Discovery of MK-5046, a potent, selective bombesin receptor subtype-3 agonist for the treatment of obesity

Article abstract of DOI:10.1021/ml100196d

We report the development and characterization of compound 22 (MK-5046), a potent, selective small molecule agonist of BRS-3 (bombesin receptor subtype-3). In pharmacological testing using diet-induced obese mice, compound 22 caused mechanism-based, dose-

Full text of DOI:10.1021/ml100196d

pubs.acs.org/acsmedchemlett
Discovery of MK-5046, a Potent, Selective Bombesin
Receptor Subtype-3 Agonist for the Treatment of Obesity
Iyassu K. Sebhat,*,† Christopher Franklin,† Michael M.-C. Lo,† David Chen,† James P. Jewell,†
Randy Miller,‡ Jianmei Pang,‡ Oksana Palyha,§ Yanqing Kan,§ Theresa M. Kelly,§
Xiao-Ming Guan,§ Donald J. Marsh,§ Jennifer A. Kosinski,§ Joseph M. Metzger,
Kathryn Lyons,‡ Jasminka Dragovic,‡ Peter R. Guzzo,^ Alan J. Henderson,^
Marc L. Reitman,§ Ravi P. Nargund,† Matthew J. Wyvratt,† and Linus S. Lin†
Departments of †Medicinal Chemistry, ‡Drug Metabolism, §Metabolic Diseases (Obesity), and Pharmacology, Merck Research
Laboratories, Rahway, New Jersey 07065, United States, and ^AMRI, 26 Corporate Circle, Albany, New York 12212, United States
ABSTRACT We report the development and characterization of compound 22
(MK-5046), a potent, selective small molecule agonist of BRS-3 (bombesin receptor
subtype-3). In pharmacological testing using diet-induced obese mice, compound
22 caused mechanism-based, dose-dependent reductions in food intake and body
weight.
KEYWORDS MK-5046, bombesin receptor subtype-3 agonist, obesity
he mammalian bombesin G-protein-coupled receptor
subfamily1 comprises three structurally related mem-
bers, the receptors for neuromedin B (NMBR or BB1),
null (KO) mice.13 Furthermore, subchronic dosing (100 mpk
BID, 8 days) led to mechanism-based body weight lowering
in eDIO mice, while it had no significant effects in KO mice.13
Undesirable features of 2 were its poor oral pharmacoki-
netics in preclinical species and a suboptimal off-target
profile that included low micromolar binding to the human
inward rectifying potassium ion channel (hERG, IC50 = 1.2 μM)
as well as the diltiazem (DLZ) site of the rabbit calcium ion
channel (IC50 = 0.5 μM).
The low unbound exposure of compound 2 was shown to
be due primarily to high intrinsic clearance. A bile duct
cannulated rat study was performed to examine the fate of
the compound after in vivo dosing. For this purpose, a tritium
radiolabel was incorporated in the 5-position of the pyridine
ring. Biliary excretion of metabolized 2 accounted for ∼65%
of the recovered radioactivity after both iv and po dosing,
suggesting good absorption. There was extensive hydroxyla-
tion and concomitant loss of the radiolabel, suggesting
oxidative metabolism of the pyridine ring. In addition,
hydroxylation of both the 2-substituent (ethyl linker) and
the 4-substituent (branched alkyl chain) of the imidazole was
observed. Together, these oxidative processes were the
cause for the poor pharmacokinetic profile of these early
leads.
T
gastrin-releasing peptide (GRPR or BB2),2 and bombesin
receptor subtype-3 (BRS-3 or BB3).3 BRS-3 is an orphan
receptor whose natural ligand is not known and which,
despite its name, does not bind bombesin with high affinity.
BRS-3 is expressed predominantly in the central nervous
system (CNS). Mice lacking functional BRS-3 develop mild
obesity and insulin resistance, suggesting a role for the
receptor in the regulation of energy homeostasis.4 In addi-
tion, BRS-3 as well as the other mammalian family members
are expressed by several human carcinoma cell lines, where
they may play a role in growth regulation.3 Until recently,
there have been limited reports of peptide5 or small-
molecule6-9 pharmacological tools to further elucidate
the role of the receptor and determine its potential as a drug
target.
Our laboratory has described the lead identification and
initial optimization of the first reported series of small-
molecule BRS-3 agonists with efficacy in body weight
lowering.10-12 Exploratory medicinal chemistry efforts fo-
cused on lead 1. The potency at the human receptor was
improved through the incorporation of branched substitu-
ents at the 4-position of the imidazole, while systemic and
CNS exposure were increased through replacement of the
benzoic acid moiety with pyridine. This work eventually led
to the identification of 2 (Bag-1), the first key pharmacolog-
ical tool in our program. Results of in vivo testing with this
compound furnished a preclinical proof of concept for the
BRS-3 mechanism: Administration of the BRS-3 agonist in
established diet-induced obese (eDIO) mice caused a sig-
nificant reduction of acute food intake and increase in fasting
metabolic rate, and these effects were not evident in Brs3
To advance the series, we searched for motifs that might
furnish more metabolically stable structures for further
structure-activity relationship (SAR) and identified ana-
logue 3 bearing a 5-fluoropyridine. When we assessed the
stability of this compound by incubation with rat liver
Received Date: August 19, 2010
Accepted Date: October 6, 2010
Published on Web Date: October 18, 2010
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Scheme 1a
Figure 1. Structures of leads 1-3.
Table 1. Human BRS-3 Activity and Rat Liver Microsomal (RLM)
Stability of 1-3
hBRS-3a
T
1/2 (RLM incubation)
(min)
a Reagents and conditions: (a) (i) TosMIC, KOtBu, THF; (ii) NH3, MeOH.
(b) Me2NSO2Cl, NMM, DME. (c) (i) n-BuLi, THF; (ii) 4-bromophenyla-
cetic acid methyl ester, THF. (d) Pd(PPh3)4, 2-bromo-5-fluoropyridine,
Me3SnSnMe3, dioxane. (e) MeMgBr, THF. (f) HCl, MeOH. (g) Chiral
separation (Chiralpak AD-H, IPA/heptane).
compd
EC50 (nM) (% act)b
1
2
3
1900 ( 760 (74 ( 33%)
47 ( 4 (99 ( 4%)
NDc
1
66 ( 16 (97 ( 4%)
3
Scheme 2a
a Data expressed as means ( SDs (g3 independent experiments).
b % act represents the maximum activation of tested compound relative
to that of the dYpeptide {[D-Tyr6,β-Ala11 ,Phe13,Nle14]bombesin(6-14)}.
c Not determined.
microsomes, it had an improved half-life in comparison with
2. However, off-target activities remained problematic, with
reduced selectivity vs the two key ion channels (hERG, Ki =
0.4 μM; DLZ, IC50 = 0.2 μM), as well as some activation of
human PXR (IC50 = 3.5 μM with 44% maximal activation).
More information was garnered from early SAR focused
on the imidazole substituent: In a survey of branched alkyl
groups, use of a 2,2-dimethylpropane in place of the 2,2-
dimethylbutane contained in compounds 2 and 3 resulted in
∼4-fold reduction in activity at human BRS-3. While a loss in
potency and selectivity is not ideal, we considered it a small
price to pay for increased metabolic stability and proceeded
to incorporate the fluoropyridine and truncated substituent.
We next aimed to improve the selectivity for BRS-3 over
off-target effects by introducing heteroatom substituents on
the central ethylene linker to increase polarity and reduce
the number of lowenergy, unfavorable configurations for the
central ethyl linker region. We envisaged that such substitu-
tions would also improve metabolic stability, given that the
benzylic C-H would no longer be available for oxidation.
Other attempts to further reduce metabolism in the pyridine
region focused on incorporation of alternate heterocycles.
Finally, we sought to further address the metabolism of the
branched alkyl chain by incorporating metabolically more
stable fluoroalkyl and cyclopropyl modifications.
a Reagents and conditions: (a) Pd(PPh3)4, 2-bromo-5-fluoropyridine,
K2CO3, toluene/MeOH. (b) MeMgBr, THF. (c) Compound 11a: 4-bro-
moacetophenone, K2CO3, rac-trans-N,N0-dimethylcyclohexane-1,
2-diamine (cat.), CuI (cat.), pyrazole, toluene; 11b: 40-bromo-2,2,2-tri-
fluoromethylacetophenone, K2CO3, rac-trans-N,N0-dimethylcyclohex-
-
ane-1,2-diamine (cat.), CuI (cat.), pyrazole, toluene (d) NOþBF4
,
MeCN. (e) TiCl3, MeOH (f) Ph3CCl, Et3N, CH2Cl2. (g) (i) n-BuLi, THF;
(ii) 9 or 11, THF. (h) HCl, dioxane. (i) Chiral separation (Chiralpak AD-H,
IPA/heptane).
Modular syntheses were developed to access the desired
compounds. Both approaches relied on the deprotonation of
suitably substituted imidazole precursors and their subse-
quent reaction with electrophiles.
To access the series of compounds with substituents attached
to the carbon adjacent to the imidazole, 2-lithiated imidazoles
were reacted with esters to generate ketone intermediates. For
the purposes of the current series, reaction of 3,3-dimethylbu-
tyraldehyde (4) with TosMIC generated 4-(2,2-dimethylpropyl)-
1H-imidazole, which was subsequently protected as the
dimethylsulfamoyl derivative 5. Lithiation of 5, followed by its
reaction with methyl 4-bromophenylacetate, led to ketone 6,
which was coupled to the stannane generated in situ from
2-bromo-5-fluoropyridine via palladium catalysis. Finally,
the addition of methylmagnesium bromide, deprotection,
and chiral separation afforded two enantiomeric alcohols.
Compound 7 is the more potent isomer.
To access the series with substituents attached to the
carbon distal from the imidazole, 1-trityl-2-methyl-4-(2,2-
dimethylpropyl)imidazole, synthesized in three steps from
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Scheme 3a
Table 3. Rat Plasma Clearance,a Protein Binding,b and Stabilityc
for Compounds 3-22
rat in vivo
rat microsomal incubation
Clp
compd (mL/min/kg)
plasma Fu
T1/2
(min)
Fu
(%)
(%)
3
7
12
14
ND
0.1
0.4
6.8
3.2
1.0
3
NDd
20
1
14
15
21
22
28
5
20
110
52
ND
ND
ND
ND
ND
ND
20
a Plasmaclearance(Clp) calculatedfollowing1 mg/kg ivdose. b Protein
binding assessed using human plasma (200 μL) or microsomal proteins
(0.2 mg/mL) and test compound (1 μM). c Microsomal stability assessed
using liver microsomes (0.2 mg protein/mL) and test compound (1 μM).
d Not determined.
a Reagents and conditions: (a) MeONHMe, EDC, HOBt, NMM, CH2Cl2.
(b) Na2CO3, I2, dioxane/H2O. (c) Na2SO3, EtOH/H2O. (d) Ph3CCl, Et3N,
CH2Cl2. (e) (i) EtMgBr, CH2Cl2; (ii) 17, CH2Cl2. (f) H2NNH2, KOH,
ethylene glycol. (g) (i) BuLi, THF; (ii) 11a or 11b, THF. (h) HCl, dioxane.
(i) Chiral separation (Chiralpak AD-H, IPA/heptane).
Table 4. Dog Pharmacokinetic Dataa for Pyrazole Analogues 21
and 22
Clp
T1/2
(h)
po AUCN
[μM h/(mg/kg)]
Foral
(%)
compd
(mL/min/kg)
21
22
10
0.3
1.4
1.4
3.7
29
52
Table 2. Human BRS-3 Binding and Functional Potency for Com-
pounds 3-22
5.6
a Plasma clearance (Clp) and half-life (T1/2) calculated following
0.5 mg/kg iv dose. Normalized oral exposure (po AUCN) and oral bio-
availability (Foral) calculated following 1 mg/kg po dose.
hBRS-3a
hBRS-3a
compd
IC50 (nM)
EC50 (nM) (% act)b
3
7
16 ( 6
82 ( 16
17 ( 5
5 ( 1
66 ( 16 (97 ( 4%)
220 ( 22 (99 ( 4%)
67 ( 13 (99 ( 2%)
16 ( 3 (94 ( 5%)
14 ( 4 (94 ( 6%)
25 ( 3 (102 ( 6%)
suggesting higher intrinsic clearance. This was reinforced in
studies assessing the stability of the compounds in incubations
with rat liver microsomes. The half-life for compound 7 was
significantly shorter than that for compound 14, indicating its
propensity for higher metabolic intrinsic clearance (the two
had similar unbound fractions under the conditions of the
assay; Fu = 20%). On this basis, we focused our efforts on
analogues of 14, leading to the identification of pyrazole 15
with improved human potency. The rat pharmacokinetic pro-
file of 15 was notable for very high total clearance, which was
largely a function of increased plasma unbound fraction as
compared to the other compounds. To further reduce the
potential for oxidative metabolism, our attention turned to
the remaining region (common to compound 2) that was
identified to be susceptible to oxidation in rats: the imidazole
4-substituent. After considerable efforts, trifluoromethyl-sub-
stituted methyl cyclopropane was incorporated to furnish
compound 21, where both the unbound fraction in rat plasma
and the plasma clearance were reduced.
The pharmacokinetic properties of 21 were also evaluated
in the dog where the half-life was found to be unacceptably
short (Table 4). Replacement of the remaining methyl group
with a trifluoromethyl group gave 22. The compound ex-
hibits overall improvements in dog pharmacokinetic proper-
ties relative to 21, while maintaining good potency and
improved selectivity over off-target activities (hERG potas-
sium channel binding Ki > 8 μM).
14
15
21
22
11 ( 2
27 ( 13
a Data expressed as means ( SDs (g3 independent experiments).
b % act represents the maximum activation of tested compounds
relative to that of dY peptide.
4,4-dimethylpentene, was lithiated in the R-position via treat-
ment with n-butyllithium and subsequently reacted with
methylaryl ketones 9 and 11a or the trifluorinated analogue
11b. As before, deprotection and chiral separation furnished
agonists 14 and 15. Similarly, the use of the trityl-protected
imidazole 20 (derived in five steps from 2-methylimidazole)
led to compounds 21 and 22.14
Human BRS-3 binding potency and functional potency for
the compounds are summarized in Table 2.15 The introduc-
tion of a tertiary alcohol on either carbon of the linker
resulted in compounds that maintained similar activity to
lead 3. Substitution distal from the imidazole (14 vs 7) gave
slightly higher potency. Compounds 7 and 14 afforded
improved off-target profiles relative to 3. In particular, bind-
ing to the hERG potassium channel was diminished (Ki = 5.9
and 4.2 μM for 7 and 14, respectively).
The initial leads 7 and 14 were assessed in rat pharmaco-
kinetic studies. Although compound 7 had lower in vivo rat
plasma clearance than 14 (14 vs 28 mL/min/kg), the un-
bound fraction (Fu) in plasma was considerably lower,
Compound 22 was evaluated in a screen of >100 recep-
tors, ion channels, and enzymes. Eight off-target activities
were identified with IC50 values in the range of 0.5-10 μM,
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Table 5. BRS-3 Potency, Selectivity,a and Pharmacokineticb Profile of 22 (MK-5046)
human
mouse
rat
dog
rhesus
BRS3 IC50 (nM)
BRS3 Ki (nM)
28 ( 13
3.7 ( 0.5
14 ( 4
5.4 ( 2
1.6 ( 0.7
21 ( 0.9
120 ( 10
1.2 ( 0.4
0.6 ( 0.2
2.2 ( 1
6.5 ( 1.9
9.9 ( 1.3
1.6 ( 0.8
50 ( 15
2.4 ( 1.3
6.9 ( 4
BRS3 EC50 (nM)
% activationc
100 ( 6
110 ( 9
100 ( 58
110 ( 8
NMBR IC50 (nM)
GRPR IC50 (nM)
hERG Ki (μM)
>10000
>10000
>8
DLZ IC50 (μM)
1.9
>25
hPXR EC50 (μM)
po AUCN [μM h/(mg/kg)]
0.1
9.6
32
1.2
64
20
0.98
0.7
3.7
52
0.8
18
F
oral (%)
Clp (mL/min/kg)
Vdss (L/kg)
5.6
8.8
2.2
1.1
0.45
1.4
0.44
1.0
T
1/2 (h)
a Data expressed as means ( SDs (g3 independent experiments) or single value. b Plasma clearance (Clp), volume of distribution (Vdss), and half-life
(T1/2) calculated following 1 mg/kg iv dose in mouse and rat and 0.5 mg/kg iv dose in dog and rhesus. Oral bioavailability (Foral) calculated following
2 mg/kg in mouse and rat and 1 mg/kg in dog and rhesus. c % activation represents the maximum activation of tested compounds relative to that of
dY-peptide for human, dog, and rhesus receptors and a reference small molecule BRS-3 agonist for mouse and rat receptors.
but none were considered significant enough to preclude the
use of the compound as a pharmacological tool or for clinical
development.
ACKNOWLEDGMENT We thank the department of Laboratory
Animal Resources for their assistance in animal dosing and sampling.
The BRS-3 potency and pharmacokinetic parameters for
22 across a range of species, as well as activity of the com-
pound in selected off-target assays are summarized in Table 5.
Potency, selectivity, and pharmacokinetics were all signif-
icantly improved as compared to compound 3, and while the
half-life remained relatively short, this was partially amelio-
rated by prolonged absorption, thereby allowing oral dosing
paradigms in pharmacological studies.
Compound 22 was assessed for acute efficacy in wild-type
(WT) and Brs3 KO mice. In WT mice, the compound caused
significant reductions in food intake and increase in fasting
metabolic rate with no effect in the KO mice. After subchronic
dosing in eDIO mice, compound 22 (25 mg/kg/day by sc
infusion) caused reductions in body weight (8-9% as com-
pared tovehicle-dosed animals) that were sustained for 14 days
with no evidence of tachyphylaxis (manuscript submitted16).
In summary, a SAR about the core of lead compound 2 has
resulted in the identification of a series of BRS-3 agonists with
improved potency, selectivity, and oral exposure in preclinical
species. On the basis of the in vitro and pharmacokinetic
profiles, compound 22 (MK-5046) was chosen for further
pharmacological study. In eDIO mice, sustained body weight
loss was observed following 14 days of treatment with the
compound. The data suggest that agonism of the BRS-3 may
present a useful treatment for obesity, and compound 22
presents a suitable candidate for clinical investigation.
REFERENCES
(1)
Jensen, R. T.; Battey, J. F.; Spindel, E. R.; Benya, R. V.
Mammalian bombesin receptors: Nomenclature, distribution,
pharmacology, signaling, and functions in normal and dis-
ease states. Pharmacol. Rev. 2008, 60, 1–42.
(2)
(3)
Battey, J.; Wada, E. Two distinct receptor subtypes for mam-
malian bombesin-like peptides. Trends Neurosci. 1991, 14,
524–528.
Fathi, Z.; Corjay, M. H.; Shapira, H.; Wada, E.; Benya, R.;
Jensen, R.; Viallet, J.; Sausville, E. A.; Battey, J. F. BRS3: A novel
bombesin receptor subtype selectively expressed in testis
and lung carcinoma cells. J. Biol. Chem. 1993, 268, 5979–
5984.
(4)
(5)
Ohki-Hamazaki, H.; Watase, K.; Yamamoto, K.; Ogura, H.;
Yamano, M.; Yamada, K.; Maeno, H.; Imaki, J.; Kikuyama, S.;
Wada, E.; Wada, K. Mice lacking bombesin receptor subtype-
3 develop metabolic defects and obesity. Nature 1997, 390,
165–169.
Boyle, R. G.; Humphries, J.; Mitchell, T.; Showell, G. A.; Apaya,
R.; Iijima, H.; Shimada, H.; Arai, T.; Ueno, H.; Usui, Y.; Sakaki,
T.; Wada, E.; Wada, K. The design of a new potent and
selective ligand for the orphan bombesin receptor subtype
3 (BRS3). J. Pept. Sci. 2005, 11, 136–141 and references
therein.
(6)
Weber, D.; Berger, C.; Heinrich, T.; Eickelmann, P.; Antel, J.;
Kessler, H. Systematic optimization of a lead-structure iden-
tities for a selective short peptide agonist for the human
orphan receptor BRS-3. J. Pept. Sci. 2002, 8, 461–475.
Weber, D.; Berger, C.; Eickelmann, P.; Antel, J.; Kessler, H.
Design of Selective Peptidomimetic Agonists for the Human
Orphan Receptor BRS-3. J. Med. Chem. 2003, 46, 1918–1930.
Carlton, D. L.; Collin-Smith, L. J.; Daniels, A. J.; Deaton, D. N.;
Goetz, A. S.; Laudeman, C. P.; Littleton, T. R.; Musso, D. L.; Ott
Morgan, R. J.; Szewczyk, J. R.; Zhang, C. Discovery of small
molecule agonists for the bombesin receptor subtype 3 (BRS-3)
SUPPORTING INFORMATION AVAILABLE Full experimen-
tal procedures and characterization data. This material is available
free of charge via the Internet at http://pubs.acs.org.
(7)
(8)
AUTHOR INFORMATION
Corresponding Author: *Tel: 732-594-0168. E-mail: iyassu_
sebhat@merck.com.
r
2010 American Chemical Society
46
DOI: 10.1021/ml100196d ACS Med. Chem. Lett. 2011, 2, 43–47
|

pubs.acs.org/acsmedchemlett
based on an omeprazole lead. Bioorg. Med. Chem. Lett. 2008,
18, 5451–5455.
HEK293AEQ cells overexpressing BRS-3, using an aequorin
bioluminescence assay.
(9)
Guo, C.; Guzzo, P. R.; Hadden, M.; Sargent, B.; Yet, L.; Kan, Y.;
Palyha, O.; Kelly, T. M.; Guan, X.-M.; Rosko, K.; Gagen, K.;
Metzger, J. M.; Dragovic, J.; Lyons, K.; Lin, L. S.; Nargund, R. P.
Synthesis of 7-benzyl-5-(piperidin-1-yl)-6,7,8,9-tetrahydro-
3H-pyrazolo[3,4-c]-[2,7]naphthyridin-1ylamine and its ana-
logs as bombesin receptor subtype-3 agonists. Bioorg. Med.
Chem. Lett. 2010, 20, 2785.
(16) Guan, X.-M. Manuscript submitted.
(10) He, S.; Dobbelaar, P. H.; Liu, J.; Jian, T.; Sebhat, I. K.; Lin, L. S.;
Goodman, A.; Guo, C.; Guzzo, P. R.; Hadden, M.; Henderson,
A. J.; Ruenz, M.; Sargent, B.; Yet, L.; Kelly, T. M.; Palyha, O.;
Kan, Y.; Pan, J.; Chen, H.; Marsh, D.; Shearman, L. P.; Strack,
A. M.; Metzger, J. M.; Feighner, S. D.; Tan, C.; Howard, A. D.;
Tamvakopoulos, C.; Peng, Q.; Guan, X.-M.; Reitman, M. L.;
Patchett, A. A.; Wyvratt, M. J.; Nargund, R. P. Discovery of
substituted biphenyl imidazoles as potent bioavailable
bombesin receptor subtype-3 agonists. Bioorg. Med. Chem.
Lett. 2010, 20, 1913–1917.
(11) Liu, J.; He, S.; Jian, T.; Dobbelaar, P. H.; Sebhat, I. K.; Lin, L. S.;
Goodman, A.; Guo, C.; Guzzo, P. R.; Hadden, M.; Henderson,
A. J.; Pattamana, K.; Ruenz, M.; Sargent, B.; Swenson, B.; Yet,
L.; Tamvakopoulos, C.; Peng, Q.; Pan, J.; Kan, Y.; Palyha, O.;
Kelly, T. M.; Guan, X.-M.; Howard, A. D.; Marsh, D.; Metzger, J.;
Reitman, M. L.; Wyvratt, M. J.; Nargund, R. P. Synthesis and
SAR of derivatives based on 2-biarylethylimidazole as bombesin
receptor subtype-3 (BRS-3) agonists for the treatment of
obesity. Bioorg. Med. Chem. Lett. 2010, 20, 2074–2077.
(12) Hadden, M.; Goodman, A.; Guo, C.; Guzzo, P. R.; Henderson,
A. J.; Pattamana, K.; Ruenz, M.; Sargent, B.; Swenson, B.; Yet,
L.; Liu, J.; He, S.; Sebhat, I. K.; Lin, L. S.; Tamvakopoulos, C.;
Peng, Q.; Kan, Y.; Palyha, O.; Kelly, T. M.; Guan, X.-M.;
Metzger, J.; Reitman, M. L.; Nargund, R. P. Synthesis and
SAR of heterocyclic carboxylic acid isosteres based on 2-bi-
arylethylimidazole as bombesin receptor subtype-3 (BRS-3)
agonists for the treatment of obesity. Bioorg. Med. Chem. Lett.
2010, 20, 2912–2915.
(13) Guan, X.-M.; Chen, H.; Dobbelaar, P. H.; Dong, Y.; Fong, T. M.;
Gagen, K.; Gorski, J.; He, S.; Howard, A. D.; Jian, T.; Jiang, M.;
Kan, Y.; Kelly, T. M.; Kosinski, J.; Lin, L. S.; Liu, J.; Marsh, D.;
Metzger, J. M.; Miller, R.; Nargund, R. P.; Palyha, O.; Shearman,
L.; Shen, Z.; Stearns, R.; Strack, A. M.; Stribling, S.; Tang, Y. S.;
Wang, S.-P.; White, A.; Yu, H.; Reitman, M. L. Regulation of
energy homeostasis by bombesin receptor subtype-3: Selective
receptor agonists for the treatment of obesity. Cell Metab.
2010, 11, 101–112.
(14) Absolute stereochemistry was determined via X-ray crystal-
lography of the hydrochloride salt of an iodinated analogue
((2S)-1,1,1-trifluoro-3-(5-iodo-4-{[1-(trifluoromethyl)cyclopropyl]-
methyl}-1H-imidazol-2-yl)-2-[4-(1H-pyrazol-1-yl)phenyl]pro-
pan-2-ol). Crystals of this salt were grown by evaporation of
an EtOAc/hexane solution over a period of approximately 1
week. Diffraction data were measured at room temperature.
Examination of the intermolecular distances to the Cl ion
as well as the electron density maps indicates the proton-
ation by HCl to form the salt occurs at N23. With the
presence of the two heavy atoms in the lattice, determina-
tion of the absolute configuration at the chiral C (C1, S) was
unambiguous.
(15) Receptor binding was performed using membranes from
CHO or HEK293 cells overexpressing the indicated receptor.
A 30 pM concentration of [125I]-[D-Tyr6,β-Ala11 ,Phe13,Nle14]-
bombesin(6-14) (human assay) and 660 pM [3H]Bag-3 (rat
and mouse assays) were used. For functional assays, agonist-
induced mobilization of intracellular Ca2þ was measured in
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