Melanin-Concentrating Hormone Receptor 1 Antagonists Lacking an Aliphatic Amine: Synthesis and Structure-Activity Relationships of Novel 1-(Imidazo[1,2-a]pyridin-6-yl)pyridin-2(1H)-one Derivatives

Article

pubs.acs.org/jmc

Melanin-Concentrating Hormone Receptor 1 Antagonists Lacking an

Aliphatic Amine: Synthesis and Structure−Activity Relationships of

Novel 1‑(Imidazo[1,2‑a]pyridin-6-yl)pyridin-2(1H)‑one Derivatives

Hideyuki Igawa,*^,†Masashi Takahashi,^†Keiko Kakegawa,^†Asato Kina,^†Minoru Ikoma,^†Jumpei Aida,^†

Tsuneo Yasuma,^‡Yayoi Kawata,^†Shuntaro Ashina,^†Syunsuke Yamamoto,^†Mrinalkanti Kundu,^∥

Uttam Khamrai,^∥Hideki Hirabayashi,^†Masaharu Nakayama,^†Yasutaka Nagisa,^§Shizuo Kasai,^†

and Tsuyoshi Maekawa^†

^†Pharmaceutical Research Division, Takeda Pharmaceutical Co., Ltd., Shonan Research Center, 26-1, Muraoka-Higashi 2-Chome,

Fujisawa, Kanagawa 251-8555, Japan

^‡CMC Center, Takeda Pharmaceutical Co., Ltd., 17-85, Jusohonmachi 2-Chome, Yodogawa-ku, Osaka 532-8686, Japan

^§CVM Marketing Japan Pharma Business Unit, Takeda Pharmaceutical Co., Ltd. 12-10, Nihonbashi 2-Chome, Chuo-ku, Tokyo

103-8686, Japan

^∥TCG Lifesciences Ltd., Block BN, Plot 7, Saltlake Electronics Complex, Sector V, Kolkata 700091, India

S

* Supporting Information

ABSTRACT: Aiming to discover melanin-concentrating hormone receptor 1 (MCHR1) antagonists with improved safety

proﬁles, we hypothesized that the aliphatic amine employed in most antagonists reported to date could be removed if the bicyclic

motif of the compound scaﬀold interacted with Asp123 and/or Tyr272 of MCHR1. We excluded aliphatic amines from our

compound designs, with a cutoﬀ value of pK_a< 8, and explored aliphatic amine-free MCHR1 antagonists in a CNS-oriented

chemical space limited by four descriptors (TPSA, ClogP, MW, and HBD count). Screening of novel bicyclic motifs with high

intrinsic binding aﬃnity for MCHR1 identiﬁed the imidazo[1,2-a]pyridine ring (represented in compounds 6a and 6b), and

subsequent cyclization of the central aliphatic amide linkage led to the discovery of a potent, orally bioavailable MCHR1

antagonist 4-[(4-chlorobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)pyridin-2(1H)-one 10a. It exhibited

low potential for hERG inhibition and phospholipidosis induction as well as suﬃcient brain concentration to exert antiobesity

eﬀects in diet-induced obese rats.

meaningful weight loss. The FDA recently approved several new

antiobesity drugs (lorcaserin, combined phentermine and

topiramate, combined bupropion and naltrexone, and liraglu-

tide), but these drugs have still not fully met doctors’ and

patients’ needs for antiobesity treatment because of concerns

regarding eﬃcacy, safety, and cost.²⁻⁵

INTRODUCTION

■

Obesity mainly results from an imbalance between energy intake

and expenditure, leading to excess energy storage in the form of

body fat, which releases numerous inﬂammatory adipokines and

contributes to the development of various disorders such as

diabetes, hypertension, dyslipidemia, depression, coronary artery

disease, and cancer.^1,2The rate of obesity, particularly morbid

obesity, is markedly increasing throughout the world, resulting in

a serious public health issue.^3,4Lifestyle modiﬁcations such as

diet, exercise, and behavior comprise the ﬁrst line of obesity

management; however, they struggle to achieve and sustain

Melanin-concentrating hormone (MCH) is a cyclic 19-amino-

acid peptide expressed predominantly in the lateral hypothalamic

Received: November 1, 2015

A

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area and zona incerta, and MCH-producing neurons project

throughout the brain.^6,7Two distinct MCH receptors, MCHR1

and MCHR2, have been identiﬁed.⁸⁻¹²MCHR1 is widely

distributed in the brain, including the hypothalamus, thalamus,

olfactory cortex, amygdala striatum, and hippocampus, in all

vertebrates.^13,14In contrast, MCHR2 is expressed only in higher

mammals and not in rodents.¹⁵⁻²⁰Furthermore, the physio-

logical function of MCHR2 is poorly understood at present.

A large number of studies have demonstrated that the MCH/

MCHR1 pathway plays a key role in the regulation of feeding

behavior and energy expenditure. MCH mRNA and peptide

levels were upregulated in dietary obese rats²¹and genetically

obese rodents including ob/ob mice,²²db/db mice,²³A^y/a

(agouti) mice,²⁴and Zucker (fa/fa) rats.²⁵Chronic intra-

cerebroventricular infusion of MCH induced hyperphagia, body

weight gain, and hyperinsulinemia, particularly under high-fat-

diet conditions.²⁶⁻²⁸Transgenic mice overexpressing MCH in

the lateral hypothalamus were more susceptible to obesity and

insulin resistance when fed a high-fat diet.²⁹In contrast, mice

lacking MCH or MCHR1 were lean with increased metabolic

rates and resistance to diet-induced obesity.³⁰⁻³³In human

studies, increased MCH expression levels were observed in the

hypothalamus in obese subjects as compared with lean

subjects.³⁴Two loss-of-function MCHR1 mutants (R210H

and P377S) were identiﬁed in markedly underweight subjects,

indicating the possibility that the lean phenotype would be linked

to deﬁcient MCHR1 signaling.³⁵

and phospholipidosis limit their clinical potential. Under these

circumstances, we initiated our activities to discover novel

MCHR1 antagonists with reduced safety concerns.

The hERG liability of MCHR1 antagonists has been

recognized as a common issue in most research programs.^45,46

Several reports have indicated the structural similarity between

MCHR1 antagonists and existing hERG channel blockers,

implying the existence of an overlapping pharmacophore for

these two targets. What makes the development of MCHR1

antagonists so challenging and keeps most programs from

advancing to the clinic is this inherent diﬃculty in separating the

aﬃnities for these two targets. It has been widely suggested that

the aliphatic amine moiety is essential for potent binding aﬃnity

via putative interactions with MCHR1; however, this positively

charged site would also be a key substructure for interactions

with the hERG channel through cation−π interactions. Indeed,

we recently reported that introducing substituents near the

cationic site eﬀectively reduced hERG inhibition.⁴⁷In some

cases, the aliphatic amine could be replaced with other functional

groups such as amides and carbamates without loss of aﬃnity for

MCHR1.⁴²These amine variants had high selectivity over

hERG; however, all were devoid of in vivo eﬃcacy potentially

due to lower CNS availability. Sasmal et al. conducted a unique

docking study for MCHR1 on the basis of the β2-adrenergic

receptor with their 2-aminoquinazoline derivative 4 and

proposed that one nitrogen atom was engaged in an ionic

interaction with Asp III:08 and the other nitrogen formed a

hydrogen bond with a Thr in the extracellular loop (ECL) 2b

(Figure 2).^48,49On the contrary, the tetraline ring does not have a

Since we ﬁrst identiﬁed MCH as an endogenous ligand for

MCHR1³⁶⁻⁴⁰and discovered the small-molecule antagonist T-

226296 (1, (−) enantiomer),⁴¹a wide variety of potent small-

molecule MCHR1 antagonists have been identiﬁed by our group

(Figure 1).⁴²⁻⁴⁴The structures can be described around a central

Figure 2. Docking model of the quinazoline derivative 4 reported by

7TM Pharma on the basis of the β2-X-ray structure. Dotted lines denote

the hydrogen-bonding interactions with the receptor. ECL refers to the

extracellular loop.

hydrogen-bonding interaction with any nearby amino acid

residue in compound 1.⁴²Inspired by this model and a recent

report of nonbasic MCHR1 antagonists by Washburn et al.,^50,51

we hypothesized that the aliphatic amine could be removed

without loss of binding aﬃnity if the bicyclic motif actively

interacts with the receptor via hydrogen bonding, thus leading to

amine-free MCHR1 antagonists without hERG liability (Figure

3). Our molecular modeling studies suggested that Asp123 and

Tyr272 were possible interaction counterparts of the bicyclic

motif in our model. On the basis of this concept, we decided to

exclude the aliphatic amine motif from our drug design and

pursue amine-free MCHR1 antagonists.

Together with the design restriction in terms of the

aforementioned chemical structure, we also set a cutoﬀ value

of basicity, pK_a, of <8, which was proposed by Ploemen et al. as a

guidance for phospholipidosis-negative chemicals.⁵²Further-

more, as MCHR1 is dominantly expressed in the brain, small-

molecule antagonists are required to penetrate the blood−brain

barrier (BBB). Compared with peripherally acting agents, CNS-

acting agents are recommended to fall into a more deﬁned

Figure 1. Chemical structures of our MCHR1 antagonists.

amide with a left-hand side (LHS) and a right-hand side (RHS),

which consists of a bicyclic and an aliphatic amine motif. A

docking study of tetraline derivative 1 with a homology model

based on bovine rhodopsin suggested that the central carbonyl

group interacts with Gln127 and the aliphatic amine in the RHS

interacts with Asn294.⁴²Although these small-molecule

MCHR1 antagonists exert potent anorectic eﬀects on obese

animal models toxicological concerns such as hERG inhibition

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Figure 3. Design concept of amine-free MCHR1 antagonists. Dotted lines denote the hydrogen-bonding interactions with the receptor.

a

Scheme 1. Synthesis of Phenoxyacetamide Derivatives 6a−d

a

Reagents and conditions: (a) corresponding RNH₂, WSC, HOBt, DMF, rt, overnight, 24%−63%.

a

Scheme 2. Synthesis of 4-Alkoxypyridone Derivatives 10a, 10c, 10d, 10g−l, and 10o−q

a

Reagents and conditions: (a) corresponding benzyl alcohol, NaH, THF, rt, 5 h, 6%−66%; (b) Ac₂O, 140 °C, 2 h, 31%−58%; (c) 11a, CuI,

DMEDA, K₂CO₃, DMSO, 110 °C, 1 h, MW irradiation, 56% (for 10c); (d) 11a, CuI, trans-N,N′-dimethylcyclohexane-1,2-diamine, K₂CO₃, DMF,

130 °C, 16 h, 11% (for 10d); (e) 11a, CuI, DMEDA, K₂CO₃, DMSO, 150 °C, 3 h, 17% (for 10a); (f) 11a−h, CuI, trans-N,N′-dimethylcyclohexane-

1,2-diamine, K₂CO₃, 1,4-dioxane, 110 °C, 11 h, 22%−50% (for 10g−l, 10o, and 10p); (g) SOCl₂, CH₂Cl₂, rt, 18 h, 96%; (h) TMSCN, TBAF, THF,

rt, 4 h, 48%.

chemical space for BBB permeability.⁵³Hitchcock et al. reported

an example consisting of four physicochemical descriptors: polar

surface area (PSA), ClogP, molecular weight (MW), and number

of hydrogen-bonding donors (HBDs).⁵⁴They deﬁned PSA < 70,

2 < ClogP < 4, MW < 450, and HBD count = 0 or 1 as a preferred

range to increase the potential for BBB penetration. This

provided us a simple guideline to design new compounds with a

high probability of CNS penetration.

In this article, we report the lead generation strategy for novel

amine-free MCHR1 antagonists with high CNS penetration and

an imidazo[1,2-a]pyridine ring as a unique bicyclic motif with

putative interactions with Asp123 and/or Tyr272 of MCHR1.

We also discuss the structure−activity relationship (SAR) of

these antagonists as well as the pharmacological action of a

representative compound to reveal that the newly designed

MCHR1 antagonist was suﬃciently exposed to the CNS and that

it exhibited a potent anorectic eﬀect based on its MCHR1

antagonistic activity.

CHEMISTRY

■

Phenoxyacetamide derivatives 6a−d were synthesized as

illustrated in Scheme 1. Carboxylic acid 5 was subject to

condensation reaction using WSC to aﬀord the desired

compounds 6a−d. The corresponding aniline reagents used in

Scheme 1 were synthesized as described in the Supporting

Information.

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a

Scheme 3. Synthesis of 5-Methylpyridone Derivative 16

a

Reagents and conditions: (a) acetyl bromide, AcOH, 80 °C, 3 h, 60%; (b) NaH, 4-chlorobenzyl alcohol, 80 °C, 16 h, 28%; (c) Ac₂O, reﬂux, 4 h,

then 1 N NaOH, MeOH, reﬂux, 1 h, 4%; (d) 11a, CuI, trans-N,N′-dimethylcyclohexane-1,2-diamine, K₂CO₃, dioxane, 110 °C, 16 h, 42%.

a

Scheme 4. Synthesis of 4-Alkoxypyridone Derivatives 10m and 10n

a

Reagents and conditions: (a) corresponding Grignard reagent, THF, −78 °C, 2 h, 39%−40%.

a

Scheme 5. Synthesis of 4-Alkoxypyridone Derivatives 10b, 10e, 10f, and 10r−z

a

Reagents and conditions: (a) H₂(1 atm), 10% Pd−C, MeOH, 1 h, 99%; (b) POBr₃, DMF, 110 °C, 1 h, 71%; (c) 4-(triﬂuoromethoxy)phenol, CuI,

DMEDA, K₂CO₃, DMSO, 150 °C, 3 h, 31%; (d) 4-bromobenzyl alcohol, bis(2-methoxyethyl) azodicarboxylate, PPh₃, THF, rt, 16 h, 63% (for 10e);

(e) corresponding benzyl bromide, K₂CO₃, DMF, rt, 16 h, 32%−39%, (for 10f and 10r); (f) corresponding benzyl alcohol, ADDP, PBu₃, THF, 60

°C, 4 h, 22%−53% (for 10s−y); (g) (2-triﬂuoromethylthiophen-4-yl)methanol, KO^tBu, toluene, 100 °C, 1 h, 33% (for 10z).

Scheme 2 depicts a general route to access 4-alkoxypyridone

derivatives (10a, 10c, 10d, 10g−l, and 10o−q). An S_NAr

reaction of a variety of benzyl alcohols to 4-chloropyridine-N-

oxide 7 gave intermediates 8a−c, which were subsequently

treated with acetic anhydride to give 1-unsubstituted pyridones

9a−c. Subsequent copper coupling reactions of 1-unsubstituted

pyridones with a variety of 6-iodoimidazopyridines 11a−h

aﬀorded the target compounds (10a, 10c, 10d, 10g−l, and 10o−

p). For these coupling reactions, we selected DMEDA or trans-

N,N′-dimethylcyclohexane-1,2-diamine as a ligand and used a

stoichiometric amount of CuI because of the low reactivity of

9a−c. The 2-cyanomethyl derivative 10q was synthesized from

the 2-hydroxymethyl derivative 10o by the chlorination reaction

followed by cyanation using TMSCN. The synthesis of 6-

iodoimidazopyridines 11a−h is summarized in the Experimental

Section.

4-Alkoxypyridone with a methyl group on its 5-position was

obtained using 4-bromo-3-methylpyridine-N-oxide 13 in a

manner analogous to Scheme 2. Intermediate 13 was prepared

from 3-methyl-4-nitropyridione-N-oxide 12 (Scheme 3).

Scheme 4 conferred further derivatization of compound 10l.

Reaction of 10l with methyl magnesium bromide or cyclopropyl

magnesium bromide at −78 °C aﬀorded compounds 10m and

10n.

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a

Scheme 6. Synthesis of 3-Alkoxypyridone Derivatives 24a and 24b

a

Reagents and conditions: (a) KOH, 4-chlorobenzyl bromide, 40 °C, 1.5 h, 80%; (b) 11a, CuI, DMEDA, K₂CO₃, DMSO, 150 °C, 1 h, 13%−15%;

(c) 4-(triﬂuoromethoxy)phenol, Cu(OAc)₂, MS3A, pyridine, CH₃CN, rt, 10 days, 47%; (d) KO^tBu, BuOH, water, 150 °C, 1 h, microwave

t

irradiation, 73%.

a

Scheme 7. Synthesis of Pyridazinone Derivative 28

a

Reagents and conditions: (a) 4-chlorobenzyl bromide, K₂CO₃, CH₃CN, DMF, rt, 16 h, 64%; (b) MeOH, conc HCl, reﬂux, 3 h, 92%; (c) 2-

cyclopropyl-6-iodo-3-methylimidazo[1,2-a]pyridine, K₂CO₃, CuI, trans-N,N′-dimethylcyclohexane-1,2-diamine, dioxane, 110 °C, 16 h, 70%.

a

Scheme 8. Synthesis of Pyrimidinone Derivative 31

a

Reagents and conditions: (a) 4-chlorobenzyl bromide, Ag₂CO₃, THF, reﬂux, 2 h, 10%; (b) (2-cycloprypyl-3-methylimidazo[1,2-a]pyridine-6-

yl)boronic acid, Cu(OAc)₂, pyridine, CH₂Cl₂, MeOH, rt, 16 h, 6%.

a

Scheme 9. Synthesis of Pyrimidinone Derivative 35

a

Reagents and conditions: (a) (4-chlorophenyl)methanol, K₂CO₃, DMF, rt, 16 h, 27%; (b) NaOH, dioxane, water, reﬂux, 3 h, 5%; (c) 11a, K₂CO₃,

CuI, trans-N,N′-dimethylcyclohexane-1,2-diamine, K₂CO₃, dioxane, 110 °C, 16 h, 18%.

Scheme 5 provided an alternative method to obtain 4-

alkoxypyridone derivatives in which the alkoxy moiety at the 4-

position could be eﬃciently screened using 4-hydroxypyridone

17 or 4-bromopyridone 18 as a key intermediate. Intermediate

17 was prepared from compound 10c via a hydrogenation

reaction, and a subsequent POBr₃bromination reaction gave

compound 18. Alkylation of intermediate 17 under a basic or the

Mitsunobu condition aﬀorded the target compounds (10e, 10f,

and 10r−y). On the contrary, intermediate 18 was subject to a

copper coupling reaction or S_NAr reaction, yielding compound

10b or 10z, respectively.

Conversely, the Chan−Lam−Evans coupling reaction of boronic

acid pinacol ester 21 with a corresponding phenol gave

intermediate 22. The chloro atom of 22 was then hydrolyzed

using KO^tBu, aﬀording the 1-unsubstituted pyridone 23, which

was converted to the target compound 24b using the copper

coupling reaction described in Scheme 2.

Schemes 7−9 illustrate the synthesis of pyridazinone

derivative 28 and pyrimidinone derivatives 31 and 35. The

synthesis of pyridazinone derivative 28 was started with a THP-

protected pyridazinone-5-ol 25. Compound 25 was alkylated

with a benzyl bromide, followed by THP deprotection and a

copper coupling reaction to aﬀord the target compound 28. The

synthetic method of the starting material 25 is summarized in the

Supporting Information. Pyrimidinone derivative 31 was

synthesized from pyrimidin-4,6-diol 29 via alkylation and the

Scheme 6 describes the synthesis of 3-alkoxypyridone

derivatives 24a and 24b. Selective alkylation of pyridine-2,3-

diol 19 at the 3-position gave 3-benzyloxypyridone 20, which was

subject to a copper coupling reaction to yield compound 24a.

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a

Scheme 10. Synthesis of Tetrahydropyranopyridone 41

a

Reagents and conditions: (a) 1-(4-chlorophenyl)prop-2-en-1-ol, Pd₂(OAc), NaHCO₃, DMF, 120 °C, 15 h, 60%; (b) LiBH₄, THF, rt, 1 h, 82%; (c)

NaH, DMF, rt, 17 h, 64%; (d) ammonium acetate, AcOH, 200 °C, 1 h, microwave irradiation, 31%; (e) 11a, CuI, N,N′-dimethylethane-1,2-diamine,

K₂CO₃, DMSO, 150 °C, 1 h, microwave irradiation, 57%.

a

Scheme 11. Synthesis of Furopyridone 45

a

Reagents and conditions: (a) isobutyl carbonochloridate, TEA, acetone, 0 °C, 1 h, then sodium azide, water, 0 °C, 30 min, quant; (b) tributylamine,

THF, diphenylether, 200 °C, 30 min, 9.9%; (c) (2-cyclopropyl-3-methylimidazo[1,2-a]pyridine-6-yl)boronic acid, Cu(OAc)₂, pyridine, MS4A,

DMF, then 50 °C, overnight, 5%.

Figure 4. Lead identiﬁcation strategy via RHS exploration. A number of bicyclic motifs were designed on the basis of the illustrated general structure.

Chan−Lam−Evans coupling reaction. The reaction time of the

alkylation of 29 should be carefully controlled to prevent the

formation of double-alkylated side products. Pyrimidinone

derivative 35 was accessed from 2,4-dichloropyrimidine 32.

The ﬁrst S_NAr reaction occurred regioselectively at the 4-position

to give intermediate 33. Then, the remaining chloro atom at the

2-position was hydrolyzed to give pyrimidinone intermediate 34,

which was subject to the copper coupling reaction to aﬀord the

target compound 35.

Scheme 10 depicts the synthesis of tetrahydropyranopyridone

41. The Heck reaction of iodopyridine 36 with propen-1-ol gave

ketone 37, which was treated with LiBH₄to yield benzylalcohol

38. Intramolecular cyclization of 38 went immediately in good

yield via the deprotonation of alcohol with NaH to give

tetrahydropyranopyridine 39. Hydrolysis of the chloro atom of

39 followed by copper coupling reaction aﬀorded the target

compound 41.

Furopyridone 45 was accessible as illustrated in Scheme 11.

Acrylic acid 42 was converted to acid azide 43 via a mixed

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anhydride in excellent yield. Then, compound 43 was heated at

200 °C under a basic condition to give the 1-unsubstituted

furopyridone 44 via Curtius rearrangement, followed by

nucleophilic attack of the furan ring to the resulting isocyanate

intermediate. Chan−Lam−Evans coupling was found to be

suitable for 44, and it aﬀorded the target compound 45.

imidazopyridine ring would actively participate in the interaction

with the receptor. Compounds 6a and 6b were well ﬁtted to the

CNS-oriented chemical space deﬁned by four physicochemical

descriptors (PSA, ClogP, MW, and HBD count). In contrast, the

indolizine derivative 6c and the benzofuran derivative 6d did not

display binding aﬃnities. These results support our design

hypothesis, indicating that the nitrogen atom at the 1-position of

the imidazopyridine ring interacts with MCHR1, presumably

using Asp123 and/or Tyr272. From this RHS exploration

imidazo[1,2-a]pyridine was identiﬁed as a novel bicyclic motif

that possessed high intrinsic binding aﬃnity and did not require

the frequently used basic aliphatic amine motif.

Because aliphatic amides are chemically and biologically

unstable due to hydrolysis, we then cyclized the aliphatic amide

linkage of 6b and designed pyridin-2(1H)-ones (Figure 5).

Conformationally, 6b was considered to take two conformers, a

closed form and an open form (Figure 5), each leading to

pyridin-2(1H)-one with a substituent at the 4- and 3-position,

respectively (I and II). The calculation using Molecular

Operating Environment software (MOE)⁵⁵suggested that the

closed form was the most stable conformer because of hydrogen

bonding between the oxygen and the amide NH (ΔE = 3.98

kcal/mol). As 6b possessed excellent binding aﬃnity, we

assumed that the active conformation was similar to the

energetically stable closed form enabling compound 6b to bind

to the receptor with less entropy loss. Indeed, 4-substituted

pyridin-2(1H)-one 10a, designed on the basis of the closed form

of 6b, exhibited potent binding aﬃnity for both human and rat

MCHR1 (Table 2), whereas 3-substituted pyridin-2(1H)-ones

(24a and 24b) designed on the basis of the open form of 6b

caused a signiﬁcant decline in potency, supporting our design

hypothesis discussed in Figure 5.

RESULTS AND DISCUSSION

■

The binding aﬃnities of the compounds prepared in this study

toward hMCHR1 and rMCHR1 were evaluated in a binding

assay using radiolabeled [¹²⁵I]MCH (4−19) and membrane

fractions prepared from Chinese hamster ovary (CHO) cell lines

stably expressing MCHR1. Lead identiﬁcation activity was

conducted in the aforementioned deﬁned chemical space as a

guideline for amine-free CNS-penetrant compounds. We

hypothesized that the aliphatic amine motif could be removed

without a loss of binding aﬃnity if the bicyclic motif itself

interacted with the receptor through putative interactions with

Asp123 and/or Tyr272 via hydrogen bonding. On the basis of

this concept, we initiated our activities to design a new bicyclic

motif with high intrinsic binding aﬃnity in which installation of

the aliphatic amine was not necessary. Design of the bicyclic

motif was based on the general formula illustrated in Figure 4.

The motif possessed a 5−5, 5−6, or 6−6 bicyclic heteroaromatic

ring with a hydrogen-bonding acceptor (X) such as a nitrogen or

an oxygen atom for the putative interactions with Asp123 and/or

Tyr272 as well as alkyl chains (R¹and R²) for lipophilic

interactions. A number of corresponding amines I were then

coupled with existing LHSs in the general formulas A and B by a

condensation reaction, and the resulting compounds were

screened to examine the inﬂuence of each bicyclic motif on

potency. The representative results of this RHS exploration are

summarized in Table 1. Imidazo[1,2-a]pyridine derivatives (6a

and 6b) displayed IC₅₀values in the single-digit nanomolar range

for both species. Given the fact that these two compounds did

not possess the aliphatic amine motif, it was assumed that the

In contrast, the 4-phenoxy derivative 10b displayed signiﬁ-

cantly decreased potency relative to the 4-benzyloxy derivative

10a. Given this result, the lowest energy conformers

corresponding to 10a′ and 10b′, in which the imidazopyridine

ring was simpliﬁed with a methyl group for calculation costs,

were generated and compared with that of compound 6b′

(Figure 6). An overlay of the structures suggested that the

terminal aryl ring in 10a′ was well superposed with the OCF₃

moiety in 6b′, whereas a large energy barrier was expected for

10b′ to ﬁt the terminal aryl ring with the OCF₃moiety in 6b′,

which explains the lower binding aﬃnities of 10b compared with

those of 10a.

Table 1. In Vitro Binding Aﬃnity of Compounds 6a−d

With the pyridin-2(1H)-one-based amine-free MCHR1

antagonist 10a in hand, we next investigated the scope of the

central ring (Table 3). The pyridone ring of 10a was replaced

with a variety of other azines (28, 31, and 35), and their potencies

and ligand lipophilicity eﬃciencies (LLEs)⁵⁷are listed in the

Table 3. Pyridazin-6-one derivative 28 and pyrimidin-4-one

derivative 31 exhibited decreased LLE values, along with a

decline in binding aﬃnities. Conversely, introduction of a

nitrogen atom at the 3-position was tolerable in terms of potency

and decreased lipophilicity [log D_7.4= 3.2 (10a) and 2.8 (35)].

As a result, pyrimidin-2-one derivative 35 had a better LLE than

10a, suggesting the superior lead-likeness of 35. From the lead

identiﬁcation study described thus far, a pyridin-2(1H)-one

(10a) and pyrimidine-2-one (35) were discovered as novel

amine-free MCHR1 antagonists via cyclization of the initial

linear amide 6b, followed by modiﬁcation of the resultant

pyridin-2(1H)-one core.

a

IC₅₀values were calculated using an experiment performed in

b

During proﬁling of the azine derivatives shown in Table 3, we

found that pyrimidin-2-one derivative 35 displayed good

duplicate, with a standard deviation of 3-fold. Binding aﬃnity for

human MCHR1. Binding aﬃnity for rat MCHR1.

c

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Figure 5. Design of cyclic amides on the basis of 2 conformers of 6b. The energy barrier between the two conformers (ΔE value) was calculated using

MOE.⁵⁵

Table 2. In Vitro Binding Aﬃnity of Compounds 10a, 10b,

24a, and 24b

Table 3. In Vitro Binding Aﬃnity, log D, and LLE of

Compounds 10a, 28, 31, and 35

a

IC₅₀(nM)

b

c

a

compd

position

R

hMCHR1

rMCHR1

IC₅₀(nM)

b

c

d

10a

10b

24a

24b

4

3

OCH₂(4-Cl-Ph)

O(4-CF₃O-Ph)

OCH₂(4-Cl-Ph)

O(4-CF₃O-Ph)

26

>1000

990

20

950

650

240

compd

X

Y

Z

hMCHR1 rMCHR1 Log D_7.4

LLE

10a

28

CH

N

CH

N

CH

N

26

92

20

110

140

30

3.2

3.7

2.9

2.8

4.4

3.3

3.9

4.6

270

31

CH

150

37

a

IC₅₀values were calculated using an experiment performed in

duplicate, with a standard deviation of 3-fold. Binding aﬃnity for

human MCHR1. Binding aﬃnity for rat MCHR1.

35

CH

b

a

IC₅₀values were calculated using an experiment performed in

duplicate, with a standard deviation of 3-fold. Binding aﬃnity for

human MCHR1. Binding aﬃnity for rat MCHR1. The log D value at

c

b

c

d

pH 7.4.⁵⁶

potency, and its IC₅₀values were superior to those of the isomeric

pyrimidin-4-one derivative 31. Given the result that the

conformation of the LHS was crucial for potent aﬃnity (Figures

5 and 6), we hypothesized that these two compounds adopted

diﬀerent LHS conformations, with the LHS conformation of 35

being closer to that of the active conformer. As illustrated in

Figure 7A, repulsion of lone pairs on the oxygen atom and the

pyrimidine nitrogen evoked conformational preference, and

these two isomeric pyrimidine derivatives adopted diﬀerent

orientations of their LHSs. On the basis of this hypothesis, we

designed a pyridin-2(1H)-one derivative with a substituent at the

5-position and bicyclic derivatives, all of which would take the

Figure 6. Overlay of the lowest energy conformers of 6b′ (yellow), 10a′ (purple), and 10b′ (orange) using MOE⁵⁵(for the calculation cost, the

imidazopyridine ring was simpliﬁed with a methyl group).

H

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Figure 7. (A) Conformational preference of compounds 31 and 35 and prediction of the active conformer. (B) Design based on the putative active

conformer. The energy barrier between the two conformers (ΔE value) was calculated using MOE.⁵⁵

putative active conformation and would be expected to enhance

potency (Figure 7).

The 5-methylpyridione derivative 16 had decreased potency

relative to the lead compound 10a, indicating that the 5-position

possessed limited capacity to accommodate a substituent (Table

4). The tetrahydropyranopyridone derivative 41, in which the

group as in 41. As a result, the furopyridone derivative 45

displayed enhanced potency with an IC₅₀value for hMCHR1 in

the single-digit nanomolar range. These results indicated that

conformational restriction of the benzyloxy group in 10a to the

active conformer was an eﬀective strategy to enhance potency,

and the resulting furopyridone core was found to be a superior

scaﬀold to control the terminal aryl group.

Table 4. In Vitro Binding Aﬃnity of Compounds 16, 41, and

45

As discussed previously, novel amine-free MCHR1 antago-

nists (10a, 35, and 45) bearing an imidazo[1,2-a]pyridine ring as

the bicyclic motif were discovered, and these compounds

possessed good binding aﬃnity as well as MCHR1 antagonistic

activity in an MCH-stimulated Ca²⁺mobilization assay using

CHO cells (Figure 8). Among them, we selected pyridone 10a

for further optimization study because it displayed the most

potent antagonistic activity in CHO cells.

The results of the initial SAR study of the terminal benzene

ring are discussed using compounds 10c−g in Table 5. We

focused the SAR study with lipophilic substituents on the para

position of the terminal benzene ring by using the results of our

previous studies on dihydronaphthalenes⁴²and quinolines,^44,47

in which the LHS was expected to be surrounded by a

hydrophobic ﬁeld formed by Phe213, Ala216, Phe217 on

TM5, and Tyr273 on TM6.⁴²The unsubstituted benzene

derivative 10c exhibited a slight decline in in vitro potency

relative to the 4-chloro analogue 10a, and introduction of a

ﬂuorine atom to 10c did not aﬀect its binding aﬃnities (10d),

indicating that an electrostatic diﬀerence in the terminal benzene

ring does not aﬀect potency. When the chlorine atom of 10a was

replaced with a bromine atom, a 2-fold increase in potency was

observed (10e). As the SAR of 4-halogenated derivatives (10d,

10a, and 10e) positively depends on the van der Waals radius, we

subsequently introduced a triﬂuoromethyl or isopropyl group

with a larger van der Waals volume (10f and 10g) to enhance

potency. However, the triﬂuoromethyl derivative 10g exhibited a

slight decrease in potency, and the isopropyl derivative 10f had

an approximately 30-fold decline relative to 10e. These results

indicated that a bromine or chlorine atom was an appropriate

substituent at this position for binding aﬃnity.

a

IC₅₀values were calculated using an experiment performed in

b

duplicate, with a standard deviation of 3-fold. Binding aﬃnity for

human MCHR1. Binding aﬃnity for rat MCHR1.

c

terminal chlorophenyl group was shown to be superposed well

with 10a using MOE, had slightly decreased potency relative to

10a. This decline in potency was considered to have arisen from

the ethylene tether of 41, which would interfere in the interaction

with the receptor. On the basis of this assumption, the

tetrahydropyran moiety of 41 was replaced with a sterically less

congested furan ring that would ﬁx the terminal chlorophenyl

The result of the SAR study at the 3-position of the

imidazopyridine ring was consistent with the ﬁndings of the

study on currently reported analogous MCHR1 antagonists,⁵⁸

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Figure 8. Chemical structures and binding aﬃnities of lead compounds 10a, 35, and 45.

that a hydrogen-bonding donor or acceptor at this position

would function as the amine motif alternative and enhance

potency through interaction with the carboxamide of Asn294.

On the basis of this hypothesis, we explored substituents at the 2-

position. Replacement of the cyclopropyl group with a methyl or

ethyl group decreased potency by 2−3-fold and 3−4-fold,

respectively (10h and 10i). Then, a variety of polar substituents,

such as nitrile, carboxamide, and ketone, were introduced directly

at the 2-position (10j, 10k, 10m, and 10n). However, all of them

caused a signiﬁcant decline in potency. The loss of potencies

observed in 10j, 10k, 10m, and 10n was caused by a decrease in

the electron density on the nitrogen atom at the 1-position via

electron-withdrawing functional groups and concomitant loss of

the intrinsic binding aﬃnity of the imidazopyridine ring. This

consideration was also consistent with the calculated electron

densities using semiempirical PM3 (MOPAC) method⁵⁹in

MOE (10d, −0.104; 10j, −0.074; 10k, −0.089; 10m, −0.081;

10n, −0.076; Figure 9). Given these results, we subsequently

designed 10o−q, which possessed a similar calculated electron

density on the nitrogen atom as 10d (10d, −0.104; 10o, −0.108;

10p, −0.103; 10q, −0.097) and presumably retained the binding

aﬃnity of the imidazopyridine ring. However, they all displayed

decreased aﬃnity for MCHR1 and only the cyanomethyl

derivative 10q had IC₅₀values in the submicromolar range,

indicating that introduction of polar functional groups at the 2-

position would have an unfavorable eﬀect on interactions with

the receptor. Consequently, the cyclopropyl group was

determined to be an optimal substituent at this position.

Introduction of polar substituents as the amine motif alternative

aimed at putative hydrogen bonding with Asn294 was not

successful.

Table 5. In Vitro Binding Aﬃnity of Compounds 10c−k and

10m−q

We subsequently replaced the terminal aryl group on the LHS

with a variety of heteroaryl groups for precise optimization

(Table 6). First, unsubstituted heteroaryl rings were introduced

(10r−w). Among the pyridyl derivatives, the 2-pyridyl derivative

10r was superior to 3-pyridyl and 4-pyridyl derivatives (10s and

10t), and 10r displayed IC₅₀values of 240 and 150 nM for

hMCHR1 and rMCHR1, respectively. The more polar

pyrimidine derivative 10u exhibited signiﬁcantly lower activities.

Although the tolerability of a nitrogen atom diﬀered depending

on the position, the overall lipophilicity preference at this

position was consistent with our previous ﬁndings.⁴²On the

contrary, thienyl derivatives (10v and 10w) had relatively good

binding aﬃnities, and the 3-thienyl derivative 10w displayed a

superior IC₅₀for hMCHR1 than the unsubstituted benzene

derivative 10c (25 vs 42 nM).

We next introduced a lipophilic substituent on these

heteroaryl rings to ﬁt LHS to the hydrophobic ﬁeld more

eﬃciently (10x−z). The 3-chloropyridine derivative 10x

exhibited superior potency to the unsubstituted derivative 10r,

and its low lipophilic nature resulted in a better LLE value than

the lead compound 10a [LLE = 5.2 (10x) and 4.4 (10a)]. The

chlorothiophene derivative 10y exhibited excellent binding

a

IC₅₀values were calculated using an experiment performed in

b

duplicate, with a standard deviation of 3-fold. Binding aﬃnity for

human MCHR1. Binding aﬃnity for rat MCHR1.

c

and it revealed that the methyl group was essential for MCHR1

binding aﬃnity. Conversely, introduction of polar functional

groups, such as an alkoxy or an alcohol group, decreased aﬃnity

(data not shown).

Next, we modiﬁed substituents on the 2-position of the

imidazopyridine ring (10h−k and 10m−q in Table 5). The

substituent at the 2-position was assumed to be exposed to the

space in which the aliphatic amines of basic compounds (1−3)

interacted with Asn294 (Figure 3). Therefore, we hypothesized

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Figure 9. Calculated electron densities on the nitrogen atom on the imidazopyridine ring of 10d, 10j, 10k, 10m, and 10n.

chlorothiophene derivative 10y with excellent in vitro potency.

This thiophene derivative also displayed excellent MCHR1

antagonistic activity in an MCH-stimulated Ca²⁺mobilization

assay using CHO cells (10y: IC₅₀= 19 nM).

Table 6. In Vitro Binding Aﬃnity of Compounds 10r−z

To validate our concept of drug design, lead compound 10a,

which displayed superior metabolic stability to the thiophene

derivative 10y, was selected for further proﬁling. Compound 10a

possesses a pK_avalue⁶⁰of 7.9 and the following physicochemical

properties: TPSA⁶¹= 49, ClogP = 5.0, MW = 405, and HBD

count = 0. These ﬁndings indicate that 10a is roughly within the

initially set target chemical space. Compound 10a did not display

phospholipidosis potential in an in vitro assay or hERG

inhibition potential in a patch clamp test (IC₅₀> 10 μM).

Pharmacokinetic screening in rats indicated that 10a was orally

available with acceptable plasma exposure (Table 7). To evaluate

its pharmacological eﬀect, compound 10a (3 and 10 mg/kg) was

orally administered to diet-induced obese (DIO) F344 rats fed a

high-fat diet ad libitum. Compound 10a exhibited a dose-

dependent anorectic eﬀect in DIO F344 rats. The change in the

cumulative food intake for 2 days compared with the vehicle

group was −15.1% and −29.6% at 3 and 10 mg/kg, respectively

(Figure 10). The plasma and brain concentrations, monitored at

24 h after dosing in day 1, were 351 and 218 ng/mL, respectively,

at 3 mg/kg and 1350 and 841 ng/mL, respectively, at 10 mg/kg

(BP ratios: 0.61 and 0.62, respectively). Because free fraction of

10a in DIO rat plasma was 0.02, plasma free concentration were

calculated to be 7.0 and 27 ng/mL (17.2 and 66.5 nM) at 3 and

10 mg/kg, respectively, meaning 0.86·IC₅₀and 3.3·IC₅₀free

concentration were maintained at 24 h in day 1. Then eﬀect of

10a on body weight reduction was assessed using 2-week

repeated dosing study (Figure 11). Signiﬁcant change in body

weight was observed from 3 mg/kg, and once daily oral

administration of 3 and 10 mg/kg of 10a resulted in 3.7% and

8.6% body weight reduction, respectively, relative to vehicle.

Cumulative food intake of these two groups were signiﬁcantly

reduced by 15.4% and 36.2%, respectively. It was noteworthy that

3 mg/kg of 10a showed equipotent antiobesity eﬀect to 1 mg/kg

of sibutramine (3.7% vs 3.4%), while anorectic eﬀect of 10a was

milder than sibutramine group (15.4% vs 21.5%). Involvement of

MCHR1 antagonists in energy expenditure have been

a

IC₅₀values were calculated using an experiment performed in

b

duplicate, with a standard deviation of 3-fold. Binding aﬃnity for

human MCHR1. Binding aﬃnity for rat MCHR1.

c

aﬃnities in both species, and its IC₅₀values successfully reached

the single-digit nanomolar range, representing an approximately

3-fold increase relative to the lead compound 10a. Replacement

of the chlorine atom on the thiophenes with a triﬂuoromethyl

group resulted in slightly decreased potency; however,

compound 10z exhibited superior potency to 10a. From an

optimization study to enhance the potency of the lead compound

10a discussed in Tables 5 and 6, we successfully discovered the

a

Table 7. Pharmacokinetic Parameters of 10a in Rats

iv (0.1 mg/kg)

po (1 mg/kg)

b

c

d

e

f

g

compd

F

(%)

58

CL_total(mL·h⁻¹·kg⁻¹

)

V_ss(mL·kg⁻¹

)

C_max(ng·mL⁻¹

)

T_max(h)

AUC_{0−8 h}(ng·h·mL⁻¹

)

10a

312

1053

313

2.0

1880

a

b

c

d

e

f

n = 3; SD rats (male, 8 weeks old). Bioavailability. Total clearance. Volume of distribution at steady state. Maximal plasma concentration. Time

g

of maximal concentration. Area under the plasma concentration−time curve (0−8 h).

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Figure 12. Results of 10a in a 3-day food intake study in MCHR1-

deﬁcient and wild-type mice. The mice were fed a high-fat diet. The

cumulative food intake was measured for 3 days. Each data represents

mean SD (n = 5 or 6 for each group). (#) p < 0.025 vs the vehicle

group (Williams test).

Figure 10. Results of 10a in a 2-day food intake study in DIO F344 rats.

Inhibition of cumulative food intake for 2 days in DIO F344 rats. The

compound was administered once daily, and food intake from the initial

administration to 2 days later was measured. The cumulative food intake

inhibition rate was calculated by dividing the average food intake of each

treatment group by that of the vehicle group. Each data represents mean

SD (n = 6 for each group). (#) p < 0.025 vs the vehicle group

(Williams test).

identiﬁcation strategy is an eﬀective approach for discovering

novel MCHR1 antagonists with superior safety proﬁles to

existing antagonists.

CONCLUSION

■

To discover a potent MCHR1 antagonist with an improved

safety proﬁle, we designed new aliphatic amine-free structures

with physicochemical properties for a high probability of CNS

penetration. We excluded the aliphatic amine motif, which has

been widely recognized as part of the pharmacophore of a small-

molecule MCHR1 antagonist, and designed structures outside

the hERG chemical space together with a cutoﬀ value of basicity,

pK_a, of <8. To compensate for the concomitant potential loss for

MCHR1, identiﬁcation of novel bicyclic motifs that can interact

with Asp123 or Tyr272 located nearby in our previously reported

MCHR1 homology model is considered critical. We set the

physicochemical descriptors (PSA < 70, 2 < ClogP < 4, MW <

450, and HBD count = 0 or 1) for CNS-penetrant compounds.

As a result, we discovered a lead compound, 4-[(4-

chlorobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-a]-

pyridin-6-yl)pyridin-2(1H)-one 10a, which was conﬁrmed to be

negative in hERG and phospholipidosis screening. Following

oral administration revealed that 10a was suﬃciently exposed to

the brain and dose-dependently exerted anorectic eﬀect as well as

body weight reduction by 3.7% and 8.6% at doses of 3 and 10

mg/kg, respectively, in a 2-week repeated dosing study using

reported,⁶²and this would bring additional beneﬁt on antiobesity

eﬀect of 10a, resulting in better body weight loss relative to

sibutramine.

To clarify the selectivity of the anorectic property to MCHR1

antagonism, the eﬀect of 10a on food intake in MCHR1-deﬁcient

and wild-type mice was examined. Compound 10a exerted no

eﬀect on food intake in MCHR-deﬁcient mice, whereas it

signiﬁcantly reduced food intake in wild-type mice in a dose-

dependent manner (Figure 12), proving that the anorectic eﬀect

and the resulting weight loss of 10a were based on its MCHR1

antagonistic activity. These ﬁndings indicated that our lead

identiﬁcation strategy using ﬁve physicochemical descriptors

(pK_a, PSA, ClogP, MW, and HBD count) as the design guideline

successfully aﬀorded the CNS-penetrant MCHR1 antagonist

10a without hERG and phospholipidosis liabilities, and the

compound displayed good brain exposure and a resulting

anorectic eﬀect. It is noteworthy that installation of the

imidazopyridine ring in RHS enabled the novel design of

amine-free MCHR1 antagonists excluding the aliphatic amine

motif. Consequently, these ﬁndings suggest that our lead

Figure 11. Results of 10a in a repeated-dose study in DIO F344 rats. (A) Body weight change from initial during 2 weeks of dosing. (B) Cumulative food

intake for 2 weeks of dosing. The compounds were administered once daily for 2 weeks, and body weight and food intake were measured before drug

administration. Each data represents mean SD (n = 6 for each group). (#) p < 0.025 vs the vehicle group (Williams test), (**) p < 0.01 vs vehicle group

(Student’s t test).

L

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122.4, 124.9, 139.3, 140.7, 142.2, 156.6, 166.4. MS (ESI/APCI) m/z

380.1 [M + H]⁺. Anal. Calcd for C₁₈H₁₆F₃N₃O₃: C, 56.99; H, 4.25; N,

11.08. Found: C, 56.92; H, 4.26; N, 11.03.

DIO F344 rats. Because 10a exerted no eﬀect on food intake in

MCHR1-deﬁcient mice, the anorectic eﬀect and the resulting

weight loss of 10a were considered to be based on its MCHR1

antagonistic activity. These results indicated that our lead

identiﬁcation strategy successfully aﬀorded potent, orally

available aliphatic amine-free MCHR1 antagonists with attractive

safety proﬁles and suﬃcient brain exposure. The newly designed

1-(imidazo[1,2-a]pyridin-6-yl)-pyridine-2(1H)-ones were novel

lead compounds in this program for further exploration of potent

MCHR1 antagonists.

N-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-2-[4-

(triﬂuoromethoxy)phenoxy]acetamide (6b). The title compound was

prepared in 63% yield using 2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-amine in an analogous manner to 6a. White solid; mp 158

°C. ¹H NMR (400 MHz, DMSO-d₆) δ 0.82−0.92 (4H, m), 1.97−2.11

(1H, m), 2.42 (3H, s), 4.78 (2H, s), 7.12 (2H, d, J = 9.4 Hz), 7.20 (1H,

dd, J = 9.4, 1.9 Hz), 7.31−7.43 (3H, m), 8.72 (1H, s), 10.22 (1H, s). ¹³C

NMR (75 MHz, DMSO-d₆) δ 7.8, 7.9, 8.0, 67.2, 113.7, 115.6, 115.9,

116.0, 118.4, 120.6 (q, J = 253.5 Hz), 122.5, 124.9, 140.8, 142.2, 144.4,

156.6, 166.4. MS (ESI/APCI) m/z 406.1 [M + H]⁺. Anal. Calcd for

C₂₀H₁₈F₃N₃O₃: C, 59.26; H, 4.48; N, 10.37. Found: C, 59.02; H, 4.49;

N, 10.29.

N-(2-Cyclopropyl-3-methylindolizin-6-yl)-2-[4-(triﬂuoromethoxy)-

phenoxy]acetamide (6c). The title compound was prepared in 24%

yield using 2-cyclopropyl-3-methylindolizin-6-amine hydrochloride in

an analogous manner to 6a. Oﬀ-white solid; mp 168−170 °C. ¹H NMR

(300 MHz, DMSO-d₆) δ 0.45−0.62 (2H, m), 0.83−0.95 (2H, m), 1.84−

1.97 (1H, m), 2.39 (3H, s), 4.75 (2H, s), 5.97 (1H, s), 6.69 (1H, dd, J =

9.5, 1.9 Hz), 7.04−7.16 (2H, m), 7.26 (1H, d, J = 9.5 Hz), 7.34 (2H, d, J

= 8.3 Hz), 8.50 (1H, s), 9.97 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ

7.4, 8.3, 9.2, 67.3, 94.7, 111.7, 112.4, 116.0, 117.6, 117.8, 120.1 (q, J =

253.5 Hz), 122.5, 123.4, 128.57, 128.61, 142.1, 142.2, 156.7, 166.1. MS

(ESI/APCI) m/z 405.2 [M + H]⁺. Anal. Calcd for C₂₁H₁₉F₃N₂O₃: C,

62.37; H, 4.74; N, 6.93. Found: C, 62.24; H, 4.71; N, 6.92.

N-(2,3-Dimethyl-1-benzofuran-5-yl)-2-[4-(triﬂuoromethoxy)-

phenoxy]acetamide (6d). The title compound was prepared in 62%

yield using 2,3-dimethyl-1-benzofuran-5-amine in an analogous manner

to 6a; mp 156−159 °C. ¹H NMR (400 MHz, DMSO-d₆) δ 2.09 (3H, s),

2.35 (3H, s), 4.74 (2H, s), 7.01 (2H, d, J = 9.0 Hz), 7.32−7.39 (4H, m),

7.81 (1H, s), 10.08 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.5, 11.6,

67.4, 109.6, 109.9, 110.1, 116.0, 116.1, 120.1 (q, J = 253.5 Hz), 122.5,

129.9, 133.2, 142.11, 142.13, 149.8, 151.3, 156.7, 165.9. MS (ESI/APCI)

m/z 380.2 [M + H]⁺. Anal. Calcd for C₁₉H₁₆F₃NO₄: C, 60.16; H, 4.25;

N, 3.69. Found: C, 59.89; H, 4.26; N, 3.69.

4-[(4-Fluorobenzyl)oxy]pyridine 1-Oxide (8a). The title compound

was prepared in 6% yield using (4-ﬂuorophenyl)methanol in an

analogous manner to 8b. ¹H NMR (300 MHz, DMSO-d₆) δ 5.15 (2H,

s), 7.04−7.13 (2H, m), 7.19−7.30 (2H, m), 7.46−7.57 (2H, m), 8.07−

8.14 (2H, m). MS (ESI/APCI) m/z 220.1 [M + H]⁺.

4-[(4-Chlorobenzyl)oxy]pyridine 1-Oxide (8b). A solution of (4-

chloropheny)methanol (49.5 g, 347 mmol) in THF (200 mL) was

added dropwise to a suspension of NaH (60% oil dispersion, 16.7 g, 419

mmol) in THF (200 mL) at 0 °C. After the mixture was stirred at 0 °C

for 30 min, 7b (45.0 g, 347 mmol) was added portionwise to the reaction

mixture. After completion of the addition, the mixture was stirred at rt

for 5 h. The mixture was quenched with water (400 mL) at 0 °C and

extracted with EtOAc/THF (1:1) four times. The organic layers were

combined, passed through NH-silica gel pad (EtOAc/MeOH), and

concentrated. The ﬁltrate was concentrated, and the residual solid was

washed with IPE and dried to give the title compound (54.3 g, 66%) as a

brown solid. ¹H NMR (400 MHz, CDCl₃) δ 5.17 (2H, s), 7.08 (2H, d, J

= 6.9 Hz), 7.48 (4H, s), 8.10 (2H, d, J = 7.0 Hz). MS (ESI/APCI) m/z

236.0 [M + H]⁺.

4-{[4-(Triﬂuoromethyl)benzyl]oxy}pyridine 1-oxide (8c). The title

compound was prepared in 29% yield using [4-(triﬂuoromethyl)-

phenyl]methanol in an analogous manner to 8b. Yellow solid. ¹H NMR

(400 MHz, DMSO-d₆) δ 5.29 (2H, s), 7.10 (2H, d, J = 7.6 Hz), 7.67

(2H, d, J = 8.0 Hz), 7.78 (2H, d, J = 8.4 Hz), 8.11 (2H, d, J = 7.6 Hz). MS

(ESI/APCI) m/z = 270.2 [M + H]⁺.

4-[(4-Fluorobenzyl)oxy]pyridine-2(1H)-one (9a). The title com-

pound was prepared in 58% yield using 8a in an analogous manner to 9b.

¹H NMR (300 MHz, DMSO-d₆) δ 5.04 (2H, s), 5.78 (1H, d, J = 2.3 Hz),

5.90 (1H, dd, J = 7.2, 2.7 Hz), 7.17−7.28 (3H, m), 7.43−7.53 (2H, m),

11.10 (1H, br s). ¹³C NMR (75 MHz, DMSO-d₆) δ 68.5, 97.9, 99.1,

115.3 (d, J = 21 Hz), 130.2 (d, J = 8.3 Hz), 132.2 (d, J = 3 Hz), 135.4,

161.9 (d, J = 242.3 Hz), 164.0, 167.3. MS (ESI/APCI) m/z = 220.1 [M +

H]⁺.

EXPERIMENTAL SECTION

■

General. Melting points were determined on a Yanaco melting point

apparatus Mp-500D and are uncorrected. H NMR and ¹³C NMR

1

spectra were recorded on a Bruker AVANCE III (300 MHz) or a Bruker

Advance III plus (400 MHz) spectrometer. Chemical shifts are given in

parts per million (ppm) downﬁeld from tetramethysilane (δ) as the

internal standard in deuterated solvent, and coupling constants (J) are in

hertz (Hz). Data are reported as follows: chemical shift, integration,

multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, quint =

quintet, m = multiplet, dd = doublet of doublets, td = triplet of doublets,

and bs = broad signal), and coupling constants. Protons of the methyl

group on the 3 position of the imidazopyridine ring were frequently

merged with solvent peak of DMSO and not observed. Reagents and

solvents were obtained from commercial sources and used without

further puriﬁcation. Reaction progress was determined by thin layer

chromatography (TLC) analysis on Merck Kieselgel 60 F254 plates or

Fuji Silysia NH plates. Chromatographic puriﬁcation was performed on

silica gel columns [(Merck Kieselgel 60, 70−230 mesh size or 230−400

mesh size, Merck) or (Chromatorex NH-DM 1020, 100−200 mesh

size)] or on Purif-Pack (SI or NH, particle size: 60 μm, Fuji Silysia

Chemical, Ltd.). LC−MS analysis was performed on a Shimadzu liquid

chromatography−mass spectrometer system, operating in APCI (+ or

−) or ESI (+ or −) ionization mode. Analytes were eluted using a linear

gradient of 0.05% TFA containing water/acetonitrile or 5 mM

ammonium acetate containing water/acetonitrile mobile phase and

detected at 220 nm. Analytical HPLC was performed with Corona

Charged Aerosol Detector (CAD) or photo diode array detector. The

column was a Capcell Pak C18AQ (50 mm × 3.0 mm I.D., Shiseido,

Japan) or L-column 2 ODS (30 mm × 2.0 mm I.D., CERI, Japan) with a

temperature of 50 °C and a ﬂow rate of 0.5 mL/min. Mobile phase A and

B under a neutral condition were a mixture of 50 mmol/L ammonium

acetate, water, and acetonitrile (1:8:1, v/v/v) and a mixture of 50 mmol/

L ammonium acetate and acetonitrile (1:9, v/v), respectively. The ratio

of mobile phase B was increased linearly from 5% to 95% over 3 min,

95% over the next 1 min. Mobile phase A and B under an acidic

condition were a mixture of 0.2% formic acid in 10 mmol/L ammonium

formate and 0.2% formic acid in acetonitrile, respectively. The ratio of

mobile phase B was increased linearly from 14% to 86% over 3 min, 86%

over the next 1 min. The purities of compounds submitted for biological

evaluation were >95% as determined by elemental analyses within

0.4% of the calculated values or analytical HPLC. Yields are not

optimized.

N-(2, 3-Dimethylimidazo[1, 2-a]pyridin-6-yl)-2-[4-

(triﬂuoromethoxy)phenoxy]acetamide (6a). A mixture of 2-[4-

(triﬂuoromethoxy)phenoxy]acetic acid (146 mg, 0.62 mmol), 2-

methyl-3-methylimidazo[1,2-a]pyridin-6-amine (100 mg, 0.62 mmol),

WSC (238 mg, 12.4 mmol), HOBt (16.8 mg, 0.12 mmol), and DMF (4

mL) was stirred at rt overnight. The mixture was poured into water and

extracted with EtOAc. The extract was washed with brine, dried over

MgSO₄, concentrated, and puriﬁed by column chromatography (NH

silica gel, hexane/EtOAc = 90/10 to 50/50) followed by recrystallization

from EtOAc−hexane to give the title compound 6a (89.2 mg, 38%) as

an oﬀ-white solid; mp 202−204 °C. ¹H NMR (300 MHz, DMSO-d₆) δ

2.29 (3H, s), 2.34 (3H, s), 4.79 (2H, s), 7.08−7.16 (2H, m), 7.22 (1H,

dd, J = 9.4, 1.9 Hz), 7.34 (2H, d, J = 8.3 Hz), 7.38−7.49 (1H, m), 8.72

(1H, d, J = 1.1 Hz), 10.23 (1H, s). ¹³C NMR (101 MHz, DMSO-d₆) δ

7.9, 13.1, 67.3, 114.0, 115.6, 116.0, 116.1, 118.5, 120.1 (q, J = 256.5 Hz),

M

DOI: 10.1021/acs.jmedchem.5b01704

J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

4-[(4-Chlorobenzyl)oxy]pyridine-2(1H)-one (9b). A mixture of 8b

(54.3 g, 230 mmol) and acetic anhydride (540 mL, 5.71 mol) was stirred

at 140 °C for 2 h. After concentration of the mixture, the residue was

dissolved in MeOH (300 mL). Water (450 mL) was added to the

mixture, followed by stirring at rt for 1 h. The resulting precipitate was

collected by ﬁltration, washed with IPA, and dried to give the title

compound (29.3 g, 54%) as a gray solid. ¹H NMR (300 MHz, CDCl₃) δ

4.99 (2H, s), 5.93 (1H, d, J = 2.3 Hz), 6.03 (1H, dd, J = 7.4, 2.5 Hz), 7.23

(1H, d, J = 7.2 Hz), 7.29−7.44 (4H, m). ¹³C NMR (75 MHz, DMSO-d₆)

δ 68.4, 98.0, 99.1, 128.5, 129.7, 132.7, 135.0, 135.5, 163.9, 167.2. MS

(ESI/APCI) m/z 236.0 [M + H]⁺.

4-{[4-(Triﬂuoromethyl)benzyl]oxy}pyridin-2(1H)-one (9c). The title

compound was prepared in 31% yield using 8d in an analogous manner

to 9b. Oﬀ-white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 5.19 (2H, s),

5.77 (1H, d, J = 2.4 Hz), 5.93 (1H, dd, J = 7.2, 2.4 Hz), 7.26 (1H, d, J =

7.6 Hz), 7.64 (2H, d, J = 8.0 Hz), 7.77 (2H, d, J = 8.4 Hz), 11.12 (1H, s).

MS (ESI/APCI) m/z 270.0 [M + H]⁺.

4-[(4-Chlorobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)pyridin-2(1H)-one (10a). Two batches of a mixture of 9b

(1.00 g, 4.24 mmol), 11a (1.27 g, 4.24 mmol), DMEDA (0.48 mL, 4.24

mmol), CuI (0.81 g, 4.24 mmol), K₂CO₃(1.76 g, 12.7 mmol), and

DMSO (15 mL) were heated at 150 °C for 3 h. The two batches were

combined and poured into 14% NH₃solution. The mixture was

extracted with EtOAc/THF (1:1). The extract was washed with brine,

dried over MgSO₄, concentrated, and puriﬁed by column chromatog-

raphy (silica gel, hexane/EtOAc = 90/10 to 0/100, then EtOAc/MeOH

= 100/0 to 85/15) followed by column chromatography (NH silica gel,

hexane/EtOAc = 90/10 to 0/100) to give a crude product (350 mg).

A mixture of 9b (200 mg, 0.85 mmol), 11a (253 mg, 0.85 mmol),

DMEDA (95.0 μL, 0.85 mmol), CuI (162 mg, 0.85 mmol), K₂CO₃(352

mg, 2.55 mmol), and DMSO (3 mL) was heated at 110 °C under

microwave irradiation for 1 h. Another two batches of a mixture of 9b

(1.00 g, 4.24 mmol), 11a (1.27 g, 4.24 mmol), DMEDA (0.48 mL, 4.24

mmol), CuI (0.81 g, 4.24 mmol), K₂CO₃(1.76 mg, 12.7 mmol), and

DMSO (13 mL) were heated at 110 °C under microwave irradiation for

1 h. The reaction mixtures were combined and poured into 14% NH₃

solution. The mixture was extracted with EtOAc/THF (1:1). The

extract was washed with brine, dried over MgSO₄, concentrated, and

puriﬁed by column chromatography (silica gel, hexane/EtOAc = 90/10

to 0/100, then EtOAc/MeOH = 100/0 to 85/15) followed by column

chromatography (NH silica gel, hexane/EtOAc = 90/10 to 0/100) to

give a crude product (950 mg).

4-(Benzyloxy)-1-(2-cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-

yl)pyridin-2(1H)-one (10c). A mixture of 11a (1.48 g, 4.97 mmol), 4-

(benzyloxy)pyridine-2(1H)-one (1.00 g, 4.97 mmol), CuI (946 mg,

4.97 mmol), DMEDA (0.534 mL, 4.97 mmol), K₂CO₃(2.06 g, 14.9

mmol), and DMSO (13 mL) was heated at 110 °C for 1 h under

microwave irradiation. The mixture was quenched with 28% NH₃

solution, and extracted with EtOAc/THF (1:1). The organic layer

was separated, washed with water and brine, dried over MgSO₄, passed

through silica gel pad (EtOAc), and concentrated in vacuo. The residue

was puriﬁed by silica gel column chromatography (silica gel, EtOAc/

MeOH = 100/0 to 85/15) and recrystallized from EtOH (80 mL) to

give the title compound (1.03 g, 56%) as an oﬀ-white solid; mp 233−235

°C. ¹H NMR (300 MHz, DMSO-d₆) δ 0.83−0.96 (4H, m), 2.02−2.12

(1H, m), 2.47 (3H, s), 5.16 (2H, s), 6.01 (1H, d, J = 2.6 Hz), 6.09−6.18

(1H, m), 7.11 (1H, dd, J = 9.4, 1.9 Hz), 7.32−7.53 (6H, m), 7.66 (1H, d,

J = 7.5 Hz), 8.37 (1H, d, J = 1.1 Hz). ¹³C NMR (75 MHz, DMSO-d₆) δ

7.8, 7.9, 8.1, 69.7, 97.7, 100.3, 114.8, 116.6, 121.9, 123.2, 127.2, 127.9,

128.2, 128.5, 135.8, 139.7, 141.8, 145.1, 162.8, 167.2. MS (ESI/APCI)

m/z 372.2 [M + H]⁺. Anal. Calcd for C₂₃H₂₁N₃O₂: C, 74.37; H, 5.70; N,

11.31. Found: C, 74.21; H, 5.65; N, 11.28.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-((4-

ﬂuorobenzyl)oxy)pyridin-2(1H)-one (10d). To a stirred mixture of 9a

(200 mg, 0.912 mmol), 11a (285 mg, 0.956 mmol), and K₂CO₃(378

mg, 2.73 mmol) in DMF (6 mL) was added CuI (34.7 mg, 0.182 mmol)

and trans-N,N′-dimethylcyclohexane-1,2-diamine (26 mg, 0.182 mmol).

The reaction vessel was sealed and heated at 130 °C for 16 h. DMF was

removed in vacuo, and then the residue was partitioned between EtOAc

(100 mL) and water (20 mL). The organic layer was washed with brine,

dried over Na₂SO₄, and concentrated in vacuo. Two batches were

combined and puriﬁed by preparative HPLC to give the title compound

1

(80 mg, 11%) as an oﬀ-white solid; mp 233−235 °C. H NMR (300

MHz, CDOD₃) δ 0.95−0.99 (4H, m), 2.06 (1H, m), 2.51 (3H, s), 5.14

(2H, s), 6.11 (1H, d, J = 2.6 Hz), 6.28 (1H, dd, J = 7.6, 2.6 Hz), 7.14 (2H,

t, J = 8.7 Hz), 7.21 (1H, dd, J = 9.4, 1.9 Hz), 7.46−7.51 (3H, m), 7.60

(1H, d, J = 7.6 Hz), 8.33 (1H, d, J = 1.3 Hz). ¹³C NMR (101 MHz,

DMSO-d₆) δ 7.8, 7.9, 8.0, 68.9, 97.6, 100.2, 115.3 (d, J = 21.2 Hz), 114.8,

116.5, 121.9, 123.1, 127.2, 130.2 (d, J = 9.1 Hz), 132.0 (d, J = 3.0 Hz),

139.7, 141.9, 145.0, 161.9 (d, J = 245.4 Hz), 162.7, 167.1. MS (ESI/

APCI) m/z 389.8 [M + H]⁺. Purity >99.9% (HPLC).

4-[(4-Bromobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)pyridin-2(1H)-one (10e). To a solution of 17 (500 mg,

1.78 mmol) in THF (5 mL) was added 4-bromobenzyl alcohol (332 mg,

1.78 mmol), (E)-bis(2-methoxyethyl) diazene-1,2-dicarboxylate (541

mg, 2.31 mmol), and PPh₃(606 mg, 2.31 mmol) at rt, and the mixture

was stirred for 16 h at rt. The mixture was poured into water and

extracted with EtOAc. The organic layer was separated, washed with

water and brine, dried over MgSO₄, and concentrated in vacuo. The

residue was puriﬁed by column chromatography (silica gel, EtOAc/

MeOH = 90/10), and the product was crystallized from EtOAc to give

the title compound (502 mg, 63%) as white crystals; mp 228−229 °C.

¹H NMR (400 MHz, DMSO-d₆) δ 0.82−0.96 (4H, m), 2.07 (1H, br s),

2.47 (3H, s), 5.15 (2H, s), 5.99 (1H, d, J = 2.4 Hz), 6.14 (1H, dd, J = 7.9,

2.5 Hz), 7.06−7.15 (1H, m), 7.43 (3H, d, J = 8.8 Hz), 7.59−7.69 (3H,

m), 8.37 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 68.8,

97.7, 100.2, 114.9, 116.6, 121.3, 121.9, 123.2, 127.2, 130.0, 131.4, 131.5,

135.3, 139.8, 141.9, 145.1. MS (ESI/APCI) m/z 450.3 [M + H]⁺. Anal.

Calcd for C₂₃H₂₀BrN₃O₂: C, 61.34; H, 4.48; N, 9.33. Found: C, 61.14;

H, 4.40; N, 9.25.

These two crude lots were combined, and recrystallized from EtOH−

water to give the title compound (1.25 g, 17% in total) as a white solid;

1

mp 230−232 °C. H NMR (300 MHz, DMSO-d₆) δ 0.84−0.96 (4H,

m), 2.01−2.13 (1H, m), 2.47 (3H, s), 5.16 (2H, s), 6.00 (1H, d, J = 2.6

Hz), 6.14 (1H, dd, J = 7.6, 2.6 Hz), 7.11 (1H, dd, J = 9.4, 1.9 Hz), 7.43

(1H, d, J = 9.4 Hz), 7.50 (4H, s), 7.66 (1H, d, J = 7.6 Hz), 8.36 (1H, d, J =

1.1 Hz). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 68.8, 97.7, 100.2,

114.9, 116.6, 121.9, 123.2, 127.2, 128.5, 129.7, 132.8, 134.9, 139.7, 141.9,

145.0, 162.8, 167.0. MS (ESI/APCI) m/z 406.1 [M + H]⁺. Anal. Calcd

for C₂₃H₂₀ClN₃O₂·H₂O: C, 65.17; H, 5.23; N, 9.91. Found: C, 65.09; H,

5.24; N, 9.87.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-[4-

(triﬂuoromethoxy)phenoxy]pyridin-2(1H)-one (10b). A mixture of 18

(100 mg, 0.29 mmol), 4-(triﬂuoromethoxy)phenol (103 mg, 0.58

mmol), DMEDA (31.0 μL, 0.29 mmol), K₂CO₃(120 mg, 0.87 mmol),

CuI (55.3 mg, 0.29 mmol), and DMSO (3 mL) was heated at 150 °C for

3 h. The mixture was puriﬁed by column chromatography (NH silica gel,

hexane/EtOAc = 90/10 to 0/100) to give the title compound (39.6 mg,

31%) as a white solid; mp 227−228 °C. ¹H NMR (400 MHz, DMSO-

d₆) δ 0.85−0.96 (4H, m), 2.08 (1H, br s), 5.58 (1H, d, J = 2.4 Hz), 6.26

(1H, dd, J = 7.4, 2.6 Hz), 7.13 (1H, d, J = 9.5 Hz), 7.35−7.48 (3H, m),

7.53 (2H, d, J = 8.7 Hz), 7.82 (1H, d, J = 7.5 Hz), 8.41 (1H, s). ¹³C NMR

(75 MHz, DMSO-d₆) δ 7.8, 7.97, 8.05, 99.5, 100.6, 115.0, 116.6, 120.0

(q, J = 254.3 Hz),122.0, 122.8, 123.0, 123.2, 127.0, 141.2, 141.9, 145.2,

145.6, 151.7, 162.4, 167.1. MS (ESI/APCI) m/z 442.3 [M + H]⁺. Anal.

Calcd for C₂₃H₁₈F₃N₃O₃: C, 62.58; H, 4.11; N, 9.52. Found: C, 62.46;

H, 4.10; N, 9.48.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-{[4-

(propan-2-yl)benzyl]oxy}pyridin-2(1H)-one (10f). To a solution of 17

(200 mg, 0.71 mmol) in DMF (3 mL) was added 1-(bromomethyl)-4-

(tert-butyl)benzene (0.131 mL, 0.78 mmol) and K₂CO₃(197 mg, 1.42

mmol), and the mixture was stirred at rt for 16 h. The mixture was

poured into water, and the resultant solid was collected by ﬁltration. The

solid was recrystallized from acetone−MeOH to give the title

1

compound (114 mg, 39%) as white crystals; mp 244−246 °C. H

NMR (400 MHz, DMSO-d₆) δ 0.84−0.99 (4H, m), 1.21 (6H, d, J = 6.9

Hz), 2.08 (1H, d, J = 5.5 Hz), 2.47 (3H, br s), 2.91 (1H, dt, J = 13.7, 6.8

Hz), 5.11 (2H, s), 6.00 (1H, d, J = 2.5 Hz), 6.12 (1H, dd, J = 7.5, 2.4 Hz),

7.05−7.16 (1H, m), 7.24−7.33 (2H, m), 7.35−7.40 (2H, m), 7.43 (1H,

N

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Journal of Medicinal Chemistry

Article

d, J = 9.5 Hz), 7.65 (1H, d, J = 7.7 Hz), 8.37 (1H, s). ¹³C NMR (101

MHz, DMSO-d₆) δ 7.8, 7.9, 8.0, 23.8, 33.1, 69.6, 97.6, 100.2, 114.8,

116.5, 121.9, 123.2, 126.4, 127.2, 128.0, 133.1, 139.6, 141.9, 145.0, 148.5,

162.8, 167.2. MS (ESI/APCI) m/z 414.4 [M + H]⁺. Anal. Calcd for

C₂₆H₂₇N₃O₂·0.1H₂O: C, 75.19; H, 6.60; N, 10.12. Found: C, 75.19; H,

6.54; N, 10.10.

7.51−7.54 (2H, m), 7.62 (1H, d, J = 9.6 Hz), 7.69 (1H, d, J = 7.6 Hz),

8.56 (1H, s). ¹³C NMR (101 MHz, DMSO-d₆) 8.8, 35.0, 38.3, 69.0, 97.6,

100.4, 115.3 (d, J = 21.2 Hz), 116.4, 123.0, 123.5, 125.5, 128.4, 130.2 (d,

J = 9.1 Hz), 132.0 (d, J = 4.0 Hz), 137.2, 139.5, 141.0, 161.9 (d, J = 245.4

Hz), 162.7, 164.7, 167.2. MS (ESI/APCI) m/z 421.4 [M + H]⁺. Anal.

Calcd for C₂₃H₂₁FN₄O₃·0.15H₂O: C, 65.29; H, 5.07; N, 13.24. Found:

C, 65.29; H, 5.02; N, 13.21.

6-{4-[(4-Fluorobenzyl)oxy]-2-oxopyridin-1(2H)-yl}-N-methoxy-

N,3-dimethylimidazo[1,2-a]pyridine-2-carboxamide (10l). To a

stirred mixture of compound 9b (500 mg, 2.28 mmol), 11f (945 mg,

2.73 mmol), K₂CO₃(945 mg, 6.84 mmol), and dioxane (20 mL) were

added CuI (174 mg, 0.91 mmol) and trans-N,N′-dimethyl-cyclohexane-

1,2-diamine (130 mg, 0.91 mmol). The reaction vessel was sealed and

heated at 110 °C for 16 h. The reaction mixture was then cooled to rt,

ﬁltered through Celite, and then concentrated. The residue was diluted

with EtOAc (150 mL), washed with brine (50 mL), dried over Na₂SO₄,

and concentrated. The crude product was puriﬁed by column

chromatography (silica gel, DCM/MeOH = 98/2) to give the title

compound (320 mg, 32%) as an oﬀ-white solid. ¹H NMR (400 MHz,

DMSO-d₆) δ 2.57 (3H, s), 3.41 (3H, s), 3.75 (3H, s), 5.15 (2H, s), 6.03

(1H, d, J = 2.6 Hz), 6.16 (1H, dd, J = 7.6, 2.6 Hz), 7.23−7.32 (3H, m),

7.51−7.54 (2H, m), 7.62 (1H, d, J = 9.6 Hz), 7.70 (1H, d, J = 7.5 Hz),

8.58 (1H, s). MS (ESI/APCI) m/z 437.4 [M + H]⁺.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-{[4-

(triﬂuoromethyl)benzy]oxy}pyridin-2(1H)-one (10g). To a stirred

mixture of 9c (110 mg, 0.409 mmol), 11a (121 mg, 0.409 mmol),

K₂CO₃(168 mg, 1.22 mmol), and dioxane (10 mL) was added CuI (15

mg, 0.079 mmol) and trans-N,N′-dimethylcyclohexane-1,2-diamine (11

mg, 0.079 mmol). The reaction vessel was sealed and heated at 110 °C

for 16 h. The mixture was ﬁltered through Celite, and the ﬁltrate was

concentrated in vacuo. The residue was diluted with water and extracted

with EtOAc. The organic layer was washed with brine, dried over

Na₂SO₄, and concentrated. The residue was puriﬁed by silica gel column

chromatography (silica gel, DCM/MeOH = 98/2) to give the title

1

compound (40 mg, 22%) as an oﬀ-white solid; mp 240−242 °C. H

NMR (400 MHz, CDOD₃) δ 0.95−1.01 (4H, m), 2.06 (1H, m), 2.51

(3H, s), 5.28 (2H, s), 6.10 (1H, d, J = 2.4 Hz), 6.33 (1H, dd, J = 7.6, 2.4

Hz), 7.21 (1H, dd, J = 9.2, 2.0 Hz), 7.47 (1H, d, J = 9.6 Hz), 7.62−7.67

(3H, m), 7.72 (2H, d, J = 8.4 Hz), 8.33 (1H, s). ¹³C NMR (75 MHz,

DMSO-d₆) δ 7.8, 8.0, 8.1, 68.7, 97.8, 100.1, 114.9, 116.6, 121.9, 123.2,

124.2 (q, J = 270.8 Hz), 125.4 (q, J = 3.8 Hz), 127.2, 128.2, 128.6 (q, J =

31.5 Hz), 139.8, 140.7, 141.9, 145.1, 162.8, 166.9. MS (ESI/APCI) m/z

440.2 [M + H]⁺. Anal. Calcd for C₂₄H₂₀F₃N₃O₂: C, 65.50; H, 4.59; N,

9.56. Found: C, 65.45; H, 4.65; N, 9.53.

1-(2-Acetyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-[(4-

ﬂuorobenzyl)oxy]pyridin-2(1H)-one (10m). To a stirred solution of 10l

(150 mg, 0.34 mmol) in THF (10 mL) was added a solution of MeMgBr

(3 M in ether, 344 μL, 1.03 mmol) at −78 °C, and the resulting mixture

was stirred at same temperature for 2 h. The reaction mixture was then

quenched with satd NH₄Cl solution (40 mL) and allowed to warm to rt.

The reaction mixture was concentrate and extracted with EtOAc (100

mL). The organic layer was washed with brine (40 mL), dried over

Na₂SO₄, and concentrated. The resulting residue was puriﬁed by

column chromatography (silica gel, DCM/MeOH = 97/3) to give the

4-(Benzyloxy)-1-(2,3-dimethylimidazo[1,2-a]pyridin-6-yl)pyridin-

2(1H)-one (10h). The title compound was prepared in 50% yield using

4-(benzyloxy)pyridine-2(1H)-one and 11b in an analogous manner to

10g. Oﬀ-white solid; mp 220−221 °C. ¹H NMR (400 MHz, DMSO-d₆)

δ 2.33 (3H, s), 2.39 (3H, s), 5.17 (2H, s), 5.99 (1H, d, J = 2.4 Hz), 6.14

(1H, dd, J = 7.6, 2.3 Hz), 7.12−7.14 (1H, m), 7.45−7.49 (5H, m), 7.67

(1H, d, J = 7.6 Hz), 8.38 (1H, s). ¹³C NMR (101 MHz, DMSO-d₆) δ 7.9,

13.1, 68.8, 97.7, 100.2, 114.9, 117.0, 122.2, 123.1, 127.2, 128.5, 129.6,

132.7, 134.8, 139.7, 140.0, 141.7, 162.7, 167.0. MS (ESI/APCI) m/z

379.8 [M + H]⁺. Anal. Calcd for C₂₁H₁₈ClN₃O₂·H₂O: C, 63.40; H, 5.07;

N, 10.56. Found: C, 63.36; H, 5.07; N, 10.50.

4-(Benzyloxy)-1-(2-ethyl-3-methylimidazo[1,2-a]pyridin-6-yl)-

pyridin-2(1H)-one (10i). The title compound was prepared in 25%

using 4-(benzyloxy)pyridine-2(1H)-one and 11c in an analogous

manner to 10g. Oﬀ-white solid; mp 218−220 °C. ¹H NMR (400

MHz, DMSO-d₆) δ 1.22 (3H, t, J = 7.5 Hz), 2.39 (3H, s), 2.69 (2H, dd, J

= 14.9, 7.3 Hz), 5.16 (2H, s), 5.99 (1H, d, J = 2.6 Hz), 6.14 (1H, dd, J =

7.6, 2.6 Hz), 7.13 (1H, dd, J = 9.4, 1.9 Hz), 7.48−7.49 (5H, m), 7.67

(1H, d, J = 7.6 Hz), 8.38 (1H, d, J = 1.3 Hz). ¹³C NMR (101 MHz,

DMSO-d₆) δ 7.8, 14.1, 20.3, 68.8, 97.7, 100.2, 115.1, 116.2, 122.3, 123.1,

127.2, 128.5, 129.6, 132.7, 134.8, 139.7, 141.8, 145.5, 162.7, 167.0. MS

(ESI/APCI) m/z 394.0 [M + H]⁺. Anal. Calcd for C₂₂H₂₀ClN₃O₂·

0.25H₂O: C, 66.33; H, 5.19; N, 10.55. Found: C, 66.22; H, 5.10; N,

10.54.

6-{4-[(4-Fluorobenzyl)oxy]-2-oxopyridin-1(2H)-yl}-3-

methylimidazo[1,2-a]pyridine-2-carbonitrile (10j). The title com-

pound was prepared in 34% yield using 9a and 11d in an analogous

manner to 10g. Oﬀ-white solid; mp 230−232 °C. ¹H NMR (400 MHz,

DMSO-d₆) δ 2.63 (3H, s), 5.15 (2H, s), 6.04 (1H, d, J = 2.4 Hz), 6.18

(1H, dd, J = 2.5, 1.6 Hz), 7.23−7.28 (2H, m), 7.47 (1H, dd, J = 9.7, 1.7

Hz), 7.51−7.54 (2H, m), 7.67−7.69 (2H, m), 8.69 (1H, s). ¹³C NMR

(101 MHz, DMSO-d₆) δ 8.7, 69.0, 97.6, 100.6, 115.1, 115.2, 115.3 (d, J =

21.2 Hz), 116.5, 123.7, 128.0, 129.6, 130.2 (d, J = 9.1 Hz), 131.2, 132.0

(d, J = 3.0 Hz), 139.3, 143.1, 161.9 (d, J = 245.4 Hz), 162.6, 167.3. MS

(ESI/APCI) m/z 375.0 [M + H]⁺. Anal. Calcd for C₂₁H₁₅FN₄O₂·

0.25H₂O: C, 66.57; H, 4.12; N, 14.79. Found: C, 66.74; H, 4.09; N,

14.78.

1

title compound (55 mg, 40%) as a white solid; mp 249−251 °C. H

NMR (400 MHz, DMSO-d₆) δ 2.61 (3H, s), 2.73 (3H, s), 5.15 (2H, s),

6.04 (1H, d, J = 2.2 Hz), 6.16 (1H, dd, J = 7.7, 2.6 Hz), 7.24−7.28 (2H,

m), 7.35 (1H, dd, J = 9.7, 1.7 Hz), 7.51−7.54 (2H, m), 7.68−7.71 (2H,

m), 8.63 (1H, s). ¹³C NMR (101 MHz, DMSO-d₆) δ 9.2, 27.7, 69.0,

97.6, 100.5, 115.3 (d, J = 21.2 Hz), 117.1, 123.3, 125.6, 126.7, 126.6,

129.2, 130.2 (d, J = 8.1 Hz), 132.0 (d, J = 3.0 Hz), 139.1, 139.4, 141.4,

161.9 (d, J = 245.4 Hz), 162.6, 167.2, 196.8. MS (ESI/APCI) m/z 392

[M + H]⁺. Purity 98.9% (HPLC).

1-[2-(Cyclopropylcarbonyl)-3-methylimidazo[1,2-a]pyridin-6-yl]-

4-[(4-ﬂuorobenzyl)oxy]pyridin-2(1H)-one (10n). The title compound

c

was prepared in 39% yield using PrMgBr in an analogous manner to

10m. White solid; mp 233−234 °C. ¹H NMR (400 MHz, DMSO-d₆) δ

1.04 (4H, m), 2.73 (3H, s), 3.34 (1H, m), 5.15 (2H, s), 6.04 (1H, d, J =

2.5 Hz), 6.16 (1H, dd, J = 7.5, 2.4 Hz), 7.24−7.28 (2H, m), 7.37 (1H, dd,

J = 9.5, 1.5 Hz), 7.51−7.54 (2H, m), 7.69 (1H, d, J = 3.9 Hz), 7.72 (1H,

d, J = 1.6 Hz), 8.64 (1H, s). ¹³C NMR (101 MHz, DMSO-d₆) δ 9.2, 11.0,

17.4, 69.0, 97.6, 100.5, 115.3 (d, J = 21.2 Hz), 117.2, 123.3, 125.4, 126.6,

129.2, 130.2 (d, J = 8.1 Hz), 132.0 (d, J = 3.0 Hz), 139.1, 139.4, 141.6,

161.9 (d, J = 245.4 Hz), 162.6, 167.2, 198.0. MS (ESI/APCI) m/z 418

[M + H]⁺. Purity 98.3% (HPLC).

4-[(4-Fluorobenzyl)oxy]-1-[2-(hydroxymethyl)-3-methylimidazo-

[1,2-a]pyridin-6-yl]pyridin-2(1H)-one (10o). The title compound was

prepared in 43% yield using 9a and 11g in an analogous manner to 10g.

White solid; mp 270−273 °C. ¹H NMR (400 MHz, CDCl₃) δ 2.46 (3H,

s), 4.58 (2H, d, J = 5.3 Hz), 5.04 (1H, m), 5.14 (2H, s), 6.02 (1H, d, J =

2.5 Hz), 6.13 (1H, dd, J = 7.6, 2.6 Hz), 7.18 (1H, dd, J = 9.5, 1.6 Hz),

7.24−7.28 (2H, m), 7.51−7.54 (3H, m), 7.68 (1H, d, J = 7.6 Hz), 8.44

(1H, s). ¹³C NMR (101 MHz, DMSO-d₆) δ 8.1, 57.0, 68.9, 97.6, 100.3,

115.3 (d, J = 22.2 Hz), 115.6, 122.6, 123.7, 127.6, 130.2 (d, J = 9.1 Hz),

132.0 (d, J = 3.0 Hz), 139.6, 141.6, 143.7, 161.9 (d, J = 245.4 Hz), 162.7,

167.1. MS (ESI/APCI) m/z 380.0 [M + H]⁺. Anal. Calcd for

C₂₁H₁₈FN₃O₃·0.5H₂O: C, 64.94; H, 4.93; N, 10.82. Found: C, 64.89;

H, 4.74; N, 10.87.

6-{4-[(4-Fluorobenzyl)oxy]-2-oxopyridin-1(2H)-yl}-N,N,3-

trimethylimidazo[1,2-a]pyridine-2-carboxamide (10k). The title

compound was prepared in 34% yield using 9a and 11e in an analogous

1

manner to 10g. White solid; mp 252−254 °C. H NMR (400 MHz,

4-[(4-Fluorobenzyl)oxy]-1-[2-(methoxymethyl)-3-methylimidazo-

[1,2-a]pyridin-6-yl]pyridin-2(1H)-one (10p). The title compound was

prepared in 26% yield using 9a and 11h in an analogous manner to 10g.

DMSO-d₆) δ 2.54 (3H, s), 3.01 (3H, s), 3.22 (3H, s), 5.15 (2H, s), 6.03

(1H, d, J = 2.4 Hz), 6.16 (1H, dd, J = 7.5, 2.4 Hz), 7.24−7.31 (3H, m),

O

DOI: 10.1021/acs.jmedchem.5b01704

J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

White solid. ¹H NMR (400 MHz, DMSO-d₆) δ 2.49 (3H, s), 3.42 (3H,

s), 4.65 (2H, s), 5.01 (2H, s), 6.04 (1H, d, J = 2.5 Hz), 6.08 (1H, dd, J =

7.5, 2.5 Hz), 7.08−7.14 (3H, m), 7.26 (1H, m), 7.39 (2H, dd, J = 8.4, 5.4

Hz), 7.63 (1H, d, J = 9.4 Hz), 7.98 (1H, s). MS (ESI/APCI) m/z 394.0

[M + H]⁺.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-(pyridin-

4-ylmethoxy)pyridin-2(1H)-one (10t). The title compound was

prepared in 28% yield using pyridin-4-yl-methanol in an analogous

manner to 10s. White solid; mp 251−253 °C. H NMR (400 MHz,

1

DMSO-d₆) δ 0.98−1.08 (4H, m), 1.98−2.02 (1H, m), 2.50 (3H, s), 5.11

(2H, s), 6.02 (1H, d, J = 2.6 Hz), 6.15 (1H, dd, J = 7.6, 2.7 Hz), 7.06 (1H,

dd, J = 9.4, 1.8 Hz), 7.31 (1H, d, J = 7.6 Hz), 7.36 (2H, d, J = 5.6 Hz),

7.57 (1H, d, J = 9.4 Hz), 7.94 (1H, s), 8.68 (2H, d, J = 5.8 Hz). ¹³C NMR

(75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 67.8, 97.8, 100.1, 114.9, 116.6,

121.8, 122.0, 123.2, 127.1, 139.9, 141.9, 144.9, 145.1, 149.8, 162.7, 166.8.

MS (ESI/APCI) m/z 373.3 [M + H]⁺. Anal. Calcd for C₂₂H₂₀N₄O₂·

0.5H₂O: C, 69.28; H, 5.55; N, 14.69. Found: C, 69.33; H, 5.50; N, 14.57.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-(pyrimi-

din-5-ylmethoxy)pyridin-2(1H)-one (10u). The title compound was

prepared in 33% yield using pyrimidin-5-yl-methanol in an analogous

manner to 10s. White solid; mp 236−238 °C.¹H NMR (400 MHz,

DMSO-d₆) δ 0.88−0.92 (4H, m), 2.05−2.09 (1H, m), 2.48 (3H, s), 5.26

(2H, s), 6.08 (1H, d, J = 2.5 Hz), 6.16 (1H, dd, J = 7.5, 2.6 Hz), 7.11 (1H,

dd, J = 9.5, 1.8 Hz), 7.43 (1H, d, J = 9.4 Hz), 7.68 (1H, d, J = 7.8 Hz),

8.38 (1H, s), 8.95 (2H, s), 9.22 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆)

δ 7.8, 8.0, 8.1, 65.1, 97.7, 100.1, 114.9, 116.6, 122.0, 123.2, 127.1, 129.7,

139.9, 141.9, 145.1, 156.5, 158.3, 162.8, 166.8. MS (ESI/APCI) m/z

374.2 [M + H]⁺. Anal. Calcd for C₂₁H₁₉N₅O_2·0.75H₂O: C, 65.19; H,

5.34; N, 18.10. Found: C, 65.32; H, 5.22; N, 18.11.

(6-{4-[(4-Fluorobenzyl)oxy]-2-oxopyridin-1(2H)-yl}-3-

methylimidazo[1,2-a]pyridin-2-yl)acetonitrile (10q). To a stirred

solution of compound 10o (120 mg, 0.31 mmol) in DCM (1 mL)

was added SOCl₂(1 mL), and the mixture was stirred at rt for 18 h. The

mixture was concentrated, diluted with DCM (100 mL), and quenched

with satd NaHCO₃solution (50 mL). The organic layer was separated,

washed successively with water (50 mL) and brine (50 mL), dried over

Na₂SO₄, and concentrated to give 1-[2-(chloromethyl)-3-

methylimidazo[1,2-a]pyridin-6-yl]-4-[(4-ﬂuorobenzyl)oxy]pyridin-

2(1H)-one (120 mg, 96%) as an oﬀ-white solid. ¹H NMR (400 MHz,

DMSO-d₆) δ 4.91 (2H, s), 5.14 (2H, s), 6.02 (1H, d, J = 2.4 Hz), 6.15

(1H, dd, J = 7.6, 2.5 Hz), 7.23−7.28 (3H, m), 7.51−7.57 (3H, m), 7.68

(1H, d, J = 7.6 Hz), 8.48 (1H, s). MS (ESI/APCI) m/z 398.2 [M + H]⁺.

To a stirred solution of 1-[2-(chloromethyl)-3-methylimidazo[1,2-

a]pyridin-6-yl]-4-[(4-ﬂuorobenzyl)oxy]pyridin-2(1H)-one (150 mg,

0.32 mmol) in THF (5 mL) were added TMSCN (115 μL, 1.13

mmol) and TBAF (1 M solution in THF, 1.13 mL, 1.13 mmol) at rt, and

the resulting mixture was stirred for 4 h. The reaction mixture was then

quenched with satd FeSO₄solution (10 mL), and extracted with DCM

(100 mL) twice. The combined organic layers were washed with brine

(30 mL), dried over Na₂SO₄, and concentrated. The crude material was

puriﬁed by column chromatography (silica gel, DCM/MeOH = 97/3)

to aﬀord the title compound (60 mg, 48%) as a white solid; mp 220−222

°C. ¹H NMR (400 MHz, DMSO-d₆) δ 2.46 (3H, s), 4.15 (2H, s), 5.14

(2H, s), 6.03 (1H, s), 6.15 (1H, d, J = 7.2 Hz), 7.24−7.28 (3H, m),

7.51−7.59 (3H, m), 7.67 (1H, d, J = 7.6 Hz), 8.48 (1H, s). ¹³C NMR (75

MHz, DMSO-d₆) δ 7.7, 16.4, 69.0, 97.6, 100.4, 115.4 (d, J = 21.0 Hz),

115.6, 118.3, 118.5, 122.9, 124.8, 128.0, 130.0 (d, J = 9.0 Hz), 132.0 (d, J

= 3.8 Hz), 133.6, 139.6, 142.2, 162.0 (d, J = 243.0 Hz), 162.7, 167.2. MS

(ESI/APCI) m/z 389.4 [M + H]⁺. Anal. Calcd for C₂₂H₁₇FN₄O₂·

0.25H₂O: C, 67.25; H, 4.49; N, 14.26. Found: C, 67.30; H, 4.46; N,

14.18.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-(thio-

phen-2-ylmethoxy)pyridin-2(1H)-one (10v). The title compound was

prepared in 22% yield using thiophen-2-yl-methanol in an analogous

manner to 10s. Oﬀ-white solid; mp 225 °C (decomposition). ¹H NMR

(400 MHz, DMSO-d₆) δ 0.88−0.92 (4H, m), 2.06−2.08 (1H, m), 5.35

(2H, s), 6.07−6.12 (2H, m), 7.07 (1H, dd, J = 4.8, 3.5 Hz), 7.09−7.13

(1H, m), 7.27 (1H, d, J = 3.1 Hz), 7.43 (1H, d, J = 9.4 Hz), 7.61 (1H, d, J

= 4.9 Hz), 7.64 (1H, d, J = 7.5 Hz), 8.37 (1H, s). ¹³C NMR (101 MHz,

DMSO-d₆) δ 7.8, 7.9, 8.0, 64.5, 97.7, 100.1, 114.8, 116.5, 121.9, 123.2,

126.9, 127.1, 127.3, 128.3, 137.7, 139.7, 141.9, 145.0, 162.7, 166.7. MS

(ESI/APCI) m/z 378.3 [M + H]⁺. Anal. Calcd for C₂₁H₁₉N₃O₂S·

0.4H₂O: C, 65.57; H, 5.19; N, 10.92. Found: C, 65.57; H, 5.01; N, 10.86.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-(thio-

phen-3-ylmethoxy)pyridin-2(1H)-one (10w). The title compound was

prepared in 40% yield using thiophen-3-yl-methanol in an analogous

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-(pyridin-

2-ylmethoxy)pyridin-2(1H)-one (10r). The title compound was

prepared in 32% yield using 2-(bromomethyl)pyridine hydrobromide

1

manner to 10s. White solid; mp 234 °C (decomposition). H NMR

(400 MHz, DMSO-d₆) δ 0.88−0.94 (4H, m), 2.04−2.10 (1H, m), 5.15

(2H, s), 6.02 (1H, d, J = 2.6 Hz), 6.10 (1H, dd, J = 7.6, 2.6 Hz), 7.07 (1H,

dd, J = 9.5, 1.8 Hz), 7.19 (1H, d, J = 4.9 Hz), 7.43 (1H, d, J = 9.4 Hz),

7.59 (1H, dd, J = 4.8, 2.9 Hz), 7.64−7.66 (2H, m), 8.37 (1H, s). ¹³C

NMR (101 MHz, DMSO-d₆) δ 7.8, 7.9, 8.0, 65.2, 97.5, 100.2, 114.8,

116.5, 121.9, 123.2, 124.7, 126.8, 127.2, 127.5, 136.6, 139.6, 141.9, 145.0,

162.8, 167.1. MS (ESI/APCI) m/z 378.3 [M + H]⁺. Anal. Calcd for

C₂₁H₁₉N₃O₂S·0.6H₂O: C, 64.96; H, 5.24; N, 10.82. Found: C, 64.84; H,

5.01; N, 10.71.

4-[(5-Chloropyridin-2-yl)methoxy]-1-(2-cyclopropyl-3-

methylimidazo[1,2-a]pyridin-6-yl)pyridin-2(1H)-one (10x). The title

compound was prepared in 53% yield using (5-chloropyridin-2-

yl)methanol in an analogous manner to 10s. White solid; mp 200−

201 °C. ¹H NMR (400 MHz, DMSO-d₆) δ 0.84−0.96 (4H, m), 2.01−

2.12 (1H, m), 2.47 (3H, s), 5.24 (2H, s), 5.99 (1H, d, J = 2.6 Hz), 6.18

(1H, dd, J = 7.6, 2.7 Hz), 7.11 (1H, dd, J = 9.5, 1.8 Hz), 7.43 (1H, d, J =

9.5 Hz), 7.61 (1H, d, J = 8.3 Hz), 7.68 (1H, d, J = 7.7 Hz), 8.03 (1H, dd, J

= 8.3, 2.4 Hz), 8.37 (1H, s), 8.67 (1H, d, J = 2.4 Hz). ¹³C NMR (75

MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 69.8, 97.8, 100.1, 114.9, 116.6, 121.9,

123.2, 123.5, 127.1, 130.5, 136.9, 139.9, 141.9, 145.1, 147.8, 154.0, 162.7,

166.9. MS (ESI/APCI) m/z 407.1 [M + H]⁺. Anal. Calcd for

C₂₂H₁₉ClN₄O₂: C, 64.94; H, 4.71; N, 13.77. Found: C, 64.80; H,

4.72; N, 13.69.

1

in an analogous manner to 10f. White crystals; mp 217−220 °C. H

NMR (400 MHz, DMSO-d₆) δ 1.05 (2H, d, J = 3.8 Hz), 1.16 (2H, d, J =

8.3 Hz), 2.23−2.32 (1H, m), 2.58 (3H, s), 5.29 (2H, s), 6.07 (1H, d, J =

2.3 Hz), 6.30 (1H, dd, J = 7.7, 2.4 Hz), 7.41−7.53 (1H, m), 7.62 (1H, d, J

= 7.9 Hz), 7.72 (1H, d, J = 7.8 Hz), 7.85−7.92 (1H, m), 7.93−8.01 (2H,

m), 8.65 (1H, d, J = 4.5 Hz), 9.00 (1H, s). ¹³C NMR (75 MHz, DMSO-

d₆) δ 5.9, 7.7, 8.1, 68.6, 97.8, 100.9, 110.5, 120.1, 123.8, 124.5, 124.8,

131.2, 132.1, 135.5, 137.0, 139.3, 141.0, 146.4, 152.9, 162.4, 167.2. MS

(ESI/APCI) m/z 373.2 [M + H]⁺. Anal. Calcd for C₂₂H₂₀N₄O₂·2HCl·

H₂O: C, 57.03; H, 5.22; N, 12.09. Found: C, 57.13; H, 5.19; N, 12.09.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-(pyridin-

3-ylmethoxy)pyridin-2(1H)-one (10s). To a solution of 17 (150 mg,

0.53 mmol), pyridin-3-yl-methanol (116 mg, 1.06 mmol), and tributyl

phosphine (322 mg, 1.59 mmol) in THF (15 mL) was added ADDP

(401 mg, 1.59 mmol). The mixture was stirred under sonication at 60 °C

for 4 h. The reaction mixture was then cooled to rt and concentrated.

The residue was diluted with DCM (60 mL), washed with water (30

mL) twice and brine (30 mL), dried over Na₂SO₄, and concentrated.

The resulting residue was puriﬁed by column chromatography (silica

gel, DCM/MeOH = 96/4) to give the title compound (61 mg, 31%) as

an oﬀ-white solid; mp 210−212 °C. ¹H NMR (400 MHz, DMSO-d₆) δ

0.88−0.92 (4H, m), 2.07 (1H, m), 5.21 (2H, s), 6.05 (1H, d, J = 2.5 Hz),

6.14 (1H, dd, J = 7.5, 2.6 Hz), 7.11 (1H, dd, J = 9.4, 1.7 Hz), 7.42−7.48

(2H, m), 7.67 (1H, d, J = 7.6 Hz), 7.89−7.91 (1H, m), 8.37 (1H, s), 8.61

(1H, br s), 8.70 (1H, br s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0,

8.1, 67.4, 97.7, 100.2, 114.9, 116.6, 122.0, 123.2, 123.6, 127.2, 131.5,

135.9, 139.8, 141.9, 145.1, 149.2, 149.5, 162.8, 167.0. MS (ESI/APCI)

m/z 373.4 [M + H]⁺. Anal. Calcd for C₂₂H₂₀N₄O₂·0.1H₂O: C, 70.61; H,

5.44; N, 14.97. Found: C, 70.75; H, 5.38; N, 14.89.

4-[(5-Chlorothiophen-3-yl)methoxy]-1-(2-cyclopropyl-3-

methylimidazo[1,2-a]pyridin-6-yl)pyridin-2(1H)-one (10y). The title

compound was prepared in 41% yield using (5-chloro-thiophen-3-yl)-

methanol in an analogous manner to 10s. White solid; mp 233−235 °C.

¹H NMR (400 MHz, DMSO-d₆) δ 0.88−0.92 (4H, m), 2.05−2.09 (1H,

m), 2.47 (3H, s), 5.06 (2H, s), 6.01 (1H, d, J = 2.6 Hz), 6.10 (1H, dd, J =

7.6, 2.7 Hz), 7.11 (1H, dd, J = 9.5, 1.9 Hz), 7.20 (1H, d, J = 1.7 Hz), 7.43

P

DOI: 10.1021/acs.jmedchem.5b01704

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Journal of Medicinal Chemistry

Article

(1H, d, J = 9.4 Hz), 7.56 (1H, d, J = 1.2 Hz), 7.65 (1H, d, J = 7.7 Hz),

8.36 (1H, d, J = 1.3 Hz). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1

65.1, 97.5, 100.2, 114.9, 116.6, 121.9, 123.2, 124.3, 127.0, 127.2, 128.9,

136.4, 139.7, 141.9, 145.1, 162.8, 166.9. MS (ESI/APCI) m/z 412.0 [M

+ H]⁺. Anal. Calcd for C₂₁H₁₈ClN₃O₂S·0.4H₂O: C, 60.18; H, 4.52; N,

10.03. Found: C, 60.01; H, 4.47; N, 10.08.

and concentrated. The residue was poured into ice-cold satd NaHCO₃

solution (100 mL). The mixture was extracted with EtOAc (150 mL)

twice, and the combined EtOAc layers were washed with water (100

mL) and brine (100 mL), dried over Na₂SO₄, and concentrated to give

1

the title compound (500 mg, 88%) as a yellow solid. H NMR (400

MHz, DMSO-d₆) δ 2.60 (3H, s), 7.46 (1H, d, J = 9.5 Hz), 7.60 (1H, d, J

= 9.3 Hz), 8.74 (1H, s). MS (ESI/APCI) m/z 283.9 [M + H]⁺.

6-Iodo-N,N,3-trimethylimidazo[1,2-a]pyridine-2-carboxamide

(11e). To a stirred solution of 6-iodo-3-methylimidazo[1,2-a]pyridine-

2-carboxylic acid (150 mg, 0.49 mmol) in DMF (5 mL) were added

HATU (283 mg, 0.74 mmol) and DIPEA (0.17 mL, 0.99 mmol) at 0 °C.

The mixture was allowed to warm to rt for 30 min, and dimethylamine

(2 N THF solution, 0.25 mL, 0.49 mmol) was added. The resultant

mixture was stirred at rt for 18 h. The mixture was concentrated, and the

residue was diluted with DCM (100 mL), washed with satd NH₄Cl

solution (40 mL), satd NaHCO₃solution (20 mL), water (30 mL), and

brine (50 mL). DCM layer was then dried over Na₂SO₄and

concentrated to give the title compound (150 mg, 93%) as brown oil.

¹H NMR (400 MHz, DMSO-d₆) δ 2.52 (3H, s), 2.99 (3H, s), 3.18 (3H,

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-{[5-

(triﬂuoromethyl)thiophen-3-yl]methoxy}pyridin-2(1H)-one (10z). A

mixture of 19 (111 mg, 0.32 mmol), [5-(triﬂuoromethyl)thiophen-3-

yl]methanol (88 mg, 0.48 mmol), and KO^tBu (109 mg, 0.97 mmol) in

toluene (3 mL) was heated at 100 °C for 1 h. The mixture was poured

into water and extracted with EtOAc/THF (1:1). The extract was

washed with brine, dried over MgSO₄, and concentrated in vacuo. The

residue was puriﬁed by column chromatography (NH silica gel, hexane/

EtOAc = 97/3 to 0/100). The solid was recrystallized from IPA−IPE to

give the title compound (47.5 mg, 33%) as a white solid; mp 209−210

°C. ¹H NMR (400 MHz, DMSO-d₆) δ 0.83−0.99 (4H, m), 2.07 (1H, br

s), 5.17 (2H, s), 6.05 (1H, s), 6.13 (1H, d, J = 5.0 Hz), 7.12 (1H, d, J = 9.9

Hz), 7.44 (1H, d, J = 9.3 Hz), 7.67 (1H, d, J = 7.5 Hz), 7.81 (1H, s), 8.07

(1H, s), 8.37 (1H, s). ¹³C NMR (101 MHz, DMSO-d₆) δ 7.8, 7.9, 8.0,

64.6, 97.5, 100.1, 114.8, 116.5, 121.9, 122.4 (q, J = 269.7 Hz), 123.1,

127.1, 129.6, 129.8 (q, J = 38.4 Hz), 130.3 (q, J = 4.0 Hz), 137.1, 139.7,

141.9, 145.1, 162.7, 166.9. MS (ESI/APCI) m/z 446.4 [M + H]⁺. Anal.

Calcd for C₂₂H₁₈F₃N₃O₂S: C, 59.32; H, 4.07; N, 9.43. Found: C, 59.46;

H, 4.20; N, 9.35.

2-Cyclopropyl-6-iodo-3-methylimidazo[1,2-a]pyridine (11a). A

mixture of 2-bromo-1-cyclopropylpropan-1-one (1.00 g, 5.65 mmol),

5-iodopyridin-2-amine (1.24 g, 5.65 mmol), and EtOH (10 mL) was

heated at 70 °C for 48 h. The mixture was poured into satd NaHCO₃

solution and extracted with EtOAc. The extract was washed with brine,

dried over MgSO₄, concentrated, and puriﬁed by column chromatog-

raphy (silica gel, hexane/EtOAc = 95/5 to 65/45) to give the title

compound (520 mg, 31%) as a pale-orange solid. ¹H NMR (300 MHz,

DMSO-d₆) δ 0.81−0.94 (4H, m), 1.96−2.11 (1H, m), 2.46 (3H, s),

7.21−7.32 (2H, m), 8.44 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 8.3,

8.4, 8.5, 75.3, 116.3, 117.5, 128.6, 130.6, 142.1, 144.9. MS (ESI/APCI)

m/z 299.0 [M + H]⁺.

s), 7.41 (1H, d, J = 9.3 Hz), 7.46 (1H, dd, J = 9.3, 1.2 Hz), 8.62 (1H, s).

MS (ESI/APCI) m/z 329.8 [M + H]⁺.

6-Iodo-N-methoxy-N,3-dimethylimidazo[1,2-a]pyridine-2-car-

boxamide (11f). To a stirred solution of 6-iodo-3-methylimidazo[1,2-

a]pyridine-2-carboxylic acid (600 mg, 1.98 mmol) in DMF (10 mL)

were added HATU (1.50 g, 3.97 mmol), DIEPA (1.30 mL, 7.94 mmol),

and N,O-dimethylhydroxylamine hydrochloride (385 mg, 3.97 mmol)

at 0 °C. The resultant mixture was stirred at rt for 16 h. The mixture was

then diluted with water (60 mL) and extracted with EtOAc (100 mL)

twice. The organic layers were successively washed with satd NaHCO₃

solution (50 mL), water (50 mL), and brine (50 mL). The EtOAc layer

was then dried over Na₂SO₄and concentrated. The crude product was

puriﬁed by column chromatography (silica gel, hexane/EtOAc = 80/20)

to give the title compound (600 mg, 88%) as an oﬀ-white solid. ¹H NMR

(400 MHz, DMSO-d₆) δ 2.56 (3H, s), 3.39 (3H, s), 3.73 (3H, s), 7.42

(1H, d, J = 9.4 Hz), 7.48 (1H, d, J = 9.4 Hz), 8.64 (1H, s). MS (ESI/

APCI) m/z 345.6 [M + H]⁺.

(6-Iodo-3-methylimidazo[1,2-a]pyridin-2-yl)methanol (11g). To a

stirred solution of ethyl 6-iodo-3-methylimidazo[1,2-a]pyridine-2-

carboxylate (2.0 g, 6.06 mmol) in DCM (20 mL) was added DIBAL-

H (1.76 M toluene solution, 7.72 mL, 13.6 mmol) at −19 °C. The

resultant mixture was stirred at the same temperature for 3 h and then at

rt for 4 h. The reaction mixture was quenched with MeOH and water (2

mL) at −40 °C. The mixture was acidiﬁed with a few drops of 5 N HCl

solution and poured into satd NaHCO₃(20 mL) solution. The mixture

was extracted with EtOAc (150 mL) three times, and the combined

EtOAc layers were washed with brine (50 mL), dried over Na₂SO₄, and

concentrated. The crude residue was puriﬁed by column chromatog-

raphy (silica gel, DCM/MeOH = 98/2) to give the title compound (1.1

g, 62%) as a yellow solid. ¹H NMR (400 MHz, DMSO-d₆) δ 2.45 (3H,

s), 4.55 (2H, s), 5.03 (1H, br s), 7.31−7.39 (2H, m), 8.53 (1H, s). MS

(ESI/APCI) m/z 288.9 [M + H]⁺.

2-Methyl-6-iodo-3-methylimidazo[1,2-a]pyridine (11b). To a

solution of 5-iodopyridin-2-amine (2.50 g, 11.4 mmol) in DMF (20

mL) was added 3-bromo-2-butanone (1.19 mL, 11.4 mmol), and the

mixture was stirred at 90 °C overnight. The mixture was neutralized with

satd NaHCO₃solution and extracted with EtOAc/THF (1:1). The

organic layer was separated, washed with water and brine, dried over

MgSO₄, and concentrated in vacuo. The residue was puriﬁed by column

chromatography (silica gel, hexane/EtOAc = 75/25 to 0/100) to give

the title compound (980 mg, 32%) as a dark-yellow solid. ¹H NMR (300

MHz, DMSO-d₆) δ 2.29 (3H, s), 2.38 (3H, s), 7.20−7.39 (2H, m), 8.46

(1H, s). MS (ESI/APCI) m/z 272.9 [M + H]⁺.

2-Ethyl-6-iodo-3-methylimidazo[1,2-a]pyridine (11c). The title

compound was prepared in 31% yield using 2-bromopentan-3-one in

an analogous manner to 11a. Yellow solid. ¹H NMR (400 MHz, DMSO-

d₆) δ 1.19 (3H, t, J = 7.2 Hz), 2.38 (3H, s), 2.65 (2H, q, J = 7.2 Hz),

7.27−7.33 (2H, m), 8.47 (1H, s). MS (ESI/APCI) m/z 286.8 [M + H]⁺.

6-Iodo-3-methylimidazo[1,2-a]pyridine-2-carbonitrile (11d). To a

stirred solution of 6-iodo-3-methylimidazo[1,2-a]pyridine-2-carboxylic

acid (1.0 g, 3.3 mmol) in DMF (15 mL) were added HATU (1.88 g, 4.92

mmol) and Et₃N (1.38 mL, 9.9 mmol) at 0 °C. The mixture was allowed

to warm to rt for 30 min, and then NH₄Cl (725 mg, 13.5 mmol) was

added. The resultant mixture was stirred at the same temperature for 18

h. The mixture was then concentrated, and the residue was diluted with

DCM (100 mL) and washed with satd NH₄Cl solution (40 mL), satd

NaHCO₃solution (20 mL), water (30 mL), and brine (50 mL)

successively. DCM layer was then dried over Na₂SO₄and concentrated

to give 6-iodo-3-methylimidazo[1,2-a]pyridine-2-carboxamide (600

6-Iodo-2-(methoxymethyl)-3-methylimidazo[1,2-a]pyridine

(11h). To a stirred solution of 11g (200 mg, 0.69 mmol) in DCM (2

mL) was added SOCl₂(56 μL, 0.78 mmol) at rt, and the mixture was

stirred at the same temperature for 4 h. SOCl₂(125 μL, 0.78 mmol) was

added, and stirred at rt for further 3 h. The reaction mixture was diluted

with DCM (100 mL), washed with satd NaHCO₃(20 mL), dried over

Na₂SO₄, and concentrated to give 2-(chloromethyl)-6-iodo-3-

1

methylimidazo[1,2-a]pyridine (150 mg, 24%) as a yellow solid. H

NMR (400 MHz, DMSO-d₆) δ 2.48 (3H, s), 4.88 (2H, s), 7.36 (1H, d, J

= 9.3 Hz), 7.42 (1H, dd, J = 9.3, 1.3 Hz), 8.56 (1H, s). MS (ESI/APCI)

m/z 306.6 [M + H]⁺.

To a stirred solution of 2-(chloromethyl)-6-iodo-3-methylimidazo-

[1,2-a]pyridine (150 mg, 0.49 mmol) and MeOH (2.5 mL) was added

NaOMe (65 mg, 1.06 mmol), and the resulting mixture was heated at

reﬂux for 3 h. The reaction mixture was then cooled to rt and

concentrated. The mixture was poured into water (50 mL) and extracted

with EtOAc (50 mL) twice. Combined EtOAc layers were washed with

brine (30 mL), dried over Na₂SO₄, and concentrated. The residue was

puriﬁed by column chromatography (silica gel, DCM/MeOH = 97/3)

1

mg, 60%) as a yellow solid. H NMR (400 MHz, DMSO-d₆) δ 2.73

(3H, s), 7.33 (1H, br s), 7.39 (1H, d, J = 9.3 Hz), 7.49 (1H, d, J = 9.4 Hz),

7.63 (1H, br s), 8.63 (1H, s). MS (ESI/APCI) m/z 301.9 [M + H]⁺.

To a stirred solution of 6-iodo-3-methylimidazo[1,2-a]pyridine-2-

carboxamide (600 mg, 1.99 mmol) was added POCl₃(40 mL), and the

mixture was heated at reﬂux for 3 h. The mixture was then cooled to rt

Q

DOI: 10.1021/acs.jmedchem.5b01704

J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

to give the title compound (100 mg, 67%) as an oﬀ-white solid. ¹H NMR

(400 MHz, DMSO-d₆) δ 2.50 (3H, s), 3.30 (3H, s), 4.49 (2H, s), 7.33−

7.38 (2H, m), 8.55 (1H, s). MS (ESI/APCI) m/z 303.0 [M + H]⁺.

4-Bromo-3-methyl-pyridine 1-Oxide (13). To a stirred solution of

12 (3.00 g, 19.4 mmol) in acetic acid (36 mL) was added dropwise acetyl

bromide (23 mL) at 0 °C. After complete addition, the reaction mixture

was heated at 80 °C for 3 h. The reaction mixture was cooled to rt, and

poured over crushed ice (200 g). The resulting mixture was neutralized

with 1 N NaOH solution and extracted with DCM (100 mL) twice. The

organic layer was washed with water (100 mL) and brine (100 mL),

dried over Na₂SO₄, and concentrated to give the title compound (2.2 g,

3-[(4-Chlorobenzyl)oxy]pyridin-2-ol (20). To a stirred solution of

KOH (0.360 g, 9.00 mmol) in MeOH (10 mL) was added pyridine-2,3-

diol (1.00 g, 9.00 mmol) portionwise. To the resulting red solution was

added 4-chlorobenzyl bromide (1.85 g, 9.00 mmol), and the reaction

mixture was stirred at rt for 30 min, then at 40 °C for 1.5 h. The mixture

was concentrated and partitioned between EtOAc, THF, and water. The

organic layer was washed with brine, dried over MgSO₄, and

concentrated. The precipitate was collected by ﬁltration and washed

with IPE to give the title compound (1.69 g, 80%) as an oﬀ-white solid.

¹H NMR (400 MHz, CDCl₃) δ 5.12 (2H, s), 6.14 (1H, d, J = 7.0 Hz),

6.74 (1H, d, J = 7.3 Hz), 7.02 (1H, d, J = 6.3 Hz), 7.31−7.40 (4H, m),

12.36−12.56 (1H, m). MS (ESI/APCI) m/z 236.0 [M + H]⁺.

2-Chloro-3-[4-(triﬂuoromethoxy)phenoxy]pyridine (22). To a

solution of 21 (2.00 g, 8.35 mmol) and 4-(triﬂuoromethoxy)phenol

(1.19 mL, 9.19 mmol) in CH₃CN (50 mL) was added Cu(OAc)₂(3.03

g, 16.7 mmol), MS3A (2.00 g), and pyridine (3.38 mL, 41.8 mmol) at rt,

and the mixture was vigorously stirred at rt for 10 days. The insoluble

material was removed by ﬁltration, and the ﬁltrate was concentrate in

vacuo. The residue was puriﬁed by column chromatography (silica gel,

hexane/EtOAc = 100/0 to 0/100) to give the title compound (1.15 g,

47%) as an orange oil. ¹H NMR (400 MHz, DMSO-d₆) δ 6.82 (1H, d, J

= 8.9 Hz), 7.16 (3H, d, J = 9.0 Hz), 7.42 (2H, d, J = 8.8 Hz), 7.46−7.54

(1H, m), 7.65 (1H, dd, J = 8.0, 1.3 Hz), 8.29 (1H, dd, J = 4.5, 1.3 Hz).

MS (ESI/APCI) m/z 289.9 [M + H]⁺.

1

60%) as a light-yellow solid. H NMR (400 MHz, DMSO-d₆) δ 2.24

(3H, s), 7.62 (1H, d, J = 6.8 Hz), 7.97 (1H, dd, J = 6.8, 1.7 Hz), 8.28 (1H,

s). MS (ESI/APCI) m/z 188.1 [M + H]⁺.

4-[(4-Chlorobenzyl)oxy]-3-methylpyridine 1-Oxide (14). The title

compound was prepared in 28% yield using 13 and (4-chlorophenyl)-

methanol in an analogous manner to 8b. Oﬀ-white solid. ¹H NMR (400

MHz, DMSO-d₆) δ 2.10 (3H, s), 5.19 (2H, s), 7.04 (1H, d, J = 7.1 Hz),

7.47 (4H, s), 8.01 (1H, d, J = 7.0 Hz), 8.07 (1H, s). MS (ESI/APCI) m/z

250.2 [M + H]⁺.

4-[(4-Chlorobenzyl)oxy]-5-methylpyridin-2(1H)-one (15). To 14

(500 mg, 1.9 mmol) was added acetic anhydride (5 mL), and the

solution was heated at reﬂux for 4 h. The reaction mixture was then

cooled to rt and concentrated. The residue was diluted with a mixture of

MeOH (20 mL) and 1 N NaOH solution (10 mL), and the resulting

solution was heated at reﬂux for 1 h. The mixture was cooled to rt and

concentrated. The residue was diluted with water (30 mL) and extracted

with DCM (75 mL) three times. The combined DCM layers were

concentrated and puriﬁed by preparative HPLC to aﬀord the title

compound (20 mg, 4%) as a white solid. ¹H NMR (400 MHz, DMSO-

d₆) δ 1.89 (3H, s), 5.08 (2H, s), 5.76 (1H, s), 7.09 (1H, s), 7.47 (4H, s),

10.95 (1H, s). MS (ESI/APCI) m/z 250.2 [M + H]⁺.

4-[(4-Chlorobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)-5-methylpyridin-2(1H)-one (16). The title compound

was prepared in 42% yield using 15 and 11a in an analogous manner to

10g. Oﬀ-white solid; mp 238−239 °C. ¹H NMR (400 MHz, DMSO-d₆)

δ 0.88−0.92 (4H, m), 1.97 (3H, s), 2.06 (1H, m), 2.47 (3H, s), 5.19 (2H,

s), 5.98 (1H, s), 7.10 (1H, dd, J = 9.4, 1.9 Hz), 7.42 (1H, d, J = 9.4 Hz),

7.50 (4H, s), 7.54 (1H, s), 8.35 (1H, d, J = 1.3 Hz). ¹³C NMR (75 MHz,

DMSO-d₆) δ 7.8, 8.0, 8.1, 112.2, 68.7, 97.2, 108.3, 114.8, 116.5, 121.9,

123.3, 127.3, 128.6, 129.3, 132.6, 135.1, 136.9, 141.9, 145.0, 162.5, 166.1.

MS (ESI/APCI) m/z 420.2 [M + H]⁺. Purity 99.8% (HPLC).

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-4-hydroxy-

pyridin-2(1H)-one (17). A mixture of 10c (2.2 g, 5.92 mmol), 10% Pd-C

(0.22 g, 2.07 mmol), and MeOH (40 mL) was stirred under H₂

atmosphere at ambient temperature for 1 h. The catalyst was removed

by ﬁltration, and the ﬁltrate was concentrated in vacuo to give the title

compound (1.65 g, 99%) as oﬀ-white amorphous solid. ¹H NMR (400

MHz, DMSO-d₆) δ 0.82−0.98 (4H, m), 2.02−2.12 (1H, m), 2.47 (3H,

s), 5.64 (1H, s), 5.97 (1H, d, J = 5.6 Hz), 7.10 (1H, d, J = 9.3 Hz), 7.41

(1H, d, J = 9.4 Hz), 7.57 (1H, d, J = 7.5 Hz), 8.33 (1H, s), 11.07 (1H, br

s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 98.1, 101.2, 114.7,

116.5, 121.8, 123.5, 127.6, 139.6, 141.9, 144.9, 163.1, 168.7. MS (ESI/

APCI) m/z 345.0 [M + H]⁺.

3-[4[(Triﬂuoromethoxy)phenoxy]pyridin-2(1H)-one (23). A mix-

ture of 22 (1.02 g, 3.52 mmol), KO^tBu (1.19 g, 10.1 mmol), water (0.19

t

mL, 10.1 mmol), and BuOH (15 mL) was heated at 150 °C for 1 h

under microwave irradiation. The mixture was poured into water and

extracted with EtOAc. The organic layer was separated, washed with

water and brine successively, dried over MgSO₄, and concentrated in

vacuo. The residue was puriﬁed by column chromatography (silica gel,

hexane/EtOAc = 100/0 to 75/25) to give the title compound (0.702 g,

73%) as white crystals. ¹H NMR (400 MHz, DMSO-d₆) δ 6.21 (1H, d, J

= 6.8 Hz), 6.97 (2H, d, J = 9.0 Hz), 7.26−7.36 (4H, m), 12.02 (1H, br s).

MS (ESI/APCI) m/z 272.9 [M + H]⁺.

3-[(4-Chlorobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)pyridin-2(1H)-one (24a). The title compound was

prepared in 15% yield using 11a and 20 in an analogous manner to

10a. White solid. 180 °C (decomposition). ¹H NMR (400 MHz,

CDCl₃) δ 0.94−1.11 (4H, m), 1.94−2.04 (1H, m), 2.48 (3H, s), 5.13

(2H, s), 6.15 (1H, t, J = 7.2 Hz), 6.73 (1H, d, J = 7.3 Hz), 7.03 (1H, d, J =

6.8 Hz), 7.08 (1H, d, J = 9.4 Hz), 7.31−7.44 (4H, m), 7.56 (1H, d, J = 9.4

Hz), 7.99 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 69.0,

104.2, 115.1, 116.0, 116.7, 122.0, 122.9, 127.4, 128.4, 129.6, 131.0, 132.5,

135.5, 141.9, 145.2, 148.1, 157.3. MS (ESI/APCI) m/z 406.1 [M + H]⁺.

Anal. Calcd for C₂₃H₂₀ClN₃O₂·0.5H₂O: C, 66.58; H, 5.10; N, 10.13.

Found: C, 66.30; H, 5.22; N, 10.21.

1-(2-Cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-3-[4-

(triﬂuoromethoxy)phenoxy]pyridin-2(1H)-one (24b). The title com-

pound was prepared in 13% yield using 11a and 23 in an analogous

1

manner to 10a. Pale-yellow crystals; mp 188−190 °C. H NMR (400

MHz, DMSO-d₆) δ 0.83−0.96 (4H, m), 2.03−2.14 (1H, m), 6.39 (1H, t,

J = 7.1 Hz), 7.09 (2H, d, J = 9.0 Hz), 7.16−7.24 (1H, m), 7.34 (2H, d, J =

8.7 Hz), 7.46 (2H, t, J = 9.5 Hz), 7.70 (1H, d, J = 5.5 Hz), 8.49 (1H, s).

¹³C NMR (101 MHz, DMSO-d₆) δ 7.8, 7.9, 8.0, 104.2, 115.0, 116.7,

4-Bromo-1-(2-cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-

pyridin-2(1H)-one (18). To a solution of 17 (1.95 g, 6.93 mmol) in

DMF (20 mL) was added phosphorus(V) tribromide oxide (1.77 mL,

17.4 mmol) at rt, and the mixture was heated at 110 °C for 1 h. The

mixture was poured into satd NaHCO₃solution and extracted with

EtOAc. The organic layer was separated, washed with water and brine,

dried over MgSO₄, and concentrated in vacuo. The residue was puriﬁed

by column chromatography (NH silica gel, hexane/EtOAc = 90/10 to

0/100) to give the title compound (1.70 g, 71%) as an oﬀ-white solid;

117.7, 120.1 (q, J = 256.5 Hz), 122.0, 122.5, 122.7, 127.0, 128.2, 136.2,

141.9, 143.1 (q, J = 2.0 Hz), 144.1, 145.3, 155.6, 157.3. MS (ESI/APCI)

m/z 442.3 [M + H]⁺. Anal. Calcd for C₂₃H₁₈F₃N₃O₃: C, 62.58; H, 4.11;

N, 9.52. Found: C, 62.43; H, 4.30; N, 9.35.

5-(4-Chlorobenzyloxy)-2-(tetrahydro-pyran-2-yl)-2H-pyridazin-3-

one (26). To a stirred solution of 25 (200 mg, 1.02 mmol) and 1-

bromomethyl-4-chlorobenzene (209 mg, 1.02 mmol) in MeCN (8 mL)

and DMF (1 mL) was added K₂CO₃(282 mg, 2.04 mmol), and the

resulting mixture was stirred at rt for 16 h. The reaction mixture was then

concentrated, diluted with water (30 mL), and extracted with EtOAc

(75 mL) three times. The organic layers were washed with water (50

mL) and brine (50 mL) successively, dried over Na₂SO₄, and

concentrated to give the title compound (210 mg, 64%) as a white

solid. ¹H NMR (400 MHz, DMSO-d₆) δ 1.44−1.49 (2H, m), 1.56−1.59

(1H, m), 1.62−1.66 (1H, m), 1.90−1.93 (1H, m), 2.00−2.10 (1H, m),

1

mp 198−200 °C. H NMR (400 MHz, DMSO-d₆) δ 0.83−0.98 (4H,

m), 2.04−2.12 (1H, m), 2.47 (3H, s), 6.60 (1H, d, J = 7.3 Hz), 6.88 (1H,

s), 7.16 (1H, d, J = 9.5 Hz), 7.46 (1H, d, J = 9.3 Hz), 7.74 (1H, d, J = 7.3

Hz), 8.47 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 7.99, 8.04,

109.3, 115.0, 116.7, 121.9, 122.1, 122.7, 126.7, 136.0, 140.3, 142.0, 145.3,

160.4. MS (ESI/APCI) m/z 345.0 [M + H]⁺.

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3.52−3.59 (1H, m), 3.91−3.94 (1H, m), 5.14 (2H, s), 5.81 (1H, dd, J =

10.5, 2.0 Hz), 6.38 (1H, d, J = 2.7 Hz), 7.48 (4H, s), 7.83 (1H, d, J = 2.7

Hz). MS (ESI/APCI) m/z 321.2 [M + H]⁺.

to give the title compound (130 mg, 5%) as an oﬀ-white solid; mp 206−

208 °C. ¹H NMR (400 MHz, DMSO-d₆) δ 5.29 (2H, s), 5.92 (1H, d, J =

7.0 Hz), 7.45 (4H, s), 7.72 (1H, d, J = 6.8 Hz), 11.36 (s, 1H). ¹³C NMR

(75 MHz, DMSO-d₆) δ 66.3, 93.5, 128.4, 130.0, 132.6, 135.2, 146.1,

156.1, 171.1. MS (ESI/APCI) m/z 237.2 [M + H]⁺. Anal. Calcd for

C₁₁H₉ClN₂O₂·0.14H₂O: C, 55.24; H, 3.91; N, 11.71. Found: C, 55.23;

H, 3.84; N, 11.92.

4-[(4-Chlorobenzyl)oxy]-1-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)pyrimidin-2(1H)-one (35). The title compound was

prepared in 18% using 11a and 34 in an analogous manner to 10g. Oﬀ-

white solid; mp 205−207 °C. ¹H NMR (400 MHz, DMSO-d₆) δ 0.85−

0.92 (4H, m), 2.05−2.09 (1H, m), 2.47 (3H, s), 5.40 (2H, s), 6.21 (1H,

d, J = 7.2 Hz), 7.19 (1H, dd, J = 9.5, 1.6 Hz), 7.44−7.52 (5H, m), 8.11

(1H, d, J = 7.2 Hz), 8.47 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 7.8,

8.0, 8.1, 66.9, 94.7, 115.0, 116.6, 122.1, 122.8, 127.1, 128.5, 130.1, 132.8,

134.9, 141.9, 145.2, 150.1, 154.9, 171.1. MS (ESI/APCI) m/z 407.2 [M

+ H]⁺. Anal. Calcd for C₂₂H₁₉ClN₄O₂·0.18H₂O: C, 64.43; H, 4.76; N,

13.66. Found: C, 64.42; H, 4.70; N, 13.57.

1-(4-Chlorophenyl)-3-(2,4-dichloropyridin-3-yl)propan-1-one

(37). To a solution of 36 (1.12 g, 4.09 mmol) in DMF (40 mL) were

added 1-(4-chlorophenyl)prop-2-en-1-ol (1.03 g, 6.13 mmol), NaHCO₃

(0.687 g, 8.18 mmol), and Pd(OAc)₂(0.092 g, 0.41 mmol), and the

mixture was heated at 120 °C for 15 h. The mixture was poured into

water and extracted with EtOAc. The organic layer was separated,

washed with water and brine, dried over MgSO₄, and concentrated. The

residue was puriﬁed by column chromatography (silica gel, hexane/

EtOAc = 100/0 to 85/15) to give the title compound (0.772 g, 60%) as a

light-brown amorphous solid. ¹H NMR (400 MHz, CDCl₃) δ 3.20−3.26

(2H, m), 3.32−3.39 (2H, m), 7.29 (1H, d, J = 5.1 Hz), 7.45 (2H, d, J =

8.5 Hz), 7.92 (2H, d, J = 8.5 Hz), 8.19 (1H, d, J = 5.1 Hz).

1-(4-Chlorophenyl)-3-(2,4-dichloropyridin-3-yl)propan-1-ol (38).

To a solution of 37 (100 mg, 0.32 mmol) in THF (3 mL) was added

LiBH₄(6.92 mg, 0.32 mmol) at rt. After 1 h, the mixture was quenched

with satd NaHCO₃solution and extracted with EtOAc. The organic

layer was separated, washed with satd NaHCO₃and brine, dried over

MgSO₄, and concentrated. The residue was puriﬁed by column

chromatography (silica gel, hexane/EtOAc = 100/0 to 70/30) to give

the title compound (83 mg, 82%) as a colorless oil. ¹H NMR (400 MHz,

CDCl₃) δ 1.93−2.10 (3H, m), 2.85−2.98 (1H, m), 3.01−3.14 (1H, m),

4.81 (1H, t, J = 7.8 Hz), 7.24 (1H, d, J = 5.1 Hz), 7.34 (4H, s), 8.13 (1 H,

d, J = 5.1 Hz).

5-(4-Chlorobenzyloxy)-2H-pyridazin-3-one (27). To a stirred

suspension of 26 (210 mg, 0.65 mmol) in MeOH (5 mL) was added

conc HCl (0.5 mL) at rt, and then the reaction mixture was heated at

reﬂux for 3 h. The mixture was cooled to rt, concentrated, and

neutralized with satd NaHCO₃(20 mL). The resulting precipitate was

collected by ﬁltration, washed with water, and dried to give the title

compound (140 mg, 91%) as an oﬀ-white solid. ¹H NMR (400 MHz,

DMSO-d₆) δ 5.12 (2H, s), 6.27 (1H, s), 7.48 (4H, s), 7.72 (1H, d, J = 2.5

Hz), 12.6 (1H, br s). MS (ESI/APCI) m/z 237.2 [M + H]⁺.

5-[(4-Chlorobenzyl)oxy]-2-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)pyridazin-3(2H)-one (28). The title compound was

prepared in 70% yield using 11a and 27 in an analogous manner to 10g.

White solid; mp 190−192 °C. ¹H NMR (400 MHz, DMSO-d₆) δ 0.88−

0.92 (4H, m), 2.06−2.09 (1H, m), 2.47 (3H, s), 5.23 (2H, s), 6.54 (1H,

d, J = 2.6 Hz), 7.23 (1H, dd, J = 9.3, 1.7 Hz), 7.46 (1H, d, J = 9.4 Hz),

7.50−7.55 (4H, m), 8.01 (1H, d, J = 2.7 Hz), 8.46 (1H, s). ¹³C NMR

(101 MHz, DMSO-d₆) δ 7.7, 7.9, 8.0, 69.5, 104.7, 114.8, 116.5, 121.3,

121.8, 127.9, 128.6, 130.0, 133.1, 133.6, 134.0, 141.8, 145.2, 159.0, 160.7.

MS (ESI/APCI) m/z 407.2 [M + H]⁺. Anal. Calcd for C₂₂H₁₉ClN₄O₂:

C, 64.94; H, 4.71; N, 13.77. Found: C, 64.88; H, 4.82; N, 13.55.

6-[(4-Chlorobenzyl)oxy]pyrimidin-4(3H)-one (30). To a stirred

solution of 29 (2.5 g, 22.3 mmol) in THF (25 mL) was added

Ag₂CO₃(15.33 g, 55.75 mmol) and 4-chlorobenzyl bromide (4.58 g,

22.3 mmol). The resultant mixture was heated at reﬂux for 2 h. The

reaction mixture was then cooled to rt, ﬁltered through Celite, and

concentrated. The crude material was puriﬁed by column chromatog-

raphy (silica gel, DCM/MeOH = 95/5 to 90/10) to aﬀord the title

compound (500 mg, 10%) as a white solid; mp 217−219 °C. ¹H NMR

(400 MHz, DMSO-d₆) δ 5.23 (2H, s), 5.60 (1H, s), 7.44 (4H, m), 8.11

(1H, s), 12.39 (1H, s). ¹³C NMR (75 MHz, DMSO-d₆) δ 67.4, 91.7,

128.4, 129.6, 132.6, 135.3, 150.4, 163.1, 169.2. MS (ESI/APCI) m/z

237.2 [M + H]⁺. Anal. Calcd for C₁₁H₉ClN₂O₂·0.6H₂O: C, 55.49; H,

3.88; N, 11.77. Found: C, 55.53; H, 3.89; N, 11.70.

6-[(4-Chlorobenzyl)oxy]-3-(2-cyclopropyl-3-methylimidazo[1,2-

a]pyridin-6-yl)pyrimidin-4(3H)-one (31). To a mixture of 30 (150 mg,

0.63 mmol) and (2-cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-

boronic acid (275 mg, 1.26 mmol) in a mixture of DCM (15 mL) and

MeOH (15 mL) were added Cu(OAc)₂(346 mg, 1.89 mmol) and

pyridine (0.5 mL, 6.3 mmol). The resulting reaction mixture was stirred

at rt for 16 h. The insoluble material was then ﬁltered through Celite and

the ﬁltrate was poured into 1 N HCl solution. The mixture was extracted

with DCM (100 mL) twice, and the combined DCM layers were washed

with satd NaHCO₃solution (50 mL) and brine (50 mL) successively,

dried over Na₂SO₄, and concentrated. The resulting residue was puriﬁed

by preparative HPLC to aﬀord the title compound (15 mg, 6%) as an oﬀ-

white solid; mp 225−226 °C. ¹H NMR (400 MHz, CD₃OD) δ 0.95−

1.02 (4H, m), 2.05 (1H, m), 2.52 (3H, s), 5.37 (2H, s), 5.88 (1H, s), 7.24

(1H, dd, J = 9.5, 1.8 Hz), 7.38−7.49 (5H, m), 8.42 (2H, d, J = 6.2 Hz).

¹³C NMR (75 MHz, DMSO-d₆) δ 7.8, 8.00, 8.04, 67.8, 91.0, 114.9,

5-Chloro-2-(4-chlorophenyl)-3,4-dihydro-2H-pyrano[3,2-c]-

pyridine (39). To a solution of 38 (678 mg, 2.14 mmol) in DMF (5 mL)

was added NaH (60% oil dispersion, 86 mg, 2.14 mmol) at 0 °C, and the

mixture was allowed to warm to rt with vigorous stirring. After 17 h, the

mixture was quenched with water and extracted with EtOAc. The

organic layer was separated, washed with 0.1 N NaOH solution and

brine, dried over MgSO₄, and concentrated. The residue was puriﬁed by

column chromatography (silica gel, hexane/EtOAc = 100/0 to 70/30)

to give the title compound (386 mg, 64%) as a pale-yellow amorphous

1

solid. H NMR (400 MHz, CDCl₃) δ 1.95−2.13 (1H, m), 2.26−2.37

(1H, m), 2.79−3.02 (2H, m), 5.21 (1H, d, J = 9.9 Hz), 6.99 (1H, d, J =

5.4 Hz), 7.29−7.47 (4H, m), 7.97−8.11 (1H, m).

116.6, 122.5, 123.0, 123.7, 128.5, 129.7, 132.7, 135.2, 142.1, 145.3, 152.4,

161.9, 168.6. MS (ESI/APCI) m/z 407.0 [M + H]⁺. Anal. Calcd for

C₂₂H₁₉ClN₄O₂: C, 64.94; H, 4.71; N, 13.77. Found: C, 64.71; H, 4.74;

N, 13.62.

2-Chloro-4-[(4-chlorobenzyl)oxy]pyrimidine (33). To a stirred

solution of 32 (10.4 g, 70.1 mmol) and (4-chlorophenyl)methanol

(10j, 70.0 mmol) in DMF (50 mL) was added K₂CO₃(14.5 g, 105.2

mmol), and the resulting mixture was stirred at rt for 16 h. The mixture

was then diluted with water (100 mL), and the resulting precipitate was

collected by ﬁltration and dried to give the title compound (4.8 g, 27%)

as an oﬀ-white solid; mp 112−120 °C. ¹H NMR (400 MHz, DMSO-d₆)

δ 5.40 (2H, s), 7.06 (1H, d, J = 5.6 Hz), 7.46−7.52 (4H, m), 8.49 (1H, d,

J = 5.7 Hz). MS (ESI/APCI) m/z 255.2 [M + H]⁺.

2-(4-Chlorophenyl)-2,3,4,6-tetrahydro-5H-pyrano[3,2-c]pyridin-

5-one (40). The mixture of 39 (384 mg, 1.37 mmol), ammonium acetate

(528 mg, 6.85 mmol), and AcOH (5 mL) was heated at 200 °C for 1 h

under microwave irradiation. The solvent was evaporated and the

residue was dissolved to EtOAc. The mixture was poured into satd

NaHCO₃solution and extracted with EtOAc. The organic layer was

separated, washed with satd NaHCO₃solution and brine, dried over

MgSO₄, and concentrated. The residue was puriﬁed by column

chromatography (silica gel, EtOAc/MeOH = 100/0 to 90/20) to give

the title compound (112 mg, 31.2%) as a pale-orange amorphous solid.

¹H NMR (400 MHz, CDCl₃) δ 1.86−2.00 (1H, m), 2.25 (1H, dt, J =

13.9, 2.7 Hz), 2.48−2.80 (2H, m), 4.99−5.06 (1H, m), 6.01 (1H, d, J =

7.3 Hz), 7.15−7.19 (1H, m), 7.27 (1H, s), 7.29−7.35 (2H, m), 7.35−

7.40 (2H, m). MS (ESI/APCI) m/z 279.8 [M + H]⁺.

2-(4-Chlorophenyl)-6-(2-cyclopropyl-3-methylimidazo[1,2-a]-

pyridin-6-yl)-2,3,4,6-tetrahydro-5H-pyrano[3,2-c]pyridin-5-one (41).

A mixture of 40 (30 mg, 0.11 mmol), 11a (68.3 mg, 0.23 mmol),

4-[(4-Chlorobenzyl)oxy]pyrimidin-2-ol (34). To a stirred mixture of

33 (2.8 g, 11.0 mmol) in dioxane and water (1:4, 30 mL) was added

NaOH (440 mg, 11 mmol), and the mixture was heated at reﬂux for 3 h.

The reaction mixture was then cooled to 0 °C, and the precipitate was

collected by ﬁltration, washed with cold water, and dried under vacuum

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DMEDA (0.018 mL, 0.17 mmol), CuI (24.0 mg, 0.13 mmol), K₂CO₃

(47.5 mg, 0.34 mmol), and DMSO (2 mL) was heated at 150 °C for 1 h

under microwave irradiation. The reaction mixture was ﬁltered through

NH silica gel (EtOAc/MeOH = 90/10). The ﬁltrate was poured into 1

N NaOH solution and extracted with EtOAc. The organic layer was

separated, washed with 0.1 N NaOH solution and brine, dried over

MgSO₄, and concentrated. The residue was puriﬁed by column

chromatography (NH silica gel, EtOAc/MeOH = 100/0 to 90/10) to

give the title compound (28.4 mg, 57%) as white crystals. ¹H NMR (400

MHz, DMSO-d₆) δ 0.81−0.97 (4H, m), 1.88−2.02 (1H, m), 2.02−2.15

(1H, m), 2.22 (1H, d, J = 14.6 Hz), 2.48 (3H, s), 2.53−2.60 (2H, m),

5.23 (1H, d, J = 8.3 Hz), 6.12 (1H, d, J = 7.5 Hz), 7.13 (1H, d, J = 9.4

Hz), 7.40−7.54 (5H, m), 7.59 (1H, d, J = 7.7 Hz), 8.37 (1H, s). ¹³C

NMR (75 MHz, DMSO-d₆) δ 7.8, 8.0, 8.1, 19.2, 27.9, 76.7, 99.9, 106.7,

114.9, 116.5, 122.0, 123.3, 127.5, 128.0, 128.5, 132.5, 137.2, 139.4, 141.9,

145.1, 161.8, 162.1. MS (ESI/APCI) m/z 432.4 [M + H]⁺. Anal. Calcd

for C₂₅H₂₂ClN₃O₂·0.55H₂O: C, 67.91; H, 5.25; N, 9.47. Found: C,

67.96; H, 5.27; N, 9.51.

7.4) supplemented with 5 mM EDTA (ethylenediaminetetraacetic acid)

were respectively suspended human MCHR1-expressing CHO cells (1

× 10⁸cells) and centrifuged. Homogenate buﬀer [10 mL, 10 mM

NaHCO₃, 5 mM EDTA, pH 7.5, 0.5 mM PMSF (phenylmethylsulfonyl

ﬂuoride), 20 mg/L leupeptin, 4 mg/L E-64, 1 mg/L pepstatin A] was

added to the pellets of the cells and, using Polytron homogenizer, the

mixture was homogenated. The supernatant obtained after centrifuga-

tion at 400g for 10 min was further centrifuged at 100000g for 1 h to give

precipitate of the membrane fraction. The precipitate were suspended in

2 mL of assay buﬀer [20 mM Tris-HCl (pH 7.5), 5 mM EDTA, 0.5 mM

PMSF, 20 mg/L leupeptin, 4 mg/L E-64, 1 mg/L pepstatin A]. The

membrane fractions were suspended in assay buﬀer to a protein

concentration of 2 mg/mL, and after dispensing, preserved at −80 °C

and used upon thawing each time when in use.

2. Binding Assay. An MCHR1-expressing CHO cellular membrane

fraction (173 μL) diluted with an assay buﬀer was dispensed to a 96-well

polypropylene plate (3363, Corning). DMSO solution (2 μL), 33 μM

cold MCH (1−19) diluted with DMSO solution (2 μL), or a test

compound solution diluted with DMSO solution to various

concentrations (2 μL) was added, and last, [¹²⁵I]-MCH(4−19) diluted

with assay buﬀer (hereinafter, sometimes to be referred to as “hot

MCH”, 25 μL) was added to each well. The mixture was reacted with

stirring at rt for 1 h, and the plate was set on FilterMate harvester

(PerkinElmer). Using a polyethylenimine-treated glass ﬁlter plate (GF/

C, PerkinElmer), which had been previously set, the plate was suction-

ﬁltered and washed three times with washing buﬀer (50 mM Tris-HCl

buﬀer pH 7.5). The glass ﬁlter plate was dried, MicroScint 0

(PerkinElmer) was added at 25 μL/well, and the resulting radioactivity

was measured by TopCount liquid scintillation counter (PerkinElmer).

The binding inhibition rate of the test compound was calculated by the

following formula.

3-[5-(4-Chlorophenyl)furan-2-yl]prop-2-enoyl azide (43). To a

solution of 42 (1 g, 4.02 mmol) and triethylamine (0.729 mL, 5.23

mmol) in acetone (20 mL) at 0 °C was added dropwise isobutyl

carbonochloridate (0.684 mL, 5.23 mmol). After stirring for 1 h at 0 °C,

sodium azide (0.340 g, 5.23 mmol) in water (4 mL) was added, and the

resultant mixture was stirred for further 30 min at 0 °C and at rt for 30

min. Water (40 mL) was added, and the resulting precipitate was

collected by ﬁltration, washed with water, and dried to give the title

1

compound (1.10 g, quant) as a yellow solid. H NMR (300 MHz,

CDCl₃) δ 6.37 (1H, d, J = 15.5 Hz), 6.71−6.82 (2H, m), 7.35−7.42 (1H,

m), 7.49 (1H, d, J = 15.5 Hz), 7.60−7.71 (2H, m). ¹³C NMR (75 MHz,

DMSO-d₆) δ 109.9, 115.5, 120.8, 126.2, 127.9, 129.1, 132.2, 133.4,

149.8, 155.3, 171.1.

2-(4-Chlorophenyl)furo[3,2-c]pyridin-4(5H)-one (44). To a stirred

mixture of diphenylether (40 mL) and tributylamine (10.0 mL, 41.7

mmol) at 200 °C was added dropwise a solution of 43 (3.83 g, 14.0

mmol) in diphenylether (60 mL) and THF (20 mL). After addition, the

resulting brown mixture was stirred for 30 min before cooling to rt.

Hexane (200 mL) was added, and the resulting suspension was ﬁltered.

The precipitate was washed with EtOH, ﬁltered, and dried to give the

Binding inhibition (%) = 100 − (radioactivity upon addition of test

compound and hot MCH − radioactivity upon addition of cold MCH

and hot MCH solution)/(radioactivity upon addition of DMSO

solution and hot MCH − radioactivity upon addition of cold MCH

and hot MCH solution) × 100

Measurement of MCH Receptor 1 Antagonistic Activity of

Test Compound Using Ca²⁺Mobilization Assay. Using an

expression vector plasmid introduced with human MCHR1 gene for

expression in animal cells, human MCHR1 gene was introduced into

CHO cells (CHO dhfr⁻) by Lipofectamine LTX (Invitrogen). The cells

were cultured in selection MEMα medium [445 mL of MEMα medium

without nucleic acid and added with 5 mL of penicillin−streptomycin

(Invitrogen) and 50 mL of dialyzed fetal bovine serum]. Colony 24

clones grown in the selection medium, which were human MCHR1

gene-expressing CHO cell candidates, were selected. From these clones,

clone no. 4 which showed the highest response to the change of Ca²⁺

concentration on stimulation by the addition of 25 nM ligand MCH (4−

19) was selected by Ca²⁺mobilization assay. In the following test, this

human MCHR1-expressing CHO cell (clone no. 4) was used. An

integrated dispe×nsing function ﬂuorometer (CellLux, PerkinElmer)

was used for Ca²⁺mobilization assay. The CHO cells were sown in a 96-

well plate (type 3904, Corning) with a black wall and clear well bottom

at a density of 20000 cells/well and cultured in an incubator for about 24

h at 5% CO₂, 37 °C. The medium was removed, and the cells were

washed with phosphate buﬀered saline (PBS). A Ca²⁺indicator dye

reagent (DOJINDO LABORATORIES, Ca screening no-wash kit

Fluo4) was added at 100 μL/well, and the dye was allowed to penetrate

into the cell for 30 min in an incubator at 5% CO₂, 37 °C. The plate was

set on a plate reader. First, a test compound solution diluted with an

assay buﬀer [10 mM HEPES (pH 7.4): 1 Assay Buﬀer (DOJINDO

LABORATORIES, attached to Ca screening no-wash kit Fluo4)

containing 0.1% BSA] or DMSO solution was added at 50 μL/well,

and then ligand MCH (4−19) peptide (ﬁnal concentration 2 nM)

diluted with assay buﬀer or DMSO was added at 50 μL/well, during

which changes in intracellular ﬂuorescence were measured at 2 second

intervals. The antagonistic activity of the test compound was calculated

by the following formula and shown as an inhibition rate (%) wherein

the intracellular ﬂuorescence activity resulting from the stimulation by

1

title compound (340 mg, 9.9%) as a light-brown amorphous solid. H

NMR (300 MHz, DMSO-d₆) δ 6.70 (1H, d, J = 7.9 Hz), 7.34 (1H, d, J =

7.2 Hz), 7.49−7.59 (3H, m), 7.87 (2H, d, J = 8.7 Hz), 11.51 (1H, br s).

¹³C NMR (75 MHz, DMSO-d₆) δ 94.4, 103.0, 117.3, 125.8, 128.0, 129.0,

132.1, 133.0, 152.2, 159.1, 159.8. MS (ESI/APCI) m/z = 246.0 [M +

H]⁺.

2-(4-Chlorophenyl)-5-(2-cyclopropyl-3-methylimidazo[1,2-a]-

pyridin-6-yl)furo[3,2-c]pyridin-4(5H)-one (45). A mixture of 44 (100

mg, 0.41 mmol), (2-cyclopropyl-3-methylimidazo[1,2-a]pyridin-6-yl)-

boronic acid (106 mmol, 0.49 mmol), Cu(OAc)₂(4.99 mg, 0.04 mmol),

pyridine (66.0 μL, 0.81 mmol), MS4A (48.9 mg), and DMF (5 mL) was

stirred at rt for 4 h and at 50 °C overnight. After ﬁltration of the reaction

mixture through Celite, the ﬁltrate was poured into 1 N HCl at rt and

extracted with EtOAc. The organic layer was separated, washed with

satd NaHCO₃solution and brine successively, dried over MgSO₄, and

concentrated in vacuo. The residue was puriﬁed by column

chromatography (silica gel, hexane/EtOAc = 20/80 to 0/100). The

solid was collected and washed with EtOH to give the title compound

1

(10 mg, 5%) as an oﬀ-white solid; mp 278−279 °C. H NMR (300

MHz, DMSO-d₆) δ 0.84−0.99 (4H, m), 2.04−2.14 (1H, m), 6.94 (1H,

dd, J = 7.6, 0.8 Hz), 7.21 (1H, dd, J = 9.4, 1.9 Hz), 7.49 (1H, d, J = 9.4

Hz), 7.53−7.59 (2H, m), 7.66 (1H, s), 7.75 (1H, d, J = 7.6 Hz), 7.88−

7.96 (2H, m), 8.48 (1H, d, J = 1.5 Hz). ¹³C NMR (75 MHz, DMSO-d₆)

δ 7.8, 8.0, 8.1, 95.3, 103.4, 115.0, 116.6, 117.1, 122.4, 123.3, 125.9, 127.3,

128.0, 129.1, 133.0, 137.1, 142.0, 145.2, 153.2, 158.4, 159.2. MS (ESI/

APCI) m/z = 416.1 [M + H]⁺. Anal. Calcd for C₂₄H₁₈ClN₃O₂: C, 69.31;

H, 4.36; N, 10.10. Found: C, 69.11; H, 4.48; N, 10.09.

Determination of hMCHR1 Competitive Inhibitory Activity of

Test Compound Using Binding Assay. 1. Preparation of

Membrane Fraction. Using hMCHR1-expressing CHO cell clone

57,⁶³MCHR1-expressing CHO cellular membrane fractions were

prepared by the following method. In phosphate buﬀered saline (pH

T

DOI: 10.1021/acs.jmedchem.5b01704

J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

the addition of ligand MCH (4−19) peptide was 100% and that of the

well added with DMSO solution alone was 0%.

period, and food intake was measured every 2 or 3 days. The change in

body weight was presented as percentage from initial body weight.

In Vivo Selectivity of Anorectic Eﬀect by Using MCHR1-

Deﬁcient Mice. Male MCHR1-deﬁcient mice and wild-type litter mate

mice (45 weeks old) loaded with a high-fat diet (D12451) from 5 weeks

of age were used. Before the start of the experiment, the mice were

independently raised, a high-fat diet (D12451) was given, and tap water

(0.5 mL) was administered for acclimation. The mice were grouped on

the basis of food intake from day−3 to day−1 and body weight of day−1

as indices. Each group was orally administered vehicle (0.5%

methylcellulose solution) or compounds suspended in vehicle at 10

mL/kg for 3 days (6 mice per group). Food intake for 3 days was

measured.

Inhibitory rate (%) = 100 − [ﬂuorescence activity upon addition of

test compound and MCH(4−19) peptide solution − ﬂuorescence

activity upon addition of DMSO solution only]/[ﬂuorescence activity

upon addition of DMSO solution and MCH(4−19) peptide solution −

ﬂuorescence activity upon addition of DMSO solution only] × 100.

Evaluation of PLsis Inducing Potential.⁶⁴DMEM medium, L-

glutamine, penicillin−streptomycin, pyruvic acid, and N-(7-nitrobenz-2-

oxa-l,3-diazol-4-yl)-l,2-hexadecanoyl-sn-glycero-3-phosphoethanol-

amine triethylammonium salt (NBD-PE) were purchased from

Invitrogen Corporation. As bovine serum albumin (BSA), a product

of Thermo Trace Ltd. (Melbourne, Australia) was used, and as

Amiodarone, a product of ICN (Costa Mesa, CA) was used. A test

compound was used in the form of a 10 mM DMSO solution.

FBS was added at a ﬁnal concentration of 5 vol % to DMEM medium

supplemented with ^L-glutamine, pyruvic acid, and penicillin−

streptomycin and subjected to the experiment. Culture was performed

using 5% carbon dioxide gas−95% air as a gas phase in a CO₂incubator

at 37 °C. HepG2 cells were suspended in a culture medium at 50 × 10⁴

cells/mL, plated in a 96-well plate at 50 μL/well, and precultured for 24

h. After preculture, the culture medium was removed, a culture medium

containing 60 μM NBD-PE was added at 50 μL/well, and a culture

medium containing 0.6 μM or 20 μM test compounds were each added

at 50 μL/well to HepG2 cells, and the cells were cultured for 24 h. As a

positive control, Amiodarone was used at a ﬁnal concentration of 10 μM.

After exposure to the test compound for 24 h, the ﬂuorescence

intensity (ex 485 nm, em 538 nm) of NBD-PE uptaken by the cells was

measured by a ﬂuorometer. The measurement value with addition of 0

μM test compound solution was subtracted as a blank, a relative value to

the measurement value with addition of 10 μM Amiodarone was

calculated, and the maximum value per unit concentration of the test

compound was obtained as a phospholipidosis inducing potential.

Estimation of log D at pH 7.4. Log D_7.4, which is a partition

coeﬃcient between 1-octanol and aqueous buﬀer at pH 7.4 of the

compounds was measured using the chromatographic procedure whose

condition was developed based on a published method.⁵⁵The

instruments were a Waters Alliance 2795 HPLC system with a 2996

UV−vis detector (Milford, MA, USA).

ASSOCIATED CONTENT

* Supporting Information

The Supporting Information is available free of charge on the

ACS Publications website at DOI: 10.1021/acs.jmed-

chem.5b01704.

■

S

Experimental procedures for the synthesis of 2,3-

dimethylimidazo[1,2-a]pyridin-6-amine, 2-cyclopropyl-3-

methylimidazo[1,2-a]pyridin-6-amine, 2-cyclopropyl-3-

methylindolizin-6-amine hydrochloride, (2-cyclopropyl-

3-methylimidazo[1,2-a]pyridin-6-yl)boronic acid, 5-hy-

droxy-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one

(25), and 6-iodo-3-methylimidazo[1,2-a]pyridine-2-car-

boxylic acid (PDF)

Molecular formula strings (CSV)

AUTHOR INFORMATION

Corresponding Author

*Phone: +81 466 32 1057. Fax: +81 466 29 4468. E-mail:

hideyuki.igawa@takeda.com.

■

Notes

The authors declare no competing ﬁnancial interest.

ACKNOWLEDGMENTS

Animal Experiments. All animal experiments were performed in

compliance with the Guidelines for the Care and Use of Laboratory

Animals of Takeda Pharmaceutical Company Ltd.

■

We acknowledge Dr. Kaneyoshi Kato, Dr. Toshiki Murata, and

Dr. Makoto Kamata for valuable suggestions for drug design. We

thank Dr. Mitsuyuki Shimada for the alliance with TCG

Lifesciences, Ltd., and Dr. Tomohiko Suzaki for arrangement

of compound shipment. We acknowledge Dr. Shoki Okuda for

valuable suggestions to make this manuscript.

Pharmacokinetic Analysis in Rat Cassette Dosing. Test

compounds were administered intravenously (0.1 mg/kg) or orally (1

mg/kg, suspended in 0.5% methylcellulose aqueous solution) by

cassette dosing to nonfasted rats. After administration, blood samples

were collected and centrifuged to obtain the plasma fraction. The plasma

samples were deproteinized by mixing with acetonitrile followed by

centrifugation. The compound concentrations in the supernatant were

measured by LC/MS/MS.

Evaluation of Anorectic Eﬀect Using Male DIO F344/Jcl Rats.

Male DIO F344/Jcl rats (50 weeks old) fed with a high-fat diet

(D12451: Research Diets) from 5 weeks old were used. From before the

start of experiment, the rats were singly housed, given a powder high-fat

diet (D12451M: Research Diets), and habituated to oral administration

with tap water. The rats were grouped based on both the food intake and

the body weight of day−1. The rats were orally administered vehicle

(0.5% methylcellulose solution) or compounds suspended in vehicle at

2 mL/kg 1 h before the onset of dark period for 2 days (6 rats per group).

The food intake for 2 days from the initial administration was measured.

The food intake inhibition rate of each compound administration group

to the vehicle group was calculated.

ABBREVIATIONS USED

■

hERG, human ether-a-go-go related gene; WSC, 1-ethyl-3-(3-

(dimethylamino)propyl) carbodiimide hydrochloride; HOBt, 1-

hydroxybenzotriazole; DMEDA, N,N′-dimethylethylenedi-

amine; DIPEA, N,N-diisopropylethylamine; ADDP, 1,1′-

(azodicarbonyl)dipiperidine; HATU, (1-[bis(dimethylamino)-

methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexa-

ﬂuorophosphate); TMSCN, trimethylsilyl cyanide; TPSA,

topological polar surface area; CHO, Chinese hamster ovary;

TM, transmembrane

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DOI: 10.1021/acs.jmedchem.5b01704

J. Med. Chem. XXXX, XXX, XXX−XXX

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DOI: 10.1021/acs.jmedchem.5b01704

J. Med. Chem. XXXX, XXX, XXX−XXX

Article Doi

Melanin-Concentrating Hormone Receptor 1 Antagonists Lacking an Aliphatic Amine: Synthesis and Structure-Activity Relationships of Novel 1-(Imidazo[1,2-a]pyridin-6-yl)pyridin-2(1H)-one Derivatives

DOI: 10.1021/acs.jmedchem.5b01704

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