Design, synthesis, and docking studies of phenylpicolinamide derivatives bearing 1H-pyrrolo[2,3-b]pyridine moiety as c-Met inhibitors

Article abstract of DOI:10.1016/j.bmc.2016.01.001

Four series of phenylpicolinamide derivatives bearing 1H-pyrrolo[2,3-b]pyridine moiety (12a-e, 13a-f, 14a-f and 15a-i) were designed, synthesized and evaluated for the IC₅₀ values against three cancer cell lines (A549, PC-3 and MCF-7) and c-Met kinase. Five selected compounds (13b, 15b, 15d, 15e and 15f) were further evaluated for the activity against HepG2 and Hela cell lines. Eighteen of the compounds showed excellent cytotoxicity activity and selectivity with the IC₅₀ valuables in single-digit μM to nanomole range. Seven of them are equal to more active than positive control Foretinib against one or more cell lines. The most promising compound 15f showed superior activity to Foretinib, with the IC₅₀ values of 1.04 ± 0.11 μM, 0.02 ± 0.01 μM and 9.11 ± 0.55 μM against A549, PC-3 and MCF-7 cell lines, which were 0.62 to 19.5 times more active than Foretinib (IC₅₀ values: 0.64 ± 0.26 μM, 0.39 ± 0.11 μM, 9.47 ± 0.22 μM), respectively. Structure-activity relationships (SARs) and docking studies indicated that replacement of quinoline nucleus of the previous active compounds with 1H-pyrrolo[2,3-b]pyridine moiety maintained even improved the potent cytotoxic activity. The results suggested that the introduction of fluoro atoms to the aminophenoxy part of target compounds or the phenyl group of pyrimidine substituted on C-4 position was benefit for the activity.

Full text of DOI:10.1016/j.bmc.2016.01.001

Accepted Manuscript
Design, synthesis, and docking studies of phenylpicolinamide derivatives bear-
ing 1H-pyrrolo[2,3-b]pyridine moiety as c-Met inhibitors
Wufu zhu, Wenhui Wang, Shan Xu, Qidong Tang, Rong Luo, Min Wang, Ping
Gong, Pengwu Zheng
PII:
S0968-0896(16)30001-3
DOI:
http://dx.doi.org/10.1016/j.bmc.2016.01.001
BMC 12744
Reference:
To appear in:
Bioorganic & Medicinal Chemistry
Received Date:
Revised Date:
Accepted Date:
5 November 2015
31 December 2015
2 January 2016
Please cite this article as: zhu, W., Wang, W., Xu, S., Tang, Q., Luo, R., Wang, M., Gong, P., Zheng, P., Design,
synthesis, and docking studies of phenylpicolinamide derivatives bearing 1H-pyrrolo[2,3-b]pyridine moiety as c-
Met inhibitors, Bioorganic & Medicinal Chemistry (2016), doi: http://dx.doi.org/10.1016/j.bmc.2016.01.001
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Design, synthesis, and docking studies of phenylpicolinamide derivatives bearing 1H-pyrrolo[2,3-b]pyridine
moiety as c-Met inhibitors
1
,† ,*
1,†
1
1
2
1
3
1
Wufu zhu , Wenhui Wang , Shan Xu ,Qidong Tang , Rong Luo ,Min Wang , Ping Gong , Pengwu Zheng
1
School of Pharmacy, Jiangxi Science & Technology Normal University, Nanchang 330013, P. R. China.
2
Jiangxi Province Institute of Materia Medica, Nanchang 330000, China;
3
Key Laboratory of Original New Drugs Design and Discovery of Ministry of Education, Shenyang Pharmaceutical
University, 103 Wenhua Road, Shenhe District, Shenyang 110016, P. R. China.
*Correspondence author.
Tel/Fax: +86 791 8380-2393;
E-mail address: zhuwf@jxstnu.edu.cn, zhuwufu-1122@163.com (W. Zhu), zhengpw@126.com (P. Zheng).
Four series of phenylpicolinamide derivatives bearing 1H-pyrrolo[2,3-b]pyridine moiety (12a–e, 13a–f, 14a–f and
1
5a–i) were designed, synthesized and evaluated for the IC50 values against three cancer cell lines (A549, PC-3 and
MCF-7) and c-Met kinase. Five selected compounds (13b, 15b, 15d, 15e and 15f) were further evaluated for the
activity against HepG2 and Hela cell lines. Eighteen of the compounds showed excellent cytotoxicity activity and
selectivity with the IC50 valuables in single-digit μM to nanomole range. Seven of them are equal to more active than
positive control Foretinib against one or more cell lines. The most promising compound 15f showed superior activity to
Foretinib, with the IC50 values of 1.04 ± 0.11 μM, 0.02 ± 0.21μM and 9.11 ± 0.55 μM against A549, PC-3 and MCF-7
cell lines, which were 0.62 to 19.5 times more active than Foretinib (IC50 values: 0.64 ± 0.26 μM, 0.39 ± 0.11 μM, 9.47
±
0.22 μM) , respectively. Structure–activity relationships (SARs) and docking studies indicated that replacement of
quinoline nucleus of the previous active compounds with 1H-pyrrolo[2,3-b]pyridine moiety maintained even improved
the potent cytotoxic activity. The results suggested that the introduction of fluoro atoms to the aminophenoxy part of
target compounds or the phenyl group of pyrimidine substituted on C-4 position was benefit for the activity.
Key words: Phenylpicolinamide; 1H-pyrrolo[2,3-b]pyridine; Synthesis; Docking; c-Met inhibitors; Antitumor activity
†
These authors contribute equally to this work

Corresponding author. Tel./fax: +86 791 83802393.
E-mail address:zhuwf@jxstnu.edu.cn, zhuwufu-1122@163.com (W. Zhu), zhengpw@126.com (P. Zheng).
1

1
Introduction
Cancer is a serious disease that threatens human health and life. There were 14.1 million new cancer cases, 8.2
[1]
million deaths and 32.6 million people living with cancer (within 5 years of diagnosis) in 2012 worldwide
.
c-Met inhibitors are a class of small molecules that inhibit the enzymatic activity of the c-Met tyrosine kinase. These
[2]
inhibitors may have therapeutic application in the treatment of various types of cancers. Cabozantinib , a small
molecule that inhibits the activity of multiple tyrosine kinases, including RET, MET, and VEGF receptor 2, was
approved by U. S. Food and Drug Administration for the treatment of patients with progressive metastatic medullary
[3]
thyroid cancer on November 29, 2012 . In recent years, many derivatives of Cabozantinib were reported, such as
[4]
[5-6]
foretinib (GSK1363089) , compounds I, II, III . The studies revealed that these compounds elicited strong
antitumor activity.
In our previous study, we reported a series of quinoline derivatives bearing phenylpicolinamide scaffold as potential
[7]
c-Met inhibitors . Most of them showed excellent in vitro antitumor activity and the structure-activity relationships
（
SARs） showed that the phenylpicolinamide scaffold is benefit to the activity. In order to investigate the influence of
the quinoline nucleus to the activity of this series of compounds, further modifications were carried out in this paper.
Firstly, keep the phenylpicolinamide scaffold unchanged, we replaced the quinoline nucleus with
1H-pyrrolo[2,3-b]pyridine moiety to get compounds 12a–e. However, these compounds showed no activity. Next, we
changed the position of the phenyl group on pyrimidine from C-4 to C-5 position to afford 13a–f. These compounds
showed excellent cytotoxicity and moderate c-Met kinase activity. What’s more, fluoro atom was added to the
aminophenoxy part of compounds 12a–e and 13a–f to study the effect to the activity and yielded 14a–f and 15a–i. To
our exciting, compounds 15a–i exhibited excellent cytotoxicity and c-Met kinase activity.
Herein we disclosed the synthesis and antitumor activity against A549, PC-3, MCF-7,HepG2 and Hela cancer cell
lines, and c-Met kinase of phenylpicolinamide derivatives bearing 1H-pyrrolo[2,3-b]pyridine moiety. Moreover,
docking studies were presented in this paper as well.
2

Figure 1 Structures of small-molecule c-Met inhibitors and target compounds.
2
. Chemistry
The preparation of target compounds 12a–e, 13a–f, 14a–f and 15a–i was described in Scheme 1.
The key intermediates 4-(4-aminophenoxy)-N-methylpicolinamide 11a,b
were achieved from
4-chloro-1H-pyrrolo[2,3-b]pyridine 8 via substitution reaction with 4-nitrophenol or 2-fluoro-4-nitrophenol 9a,b and
reduction with hydrazine hydrate in sequence as shown in Scheme 1.
-Bromo-pyridine-carboxylic acid 1 or 5-bromo-pyridine-carboxylic acid 2 was reacted with substituted
4
phenylboronic acids 3a–k through Suzuki-coupling reaction with bis(triphenylphosphine)palladium(II) dichloride as
catalyst to get 4a–k and 5a–k, which were then chlorination with oxalyl chloride to obtain 6a–k and 7a–k, respectively.
Finally, reaction of amides 11a,b with acyl chlorides 6a–k or 7a–k promoted by DIPEA in dichloromethane at room
temperature yielded target compounds 12a–e, 13a–f, 14a–f and 15a–i, respectively.
3

Scheme 1 Synthetic route of target compounds. Reagents and conditions: (a) [(C
,4-dioxane, 5-8h; (b) (COCl) , DMF(cat.), CH Cl , 0.5h; (c) Phenyl ether, 190℃, 1 h; (d) N
carbon, EtOH, 10 min; (e) DIPEA, CH Cl , 0.5h.
6
H
5
)
3
P]
2
PdCl
2
, Na
2
CO
3
, H
2
O,
.
1
2
2
2
2
H
4
H
2
O, FeCl
3
, activated
2
2
3
. Results and discussion
3
.1 Biological evaluation
Taking c-Met inhibitor Foretinib as reference compound, the target compounds (12a–e, 13a–f, 14a–f and 15a–i) were
evaluated for the cytotoxicity against three cancer cell lines A549 (human lung cancer), PC-3 (human prostatic cancer)
and MCF-7(human breast cancer) were evaluated by 3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide
(MTT) cell proliferation assay. In addition, these compounds were evaluated for the IC50 values against c-Met kinase in
vitro by the Mobility shift assay with ATP concentration at Km, together with reference compound Foretinib. Moreover,
five selected compounds (13b, 15b, 15d, 15e and 15f) were further evaluated for the activity against HepG2 and Hela
cell lines. The results expressed as inhibition rates or IC50 values were summarized in Tables 1–5 and the values are the
average of at least two independent experiments.
Table 1 Structures, cytotoxicity and c-Met kinase inhibitory activity of compounds 12a-e
4

a
IC50 (μM)
PC-3
Compd. N.O.
R
A549
MCF-7
>10
c-Met
12a
2b
4-Cl
>10
>10
>10
>10
>10
7.3
1
4-CH
3
>10
1
2c
2d
2e
Foretinib
4-OCH
3
>10
>10
>10
8.2
1
2,4difluo
>10
>10
>10
>10
>10
0.014
1
4-CH
2
CH
3
>10
>10
>10
b
-
0.64 ± 0.26
0.39 ± 0.11
9.47 ± 0.22
a
The values are an average of two separate determinations.
Used as a positive control
b
Table 2 Structures, cytotoxicity and c-Met kinase inhibitory activity of compounds 13a-f
a
IC50 (μM)
Compd. N.O.
R
A549
PC-3
MCF-7
>10
c-Met
1
3a
3b
3c
3d
3e
3f
Foretinib
4-CH
4-OCH
4-CF
4-Cl
3
>10
>10
2.3
1.6
>10
4.6
1
3
6.97 ± 1.01
3.95 ± 1.11
4.31 ± 1.03
7.01 ± 1.10
2.92 ± 1.17
0.64 ± 0.26
9.62±1.01
14.22±0.99
25.11±0.41
5.47±0.78
3.27±1.19
0.39 ±0.11
8.21 ± 0.45
>10
1
3
1
>10
c
1
3-CH
3
7.47 ± 0.21
3.75 ± 0.45
9.47 ± 0.22
ND
ND
1
H
-
b
0.014
a
b
c
The values are an average of two separate determinations.
Used as a positive control
ND: Not determined
5

Table 3 Structures, cytotoxicity and c-Met kinase inhibitory activity of compounds 14a-f
a
IC50 (μM)
Compd. No
R
H
A549
PC-3
MCF-7
c-Met
1
4a
4b
4c
4d
4e
4f
Foretinib
1.25 ± 0.13
2.30 ± 0.14
2.24 ± 0.14
6.01 ± 1.14
10.00 ± 1.85
3.10 ± 0.169
0.64 ± 0.26
0.76 ± 0.12
2.64 ± 1.00
0.65 ± 0.18
13.36 ± 0.75
3.03 ± 1.27
4.72 ± 1.51
0.39 ± 0.11
4.39 ± 1.08
8.02 ± 1.05
10.09 ± 1.02
>10
>10
6.6
1
4-CH
3
1
4-OCH
3
>10
>10
8.9
1
4-F
1
4-Cl
2,4-di F
-
>10
1
>10
5.9
b
9.47 ± 0.22
0.014
a
The values are an average of two separate determinations.
Used as a positive control
b
Table 4 Structures, cytotoxicity and c-Met kinase inhibitory activity of compounds 15a-i
a
IC50 (μM)
Compd. No.
R
H
A549
PC-3
MCF-7
>10
c-Met
>10
1
5a
5b
5c
5d
5e
5f
4.95 ± 1.08
1.75 ± 0.12
3.12 ± 1.35
4.35 ± 0.15
3.37 ± 0.15
1.04 ± 0.11
>10
1
3-CH
4-CH
2,4-di CH
4-CH CH
4-OCH
3
0.86 ± 0.17
3.97 ± 0.94
4.01 ± 1.36
2.96 ± 1.48
0.02 ± 0.21
9.12 ± 0.56
8.43 ± 0.99
6.22 ± 1.41
7.47 ± 0.51
9.11 ± 0.55
0.61
5.23
0.89
1.21
0.69
1
3
1
3
1
2
3
1
3
6

1
5g
5h
5i
Foretinib
4-F
1.09 ± 0.15
4.92 ± 1.41
>10
0.71 ± 0.14
4.35 ± 1.16
>10
9.52 ± 0.21
8.55 ± 0.44
>10
2.2
>10
1
4-CF
3
1
2,4-di F
-
>10
b
0.64 ± 0.26
0.39 ± 0.11
9.47 ± 0.22
0.014
a
The values are an average of two separate determinations.
Used as a positive control
b
Table 5 Cytotoxicity of selected compounds 13b, 15b, 15d-f.
a
IC50 (μM)
Compd. No
HepG2
Hela
1
1
1
3b
5b
5d
11.65±1.06
11.16 ±0.95
13.32 ±1.52
1038±0.97
7.53±0.95
10.32±1.14
11.89±1.75
8.63±1.12
1
5e
15f
13.26±1.32
5.89±1.02
a
The values are an average of two separate determinations.
As shown in Tables 1–5, eighteen of the compounds exhibited excellent cytotoxicity activity against different cancer
cells with potency from the single-digit μM to single-digit nanomole range. In general, the third and the fourth series of
compounds (14a–f and 15a–i) which were substituted by fluoro atom on aminophenoxy part are more active than that
of the first and the second series (12a–e and 13a–f). What’s more, compounds 12a–e, the phenyl group on pyrimidine
of which was C-5 position showed bad activity. However, when fluoro atoms were added to the aminophenoxy part of
these compounds or the phenyl group on pyrimidine was changed to C-5 position, the target compounds 14a–f and
1
3a–f were more active than compounds 12a–e. Similarly, compounds 15a–i were more potent than compounds 14a–f
and 13a–f. The results suggested that the introduction of fluoro atoms to the aminophenoxy part of target compounds or
the phenyl group of pyrimidine substituted on C-4 position is benefit for the activity. That’s why compounds 15a–i
showed the best activity among these four series compounds.
The cytotoxicity activity against A549 and PC-3 cancer cell lines was well while it’s moderate or bad against MCF-7,
HepG2 and Hela cell lines. It’s indicated that the target compounds showed excellent selectivity toward A549 and PC-3
to MCF-7, HepG2 and Hela. Among the eighteen active compounds, seven of them (compounds 14a, 14c, 15b, 15d,
1
5f-g and 15i) were equal to more potent than Foretinib against one or more cell lines. Especially, compound 15f
showed superior activity to Foretinib, with the IC50 values of 1.04 ± 0.11 μM, 0.02 ± μM and 9.11 ± 0.55 μM against
A549, PC-3 and MCF-7 cell lines, which were 0.62 to 19.5 times more active than Foretinib (IC50 values: 0.64 ± 0.26
7

μM, 0.39 ± 0.11 μM, 9.47 ± 0.22 μM) , respectively. The results suggested that replacement of quinoline nucleus with
1H-pyrrolo[2,3-b]pyridine moiety maintained even improved the potent cytotoxic activity.
Furthermore, different substitutions of aryl group affected the cytotoxicity of target compounds. For the second
series (13a-f), substituted on C-4 position is not beneficial to the activity (13a-d) and substituted on C-3 position or
none substituents (13e-f) is more preferred. For the third series (14a-f), it’s seem to be that electron withdrawing groups
such as halogen atoms (-F,-Cl) in C-2 position or C-4 position is not favor to the activity and electron-donating group is
advantageous to the activity (14a-c vs 14d-f). For the fourth series (15a-i), substituted on C-2 position is not beneficial
to the activity (15d, 15f) and substituted on C-3 position or C-4 position is more preferred.
Activity against c-Met kinase of target compounds was further carried out in this paper to investigate the target of
these compounds. According to the results of Tables 1–4, we can easily find that the result is similar to the cytotoxicity
activity. Compounds 14a–f and 15a–i are more active than that of 12a–e and 13a–f against c-Met kinase, especially
compounds 15b, 15d and15f, with IC50 values in nanomole range. The results prompt us that the compounds are series
of excellent c-Met kinase inhibitors.
3
.2 Molecular docking study
To explore the binding modes of target compounds with the active site of c-Met, molecular docking simulation
studies were carried out by using SURFLEX-DOCK module of SYBYL package version. Based on the in vitro
inhibition results, we selected compound 15f, our best c-Met inhibitor in this study, as ligand example, and the structure
[5]
of c-Met was selected as the docking model (PDB ID code : 3LQ8 ).
Fig. 2 Binding poses of compound 15f with c-Met. The proteins were displayed by cyan ribbon. Compound 15f and
8

lead compound were displayed by orange and green sticks, respectively. H-bonding interactions between the 15f, lead
compound and c-Met were indicated with dashed lines in yellow.
The binding modes of compound 15f and lead compound Foretinib were shown in Fig. 2. As depicted in Fig. 2,
compound 15f and lead compound can nearly overlap in the binding model and pyrrolo[2,3-b]pyridine group and
phenylpicolinamide group formed three hydrogen bonds with residues MET1160 and LYS1110, respectively. Analysis
of compound 15f’s binding mode in the active binding site demonstrated that the docking mode of the 15f is similar to
the lead compound with the same H-bond between pyrrolo[2,3-b]pyridine group, phenylpicolinamide group and
MET1160, LYS1110. The three hydrogen bonds really play an important role in increasing the inhibitory potency of
pyrrolo[2,3-b]pyridine derivatives against c-Met kinase according to the docking results. Furthermore, the docking
results also give us a new direction to design new c-Met inhibitors. The above-mentioned results of SARs analysis and
molecular docking study may allow the rational design of more potent c-Met inhibitors.
4
. Conclusions
In summary, we designed and synthesized four series of phenylpicolinamide derivatives bearing
H-pyrrolo[2,3-b]pyridine moiety and evaluated for the IC50 values against three cancer cell lines and c-Met kinase.
1
Eighteen of the compounds showed excellent cytotoxicity activity with the IC50 valuables in single-digit μM to
nanomole range. Seven of them are equal to more active than positive control Foretinib against one or more cell lines.
The target compounds showed excellent selectivity toward A549 and PC-3 to MCF-7, HepG2 and Hela. The most
promising compound 15f showed superior activity to Foretinib, with the IC50 values of 1.04 ± 0.11 μM, 0.02 ± 0.21 μM
and 9.11 ± 0.55μM against A549, PC-3 and MCF-7 cell lines, which were 0.62 to 19.5 times more active than Foretinib,
respectively. Structure–activity relationships (SARs) and docking studies indicated that replacement of quinoline
nucleus with 1H-pyrrolo[2,3-b]pyridine moiety maintained even improved the potent cytotoxic activity. The results
suggested that the introduction of fluoro atoms to the aminophenoxy part of target compounds or the phenyl group of
pyrimidine substituted on C-4 position is benefit for the activity. The fourth series (15a-i) exhibited the best activity.
Furthermore, different substituted of aryl group affected the cytotoxicity of target compounds. Further study will be
carried out to identify the exact action mechanism in near future.
5
. Experimental
5
.1. Chemistry
All melting points were obtained on a Büchi Melting Point B-540 apparatus (Büchi Labortechnik, Flawil,
Switzerland) and were uncorrected. NMR spectra were performed using Bruker 400 MHz spectrometers (Bruker
Bioscience, Billerica, MA, USA) with TMS as an internal standard. Mass spectra (MS) were taken in ESI mode on
Agilent 1100 LCMS (Agilent, Palo Alto, CA, USA). The IR spectra were recorded by means of the KBr pellet
9

technique on WGH-30A drug-beam infrared spectrophotometer. All the materials were obtained from commercial
suppliers and used without purification, unless otherwise specified. Yields were not optimized. TLC analysis was
carried out on silica gel plates GF254 (Qindao Haiyang Chemical, China). All the materials were obtained from
commercial suppliers and used without purification, unless otherwise specied. Yields were not optimized.
5
5
.2 General procedure for the preparation of compounds 4a–k and 5a–k.
To the mixture of an appropriate amount of 4-bromo-pyridine-carboxylic acid 1 (0.90 mmol) or
-bromo-pyridine-carboxylic acid 2 (0.90 mmol), substituted phenylboronic acid 3a–k (0.18 mmol), anhydrous sodium
2
carbonate (0.29 g, 2.70 mmol), 1,4-dioxane (30 mL), H O(10 mL), and bis(triphenylphosphine)palladium(II) dichloride
(0.09 g, 0.135 mmol) were added under an atmosphere of nitrogen at room temperature. The reaction mixture then was
stirred at 100℃for 5-8 h and monitored by thin-layer chromatography (TLC). The reaction mixture was cooled to room
temperature and the resulting precipitate was isolated by filtration to furnish the acid intermediate as the sodium salt,
and then the solution was acidified to pH 5 to furnish the desired target compounds 4a–k and 5a–k.
5
.3 Preparation of phenylpyridine chloride 6a–k and 7a–k
Oxalyl chloride (0.5 mmol) was added drop-wise to a stirred mixture of compounds 4a–k or 5a–k (0.2 mmol) and
DMF( 0.01 mmol) in dichloromethane (10 mL) in room temperature for 10 min, and the mixture was distilled and
dissolved in dichloromethane (10 mL) immediately. The solution was used for the next step without further purification.
5
.4. General procedure for the key intermediates 11a,b
The mixture of 4-chloro-1H-pyrrolo[2,3-b]pyridine (10 g, 66 mmol) and 4-nitrophenol 9a (13.62 g, 98 mmol) or
-fluoro-4-nitrophenol 9b ( 15.56 g, 99 mmol) were refluxed in ether at 190℃ for 1 h and monitored by TLC. The
2
mixture was cooled to 70 ℃ and poured into 400 mL ethyl acetate in an ice bath for 30 mins, light yellow solid was
precipitated, filtered off and dried to obtain the title compound 10a or 10b.
Then 10a or 10b (20 mmol) was refluxed with hydrazine hydrate (200 mmol), Ferric chlorid（10 mmol) and an
appropriate amount of activated carbon in ethanol (100 mL) for 10min and monitored by TLC. The reaction solution
was cooled. Then the insoluble was filtered, the filtrate was evaporated, after that water was added and stirred at room
temperature for 30 min. Afterward, the yellow solid was filtered off and dried to obtain the desired target compounds
11a or 11b.
5
.5 General procedure for the preparation of compounds 12a–e, 13a–f, 14a–f and 15a–i
A solution of phenylpyridine chloride 6a–k or 7a–k (0.82 mmol) in dichloromethane (10 mL) was added drop-wise
to a solution of aniline 11a,b (0.41 mmol) and diisopropylethylamine (0.49 mmol) in dichloromethane (10 mL) in an ice
10

bath. Upon completion of the addition, the reaction mixture was removed from the ice bath and placed in room
temperature for 30min and monitored by TLC. The mixture was washed with 10% K CO (50 mL ×3) followed by brine
50 mL ×1), and the organic phase was separated, dried, and evaporated to yield 12a–e, 13a–f, 14a–f and 15a–i which
were purified by isopropanol.
2
3
(
5
.5.1 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-5-(4-chlorophenyl)picolinamide (12a)
+
This compound was obtained as reddish brown powder in 43.8% yield; ESI-MS m/z: 441.1[M+H] ; m.p. 254.8 ~
1
2
59.6 ℃; H NMR (400 MHz, DMSO) δ 11.73 (s, 1H), 10.84 (s, 1H), 9.05 (s, 1H), 8.38 (dd, J = 8.2, 2.0 Hz, 1H), 8.25
(d, J = 8.2 Hz, 1H), 8.09 (d, J = 5.4 Hz, 1H), 8.04 (d, J = 8.9 Hz, 2H), 7.89 (d, J = 8.4 Hz, 2H), 7.63 (d, J = 8.4 Hz, 2H),
7
.36 (s, 1H), 7.22 (d, J = 8.9 Hz, 2H), 6.43 (d, J = 5.4 Hz, 1H), 6.23 (s, 1H).
5
.5.2 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-5-p-tolylpicolinamide (12b)
+
1
This compound was obtained as white powder in 56.4% yield; ESI-MS m/z: 421.2[M+H] ; m.p. 269.5 ~ 270.6℃; H
NMR (400 MHz, DMSO) δ 11.73 (s, 1H), 10.81 (s, 1H), 9.03 (s, 1H), 8.34 (d, J = 8.1 Hz, 1H), 8.23 (d, J = 8.1 Hz, 1H),
8
6
.12 – 8.07 (m, 1H), 8.04 (d, J = 8.7 Hz, 2H), 7.75 (d, J = 7.7 Hz, 2H), 7.38 (d, J = 8.4 Hz, 3H), 7.22 (d, J = 8.6 Hz, 2H),
.43 (d, J = 5.2 Hz, 1H), 6.23 (s, 1H), 2.39 (s, 3H).
5
.5.3 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-5-(4-methoxyphenyl)picolinamide (12c)
+
This compound was obtained as light yellow powder in 55.3% yield; ESI-MS m/z: 437.2[M+H] ; m.p. 238.3 ~ 249.8℃;
1
H NMR (400 MHz, DMSO) δ 11.59 (s, 1H), 10.65 (s, 1H), 8.85 (s, 1H), 8.16 (d, J = 8.1 Hz, 1H), 8.05 (d, J = 8.2 Hz,
1
6
H), 7.92 (t, J = 5.8 Hz, 1H), 7.88 (d, J = 8.8 Hz, 2H), 7.65 (d, J = 8.6 Hz, 2H), 7.20 (s, 1H), 7.07 (d, J = 8.8 Hz, 2H),
.96 (d, J = 8.6 Hz, 2H), 6.27 (d, J = 5.4 Hz, 1H), 6.08 (s, 1H), 3.68 (s, 3H).
5
.5.4 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-5-(2,4-difluorophenyl)picolinamide (12d)
+
This compound was obtained as light brown powder in 51.4% yield; ESI-MS m/z: 443.1[M+H] ; m.p. 250.1 ~ 250.8℃;
1
H NMR (400 MHz, DMSO) δ 11.74 (s, 1H), 10.87 (s, 1H), 8.91 (s, 1H), 8.26 (q, J = 8.3 Hz, 2H), 8.09 (d, J = 5.4 Hz,
1
7
H), 8.04 (d, J = 8.8 Hz, 2H), 7.75-7.85 (m, 1H), 7.54 – 7.46 (m, 1H), 7.35 (d, J = 8.4 Hz, 1H), 7.31 (d, J = 8.4 Hz, 1H),
.22 (d, J = 8.8 Hz, 2H), 6.43 (d, J = 5.4 Hz, 1H), 6.23 (s, 1H).
5
.5.5 N-(4-((1H-pyrrolo[2,3-b]pyridin-4-yl)oxy)phenyl)-5-(4-ethylphenyl)picolinamide (12e)
11

+
1
This compound was obtained as white powder in 48.4% yield; ESI-MS m/z: 435.2[M+H] ; m.p.261.7 ~ 266.1℃; H
NMR (400 MHz, DMSO) δ 11.76 (s, 1H), 10.82 (s, 1H), 9.03 (s, 1H), 8.34 (d, J = 8.1 Hz, 1H), 8.23 (d, J = 8.2 Hz, 1H),
8
1
.09 (d, J = 5.4 Hz, 1H), 8.04 (d, J = 8.8 Hz, 2H), 7.77 (d, J = 8.0 Hz,2H), 7.41 (d, J = 7.9 Hz, 2H), 7.35 (t, J = 4.9 Hz,
H), 7.22 (d, J = 8.8 Hz,2H), 6.43 (d, J = 5.4 Hz, 1H), 6.23 (d, J = 3.2 Hz, 1H), 2.69 (q, J = 7.4 Hz, 2H), 1.23 (t, J = 7.5
Hz, 3H).
5
.5.6 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-4-p-tolylpicolinamide (13a)
+
This compound was obtained as white powder in 49.9% yield; ESI-MS m/z: 421.2[M+H] ; m.p. 262.1 ~ 266.2 ℃; IR
-1
(
KBr) cm : 3490.0, 3394.0, 3100.0, 3046.0, 3016.0, 3010.0, 1644.0, 1602.0, 1572.0, 1509.0, 1461.0, 1455.0, 1413.0,
1
1
5
380.0, 1350.0, 1257.0, 858.0, 831.0, 810.0. H NMR (400 MHz, DMSO) δ 11.73 (s, 1H), 10.84 (s, 1H), 8.79 (d, J =
.2 Hz, 1H), 8.41 (d, J = 1.3 Hz, 1H), 8.09 (d, J = 5.4 Hz, 1H), 8.06 – 8.02 (m, 2H), 8.00 (dd, J = 5.2, 1.9 Hz, 1H), 7.82
(d, J = 8.2 Hz, 2H), 7.39 (d, J = 8.0 Hz, 2H), 7.37 – 7.34 (m, 1H), 7.25 – 7.20 (m, 2H), 6.43 (d, J = 5.4 Hz, 1H), 6.23
+
(dd, J = 3.4, 2.0 Hz, 1H), 2.40 (s, 3H). ESI-HRMS m/z: calcd for C26
H
20
N
4
O
2
[M+H] :421.4396; found 421.4389.
5
.5.7 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-4-(4-methoxyphenyl)picolinamide (13b)
+
This compound was obtained as pale yellow powder in 61.5% yield; ESI-MS m/z: 437.2[M+H] ; m.p.229.8 ~ 233.4℃;
-1
IR (KBr) cm : 3478.0, 3370.0, 3100.0, 3058.0, 3022.0, 3010.0, 1674.0, 1584.0, 1539.0, 1497.0, 1461.0, 1455.0, 1413.0,
1
1
380.0, 1347.0, 1266.0, 1050.0, 852.0, 813.0, 804.0. H NMR (400 MHz, DMSO) δ 11.73 (s, 1H), 10.83 (s, 1H), 8.75
(d, J = 5.2 Hz, 1H), 8.39 (d, J = 1.4 Hz, 1H), 8.09 (d, J = 5.4 Hz, 1H), 8.06 – 8.01 (m, 2H), 7.97 (dd, J = 5.2, 1.9 Hz,
1
1
1
H), 7.92 – 7.86 (m, 2H), 7.38 – 7.33 (m, 1H), 7.25 – 7.20 (m, 2H), 7.14 (dd, J = 6.9, 4.9 Hz, 2H), 6.44 (d, J = 5.4 Hz,
13
H), 6.23 (dd, J = 3.4, 2.0 Hz, 1H), 3.84 (d, J = 5.6 Hz, 3H). C NMR (101 MHz, DMSO) δ 162.64, 160.75, 157.49,
51.26, 150.71, 149.11, 148.53, 144.33(2C), 135.54, 128.60, 128.43(2C), 124.72, 123.40, 121.97(2C), 120.80(2C),
+
118.87, 114.85(2C), 110.23, 102.11, 97.26, 55.40. ESI-HRMS m/z: calcd for C26
H
20
N
4
O
3
[M+H] :437.4370; found
4
37.4379.
5
.5.8 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-4-(4-(trifluoromethyl)phenyl)picolinamide (13c)
+
1
This compound was obtained as white powder in 55.6% yield; ESI-MS m/z: 475.2[M+H] ; m.p.225.8 ~ 229.7℃; H
NMR (400 MHz, DMSO) δ 11.73 (s, 1H), 10.88 (s, 1H), 8.88 (d, J = 5.1 Hz, 1H), 8.48 (d, J = 1.3 Hz, 1H), 8.15 (d, J =
8
2
.2 Hz, 2H), 8.11 – 8.07 (m, 2H), 8.07 – 8.02 (m, 2H), 7.94 (d, J = 8.3 Hz, 2H), 7.37 – 7.33 (m, 1H), 7.25 – 7.20 (m,
H), 6.44 (d, J = 5.4 Hz, 1H), 6.25 – 6.21 (m, 1H).
12

5
.5.9 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)phenyl)-4-(4-chlorophenyl)picolinamide (13d)
+
This compound was obtained as light brown powder in 48.6% yield; ESI-MS m/z: 441.1[M+H] ; m.p.258.5 ~ 262.4℃;
1
H NMR (400 MHz, DMSO) δ 11.73 (s, 1H), 10.86 (s, 1H), 8.83 (d, J = 5.3 Hz, 1H), 8.42 (d, J = 1.3 Hz, 1H), 8.09 (d, J
=
5.4 Hz, 1H), 8.05 (d, J = 2.1 Hz, 1H), 8.04 – 8.01 (m, 2H), 7.98 – 7.93 (m, 2H), 7.66 – 7.61 (m, 2H), 7.37 – 7.34 (m,
1
H), 7.25 – 7.20 (m, 2H), 6.44 (d, J = 5.4 Hz, 1H), 6.23 (dd, J = 3.4, 2.0 Hz, 1H).
5
.5.10 N-(4-((1H-pyrrolo[2,3-b]pyridin-4-yl)oxy)phenyl)-4-(m-tolyl)picolinamide (13e)
+
This compound was obtained as light brown powder in 49.5% yield; ESI-MS m/z: 421.2[M+H] ; m.p 211.5 ~ 212.6℃;
1
H NMR (400 MHz, DMSO) δ 11.74 (s, 1H), 10.86 (s, 1H), 8.80 (d, J = 5.1 Hz, 1H), 8.41 (s, 1H), 8.11 – 8.07 (m, 1H),
8
.04 (t, J = 8.3 Hz, 2H), 7.99 (t, J = 5.9 Hz, 1H), 7.73 (s, 1H), 7.69 (d, J = 7.7 Hz, 1H), 7.46 (t, J = 7.6 Hz, 1H), 7.35 (d,
J = 7.4 Hz, 2H), 7.23 (d, J = 8.8 Hz, 2H), 6.43 (d, J = 5.4 Hz, 1H), 6.23 (s, 1H), 2.43 (s, 3H).
5
.5.11 N-(4-((1H-pyrrolo[2,3-b]pyridin-4-yl)oxy)phenyl)-4-phenylpicolinamide (13f)
+
1
This compound was obtained as white powder in 46.9% yield; ESI-MS m/z: 407.1[M+H] ; m.p 205.8 ~ 206.5℃; H
NMR (400 MHz, DMSO) δ 11.74 (s, 1H), 10.86 (s, 1H), 8.82 (d, J = 5.1 Hz, 1H), 8.43 (s, 1H), 8.08 (t, J = 4.6 Hz, 1H),
8
1
.06 (s, 1H), 8.05 – 8.01 (m, 2H), 7.91 (d, J = 7.3 Hz, 2H), 7.59 (t, J = 7.1 Hz, 2H), 7.55 (d, J = 6.9 Hz, 1H), 7.36 (s,
H), 7.23 (d, J = 8.8 Hz, 2H), 6.43 (d, J = 5.4 Hz, 1H), 6.23 (s, 1H).
5
.5.12 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-5-phenylpicolinamide (14a)
+
1
This compound was obtained as white powder in 44.4% yield; ESI-MS m/z: 425.1[M+H] ; m.p.231.5 ~ 232.9℃; H
NMR (400 MHz, DMSO) δ 11.78 (s, 1H), 11.05 (s, 1H), 9.06 (s, 1H), 8.38 (d, J = 7.9 Hz, 1H), 8.27 (d, J = 8.1 Hz, 1H),
8
7
.19 - 8.13 (m, 1H), 8.10 (d, J = 5.2 Hz, 1H), 7.91 (d, J = 9.0 Hz, 1H), 7.86 (d, J = 7.4 Hz, 2H), 7.62 – 7.54 (m, 2H),
.52 (d, J = 7.2 Hz, 1H), 7.45 - 7.38 (m, 2H), 6.41 (d, J = 5.4 Hz, 1H), 6.28 (s, 1H).
5
.5.13 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-5-p-tolylpicolinamide (14b)
+
This compound was obtained as yellow powder in 48.9% yield; ESI-MS m/z: 477.2[M+K] ; m.p.248.5 ~ 250.4℃; IR
-
1
(
KBr) cm : 3478.0, 3160.0, 3100.0, 3064.0, 3058.0, 3010.0, 1656.0, 1593.0, 1539.0, 1497.0, 1461.0, 1455.0, 1407.0,
1
1
380.0, 1350.0, 1236.0, 840.0. H NMR (400 MHz, DMSO) δ 11.78 (s, 1H), 11.02 (s, 1H), 9.03(s, 1H), 8.35 (d, J = 8.0
Hz, 1H), 8.27 – 8.20 (m, 1H), 8.18 - 8.13 (m, 1H), 8.09 (d, J = 5.4 Hz, 1H), 7.90 (d, J = 8.7 Hz, 1H), 7.74 (d, J = 7.7 Hz,
1
3
1
H), 7.42 (t, J = 6.9 Hz, 1H), 7.38 (d, J = 7.3 Hz, 3H), 6.41 (d, J = 5.2 Hz, 1H), 6.28 (s, 1H), 2.40 (s, 3H). C NMR
13

(101 MHz, DMSO) δ 162.63, 157.05, 152.11, 150.97, 147.99, 146.07, 144.12, 138.48, 138.30, 135.40(2C), 132.98,
1
29.77(2C), 126.95(2C), 124.73, 123.50, 122.67, 116.87, 109.24, 108.93, 108.70, 100.54, 96.65, 20.64. ESI-HRMS m/z:
+
calcd for C26
H19FN
4
O
2
[M+H] :439.4285; found 439.4290.
5
.5.14 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-5-(4-methoxyphenyl)picolinamide (14c)
+
1
This compound was obtained as white powder in 56.9% yield; ESI-MS m/z: 493.1[M+K] ; m.p.241.7 ~ 243.6℃; H
NMR (400 MHz, DMSO) δ 11.78 (s, 1H), 11.01 (s, 1H), 9.02 (s, 1H), 8.33 (d, J = 8.2 Hz, 1H), 8.23 (d, J = 8.0 Hz, 1H),
8
1
.18 - 8.13 (m, 1H),, 8.09 (s, 1H), 7.90 (d, J = 8.8 Hz, 1H), 7.82 (d, J = 8.0 Hz, 2H), 7.44 (d, J = 9.2 Hz, 1H), 7.39 (s,
H), 7.13 (d, J = 8.3 Hz, 2H), 6.41 (d, J = 4.8 Hz, 1H), 6.28 (s, 1H), 3.85 (s, 3H).
5
.5.15 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-5-(4-fluorophenyl)picolinamide (14d)
+
1
This compound was obtained as yellow powder in 54.3% yield; ESI-MS m/z: 481.1[M+K] ; m.p.285.7 ~ 287.1℃; H
NMR (400 MHz, DMSO) δ 11.77 (s, 1H), 11.03 (s, 1H), 9.05 (s, 1H), 8.37 (d, J = 8.3 Hz, 1H), 8.26 (d, J = 7.9 Hz, 1H),
8
5
.18-8.13 (d, 1H), 8.09 (d, J = 5.3 Hz, 1H), 7.92 (t, J = 9.7 Hz, 3H), 7.43 (d, J = 8.8 Hz, 2H), 7.39 (s, 2H), 6.41 (d, J =
.3 Hz, 1H), 6.28 (s, 1H).
5
.5.16 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-5-(4-chlorophenyl)picolinamide (14e)
+
1
This compound was obtained as yellow powder in 53.3% yield; ESI-MS m/z: 459.1[M+H] ; m.p.242.5 ~ 243.7℃; H
NMR (400 MHz, DMSO) δ 11.77 (s, 1H), 11.06 (s, 1H), 9.04 (s, 1H), 8.39 (d, J = 8.1 Hz, 1H), 8.27 (d, J = 8.1 Hz, 1H),
8
.18 -8.12 (m, 1H), 8.08 (t, J = 7.5 Hz, 1H), 7.90 (d, J = 8.3 Hz, 3H), 7.64 (d, J = 8.4 Hz,2H), 7.45 – 7.37 (m, 2H), 6.41
(d, J = 5.2 Hz, 1H), 6.28 (s, 1H).
5
.5.17 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-5-(2,4-difluorophenyl)picolinamide (14f)
+
This compound was obtained as light yellow powder in 58.7% yield; ESI-MS m/z: 461.1[M+H] ; m.p.266.4 ~ 267.1℃;
-
1
IR (KBr) cm : 3472.0, 3292.0, 3082.0, 3034.0, 3028.0, 3010.0, 1635.0, 1578.0, 1533.0, 1491.0, 1452.0, 1407.0,
1
1
8
7
1
320.0, 1245.0, 798.0, 693.0. H NMR (400 MHz, DMSO) δ 11.77 (s, 1H), 11.06 (s, 1H), 8.93 (s, 1H), 8.29 (s, 2H),
.13 -8.18 (m, 1H), 8.09 (d, J = 5.4 Hz, 1H), 7.90 (d, J = 8.4 Hz, 1H), 7.81 (d, J = 8.6 Hz, 1H), 7.53 – 7.48 (m, 1H),
1
3
.42 (s, 1H), 7.39 (s, 1H), 7.32 (s, 1H), 6.41 (d, J = 5.3 Hz, 1H), 6.28 (s, 1H). C NMR (101 MHz, DMSO) δ 162.64,
57.23, 154.74, 152.31, 151.17, 148.84, 148.07, 144.31, 138.17, 137.06, 133.22, 132.39, 124.93, 123.68, 122.64,
15 3 4 2
117.10, 112.73, 112.52, 109.45, 109.19, 108.96, 105.21, 104.94,100.77, 96.84. ESI-HRMS m/z: calcd for C25H F N O
+
[
M+H] :461.1225; found 461.1220.
14

5
.5.18 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-phenylpicolinamide (15a)
+
This compound was obtained as light yellow powder in 42.7% yield; ESI-MS m/z: 425.1[M+H] ; m.p.263.6 ~
+
1
2
4
71.3℃;ESI-MS m/z: [M+K] 463.1; H NMR (400 MHz, DMSO) δ 11.67 - 11.62 (d, 1H), 11.07 (s, 1H), 8.81 (d, J =
.5 Hz, 1H), 8.41 (s, 1H), 8.16-8.12 (d, 1H), 8.07 (d, J = 5.1 Hz, 1H), 8.01 (s, 1H), 7.89 (d, J = 5.8 Hz, 3H), 7.62 – 7.50
(m, 3H), 7.42 (d, J = 8.8 Hz, 1H), 7.36 (s, 1H), 6.38 (d, J = 5.1 Hz, 1H), 6.24 (d, J = 14.0 Hz, 1H).
5
.5.19 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-m-tolylpicolinamide (15b)
This compound was obtained as white powder in 51.3% yield; ESI-MS m/z: 439.2[M+H]+; m.p.224.1 ~ 231.4℃; IR
-1
(
KBr) cm : 3496.0, 3430.0,,3100.0, 3010.0, 1680.0, 1581.0, 1488.0, 1476.0, 1461.0, 1410.0, 1380.0, 1350.0, 1341.0,
1
1
8
1
1
1
2
269.0, 1050.0, 771.0, 750.0. H NMR (400 MHz, DMSO) δ 11.78 (s, 1H), 11.07 (s, 1H), 8.82 (d, J = 5.1 Hz, 1H),
.42 (d, J = 1.3 Hz, 1H), 8.18 – 8.13 (m, 1H), 8.09 (d, J = 5.5 Hz, 1H), 8.03 – 8.00 (m, 1H), 7.93 – 7.88 (m, 1H), 7.74 (s,
H), 7.70 (d, J = 7.8 Hz, 1H), 7.49-7.42 (m, 2H), 7.41 – 7.33 (m, 2H), 6.40 (d, J = 5.4 Hz, 1H), 6.28 (dd, J = 3.5, 2.0 Hz,
1
3
H), 2.44 (s, 3H). C NMR (101 MHz, DMSO) δ 163.15, 157.43(2C), 151.37, 150.68, 149.46, 144.51(3C), 139.03,
36.69, 130.71, 129.56, 127.85, 125.13, 124.58, 124.41, 123.91, 119.95, 117.29, 109.64, 109.36, 109.12, 100.97, 97.04,
+
1.26. ESI-HRMS m/z: calcd for C26
H19FN
4
O
2
[M+H] :439.1570; found 439.1578.
5
.5.20 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-p-tolylpicolinamide (15c)
+
1
This compound was obtained as white powder in 53.0% yield; ESI-MS m/z: 477.2[M+K] ; m.p.242.9 ~ 246.7℃; H
NMR (400 MHz, DMSO) δ 11.40 (s, 1H), 11.22 – 11.00 (m, 1H), 8.89 (d, J = 4.7 Hz, 1H), 8.50 (s, 1H), 8.25 (d, J =
1
3.1 Hz, 1H), 8.18 (d, J = 5.2 Hz, 1H), 8.10 (d, J = 3.2 Hz, 1H), 7.99 (d, J = 8.7 Hz, 1H), 7.91 (d, J = 7.6 Hz, 1H), 7.52
(d, J = 6.8 Hz, 1H), 7.48 (d, J = 7.2 Hz, 3H), 6.49 (d, J = 5.1 Hz, 1H), 6.36 (s, 1H), 6.36 (s, 1H), 2.49 (s, 3H).
5
.5.21 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-(2,4-dimethylphenyl)picolinamide (15d)
+
This compound was obtained as white powder in 52.6% yield; ESI-MS m/z: 491.2[M+K] ; ESI-MS m/z:
-
1
[
M+H]+453.2; m.p.227.6 ~ 230.2℃; IR (KBr) cm : 3496.0, 3244.0, 3100.0, 3040.0, 3010.0, 1650.0, 1599.0,
1
1
530.0, 1470.0, 1461.0, 1452.0, 1419.0, 1380.0, 1350.0, 1269.0, 810.0, 690.0. H NMR (400 MHz, CDCl
3
) δ
11.76 (s, 1H), 11.04 (s, 1H), 8.78 (d, J = 4.6 Hz, 1H), 8.15-8.10 (m, 1H), 8.07 (s, 2H), 7.87 (d, J = 8.4 Hz, 1H),
7
7
1
.68 (d, J = 3.2 Hz, 1H), 7.41 (d, J = 8.9 Hz, 1H), 7.37 (s, 1H), 7.22 (d, J = 7.6 Hz, 1H), 7.19 (s, 1H), 7.15 (d, J =
13
.3 Hz, 1H), 6.38 (d, J = 5.1 Hz, 1H), 6.25 (s, 1H), 2.33 (s, 3H), 2.26 (s, 3H). C NMR (101 MHz, DMSO) δ
62.94, 157.22, 154.74, 152.31, 151.18, 150.79, 149.81, 148.56, 144.31, 138.34, 137.05, 135.14, 134.59, 131.51,
15

129.33, 127.31, 127.11, 124.93, 123.67, 122.66, 117.12, 109.19, 100.77, 96.84,20.74, 19.92, 9.6. ESI-HRMS m/z:
+
calcd for C27
H
21FN
4
O
2
[M+H] :453.1727; found 453.1732.
5
.5.22 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-(4-ethylphenyl)picolinamide (15e)
+
This compound was obtained as light yellow powder in 50.2% yield; ESI-MS m/z: 453.2[M+H] ; m.p.225.9 ~ 227.8℃;
-
1
IR (KBr) cm :3496.0, 3190.0, 3100.0, 3070.0, 3040.0, 3010.0, 1680.0, 1590.0, 1572.0, 1530.0, 1500.0, 1464.0,
1
1
1
7
1
461.0, 1410.0, 1380.0, 1350.0, 1269.0, 843.0, 801.0, 696.0. H NMR (400 MHz, Acetone) δ 11.78 (s, 1H), 11.05 (s,
H), 8.81 (d, J = 4.8 Hz, 1H), 8.42 (s, 1H), 8.19-8.14 (m, 1H), 8.09 (d, J = 5.2 Hz, 1H), 8.02 (d, J = 4.2 Hz, 1H), 7.93 –
.87 (m, 1H), 7.84 (d, J = 7.7 Hz, 2H), 7.46 – 7.36 (m, 4H), 6.41 (d, J = 5.2 Hz, 1H), 6.28 (s, 1H), 2.74-2.66 (m, 2H),
.24 (t, J = 7.4 Hz, 3H).
5
.5.23 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-(4-methoxyphenyl)picolinamide (15f)
+
This compound was obtained as light yellow powder in 55.7% yield; m.p.213.5 ~ 215.9℃; ESI-MS m/z: [M+H] 455.2;
-
1
IR (KBr) cm : 3484.0, 3196.0, 3100.0, 3076.0, 3010.0, 1680.0, 1590.0, 1569.0, 1470.0, 1461.0, 1452.0, 1407.0,
1
1
9
1
1
1
1
4
380.0, 1260.0, 1050.0, 1341.0, 837.0, 807.0, 717.0. H NMR (400 MHz, DMSO) δ 11.76 (s, 1H), 11.03-10.97 (m, 1H),
.00 (s, 1H), 8.75 (d, J = 5.0 Hz, 1H), 8.37 (s, 1H), 8.29 (s, 1H), 8.22 – 8.16 (m, 1H), 8.12 (s, 1H), 8.07 (d, J = 5.3 Hz,
H), 7.99 – 7.93 (m, 1H), 7.87 (d, J = 8.5 Hz, 1H), 7.79 (d, J = 8.5 Hz, 1H), 7.45 – 7.32 (m, 1H), 7.11 (d, J = 8.4 Hz,
13
H), 6.38 (d, J = 5.2 Hz, 1H), 6.26 (s, 1H), 3.83 (s, 3H). C NMR (101 MHz, DMSO) δ 163.01, 160.78, 160.13,
57.23，152.31, 151.17, 150.40, 149.17, 148.58, 147.73, 145.97, 144.31, 138.24, 137.17, 135.13, 128.45,124.92, 123.67,
+
22.81, 118.99, 117.04, 114.86, 109.12, 100.76, 96.84, 55.41.. ESI-HRMS m/z: calcd for C26
55.1519; found 455.1514.
H19FN
4
O
3
[M+H] :
5
.5.24 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-(4-fluorophenyl)picolinamide (15g)
+
This compound was obtained as light yellow powder in 52.2% yield; ESI-MS m/z: 443.1[M+H] ; m.p. 269.8 ~ 276.5℃;
-
IR (KBr) cm 1: 3478.0, 3226.0, 3100.0, 3082.0, 3052.0, 3010.0, 1671.0,
1593.0, 1542.0, 1485.0, 1413.0, 1320.0,
1
1
1
1
1
254.0, 789.0, 699.0, 846.0. H NMR (400 MHz, CDCl
3
) δ 11.75 (s, 1H), 11.03 (s, 1H), 8.80 (d, J = 4.7 Hz, 1H), 8.40 (s,
H), 8.17-8.10 (m, 1H), 8.07 (d, J = 5.2 Hz, 1H), 8.00 (d, J = 5.0 Hz, 1H), 7.96 (d, J = 6.9 Hz, 2H), 7.88 (d, J = 9.0 Hz,
1
3
H), 7.38 (d, J = 9.4 Hz, 4H), 6.38 (d, J = 5.2 Hz, 1H), 6.26 (s, 1H). C NMR (101 MHz, DMSO) δ162.88, 157.20,
54.73, 151.16, 150.53, 149.30, 148.02, 144.29(2C), 137.02, 133.01, 129.54, 129.45, 124.91, 124.30, 123.68, 119.69,
+
117.07, 116.46, 116.24, 109.43, 109.15, 108.92, 100.77, 96.81. ESI-HRMS m/z: calcd for C25
H
16
F
2
N
4
O
2
[M+H] :
4
43.1320; found 443.1328.
16

5
.5.25 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-(4-(trifluoromethyl)phenyl)picolinamide (15h)
+
1
This compound was obtained as white powder in 51.0% yield; ESI-MS m/z: 493.1[M+H] ; m.p. 243.1 ~ 244.3℃; H
NMR (400 MHz, DMSO) δ 11.86 (s, 1H), 11.17 (s, 1H), 8.98 (d, J = 4.4 Hz, 1H), 8.58 (s, 1H), 8.28 – 8.21 (m, 3H),
8
.19 (s, 2H), 8.05 – 7.96 (m, 3H), 7.52 (t, J = 8.8 Hz, 1H), 7.48 (s, 1H), 6.50 (d, J = 4.9 Hz, 1H), 6.37 (s, 1H).
5
.5.26 N-(4-(1H-pyrrolo[2,3-b]pyridin-4-yloxy)-3-fluorophenyl)-4-(2,4-difluorophenyl)picolinamide (15i)
+
This compound was obtained as light yellow powder in 51.8% yield; ESI-MS m/z: 461.1[M+H] ; m.p. 279.0 ~ 280.1℃;
1
3
H NMR (400 MHz, CDCl ) δ 11.76 (s, 1H), 11.06 (s, 1H), 8.85 (d, J = 4.8 Hz, 1H), 8.30 (s, 1H), 8.14-8.10 (d, 1H),
8
1
.07 (d, J = 5.3 Hz, 1H), 7.87 (d, J = 9.4 Hz, 2H), 7.85 – 7.79 (m, 1H), 7.48 (t, J = 10.0 Hz, 1H), 7.42 (d, J = 9.0 Hz,
H), 7.37 (s, 1H), 7.29 (t, J = 7.9 Hz, 1H), 6.38 (d, J = 5.2 Hz, 1H), 6.25 (s, 1H).
5
. 6 Cytotoxicity assay in vitro
The cytotoxic activities of target compounds (12a–e, 13a–f, 14a–f and 15a–i) were evaluated with A549, PC-3 and
MCF-7 cell lines and the five selected compounds (13b, 15b, 15d, 15e and 15f) were further evaluated for the activity
against HepG2 and Hela cell lines by the standard MTT assay in vitro, with compounds c-Met inhibitors Foretinib as
positive control. The cancer cell lines were cultured in minimum essential medium (MEM) supplement with 10% fetal
3
bovine serum (FBS). Approximately 4×10 cells, suspended in MEM medium, were plated onto each well of a 96-well
2
plate and incubated in 5% CO at 37°C for 24 h. The test compounds at indicated final concentrations were added to the
culture medium and the cell cultures were continued for 72 h. Fresh MTT was added to each well at a terminal
concentration of 5μg/mL and incubated with cells at 37°C for 4 h. The formazan crystals were dissolved in 100 μL
DMSO each well, and the absorbency at 492 nm (for absorbance of MTT formazan) and 630 nm (for the reference
wavelength) was measured with the ELISA reader. All of the compounds were tested three times in each of the cell
lines. The results expressed as inhibition rates or IC50 (half-maximal inhibitory concentration) were the averages of two
determinations and calculated by using the Bacus Laboratories Incorporated Slide Scanner (Bliss) software.
5
. 7 c-Met kinase assay in vitro
The target compounds (12a–e, 13a–f, 14a–f and 15a–i) are tested for their activity against c-Met kinase through the
[
8-9]
mobility shift assay
. All kinase assays were performed in 96-well plates in a 50 μL reaction volume. The kinase
, 0.0015% Brij-35 and 2 mM DTT. The stop buffer contains 100
mM HEPES, pH 7.5, 0.015% Brij-35, 0.2% Coating Reagent #3 and 50 mM EDTA. Dilute the compounds to 500 μM
buffer contains 50 mM HEPES, pH 7.5, 10 mM MgCl
2
17

by 100% DMSO, then transfer 10 μL of compound to a new 96-well plate as the intermediate plate, add 90 μL kinase
buffer to each well. Transfer 5 μL of each well of the intermediate plate to 384-well plates. The following amounts of
enzyme and substrate were used per well: kinase base buffer, FAM-labeled peptide, ATP and enzyme solution. Wells
containing the substrate, enzyme, DMSO without compound were used as DMSO control. Wells containing just the
substrate without enzyme were used as low control. Incubate at room temperature for 10 min. Add 10 μL peptide
solution to each well. Incubate at 28 °C for specified period of time and stop reaction by 25 μL stop buffer. At last
collect data on Caliper program and convert conversion values to inhibition values. Percent inhibition = (max −
conversion)/(max − min) × 100. “max” stands for DMSO control; “min” stands for low control.
5
. 8 Docking studies
For docking purposes, the three-dimensional structure of the c-Met (PDB code: 3LQ8) were obtained from RCSB
[5]
Protein Data Bank . Hydrogen atoms were added to the structure allowing for appropriate ionization at physiological
pH. The protonated state of several important residues, such as CYS919, and ASP1046 were adjusted by using
SYBYL6.9.1 (Tripos, St. Louis, USA) in favor of forming reasonable hydrogen bond with the ligand. Molecular
docking analysis was carried out by the SURFLEX-DOCK module of SYBYL 6.9.1 package to explore the binding
model for the active site of c-Met with its ligand. All atoms located within the range of 5.0 Å from any atom of the
cofactor were selected into the active site, and the corresponding amino acid residue was, therefore, involved into the
active site if only one of its atoms was selected. Other default parameters were adopted in the SURFLEX-DOCK
calculations. All calculations were performed on Silicon Graphics workstation.
Acknowledgments
We gratefully acknowledge the generous support provided by The National Natural Science Funds (No.
8
1460527), Top-notch talent project of Jiangxi Science & Technology Normal University (2015QNBJRC001),
Program of Key Laboratory of Drug Design and Optimization, Jiangxi Science & Technology Normal University
300098010306）, College Students' Science and Technology Innovation Project of Jiangxi Province and the
（
Innovation Fund Designated for Graduate Students of Jiangxi Province(YC2015-X28).
.
18

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[
[
[
[
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[
[
[
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5
7,162.
19

Graphical Abstract
5
4
R
H,F
N
H
X
N
bioisostere
of quinoline
Substituted on
C-4 or C-5 of
pyridine
O
O
N
1
1
2a-e,X=H 15f: R=-OCH
3
3a-f,X=H
14a-f,X=F
5a-i,X=F
IC :c-Met 0.69μM
5
0
N
H
A549 1.04μM
PC-3 0.02μM
MCF-7 9.11μM
1
20