9
[66%]
735.0
[38]
24
52.5
HHCl
N
O
F
a
9.8
[105]
295.0
[55]
b, c
12
25
26
27
6.6
nd
nd
nd
N
O
F
O
33.6
[115]
465
[63]
11.9
44.3
O
F
19
OH
73.2
[95]
615
[36]
O
11
F
a
Compounds were dosed in Sprague-Dawley rats as a solution in EtOH/PEG/
Scheme 4. Reagents and conditions: (a) SOCl2, MeOH, reflux, 2 h; (b) tetrahydro-
saline (1:4:5) at 1 mg/kg, iv (n = 2) and 4 mg/kg, po (n = 3).
4H-pyran-4-one, DIEA, 4 Å molecular sieve, CH2Cl2, rt, O/N; (c) NaOH, THF–H2O, rt,
4 h.
b
nd = not determined.
Compound 20a showed potent MC4R binding and functional
activity and good oral bioavailability in rats (F = 32%), however, it
had modest MC4R selectivity against MC1b (ꢁ100-fold). The
effects of modifications to the N-substitution of the pyrrolidine of
20A are highlighted in Table 2. Compound 2, containing an N-
THP pyrrolidine was revealed as a potent MC4R full agonist with
much improved selectivity against MC1b (>600-fold) and good
PK profiles in rats (F = 64%, AUCN = 0.44 lM h, CLp = 42.9 ml/min/

L. Guo et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4895–4900
4899
Table 3
Effects of substitutions at benzene ring of benzofuran on oral bioavailability and binding affinity and functional activity at the human MC4R7
X
Y
O
N
O
N
CN
F
O
F
Compd
X, Y
MC4R binding IC50 (nM)
MC4R cAMP EC50 (nM) [%Max]
F%a
cLog P
2
28
29
Me, Cl
F, F
Cl, F
2.0
7.6
7.3
2.7 [106]
8.4 [99]
17.1 [109]
64
7
23
6.9
5.9
6.6
a
Compounds were dosed in Sprague-Dawley rats as a solution in EtOH/PEG/saline (1:4:5) at 1 mg/kg, iv (n = 2) and 4 mg/kg, po (n = 3).
Table 4
Effect of substitution at benzene ring of isobenzofuran on binding affinity and functional activity at the human MC4R7
O
Cl
N
F
N
*
O
O
R
F
Compda
R
MC4R binding IC50 (nM)
MC4R cAMP EC50 (nM) [%Max]
F%
16A
16Bb
30
31
32
NHAc
NHAc
NMe2
NHMe
NHEt
6.2
150.7
2.4
3.6
2.3
7.4 [99]
26
nd
nd
46
nd
351.7 [92]
1.5 [101]
3.8 [101]
3.3 [100]
33
34
1.0
1.6
0.6 [107]
3.1 [109]
nd
nd
N
O
N
N
35
36
1.4
2.4
0.8 [107]
2.9 [100]
nd
nd
OH
F
N
F
a
All the compounds, except 16B, were derived from intermediate 8A, and hold ‘S’ configuration.
Compound 16B, derived from 8B, holds ‘R’ configuration.
b
Table 5
MCR activity profiles of 2
Table 6
Brain penetration study of 2
Receptor
Binding IC50 (nM)
cAMP EC50 (nM)
% Acti. at 10 l
Ma
Time point
Plasma (lM)
Brain (lM)
b/p ratio
hMC1b
hMC3
hMC4
hMC5
rMC4
1523
482.7
2.0
122.7
nd
1697
766.7
2.7
—
1.1
37
82
106
1
101
105
100
15 min
1 h
4 h
0.425
0.352
0.139
1.212
0.666
0.299
2.848
1.890
2.151
f MC4
mMC4
nd
nd
3.1
3.4
tion of the hydroxyl functionality affords a number of benefits:
lipophilicity is lowered; and the structure holds potential in
addressing a metabolism issue associated with other compounds
bearing an N-tert-butyl substituent.8
Attempts were also made to vary the substituents on the iso-
benzofuran benzene ring with a goal of further lowering the polar-
a
Percentage of cAMP accumulation at 10 lM relative to a-MSH.
kg, t1/2 = 5.4 h). Another interesting finding from this study was
that, incorporating N-tert-butyl hydroxyl pyrrolidine (23), main-
tained similar MC4R potency and rat PK profile to 20A. Incorpora-

4900
L. Guo et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4895–4900
agonist with good efficacy of lowering food intake and body weight
in DIO mice and rats.
WT-veh
p=0.000 (24%)
WT-2, 10mpk
KO-veh
4
3
2
1
0
KO-2, 10mpk
*
References and notes
p=0.000 (43%)
**
1. (a) Emmerson, P. J.; Fisher, M. J.; Yan, L. Z.; Mayer, J. P. Curr. Top. Med. Chem.
2007, 7, 1121; (b) Nargund, R. P.; Strack, A. M.; Fong, T. M. J. Med. Chem. 2006, 49,
4035; (c) Zimanyi, I. A.; Pelleymounter, M. A. Curr. Pharm. Des. 2003, 9, 627; (d)
Van der Ploeg, L. H. T.; Howard, A. D.; Nargund, R. P.; Austin, C. P.; Guan, X.;
Drisko, J.; Cashen, D.; Sebhat, I.; Pattchett, A. A.; Figueroa, D. J.; DiLella, A. G.;
Connolly, D. H.; Weinberg, D. H.; Tan, C. T.; Palyha, O. C.; Pong, S.; MacNeil, T.;
Rosenblum, C.; Vongs, A.; Tang, R.; Yu, H.; Sailer, A. W.; Fong, T. M.; Huang, C.;
Tota, M.; Chang, R. S.; Sterns, R.; Tamvakopoulos, T.; Christ, G.; Drazen, D. L.;
Spar, B. D.; Nelson, R. J.; Maclntyre, D. E. Proc. Natl. Acad. Sci. U.S.A. 2002, 99,
11381; (e) Eberle, A. N. In The Melanocortin Receptors; Cone, R. D., Ed.; Humana
Press, 2000. p 3; (f) Gu, W.; Tu, Z.; Kleyn, P. W.; Kissebah, A.; Duprat, L.; Lee, J.;
Chin, W.; Maruti, S.; Deng, N.; Fisher, S. L.; Franco, L. S.; Burn, P.; Yagaloff, K. A.;
Nathan, J.; Heymsfield, S.; Albu, J.; Pi-Sunyer, F. X.; Allison, D. B. Diabetes 1999,
48, 635; (g) Hinney, A.; Schmidt, A.; Nottebom, K.; Heibult, O.; Becker, I.; Ziegler,
A.; Gerber, G.; Sina, M.; Gorg, T.; Mayer, H.; Hebebrand, J. J. Clin. Endocrinol.
Metab. 1999, 84, 1483; (h) Huszar, D.; Lynch, C. A.; Fairchild-Huntress, V.;
Dunmore, J. H.; Fang, Q.; Berkemeier, L. R.; Gu, W.; Kesterson, R. A.; Boston, B. A.;
Cone, R. D.; Smith, F. J.; Campfield, L. A.; Burn, P.; Lee, F. Cell 1997, 88, 131; (i)
Fan, W.; Boston, B. A.; Kesterson, R. A.; Hruby, V. J.; Cone, R. D. Nature 1997, 385,
165.
4-hr FI
18-hr FI
0.75
0.5
0.25
0
1
WT-veh
-0.25
-0.5
-0.75
-1
WT-2, 10mpk
KO-veh
**
KO-2, 10mpk
***
2. (a) Ujjainwalla, F.; Sebhat, I. K. Curr. Top. Med. Chem. 2007, 7, 1068; (b) Sebhat, I.
K.; Lai, Y.; Barakat, K.; Ye, Z.; Tang, R.; Kalyani, R. N.; Vongs, A.; MacNeil, T.;
Weinberg, D. H.; Cabello, M. A.; Maroto, M.; Teran, A.; Fong, T. M.; Van der Ploeg,
L. H. T.; Patchett, A. A.; Nargund, R. P. Bioorg. Med. Chem. Lett. 2007, 17, 5720; (c)
Ye, Z.; Guo, L.; Barakat, K. J.; Pollard, P. G.; Palucki, B. L.; Sebhat, I. K.; Bakshi, R.
K.; Tang, R.; Kalyani, R. N.; Vongs, A.; Chen, A. S.; Chen, H. Y.; Rosenblum, C. I.;
MacNeil, T.; Weinberg, D. H.; Peng, Q.; Tamvakopoulos, C.; Miller, R. R.; Stearns,
R. A.; Cashen, D. E.; Martin, W. J.; Metzger, J. M.; Strack, A. M.; MacIntyre, D. E.;
Van der Ploeg, L. H. T.; Patchett, A. A.; Wyvratt, M. J.; Nargund, R. P. Bioorg. Med.
Chem. Lett. 2005, 15, 3501; (d) Ujjainwalla, F.; Warner, D.; Snedden, C.; Grisson,
R. D.; Walsh, T. F.; Wyvratt, M. J.; Kalyani, R. N.; Mac Neil, T.; Tang, R.; Weinberg,
D. H.; Van der Ploeg, L.; Goulet, M. T. Bioorg. Med. Chem. Lett. 2005, 15, 4023; (e)
Pan, K.; Scott, M. K.; Lee, D. H. S.; Fitzpatrick, L. J.; Crooke, J. J.; Rivero, R. A.;
Rosenthal, D. I.; Vaidya, A. H.; Zhao, B.; Reitz, A. B. Bioorg. Med. Chem. 2003, 11,
185; (f) Sebhat, I. K.; Martin, W. J.; Ye, Z.; Barakat, K.; Mosley, R. T.; Johnston, D.
B. R.; Bakshi, R.; Palucki, B.; Weinberg, D. H.; MacNeil, T.; Kalyani, R. N.; Tang, R.;
Sterns, R. A.; Miller, R.; Tamvakopoulos, C.; Strack, A.; McGowan, E.; Cashen, D.
E.; Drisko, J. E.; Hom, G. J.; Howard, A. D.; MacIntyre, D. E.; Van der Ploeg, L. H. T.;
Patchett, A. A.; Nargund, R. A. J. Med. Chem. 2002, 45, 4589.
3. (a) Guo, L.; Ye, Z.; Ujjainwalla, F.; Sings, H. L.; Sebhat, I. K.; Huber, J.; Weinberg,
D. H.; Tang, R.; MacNeil, T.; Tamvakopoulos, C.; Peng, Q.; MacIntyre, E.; van der
Ploeg, L. H. T.; Goulet, M. T.; Wyvratt, M. J.; Nargund, R. P. Bioorg. Med. Chem. Lett.
2008, 18, 3242; (b) Ujjainwalla, F. Presented at the 230th National Meeting of
American Chemical Society, Washington, DC, August 2005; paper MEDI-275.
4. (a) Guo, L. et al. unpublished results.; (b) He, S.; Ye, Z.; Dobbelaar, P. H.; Sebhat, I.
K.; Guo, L.; Liu, J.; Jian, T.; Lai, Y.; Franklin, C. L.; Bakshi, B. K.; Dellureficio, J. P.;
Hong, Q.; Tsou, N. N.; Ball, R. G.; Sashen, E. E.; Martin, W. J.; Weinberg, D. H.;
MacNeil, T.; Tang, T.; Tamvakopoulos, C.; Peng, Q.; Miller, R. R.; Stearns, R. A.;
Chen, H. Y.; Chen, A. S.; Strack, A. M.; Fong, T. M.; MacIntyre, D. E.; Wyvratt, M. J.;
Nargund, R. P. Bioorg. Med. Chem. Lett. 2010, 20, 2106.
Figure 4. Effects of 2 on FI and BW in MC3/4R KO and WT DIO mice.
ity (lower calculated Log P) and/or increasing the metabolic stabil-
ity of the compounds. However, this approach resulted in less po-
tent and less orally bioavailable MC4R agonists (Table 3).
Another approach to further optimize lead compound 2 focused
on replacing the nitrile group. As shown in Table 4, a number of
functional groups were well tolerated, including the hydroxyl
amine (35). In addition, most amide analogs of 2 showed compara-
ble potency to 2. Methyl amide 31 also exhibited very promising
rat pharmacokinetics profile with higher drug level and lower
clearance compared to 2 (F% = 46, AUCN = 1.0 lM h kg/mg, Cl =
12.1 ml/min/kg, Vdss = 1.5 L/kg). However, 29A bearing a gem-di-
methyl acetamide exhibited decreased MC4R potency and modest
oral bioavailability. Compound 29B, the diastereomer of 29A, was
less potent than 29A, suggesting that the S stereo configuration
at the 3-position of the spiro isobenzofuran is preferred for
MC4R activity regardless of substitution.
Compound 2 was the most promising lead in this series. Potent
functional activity and selectivity was maintained at melanocortin-
4 receptors across a number of species (ferret, rat, and mouse)
(Table 5).
5. The supplementary crystallographic data for compound 8A has been deposited
with The Cambridge Crystallographic Data Centre as CCDC 780119. These data
can be obtained free of charge from The Cambridge Crystallographic Data Centre
via www.ccdc.cam.ac.uk/data_request/cif.
The compound also exhibited high exposure in the brain of
Sprague-Dawley rats (brain/plasma ratio >2 @ 4 h time point fol-
lowing 1 mg/kg IV administration) (Table 6).
6. (a) Chung, J. Y. L.; Cvetovich, R.; Amato, J.; McWilliams, J. C.; Reamer, R.;
DiMichele, L. J. Org. Chem. 2005, 70, 3592; (b) Compound 12 was provided by
Merck process research labs and synthesized in a similar way as shown in
Scheme 3 and literature 6a, in which benzyl amine was used in the first step,
and hydrogenation and Boc formation were involved at the end of synthesis.
7. (a) MC4R binding IC50 was defined as the concentration of compound that can
Compound 2 was further evaluated in in vivo rodent obesity
models. It demonstrated mechanism-based acute food-intake and
body-weight reduction in DIO mice (Fig. 4), significantly lowering
food intake (43% and 24% at 4 h and 18 h, respectively) and body
weight in WT DIO mice with no appreciable effects these parame-
ters in MC3R/4R knockout mice. Further, dose-dependent food-in-
take and body-weight reduction was observed in an 18-h acute
food-intake study in DIO rats following orally administration of 2
(3 and 10 mpk). Compared to vehicle (0.5% methylcellulose), the
compound lowered food intake by 63% at 18 h post-oral dosing
at 10 mpk.9
inhibit binding of [125I]-NDP-
a-MSH by 50% from membranes prepared from
CHO cells expressing human MC4R. Agonist potency was determined in cAMP
release assays using CHO cells expressing the relevant receptors. MC4R cAMP
EC50 was defined as the inflection point of the cAMP dose–response curve for
any given compounds. Maxi percentage activation [%Max] is the percentage of
cAMP accumulation at 10 lM of compound relative to a-MSH; (b) for details
about assay protocols see: (i) Bednarek, M. A.; MacNeil, T.; Kalyani, R. N.; Tang,
R.; Van der Ploeg, L. H. T.; Weinberg, D. H. J. Med. Chem. 2001, 44, 3665; (ii)
Bednarek, M. A.; Silva, M. V.; Arison, B.; MacNeil, T.; Kalyani, R. N.; Huang, R. R.
C.; Weinberg, D. H. Peptides 1999, 20, 401.
In summary, we have described the design, synthesis, SAR, and
evaluation of a series of MC4R selective agonists containing a spiro
isobenzofuran core structure. This study led to the discovery of 2 as
a potent, selective, orally bioavailable, and brain penetrable MC4R
8. Miller, R. R. et al., unpublished results.
9. Strack, A. et al., unpublished results.