Lead optimization of a novel series of imidazo[1,2-a]pyridine amides leading to a clinical candidate (Q203) as a multi- and extensively-drug- resistant anti-tuberculosis agent

Article abstract of DOI:10.1021/jm5003606

A critical unmet clinical need to combat the global tuberculosis epidemic is the development of potent agents capable of reducing the time of multi-drug-resistant (MDR) and extensively-drug-resistant (XDR) tuberculosis therapy. In this paper, we report on the optimization of imidazo[1,2-a]pyridine amide (IPA) lead compound 1, which led to the design and synthesis of Q203 (50). We found that the amide linker with IPA core is very important for activity against Mycobacterium tuberculosis H37Rv. Linearity and lipophilicity of the amine part in the IPA series play a critical role in improving in vitro and in vivo efficacy and pharmacokinetic profile. The optimized IPAs 49 and 50 showed not only excellent oral bioavailability (80.2% and 90.7%, respectively) with high exposure of the area under curve (AUC) but also displayed significant colony-forming unit (CFU) reduction (1.52 and 3.13 log₁₀ reduction at 10 mg/kg dosing level, respectively) in mouse lung.

Full text of DOI:10.1021/jm5003606

Subscriber access provided by the Henry Madden Library | California State University, Fresno
Article
Lead Optimization of a Novel series of Imidazo[1,2-a]pyridine
Amides Leading to a Clinical Candidate (Q203) as a Multi-
and Extensively-Drug Resistant Antituberculosis Agent
Sunhee Kang, Ryang Yeo Kim, Min Jung Seo, Saeyeon Lee, Young Mi Kim, Mooyoung Seo, Jeong
Jea Seo, Yoonae Ko, Inhee Choi, Jichan Jang, Jiyoun Nam, Seijin Park, Hwankyu Kang, Hyung
Jun Kim, Jungjun Kim, Sujin Ahn, Kevin Pethe, Kiyean Nam, Zaesung No, and Jaeseung Kim
J. Med. Chem., Just Accepted Manuscript • DOI: 10.1021/jm5003606 • Publication Date (Web): 28 May 2014
Downloaded from http://pubs.acs.org on May 30, 2014
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a free service to the research community to expedite the
dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts
appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been
fully peer reviewed, but should not be considered the official version of record. They are accessible to all
readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered
to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published
in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just
Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor
changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers
and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors
or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
Journal of Medicinal Chemistry is published by the American Chemical Society. 1155
Sixteenth Street N.W., Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 55
Journal of Medicinal Chemistry
1
Lead Optimization of a Novel series of Imidazo[1,2-a]pyridine Amides Leading to a Clinical Candidate
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
(
Q203) as a Multi- and Extensively-Drug Resistant Antituberculosis Agent
1
2
1
1
1
1
Sunhee Kang , Ryang Yeo Kim , Min Jung Seo , Saeyeon Lee , Young Mi Kim , Mooyoung Seo , Jeong Jea
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
1
1
2
3
3
3
Seo , Yoonae Ko , Inhee Choi , Jichan Jang , Jiyoun Nam , Seijin Park , Hwankyu Kang , Hyung Jun Kim ,
2
4
Jungjun Kim , Sujin Ahn , Kevin Pethe , Kiyean Nam , Zaesung No *, Jaeseung Kim *
3
2
4
1,#,
1
1
Medicinal & Bioorganic Chemistry group, Institut Pasteur Korea, 16, Daewangpango-ro 712Beon-gil,
2
Bundang-gu, Seongnam-si, Gyeonggi-do, 463-400 Korea; Antibacteial Drug Discovery group, Institut Pasteur
3
Korea, 16, Daewangpango-ro 712Beon-gil, Bundang-gu, Seongnam-si, Gyeonggi-do, 463-400 Korea; DMPK
group, Institut Pasteur Korea, 16, Daewangpango-ro 712Beon-gil, Bundang-gu, Seongnam-si, Gyeonggi-do,
4
63-400 Korea; Qurient Co. Ltd, 16, Daewangpango-ro 712Beon-gil, Bundang-gu, Bundang-gu, Seongnam-si,
4
Gyeonggi-do, 463-400 Korea;
#
Present address: Gyeonggi Biocenter, 147 Gwanggyo-ro, Youngtong-gu, Suwon-si, Gyeonggi-do, 443-270
Korea.
RECEIVED DATE (to be automatically inserted after your manuscript is accepted if required according
to the journal that you are submitting your paper to)
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 2 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
ABSTRACT
A critical unmet clinical need to combat the global tuberculosis epidemic is the development of potent agents
capable of reducing the time of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis
therapy. In this paper, we report on the optimization of the imidazo[1,2-a]pyridine amide (IPA) lead compound
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
, which led to the design and synthesis of Q203 (50). We found that the amide linker with IPA core is very
important for activity against Mycobacterium tuberculosis H37Rv. Linearity and lipophilicity of the amine part
in the IPA series play a critical role in improving in vitro and in vivo efficacy and pharmacokinetic profile. The
optimized IPAs, 49 and 50 showed not only excellent oral bioavailability (80.2 and 90.7%, respectively) with
high exposure of the area under curve (AUC) but also displayed significant colony-forming unit (CFU)
reduction (1.52 and 3.13 log reduction at 10 mg/kg dosing level, respectively) in the lung of mice.
1
0
ACS Paragon Plus Environment

Page 3 of 55
Journal of Medicinal Chemistry
1
INTRODUCTION
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Tuberculosis (TB) is a chronic infectious disease caused by Mycobacterium tuberculosis. TB remains a serious
global health problem positioned as the second leading cause of death from a single infectious agent worldwide
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
with an incidence rate of almost 9 million cases and a fatality rate of 1.4 million in 2011 . Notwithstanding the
successful implementation of Directly Observed Treatment Short course (DOTS) in many countries, high
prevalence of MDR and XDR tuberculosis has intensified the urgent need for new anti-tubercular drugs. Several
2
new classes of compounds have been discovered for treatment of tuberculosis in the last decade . A few of
-4
5
-7
them are currently in clinical trials and one of them, bedaquiline, was recently approved for the treatment of
MDR tuberculosis (Figure 1). However, given the emergence of drug resistance and the low success rate
encountered in clinical development, there is still a need to develop additional drug candidates for TB treatment.
8
We recently reported a novel chemical entity named Q203 (50) as a promising anti-TB drug candidate using
9
phenotypic high-content screening (HCS) technology inside infected macrophages . The IPA series targets
8,10
QcrB , which encodes the b subunit of the electron transport ubiquinol cytochrome C reductase. The IPA
1
1-13
series was reported by others as an attractive anti-TB lead series
. Here, we report on the lead optimization
that led to the design the potential clinical candidate 50.
5
Figure 1. Structures of bedaquiline, PA-824 and delamanid.
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 4 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Chemistry. General amide coupling of appropriately substituted imidazo[1,2-a]pyridine-3-carboxylic acid (4a-
4
g) with corresponding amines R3 afforded target compound in over 60 % yield (Scheme 1). One of the
precursor, the imidazo[1,2-a]pyridine-3-carboxylic acid (4a-4g) was prepared starting from bromination of
various β-keto esters except for commercially available ethyl 2-chloro-3-oxobutanoate (2a). Unsubstituted β-
keto esters were treated with N-bromosuccinimide and over two equivalent of NH OAc in Et O to afford the 2-
4
2
1
4
monobrominated product (2b) . Alternatively, β-keto esters could be transformed to 2-bromo products by
bromine (2c-2d) which resulted in comparable yields. The adequate monobrominated β-keto esters were
condensed with the substituted 2-aminopyridines via imine-enamine formation in absolute ethanol at reflux
15
temperature and the resulting esters (3a-3g) were hydrolyzed to acids (4a-4g).
The counter parts, methanamine derivatives, can be classified into three functional groups; i) bi-aryl, ii) 2-
ring system having a saturated cyclic amine, iii) 3-ring system having a saturated cyclic amine between two aryl
rings. The synthetic route of the methanamine is shown in Scheme 2. Suzuki coupling of 4-chlorobenzonitrile
1
3
6
and substituted phenylboronic acid using PdCl (dppf) and aqueous Na CO
2
followed by reduction with
2
ACS Paragon Plus Environment

Page 5 of 55
Journal of Medicinal Chemistry
1
lithium aluminum hydride gave biphenyl methanamines 7a-7f. Exceptionally, biphenyl methanamines having a
cyano group and tert-butyl group, 7f and 7g were synthesized by direct Suzuki coupling with 4-
bromobenzylamine. The benzylamines which have saturated ring contained bis- and tris-ring were prepared in a
straightforward manner as shown in Scheme 3. Commercially available 4-fluorobenzonitrile was reacted with
appropriate cyclic amine and K CO by heating in dimethyl sulfoxide and then reduced with lithium aluminum
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
3
1
hydride to produce benzylamines 8a-8n .
7
The synthetic route for the linker modification is shown in Scheme 4. Compound 9, which has no benzylic
carbon next to amide was prepared via acid chloride activation of 4f and N-methylated compound 10 was
synthesized by treating sodium hydride and iodomethane from compound 1. Amide reduction of 1 in a mild
condition using boron trifluoride etherate and NaBH provided 11 with moderate yield.
4
In the case of 14 and 15, which have one more carbon between the imidazo[1,2-a]pyridine ring and the
carbonyl group of amide bond, 4-brominated β-keto ester is required. Interestingly, the desired 4-brominated β-
keto ester 12 was regioselectively synthesized by treating N-bromosuccinimide with 0.1 equivalent of neutral
catalyst, NH OAc compared to the presence of over 2 equivalent of NH OAc for the synthesis of 2-brominated
4
4
β-keto ester 2b. In this reaction, 2-brominated β-keto ester, 2b was generated initially and then the bromo group
was migrated to 4-position after overnight reaction. However, the migration of the bromo group was not
occurred in the presence of excess amount of NH OAc. 12 was condensed with 2-amino-5-chloropyridine and
4
saponified to afford the intermediate acid 13 followed by general amide coupling afforded 14 and 15. The
reverse-amide compound 17 was synthesized via curtius rearrangement using diphenylphosporyl azide and
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 6 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
triethylamine in t-BuOH. Subsequent de-protection of Boc group by trifluoroacetic acid and amide coupling via
acid chloride activation afforded the target compound.
a
Scheme 1. Synthesis of imidazo[1,2-a]pyridine-3-carboxylic acids 4a-g
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
a
a
b
Reagents and conditions: (i ) NBS, NH OAc (over 2eq.), ether, rt, 6h; (i ) Br , CHCl , 0℃ – rt, 20min; (ii) 2a-
4
2
3
2d, EtOH, reflux, overnight; (iii) LiOH, EtOH/H O (3:1, v/v), rt, overnight; (iv) corresponding amine, EDC,
2
HOBt, TEA, DMF, 80℃, 2-4h
a
Scheme 2. Synthesis of [1,1'-biphenyl]-4-ylmethanamine analogues 7a-g
ACS Paragon Plus Environment

Page 7 of 55
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
a
Reagents and conditions: (i) Pd(dppf)Cl , Na CO , DME/H O (3:1, v/v), 150℃, 1h – 3h; (ii) LAH, THF, 0℃ -
2
2
3
2
reflux, 1h
a
Scheme 3. Synthetic scheme for benzylamine tails 8a-8n
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
a
Reagents and conditions: (i) K CO , corresponding amine, 90-120℃, 3h; (ii) LAH, THF, reflux, 1h
2
3
a
Scheme 4. Synthetic scheme for linker modification
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 8 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
a
Reagents and conditions: (i) SOCl , 100 ℃, 1h, then [1,1'-biphenyl]-4-amine, TEA, MC, rt, 1h; (ii) NaH,
2
CH I, DMF, 0℃ - rt, 1h; (iii) NaBH , BF ·etherate, THF, reflux, 2h; (iv) NBS, NH OAc (0.1 eq.), Et O, rt,
3
4
3
4
2
overnight; (v) 2-amino-5-chloropyridine, ethanol, reflux, overnight; (vi) LiOH, MeOH/H O (3:1, v/v), rt,
2
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
overnight; (vii) EDC, [1,1'-biphenyl]-4-ylmethanamine, HOBt, TEA, DMF, 80℃, 3h; (viii) [1,1’-biphenyl]-4-
amine, EDC, HOBt, TEA, DMF, 80℃, 3h; (viiii) DPPA, TEA, t-BuOH, reflux, overnight; (x) TFA, MC, rt, 1h;
(xi) 2-([1,1'-biphenyl]-4-yl)acetic acid, SOCl , TEA, MC, rt, 1h
2
RESULTS AND DICUSSION
The activity of IPA derivatives was tested against M. tuberculosis replicating inside macrophages (intracellular
8
MIC ) and in liquid broth culture medium (extracellular MIC ) (Tables 1-4), as previously described . In
8
0
80
addition, the metabolic stability of the compounds was evaluated in mouse and human liver microsomal
preparations to study structure-property relationships (SPR) in order to prioritize compounds for in vivo
pharmacokinetic evaluation. The initial structure-activity relationship (SAR) studies evaluated a set of
analogues that contained replacement of the 3-carboxamide linker in an attempt to affect potency of compound
1 (Table 1). Replacement of hydrogen on NH with methyl (10) significantly decreased the activity against M.
tuberculosis H37Rv replicating outside and inside macrophages by approximately 190 and 690-fold.
Surprisingly, the modification of the length of the amide linker with one carbon (14 and 15) between the
imidazo[1,2-a]pyridine ring and the carbonyl group of amide bond abolished the potency against M.
tuberculosis. Introduction of N-phenyl group (9) at the amine position did not give any activity as well. We then
ACS Paragon Plus Environment

Page 9 of 55
Journal of Medicinal Chemistry
1
investigated the contribution of the H-bonding acceptor of the carbonyl group. Modification by a reversed
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
amide (17) or removal of the oxygen on carbonyl group (11) reduced the activity to sub-micromolar range
(intracellular MIC = 200 nM and 690 nM, respectively) compared to the parent compound 1. Accordingly,
80
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
this set of modifications revealed that the carboxamide linker with the N-benzylic group is critical for anti-
mycobacterial activity.
Table 1. Activity of linker modified IPA analogues against M. tuberculosis H37Rv
X
Cl
N
N
Antimycobacterial activity against M. tuberculosis H37Rv
aextracellular
MIC80 (nM)
bintracellular
MIC80 (nM)
X
Compound
1
O
O
NH
N
45
1.39
970
10
9
8690
O
>10000
>10000
>10000
NH
H
N
>10000
15
O
H
N
14
>10000
>10000
O
1
1
N
H
810
200
690
H
1
7
N
1670
O
a
b
extracellular MIC : the inhibitory activity against M. tuberculosis H37Rv replicating in culture broth medium;
8
0
intracellular MIC : the inhibitory activity against M. tuberculosis H37Rv replicating inside macrophages;
80
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 10 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
MIC is the minimum concentration required to inhibit growth of 80 %; MIC indicates average value of two
80 80
independent measurement.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Next, the optimization was focused on R1, R2 and R3 modifications (Table 2). During the exploration of SAR
at R2 and R3 positions, the substituent’s at R1 was limited to H, 6-Cl, and 7-Cl. This is because we found early
on that 6 or 7-Cl substitutions improved antibacterial activity and increased the metabolic stability compared to
other substituent groups (data not shown). To investigate the hydrophobic interaction of R2 alkyl groups, the
four compounds 18-21 were designed to alter the size of R2 position that might disturb positioning of the 3-
carboxamide. As expected, the smaller size groups, methyl and ethyl (18 and 19) showed better activity
(intracellular MIC = 20 nM and 97 nM, respectively) than longer and bulky groups, propyl and iso-propyl (20
80
and 21). Furthermore the sterically hindered compound 21 (intracellular MIC = 3130 nM) was much less
8
0
potent than the linear compound 20 (intracellular MIC = 50 nM). To confirm the improved potency of methyl
8
0
and ethyl group at the R2 position, the compound 1 and 22 were prepared and evaluated. Compound 1, which
possesses an ethyl group, showed approximately 30-fold greater potency against intracellular mycobacteria
compared to methyl substituted compound 22. However, the methyl substitution had a better metabolic stability
compared to the ethyl substitution.
Further exploration of the SAR at the R3 position of the phenyl group having 6- or 7-Cl of imidazo[1,2-
a]pyridine core was conducted by substitution of various functional groups such as donating, withdrawing and
carboxylic acid. For the effect of the position on the benzene ring, a para-chloro group (24) offered much more
potency and metabolic stability than the ortho-chloro group (23). Encouraged by the positive effect of the para-
ACS Paragon Plus Environment

Page 11 of 55
Journal of Medicinal Chemistry
1
substitution on antibacterial effect, we focused on screening several analogues at this position to investigate the
activity and metabolic stability. Intracellular activity of all substituents such as donating and withdrawing
groups (24-28), except a hydrophilic acid, were very similar within a 2-fold range of MIC = 0.4 ~1.3 nM. Even
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
8
0
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
the more lipophilic and sterically hindered compound 30 showed similar intracellular activity comparable to
compound 27. On the other hand, compound 29 that has a hydrophilic carboxylic acid group, gave a deleterious
effect on the activity. In terms of metabolic stability, the alkyl group such as methyl (26 and 27) and tert-butyl
(30) showed much lower microsomal stability than the other groups Cl, CN and acid (24, 25, 28 and 29) in
human and mouse liver microsomes. This again suggested that the position of R3 on the benzene ring may play
a role in potency and microsomal stability.
From our initial SAR study, compound 24, 25 and 28 showed desirable extra- and intracellular potency as
well as metabolic stability in human and mouse liver microsomes to perform in vivo pharmacokinetic (PK)
experiments. However, compound 25 was too insoluble in aqueous and organic solutions caused by extension
of aromatic character to conduct in vivo experiment. Thus, compound 24 was selected and in vivo PK profile
was evaluated in Spraque-Dawley rat after administration by oral (p.o.) and intravenous (i.v.) routes. However,
the half-life and AUC after oral administration could not be calculated because the concentration of compound
detected in the plasma remained constant (or even slightly increased) up to 16 hours after dosing (supporting
information, Table S1). This result suggested that compound 24 was rebounded in absorption after 4 h, could be
subject to compound precipitation in gut and extended absorption due to the highly lipophilic nature of the
compound 24. Therefore, our next optimization strategy was focused on replacement of the second phenyl
group with various ring systems to improve solubility and reduce lipophilicity (Table 3).
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 12 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Table 2. Activity and metabolic stability of bis-phenyl IPA analogues against M. tuberculosis H37Rv
R3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
O
5
4
3
NH
6
N
R1 7
R2
2
N
1
8
Antimycobacterial activity against
Microsomal stability (t1/2, min)
M. tuberculosis H37Rv
extracellular
MIC80 (nM)
intracellular
MIC80 (nM)
Human
Mouse
Compound
R1
R2
R3
1
8
H
H
H
H
Me
Et
H
H
H
H
250
63
20
97
aND
27.1
22.4
31.6
aND
aND
8.7
19
2
0
Pr
1180
3130
50
21
iPr
3130
aND
2
2
6-Cl
6-Cl
6-Cl
Me
Et
H
175
45
42
1.39
9.3
>120
22.8
38.9
>60
19.3
13.1
1
H
23
Et
2-Cl
43
24
6-Cl
6-Cl
Et
Et
4-Cl
0.9
0.45
0.68
83
>60
>60
25
4-CN
<0.5
>120
2
6
6-Cl
7-Cl
7-Cl
7-Cl
Et
Et
Et
Et
4-Me
4-Me
4-Cl
0.7
<0.5
1.3
0.43
1.35
1.01
217
30.5
25.4
>120
>120
40.6
23.1
>60
27
28
29
4-CO H
250
>120
2
t
30
7-Cl
Et
4- Bu
12
0.46
617
180
18.5
50.7
449
Isoniazid(INH)
Rifampicin(RIF)
26.6
a
ND, Not Determined.
The changes were focused on the second phenyl ring on right-hand side while keeping the 7-chloro-2-
ethylimidazo[1,2-a]pyridine-3-carboxamide at the left-hand side. Firstly, we introduced hetero aromatic groups
that were favorable to make a salt formation to improve solubility. However, introduction of heteroaromatic
ACS Paragon Plus Environment

Page 13 of 55
Journal of Medicinal Chemistry
1
rings led to a dramatic reduction in potency (31 and 32, intracellular MIC of 140 and 740 nM, respectively).
80
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Another strategy involved the introducing of nitrogen containing saturated ring next to the first phenyl ring,
such as a piperidine, azepane and piperazine. The nitrogen atom, if appropriately acidic, could potentially
provide an ionizable site. When the piperidine and azepane moieties (33 and 34) were placed at the end, the
cellular activity was maintained compared to heteroaromatic rings, 31 and 32. However, not only the
compounds were heavily metabolized in human and mouse liver microsomes, they also had decreased
antibacterial activity in macrophages compared to the lead compound 1. Furthermore we did not find any sign
of improvement of microsomal stability by substitution at the 4-position on the piperidine ring (35 and 36).
A series of analogues containing piperazine at the end of the phenyl ring were also synthesized and evaluated.
From the previous SAR studies, it was shown that a hydrophilic character on the second ring led to decreased
potency. Thus, we applied small alkyl group such as methyl and iso-propyl at 4-position of piperazine to protect
activity loss. Unlike piperidine compounds 35 and 36, compounds 37 and 38 lost their potency against M.
tuberculosis H37Rv replicating inside and outside macrophage (intracellular MIC > 140 nM). On the other
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
8
0
hand, compounds 39 and 40 with the fused ring showed a potency comparable to compound 1, whether or not
the fused ring had an aromaticity. The excellent potency of the lipophilic and bulky analogues 39 and 40
suggested that there is more lipophilic space and that the two-ring system could be extended to a three-ring
system on the right-hand side. In addition, compound 25 having a nitrile group that is representative linearity or
aromaticity next to the biphenyl ring, showed a superior potency with MIC value of less than 1 nM against M.
80
tuberculosis H37Rv replicating outside and inside macrophages, as well as good metabolic stability in human
and mouse liver microsomes (Table 2).
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 14 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Table 3. Activity of IPA analogues containing two or three rings against M. tuberculosis H37Rv
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
a
ND, Not Determined.
With this structure-activity relationship (SAR) analysis, two analogues 41 and 42 were designed and
evaluated against M. tuberculosis H37Rv replicating outside and inside macrophage. Compound 41 with a 4-
ACS Paragon Plus Environment

Page 15 of 55
Journal of Medicinal Chemistry
1
fluorophenyl piperazine showed not only dramatically increased with MIC values of 0.3 nM compared to
80
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
analogue 38 (MIC =140 nM) against bacteria replicating inside macrophages but also approximately 3-fold
8
0
improved stability in mouse liver microsomes. In the same manner, the 4-fluorophenyl piperidine analogue 42
showed over 10-fold better potency by incorporation of phenyl ring than analogue 33 (intracellular MIC = 8.2
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
8
0
nM) with better microsomal stability in mouse. However, the microsomal stability still required to be more
improved.
Table 4. Activity of IPA analogues containing three ring system against M. tuberculosis H37Rv
a
b
Values were determined by LC/MS method; The assay was performed using recombinant CYP enzymes and
analyzed by LC/MS/MS
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 16 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
To improve the microsomal stability, substituents on the third aromatic ring were investigated with 7-chloro-
or 6-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxamide as a left-hand side. As shown in Table 4, the
analogues 47 and 48, in which the fluorine was replaced by chlorine, had a retained potency (intracellular
MIC = 0.46 nM and 1.3 nM, respectively) and improved microsomal stability (t > 60 min, mouse
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
8
0
1/2
microsomes). Other analogues (44-45 and 49-50) bearing a trifluoromethoxy substituent showed a similar
activity and good stability in human and mouse microsome. All analogues in Table 4 displayed unprecedented
MIC values in single-digit nanomolar or sub-nanomolar range against M. tuberculosis H37Rv replicating both
80
outside and inside macrophage. In terms of metabolic stability, the stability-substituent correlation was followed
in order of trifluoromethoxy > chlorine > fluorine and most compounds had good metabolic stability to perform
in vivo experiment except 4-fluorophenyl piperidine analogues 42 and 46. In addition to the stability, the
incorporation of ionizable saturated ring between two aryl-rings compared to bis-phenyl IPA analogues (24-26)
resulted in improved solubility under acidic condition without activity loss (supporting information, Table S2).
All analogues were also evaluated for cytochrome P450 inhibition with 5-different isozymes in order to
prioritize compounds for in vivo PK experiments. Drug-drug interaction is a critical factor to develop an anti-
TB agent due to combination therapy with other TB drugs or HIV drugs for co-infected treatment. Based on
their overall properties, compounds 41, 49 and 50 were shortlisted for in vivo PK experiments.
Table 5. In vivo pharmacokinetic values in mice of selected compounds, 41, 49 and 50
ACS Paragon Plus Environment

Page 17 of 55
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
a
PK values for compound 50 were adapted from reference 8 and presented for the sake of comparison.
The in vivo PK properties of compounds 41, 49 and 50 were evaluated in mice after intravenous (i.v.) and oral
(p.o.) administration of 2 and 10 mg/kg, respectively. As shown in Table 5, those compounds displayed good
PK properties with long half-life, low systemic clearance and moderate to high volume of distributions. After
oral dosing, all compounds reached a maximum concentration in plasma within 2 hours, their elimination half-
life was favorable (9.4, 21.3 and 23.4 hours, respectively) and the area under curve (AUC) was 59,576, 27,349
.
and 44,100 ng h/mL, respectively. Overall, they achieved good oral exposure in systemic circulation that
resulted in superior oral bioavailability (69.9, 80.2 and 90.7%, respectively).
On the basis of the promising in vivo PK profile, we conducted in vivo efficacy studies for compound 49 in
8
an established mouse model under the same condition as previously described . BALB/c mice were infected
2
3
with 2 x10 to 2 x 10 CFU of M. tuberculosis H37Rv by the intranasal route. Compound treatment was
initiated three weeks after infection. Compound 49 or the reference drug isoniazid (INH) was administered by
oral gavage for 28 days, five times per week. Bacterial load in the lungs of infected mice was determined by
CFU enumeration as shown in Table 6. Both compounds displayed potential efficacy results in a dose-
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 18 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
dependent manner. The compound 49 was active and promoted a significant reduction of the bacterial burden in
the lungs of infected animals. The reduction in CFU was proportional to the administered dose, but was less
pronounced than for isoniazid despite better pharmacodynamic indices. The compound 49 (1.52 log CFU
1
0
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
8
reduction at 10 mg/kg) was also less efficacious than 50 (3.13 log CFU reduction at 10 mg/kg) , the drug
1
0
candidate that was selected for clinical evaluation.
Table 6. In vivo efficacy of compound 49 against M. tuberculosis in an established mouse model
Dose
(mg/kg)
Compd.
CFU (Log10)/lung
4
9
2
6.23 ± 0.30
5.72 ± 0.40
5.65 ± 0.40
5.01 ± 0.14
7.24 ± 0.17
1
5
0
0
INH
15
Untreated
CONCLUSIONS
In this study, we report on the optimization of the lead compound 1 that led to 50 including SAR studies to
improve an activity against M. tuberculosis H37Rv replicating inside and outside macrophage assay and SPR
studies for development potential. We found that 3-carboxamide linker of this series played a critical role in
anti-tuberculosis activity. The issue of accumulation in plasma of bi-phenyl analogue 24 from in vivo PK study
was investigated by replacement of the last phenyl ring with heterocyclic groups. Further SAR investigations
with three ring systems, which had a linear and long hydrophobic groups revealed that the incorporation of a
piperidine or piperazine group in middle of two phenyl rings showed enhanced potency (intracellular MIC < 1
8
0
ACS Paragon Plus Environment

Page 19 of 55
Journal of Medicinal Chemistry
1
nM), improved metabolic stability (t1/2 > 60 min) and no inhibitory profile against 5 cytochrome P450 isozymes
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
(
IC > 40 µM). Ultimately, the optimization process led to compounds 49 and 50 that were performed in vivo
5
0
PK and efficacy in a mouse model and 50 was selected as a final candidate for further evaluation as a clinical
candidate based on its overall properties and high potency in the mouse model of tuberculosis. Preclinical study
and target engagement study of 50 will be reported in due course.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
EXPERIMENTAL SECTION
Chemistry. All reactions were carried out under an argon atmosphere in oven-dried glassware with magnetic
stirring and the reaction solvents were purified by passage through a bed of activated alumina. Purification of
reaction products was carried out by flash chromatography using silica gel 60 (Merck, 230-400 mesh).
Analytical thin layer chromatography was performed on 0.25 mm silica gel 60-F254 plates (Merck).
Visualization was accomplished with 254 nm of UV light and PMA or potassium permanganate staining
1
13
19
followed by heating. H-NMR (at 400 MHz), C-NMR (at 100 MHz) and F-NMR (at 376 MHz) spectra were
1 13
reported on a Varian 400 MHz spectrometer. H-NMR spectra (CDCl at 7.26 ppm) and C-NMR spectra
3
19
CDCl at 77.2 ppm) were recorded in ppm using solvent as an internal standard. F-NMR spectra were
(
3
recorded in ppm using α,α,α-trifluorotoluene as an external standard (at -64.72 ppm). Data are reported as (ap =
apparent, s = singlet, d= doublet, t = triplet, q = quartet, m = multiplet, br = broad; coupling constant(s) in Hz;
integration). LC/MS data were obtained using a Waters 2695 LC and Micromass ZQ spectrometer. The purity
of all biologically tested compounds was ≥95 % by HPLC. Yields refer to purified products and are not
optimized.
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 20 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Minimum inhibitory concentration determination, metabolic stability, CYP inhibition assay (Fluorescence
8
method), in vivo pharmacokinetics and in vivo efficacy were performed as previously descibed .
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Compound 50 has been tested using recombinant CYP enzymes to confirm its inhibitory activities.
Recombinant CYP inhibition assay:
2
ꢀL test compound/positive controls working solution are incubated with 100 ꢀL substrate and recombinant
CYP enzyme mixture working solution in the absence and presence of 98 ꢀL cofactor solutions for 10 minutes
for CYP1A2 and CYP2C9, 20 min for CYP2C19 and CYP2D6 and 3 min for CYP3A4. The final incubations
are terminated by 200 ꢀL cold IS-fortified (100 ng/mL tolbutamide) stop solution and the samples analyzed by
LC-MS/MS. The reaction mixtures (200 ꢀL final volume) contained approximately 100 mM potassium
phosphate buffer (pH 7.4), 3.3 mM MgCl , 1 mM NADPH and 5 pmol/mL recombinant CYP enzymes.
2
General Procedure for the Preparation of 2b-d. Method A: Ethyl 2-bromo-3-oxopentanoate (2b). To a
stirred solution of ethyl 3-oxopentanoate (13.8 mmol) in Et O (70 mL) were added ammonium acetate (41.4
2
mmol) and N-bromosuccinimide (13.8 mmol) and the reaction mixture was stirred at room temperature for 6
hours. The reaction mixture was diluted with Et O (20 mL) and washed with water (50 mL × 2). The organic
2
phase was dried over anhydrous MgSO and concentrated in vacuo to give a title a compound as a clear oil that
4
was used for next reaction without further purification.
ACS Paragon Plus Environment

Page 21 of 55
Journal of Medicinal Chemistry
1
Method B: Ethyl 2-bromo-3-oxohexanoate (2c) and ethyl 2-bromo-4-methyl-3-oxopentanoate (2d). To a
stirred solution of ethyl 3-oxohexanoate (6.3 mmol) in chloroform (5 mL) was added a solution of bromine (6.3
mmol) in chloroform (30 mL) dropwise under ice-bath and the resulting solution was allowed to room
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
temperature and stirred for 20 min. The reaction was quenched with saturated NaHCO (aq. 10 mL) and
3
extracted with chloroform. The resulting organic phase was washed with brine (20 mL), dried MgSO and
4
concentrated in vacuo to give a title compound 2c. The resulting crude residue was used for next reaction
without further purification.
In a similar manner, 2d was synthesized according to method B.
General Procedure for the Preparation of 3a-3g. To a solution of ethyl 2-bromo-3-oxopentanoate (2b, 12.9
mmol) in EtOH (25 mL) was added 2-amino-5-chloropyridine (12.9 mmol). The mixture was stirred at reflux
temperature for overnight. After cooling, the reaction mixture was concentrated. The resulting dark residue was
dissolved in EtOAc (20 mL) and washed with water (20 mL). The organic phase was washed with brine (20
mL), dried MgSO and concentrated in vacuo. The crude residue was purified by flash column chromatography
4
(n-hexane: EtOAc = 4:1) to give 3f as a pale yellow solid.
1
Ethyl 2-methylimidazo[1,2-a]pyridine-3-carboxylate (3a). H NMR (400 MHz, CDCl ); δ 1.29 (t, J = 7.2 Hz,
3
3
H), 2.56 (s, 3H), 4.27 (q, J = 7.2 Hz, 2H), 6.77 – 6.81 (m, 1H), 7.17 – 7.22 (m, 1H), 7.42 – 7.45 (m, 1H), 9.11
–
9.13 (m, 1H).
1
Ethyl 2-ethylimidazo[1,2-a]pyridine-3-carboxylate (3b). H NMR (400 MHz, CDCl ); δ 1.36 (t, J = 7.6 Hz,
3
3H), 1.43 (t, J = 7.2 Hz, 3H), 3.12 (q, J = 7.6 Hz, 2H), 4.43 (q, J = 7.2 Hz, 2H), 6.95 – 6.98 (m, 1H), 7.35 – 7.39
(m, 1H), 7.63 – 7.65 (m, 1H), 9.31 – 9.33 (m, 1H).
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 22 of 55
1
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Ethyl 2-propylimidazo[1,2-a]pyridine-3-carboxylate (3c). H NMR (400 MHz, CDCl ); δ 1.02 (t, J = 7.6 Hz,
3
3
H), 1.44 (t, J = 7.2 Hz, 3H), 1.76 – 1.85 (m, 2H), 3.08 (t, J = 7.6 Hz, 2H), 4.43 (q, J = 7.2 Hz, 2H), 6.95 – 6.99
(
m, 1H), 7.35 – 7.39 (m, 1H), 7.62 – 7.64 (m, 1H), 9.32 – 9.34 (m, 1H).
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
Ethyl 2-isopropylimidazo[1,2-a]pyridine-3-carboxylate (3d). H NMR (400 MHz, CDCl ); δ 1.38 (d, J = 6.8
3
Hz, 6H), 1.44 (t, J = 7.2 Hz, 3H), 3.80 – 3.87 (m, 1H), 4.43 (t, J = 7.2 Hz, 2H), 6.94 – 6.97 (m, 1H), 7.34 – 7.38
(
m, 1H), 7.66 – 7.69 (m, 1H), 9.32 – 9.34 (m, 1H).
1
Ethyl 6-chloro-2-methylimidazo[1,2-a]pyridine-3-carboxylate (3e). H NMR (400 MHz, CDCl ); δ 1.43 (t, J
3
= 7.2 Hz, 3H), 2.69 (s, 3H), 4.42 (q, J = 7.2 Hz, 2H), 7.34 (dd, J = 9.2, 2.0 Hz, 1H), 7.54 (d, J = 9.2 Hz, 1H),
9.38 (d, J = 2.0 Hz, 1H).
1
Ethyl 6-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxylate (3f). H NMR (400 MHz, CDCl ); δ 1.35 (t, J =
3
7
.6 Hz, 3H), 1.44 (t, J = 7.2 Hz, 3H), 3.11 (q, J = 7.6 Hz, 2H), 4.44 (q, J = 7.2 Hz, 2H), 7.35 (dd, J = 9.6, 2.0 Hz,
1
H), 7.58 (d, J = 9.6 Hz, 1H), 9.42 (d, J = 2.0 Hz, 1H).
1
Ethyl 7-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxylate (3g). H NMR (400 MHz, CDCl ); δ 1.34 (t, J =
3
7
.6 Hz, 3H), 1.43 (t, J = 7.2 Hz, 3H), 3.09 (q, J = 7.6 Hz, 2H), 4.43 (q, J = 7.2 Hz, 2H), 6.95 (dd, J = 7.6, 2.0 Hz,
H), 7.62 (d, J = 2.0 Hz, 1H), 9.26 (d, J = 7.6 Hz, 1H).
1
General Procedure for the Preparation of 4a-4g. To a solution of 3f (4.3 mmol) in EtOH (30 mL) was
added an aqueous solution of lithium hydroxide (13.0 mmol in 10 mL of water) and the mixture was stirred at
room temperature for overnight. The organic solvent was evaporated and 1N HCl was added until pH was
reached to 4. The residual pale solid was collected by filtration, washed with water and dried to give 4f as a
white solid.
ACS Paragon Plus Environment

Page 23 of 55
Journal of Medicinal Chemistry
1
In similar manner, 4a-4e and 4g were synthesized.
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
General Procedure for the Preparation of 7a-7g. Method A (7a-7e): To a solution of 4-chlorobenzonitrile
(
3.63 mmol) in dimethoxyethane (9 mL) were added 4-chlorophenylboronic acid (4.36 mmol), 1,1’-
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
bis(diphenylphosphino)ferrocene)-dichloropalladium(II) (0.11 mmol), Na CO (7.26 mmol in 3 mL of water)
2
3
and the mixture was stirred at 150℃. After 1h, the mixture was cooled to room temperature, then the mixture
was extracted with EtOAc (20 mL), washed with sat. NaHCO (aq. 15 mL) and brine (15 mL) and dried over
3
MgSO and concentrated. The resulting residue was purified by flash column chromatography (n-
4
hexane:EtOAc = 10:1) to give 6b as a white solid (67% yield). To a solution of 6b (2.38 mmol) in THF (24 mL)
was added lithium aluminum hydride (7.14 mmol) under ice-bath and then the resulting mixture was refluxed
for an hour. The reaction mixture was cooled to room temperature, quenched with water, added sat. Na CO (aq.
2
3
1
5 mL) and extracted with EtOAc (30 mL × 2). The combined organic layers were washed with brine (20 mL),
dried over MgSO and concentrated in vacuo to give 7b as a pale yellow solid (89%). The resulting residue was
4
used for next reaction without further purification.
In a similar manner, 7a and 7c-7e were synthesized according to method A.
Method B (7f-7g): A solution of 4-bromobenzyl amine (0.32 mmol) in dimethoxyethane (1 mL) were added
(
4-(tert-butyl)phenyl)boronic acid (0.39 mmol), 1,1’-bis(diphenylphosphino)ferrocene)-dichloropalladium(II)
(
0.01 mmol), Na CO (0.64 mmol in 350 uL of water) and the mixture was stirred at 150℃. After 1h, the
2
3
mixture was cooled to room temperature, then the mixture was extracted with EtOAc (10 mL), washed with
saturated NaHCO (aq. 10 mL) and brine (10 mL), dried over MgSO and concentrated in vacuo to give crude
3
4
7f. The resulting residue was used for next reaction without further purification.
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 24 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
In a similar manner, 7g was synthesized according to method B.
General Procedure for the Preparation of 8a-8n. A mixture of 4-fluorobenzonitrile (3.0 mmol), 4-(4-
trifluoromethoxy)piperidine (3.3 mmol) and K CO (6.0 mmol) in DMSO (5 mL) was heated to 120℃ for 4
2
3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
hours. After the cooling, the mixture was poured to the water and then generating solid was filtered, washed
with water and dried. The resulting crude product (2.1 mmol) was dissolved in THF (10 mL) and then lithium
aluminum hydride (6.2 mmol) was added slowly. After the refluxing for an hour, the reaction mixture was
cooled to room temperature, quenched with water and filtered off the insoluble aggregates using of cellite. The
filtrate was basified with saturated Na CO (aq. 20 mL) and then extracted with EtOAc (20 mL × 2). The
2
3
combined organic layers were washed with brine (15 mL), dried over MgSO and concentrated in vacuo to give
4
8m as a pale yellow solid (64%, 2 steps).
In a similar manner, 8a-8l and 8n were synthesized according to procedure above.
General Procedure for amide coupling. To a stirred solution of 6-chloro-2-ethylimidazo[1,2-a]pyridine-3-
carboxylic acid (4f, 2.83 mmol) in anhydrous DMF (10 mL) was added 1-(3-dimethylaminopropyl)-3-
ethylcarbodiimide hydrochloride (3.84 mmol), 1-hydroxybenzotriazole (1.54 mmol), triethylamine (5.12 mmol)
and 4-(4-(4-(trifluoromethoxy)phenyl)piperidin-1-yl)phenyl)methanamine (8n, 2.56 mmol) at room temperature,
then the resulting solution was heated to 70℃ with stirring. After 2 hours, the reaction mixture was cooled to
room temperature and evaporated. Water (50 mL) was added into the crude residue, the resulting solid was
collected by filtration, the filter cake was washed with water (50 mL) and dried to afford crude product. The
resulting crude compound was purified by flash column chromatography (n-hexane:EtOAc:methylene chloride
= 1:1:1), then recrystallized from EtOAc to give title compound, 50 as a white solid.
ACS Paragon Plus Environment

Page 25 of 55
Journal of Medicinal Chemistry
1
6
-Chloro-2-ethyl-N-(4-(4-(4-(trifluoromethoxy)phenyl)piperidin-1-yl)benzyl)imidazo[1,2-a]pyridine-3-
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
1
carboxamide (50). mp = 164.0 ℃; H NMR (400 MHz, CDCl ); δ 1.37 (t, J = 7.6 Hz, 3H), 1.82 – 1.97 (m, 4H),
3
2
6
0
1
.64 – 2.70 (m, 1H), 2.80 – 2.87 (m, 2H), 2.93 (q, J = 7.6 Hz, 2H), 3.80 – 3.83 (m, 2H), 4.61 (d, J = 5.2 Hz, 2H),
.00 (brt, J = 5.2 Hz, 1H), 6.96 – 6.99 (m, 2H), 7.15 (d, J = 8.0 Hz, 2H), 7.24 – 7.30 (m, 5H), 7.52 (dd, J = 9.6,
.8 Hz, 1H), 9.53 (dd, J = 2.0, 0.8 Hz, 1H); 13C NMR (100 MHz, CDCl3) δ 13.3, 23.6, 33.4, 42.0, 43.3, 50.4,
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
15.4, 117.0, 121.2, 121.6, 121.9, 126.3, 128.2, 128.3, 128.7, 128.9, 144.5, 144.7, 147.7, 151.4, 151.5, 161.2;
1
9
+
F NMR (376 MHz, CDCl ) δ 58.31 (s, 3F); LCMS (ESI) m/z 557 [M + H] ; HRESIMS calcd for
3
+
C H ClF N O [M + H] 557.1926, found 557.1918.
29
29
3
4
2
1
N-([1,1'-Biphenyl]-4-ylmethyl)-6-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (1). H NMR (400
MHz, CDCl ) δ 1.41 (t, J = 7.6 Hz, 3H), 2.98 (q, J = 7.6 Hz, 2H), 4.74 (d, J = 5.6 Hz, 2H), 6.15 (brs, 1H), 7.29
3
(
dd, J = 9.6, 2.0 Hz, 1H), 7.35 - 7.37 (m, 1H), 7.43 – 7.47 (m, 4H), 7.55 (d, J = 9.2 Hz, 1H), 7.58 – 7.62 (m, 4H),
+
.56 (d, J = 2.0 Hz, 1H); LCMS (ESI) m/z 390 [M + H] .
9
1
N-([1,1'-Biphenyl]-4-ylmethyl)-2-methylimidazo[1,2-a]pyridine-3-carboxamide (18). H NMR (400 MHz,
DMSO-d ) δ 2.62 (s, 3H), 4.57 (d, J = 6.0 Hz, 2H), 7.00 (dd, J = 6.8, 6.8 Hz, 1H), 7.33 – 7.40 (m, 2H), 7.45 –
6
7
.48 (m, 4H), 7.56 (d, J = 8.8 Hz, 1H), 7.64 – 7.66 (m, 4H), 8.38 (brt, J = 6.0 Hz, 1H), 9.04 (d, J = 6.8 Hz, 1H);
+
MS (ESI) m/z 342 [M + H] .
1
N-([1,1'-Biphenyl]-4-ylmethyl)-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (19). H NMR (400 MHz,
CDCl ) δ 1.42 (t, J = 7.6 Hz, 3H), 3.02 (q, J = 7.6 Hz, 2H), 4.75 (d, J = 5.6 Hz, 2H), 6.19 (brs, 1H), 6.92 (dd, J
3
= 6.4, 6.4 Hz, 1H), 7.32 (d, J = 7.6 Hz, 1H), 7.35 (d, J = 7.2 Hz, 1H), 7.45 (d, J = 8.0 Hz, 4H), 7.58 – 7.59 (m,
+
5H), 9.41 (d, J = 6.8 Hz, 1H); LCMS (ESI) m/z 356 [M + H] .
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 26 of 55
1
N-([1,1'-Biphenyl]-4-ylmethyl)-2-propylimidazo[1,2-a]pyridine-3-carboxamide (20). H NMR (400 MHz,
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
CDCl ) δ 0.98 (t, J = 7.4 Hz, 3H), 1.80 – 1.89 (m, 2H), 2.93 (t, J = 7.8 Hz, 2H), 4.73 (d, J = 5.6 Hz, 2H), 6.29 (t,
3
J = 5.6 Hz, 1H), 6.89 (dd, J = 6.8, 1.2 Hz, 1H), 7.27 – 7.37 (m, 2H), 7.42 – 7.46 (m, 4H), 7.56 – 7.61 (m, 5H),
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
.35 (d, J = 6.8 Hz, 1H). ; LCMS (ESI) m/z 370 [M + H] .
9
1
N-([1,1'-Biphenyl]-4-ylmethyl)-2-isopropylimidazo[1,2-a]pyridine-3-carboxamide (21). H NMR (400 MHz,
CDCl ) δ 1.44 (d, J = 6.4 Hz, 6H), 3.34 – 3.41 (m, 1H), 4.76 (d, J = 5.6 Hz, 2H), 6.16 (m, 1H), 6.90 (dd, J = 7.2,
3
7
.2 Hz, 1H), 7.29 – 7.37 (m, 2H), 7.42 – 7.47 (m, 4H), 7.60 – 7.64 (m, 5H), 9.32 (d, J = 7.2 Hz, 1H); LCMS
+
(ESI) m/z 370 [M + H] .
1
N-([1,1'-Biphenyl]-4-ylmethyl)-6-chloro-2-methylimidazo[1,2-a]pyridine-3-carboxamide (22). H NMR (400
MHz, DMSO-d ) δ 2.61 (s, 3H), 4.56 (d, J = 5.6 Hz, 2H), 7.32 – 7.34 (m, 1H), 7.41 – 7.46 (m, 5H), 7.62 – 7.64
6
+
m, 5H), 8.47 (brt, J = 5.6 Hz, 1H), 9.13 – 9.14 (m, 1H); LCMS (ESI) m/z 376 [M+H] .
(
1
-Chloro-N-((2'-chloro-[1,1'-biphenyl]-4-yl)methyl)-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (23). H
6
NMR (400 MHz, CDCl ) δ 1.44 (t, J = 7.6 Hz, 3H), 3.02 (q, J = 7.6 Hz, 2H), 4.77 (d, J = 6.0 Hz, 2H), 6.18 (m,
3
1
H), 7.27 – 7.35 (m, 4H), 7.43 – 7.48 (m, 5H), 7.56 (d, J = 9.6 Hz, 1H), 9.56 (d, J = 2.0 Hz, 1H); LCMS (ESI)
+
m/z 424 [M + H] .
1
-Chloro-N-((4'-chloro-[1,1'-biphenyl]-4-yl)methyl)-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (24). H
6
NMR (400 MHz, CDCl ) δ 1.43 (t, J = 7.6 Hz, 3H), 3.01 (q, J = 7.6 Hz, 2H), 4.71 (d, J = 6.0 Hz, 2H), 6.13 (m,
3
1
H), 7.31 (dd, J = 9.6, 2.0 Hz, 1H), 7.41 (d, J = 8.8 Hz, 2H), 7.45 (d, J = 8.0 Hz, 2H), 7.51 (d, J = 8.4 Hz, 2H),
+
7.54 – 7.58 (m, 3H), 9.55 (d, J = 2.0 Hz, 1H); LCMS (ESI) m/z 424 [M+H] ; HRESIMS calcd for
+
C H Cl N O [M + H] 424.0978, found 424.0989.
23
20
2
3
ACS Paragon Plus Environment

Page 27 of 55
Journal of Medicinal Chemistry
1
1
6
-Chloro-N-((4'-cyano-[1,1'-biphenyl]-4-yl)methyl)-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (25).
H
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
NMR (400 MHz, CDCl ) δ 1.42 (t, J = 7.6 Hz, 3H), 2.992 (q, J = 7.2 Hz, 2H), 4.75 (d, J = 5.6 Hz, 2H), 6.18
3
(brt, J = 5.6 Hz, 1H), 7.30 (dd, J = 9.6, 2.0 Hz, 1H), 7.49 (d, J = 8.0 Hz, 2H), 7.56 (d, J = 9.6 Hz, 1H), 7.59 (d, J
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
=
8.0 Hz, 2H), 7.67 (d, J = 8.4 Hz, 2H), 7.72 (d, J = 8.4 Hz, 2H), 9.55 (d, J = 2.0 Hz, 1H); LCMS (ESI) m/z 415
+
M + H] .
[
1
-Chloro-2-ethyl-N-((4'-methyl-[1,1'-biphenyl]-4-yl)methyl)imidazo[1,2-a]pyridine-3-carboxamide (26). H
6
NMR (400 MHz, CDCl ) δ 1.42 (t, J = 7.6 Hz, 3H), 3.00 (q, J = 7.6 Hz, 2H), 2.40 (s, 3H), 4.74 (d, J = 5.6 Hz,
3
2H), 6.16 (m, 1H), 7.25 (d, J = 7.2 Hz, 2H), 7.30 (dd, J = 9.6, 2.0 Hz, 1H), 7.44 (d, J = 8.0 Hz, 2H), 7.49 (d, J =
8.0 Hz, 2H), 7.54 (d, J = 9.6 Hz, 1H), 7.59 (d, J = 8.4 Hz, 2H), 9.55 (d, J = 2.0 Hz, 1H); LCMS (ESI) m/z 404
+
[M + H] .
1
-Chloro-2-ethyl-N-((4'-methyl-[1,1'-biphenyl]-4-yl)methyl)imidazo[1,2-a]pyridine-3-carboxamide (27). H
7
NMR (400 MHz, CDCl ) δ 1.41 (t, J = 7.6 Hz, 3H), 2.40 (s, 3H), 2.99 (q, J = 7.6 Hz, 2H), 4.73 (s, 2H), 6.91 (dd,
3
J = 7.6, 2.0 Hz, 1H), 7.25 (d, J = 8.4 Hz, 2H), 7.43 (d, J = 8.0 Hz, 2H), 7.48 (d, J = 8.0 Hz, 2H), 7.58 – 7.60 (m,
+
H), 9.38 (d, J = 7.2 Hz, 1H) ; LCMS (ESI) m/z 404 [M + H] .
3
1
-Chloro-N-((4'-chloro-[1,1'-biphenyl]-4-yl)methyl)-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (28). H
7
NMR (400 MHz, CDCl ) δ 1.42 (t, J = 7.6 Hz, 3H), 2.99 (q, J = 7.6 Hz, 2H), 4.73 (s, 2H), 6.15 (brs, 1H), 6.91
3
(dd, J = 7.6, 2.0 Hz, 1H), 7.40 (d, J = 8.4 Hz, 2H), 7.44 (d, J = 8.0 Hz, 2H), 7.51 (d, J = 8.4 Hz, 2H), 7.56 (d, J
+
8.0 Hz, 2H), 7.60 (d, J = 1.6 Hz, 1H), 9.38 (d, J = 7.2 Hz, 1H) ; LCMS (ESI) m/z 424 [M + H] ; HRESIMS
=
+
calcd for C H Cl N O [M + H] 424.0978, found 424.0983.
2
3
20
2
3
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 28 of 55
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
4
'-((7-Chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxamido)methyl)-[1,1'-biphenyl]-4-carboxylic acid (29).
1
H NMR (400 MHz, DMSO-d ) δ 1.26 (t, J = 7.6 Hz, 3H), 2.98 (q, J = 7.6 Hz, 2H), 4.58 (d, J = 6.0 Hz, 2H),
6
7
.08 (dd, J = 7.2, 2.4 Hz, 1H), 7.48 (d, J = 8.4 Hz, 2H), 7.72 (d, J = 8.0 Hz, 2H), 7.78 - 7.80 (m, 3H), 8.00 (d, J
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
8.4 Hz, 2H), 8.51 (brt, J = 6.0 Hz, 1H), 8.97 (d, J = 7.2 Hz, 1H); LCMS (ESI) m/z 434 [M + H] .
=
N-((4'-(tert-Butyl)-[1,1'-biphenyl]-4-yl)methyl)-7-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (30).
1
H NMR (400 MHz, CDCl ) δ 1.36 (s, 9H), 1.41 (t, J = 7.6 Hz, 3H), 2.99 (q, J = 7.6 Hz, 2H), 4.73 (d, J = 5.6
3
Hz, 2H), 6.13 (brs, 1H), 6.91 (dd, J = 7.2, 2.0 Hz, 1H), 7.43 (d, J = 8.0 Hz, 2H), 7.47 (d, J = 8.4 Hz, 2H), 7.53
+
(d, J = 8.4 Hz, 2H), 7.59 – 7.61 (m, 3H), 9.38 (d, J = 7.2 Hz, 1H); LCMS (ESI) m/z 446 [M + H] .
1
N-(4-(1H-Pyrrol-2-yl)benzyl)-7-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (31). H NMR (400
MHz, CDCl ) δ 1.37 (t, J = 7.6 Hz, 3H), 2.94 (q, J = 7.6 Hz, 2H), 4.67 (d, J = 6.0 Hz, 2H), 6.10 (brs, 1H), 6.29
3
–
2
6.32 (m, 1H), 6.52 – 6.54 (m, 1H), 6.86 – 6.88 (m, 1H), 6.89 (dd, J = 7.2, 2.0 Hz, 1H), 7.35 (d, J = 8.0 Hz,
H), 7.47 (d, J = 8.4 Hz, 2H), 7.58 (d, J = 2.0 Hz, 1H), 8.51 (brs, 1H), 9.35 (d, J = 7.2 Hz, 1H); LCMS (ESI)
+
m/z 379 [M + H] .
1
-Chloro-2-ethyl-N-(4-(pyridin-4-yl)benzyl)imidazo[1,2-a]pyridine-3-carboxamide (32). H NMR (400 MHz,
7
CDCl ) δ 1.42 (t, J = 7.6 Hz, 3H), 3.01 (q, J = 7.6 Hz, 2H), 4.76 (d, J = 5.6 Hz, 2H), 6.20 (brs, 1H), 6.91 (dd, J
3
=
7.6, 2.0 Hz, 1H), 7.26 – 7.51 (m, 4H), 7.61 (d, J = 2.0 Hz, 1H), 7.65 (d, J = 7.6 Hz, 2H), 8.65 (brs, 2H), 9.37
+
d, J = 7.6 Hz, 1H); LCMS (ESI) m/z 391 [M + H] .
(
1
-Chloro-2-ethyl-N-(4-(piperidin-1-yl)benzyl)imidazo[1,2-a]pyridine-3-carboxamide (33). H NMR (400
7
MHz, CDCl ) δ 1.35 (t, J = 7.6 Hz, 3H), 1.55 – 1.57 (m, 2H), 1.66 – 1.70 (m, 4H), 2.91 (q, J = 7.6 Hz, 2H),
3
3.12 – 3.15 (m, 4H), 4.56 (d, J = 5.6 Hz, 2H), 6.07 (brs, 1H), 6.86 (dd, J = 7.6, 2.0 Hz, 1H), 6.90 (d, J = 8.4 Hz,
ACS Paragon Plus Environment

Page 29 of 55
Journal of Medicinal Chemistry
1
2
H), 7.22 (d, J = 8.4 Hz, 2H), 7.54 (d, J = 2.0 Hz, 1H), 9.30 (d, J = 7.6 Hz, 1H) ; LCMS (ESI) m/z 397 [M +
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
H] .
1
N-(4-(Azepan-1-yl)benzyl)-7-chloro-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (34). H NMR (400 MHz,
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
CDCl ) δ 1.38 (t, J = 7.6 Hz, 3H), 1.52-1.55 (m, 4H), 1.78 (m, 4H), 2.94 (q, J = 7.6 Hz, 2H), 3.45 (t, J = 6.0 Hz,
3
4
H), 4.56 (d, J = 5.2 Hz, 2H), 5.95 (brs, 1H), 6.67 (d, J = 8.8 Hz, 2H), 6.89 (dd, J = 7.6, 2.4 Hz, 1H), 7.20 (d, J
+
8.8 Hz, 2H), 7.57 (d, J = 2.4 Hz, 1H), 9.36 (d, J = 7.6 Hz, 1H); LCMS (ESI) m/z 411 [M+H] .
=
7
-Chloro-2-ethyl-N-(4-(4-(trifluoromethyl)piperidin-1-yl)benzyl)imidazo[1,2-a]pyridine-3-carboxamide (35).
1
mp = 209.4 ℃; H NMR (400 MHz, CDCl ); δ 1.34 (t, J = 7.6 Hz, 3H), 1.68 – 1.78 (m, 2H), 1.94 – 1.98 (m,
3
2H), 2.11 – 2.20 (m, 1H), 2.66 – 2.73 (m, 2H), 2.90 (q, J = 7.6 Hz, 2H), 3.73 – 3.77 (m, 2H), 4.58 (d, J = 5.2 Hz,
2H), 6.03 (brt, J = 5.2 Hz, 1H), 6.86 (dd, J = 7.6, 2.4 Hz, 1H), 6.91 (d, J = 8.8 Hz, 2H), 7.25 (d, J = 8.8 Hz, 2H),
+
.56 (d, J = 2.4 Hz, 1H), 9.32 (d, J = 7.6 Hz, 1H); LCMS (ESI) m/z 465 [M + H] .
7
1
-Chloro-N-(4-(4-chloropiperidin-1-yl)benzyl)-2-ethylimidazo[1,2-a]pyridine-3-carboxamide (36). H NMR
7
(400 MHz, CDCl ); δ 1.36 (t, J = 7.2 Hz, 3H), 1.99 – 2.04 (m, 2H), 2.17 – 2.20 (m, 2H), 2.92 (q, J = 7.2 Hz,
3
2
H), 3.05 – 3.10 (m, 2H), 3.50 – 3.52 (m, 2H), 4.20 – 4.23 (m, 1H), 4.59 (d, J = 5.6 Hz, 2H), 5.99 (brs, 1H),
+
.89 – 6.94 (m, 3H), 7.26 – 7.27 (m, 2H), 7.58 (s, 1H), 9.35 (d, J = 6.8 Hz, 1H); LCMS (ESI) m/z 431 [M + H] .
6
1
7
-Chloro-2-ethyl-N-(4-(4-methylpiperazin-1-yl)benzyl)imidazo[1,2-a]pyridine-3-carboxamide (37).
H
NMR (400 MHz, CDCl ) δ 1.37 (t, J = 7.6 Hz, 3H), 2.35 (s, 3H), 2.57 – 2.59 (m, 4H), 2.94 (q, J = 7.6 Hz, 2H),
3
3
.20 – 3.23 (m, 4H), 4.59 (d, J = 5.2 Hz, 2H), 6.00 (brs, 1H), 6.88 – 6.94 (m, 3H), 7.27 (d, J = 8.4 Hz, 2H), 7.58
+
(d, J = 2.0 Hz, 1H), 9.35 (d, J = 7.6 Hz, 1H) ; LCMS (ESI) m/z 412 [M + H] .
ACS Paragon Plus Environment