Synthesis and antiproliferative activity of 6,7-disubstituted-4-phenoxyquinoline derivatives bearing the 1,8-naphthyridin-2-one moiety

Article abstract of DOI:10.1016/j.ejmech.2018.08.066

A series of 6,7-disubstituted-4-phenoxyquinoline derivatives bearing the 1,8-naphthyridin-2-one moiety were designed, synthesized and evaluated for their biological activities. The target compounds exhibited moderate to high antiproliferative activity against three cancer cell lines (A549, HepG2 and MCF-7) and several compounds (25, 27, 33, 37, 41, 43, 49 and 53) were evaluated for the activity against c-Met kinase. The most promising compound 33 (IC₅₀ c-Met = 2.36 nM) showed excellent activity against A549, HepG2 and MCF-7 cell lines with IC₅₀ values of 0.23 μM, 0.42 μM and 0.21 μM, respectively, which was 1.5–2.1 times of the positive control. Furthermore, compound 33 was evaluated for the activity against Flt3, PDGFR-α PDGFR-β c-Kit, Flt4, ALK and EGFR kinase. Structure activity relationship studies indicated that mono-EWGs (such as R₂ = F) at 4-position of moiety C was a key factor in improving the antitumor activity. In addition, further research on compound 33 was mainly including concentration dependence, apoptosis (acridine orange staining), apoptosis result analyzing and molecular docking.

Full text of DOI:10.1016/j.ejmech.2018.08.066

Accepted Manuscript
Synthesis and antiproliferative activity of 6,7-disubstituted-4-phenoxyquinoline
derivatives bearing the 1,8-naphthyridin-2-one moiety
Qidong Tang, Yongli Duan, Hehua Xiong, Ting Chen, Zhen Xiao, Linxiao Wang,
Yueyue Xiao, Shunmin Huang, Yinhua Xiong, Wufu Zhu, Ping Gong, Pengwu Zheng
PII:
S0223-5234(18)30738-4
10.1016/j.ejmech.2018.08.066
EJMECH 10677
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 30 April 2018
Revised Date: 23 August 2018
Accepted Date: 24 August 2018
Please cite this article as: Q. Tang, Y. Duan, H. Xiong, T. Chen, Z. Xiao, L. Wang, Y. Xiao, S. Huang,
Y. Xiong, W. Zhu, P. Gong, P. Zheng, Synthesis and antiproliferative activity of 6,7-disubstituted-4-
phenoxyquinoline derivatives bearing the 1,8-naphthyridin-2-one moiety, European Journal of Medicinal
Chemistry (2018), doi: 10.1016/j.ejmech.2018.08.066.
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ACCEPTED MANUSCRIPT
Graphical abstract
cytotoxic activities
Mechanism
cell cycle
A series of 6,7-disubstituted-4-phenoxyquinoline derivatives bearing the
1,8-naphthyridin-2-one moiety (25–56) were designed, synthesized and evaluated for
their activity against c-Met kinase and cancer cell lines. The most promising
compound 33 showed excellent in vitro activity.

ACCEPTED MANUSCRIPT
Synthesis and antiproliferative activity of 6,7-disubstituted-4-phenoxyquinoline derivatives
bearing the 1,8-naphthyridin-2-one moiety
Qidong Tang a, b, *, †
,
Yongli Duan a, †
,
, ,
Hehua Xiong a, Ting Chen a, Zhen Xiao a, Linxiao Wanga Yueyue Xiaoa
Shunmin Huang a Yinhua Xiong a
,
,
Wufu Zhu a
,
Ping Gong b, Pengwu Zheng a,
a Jiangxi Provincial Key Laboratory of Drug Design and Evaluation, School of Pharmacy, Jiangxi Science &
Technology Normal University, Nanchang 330013, PR China
b
Key Laboratory of Structure-Based Drug Design and Discovery of Ministry of Education, School of
Pharmaceutical Engineering, Shenyang Pharmaceutical University, Shenyang 110016, PR China
Abstract
A
series
of
6,7-disubstituted-4-phenoxyquinoline
derivatives
bearing
the
1,8-naphthyridin-2-one moiety were designed, synthesized and evaluated for their biological
activities. The target compounds exhibited moderate to high antiproliferative activity against three
cancer cell lines (A549, HepG2 and MCF-7) and several compounds (25, 27, 33, 37, 41, 43, 49 and
53) were evaluated for the activity against c-Met kinase. The most promising compound 33 (IC50
c-Met
= 2.36 nM) showed excellent activity against A549, HepG2 and MCF-7 cell lines with IC50
values of 0.23 µM, 0.42 µM and 0.21 µM, respectively, which was 1.5-2.1 times of the positive
control. Furthermore, compound 33 was evaluated for the activity against Flt3, PDGFR-α,
PDGFR-β, c-Kit, Flt4, ALK and EGFR kinase. Structure activity relationship studies indicated that
mono-EWGs (such as R2 = F) at 4-position of moiety C was a key factor in improving the antitumor
activity. In addition, further research on compound 33 was mainly including concentration
dependence, apoptosis (acridine orange staining), apoptosis result analyzing and molecular docking.
Keywords: Synthesis; quinoline derivatives; antiproliferative activity; c-Met
1. Introduction
Cancer is a widespread, complex and fatal disease that threaten human health and is the second
leading cause of death worldwide after cardiovascular disease [1-3]. At present, the main cancer
treatment includes surgery, chemotherapy, radiotherapy and small molecular targeted therapy.
Among of those, molecular targeted therapy is a new direction for tumor therapy [4-5]. c-Met is a
Corresponding authors. Tel./fax: +86 791 83802393.
E-mail addresses: tangqidongcn@126.com (Q. Tang), zhengpw@126.com (P. Zheng).
† These authors contribute equally to this work
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member of receptor tyrosine kinases (RTKs), also termed as hepatocyte growth factor receptor
(HGFR) [6-8]. The HGF/c-Met kinase pathway at the extracellular domain induce several complex
signaling pathways, which play an important role [9-10]. However, the abnormal activation of
c-Met occurs in many types of human cancers like liver, breast, gastric cancers and so on. Recently,
a respectable number of c-Met inhibitors with different molecular scaffolds have been discovered
and some derivatives bearing 6,7-disubstituted (or 7-substituted)-4-aryloxyquinoline moieties are
already being marketed or in clinical trials, such as Foretinib, Cabozantinib, Amgen, AM 7, MG 10
and Kirin (1-6, Fig. 1) [11-15]. We found most of those compounds are multikinase c-Met
inhibitors showed outstanding c-Met inhibition activity, which belong to Class II c-Met inhibitors.
Foretinib is a good example of these inhibitors of Class II. Structurally, those inhibitors are
composed of three moieties (Fig. 1): The first moiety is the phenoxy quinoline (moiety A). The
nitrogen in moiety A can form a hydrogen bond with Met1160, which enhances its inhibiting
activity and effectively increases the optimal selectivity of c-Met kinase. The second moiety is the
linker group (moiety B), which has two obvious structural characteristics: one is the ‘5 atoms
regulation’, which means six chemical bonds distance existing between moiety A and moiety C; the
other is the linker containing hydrogen, oxygen, and nitrogen atoms which could form
hydrogen-bond donor or acceptor with c-Met (Lys1110 and Asp1222). The third moiety is
substituted phenyl (moiety C). The binding mode of Foretinib with c-Met kinase is shown in Fig. 2.
(Figure 1. should listed here)
(Figure 2. should listed here)
In our previous studies, we described quinoline and 1,4-dihydroindoline fragments as part of a
5-atom linker and the result derivatives 7 and 8 (Fig. 3) showed excellent efficacy [16-17]. It is
worth noting that the 1,2-dihydro-1,8-naphthyridine-3-carbonyl fragment was widely used as a
building block in drug design due to its ability to be targeted to drug binding, such as the key
intermediate 9 [18]. So, the introduction of 1,2-dihydro-1,8-naphthyridin-3-carbonyl introduces a
5-atom linkage in moiety B while retaining the amide bond since the carbonyl oxygen and the two
nitrogen atoms in the 1,2-dihydro-1,8-naphthyridin-3-carbonyl have a high ability to form
hydrogen-bonding interactions with c-Met.
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In addition, in order to investigate the effect of the 4-carbon tether on the activity of the target
compounds at the 7-position of the moiety A, four water-soluble substituents were introduced,
including morpholinyl, 4-methylpiperazinyl, piperidinyl and 4-methylpiperidinyl. Furthermore,
various substituents were introduced into the benzene ring (moiety C) to investigate the effect on
inhibitory activity. Accordingly, we designed and synthesized
a
series of
6,7-disubstituted-4-phenoxyquinoline derivatives with a 1,8-naphthyridin-2-one moiety (Fig. 4).
(Figure 3. should listed here)
(Figure 4. should listed here)
2. Chemistry
2.1 Synthesis of 6,7-disubstituted-4-phenoxyquinolines (17a-d)
Starting from commercially available 1-(4-hydroxy-3-methoxyphenyl)ethaone, the key
intermediate 4-(2-fluorophenoxy) quinoline was obtained by a simple 8-step synthesis as shown in
Scheme 1, which was illustrated in detail in our previous study [17].
(Scheme 1. should be listed here)
2.2 Synthesis of the target compounds of 25-56
The synthesis of target compounds 25-56 were prepared as illustrated in Scheme 2.
2-chloronicotinic acid and the appropriately corresponding aniline were mixed in acetic acid at
100°C to afford intermediates 19a-h as a white solid. Reduction of 19a-h with LiAIH4 in THF gives
intermediates 20a-h as a yellow solid, which were reacted with pyridinium in dichloromethane to
obtain intermediates 21a-h. 21a-h and dimethyl malonate were acylated and closed at 110°C in
EtOH
to
give
the
key
intermediate
2-oxo-1-substituted
phenyl-1,2-dihydro-1,8-naphthyridine-3-carboxylate 22a-h. The corresponding acid chlorides
24a-h were obtained by hydrolysis and carboxylation; the reaction was carried out with K2CO3 and
thionyl chloride, respectively. The target compounds 25-56 were obtained at room temperature and
the reaction of the anilines 17a-d with the acid chlorides 24a-h were facilitated by DIPEA in
dichloromethane.
(Scheme 2. should be listed here)
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3 Results and Discussion
3.1 In vitro cytotoxic activities and SAR
The target compounds were evaluated for the cytotoxicity against three cancer cell lines
including HepG2, A549 and MCF-7 by MTT cell proliferation assay taken Foretinib as a reference.
Among them, A549 and HepG2 cells are belong to with high expression of c-Met, while MCF-7
cell line was not overexpressed in c-Met. In this research we chosen MCF-7 cell lines to study
whether these compounds show effective for low expression cell lines anti-proliferative effect [19].
Three parallel experiments were performed and the IC50 values were summarized in Table 1. As
learned from the data in Table 1, most of the target compounds showed moderate to excellent
cytotoxic activity against three cell lines, especially compound 33 exhibits the best activity against
HepG2, A549 and MCF-7 cell lines with the IC50 values of 0.23 ± 0.01 µM, 0.42 ± 0.06 µM and
0.21 ± 0.02 µM, which was more active than the reference Foretinib. It is unexpected discovery
that most compounds have significant selectivity for HepG2, which will make some contribution to
the field of anti-liver cancer, such as representative compounds 25 (IC50 = 0.76 ± 0.02), 30 (IC50 =
4.17 ± 0.87), 33 (IC50 = 0.23 ± 0.01), 34 (IC50 = 2.35 ± 0.92), 37 (IC50 = 0.36 ± 0.01), 41 (IC50 =
0.33 ± 0.01), 42 (IC50 = 1.49 ± 0.41), 43 (IC50 = 0.46 ± 0.12) and 52 (IC50 = 2.39 ± 0.72) against
HepG2 cell. Subsequent studies focused on the mechanism of compounds on HepG2 cells.
As showed in Table 1, when R1 was a different water-soluble group, it has a significant effect
on the inhibitory activity of the target compounds. It is worth mentioning that the
1-methylpiperazine group (R1 group) is superior to 4-methylpiperidine, piperidine and morpholine.
Such as compound 33 (R1 = 1-methylpiperazine, IC50 = 0.23 ± 0.01 µM), the inhibitory effect leads
25 (R1 = morpholine, IC50 = 0.76 ± 0.02 µM), 49 (R1 = piperidine, IC50 = 0.84 ± 0.37 µM) and 41
(R1 = 4-methylpiperidine, IC50 = 0.33 ± 0.01 µM) respectively against HepG2 cells. When R1 was
1-methylpiperazine which contains two hydrogen bonds accept (HBA), it could effectively increase
the interaction with the c-Met structural protein. Besides, the presence of the tertiary nitrogen
methyl group enables the molecule to rotate well, which provide a variety of effective direction of
action effectively to improve the anti-proliferative activity.
As found in Table 1, it is readily observed that there is a significant difference in cytotoxic
activity when various R2 groups are introduced into the phenyl ring (moiety C). The introduction of
mono-electron-withdrawing groups (mono-EWGs) at the 4-position of the phenyl ring showed
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strong cytotoxicity against HepG2 cells, such as representative compound 25 (IC50 = 0.76 ± 0.02
µM), 33 (IC50 = 0.23 ± 0.01 µM), 41 (IC50 = 0.33 ± 0.01 µM) and 49 (IC50 = 0.84 ± 0.37 µM).
Whereas the introduction of double-EWGs electron-donating at the 2nd / 3rd and 4th positions of
the phenyl ring resulted in a less negative on antitumor activity, such as compound 30 (IC50 = 4.17
± 0.87 µM), 37 (IC50 = 0.36 ± 0.01 µM), 44 (IC50 = 7.1 ± 1.69 µM) and 54 (IC50 = 4.92 ± 0.98 µM)
against HepG2 cells. It was observed that there was a significant negative effect of antitumor
activity during introduction of the mono-electron donor group (EDG) at the 4-position of the
benzene ring. It is noteworthy that when there is no substitution at 4-position of the phenyl ring,
tumor activity was significantly improved than the introduction of single electron donor groups, for
instance compounds 39 (R1 = 1-methylpiperazine, R2 = OCH3, IC50 = 7.92 ± 2.31 µM) and 40 (R1 =
1-methylpiperazine, R2 = H, IC50 = 3.237 ± 0.84 µM) against HepG2 cells.
(Table 1. should be listed here)
3.2 The relationship between the concentration of compound 33 and the cytotoxicity of three cancer
cell lines
In order to effectively studied whether compound 33 is concentration-dependent, the cells were
treated by MTT methods and seven concentration gradients were selected. After 72 hours of
interaction, the inhibition rate was determined. Not surprisingly, compound 33 was observed in a
concentration-dependent manner and was highly selective to HepG2 cells. The result was shown in
Fig. 5.
(Figure 5. should be listed here)
3.3 Morphological changes of HepG2 cells were observed under microscope and fluorescence
microscope
Based on the higher selectivity of the compound 33 on HepG2 cell line, the specific
mechanism of compound 33 on cytotoxicity was explored by Acridine Orange (AO) single staining.
As showed in Fig. 6, cells in the control group (Fig. 6a) were stained with acridine orange (AO).
The cells were oval in shape with clear margins and were seldom separated from the cell mass, none
of those are abnormal. However, under the conditions of compound 33 concentration of 1.1 µM and
acridine orange (AO) staining, cell shape was significantly abnormal and aggregated with each
other (Fig. 6b). The nucleus breaks down and disperses simultaneously, and the chromatin is
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concentrated. Compound 33 was identified as have a stronger apoptosis-inducing effect on HepG2
cells.
(Figure 6. should be listed here)
3.4 Apoptosis result analyzing
To further reveal the mechanism of compound 33 induced HepG2 apoptosis, the experiment of
Annexin V/PI staining was carried out [20-21]. As shown in Fig.7, compared with control group,
the compound 33 significantly increased the number of late apoptosis (20.35%) and its induction
effect is superior to the apoptosis rate of Foretinib (18.51%). The early apoptosis rate of compound
33 was greater than that of Foretinib. It could be concluded that the target compound 33 can induce
the apoptosis of HepG2 and is more effective than Foretinib.
(Figure 7. should be listed here)
3.5 In vitro enzymatic assays
The c-Met enzymatic assays of 6,7-disubstituted-4-phenoxyquinoline derivatives were
evaluated using homogeneous time-resolved fluorescence (HTRF) assay [21-22]. The results
expressed as the half maximal inhibitory concentration IC50 values presented in Table 2. The results
suggested that the inhibition of c-Met may be one mechanism of the antitumor effect of these
derivatives. The results show that these derivatives are closely related to the anti-tumor mechanism
of c-Met inhibitors (Table 2). Compound 33 showed the most potent activity with an IC50 value of
2.36 nM, which was comparable to the positive control Foretinib (IC50 = 1.83 nM) and this
compound should be studied further. Besides, enzymatic assays derived from these derivatives
show that: a single EWG introduced to R2 has a positive effect on inhibition and N-methyl
piperazinyl exerts an advantageous effect, which are consistent with the cells toxicity test results.
(Table 2. should be listed here)
3.6 Enzymatic selectivity assays
To explore the selectivity of compound 33 for other tyrosine kinases, which was screened
against the other 7 tyrosine kinases. As shown in Table 3, compound 33 showed comparable
potency to c-Met (IC50 = 2.36 nM) against PDGFR-α (IC50 = 4.35 nM) and Flt3 (IC50 = 5.31 nM).
While the inhibitory activities on c-Kit (IC50 = 10.12 nM), PDGFR-β (IC50 = 19.46 nM), Flt4 (IC50
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= 183.1 nM) and ALK (IC50 = 199.7 nM) were relatively drawback. These data indicated that
compound 33 is a multi-target c-Met kinase inhibitor.
(Table 3. should be listed here)
4. Binding model analysis
To explore the binding modes of target compounds with the active site of c-Met, molecular
docking simulation studies were carried out by using AutoDock vina 1.1.2, Open Babel and
Discover Studio 3.5 visualizer. Considerable research found that a new c-Met inhibitor
pharmacophore was found: Hydrogen bond acceptor (HBA), hydrophobic center (HY) and aromatic
(RA) as a prerequisite [23-24]. The promising target compound 33 was chosen as the docking
ligand and the c-Met structural protein (PDB ID code: 3LQ8) was chosen as docking model. The
docking results are shown in Fig.8. It is clear that the target compound 33 was well aligned with the
c-Met structural protein and the 4-methylpiperazine in the hydrophobic center of moiety A acts as
an HBA with His1162 form a hydrogen bond, however, the morpholino group in the lead
compound does not well interact well with c-Met as a water-soluble group, so the interaction
between compound 33 and c-Met was stronger than the lead compound. The quinoline ring in
moiety A forms an π-π system with the Tyr1159 and the nitrogen atom in the quinoline ring acts as
an HBA with methionine Met1160 form a hydrogen bond. The two oxygen atoms on the amide
bonds between moiety A and moiety C as an HBA forms two hydrogen bonds with Lys1110. It is
worth mentioning that due to tyrosine (Tyr) unique in the c-Met kinase domain [25], the compound
shows good selectivity for its interaction with it, which also well explains that the target compound
has a higher selective inhibition on HepG2. However, moiety B as a hydrophobic center does not
interact with c-Met structural protein and further optimization will be reported later.
(Figure 8. should be listed here)
5. Conclusions
In summary, a series of 6,7-disubstituted-4-phenoxyquinoline derivatives bearing the
1,8-naphthyridin-2-one moiety was designed, synthesized and evaluated for the IC50 values against
three cancer cell lines (A549, HepG2, MCF-7). According to the results of IC50 values, some
selected compounds (25, 27, 33, 37, 41, 43, 49, 53) were evaluated for activity against c-Met kinase.
Afterwards, based on the results of the IC50 values on c-Met kinase assays, the promising compound
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33 (c-Met IC50 = 2.36 nM) was selected for the selective determination of c-Met against other
kinases (PDGFR-α, Flt3, c-Kit, PDGFR-β, Flt4, ALK and EGFR), the measurement dependence
and the cytomorphological validation. What’s more, analysis of SARs indicated that introduction of
mono-electron-withdrawing groups (mono-EWGs) was ahead of introduction of double-EWGs
electron-donating and the mono-electron donor group (EDG) and the 2nd/3rd and 4th positions of
the phenyl ring (moiety C). Moreover, the most obvious positive effect on the antiproliferative
activity of the compound when R1 was 1-methylpiperazinyl. Further studies on the mechanism of
action of these compounds are in progress and their results will be reported in the future.
6. Experimental
6.1 Chemistry
All melting points were obtained on a Büchi Melting Point B-540 apparatus (Büchi Labor
technik, 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 LCeMS (Agilent, Palo Alto, CA,
USA). 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 specified. Yields were not optimized.
6.2 Preparation of 3-fluoro-4-(6,7-disubstituted quinolin-4-yloxy) anilines (17a-d)
The preparation of the key intermediate 17a-d has been illustrated in detail in our previous work,
and so the synthesis method would not be listed here.
6.2.1.3-Fluoro-4-(6-methoxy-7-(3-morpholinopropoxy)quinolin-4-yloxy)aniline (17a)
1
Light yellow solid; yield: 82.6%; m.p.: 216.1-217.2°C; H NMR(400 MHz, DMSO-d6) δ 8.43
(d, J = 5.2 Hz, 1H), 7.49 (s, 1H), 7.36 (s, 1H), 7.05 (t, J = 9.0 Hz, 1H), 6.54 (dd, J = 9.2, 7.3 Hz,
1H), 6.46 (d, J = 8.6 Hz, 1H), 6.37 (d, J = 7.1 Hz, 1H), 5.47 (s, 2H), 4.16 (t, J = 7.2 Hz, 2H), 3.92 (s,
3H), 3.57 (d, J = 4.1 Hz, 4H), 2.44 (t, J = 7.0 Hz, 2H), 2.36 – 2.29 (m, 4H), 2.00 – 1.90 (m, 2H).
6.2.2.3-Fluoro-4-(6-methoxy-7-(3-(4-methylpiperazine-1-yl)propoxy)quinolin-4-yl-oxy)anilin (17b)
8

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1
White solid; yield: 72%; m.p.: 203.4-202.1°C; H NMR (400 MHz, DMSO-d6) δ 8.43 (d, J =
5.1 Hz, 1H), 7.49 (s, 1H), 7.34 (s, 1H), 7.05 (t, J = 8.9 Hz, 1H), 6.61 – 6.49 (m, 1H), 6.45 (d, J = 8.4
Hz, 1H), 6.37 (d, J = 4.7 Hz, 1H), 5.47 (s, 2H), 4.16 (d, J = 5.8 Hz, 2H), 3.92 (s, 3H), 2.49 (s, 2H),
2.40 – 2.23 (m, 8H), 2.12 (s, 3H), 2.00 – 1.88 (m, 2H).
6.2.3.3-Fluoro-4-(6-methoxy-7-(3-(piperdine-1-yl)propoxy)quinolin-4-yloxy)aniline (17c)
1
White solid; yield: 56%; m.p.: 188.5-191.2°C; H NMR (400 MHz, DMSO-d6) δ 8.43 (d, J =
5.1 Hz, 1H), 7.49 (s, 1H), 7.34 (s, 1H), 7.05 (t, J = 9.0 Hz, 1H), 6.54 (d, J = 10.9 Hz, 1H), 6.45 (d, J
= 8.4 Hz, 1H), 6.37 (d, J = 6.2 Hz, 1H), 5.47 (s, 2H), 4.15 (s, 2H), 3.92 (s, 3H), 2.39 (d, J = 7.1 Hz,
2H), 2.32 (s, 3H), 1.94 (d, J = 6.7 Hz, 2H), 1.48 – 1.42 (m, 4H), 1.37 (s, 2H).
6.2.4.3-Fluoro-4-(6-methoxy-7-(3-(4-methylpiperdine-1-yl)propoxy)quinolin-4-yl-oxy)aniline (17d)
Light yellow solid; yield: 53%; m.p.: 195.1-194.2°C; 1H NMR (400 MHz, DMSO-d6) δ 8.43 (s,
1H), 7.49 (s, 1H), 7.34 (s, 1H), 7.05 (s, 1H), 6.54 (d, J = 9.6 Hz, 1H), 6.46 (s, 1H), 6.37 (s, 1H),
5.47 (s, 2H), 4.15 (s, 2H), 3.92 (s, 3H), 2.83 (s, 2H), 2.41 (s, 2H), 1.89 (d, J = 5.8 Hz, 4H), 1.56 (s,
2H), 1.30 (s, 1H), 1.13 (s, 2H), 0.87 (s, 3H).
6.3 General procedure for the preparation of compounds 25-56
Corresponding anilines 18a-h (0.112 mol) and 2-chloronicotinic acid (0.064 mol) in glacial
acetic acid (75 mL) were added at room temperature and the reaction mixture was stirred at 100°C
for 3-6 h and analyzed by thin layer chromatography (TLC). The reaction mixture was cooled to
room temperature, added with the appropriate amount of water, basified with potassium hydroxide
to pH = 12 and filtered. The filtrate was acidified to pH = 3 with hydrochloric acid and stirred for
0.5 h. The precipitate was collected by filtration and dried to give the corresponding products 19a-h
as a white solid.
Lithium aluminum hydride (LiAlH4, 0.228 mol) was added portion wise to tetrahydrofuran
(THF, 40 mL). The mixture was stirred in an ice bath for 30 minutes under a nitrogen atmosphere at
0°C. The intermediates 19a-h (0.054 mol) dissolved in THF was slowly added and stirred at room
temperature for 3.5 hours. The reaction was added to ethyl acetate, stirred for 30 minutes and
basified to pH = 12 with potassium hydroxide and filtered. The filtrate was concentrated in vacuum
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and stirred the residue in petroleum ether. The precipitate was collected by filtration and dried to
give the corresponding intermediates 20a-h as a white solid.
Intermediates 20a-h (0.072 mol), pyridinium dichromate on silica gel and added to CH2Cl2 (75
mL) at room temperature. The reaction mixture is stirred at room temperature for 5-7 hours and
monitored by thin layer chromatography (TLC). The reaction mixture was filtered, the filtrate was
concentrated in vacuo and the residue stirred in water (70 mL) for 0.5h. The precipitate was
collected by filtration and dried to give the corresponding intermediates 21a-h as a pale yellow
solid.
To the intermediates 21a-h, diethyl malonate (0.022 mol) and piperidine (0.009 mol) were
added EtOH (75 mL), refluxed for 30-35 hours then the solvent was concentrated in vacuo and the
residue was treated with H2O (70 mL) for 0.5 h at rt. The precipitate was collected by filtration and
dried to give the key intermediates 22a-h.
To a mixture of dioxane (35 mL) and water (35 mL) was added the key intermediates 22a-h
(16 mmol) and potassium carbonate (45 mmol). The reaction mixture is heated at 80°C for 4-5
hours. Water (150 mL) was added to the mixture and the pH = 6 with hydrochloric acid. A large
amount of solid was aspirated and the precipitate collected by filtration and dried to give the
corresponding the key intermediates 23a-h as a yellow solid.
The key intermediates 23a-h (4 mmol) was added to thionyl chloride (10 mL), a proper
amount of DMF was added dropwise and refluxed for 0.5 hour. The reaction mixture is evaporation
produces the corresponding intermediates 24a-h, which used immediately for the next step without
further purification.
6.3.1. 1-(4-fluorophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxylic acid (24b)
White solid; yield: 72.6%; 1H NMR (400 MHz, DMSO-d6) δ 13.69 (s, 1H), 9.04 (d, J = 9.5 Hz,
1H), 8.64 – 8.52 (m, 2H), 8.06 (s, 1H), 7.98 (d, J = 8.5 Hz, 1H), 7.87 (d, J = 9.4 Hz, 1H), 7.85 (s,
1H), 7.50 (d, 8.7 Hz, 1H).
6.3.2. 1-(4-chlorophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxylic acid (24c)
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White solid; yield: 79.3%; 1H NMR (400 MHz, DMSO-d6) δ 13.75 (s, 1H), 8.95 (s, 1H), 8.53
(d, J = 9.3 Hz, 1H), 8.48 (d, J = 6.6 Hz, 1H), 8.45 (d, J = 7.6 Hz, 1H), 7.57 (d, 1H), 7.55 (s, 1H),
7.35 (d, J = 8.5 Hz, 1H).
6.3.3. 1-(4-bromophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxylic acid (24d)
White solid; yield: 84.3%; 1H NMR (400 MHz, DMSO-d6) δ 13.84 (s, 1H), 9.01 (s, 1H), 8.62
– 8.51 (m, 2H), 7.75 (t, J = 8.8 Hz, 2H), 7.48 (dd, J = 7.7, 6.2 Hz, 1H), 7.35 (d, J = 8.5 Hz, 2H).
6.3.4. 1-(4-methoxyphenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxylic acid (24e)
1
White solid; yield: 82.7%; H NMR (400 MHz, DMSO-d6) δ 14.05 (s, 1H), 9.05 (s, 1H), 8.64
(dd, J = 7.9, 7.3 Hz, 1H), 8.57 (dd, J = 7.8, 6.6 Hz, 1H), 7.49 (dd, J = 7.8, 5.7 Hz, 1H), 7.28 (d, J =
8.8 Hz, 2H), 7.11 (d, J = 8.9 Hz, 2H), 3.85 (s, 3H).
6.3.5. 1-(3-chloro-4-fluorophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxylic acid (24g)
White solid; yield: 88.3%; 1H NMR (400 MHz, DMSO-d6) δ 13.67 (s, 1H), 9.01 (s, 1H), 8.60
(d, J = 7.5 Hz, 1H), 8.55 (d, J = 7.8 Hz, 1H), 8.05 (s, 1H), 7.96 (d, J = 8.4 Hz, 1H), 7.78 (d, J = 7.5
Hz, 1H), 7.48 (dd, J = 7.7, 9.7 Hz, 1H).
6.4.6.1-(2-chloro-4-(trifluoromethyl)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxylic
acid (24h)
White solid; yield: 78.9%; 1H NMR (400 MHz, DMSO-d6) δ 13.67 (s, 1H), 9.01 (s, 1H), 8.60
(d, J = 7.1 Hz, 1H), 8.55 (d, J = 7.8 Hz, 1H), 8.05 (s, 1H), 7.96 (d, J = 8.4 Hz, 1H), 7.78 (d, J = 8.5
Hz, 1H), 7.48 (dd, J = 7.7, 9.7 Hz, 1H).
The appropriate anilines 17a-d (6 mmol) and DMF (2 mmol) were dissolved in 25 mL
dichloromethane, the product of the previous step was dissolved in anhydrous dichloromethane (20
mL). Is was added dropwise to the former 24a-h in an ice bath. After the addition was completed,
the reaction was moved to room temperature for 3-5 hours and monitored by TLC. The mixture was
washed with 10% K2CO3 (60 mL) then brine (40 mL), the organic phase was separated, dried and
concentrated in vacuum. The crude product was purified by silica gel chromatography using MeOH
/ CH2Cl2 to give the target compound 25-56 as a white solid.
11

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6.3.7.N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-yl)oxy)phenyl)-1-(4-fluorophe
nyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (25)
1
Yield: 67%; m.p.: 178.8–179.6°C; H NMR (400 MHz, DMSO-d6) δ 11.95 (s, 1H), 9.17 (s,
1H), 8.63 (s, 1H), 8.61 (d, J = 7.3 Hz, 1H), 8.58 (d, J = 7.5 Hz, 1H), 8.11 (dd, J = 10.9, 7.1 Hz, 1H),
7.67 (d, J = 8.9 Hz, 1H), 7.62 (s, 1H), 7.51 (dd, J = 9.8, 7.7 Hz, 2H), 7.50 – 7.47 (m, 2H), 7.46 –
7.42 (m, 2H), 6.63 (d, J = 7.2 Hz, 1H), 4.28 (d, J = 8.7 Hz, 2H), 3.98 (s, 3H), 3.51 (d, J = 9.8 Hz,
2H), 2.99 – 2.87 (m, 2H), 2.32 (s, 2H), 1.80 (d, J = 9.4 Hz, 2H), 1.55 – 1.42 (m, 2H), 0.93 (d, J =
6.3 Hz, 2H); Anal. Calcd. for C37H32F2N6O (%): C, 65.74; H, 4.78; N, 19.10; Found (%): C, 65.79;
H, 4.81; N, 19.12.
6.3.8.1-(4-chlorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-yl)oxy)phe
nyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (26)
Yield: 56%; m.p.: 214.1–216.3°C; 1H NMR (400 MHz, DMSO) δ 11.93 (s, 1H), 11.22 (s, 1H),
9.34 (s, 1H), 9.16 (s, 1H), 8.67 (d, J = 5.6 Hz, 1H), 8.62 – 8.59 (m, 2H), 8.14 – 8.09 (m, 1H), 7.66
(s, 2H), 7.64 (s, 1H), 7.55 (t, J = 8.9 Hz, 1H), 7.49 – 7.47 (m, 1H), 7.46 (s, 1H), 7.44 (s, 1H), 6.77 (s,
1H), 5.75 (s, 1H), 4.30 (t, J = 5.7 Hz, 2H), 3.99 (s, 3H), 3.83 (t, J = 6.7 Hz, 2H), 3.60 – 3.57 (m,
2H), 3.57 – 3.54 (m, 2H), 2.42 (d, J = 8.1 Hz, 2H), 1.85 (d, J = 9.4 Hz, 2H), 1.65 – 1.57 (m, 2H),
0.89 (d, J = 4.5 Hz, 2H); Anal. Calcd. for C37H32ClFN6O (%): C, 64.26; H, 4.27; N, 9.85; Found
(%): C, 64.27; H, 4.28; N, 9.86.
6.3.9.1-(4-bromophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-yl)oxy)phe
nyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (27)
1
Yield: 58%; m.p.: 264.1–265.3°C; H NMR (400 MHz, DMSO-d6) δ 11.97 (s, 1H), 9.24 (s,
1H), 8.68 (d, J = 8.7 Hz, 2H), 8.55 (d, J = 7.1 Hz, 1H), 8.16 (d, J = 8.8 Hz, 1H), 7.87 (d, J = 8.5 Hz,
1H), 7.69 (s, 1H), 7.61 (s, 1H), 7.56 (d, J = 8.5 Hz, 2H), 7.49 (s, 2H), 7.46 (s, 1H), 6.56 (d, J = 7.0
Hz, 1H), 4.30 (s, 3H), 4.03 (s, 2H), 3.79 (s, 2H), 3.69 – 3.64 (m, 2H), 3.19 (d, J = 7.2 Hz, 2H), 2.58
(s, 2H), 2.19 (s, 2H), 1.37 (s, 1H), 1.35 (s, 2H), 1.34 (s, 1H); Anal. Calcd. for C37H32BrFN6O6 (%):
C, 61.50; H, 3.53; N, 12.46; Found (%): C, 61.51; H, 3.54; N, 12.47.
6.3.10.1-(3-chloro-4-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-
yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (28)
12

ACCEPTED MANUSCRIPT
1
Yield: 68%; m.p.: 190.8–191.6°C; H NMR (400 MHz, DMSO-d6) δ 11.84 (s, 1H), 9.17 (s,
1H), 8.62 (s, 2H), 8.47 (d, J = 8.4 Hz, 1H), 8.08 (d, J = 9.6 Hz, 1H), 7.81 (d, J = 7.5 Hz, 1H), 7.65
(dd, J = 9.8, 7.8 Hz, 2H), 7.51 (d, J = 7.9 Hz, 3H), 7.39 (s, 1H), 6.47 (d, J = 7.4 Hz, 1H), 4.19 (s,
2H), 3.94 (s, 3H), 3.60 – 3.51 (m, 5H), 2.44 – 2.39 (m, 4H), 1.99 (s, 3H), 1.45 – 1.51 (m, 2H), 0.91
(d, J = 5.1 Hz, 2H); Anal. Calcd. for C36H31ClF2N6O6 (%): C, 62.47; H, 4.70; N, 9.57; Found (%): C,
62.48; H, 4.71; N, 9.58.
6.3.11.1-(4-bromo-2-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-
yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (29)
1
Yield: 71%; m.p.: 166.4–168.8°C; H NMR (400 MHz, DMSO-d6) δ 11.66 (s, 1H), 9.18 (d, J
= 7.8 Hz, 1H), 8.63 (d, J = 9.9 Hz, 2H), 8.48 (d, J = 7.1 Hz, 1H), 8.07 (d, J = 10.8 Hz, 1H), 7.89 (d,
J = 9.4 Hz, 1H), 7.67 (d, J = 9.5 Hz, 1H), 7.63 (d, J = 8.2 Hz, 1H), 7.58 (d, J = 8.1 Hz, 1H), 7.54 (s,
1H), 7.50 (d, J = 7.5 Hz, 1H), 7.43 (s, 1H), 6.49 (d, J = 8.5 Hz, 1H), 4.25 (s, 2H), 3.95 (s, 3H), 3.83
(s, 2H), 3.58 (s, 1H), 3.20 (s, 2H), 3.12 (s, 2H), 2.26 (s, 2H), 1.28 (s, 1H), 1.27 (s, 1H), 1.25 (s, 1H),
1.23 (s, 1H); Anal. Calcd. for C37H31BrF2N6O6 (%): C, 62.67; H, 4.35; N, 9.61; Found (%): C, 62.68;
H, 4.34; N, 9.62.
6.3.12.1-(2-chloro-4-(trifluoromethyl)phenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)
quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (30)
Yield: 68%; m.p.: 234.4–235.5°C; 1H NMR (400 MHz, DMSO-d6) δ 8.45 (d, J = 5.2 Hz, 1H),
7.51 (s, 1H), 7.38 (s, 1H), 7.07 (t, J = 8.0 Hz, 1H), 6.56 (dd, J = 9.2, 8.4 Hz, 1H), 6.47 (dd, J = 8.7,
6.3 Hz, 1H), 6.39 (d, J = 7.2 Hz, 1H), 5.76 (s, 4H), 5.50 (s, 2H), 4.19 (t, J = 6.4 Hz, 2H), 3.94 (s,
3H), 3.59 (t, J = 4.4 Hz, 4H), 2.50 – 2.49 (m, 1H), 2.47 (t, J = 7.1 Hz, 2H), 2.39 – 2.32 (m, 4H),
1.98 (d, J = 6.6 Hz, 2H); 13C NMR (100 MHz, DMSO-d6) δ 161.92, 160.25, 158.88, 152.14, 151.15,
150.16(2C), 149.00, 148.10, 145.30, 143.44, 138.93, 136.45, 136.37, 135.69, 135.61, 135.24,
132.58, 130.48, 130.33, 128.77, 123.70, 121.88, 119.64, 119.40, 116.33, 114.15, 108.45, 108.23,
107.94, 101.75, 98.65, 65.61, 55.33(2C), 53.01, 52.72, 51.35(2C), 30.19; Anal. Calcd. for
C37H31ClF5N6O6 (%): C, 59.09; H, 4.18; N, 9.07; Found (%): C, 59.08; H, 4.17; N, 9.06.
6.3.13.N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-yl)oxy)phenyl)-1-(4-methoxy
phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (31)
13

ACCEPTED MANUSCRIPT
1
Yield: 54%; m.p.: 198.5–200.4°C; H NMR (400 MHz, DMSO-d6) δ 12.01 (s, 1H), 9.64 (s,
1H), 9.13 (s, 1H), 8.59 (dd, J = 10.7, 8.5 Hz, 2H), 8.09 (d, J = 7.7 Hz, 1H), 7.67 – 7.56 (m, 2H),
7.47 (d, J = 9.4, 2H), 7.30 (d, J = 8.6 Hz, 1H), 7.12 (d, J = 8.6 Hz, 1H), 6.58 (d, J = 7.3 Hz, 1H),
5.76 (s, 2H), 4.29 (s, 1H), 3.96 (d, J = 9.3 Hz, 3H), 3.85 (d, J = 4.5 Hz, 3H), 3.11 (dd, J = 7.0, 5.1
Hz, 1H), 2.35 (s, 1H), 1.37 – 1.32 (m, 5H), 1.29 – 1.22 (m, 5H), 1.24 (s, 1H); Anal. Calcd. for
C38H31FN6O7 (%): C, 64.58; H, 4.99; N, 11.89; Found (%): C, 64.56; H, 4.97; N, 11.88.
6.3.14.N-(3-fluoro-4-((6-methoxy-7-(3-morpholinopropoxy)quinolin-4-yl)oxy)phenyl)-2-oxo-1-phen
yl-1,2-dihydro-1,8-naphthyridine-3-carboxamide (32)
1
Yield: 65%; m.p.: 211.3–212.5°C; H NMR (400 MHz, DMSO-d6) δ 11.93 (s, 1H), 9.15 (s,
1H), 8.58 (d, J = 7.6 Hz, 2H), 8.45 (d, J = 8.2 Hz, 1H), 8.06 (dd, J = 9.8, 7.7 Hz, 1H), 7.60 – 7.55
(m, 3H), 7.51 (s, 1H), 7.48 – 7.44 (m, 2H), 7.40 – 7.36 (m, 3H), 6.46 (d, J = 7.1 Hz, 1H), 4.19 (t, J
= 6.0 Hz, 2H), 4.01 (d, J = 7.1 Hz, 2H), 3.93 (s, 3H), 3.62 (s, 3H), 1.97 (s, 2H), 1.25 (d, J = 6.6 Hz,
13
2H), 1.16 (t, J = 7.1 Hz, 3H); C NMR (100 MHz, DMSO-d6) δ 162.03, 160.29, 158.65, 154.20,
152.16, 151.34, 150.28, 149.05, 148.30, 145.83, 143.38, 138.86, 136.36, 128.69(2C), 128.45(2C),
127.83, 123.64, 121.82, 119.27, 116.29, 114.08, 108.48, 108.25, 108.10, 101.65, 98.57, 66.03,
65.06, 55.26(2C), 54.04(2C), 52.79, 52.38, 24.57; Anal. Calcd. for C37H33FN6O6 (%): C, 65.67; H,
4.92; N, 12.42; Found (%): C, 65.68; H, 4.93; N, 12.43.
6.3.15.N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-
1-(4-fluorophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (33)
1
Yield: 61%; m.p.: 224.4–225.9°C; H NMR (400 MHz, DMSO-d6) δ 11.91 (s, 1H), 9.14 (s, 1H),
8.59 (t, J = 7.6 Hz, 2H), 8.47 (d, J = 8.1 Hz, 1H), 8.06 (dd, J = 9.8, 8.0 Hz, 1H), 7.61 (d, J = 8.9 Hz,
1H), 7.53 (s, 1H), 7.48 – 7.45 (m, 3H), 7.43 (s, 1H), 7.39 (t, J = 7.5 Hz, 2H), 6.47 (d, J = 7.0 Hz,
1H), 4.21 (s, 2H), 3.95 (s, 3H), 3.69 (s, 3H), 2.79 – 2.76 (m, 4H), 2.08 (s, 2H), 1.36 – 1.30 (m, 5H),
1.04 (d, J = 6.1 Hz, 1H); 13C NMR (100 MHz, DMSO-d6) δ δ 162.31, 161.21, 160.11, 159.36,
156.85, 155.26, 153.57, 152.82, 152.33, 150.49, 150.18, 149.21, 146.29, 145.04 , 140.35, 133.39,
131.70, 129.08, 124.79(2C), 123.30, 121.05, 120.55, 120.32, 117.59, 115.30(2C), 109.61, 108.98 ,
102.96 , 99.83 , 66.70 , 64.03 , 56.48(2C) , 54.40(2C) , 51.93 , 23.64; Anal. Calcd. for
C37H36F2N7O5 (%): C, 66.17; H, 5.26; N, 14.22; Found (%):C, 66.18; H, 5.27; N, 14.23.
14

ACCEPTED MANUSCRIPT
6.3.16.1-(4-chlorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propoxy)quinoli
n-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (34)
1
Yield: 62%; m.p.: 194.8–195.9°C; H NMR (400 MHz, DMSO-d6) δ 11.91 (s, 1H), 10.08 (s,
1H), 9.16 (s, 1H), 8.60 (d, J = 7.6 Hz, 2H), 8.57 (d, J = 8.1 Hz, 1H), 8.10 (dd, J = 8.8, 6.5 Hz, 1H),
7.65 (d, J = 8.6 Hz, 3H), 7.60 (s, 1H), 7.53 – 7.50 (m, 2H), 7.46 (s, 1H), 7.44 (s, 1H), 6.62 (d, J =
7.9 Hz, 1H), 5.75 (s, 1H), 4.27 (t, J = 8.7 Hz, 2H), 3.97 (s, 2H), 3.48 (d, J = 8.9 Hz, 3H), 3.21 (s,
2H), 2.90 (d, J = 9.8 Hz, 2H), 2.30 (s, 2H), 1.78 – 1.75 (m, 4H), 1.26 (dd, J = 8.3, 6.6 Hz, 3H); 13C
NMR (100 MHz, DMSO-d6) δ 162.41, 160.85, 152.98, 152.67, 152.06, 150.66, 150.17, 146.67,
143.95, 139.45, 137.31, 137.21, 135.70, 133.09, 130.97(2C), 130.81, 129.28(2C), 124.17, 122.44,
119.94, 116.92, 114.75, 114.64, 109.01, 108.79, 105.70, 102.47, 99.50, 66.60, 56.06(2C),
53.33(2C), 51.09, 48.83, 17.95, 16.67; Anal. Calcd. for C37H36ClFN7O5 (%): C, 63.02; H, 4.87; N,
13.54; Found (%): C, 63.03; H, 4.89; N, 13.56.
6.3.17.1-(4-bromophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propoxy)quinoli
n-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (35)
1
Yield: 57%; m.p.: 178.3–179.9°C; H NMR (400 MHz, DMSO-d6) δ 11.95 (s, 1H), 9.15 (s,
1H), 8.54 (d, J = 9.7 Hz, 3H), 8.08 (d, J = 8.9 Hz, 1H), 7.60 – 7.54 (m, 3H), 7.46 (d, J = 7.3 Hz, 3H),
7.40 (d, J = 7.3 Hz, 2H), 6.51 (s, 1H), 4.21 (s, 2H), 3.95 (s, 3H), 3.57 (s, 2H), 3.10 – 2.87 (m, 5H),
2.66 (s, 3H), 2.07 (s, 2H), 1.30 – 1.25 (m, 3H); Anal. Calcd. for C37H36BrFN7O5 (%): C, 61.01; H,
4.72; N, 10.94; Found (%): C, 61.02; H, 4.73; N, 10.95.
6.3.18.1-(3-chloro-4-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propox
y)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (36)
1
Yield: 65%; m.p.: 198.4–199.5°C; H NMR (400 MHz, DMSO-d6) δ 11.90 (s, 1H), 9.16 (s,
1H), 8.83 (d, J = 7.8 Hz, 1H), 8.63 (s, 2H), 8.16 (d, J = 8.5 Hz, 1H), 7.79 (d, J = 8.9 Hz, 3H), 7.66
(d, J = 8.9 Hz, 3H), 7.49 (s, 2H), 6.99 (d, J = 7.1 Hz, 1H), 4.37 (s, 2H), 4.05 (s, 3H), 3.70 (s, 2H),
3.42 (s, 3H), 2.84 (s, 3H), 2.39 (s, 2H), 1.35 – 1.19 (m, 2H), 1.02 (d, J = 7.7 Hz, 2H), 0.84 (s, 1H);
Anal. Calcd. for C38H34ClF2N7O5 (%): C, 61.50; H, 4.62; N, 13.21; Found (%): C, 61.51; H, 4.63; N,
13.22.
6.3.19.1-(4-bromo-2-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propox
y)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (37)
15

ACCEPTED MANUSCRIPT
Yield:76%; m.p.: 215.3–216.6°C; 1H NMR (400 MHz, DMSO-d6) δ 11.58 (s, 1H), 9.06 (d, J =
7.2 Hz, 1H), 8.57 – 8.32 (m, 4H), 7.96 (d, J = 8.6 Hz, 1H), 7.78 (d, J = 8.9 Hz, 1H), 7.57 (d, J = 8.4
Hz, 1H), 7.49 (d, J = 7.8 Hz, 1H), 7.41 (d, J = 6.5 Hz, 1H), 7.37 (d, J = 8.8 Hz, 1H), 7.29 (s, 1H),
6.37 (d, J = 5.2 Hz, 1H), 4.09 (s, 2H), 3.83 (s, 3H), 2.80 – 2.78 (m, 4H), 2.60 (s, 1H), 2.40 (d, J =
9.5 Hz, 7H), 1.92 (d, J = 6.1 Hz, 2H); Anal. Calcd. for C38H34BrF2N7O5 (%): C, 58.02; H, 4.36; N,
12.46; Found (%): C, 58.03; H, 4.37; N, 12.47.
6.3.20.1-(2-chloro-4-(trifluoromethyl)phenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1
-yl)propoxy)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide
(38)
1
Yield: 64%; m.p.: 218.8–220.3°C; H NMR (400 MHz, DMSO-d6) δ 11.89 (s, 1H), 9.18 (s,
1H), 8.79 (d, J = 7.4 Hz, 1H), 8.64 (d, J = 9.2 Hz, 2H), 8.20 – 8.07 (m, 2H), 8.00 (d, J = 8.8 Hz, 1H),
7.84 (d, J = 7.5 Hz, 2H), 7.73 (d, J = 9.4 Hz, 2H), 7.61 (t, J = 8.5 Hz, 1H), 7.50 (d, J = 9.3 Hz, 1H),
6.93 (s, 1H), 4.34 (s, 2H), 4.04 (s, 3H), 3.65 (br, 6H), 3.15 (s, 1H), 2.94 (d, J = 5.9 Hz, 1H), 2.82 (s,
3H), 2.46 (d, J = 6.3 Hz, 1H), 2.36 (s, 2H), 1.35 (s, 1H); Anal. Calcd. for C39H3ClF4N7O5 (%): C,
59.13; H, 4.33; N, 12.38; Found (%): C, 59.14; H, 4.34; N, 12.39.
6.3.21.N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-
1-(4-methoxyphenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (39)
1
Yield: 63%; m.p.: 196.2–198.3°C; H NMR (400 MHz, DMSO-d6) δ 12.00 (s, 1H), 9.13 (s,
1H), 8.61 (d, J = 9.5 Hz, 1H), 8.50 (d, J = 9.3 Hz, 1H), 8.08 (d, J = 8.8 Hz, 1H), 7.62 (d, J = 7.8 Hz,
1H), 7.56 (s, 1H), 7.44 (s, 1H), 7.30 (d, J = 8.7 Hz, 2H), 7.12 (d, J = 8.8 Hz, 2H), 6.52 (d, J = 9.2
Hz, 1H), 5.76 (s, 2H), 4.22 (s, 2H), 3.96 (s, 3H), 3.86 (s, 3H), 2.90 (d, J = 6.3 Hz, 1H), 2.67 (s, 3H),
2.08 (s, 2H), 1.32 – 1.22 (m, 6H), 0.94 (d, J = 5.6 Hz, 1H), 0.85 (t, J = 6.7 Hz, 2H); 13C NMR (100
MHz, DMSO-d6) δ 161.92, 160.25, 158.88, 152.14, 151.15, 150.16, 149.00, 148.10, 145.30, 143.44,
138.93, 136.45, 136.37(2C), 135.69, 135.61, 135.24, 132.58, 130.48, 130.33, 128.77, 123.70,
121.88, 119.64, 119.40, 116.33, 114.15(2C), 108.45, 108.23, 107.94, 101.75, 98.65, 65.61, 55.33,
53.01, 52.72, 51.89(2C), 30.19; Anal. Calcd. for C39H38FN7O6 (%): C, 65.08; H, 5.32; N, 13.62;
Found (%):C, 65.09; H, 5.33; N, 13.63.
6.3.22.N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperazin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-
2-oxo-1-phenyl-1,2-dihydro-1,8-naphthyridine-3-carboxamide (40)
16

ACCEPTED MANUSCRIPT
1
Yield: 62%; m.p.: 244.6–245.8°C; H NMR (400 MHz, DMSO-d6) δ 11.94 (s, 1H), 9.16 (s,
1H), 8.55 (d, J = 9.4 Hz, 3H), 8.08 (d, J = 8.7 Hz, 1H), 7.68 – 7.53 (m, 5H), 7.45 (d, J = 8.6 Hz, 3H),
7.39 (d, J = 6.8 Hz, 2H), 6.50 (s, 1H), 4.26 (s, 2H), 3.96 (s, 3H), 2.50 – 2.39 (m, 5H), 2.28 (s, 2H),
1.79 (d, J = 5.2 Hz, 2H), 1.48 (s, 2H), 1.25 (d, J = 6.0 Hz, 2H), 0.94 (s, 3H); 13C NMR (100 MHz,
DMSO-d6) δ 163.54, 161.93, 160.40, 157.80, 155.33, 154.27, 152.25, 150.63, 150.20, 143.63,
142.66, 139.03, 137.06, 134.58, 133.22, 133.22, 130.88, 129.75(2C), 129.67(2C), 123.65, 121.91,
119.50, 117.17, 116.84, 116.60, 114.54, 114.17, 108.57, 102.42, 99.63, 66.46, 56.00, 52.75,
52.70(2C), 51.36(2C), 40.99, 30.13; Anal. Calcd. for C38H36FN7O5 (%): C, 66.17; H, 5.26; N, 14.22;
Found (%): C, 66.18; H, 5.27; N, 14.23.
6.3.23.N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-1-(4-fluor
ophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (41)
1
Yield: 66%; m.p.: 164.5–165.8°C; H NMR (400 MHz, DMSO-d6) δ 11.91 (s, 1H), 9.16 (s,
1H), 8.61 (s, 2H), 8.47 (d, J = 8.0 Hz, 1H), 8.08 (d, J = 9.7 Hz, 1H), 7.62 (d, J = 8.5 Hz, 1H), 7.52
(s, 1H), 7.46 (dd, J = 9.6, 7.7 Hz, 6H), 7.38 (s, 1H), 6.47 (d, J = 7.8 Hz, 1H), 4.16 (d, J = 5.6 Hz,
2H), 3.94 (s, 3H), 2.42 (t, J = 6.7 Hz, 2H), 2.34 – 2.30 (m, 4H), 2.00 – 1.90 (m, 2H), 1.50 – 1.42 (m,
4H), 1.38 (s, 2H); Anal. Calcd. for C38H34F2N6O5 (%): C, 65.89; H, 4.95; N, 12.13; Found (%): C,
65.90; H, 4.96; N, 12.14.
6.3.24.1-(4-chlorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinolin-4-yl)ox
y)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (42)
1
Yield: 61%; m.p.: 150.3–152.4°C; H NMR (400 MHz, DMSO-d6) δ 11.93 (s, 1H), 9.16 (s,
1H), 8.60 (d, J = 7.9 Hz, 2H), 8.52 (d, J = 6.1 Hz, 1H), 8.09 (d, J = 9.6 Hz, 1H), 7.78 (d, J = 8.5 Hz,
2H), 7.63 (d, J = 7.5 Hz, 1H), 7.57 (d, J = 8.6 Hz, 1H), 7.49 (s, 1H), 7.45 (d, J = 7.8 Hz, 1H), 7.38
(d, J = 8.4 Hz, 2H), 6.56 (s, 1H), 4.22 (s, 2H), 3.95 (s, 3H), 2.97 – 2.94 (m, 5H), 2.71 – 2.63 (m,
5H), 2.31 (s, 1H), 2.04 (s, 3H), 1.30 (s, 1H); Anal. Calcd. for C38H34BrFN6O5 (%): C, 60.56; H, 4.55;
N, 11.15; Found(%): C, 60.57; H, 4.56; N, 11.16.
6.3.25.1-(4-bromophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinolin-4-yl)ox
y)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (43)
1
Yield: 62%; m.p.: 205.8–206.3°C; H NMR (400 MHz, DMSO-d6) δ 11.89 (s, 1H), 10.58 (s,
1H), 9.14 (s, 1H), 8.57 (dd, J = 9.7, 7.3 Hz, 3H), 8.08 (d, J = 7.8 Hz, 1H), 7.79 (d, J = 8.2 Hz, 2H),
17

ACCEPTED MANUSCRIPT
7.64 – 7.56 (m, 2H), 7.48 (d, J = 9.1 Hz, 2H), 7.39 (d, J = 8.2 Hz, 2H), 6.57 (d, J = 4.6 Hz, 1H),
4.27 (s, 2H), 3.97 (s, 3H), 3.11 (d, J = 4.0 Hz, 2H), 2.91 (s, 2H), 2.33 (s, 2H), 1.80 (s, 3H), 1.69 (s,
1H), 1.57 (s, 1H), 1.47 (s, 1H), 1.08 (d, J = 9.0 Hz, 1H), 0.93 (s, 1H); 13C NMR (100 MHz,
DMSO-d6) δ 163.13, 161.35, 159.78, 156.49, 153.32, 152.90, 152.52, 151.35, 150.22, 149.44,
146.99, 144.75, 140.12, 137.49, 134.40, 132.03, 130.86, 130.80, 124.81, 123.06, 120.61, 118.15,
117.93, 117.52, 115.34, 115.17, 109.68, 109.45, 109.24, 102.83, 99.71, 67.24, 66.45, 56.41, 55.29,
53.70, 25.94; Anal. Calcd. for C38H34ClF2N6O5 (%): C, 62.77; H, 4.57; N, 11.56; Found (%): C,
62.78; H, 4.58; N, 11.57.
6.3.26.1-(3-chloro-4-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinoli
n-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (44)
1
Yield: 63%; m.p.: 204.7–205.9°C; H NMR (400 MHz, DMSO-d6) δ 11.94 (s, 1H), 10.79 (s,
1H), 9.13 (s, 1H), 8.61 – 8.57 (m, 2H), 8.54 (d, J = 7.4 Hz, 1H), 8.08 (d, J = 7.8 Hz, 1H), 7.56 (d, J
= 5.3 Hz, 2H), 7.50 (d, J = 7.2 Hz, 2H), 7.37 (d, J = 7.5 Hz, 2H), 6.58 (d, J = 7.0 Hz, 1H), 4.25 (s,
2H), 3.95 (s, 3H), 2.49 – 2.38 (m, 5H), 1.96 (s, 2H), 1.15 (t, J = 7.1 Hz, 4H), 0.90 (s, 3H); Anal.
Calcd. for C38H33BrF2N6O5 (%): C, 59.15; H, 4.31; N, 10.89; Found (%): C, 59.16; H, 4.32; N,
10.90.
6.3.27.1-(4-bromo-2-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinoli
n-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (45)
Yield: 63%; m.p.: 197.3–198.8°C; 1H NMR (400 MHz, DMSO-d6) δ 9.34 (d, J = 8.7 Hz, 1H),
10.80 (s, 1H), 9.19 (d, J = 7.8 Hz, 1H), 8.82 (d, J = 7.5 Hz, 1H), 8.64 (d, J = 8.3 Hz, 2H), 8.16 (d, J
= 9.8 Hz, 1H), 7.89 (d, J = 9.1, 7.5 Hz, 1H), 7.84 (s, 1H), 7.76 (s, 1H), 7.69 (d, J = 7.8 Hz, 1H),
7.59 (d, J = 7.2 Hz, 1H), 7.52 (t, J = 7.9 Hz, 1H), 6.97 (d, J = 6.3 Hz, 1H), 4.34 (t, J = 6.7 Hz, 2H),
4.04 (s, 3H), 3.22 (s, 2H), 2.97 – 2.84 (m, 2H), 2.38 (s, 2H), 1.80 (s, 2H), 1.23 (s, 1H), 1.17 (t, J =
7.1 Hz, 3H), 1.03 (s, 1H), 0.84 (d, J = 8.3 Hz, 1H); Anal. Calcd. for C39H33ClF4N6O5 (%): C, 60.27;
H, 4.28; N, 10.81; Found (%): C, 60.28; H, 4.29; N, 10.82.
6.3.28.1-(2-chloro-4-(trifluoromethyl)phenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propo
xy)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (46)
Yield: 62%; m.p.: 202.3–204.2°C; 1H NMR (400 MHz, DMSO-d6) δ 8.45 (d, J = 7.2 Hz, 1H),
7.51 (s, 1H), 7.37 (s, 1H), 7.07 (t, J = 9.0 Hz, 1H), 6.60 (s, 1H), 6.48 (d, J = 8.6 Hz, 1H), 6.39 (d, J
18

ACCEPTED MANUSCRIPT
= 5.0 Hz, 1H), 5.76 (s, 2H), 5.50 (s, 2H), 4.17 (t, J = 7.2 Hz, 2H), 3.95 (s, 3H), 2.42 (t, J = 7.0 Hz,
13
2H), 2.34 – 1.98 (m, 4H), 2.00 – 1.90 (m, 2H), 1.49 – 1.43 (m, 4H), 1.38 (d, J = 3.7 Hz, 2H); C
NMR (100 MHz, DMSO-d6) δ 162.37, 160.74, 159.18, 154.70, 152.63, 152.25, 151.72, 150.78,
150.62, 149.53, 148.81, 146.26, 143.92, 139.55, 139.42, 136.90, 136.86, 136.80, 136.41, 136.28,
136.21, 132.23, 131.33, 129.20, 128.96, 124.16, 122.43, 121.58, 119.90, 116.82, 114.65, 108.71,
102.21, 99.12, 66.34, 64.50, 55.80, 54.12, 53.24, 52.08, 24.10, 18.60, 17.94, 16.73; Anal. Calcd. for
C39H37FN6O6 (%): C, 66.47; H, 5.29; N, 11.92; Found (%): C, 66.48; H, 5.30; N, 11.93.
6.3.29.N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-1-(4-meth
oxyphenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (47)
1
Yield: 57%; m.p.: 208.9–209.2°C; H NMR (400 MHz, DMSO-d6) δ 12.00 (s, 1H), 9.13 (s,
1H), 8.60 (dd, J = 9.4, 8.7 Hz, 1H), 8.49 (d, J = 7.1 Hz, 1H), 8.08 (d, J = 8.8 Hz, 1H), 7.62 (d, J =
8.9 Hz, 1H), 7.56 (s, 1H), 7.46 (d, J = 8.4 Hz, 2H), 7.30 (d, J = 8.6 Hz, 1H), 7.12 (d, J = 8.2 Hz,
1H), 6.49 (d, J = 4.9 Hz, 1H), 5.76 (s, 3H), 4.26 (s, 1H), 3.96 (s, 3H), 3.86 (s, 3H), 3.59 (d, J = 6.3
Hz, 1H), 3.21 (s, 2H), 3.11 (d, J = 6.7 Hz, 1H), 2.33 (s, 1H), 1.82 (s, 3H), 1.30 – 1.23 (m, 5H), 1.04
(d, J = 6.1 Hz, 1H), 0.85 (s, 1H); Anal. Calcd. for C38H35FN6O6 (%): C, 67.64; H, 5.23; N, 12.46;
Found (%): C, 67.64; H, 5.23; N, 12.46.
6.3.30.N-(3-fluoro-4-((6-methoxy-7-(3-(piperidin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-2-oxo-1-p
henyl-1,2-dihydro-1,8-naphthyridine-3-carboxamide (48)
1
Yield: 59%; m.p.: 225.6–226.7°C; H NMR (400 MHz, DMSO-d6) δ 11.96 (s, 1H), 9.15 (s,
1H), 8.60 (d, J = 5.8 Hz, 2H), 8.53 (d, J = 5.0 Hz, 1H), 8.08 (d, J = 10.6 Hz, 1H), 7.58 (d, J = 5.0
Hz, 3H), 7.49 (t, J = 8.8 Hz, 4H), 7.39 (d, J = 7.6 Hz, 2H), 6.55 (d, J = 4.8 Hz, 1H), 4.27 (s, 2H),
3.97 (s, 3H), 2.90 (d, J = 9.2 Hz, 2H), 2.33 (s, 1H), 1.98 (s, 1H), 1.80 (s, 3H), 1.28 – 1.19 (m, 6H),
1.17 (s, 1H); Anal. Calcd. for C39H37FN6O6 (%): C, 66.47; H, 5.29; N, 11.92; Found (%): C, 66.48;
H, 5.30; N, 11.93.
6.3.31.N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propoxy)quinolin-4-yl)oxy)phenyl)-1
-(4-fluorophenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (49)
1
Yield: 56%; m.p.: 254.5–255.7°C; H NMR (400 MHz, DMSO-d6) δ 11.97 (s, 1H), 10.56 (s,
1H), 9.16 (s, 1H), 8.78 (d, J = 7.9 Hz, 1H), 8.62 (s, 2H), 8.16 (d, J = 9.7 Hz, 1H), 7.75 (d, J = 8.7
Hz, 2H), 7.70 (d, J = 9.0 Hz, 1H), 7.60 (t, J = 8.0 Hz, 1H), 7.47 – 7.42 (m, 3H), 6.93 (d, J = 7.9 Hz,
19

ACCEPTED MANUSCRIPT
1H), 4.33 (s, 2H), 4.03 (s, 3H), 3.22 (s, 3H), 2.93 (d, J = 8.7 Hz, 2H), 2.37 (s, 2H), 1.78 (d, J = 9.3
Hz, 2H), 1.60 – 1.50 (m, 2H), 1.24 (s, 1H), 0.93 (s, 3H), 0.85 (s, 1H); 13C NMR (100 MHz,
DMSO-d6) δ δ 162.31, 161.21, 160.11, 156.85, 155.26, 153.57, 152.82, 152.33, 150.49, 150.18,
149.21, 146.29, 145.04, 140.35, 133.39, 129.08, 124.79(2C), 121.05, 120.55, 120.32, 117.59,
115.36(2C), 115.24, 109.72, 109.49, 108.98, 102.96, 99.83, 70.40, 66.70, 64.03, 56.48, 54.40, 51.93,
23.64, 18.59, 17.28, 13.37; Anal. Calcd. for C39H36F2N6O5 (%): C, 66.28; H, 5.13; N, 11.89; Found
(%): C, 66.29; H, 5.14; N, 11. 90.
6.3.32.1-(4-chlorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propoxy)quinoli
n-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (50)
1
Yield: 59%; m.p.: 268.5–269.9°C; H NMR (400 MHz, DMSO-d6) δ 11.89 (s, 1H), 10.09 (s,
1H), 9.15 (s, 1H), 8.60 (d, J = 7.9 Hz, 2H), 8.48 (d, J = 8.3 Hz, 1H), 8.07 (d, J = 8.9, 1H), 7.66 (s,
1H), 7.64 (s, 1H), 7.61 (d, J = 8.4 Hz, 1H), 7.54 (s, 1H), 7.47 (s, 1H), 7.46 (s, 1H), 7.44 (s, 1H),
6.49 (d, J = 5.1 Hz, 1H), 4.24 (s, 1H), 3.94 (s, 3H), 3.49 (d, J = 10.9 Hz, 2H), 3.20 (s, 2H), 3.10 (dd,
J = 7.5, 6.8 Hz, 1H), 2.92 (s, 2H), 2.29 (s, 2H), 1.78 (d, J = 8.3 Hz, 2H), 1.53 – 1.42 (m, 2H), 1.29
13
(s, 1H), 1.27 (s, 1H), 1.25 (s, 1H), 1.24 (s, 1H), 0.91 (s, 3H); C NMR (100 MHz, DMSO-d6) δ
162.48, 160.78, 159.86, 154.59, 152.14, 151.97, 150.70, 149.66, 148.02, 144.89, 143.83, 139.35,
137.06, 136.80, 129.23(2C), 128.91(2C), 128.75, 128.37, 124.14, 122.23, 119.80, 116.83, 114.73,
114.51, 108.91, 108.71, 107.63, 102.29, 99.26, 66.19, 55.89, 53.45(2C), 53.31, 52.08, 23.10,
22.35(2C), 21.32; Anal. Calcd. for C39H36ClFN6O5 (%): C, 64.77; H, 5.02; N, 11.62; Found (%): C,
64.78; H, 5.03; N, 11.63.
6.3.33.1-(4-bromophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propoxy)quinoli
n-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (51)
Yield: 57%; m.p.: 178.9–179.9°C; 1H NMR (400 MHz, DMSO-d6) δ 11.92 (s, 1H), 9.16 (s,
1H), 8.61 (s, 2H), 8.46 (d, J = 7.5 Hz, 1H), 8.08 (d, J = 9.3 Hz, 1H), 7.63 (d, J = 8.6 Hz, 1H), 7.52
(s, 1H), 7.44
–
7.29 (m, 5H), 7.38 (s, 1H), 6.47 (s, 1H), 4.17 (s, 2H), 3.94 (s, 3H), 2.50 (br, 4H),
1.43 (m, 5H), 1.38 (s, 2H); 13C NMR (100 MHz, DMSO-d6)
2.38 2.24 (m, 6H), 1.95 (s, 2H), 1.50
–
–
δ 163.48, 163.11, 161.33, 161.06, 159.64, 153.16, 152.48, 151.37, 150.12, 149.30, 146.90, 144.44,
139.93, 131.69(2C), 131.57(2C), 124.68, 120.34, 117.37, 116.67, 116.45, 115.19, 114.98, 109.52,
20

ACCEPTED MANUSCRIPT
109.29, 109.01, 102.62, 101.99, 99.54, 67.34, 56.27, 55.58, 54.61, 26.62, 26.12, 24.65; Anal. Calcd.
for C39H36BrFN6O5 (%): C, 61.02; H, 4.73; N, 10.95; Found (%): C, 61.03; H, 4.74; N,10.96.
6.3.34.1-(3-chloro-4-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propox
y)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (52)
Yield: 61%; m.p.: 184.4–185.5°C; 1H NMR (400 MHz, DMSO-d6) δ 11.89 (s, 1H), 10.73 (d, J
= 8.1 Hz, 1H), 9.14 (s, 1H), 8.81 (d, J = 9.3 Hz, 1H), 8.60 (d, J = 8.1 Hz, 2H), 8.19 (s, 1H), 7.81 (s,
1H), 7.73 (d, J = 8.5 Hz, 1H), 7.70 – 7.62 (m, 2H), 7.48 (d, J = 7.3, 2H), 6.96 (d, J = 6.2 Hz, 1H),
4.32 (s, 2H), 4.03 (s, 3H), 1.96 (s, 3H), 1.82 – 1.71 (m, 2H), 1.62 – 1.53 (m, 2H), 1.29 (d, J = 6.5
Hz, 2H), 1.25 (d, J = 6.5 Hz, 2H), 1.15 (s, 2H), 0.91 (s, 3H); Anal. Calcd. for C39H36ClF2N6O5 (%):
C, 63.20; H, 4.76; N, 11.34; Found (%): C, 63.21; H, 4.77; N, 11.35.
6.3.35.1-(4-bromo-2-fluorophenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propox
y)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (53)
1
Yield: 64%; m.p.: 218.2–220.3°C; H NMR (400 MHz, DMSO-d6) δ 11.73 (s, 1H), 10.55 (s,
1H), 9.17 (d, J = 7.4 Hz, 1H), 8.71 (d, J = 8.6 Hz, 1H), 8.63 (d, J = 8.2 Hz, 2H), 8.12 (d, J = 9.7 Hz,
1H), 7.88 (d, J = 9.4 Hz, 1H), 7.67 (d, J = 7.3 Hz, 3H), 7.58 (d, J = 8.3 Hz, 2H), 7.52 (d, J = 6.8 Hz,
1H), 6.83 (d, J = 7.3 Hz, 1H), 4.30 (s, 2H), 4.00 (s, 3H), 3.20 (s, 3H), 2.92 (d, J = 8.2 Hz, 2H), 2.35
(s, 2H), 1.76 (d, J = 4.1 Hz, 2H), 1.68 – 1.44 (m, 3H), 1.21 (s, 1H), 1.05 (s, 1H), 0.91 (d, J = 5.4 Hz,
2H); Anal. Calcd. for C39H36ClF2N6O5 (%): C, 59.62; H, 4.49; N, 10.70; Found (%): C, 59.63; H,
4.50; N, 10.71.
6.3.36.1-(2-chloro-4-(trifluoromethyl)phenyl)-N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1
-yl)propoxy)quinolin-4-yl)oxy)phenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide
(54)
1
Yield: 70%; m.p.: 261.4–262.3°C; H NMR (400 MHz, DMSO-d6) δ 11.80 (s, 1H), 9.20 (s,
1H), 8.64 (d, J = 8.0 Hz, 2H), 8.48 (d, J = 5.2 Hz, 1H), 8.09 (d, J = 9.7 Hz, 2H), 8.01 (d, J = 8.5 Hz,
1H), 7.85 (d, J = 8.3 Hz, 1H), 7.65 (d, J = 8.9 Hz, 1H), 7.55 – 7.47 (m, 3H), 7.39 (s, 1H), 6.48 (d, J
= 5.1 Hz, 1H), 4.19 (t, J = 6.3 Hz, 2H), 3.95 (s, 3H), 2.87 (d, J = 10.1 Hz, 2H), 2.47 (s, 1H), 1.96
(dd, J = 7.2, 6.1 Hz, 4H), 1.59 (d, J = 5.7 Hz, 2H), 1.30 (s, 1H), 1.25 (d, J = 3.1 Hz, 1H), 1.21 –
1.07 (m, 2H), 0.89 (s, 3H); Anal. Calcd. for C40H35ClF4N6O5 (%): C, 60.72; H, 4.46; N, 10.62;
Found (%): C, 60.73; H, 4.47; N, 10.63.
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6.3.37.N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propoxy)quinolin-4-yl)oxy)phnyl)-1-
(4-methoxyphenyl)-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamide (55)
1
Yield: 69%; m.p.: 225.8–226.7°C; H NMR (400 MHz, DMSO-d6) δ 12.02 (s, 1H), 9.89 (s,
1H), 9.16 (s, 1H), 8.61 (t, J = 7.3 Hz, 2H), 8.53 (d, J = 8.3 Hz, 1H), 8.10 (d, J = 9.8 Hz, 1H), 7.64 (d,
J = 8.3 Hz, 1H), 7.59 (s, 1H), 7.49 (t, J = 8.4 Hz, 3H), 7.30 (d, J = 8.7 Hz, 1H), 7.12 (d, J = 8.8 Hz,
1H), 6.56 (d, J = 5.1 Hz, 1H), 4.28 (s, 2H), 3.98 (s, 3H), 3.86 (s, 3H), 3.53 (d, J = 7.2 Hz, 2H), 2.95
(d, J = 6.9 Hz, 2H), 2.31 (s, 2H), 1.81 (d, J = 7.8 Hz, 2H), 1.63 (s, 1H), 1.53 – 1.43 (m, 2H), 1.28 (s,
3H), 0.94 (d, J = 6.3 Hz, 2H); Anal. Calcd. for C40H39FN6O5 (%): C, 66.84; H, 5.47; N, 11.69;
Found (%): C, 66.84; H, 5.47; N, 11.69.
6.3.38.N-(3-fluoro-4-((6-methoxy-7-(3-(4-methylpiperidin-1-yl)propoxy)quinolin-4-yl)oxy)phnyl)-2-
oxo-1-phenyl-1,2-dihydro-1,8-naphthyridine-3-carboxamide (56)
1
Yield: 60%; m.p.: 175.2–177.8°C; H NMR (400 MHz, DMSO-d6) δ 11.94 (s, 1H), 9.16 (s,
1H), 8.55 (d, J = 7.4 Hz, 3H), 8.08 (d, J = 8.7 Hz, 1H), 7.68 – 7.53 (m, 5H), 7.45 (d, J = 9.6 Hz, 3H),
7.39 (d, J = 6.8 Hz, 2H), 6.50 (s, 1H), 4.26 (s, 2H), 3.96 (s, 3H), 2.50 – 2.41 (m, 5H), 2.28 (s, 2H),
1.79 (d, J = 6.2 Hz, 2H), 1.48 (s, 2H), 1.25 (d, J = 8.3 Hz, 2H), 0.94 (s, 3H); Anal. Calcd. for
C39H37FN6O5 (%): C, 68.01; H, 5.41; N, 12.20; Found (%): C, 68.02; H, 5.43; N, 12.21.
6.4 Pharmacology
6.4.1 MTT assay in vitro
The anti-proliferative activities of target compounds 25-56 were evaluated with HepG2,
MCF-7 and A549 cell lines by the standard MTT assay in vitro with c-Met inhibitors, Foretinib as
the positive control. The cancer cell lines were cultured in minimum essential medium (MEM)
supplement with 10% fetal bovine serum (FBS). Approxi-mately 4×103 cells, suspended in MEM
medium, were plated onto each well of a 96-well plate and incubated in 5% CO2 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
5l g/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 [26-27]. All of the target
compounds were tested three times in each of the cell lines. The results expressed as inhibition rates
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or IC50 (half-maximal inhibitory concentration) were the averages of three determinations and
calculated by using the Bacus Laboratories Incorporated Slide Scanner (Bliss) software.
6.4.2 Cell apoptosis assay by flow cytometry
HepG2 cells were seeded in 16-well plates at a density of 1×106 cells/well in RPMI 1640
medium and grown for 24 h at 37 °C in 5% CO2. Then treated with 0.5 µM Foretinib and compound
33 for 48 h. Cultured cells were stained with Annexin V-FITC and propidium iodide (PI) in the
dark at 4°C for 30 min and analyzed by FACS Calibur flow cytometer (Becton Dickinson, San Jose,
CA) using Cell Quest software.
6.4.3. Enzymatic selectivity assays
Through the study of anti-proliferative activity, select some expect compounds (25, 27, 33, 37,
41, 43, 49 and 53) for kinase assay experiments. As previously reported protocol, the c-Met kinase
activities were evaluated using homogeneous time-resolved fluorescence (HTRF) assays [28-29].
All kinase assays were performed in 96-well plates in a 50 µL reaction volume. The kinase buffer
contains 50mM HEPES, pH 7.5, 10mM MgCl2, 0.0015% Brij-35 and 2 mM DTT. The stop buffer
contains100mM HEPES, pH 7.5, 0.015% Brij-35, 0.2% Coating Reagent #3and 50 mM EDTA.
Dilute the compounds to 500µM by 100% DMSO, then transfer 10µL of compound to a new
96-well plate as the intermediate plate, add 90 µL kinase buffer to each well. Transfer 5µL 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 10min. Add 10µL peptide solution to each well. Incubate at 28°C for specified
period of time and stop reaction by 25µL stop buffer. IC50 values were calculated from the
inhibition curves.
7. Acknowledgements
We gratefully acknowledge the generous support provided by National Natural Science
Foundation of China (NSFC No. 81660572；NSFC No. 81803369), Natural Science Foundation of
Jiangxi Province (20171BAB215071), Top-notch talent project of Jiangxi Science & Technology
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Normal University (2016QNBJRC002), and Jiangxi Provincial Key Laboratory of Drug Design and
Evaluation (20171BCD40015).
References
[1] Okwuosa, I. S.; Lewsey, S. C.; Adesiyun, T.; Blumenthal, R. S.; Yancy, C. W., Worldwide
disparities in cardiovascular disease: Challenges and solutions. International Journal of Cardiology
2016, 20(2), 433-440.
[2] Bowry, A. D.; Lewey, J.; Dugani, S. B.; Choudhry, N. K., The Burden of Cardiovascular
Disease in Low- and Middle-Income Countries: Epidemiology and Management. Canadian Journal
of Cardiology 2015, 31 (9), 1151-1159.
[3] Borghi, C.; Verardi, F. M.; Pareo, I.; Bentivenga, C.; Cicero, A. F., Hyperuricemia and
cardiovascular disease risk. Expert Review of Cardiovascular Therapy 2014, 12 (10), 1219-1225.
[4]Rosã, s. E.; Santomã, P.; Duran-Frigola, M.; Hernandez, B.; Llinà s, M. C.; Ruiz-Gonzã, l. R.;
Nonell, S.; Sã, n.-G. a. D.; Edelman, E. R.; Balcells, M., Modifications of microvascular EC surface
modulate phototoxicity of a porphycene anti-ICAM-1 immunoconjugate; therapeutic implications.
Langmuir the Acs Journal of Surfaces & Colloids 2013, 29 (31), 9734-9743.
[5] Pasero, C.; Speiser, D. E.; Derré, L.; Olive, D., The HVEM network: new directions in
targeting novel costimulatory/co-inhibitory molecules for cancer therapy. Current Opinion in
Pharmacology 2012, 12 (4), 478-485.
[6] Anum, M.; Muhammad, A. H.; Asfandyar, S.; Hammad, K. A. M.; Shafaq, S., c-Met inhibitors.
Infectious Agents & Cancer 2013, 8 (1), 13-21.
[7] Bladt, F.; Faden, B.; Friese-Hamim, M.; Knuehl, C.; Wilm, C.; Fittschen, C.; Grã¤Dler, U.;
Meyring, M.; Dorsch, D.; Jaehrling, F., EMD 1214063 and EMD 1204831 constitute a new class of
potent and highly selective c-Met inhibitors. Clinical Cancer Research 2013, 19 (11), 2941-2951.
[8] Varkaris, A.; Corn, P. G.; Gaur, S.; Dayyani, F.; Logothetis, C. J.; Gallick, G. E., The Role of
HGF/c-Met Signaling in Prostate Cancer Progression and c-Met Inhibitors in Clinical Trials. Expert
Opinion on Investigational Drugs 2011, 20 (12), 1677-1681.
[9] Clavijo-Cornejo, D.; Enriquez-Cortina, C.; López-Reyes, A.; Domínguez-Pérez, M.; Nuño, N.;
Domínguez-Meraz, M.; Bucio, L.; Souza, V.; Factor, V. M.; Thorgeirsson, S. S., Biphasic
regulation of the NADPH oxidase by HGF/c-Met signaling pathway in primary mouse hepatocytes.
Biochimie 2013, 95 (6), 1177-1184.
[10]Papa, E.; Weller, M.; Weiss, T.; Ventura, E.; Burghardt, I.; Szabó, E., Negative control of the
HGF/c-MET pathway by TGF-β: a new look at the regulation of stemness in glioblastoma. Cell
Death & Disease 2017, 8 (12), 3210-3221.
[11]Yakes, F. M.; Chen, J.; Tan, J.; Yamaguchi, K.; Shi, Y.; Yu, P.; Qian, F.; Chu, F.; Bentzien, F.;
Cancilla, B., Cabozantinib (XL184), a Novel MET and VEGFR2 Inhibitor, Simultaneously
Suppresses Metastasis, Angiogenesis, and Tumor Growth. Molecular Cancer Therapeutics 2011,
10 (12), 2298-2311.
[12]Yu, K.; Mukohara, T.; Tomioka, H.; Funakoshi, Y.; Kiyota, N.; Fujiwara, Y.; Yashiro, M.;
Hirakawa, K.; Hirai, M.; Minami, H., Foretinib (GSK1363089), a multi-kinase inhibitor of MET
and VEGFRs, inhibits growth of gastric cancer cell lines by blocking inter-receptor tyrosine kinase
networks. Investigational New Drugs 2012, 30 (4), 1352-1360.
[13]Underiner, T. L.; Herbertz, T.; Miknyoczki, S. J., Discovery of small molecule c-Met
inhibitors: Evolution and profiles of clinical candidates. Anticancer Agents Med Chem 2010, 10
(1),176-185.
24

ACCEPTED MANUSCRIPT
[14]Norman, M. H.; Liu, L.; Lee, M.; Xi, N.; Fellows, I.; D'Angelo, N. D.; Dominguez, C.; Rex, K.;
Bellon, S. F.; Kim, T. S., Structure-based design of novel class II c-Met inhibitors: 1. Identification
of pyrazolone-based derivatives. Journal of Medicinal Chemistry 2012, 55 (5), 1858-1867.
[15]Parikh, P. K.; Ghate, M. D., Recent advances in the discovery of small molecule c-Met Kinase
inhibitors. European Journal of Medicinal Chemistry 2017, 14 (3), 134-145.
[16]Tang, Q.; Xin, Z.; Tu, Y.; Ping, W.; Wang, L.; Wu, C.; Wang, W.; Xie, H.; Ping, G.; Zheng, P.,
Synthesis and antiproliferative activity of 6,7-disubstituted-4-phenoxyquinoline derivatives bearing
the 2-oxo-4-chloro-1,2-dihydroquinoline-3-carboxamide moiety. Bioorganic
Chemistry Letters 2016, 26 (7), 1794-1798.
&
Medicinal
[17]Huang, Y.; Lu, S. F.; Hu, C. J.; Fu, S. P.; Shen, W. X.; Liu, W. X.; Li, Q.; Wang, N.; He, S. Y.;
Liang, F. R., Electro-acupuncture at Neiguan pretreatment alters genome-wide gene expressions
and protects rat myocardium against ischemia-reperfusion. Molecules 2014, 19 (10), 1658-1678.
[18]Kuroda, T.; Suzuki, F.; Tamura, T.; Ohmori, K.; Hosoe, H., A novel synthesis and potent
antiinflammatory activity of 4-hydroxy-2(1H)-oxo-1-phenyl-1,8-naphthyridine-3-carboxamides.
Journal of Medicinal Chemistry 1992, 35 (6), 1130-1146.
[19]Tang, Q.; Zhao, Y.; Du, X.; Chong, L. E.; Gong, P.; Guo, C., Design, synthesis, and
structure–activity relationships of novel 6,7-disubstituted-4-phenoxyquinoline derivatives as
potential antitumor agents. European Journal of Medicinal Chemistry 2013, 69 (7), 77-89.
[20]Wang, L.; Xu, S.; Chen, X.; Liu, X.; Duan, Y.; Kong, D.; Zhao, D.; Zheng, P.; Tang, Q.; Zhu,
W., Synthesis and bioevaluation study of novel N -methylpicolinamide and thienopyrimidine
derivatives as selectivity c-Met kinase inhibitors. Bioorganic & Medicinal Chemistry 2017, 26 (1),
245-256.
[21]Sun, Z. H.; Bu, P., Downregulation of phosphatase of regenerating liver-3 is involved in the
inhibition of proliferation and apoptosis induced by emodin in the SGC-7901 human gastric
carcinoma cell line. Experimental & Therapeutic Medicine 2012, 3 (6), 1077-1081.
[22]Chaudhry, C.; Davis, J.; Zhang, Y.; Posy, S.; Lei, M.; Shen, H.; Yan, C.; Devaux, B.; Zhang,
L.; Blat, Y., Building homogeneous time-resolved fluorescence resonance energy transfer assays for
characterization of bivalent inhibitors of an inhibitor of apoptosis protein target. Analytical
Biochemistry 2016, 49 (7), 8-17.
[23]Yuan, H.; Liu, Q.; Zhang, L.; Hu, S.; Chen, T.; Li, H.; Chen, Y.; Xu, Y.; Lu, T., Discovery,
optimization and biological evaluation for novel c-Met kinase inhibitors. European Journal of
Medicinal Chemistry 2018, 14(3), 491-502.
[24]Sarma, R.; Sinha, S.; Ravikumar, M.; Kishore, K. M.; Mahmood, S. K., Pharmacophore
modeling of diverse classes of p38 MAP kinase inhibitors. European Journal of Medicinal
Chemistry 2008, 43 (12), 2870-2876.
[25]Burbridge, M. F.; Bossard, C. J.; Saunier, C.; Fejes, I.; Bruno, A.; Léonce, S.; Ferry, G.; Da, V.
G.; Bouzom, F.; Cattan, V., S49076 is a novel kinase inhibitor of MET, AXL, and FGFR with
strong preclinical activity alone and in association with bevacizumab. Molecular Cancer
Therapeutics 2013, 12 (9), 1749-1762.
[26]Yu, X.; Zhao, X.; Zhu, L.; Zou, C.; Liu, X.; Zhao, Z.; Huang, J.; Li, H., Discovery of novel
inhibitors for human farnesyltransferase (hFTase) via structure-based virtual screening.
Medchemcomm 2013, 4 (6), 962-971.
[27]Zhai, X.; Zhao, Y. F.; Liu, Y. J.; Zhang, Y.; Xun, F. Q.; Liu, J.; Gong, P., ChemInform
Abstract: Synthesis and Cytotoxicity Studies of Novel [1,2,4]Triazolo[1,5-a]pyrimidine-7-amines.
Chemical & Pharmaceutical Bulletin 2008, 39 (5), 941-945.
25

ACCEPTED MANUSCRIPT
[28]Jia, Y.; Quinn, C. M.; Gagnon, A. I.; Talanian, R., Homogeneous time-resolved fluorescence
and its applications for kinase assays in drug discovery. Analytical Biochemistry 2006, 56 (2),
273-281.
[29]Clarke, E. E.; Shearman, M. S., Quantitation of amyloid-beta peptides in biological milieu
using a novel homogeneous time-resolved fluorescence (HTRF) assay. Journal of Neuroscience
Methods 2000, 102 (31), 61-68.
Legends
Fig. 1 The representative small-molecule c-Met kinase inhibitors.
Fig. 2 The docking mode of Foretinib with c-Met protein.
Fig. 3. Our previous work on antiproliferative agents (7 and 8) bearing quinoline scaffolds and
potent drugs (9) bearing 1,8-naphthyridin-2-one.
Fig. 4 Design strategy for the quinoline derivatives bearing1,8-naphthyridin-2-one moiety
Fig. 5 The relationship between different concentrations of compound 33 and cytotoxicity
Fig. 6 Morphologic changes of HepG2 cells under inverted microscopy and fluorescence
microscopy. (6a) The control group cell treated with nothing; (6b) Experimental group treated with
1.1 µM concentration of compound 33.
Fig. 7 Representative dot plot of Annexin V (x-axis) versus PI (y-axis) analyses. Q2-1: Mechanical
damage; Q2-2: Early apoptotic; Q2-3: Living cells; Q2-4: Late apoptotic and death.
Fig. 8 The 3D interaction map between the 33 and c-Met, the hydrogen bonds was colored with red
and the pi-pi bonds was colored with black, respectively (7a). 2D interaction map between the 33
and c-Met, the hydrogen bonds and the pi-pi bonds was colored light blue (7b)
Scheme 1. Reagents and conditions: (i) Br(CH2)3Cl, acetone, 0°C,30 min, 12 h; (ii) 98% HNO3,
CH2Cl2, 0°C, 4 h; (iii) DMF–DMA, toluene, 110°C, 10 h; (iv) Fe powder, AcOH, 30 min, 80°C, 2 h;
(v) secondary amines, CH3CN, 85°C, 10 h; (vi) POCl3, 85°C, 6 h; (vii) 2-fluoro-4-nitrophenol, PhCl,
140°C, 30 h; (viii) Fe powder, NH4Cl (cat.), EtOH/H2O, reflux, 5 h.
Scheme 2. Reagents and conditions: (i) corresponding aniline, CH3COOH, 100°C,5 h; (ii) THF,
AlLiH4, N2 ,6 h; (iii) CH2Cl2, PCC, 0°C, 6 h. (iv) CH2(COOC2H5)2, C2H5OH, 110°C 34 h; (v)
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K2CO3, 1,4-Dioxane, H2O, 8 h (vi) SOCl2, reflux, 1 h; (vii) corresponding aniline, DIPEA, CH2Cl2,
rt, 1 h.
Table 1
Cytotoxic activities of compounds 25-56 against HepG2, MCF-7, A549 cell lines in vitro
Table 2
c-Met kinase activity of selected compounds 25, 27, 33, 37, 41, 43, 49, 53 and Foretinib in vitro
Table 3
Inhibition of tyrosine kinases by compound 33
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Table 1
Cytotoxic activities of compounds 25-56 against HepG2, MCF-7, A549 cell lines in vitro
IC50 a (µM)
R1
R2
Compd.
HepG2
0.76 ± 0.02b
NAc
MCF-7
1.78 ± 0.34
3.21 ± 1.12
0.61 ± 0.38
1.12 ± 0.56
9.75 ± 1.02
9.41 ± 0.97
10.49 ± 0.89
NA
A549
2.41 ± 0.08
4.23 ± 0.21
4.17 ± 0.32
NA
4-F
25
4-Cl
26
4-Br
1.98 ± 0.18
9.68 ± 1.08
NA
27
3-Cl-4-F
4-Br-2-F
2-Cl-4-CF3
4-OCH3
4-H
28
NA
29
4.17 ± 0.87
9.16 ± 0.09
11.6 ± 2.71
0.23 + 0.01
2.35 ± 0.92
9.49 ± 2.15
6.36 ± 1.09
0.36 ± 0.01
3.98 ± 1.04
7.92 ± 2.31
3.23 ± 0.84
0.33 ± 0.01
1.49 ± 0.41
0.46 ± 0.12
7.14 ± 1.69
NA
4.19 ± 1.01
19.47 ± 1.98
16.76 ± 2.21
0.21 ± 0.02
NA
30
31
32
4-F
33
0.42 ± 0.06
25.21 ± 0.74
10.62 ± 1.76
12.32 ± 1.09
2.49 ± 0.87
4.34 ± 0.58
6.45 ± 0.93
2.62 ± 0.64
0.45 ± 0.01
1.83 ± 0.89
NA
4-Cl
34
4-Br
15.83 ± 1.75
19.76 ± 1.41
0.29 ± 0.01
9.13 ± 0.76
1.28 ± 0.51
NA
35
3-Cl-4-F
4-Br-2-F
2-Cl-4-CF3
4-OCH3
4-H
36
37
38
39
40
4-F
1.26 ± 0.32
1.49 ± 0.21
0.21 ± 0.13
NA
41
4-Cl
42
4-Br
43
3-Cl-4-F
4-Br-2-F
2-Cl-4-CF3
4-OCH3
4-H
5.60 ± 1.31
12.21 ± 1.73
NA
44
18.98 ± 2.15
NA
45
9.01 ± 2.51
9.45 ± 2.13
6.25 ± 0.85
0.84 ± 0.37
4.86 ± 0.51
3.63 ± 0.91
2.39 ± 0.72
1.56 ± 0.82
4.92 ± 0.98
9.47 ± 1.23
NA
46
9.18 ± 0.98
3.27 ± 1.23
0.68 ± 0.04
15.87 ± 1.15
3.47 ± 0.65
3.92 ± 1.12
0.61 ± 0.02
0.71 ± 0.01
8.41 ± 1.36
7.98 ± 1.08
0.76 ± 0.01
8.11 ± 1.12
11.93 ± 2.34
0.31 ± 0.01
9.12 ± 1.12
2.71 ± 0.98
1.01 ± 0.75
NA
47
48
4-F
49
4-Cl
50
4-Br
51
3-Cl-4-F
4-Br-2-F
2-Cl-4-CF3
4-OCH3
4-H
52
53
15.71 ± 2.65
11.49 ± 1.53
9.07 ± 1.36
0.32 ± 0.03
54
55
56
Foretinibd
0.48 ± 0.07
a The half maximal inhibitory concentration (IC50) values are an average of three separate determinations
b Bold values show the IC50 values of target compounds lower than the values of the positive control
c NA: No Activity (IC50 > 40 µM)
d Used as a positive control