Design, synthesis, antiproliferative activity and docking studies of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline as potential EGFR inhibitors

Article abstract of DOI:10.1016/j.ejmech.2018.05.006

Eight series of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline were designed, synthesized and evaluated for the IC₅₀ values against three cancer cell lines (A549, MCF-7 and PC-3). Most of the forty nine target compounds showed excellent antiproliferative activity against one or several cancer cell lines. The compound 13a showed the best activity against A549, MCF-7 and PC-3 cancer cell lines, with the IC₅₀ values of 1.09 ± 0.04 μM, 1.34 ± 0.13 μM and 1.23 ± 0.09 μM, respectively. Eight selected compounds were further selected to evaluated for the inhibitory activity against EGFR kinase. Three of them showed equal activity against EGFR kinase to positive control afatinib. AnnexinV-FITC, propidium iodide (PI) double staining and acridine orange single staining results indicated that the compound 13a could induce apoptosis of human lung cancer A549 cells.

Full text of DOI:10.1016/j.ejmech.2018.05.006

Accepted Manuscript
Design, synthesis, antiproliferative activity and docking studies of quinazoline
derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline as potential
EGFR inhibitors
Yiqiang OuYang, Wensheng Zou, Liang Peng, Zunhua Yang, Qidong Tang, Mengzi
Chen, Shuang Jia, Hong Zhang, Zhou Lan, Pengwu Zheng, Wufu Zhu
PII:
S0223-5234(18)30410-0
10.1016/j.ejmech.2018.05.006
EJMECH 10416
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 24 January 2018
Revised Date: 16 April 2018
Accepted Date: 5 May 2018
Please cite this article as: Y. OuYang, W. Zou, L. Peng, Z. Yang, Q. Tang, M. Chen, S. Jia, H. Zhang,
Z. Lan, P. Zheng, W. Zhu, Design, synthesis, antiproliferative activity and docking studies of quinazoline
derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline as potential EGFR inhibitors,
European Journal of Medicinal Chemistry (2018), doi: 10.1016/j.ejmech.2018.05.006.
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Graphical abstract

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Design, synthesis, antiproliferative activity and docking studies of quinazoline derivatives bearing
2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline as potential EGFR inhibitors
Yiqiang OuYang1,†, Wensheng Zou1,†, Liang Peng1, Zunhua Yang2, Qidong Tang1, Mengzi Chen1, Shuang Jia1, Hong
Zhang1, Zhou Lan1, Pengwu Zheng1,*, Wufu Zhu1,*
1 Jiangxi Provincial Key Laboratory of Drug Design and Evaluation, School of Pharmacy, Jiangxi Science & Technology
Normal University, Nanchang 330013
2College of Pharmacy, Jiangxi University of Traditional Chinese Medicine, Nanchang 330004, China
*Correspondence author.
Tel/Fax: +86 791 8380-2393; E-mail address: zhengpw@126.com (P. Zheng), zhuwufu-1122@163.com (W. Zhu)
Abstract: Eight series of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline were
designed, synthesized and evaluated for the IC50 values against three cancer cell lines (A549, MCF-7 and PC-3). Most of
the forty nine target compounds showed excellent antiproliferative activity against one or several cancer cell lines. The
compound 13a showed the best activity against A549, MCF-7 and PC-3 cancer cell lines, with the IC50 values of 1.09 ±
0.04 µM, 1.34 ± 0.13 µM and 1.23 ± 0.09 µM, respectively. Eight selected compounds were further selected to evaluated
for the inhibitory activity against EGFR kinase. Three of them showed equal activity against EGFR kinase to positive
control afatinib. AnnexinV-FITC, propidium iodide (PI) double staining and acridine orange single staining results
indicated that the compound 13a could induce apoptosis of human lung cancer A549 cells.
Key words: Quinazoline derivatives; Synthesis; Antiproliferative activity; EGFR inhibitors
† These authors contribute equally to this work
1
Corresponding author. Tel./fax: +86 791 83802393.
E-mail address: zhengpw@126.com (P. Zheng), zhuwf@jxstnu.edu.cn, zhuwufu-1122@163.com (W. Zhu).

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1 Introduction
Cancer is still one of the most serious diseases of humans. At present, statistics show that approximately 14 million
new cancer patients are treated globally, and 8.2 million people died from cancer [1]. Many signaling pathways were
involved in the development of cancer, among them the Epidermal Growth Factor Receptor (EGFR) pathway plays a
crucial role in the apoptosis, proliferation, differentiation, migration and cell cycle of cancer cells [2-4]. Therefore,
designing small molecule inhibitors targeting EGFR signaling pathway can effectively treat cancer further.
In recent years, many quinazoline derivatives were reported as EGFR inhibitors such as gefitinib (1) [5], canertinib
(2) [6] and afatinib (3) [7] (Fig. 1). Afatinib, a second-generation epidermal growth factor receptor (EGFR) tyrosine kinase
inhibitor, has been approved worldwide as a first-line treatment for advanced non-small cell lung cancer (NSCLC) that
harbors activating EGFR mutations. The structure-activity relationships studies of these compounds revealed that the
quinazoline skeleton plays a key role in their antitumor activities, especially 4-anilinoquinazoline core (4). Therefore, the
quinazoline skeleton was kept as a privileged scaffold, and the C-4 position of quinazoline derivatives was substituted by
aniline structures. In addition, in order to investigate the effect of N-H at C-4 position of quinazoline on the
antiproliferative activity, a five-atom or six-atom liposoluble ring structure between the N-H and C-2 position of phenyl
group were constructed resulting in the 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline unit, just as shown in Fig 2.
Therefore, several series of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline were
achieved.
Furthermore, it was reported that most of quinazoline derivatives rely on an acrylamide electrophilic warhead which
can form a covalent bond with a conserved cysteine residue in EGFR and enable a great occupancy of the ATP binding
site [8-9]. Inspired by canertinib and gefitinib, we decided to introduce acrylamide group into the target compounds and the
tetrahydrofuran unit was replaced with methyl group. Therefore, we designed and synthesized four series of compounds
(8a–g, 9a–g, 10a–e and 11a–e). However, these compounds (8a–g, 9a–g, 10a–e and 11a–e) show unsatisfactory
solubility which affects the pharmacokinetics in vitro. In order to increase the water-soluble and inspired by afatinib, the
small molecule dimethylamine was substituted with different small molecule secondary amines to obtain the other four
series (12a–g, 13a–f, 14a–f and 15a–f). The design strategy for all target compounds is shown in Fig. 2.
Hence we disclosed the design, synthesis and antiproliferative activity of novel quinazoline analogues against EGFR
overexpression cancer cells A549 (human lung cancer), MCF-7 (human breast cancer) and PC-3 (human prostate cancer)
[10-12]
and as well as the inhibitory on EGFR kinase. Moreover, flow cytometry, acridine orange single staining and
docking studies were presented in this paper as well.

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Fig. 1. Structures of small-molecule EGFR inhibitors based on the quinazoline scaffold and the target compounds 8a–g,
9a–g, 10a–e, 11a–e, 12a–g, 13a–f, 14a–f and 15a–f.
Fig. 2. Structures and design strategy for target compounds 8a–g, 9a–g, 10a–e, 11a–e, 12a–g, 13a–f, 14a–f and 15a–f.
2 Results and discussion
2.1 Chemistry
The structures and preparation of target compounds 8a–g, 9a–g, 10a–e, 11a–e, 12a–g, 13a–f, 14a–f and 15a–f are
described in Schemes 1-2. These target compounds were synthesized from commercially available
2-amino-4-fluorobenzoic acid through seven steps or eight steps, respectively. Cyclization reaction of
2-amino-4-fluorobenzoic acid in the present of formamidine acetate obtained compound 1 which was a white solid.
Subsequently, compound 1 was nitrified with concentrated sulfuric acid and fuming nitric acid refluxed for 2 h to obtain
compound 2. Compound 2 was chlorinated with SOCl2 and DMF for 4 h at 90 °C to obtain compound 3. Nucleophilic
displacement of the compound 3 with 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline gave access to intermediates 4a
and 4b, respectively. Compound 4a or 4b was treated with methanol, 50% sodium hydroxide as base catalyst to furnish

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5a and 5c, respectively. Compound 4a or 4b was condensed with (S)-tetrahydrofuran-3-ol in the present of 60% NaH
obtaining 5b and 5d, respectively. Then compounds 5a–d were reduced by NH2NH2•H2O (80%), FeCl3 and activated
carbon to generate key intermediates 6a–d, respectively. Finally, intermediates 6a–d were condensed with the
corresponding acrylic acid derivatives (Acrylic acid, Methacrylate, 3,3-Dimethacrylic acid, Trans-2-butenoic acid,
Trans-2-pentenoic acid, 4-methyl-2-pentenoic acid, Trans-2-hexenoic acid) to get target compounds 8a–g, 9a–g, 10a–e
and 11a–e, respectively. Intermediates 6a–d were condensed with diethylphosphonoacetic acid to get intermediates 7a–d,
respectively. Finally, intermediates 7a–d were condensed with the corresponding aldehydes (B1-B7) to get target
compounds 12a–g, 13a–f, 14a–f and 15a–f, respectively.
Scheme 1. Synthetic route of target compounds 8a–g, 9a–g, 10a–e and 11a–e.
Reagents and conditions: (a) ethanol, 24 h; (b) Con.H2SO4, fuming HNO3, 2 h; (c) SOCl2, DMF, 4 h; (d)
2,3-dihydroindole or 1,2,3,4-tetrahydroindole, isopropanol, triethylamine, 1.5 h; (e) CH3OH, 50%NaOH 3 h;
(S)-tetrahydrofuran-3-ol, 60%NaH, THF; (f) 80% hydrazine monohydrate, FeCl3, activated carbon, ethanol, 1 h; (g)
oxalyl chloride, DIPEA, CH2Cl2, 1 h.
Scheme 2. Synthetic route of target compounds 12a–g, 13a–f, 14a–f and 15a–f.
Reagents and conditions: (a) N,N'-Carbonyldiimidazole, THF, 2 h; (b) K2CO3, 1,4-dioxane, 4 h; (c) conc HCl, 4 h;

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(d)50%NaOH, THF, 1 h;
2.2 Biological evaluation
All the target compounds (8a–g, 9a–g, 10a–e, 11a–e, 12a–g, 13a–f, 14a–f and 15a–f) were tested for
antiproliferative activity against EGFR overexpression cancer cells A549, MCF-7 and PC-3 cancer cell lines through
3-(4,5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium Bromide (MTT) assay, using EGFR inhibitor afatinib as positive
control. In addition, some selected compounds were evaluated for the IC50 values against EGFR kinase by the mobility
shift assay, together with reference compound afatinib. The results expressed as IC50 values were summarized in Tables
1–2 and the values are the average of at least two independent experiments.
Table 1. Structures and activity of target compounds 8a–g, 9a–g, 10a–e and 11a–e.
a
IC50 (µM)
Compounds
No.
n
R1
R2
A549
MCF-7
PC-3
1
1
1
1
1
1
1
1
1
1
1
1
1
1
2
2
2
-CH3
-CH3
-CH3
-CH3
-CH3
-CH3
-CH3
-CH=CH2
5.58 ± 0.62
7.32 ± 0.86
6.27 ± 0.72
8.29 ± 0.58
2.47 ± 0.31
6.41 ± 0.51
2.92 ± 0.36
9.32 ± 0.97
16.58 ± 1.22
3.67 ± 0.56
2.61 ± 0.32
10.92 ± 1.14
8.52 ± 0.59
9.11 ± 0.79
5.41 ± 0.63
8.19 ± 0.91
5.03 ± 0.60
11.68 ± 1.06
11.11 ± 1.05
6.39 ± 0.76
4.03 ± 0.61
4.03 ± 0.38
6.78 ± 0.79
3.16 ± 0.34
5.95 ± 0.77
14.68 ± 1.16
15.67 ± 1.12
4.90 ± 0.51
8.84 ± 0.95
9.88 ± 0.91
4.55 ± 0.66
6.87 ± 0.81
12.66 ± 1.10
2.87 ± 0.36
5.94 ± 0.75
4.83 ± 0.68
7.84 ± 0.81
8.12 ± 0.91
5.89 ± 0.48
6.53 ± 0.52
4.24 ± 0.52
2.71 ± 0.43
4.91 ± 0.69
4.62 ± 0.51
5.22 ± 0.48
9.59 ± 1.04
11.23 ± 0.98
9.23 ± 0.88
6.13 ± 0.72
7.52 ± 0.85
6.31 ± 0.57
8a
8b
8c
-C(CH3)=CH2
-CH=C(CH3)2
-CH=CHCH3
8d
8e
-CH=CHCH2CH3
-CH=CHCH(CH3)2
-CH=CHCH2CH2CH3
-CH=CH2
8f
8g
9a
9b
9c
-C(CH3)=CH2
-CH=C(CH3)2
-CH=CHCH3
9d
9e
-CH=CHCH2CH3
-CH=CHCH(CH3)2
-CH=CHCH2CH2CH3
-CH=C(CH3)2
9f
9g
10a
10b
10c
-CH3
-CH3
-CH3
-CH=CHCH3
-CH=CHCH2CH3

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2
2
2
2
2
2
2
-CH3
-CH3
-CH=CHCH(CH3)2
-CH=CHCH2CH2CH3
-CH=C(CH3)2
3.45 ± 0.42
1.80 ± 0.15
7.91 ± 0.89
10.77 ± 1.03
14.09 ± 1.31
6.82 ± 0.51
7.32 ± 0.62
0.71 ± 0.05
4.29 ± 0.51
1.21 ± 0.08
11.14 ± 0.98
5.93 ± 0.67
14.37 ± 1.25
4.68 ± 0.68
4.23 ± 0.62
0.93 ± 0.09
8.43 ± 0.79
2.24 ± 0.35
9.02 ± 0.98
9.35 ± 0.99
14.29 ± 1.38
8.51 ± 0.93
11.84 ± 1.06
2.51 ± 0.18
10d
10e
11a
-CH=CHCH3
11b
-CH=CHCH2CH3
-CH=CHCH(CH3)2
-CH=CHCH2CH2CH3
11c
11d
11e
Afatinibb
The bold values showed excellent antiproliferative activity (1.21 - 2.92 µM)
a The values are an average of two separate determinations
b Used as a positive controls
Table 2. Structures and activity of target compounds 12a–g, 13a–f, 14a–f and 15a–f.
IC50a(µM)
Compounds
No.
n
R
A549
MCF-7
PC-3
1
1
1
1
1
1
1
-CH3
-CH3
-CH3
-CH3
-CH3
-CH3
-CH3
3.37 ± 0.53
1.99 ± 0.21
1.51 ± 0.18
1.68 ± 0.22
6.43 ± 0.81
8.45±0.93
2.62 ± 0.31
2.19 ± 0.16
3.69 ± 0.37
2.62 ± 0.42
4.18 ± 0.62
9.18 ± 0.96
8.29 ± 0.92
2.65 ± 0.27
12a
12b
12c
12d
12e
12f
2.04 ± 0.25
1.69 ± 0.22
1.21 ± 0.09
2.54 ± 0.41
5.36 ± 0.73
9.25 ± 1.05
1.98 ± 0.21
12g
1
1
1
1
1
1
2
2
2
13a
13b
13c
13d
13e
13f
1.09 ± 0.04
5.22 ± 0.72
2.23 ± 0.35
4.66 ± 0.67
6.09 ± 0.79
3.67 ± 0.56
1.65 ± 0.21
6.41 ± 0.81
2.23 ± 0.35
1.34 ± 0.13
4.98 ± 0.70
2.31 ± 0.36
1.89 ± 0.25
5.35 ± 0.73
4.69 ± 0.57
1.77 ± 0.24
5.31 ± 0.73
3.19 ± 0.51
1.23 ± 0.09
5.78 ± 0.76
1.51 ± 0.18
4.51 ± 0.65
4.21 ± 0.63
3.31 ± 0.52
1.75 ± 0.24
6.89 ± 0.84
1.74 ± 0.24
-CH3
-CH3
-CH3
14a
14b
14c

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2
2
2
2
2
2
2
2
2
-CH3
-CH3
-CH3
3.93 ± 0.59
8.56 ± 0.81
5.74 ± 0.76
2.85 ± 0.45
3.87 ± 0.49
4.25 ± 0.52
4.45 ± 0.55
7.18 ± 0.85
4.96 ± 0.59
0.93 ± 0.09
5.78 ± 0.76
6.26 ± 0.82
7.15 ± 0.79
4.17 ± 0.62
4.27 ± 0.53
5.78 ± 0.61
3.63 ± 0.41
9.45 ± 0.99
5.46 ± 0.62
2.51 ± 0.18
14d
14e
2.88 ± 0.36
7.83 ± 0.89
6.97 ± 0.74
2.32 ± 0.32
5.65 ± 0.75
6.51 ± 0.81
3.67 ± 0.46
5.87 ± 0.55
5.30 ± 0.72
0.71 ± 0.05
14f
15a
15b
15c
15d
15e
15f
Afatinibb
The bold values showed excellent antiproliferative activity (1.09 - 2.85 µM)
a The values are an average of two separate determinations
b Used as a positive controls
Fig. 3 Statistical results of the cell proliferation inhibition in A549, MCF-7 and PC-3 cells treated with different
concentrations of target compound 13a.
Table 3 EGFRWT and EGFRT790M kinase inhibitory activity of compounds 8e, 9d, 10e, 12c, 12d, 12g, 13a, 14a and
afatinib.
IC50 (nM)
Compounds No.
EGFRWT
17
EGFRT790M
186
8e

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29
33
3
192
9d
10e
162
34
12c
45.4
4
115
21
12d
12g
5
26
13a
72
5
107
7
14a
Afatiniba
a Used as a positive controls
As illustrated in Tables 1-2, most of the forty nine target compounds showed excellent antiproliferative activity
against the different tested cancer cells. In general, the antiproliferative activity of target compounds against A549 cell
line were more potent than against MCF-7 and PC-3 cell lines. What’s more, we could easily see that the four series of
compounds 12a–g, 13a–f, 14a–f and 15a–f were much more active than the other series of compounds 8a–g, 9a–g,
10a–e and 11a–e against the three cancer cell lines. The results suggested that the compounds substituted with secondary
amines were beneficial to the antiproliferative activity. We hypothesized that secondary amines may contribute to the
increasing of water-soluble and high binding affinity. On the whole, when the C-4 position of quinazoline was substituted
by 2,3-dihydroindole or 1,2,3,4-tetrahydroquinoline, there would be no significant effect on antiproliferative activity.
Among the selected compounds, 10e, 12c, 12g, 13a, 13c and 14a showed excellent antiproliferative activity (1.09 -
2.85 µM) against A549, MCF-7 and PC-3 cancer cell lines. As illustrated in Table 1, it can be seen that the compounds
substituted with 2,3-dihydroindole and 1,2,3,4-tetrahydroquinoline at C-4 position of quinazoline had almost the same
influence of the antiproliferative activity. It was noticeable that compounds (8a–g and 10a–e) with the increasing of alkyl
chain length had a slightly impact on the activity. Notably, hex-2-enamide group substituted at C-7 position of
quinazoline was more preferred, such as compounds 8g and 10e. We speculated that the increasing of the alkyl chain
length may contribute to cell binding capacity in vitro. Compound 10e showed superior inhibitory activity against three
tested cells, with IC50 values of 1.80 ± 0.15 µM, 1.21 ± 0.08 µM, and 2.24 ± 0.35 µM against A549, MCF-7 and PC-3
cell lines, respectively. In addition, the results indicated that the replacement of the small molecule tetrahydrofuran
moiety by methyl group could maintained the antitumor activity. In Table 2, It is worth mentioning that the substituents
of the secondary amines had a slightly influence on antiproliferative activity. The compounds (13a and 14a) including
dimethylamine could improve the antiproliferative activity against A549, MCF-7 and PC-3 cell lines, which were more
remarkable than that of compounds containing other secondary amines. We speculated that dimethylamine unit may
contribute to high binding affinity. On the whole, the compounds substituted with methoxy or tetrahydrofuran-3-oxy at

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C-8 position of quinazoline showed slightly influence of the antiproliferative activity. What’s more, compound 13a
showed the best antiproliferative activity against the three tested human cancer cell lines. The results of the
dosage-dependent effects on cell lines were showed in Figs. 3. It was found that compound 13a exerted antiproliferative
effects in a dosage-dependent manner and the IC50 values of compound 13a were 1.09 µM, 1.34 µM and 1.23 µM against
A549, MCF-7 and PC-3, which were equal or more active than afatinib (0.71 µM, 0.93 µM and 2.51 µM), respectively.
Experiments against EGFRWT and EGFRT790M kinases of eight selected compounds (8e, 9d, 12c, 12d, 12g, 13a and
14a) were carried out in this paper for further study. In Table 3, obviously, it was found that the overall activity against
EGFRWT kinase of compounds 12c, 12g and 13a was equal to the reference compound afatinib, with the IC50 values of 3
nM, 4 nM and 5 nM against EGFRWT kinase, respectively. Most of the eight selected compounds showed excellent
activity against EGFRT790M kinase. In particular, the compound 12g showed excellent inhibitory activity to EGFRT790M
with the IC50 values of 21 nM. On the whole, it was found that the overall activity against EGFRT790M kinase of the
selected compounds was a little bit lower than the reference compound afatinib. In a word, the results prompted us that
this series of compounds may be a kind of potential EGFRT790M kinase inhibitors.
2.3 Apoptosis result analyzing
There are three types of programmed cell death in biological systems, namely autophagy, necrosis and apoptosis.
Apoptosis is a series of biochemical events leading to morphological changes and cell death. It has been observed that
many cytotoxic drugs show antiproliferative activities by inducing cell apoptosis. To further explain the inhibition of cell
growth, we first evaluated the effect of compound 13a induced apoptosis on A549 cells using AnnexinV-FITC and
propidium iodide (PI) double staining by flow cytometry, which stained phosphatidylserine residues and DNA,
respectively. The results were shown in Fig. 4. As can be seen, compared with the control group, the number of early
apoptotic cells of afatinib and compound 13a was increased, the number of late apoptotic and death cells was increased
as well. The number of mechanical damage cells was within the error range, which claimed that our data was credible.
In addition, the number of early apoptotic cells of compound 13a was equal to the number of afatinib. What’s more, the
number of late apoptotic and death cells (14.59%) was higher than number of afatinib (13.53%). It could be concluded
that the target compound 13a can induce apoptosis of A549 and it was more efficient than afatinib.

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Fig. 4. 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.
2.4 Morphology of cells by fluorescence microscopy
To further confirm the induction of apoptosis, we monitored the morphological changes of A549 cells induced by
compound 13a by acridine orange single staining. After exposure to a concentration of 1.09 µM of compound 13a for 12
h, cell morphology was investigated using fluorescence microscopy. As reflected in Fig. 5a, it can be observed that the
control cells showing the nucleus of A549 cells were stained into green from fluorescence microscopy, the cells grew
well, the edges were clear and the refraction was great in the control group. However, when the cells were treated with
compound 13a, it can be seen in 5b, that the A549 cells showed significant evidence of apoptosis mediated cell death,
including that the nuclei were dyed red, and cells shrunk and chromatin condensed. It suggested that compound 13a
could induce the cellular apoptosis, which was consistent with the former result of antiproliferative activity assay.
Fig. 5 Morphologic changes of A549 cells under inverted microscopy and fluorescence Microscopy. (5a) The control
group cell treated with nothing; (5b) Experimental group treated with 1.09 µM concentration of compound 13a.
2.5 Molecular docking study

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To explore the binding modes of target compounds with the active site of EGFR, molecular docking simulation
studies were carried out by using SURFLEX-DOCK module of SYBYL package version. Based on the inhibition results,
we selected 13a, our best EGFR inhibitor in this study, as ligand example, and the structure of EGFR was selected as the
docking model (PDB ID code :4G5P).
Fig. 6. Molecular docking analysis was carried out by the SYBYL to explore the binding model for the active site of
EGFR with its ligand. Docking simulations show that the compound 13a occupy the ATP site of EGFR kinase (Fig. 6a).
Binding model of afatinib and compound 13a with EGFR (Fig. 6b). The proteins were displayed by colors ribbon.
Compound 13a and afatinib were displayed by white and wine red sticks, respectively. H-bonding interactions between
the 13a, afatinib and EGFR were indicated with dashed lines in yellow.
The binding modes of afatinib and compound 13a were depicted in Fig. 6. Visual inspection of the pose of the
compound 13a was tightly embedded into the active binding pocket in Fig. 6a. In Fig. 6b, compound 13a and afatinib
can nearly overlap in the binding model. In addition, afatinib can formed hydrogen bonds with residues LYS-860,
LEU-861 and LYS-949, compound 13a can formed hydrogen bonds with residues LEU-861 and LYS-949. In general,
these results of the molecular docking study showed that quinazoline derivatives bearing 2,3-dihydro-indole or
1,2,3,4-tetrahydroquinoline moiety could act synergistically to interact with the active binding site of EGFR. The results
suggested that compound 13a may be a potential inhibitor of EGFR. Analysis of the binding mode in the active binding
site demonstrated that the docking mode of the 13a was similar to the afatinib in the respect of binding site. The Fig. 6b
showed that compound 13a and afatinib can bind to the ATP-binding pocket of the EGFR protein. Combining with the
antiproliferative activity results of compound 13a, whose antiproliferative activity against the tested three human cancer
cell lines was equal to or even higher than that of afatinib, the results suggested that the results of molecular docking and
anti-proliferative activity were consistent. In Fig. 6a, It can be seen that the dimethylamine group extends into the
hydrophilic pocket, it was the reason that the four series of compounds 12a–g, 13a–f, 14a–f and 15a–f were much more
active than the other series of compounds 8a–g, 9a–g, 10a–e and 11a–e against the three cancer cell lines. It means our

ACCEPTED MANUSCRIPT
deduction may be correct which secondary amines may contribute to increasing the water-soluble. Furthermore, the
above-mentioned results of structure-activity relationships analysis and molecular docking study may allow the rational
design of more potential EGFR inhibitors.
3 Conclusions
Eight series of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline moiety (8a–g,
9a–g, 10a–e, 11a–e, 12a–g, 13a–f, 14a–f and 15a–f) were designed, synthesized and evaluated for the IC50 values
against three cancer cell lines (A549, MCF-7 and PC-3). Most of the compounds showed excellent antiproliferative
activity against the different cancer cells. Notably, compound 13a showed the best activity against A549, MCF-7 and
PC-3 cancer cell lines and exerted antiproliferative effects in a dosage-dependent manner. The antiproliferative activity
data showed that the eight series of compounds possessed more selectivity for A549 cell line than MCF-7 and PC-3 cell
lines. That meant these compounds had better treatment effect on lung cancer. On the whole, the compounds substituted
with secondary amines were beneficial to the antiproliferative activity. What’s more, the compound 13a could induce
apoptosis of A549 cells and it’s binding model to the EGFR was similar to afatinib in the respect of H-bond. The activity
against EGFR kinase of compounds 12c, 12g and 13a were equal to the reference compound afatinib. The results
prompted us that compounds 12c, 12g and 13a may be potential EGFR kinase inhibitors.
4 Experimental
4.1. 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 electrospray
ionization (ESI) mode on Agilent 1100 LC–MS (Agilent, Palo Alto, CA, USA). Thin layer chromatography (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.
4.2 Preparation of 4-chloro-7-fluoro-6-nitroquinazoline (3)
Compound 3 was synthesized according to the procedures in our previous research [13]
4.3 Preparation of 4-(2,3-dihydro-indol-1-yl)-7-fluoro-6-nitro-quinazoline (4a)
.
To the solution of compound 3 (50.0 g, 0.220 mol), isopropanol (400 mL), triethylamine (35 mL),
2,3-dihydro-indole (31.5 g, 0.265 mol) was added at room temperature. Then the reaction mixture was stirred at room
temperature for 1.5 h. After completion of reaction as indicated by TLC, the mixture was filtered, washed with

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isopropanol and water, dried to compound 4a as orange solid (65.0 g, 95.4%). m.p. 261.4-262.5 °C. ESI-MS m/z:
[M+H]+311.2. 1H NMR (400 MHz, CDCl3) δ 9.02 (d, J = 8.1 Hz, 1H), 8.74 (s, 1H), 7.90 (d, J = 8.1 Hz, 1H), 7.85 (d, J =
12.5 Hz, 1H), 7.36 (d, J = 7.3 Hz, 1H), 7.20 (t, J = 7.8 Hz, 1H), 7.08 (t, J = 7.4 Hz, 1H), 4.59 (t, J = 7.8 Hz, 2H), 3.21 (t,
J = 7.7 Hz, 2H).
4.4 Preparation of 4-(3,4-dihydroquinolin-1(2H)-yl)-7-fluoro-6-nitroquinazoline (4b)
To the solution of compound 3 (20.0 g, 0.088 mol), isopropanol (160 mL), triethylamine (14 mL),
1,2,3,4-tetrahydroquinoline (14.1 g, 0.106 mol) was added at room temperature. Then the reaction mixture was stirred at
room temperature for 1.5 h. After completion of reaction as indicated by TLC, the mixture was filtered, washed with
isopropanol and water, dried to compound 4b as orange solid (26.2 g, 92.1%). m.p. 252.4-254.6 °C. ESI-MS m/z:
[M+H]+324.3. 1H NMR (400 MHz, DMSO-d6) δ 9.04 (s, 1H), 7.98 (s, 1H), 7.39 (d, J = 7.4 Hz, 1H), 7.21 (t, J = 7.4 Hz,
1H), 7.05 (t, J = 7.6 Hz, 1H), 6.97 (s, 1H), 6.93 (d, J = 8.0 Hz, 1H), 4.15 (t, J = 6.3 Hz, 2H), 2.92 (t, J = 6.6 Hz, 2H), 2.09
(p, J = 6.5 Hz, 2H).
4.5 Preparation of 4-(2,3-dihydro-indol-1-yl)-7-methoxy-6-nitro-quinazoline (5a)
To the mixture of compound 4a (10.0 g, 0.032 mol), methanol (227 mL), 50 percent sodium hydroxide (15 mL) was
added at room temperature. Then the reaction mixture was refluxed for 1 h and monitored by TLC. The reaction mixture
was cooled to room temperature and poured into sodium bicarbonate solution, yielding a precipitate. Then the mixture
was separated by filtration and washed with water, dried to afford compound 5a as yellow solid (9.1 g, 87.6%). m.p.
292.3-293.0 °C. ESI-MS m/z: [M+H]+323.1.
4.6 Preparation of 4-(2,3-dihydro-indol-1-yl)-6-nitro-7-(tetrahydro-furan-3-yloxy)-quinazoline (5b)
To the mixture of 60%NaH (7.68 g, 0.053 mol), anhydrous THF (400 mL), (S)-tetrahydrofuran-3-ol (8.45 g, 0.096
mol) was added drop-wise at room temperature. The mixture was stirred at room temperature for 1 h. a solution of
compound 4a (10.0 g, 0.032 mol) in anhydrous THF (200 mL) was added via a dropping funnel over 1 h (the dropping
funnel was rinsed with 15 mL of THF) at room temperature and the reaction mixture was stirred at room temperature for
2 h, the reaction was monitored by TLC. Then the solution was concentrated under a reduced pressure and the residue
was poured into water with stirring, yielding a precipitate, then the mixture was filtered and washed with ice-water and
ethanol, dried to afford compound 5b as yellow solid (9.8 g, 81.0%). m.p. 215.1-217.0 °C. ESI-MS m/z: [M+H]+379.2.
4.7 Preparation of 4-(2,3-dihydro-indol-1-yl)-7-methoxy-6-nitro-quinazoline (5c)
The synthetic method of compound 5c was the same to compound 5a. Compound 5c was obtained as yellow solid

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(9.1 g, 81.7%). m.p. 272.1-273.4 °C. ESI-MS m/z: [M+H]+337.2.
5.8 Preparation of (R)-4-(3,4-dihydroquinolin-1(2H)-yl)-6-nitro-7-((tetrahydrofuran-3-yl)oxy)quinazoline (5d)
The synthetic method of compound 5d was the same to compound 5b. Compound 5d was obtained as yellow solid
(8.7 g, 76.9%). m.p. 294.1-295.4 °C. ESI-MS m/z: [M+H]+393.2.
4.9 Preparation of 4-(2,3-dihydro-indol-1-yl)-7-methoxy-quinazolin-6-ylamine (6a)
The solution of compound 5a (9.1 g, 0.028 mol) and ethanol (370 mL) was stirred at 60 °C, an appropriate amount
of activated carbon (3.5 g) and ferric chloride (1.3 g) were added at the temperature, the mixture was heated to 80 °C and
80 percent hydrazine hydrate (7 mL) was added to the solution. The reaction mixture was refluxed for 1.5 h and
monitored by TLC. Then the mixture was filtered and the precipitate was washed with ethanol. The filtrate was
concentrated under reduce pressure and the residue was poured into water with stirred for 30 min. The precipitate was
filtered and dried to obtain compound 6a as yellow solid (7.6 g, 91.8%). m.p. 258.4-259.7 °C. ESI-MS m/z:
[M+H]+293.3 . 1H NMR (400 MHz, DMSO-d6) δ 8.50 (s, 1H), 7.27 (d, J = 7.1 Hz, 1H), 7.17 (s, 1H), 7.03 (d, J = 7.4 Hz,
1H), 7.00 (s, 1H), 6.87 (s, 2H), 5.51 (s, 2H), 4.23 (t, J = 7.6 Hz, 2H), 3.98 (s, 3H), 3.15 (d, J = 8.1 Hz, 2H).
4.10 Preparation of 4-(2,3-dihydro-indol-1-yl)-7-(tetrahydro-furan-3-yloxy)-quinazolin-6-ylamine (6b)
The synthetic method of compound 6b was the same to compound 6a. Compound 6b as light yellow solid (8.0 g,
1
89.0%). m.p. 239.1-240.1 °C. ESI-MS m/z: [M+H]+349.1. H NMR (400 MHz, DMSO-d6) δ 8.50 (s, 1H), 7.27 (d, J =
7.1 Hz, 1H), 7.17 (s, 1H), 7.03 (d, J = 7.4 Hz, 1H), 7.00 (s, 1H), 6.80 (t, 2H), 5.51 (s, 2H), 4.95 (s, 1H), 4.02 – 3.93 (m,
2H), 3.89 (dd, J = 15.4, 7.5 Hz, 2H), 3.82 – 3.78 (m, 2H), 2.87 (t, J = 6.4 Hz, 2H), 2.31 (td, J = 14.1, 7.7 Hz, 1H), 2.15 –
2.07 (m, 1H).
4.11 Preparation of 4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-amine (6c)
The synthetic method of compound 6c was the same to compound 6a. Compound 6c as light yellow solid (8.0 g,
1
89.0%). m.p. 228.2-229.4 °C. ESI-MS m/z: [M+H]+307.1. H NMR (400 MHz, DMSO-d6) δ 8.57 (s, 1H), 7.29 (d, J =
7.1 Hz, 1H), 7.13 (s, 1H), 7.03 (d, J = 7.4 Hz, 1H), 7.00 (s, 1H), 6.75 (t, J = 7.4 Hz, 1H), 6.61 (t, J = 7.3 Hz, 1H), 6.83 (s,
1H), 5.51 (s, 2H), 3.98 (s, 3H), 3.82 – 3.78 (m, 2H), 2.87 (t, J = 6.4 Hz, 2H), 2.05 – 1.98 (m, 2H).
4.12 Preparation of (R)-4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-amine (6d)
The synthetic method of compound 6d was the same to compound 6a. Compound 6d as light yellow solid (8.0 g,
1
89.0%). m.p. 231.1-242.5 °C. ESI-MS m/z: [M+H]+363.2. H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.10 (d, J =

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6.9 Hz, 1H), 7.01 (s, 1H), 6.90 – 6.80 (m, 2H), 6.67 (s, 1H), 6.34 (d, J = 7.8 Hz, 1H), 5.53 (s, 2H), 5.23 (s, 1H), 4.07 –
3.95 (m, 2H), 3.89 (dd, J = 15.4, 7.5 Hz, 2H), 3.82 – 3.78 (m, 2H), 3.17 (t, J = 6.4 Hz, 2H), 2.31 (td, J = 14.1, 7.7 Hz,
1H), 2.25 – 2.15 (m, 1H), 2.05 – 1.98 (m, 2H).
4.13 Preparation of diethyl (2-((4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)amino)-2-oxoethyl)phosphonate(7a)
To the solution of diethoxy phosphorylacetic acid (1.5 g, 7.4 mmol) are placed in 100 ml of THF and combined with
N,N'-carbonyldiimidazole (1.2 g, 7.4 mmol) at ambient temperature. After the development of gas has ceased, then
compound 6a (2.0 g, 6.7 mmol) are added and the reaction mixture was heated to 40 °C, the reaction mixture was
refluxed for 2 h and monitored by TLC. Then the mixture was concentrated under reduce pressure and the residue was
poured into water with stirred for 30 min. The precipitate was filtered and dried to obtain compound 7a as white solid
1
(2.3 g, 71.9%). m.p. 188.2-189.7 °C. ESI-MS m/z: [M+H]+471.5. H NMR (400 MHz, CDCl3) δ 9.22 (s, 1H), 9.18 (s,
1H), 8.67 (s, 1H), 7.62 (d, J = 7.9 Hz, 1H), 7.38 (s, 1H), 7.26 (s, 1H), 7.18 (t, J = 7.8 Hz, 1H), 7.00 (t, J = 7.2 Hz, 1H),
4.53 (t, J = 7.8 Hz, 2H), 4.18 (dd, J = 14.7, 7.4 Hz, 4H), 4.09 (s, 3H), 3.23 (t, J = 7.8 Hz, 2H), 3.06 (d, J = 20.8 Hz, 2H),
1.35 (t, J = 6.9 Hz, 6H).
4.14
Preparation
of
diethyl
(R)-(2-((4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)amino)-2-oxoethyl)phosphonate (7b)
The synthetic method of compound 7b was the same to compound 7a. Compound 7b as white solid (2.0 g, 66.7%).
m.p. 199.4-200.5 °C. ESI-MS m/z: [M+H]+527.5.
4.15
Preparation
of
diethyl
(2-((4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)amino)-2-oxoethyl)phosphonate (7c)
The synthetic method of compound 7c was the same to compound 7a. Compound 7c as white solid (2.1 g, 65.6%).
m.p. 203.1-205.7 °C. ESI-MS m/z: [M+H]+485.7.
4.16
Preparation
of
diethyl
(R)-(2-((4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)amino)-2-oxoethyl)phosphonate
(7d)
The synthetic method of compound 7d was the same to compound 7a. Compound 7d as white solid (1.9 g, 63.6%).
m.p. 183.1-184.6 °C. ESI-MS m/z: [M+H]+541.6.
4.17 Preparation of 2,2-dimethoxy-N,N-dimethylethan-1-amine (A1)

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To a solution of 2-bromo-1,1-dimethoxyethane (2.0 g, 0.012 mol), Dimethylamine (0.54 g, 0.012 mol) and K2CO3
(2.48 g, 0.018 mol) in 1,4-dioxane (20 mL). The reaction mixture was heated to 100 °C for 4 h and monitored by TLC.
After completion, the reaction was quenched with saturated aqueous NaCl and then extracted with DCM (30 mL × 3).
The combined organic layer was dried over anhydrous Na2SO4, filtered and concentrated to get colorless oil directly for
further reaction. ESI-MS m/z: [M+H]+134.1.
4.18 Preparation of N,N-diethyl-2,2-dimethoxyethan-1-amine (A2), 1-(2,2-dimethoxyethyl)pyrrolidine (A3),
1-(2,2-dimethoxyethyl)piperidine
(A4),
1-(2,2-dimethoxyethyl)-4-methylpiperazine
(A5),
4-(2,2-dimethoxyethyl)morpholine (A6) and (4-(2,2-dimethoxyethyl)piperazin-1-yl)methanol (A7),
The synthetic method of compounds A2–A7 was the same to compound A1. Compounds A2–A7 as yellow liquid.
4.19 Preparation of 2-(dimethylamino)acetaldehyde (B1)
To a solution of 2,2-dimethoxy-N,N-dimethylethan-1-amine (1.5 g, 0.009 mol) dissolved in concentrated 30% HCl
(10 mL) was refluxed at 110 °C for 4 h. The reaction mixture was concentrated in reduced pressure to obtain dark oil,
which was used without purification.
4.20
Preparation
of
2-(diethylamino)acetaldehyde
(B2),
2-(pyrrolidin-1-yl)acetaldehyde
(B3),
2-(piperidin-1-yl)acetaldehyde (B4), 2-(4-methylpiperazin-1-yl)acetaldehyde (B5), 2-morpholinoacetaldehyde (B6) and
2-(4-(hydroxymethyl)piperazin-1-yl)acetaldehyde (B7)
The synthetic method of compounds B2–B7 was the same to compound B1. Compounds B2–B7 as yellow liquid.
4.21 General procedure for the preparation of compounds 8a–g, 9a–g, 10a–e and 11a–e
Oxalyl chloride (2.0 mmol) was added drop-wise to a stirred mixture of the corresponding acrylic acid derivatives
(Acrylic acid, Methacrylic acid, 3,3-Dimethacrylic acid, Trans-2-butenoic acid, Trans-2-pentenoic acid,
4-methyl-2-pentenoic acid, Trans-2-hexenoic acid) (1.0 mmol) and DMF( 0.02 mmol) in dichloromethane (16 mL) at
room temperature for 10 min, and the mixture was distilled and dissolved in dichloromethane (16 mL) immediately. The
solution was used for the next step without further purification. Then, the solution was added drop-wise to a solution of
compounds 6a–d (0.4 mmol) and diisopropylethylamine (0.4 mmol) in dichloromethane (15 mL) in an ice bath.
Upon completion of the addition, the reaction mixture was removed from the ice bath and placed at room
temperature for 30 min and monitored by TLC. The mixture was washed with 10% K2CO3 (50 mL × 3) followed
by saturated aqueous NaCl (50 mL × 1), and the organic phase was separated, dried, and evaporated to yield 8a–g,
9a–g, 10a–e and 11a–e which were purified by dichloromethane.

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4.22 General procedure for the preparation of compounds 12a–g, 13a–f, 14a–f and 15a–f
50%NaOH (1 mL) was dissolved in THF (5 mL), the corresponding compound 7a–d (0.5 mmol) was added to the
above solution. After the mixture with stirred for 10min, the corresponding aldehyde B1–B7 (2.0 mmol) was added at
0−5 °C in an ice bath. The reaction mixture was stirred at room temperature for the indicated time. Then the reaction
mixture was adjusted to pH = 1−2 with 2 N HCl and washed with EtOAc (3 × 30 mL) three times. The aqueous layers
were adjusted to pH = 10.0 with 4N NaOH and extracted with EtOAc (3 × 30 mL) three times. The combined organic
layer was washed with saturated aqueous NaCl, dried over anhydrous Na2SO4, and concentrated in reduced pressure to
obtain the crude product. The crude produc was purified through a column chromatography on silica with
dichloromethane/methanol (V:V ¼ 30:1) as eluent to produce 12a–g, 13a–f, 14a–f and 15a–f as white solid.
N-[4-(2,3-dihydro-indol-1-yl)-7-methoxy-quinazolin-6-yl]-acrylamide (8a)
White solid in 65% yield. m.p. 165.2-166.9 °C. ESI-MS m/z: [M+H]+ 347.1. 1H NMR (400 MHz, DMSO-d6) δ 9.76
(s, 1H), 9.03 (s, 1H), 8.61 (s, 1H), 7.42 (d, J = 8.1 Hz, 1H), 7.35 (s, 1H), 7.30 (d, J = 7.2 Hz, 1H), 7.11 (t, J = 7.7 Hz, 1H),
6.95 (t, J = 7.4 Hz, 1H), 6.80 (dd, J = 17.0, 10.2 Hz, 1H), 6.26 (d, J = 16.9 Hz, 1H), 5.76 (d, J = 10.2 Hz, 1H), 4.41 (t, J =
8.0 Hz, 2H), 4.06 (s, 3H), 3.20 (t, J = 7.9 Hz, 2H).
N-[4-(2,3-dihydro-indol-1-yl)-7-methoxy-quinazolin-6-yl]-2-methyl-acrylamide (8b)
White solid in 64% yield. m.p. 163.1-164.2 °C. ESI-MS m/z: [M+H]+ 361.2. 1H NMR (400 MHz, DMSO-d6) δ 9.25
(s, 1H), 8.77 (s, 1H), 8.60 (s, 1H), 7.38 (d, J = 8.0 Hz, 1H), 7.35 (s, 1H), 7.29 (d, J = 7.3 Hz, 1H), 7.08 (t, J = 7.6 Hz, 1H),
6.94 (t, J = 7.4 Hz, 1H), 4.37 (t, J = 8.0 Hz, 2H), 4.07 (s, 3H), 3.18 (t, J = 7.8 Hz, 2H), 2.72 (ddd, J = 12.9, 6.6, 2.8 Hz,
1H), 2.36 – 2.29 (m, 1H), 1.62 (s, 3H).
N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)-3-methylbut-2-enamide (8c)
White solid in 67% yield. m.p. 137.1-139.4 °C. ESI-MS m/z: [M+H]+ 375.1. 1H NMR (400 MHz, DMSO-d6) δ 9.46
(s, 1H), 9.15 (s, 1H), 8.78 (s, 1H), 7.90 (d, J = 7.8 Hz, 1H), 7.45 (s, 1H), 7.38 (d, J = 7.1 Hz, 1H), 7.24 (t, J = 7.5 Hz, 1H),
7.13 (t, J = 7.2 Hz, 1H), 6.21 (s, 1H), 4.61 (t, J = 7.2 Hz, 2H), 4.05 (s, 3H), 3.26 (t, J = 7.0 Hz, 2H), 2.14 (s, 3H), 1.87 (s,
3H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)but-2-enamide (8d)
White solid in 65% yield. m.p. 151.6-153.1 °C. ESI-MS m/z: [M+H]+ 361.3. 1H NMR (400 MHz, CDCl3) δ 9.30 (s,
1H), 8.66 (s, 1H), 7.97 (s, 1H), 7.69 (d, J = 8.1 Hz, 1H), 7.30 (s, 1H), 7.26 (d, J = 3.3 Hz, 1H), 7.19 (t, J = 7.7 Hz, 1H),

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7.05 – 6.95 (m, 2H), 6.03 (d, J = 15.0 Hz, 1H), 4.56 (t, J = 8.0 Hz, 2H), 4.06 (s, 3H), 3.22 (t, J = 7.9 Hz, 2H), 1.93 (d, J =
6.8 Hz, 3H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)pent-2-enamide (8e)
White solid in 61% yield. m.p. 129.2-130.8 °C. ESI-MS m/z: [M+H]+ 375,2. 1H NMR (400 MHz, DMSO-d6) δ 9.77
(s, 1H), 9.30 (s, 1H), 8.84 (s, 1H), 8.05 (d, 1H), 7.52 (s, 1H), 7.41 (d, J = 6.4 Hz, 1H), 7.29 (t, 1H), 7.19 (t, J = 7.0 Hz,
1H), 6.93 (m, J = 15.1 Hz, 1H), 6.51 (d, J = 15.1 Hz, 1H), 4.68 (t, 2H), 4.08 (s, 3H), 3.28 (t, 2H), 2.23 (m, 2H), 1.05 (t, J
= 7.0 Hz, 3H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)-4-methylpent-2-enamide (8f)
White solid in 66% yield. m.p. 130.7-131.9 °C. ESI-MS m/z: [M+H]+ 389.2. 1H NMR (400 MHz, CDCl3) δ 9.76 (s,
1H), 9.26 (s, 1H), 8.80 (s, 1H), 7.98 (d, J = 6.8 Hz, 1H), 7.50 (s, 1H), 7.39 (d, J = 6.2 Hz, 1H), 7.26 (t, 1H), 7.15 (t, 1H),
6.86 (dd, J = 14.9, 5.6 Hz, 1H), 6.47 (d, J = 15.3 Hz, 1H), 4.64 (t, 2H), 4.07 (s, 3H), 3.26 (t, 2H), 2.48 – 2.42 (m, 1H),
1.05 (d, J = 6.0 Hz, 6H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)hex-2-enamidee (8g)
White solid in 65% yield. m.p. 149.9-151.4 °C. ESI-MS m/z: [M+H]+ 389.1. 1H NMR (400 MHz, CDCl3) δ 9.43 (s,
1H), 8.92 (s, 1H), 8.51 (s, 1H), 7.32 (d, J = 7.9 Hz, 1H), 7.24 (s, 1H), 7.20 (d, J = 6.7 Hz, 1H), 7.05 – 6.98 (t, 1H), 6.88 –
6.82 (t, 1H), 6.74 – 6.69 (m, 1H), 6.39 (d, J = 15.3 Hz, 1H), 4.32 (t, J = 7.2 Hz, 2H), 3.96 (s, 3H), 3.11 (t, J = 7.2 Hz, 2H),
2.08 (m, J = 6.9 Hz, 2H), 1.38 – 1.34 (m, 2H), 0.83 – 0.80 (t, 3H).
N-[4-(2,3-dihydro-indol-1-yl)-7-(tetrahydro-furan-3-yloxy)-quinazolin-6-yl]-acrylamide (9a)
White solid in 67% yield. m.p. 169.1-170.1 °C. ESI-MS m/z: [M+H]+ 403.0. 1H NMR (400 MHz, DMSO-d6) δ 9.44
(s, 1H), 8.64 (d, J = 3.0 Hz, 1H), 8.07 (s, 1H), 7.83 (d, J = 8.1 Hz, 1H), 7.54 (s, 1H), 7.30 (d, J = 7.1 Hz, 1H), 7.23 (d, J =
7.7 Hz, 1H), 7.07 (t, J = 7.2 Hz, 1H), 6.46 (d, J = 16.8 Hz, 1H), 6.33 (dd, J = 16.9, 10.1 Hz, 1H), 5.85 (d, J = 10.1 Hz,
1H), 5.26 (s, 1H), 4.67 – 4.57 (m, 2H), 4.19 (d, J = 10.6 Hz, 1H), 4.11 – 4.04 (m, 2H), 3.92 (dd, J = 13.3, 5.6 Hz, 1H),
3.27 (t, J = 7.9 Hz, 2H), 2.55 – 2.47 (m, 1H), 2.29 – 2.22 (m, 1H).
N-[4-(2,3-dihydro-indol-1-yl)-7-(tetrahydro-furan-3-yloxy)-quinazolin-6-yl]-2-methyl-acrylamide (9b)
White solid in 65% yield. m.p. 208.2-209.3 °C. ESI-MS m/z: [M+H]+ 417.1. 1H NMR (400 MHz, CDCl3) δ 9.29 (d,
J = 16.7 Hz, 1H), 9.12 (s, 1H), 8.65 (s, 1H), 7.69 (s, 1H), 7.37 (s, 1H), 7.29 (d, J = 7.5 Hz, 1H), 7.19 (d, J = 7.4 Hz, 1H),
7.05 (d, J = 7.9 Hz, 1H), 5.22 (s, 1H), 4.54 (d, J = 7.6 Hz, 2H), 4.19 (d, J = 5.8 Hz, 2H), 4.04 (s, 1H), 3.26 (t, J = 7.7 Hz,

ACCEPTED MANUSCRIPT
2H), 2.94 (s, 1H), 2.43 (s, 2H), 2.40 – 2.36 (m, 1H), 2.06 (d, J = 17.6 Hz, 1H), 1.71 (s, 3H).
(R)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-3-methylbut-2-enamide (9c)
White solid in 60% yield. m.p. 142.3-143.9 °C. ESI-MS m/z: [M+H]+ 431.1. 1H NMR (400 MHz, DMSO-d6) δ 9.05
(s, 1H), 8.94 (s, 1H), 8.60 (s, 1H), 7.37 (d, J = 8.1 Hz, 1H), 7.30 (d, J = 7.4 Hz, 1H), 7.27 (s, 1H), 7.10 (t, J = 7.6 Hz, 1H),
6.95 (t, J = 7.3 Hz, 1H), 6.17 (s, 1H), 5.32 (m, 1H), 4.39 (t, J = 7.9 Hz, 2H), 4.03 (dt, J = 10.2, 7.2 Hz, 2H), 3.98 – 3.91
(m, 2H), 3.20 (t, J = 7.6 Hz, 2H), 2.35 (dd, J = 13.8, 6.7 Hz, 1H), 2.19 (dd, J = 13.2, 5.8 Hz, 1H), 2.12 (s, 3H), 1.88 (s,
3H).
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)but-2-enamide (9d)
White solid in 65% yield. m.p. 187.1-188.7 °C. ESI-MS m/z: [M+H]+ 417.3. 1H NMR (400 MHz, CDCl3) δ 9.32 (s,
1H), 8.64 (s, 1H), 7.91 (s, 1H), 7.69 (d, J = 8.1 Hz, 1H), 7.25 (d, J = 7.5 Hz, 1H), 7.21 (s, 1H), 7.17 (d, J = 7.9 Hz, 1H),
7.04 – 6.93 (m, 2H), 6.01 (d, J = 15.1 Hz, 1H), 5.18 (s, 1H), 4.55 (t, J = 7.9 Hz, 2H), 4.16 (d, J = 10.6 Hz, 1H), 4.08 –
4.02 (m, 2H), 3.93 (dt, J = 13.6, 6.9 Hz, 1H), 3.21 (t, J = 8.0 Hz, 2H), 2.42 (td, J = 14.3, 7.4 Hz, 1H), 2.29 – 2.21 (m, 1H),
1.93 (d, J = 6.9 Hz, 3H).
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)pent-2-enamide (9e)
White solid in 66% yield. m.p. 137.7-139.1 °C. ESI-MS m/z: [M+H]+ 431.1. 1H NMR (400 MHz, DMSO-d6) δ 9.34
(s, 1H), 9.12 (s, 1H), 8.68 (s, 1H), 7.62 (d, J = 8.1 Hz, 1H), 7.34 (d, J = 7.8 Hz, 1H), 7.32 (s, 1H), 7.17 (t, J = 7.7 Hz, 1H),
7.03 (t, J = 7.4 Hz, 1H), 6.90 (dt, J = 15.2, 6.4 Hz, 1H), 6.48 (d, J = 15.4 Hz, 1H), 5.32 (s, 1H), 4.49 (t, J = 7.9 Hz, 2H),
4.10 (d, J = 10.3 Hz, 1H), 4.02 (dd, J = 10.5, 4.9 Hz, 1H), 3.99 – 3.94 (m, 1H), 3.81 (dt, J = 13.2, 6.6 Hz, 1H), 3.22 (t, J
= 7.7 Hz, 2H), 2.42 – 2.34 (m, 1H), 2.27 – 2.22 (m, 2H), 2.19 (d, J = 7.8 Hz, 1H), 1.06 (t, J = 7.4 Hz, 3H).
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-methylpent-2-enamide (9f)
White solid in 65% yield. m.p. 135.9-137.1 °C. 1H NMR (400 MHz, DMSO-d6) δ 9.24 (s, 1H), 9.02 (s, 1H), 8.56 (s,
1H), 7.42 (d, J = 8.0 Hz, 1H), 7.27 (d, J = 7.3 Hz, 1H), 7.24 (s, 1H), 7.09 (t, J = 7.5 Hz, 1H), 6.93 (t, J = 7.4 Hz, 1H),
6.80 (dd, J = 15.4, 6.5 Hz, 1H), 6.41 (d, J = 15.3 Hz, 1H), 5.29 (s, 1H), 4.38 (t, J = 8.0 Hz, 2H), 4.06 (d, J = 9.9 Hz, 1H),
3.99 (dd, J = 10.3, 4.8 Hz, 1H), 3.93 (t, J = 7.7 Hz, 1H), 3.77 (td, J = 8.1, 5.2 Hz, 1H), 3.17 (t, J = 7.8 Hz, 2H), 2.48 (d, J
= 5.9 Hz, 1H), 2.35 (td, J = 14.2, 7.6 Hz, 1H), 2.23 – 2.15 (m, 1H), 1.04 (s, 3H), 1.03 (s, 3H); 13C NMR (101 MHz,
CDCl3) δ 165.03, 159.70, 153.92, 152.73, 152.49, 150.73, 145.32, 133.19, 127.88, 126.97, 125.44, 122.93, 122.57,
116.60, 116.33, 111.56, 108.23, 79.68, 72.63, 67.32, 54.49, 33.05, 30.91, 29.39, 22.06(2C). HRMS (ESI): m/z: [M+H]+
calcd for C26H28N4O3, 445.2171; found 445.2175.

ACCEPTED MANUSCRIPT
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)hex-2-enamide (9g)
White solid in 62% yield. m.p. 142.4-144.1 °C. ESI-MS m/z: [M+H]+ 445.3. 1H NMR (400 MHz, CDCl3) δ 9.33 (s,
1H), 8.61 (s, 1H), 7.96 (s, 1H), 7.72 (d, J = 8.1 Hz, 1H), 7.30 (s, 1H), 7.24 (d, J = 7.4 Hz, 1H), 7.18 (t, J = 7.7 Hz, 1H),
7.02 – 6.94 (m, 2H), 5.99 (d, J = 15.2 Hz, 1H), 5.19 (s, 1H), 4.55 (t, J = 8.0 Hz, 2H), 4.16 (d, J = 10.6 Hz, 1H), 4.04 (dt,
J = 7.6, 6.3 Hz, 2H), 3.91 (td, J = 8.5, 5.1 Hz, 1H), 3.20 (t, J = 8.0 Hz, 2H), 2.42 (td, J = 14.1, 7.8 Hz, 1H), 2.28 – 2.17
(m, 3H), 1.56 – 1.46 (m, 2H), 0.97 – 0.92 (t, 3H); 13C NMR (101 MHz, CDCl3) δ 164.35, 162.76, 154.98, 154.31, 150.87,
142.64, 141.08, 129.65, 128.71, 128.42, 126.41(2C), 122.62, 120.59, 115.94, 113.42, 106.84, 56.93, 49.29, 26.96, 23.62,
18.10.
N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-3-methylbut-2-enamide (10a)
White solid in 64% yield. m.p. 139.4-140.8 °C. ESI-MS m/z: [M+H]+ 389.4. 1H NMR (400 MHz, DMSO-d6) δ 9.31
(s, 1H), 8.73 (s, 1H), 8.42 (s, 1H), 7.33 (s, 1H), 7.19 (t, 1H), 6.90 (t, J = 3.1 Hz, 2H), 6.62 (m, 1H), 4.81 (d, J = 14.7 Hz,
1H), 4.02 (s, 3H), 3.90 (t, 2H), 3.11 (t, 2H), 2.86 (m, J = 5.7 Hz, 2H), 2.02 (s, 3H), 1.75 (s, 3H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)but-2-enamide (10b)
White solid in 67% yield. m.p. 161.1-162.4 °C. ESI-MS m/z: [M+H]+ 375.3. 1H NMR (400 MHz, DMSO-d6) δ 9.35
(s, 1H), 8.71 (s, 1H), 8.56 (s, 1H), 7.34 (s, 1H), 7.17 (s, 1H), 6.89 (s, 2H), 6.73 (dd, J = 14.1, 6.9 Hz, 1H), 6.60 (s, 1H),
6.39 (d, J = 14.8 Hz, 1H), 4.02 (s, 3H), 3.89 (t, 2H), 2.86 (t, 2H), 1.97 (m, J = 4.4 Hz, 2H), 1.82 (d, J = 5.5 Hz, 3H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)pent-2-enamide (10c)
White solid in 65% yield. m.p. 140.3-142.1 °C. ESI-MS m/z: [M+H]+ 389.1. 1H NMR (400 MHz, DMSO-d6) δ 9.51
(s, 1H), 8.99 (s, 1H), 8.48 (s, 1H), 7.55 (s, 1H), 7.36 (d, J = 7.4 Hz, 1H), 7.25 (s, 1H), 7.16 (t, 1H), 7.05 (s, 2H), 6.87 –
6.73 (m, 1H), 6.38 (d, J = 15.3 Hz, 1H), 4.18 (t, 2H), 4.05 (s, 3H), 2.89 (t, 2H), 2.19 (dd, J = 13.3, 6.5 Hz, 2H), 2.07 (m,
2H), 1.01 (t, J = 7.6 Hz, 3H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-4-methylpent-2-enamide (10d)
White solid in 66% yield. m.p. 152.8-153.9 °C. ESI-MS m/z: [M+H]+ 403.1. 1H NMR (400 MHz, CDCl3) δ 8.87 (s,
1H), 8.75 (s, 1H), 7.95 (s, 1H), 7.31 (s, 1H), 7.21 – 7.16 (t, 1H), 6.97 – 6.93 (m, 2H), 6.92 – 6.86 (t, 1H), 6.84 – 6.80 (m,
1H), 5.88 (d, J = 15.3 Hz, 1H), 4.08 – 4.04 (t, 2H), 4.04 (s, 3H), 2.93 (t, J = 6.6 Hz, 2H), 2.47 (dq, J = 13.4, 6.8 Hz, 1H),
2.09 – 2.01 (m, 2H), 1.07 (s, 3H), 1.06 (s, 3H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)hex-2-enamide (10e)

ACCEPTED MANUSCRIPT
White solid in 68% yield. m.p. 146.3-147.8 °C. ESI-MS m/z: [M+H]+ 403.2. 1H NMR (400 MHz, CDCl3) δ 9.49 (s,
1H), 8.95 (s, 1H), 8.48 (s, 1H), 7.50 (s, 1H), 7.34 (d, J = 7.0 Hz, 1H), 7.13 (t, 1H), 7.01 (s, 2H), 6.75 – 6.66 (m, 1H), 6.38
(d, J = 15.4 Hz, 1H), 4.14 (t, 2H), 4.04 (s, 3H), 2.88 (t, 2H), 2.13 (m, J = 6.0 Hz, 2H), 2.05 (t, 2H), 1.43 (m, J = 6.7 Hz,
2H), 0.88 (t, J = 6.9 Hz, 3H).
(R)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-3-methylbut-2-enamide (11a)
White solid in 65% yield. m.p. 148.9-150.2 °C. ESI-MS m/z: [M+H]+ 445.1. 1H NMR (400 MHz, DMSO-d6) δ 8.86
(s, 1H), 8.63 (s, 1H), 8.44 (s, 1H), 7.22 (s, 1H), 7.17 – 7.13 (m, 1H), 6.91 – 6.85 (m, 2H), 6.59 – 6.53 (m, 1H), 5.99 (s,
1H), 5.23 (s, 1H), 3.97 (dd, J = 13.5, 7.6 Hz, 2H), 3.90 – 3.83 (m, 4H), 2.83 (t, J = 6.5 Hz, 2H), 2.32 (td, J = 14.0, 7.5 Hz,
1H), 2.15 – 2.09 (m, 1H), 2.02 (s, 3H), 1.94 (t, J = 6.0 Hz, 2H), 1.81 (s, 3H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)but-2-enamide (11b)
White solid in 67% yield. m.p. 141.6-143.2 °C. ESI-MS m/z: [M+H]+ 431.1. 1H NMR (400 MHz, CDCl3) δ 8.86 (s,
1H), 8.74 (s, 1H), 7.87 (s, 1H), 7.23 (s, 1H), 7.22 – 7.18 (m, 1H), 6.97 (dd, J = 5.7, 3.5 Hz, 2H), 6.95 – 6.86 (m, 1H),
6.86 – 6.80 (m, 1H), 5.99 – 5.92 (m, 1H), 5.17 (m, J = 5.8 Hz, 1H), 4.15 (d, J = 10.6 Hz, 1H), 4.07 – 4.02 (m, 4H), 3.93
(ddd, J = 13.3, 8.6, 4.4 Hz, 1H), 2.93 (t, J = 6.6 Hz, 2H), 2.42 (dt, J = 14.1, 7.8 Hz, 1H), 2.28 – 2.20 (m, 1H), 2.09 – 2.02
(m, 2H), 1.89 (d, J = 13.7, 6.9 Hz, 3H); 13C NMR (101 MHz, DMSO-d6) δ 165.30, 162.62, 154.05, 153.03, 152.80,
150.24, 142.31, 129.53, 129.06, 128.58, 126.36, 122.83, 120.74, 119.32, 116.04, 113.10, 107.78, 79.41, 72.36, 67.03,
49.25, 32.60, 27.28, 26.78, 23.49, 20.10.
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)pent-2-enamide (11c)
White solid in 65% yield. m.p. 148.6-149.9 °C. ESI-MS m/z: [M+H]+ 445.2. 1H NMR (400 MHz, CDCl3) δ 8.83 (s,
1H), 8.70 (s, 1H), 7.80 (s, 1H), 7.21 (s, 1H), 7.20 – 7.16 (m, 1H), 6.94 (dd, J = 10.7, 4.5 Hz, 3H), 6.83 – 6.78 (m, 1H),
5.86 (d, J = 15.3 Hz, 1H), 5.15 (m, 1H), 4.12 (d, J = 10.8 Hz, 1H), 4.02 (dd, J = 13.7, 7.6 Hz, 5H), 3.92 – 3.85 (m, 1H),
2.90 (t, J = 6.5 Hz, 2H), 2.44 – 2.35 (m, 2H), 2.24 – 2.19 (m, 2H), 2.02 (dt, J = 11.8, 5.9 Hz, 2H), 1.04 (t, J = 7.4 Hz,
3H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-methylpent-2-enamide
(11d)
White solid in 62% yield. m.p. 159.6-161.0 °C. ESI-MS m/z: [M+H]+ 459.3. 1H NMR (400 MHz, DMSO-d6) δ 9.22
(s, 1H), 8.93 (s, 1H), 8.54 (s, 1H), 7.38 (s, 1H), 7.34 (d, J = 7.1 Hz, 1H), 7.12 (t, J = 7.1 Hz, 1H), 7.03 (t, J = 7.6 Hz, 1H),
6.97 (d, J = 8.1 Hz, 1H), 6.73 (dd, J = 15.1, 6.1 Hz, 1H), 6.34 (d, J = 15.5 Hz, 1H), 5.28 (m, 1H), 4.12 (t, 2H), 4.09 (s,

ACCEPTED MANUSCRIPT
1H), 4.03 – 3.98 (m, 1H), 3.95 (t, J = 7.3 Hz, 1H), 3.80 (t, J = 5.1 Hz, 1H), 2.89 (t, 2H), 2.48 – 2.43 (m, 1H), 2.38 (dd, J
= 13.7, 7.0 Hz, 1H), 2.22 (s, 1H), 2.09 – 2.01 (m, 2H), 1.04 (d, J = 2.9 Hz, 3H), 1.03 (s, 3H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)hex-2-enamide (11e)
White solid in 66% yield. m.p. 138.9-140.3 °C. ESI-MS m/z: [M+H]+ 458.1. 1H NMR (400 MHz, DMSO-d6) δ 9.23
(s, 1H), 8.92 (s, 1H), 8.51 (s, 1H), 7.51 (s, 1H), 7.33 (d, J = 7.0 Hz, 1H), 7.11 (t, 1H), 6.99 (dd, J = 15.0, 7.2 Hz, 2H),
6.78 – 6.67 (m, 1H), 6.39 (d, J = 15.3 Hz, 1H), 5.26 (s, 1H), 4.11 (d, J = 5.7 Hz, 2H), 4.08 (s, 1H), 4.01 (s, 1H), 3.94 (t, J
= 7.2 Hz, 2H), 2.88 (t, 2H), 2.44 – 2.34 (m, 1H), 2.25– 2.19 (m, 1H), 2.19 – 2.12 (m, 2H), 2.08 – 2.01 (m, 2H), 1.49 –
1.39 (m, 2H), 0.89 (t, J = 6.7 Hz, 3H); 13C NMR (101 MHz, DMSO-d6) δ 164.57, 162.23, 154.14, 149.98, 146.39, 140.67,
132.41, 129.81, 129.14, 126.94(2C), 126.20, 125.07, 122.84, 117.36, 110.27, 102.85, 80.65, 72.47, 67.34, 49.50, 34.11,
32.94, 26.52, 24.11, 21.71, 14.28.
(E)-4-(dimethylamino)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)but-2-enamide (12a)
White solid in 67% yield. m.p. 148.6-149.1 °C. ESI-MS m/z: [M+H]+ 404.1. 1H NMR (400 MHz, CDCl3) δ 9.26 (s,
1H), 8.64 (s, 1H), 8.10 (s, 1H), 7.63 (d, J = 8.0 Hz, 1H), 7.23 (s, 2H), 7.16 (t, J = 7.5 Hz, 1H), 6.97 (d, J = 7.0 Hz, 1H),
6.95 – 6.91 (m, 1H), 6.28 (d, J = 15.3 Hz, 1H), 4.52 (t, J = 8.0 Hz, 2H), 4.04 (s, 3H), 3.25 (d, J = 5.8 Hz, 2H), 3.20 (t, J =
7.9 Hz, 2H), 2.38 (s, 6H); 13C NMR (101 MHz, DMSO-d6) δ 164.30, 159.57, 154.71, 153.75, 150.84, 145.17, 142.24,
132.99, 127.45, 126.77, 126.60, 125.26, 122.69, 116.25, 115.86, 111.40, 106.72, 60.17, 56.78, 54.27, 45.51(2C), 29.15.
(E)-4-(diethylamino)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)but-2-enamide (12b)
White solid in 65% yield. m.p. 138.8-140.2 °C. ESI-MS m/z: [M+H]+ 432.1. 1H NMR (400 MHz, CDCl3) δ 9.27 (s,
1H), 8.66 (s, 1H), 8.15 (s, 1H), 7.64 (d, J = 8.1 Hz, 1H), 7.25 (d, J = 3.6 Hz, 2H), 7.18 (t, J = 7.7 Hz, 1H), 7.03 – 6.95 (m,
2H), 6.34 (d, J = 15.2 Hz, 1H), 4.53 (t, J = 8.0 Hz, 2H), 4.06 (s, 3H), 3.46 (d, J = 5.4 Hz, 2H), 3.21 (t, J = 7.9 Hz, 2H),
2.74 (dd, J = 7.0 Hz, 4H), 1.17 (t, J = 7.1 Hz, 6H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)-4-(pyrrolidin-1-yl)but-2-enamide (12c)
White solid in 61% yield. m.p. 132.5-134.0 °C. ESI-MS m/z: [M+H]+ 430.2. 1H NMR (400 MHz, CDCl3) δ 9.31 (s,
1H), 8.68 (s, 1H), 8.10 (s, 1H), 7.67 (d, J = 8.1 Hz, 1H), 7.27 (s, 2H), 7.20 (t, J = 7.7 Hz, 1H), 7.02 (dt, J = 14.6, 6.5 Hz,
2H), 6.24 (d, J = 15.3 Hz, 1H), 4.56 (t, J = 8.0 Hz, 2H), 4.07 (s, 3H), 3.35 (d, J = 5.6 Hz, 2H), 3.24 (t, J = 8.0 Hz, 2H),
2.62 (s, 4H), 1.85 (s, 4H); 13C NMR (101 MHz, DMSO-d6) δ 164.49, 159.55, 154.69, 153.72, 150.82, 145.17, 143.03,
132.97, 127.50, 126.76, 125.65, 125.24, 122.66, 116.24, 115.80, 111.40, 106.70, 56.76, 56.65, 54.26, 54.06(2C), 29.15,
23.75(2C).

ACCEPTED MANUSCRIPT
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)-4-(piperidin-1-yl)but-2-enamide (12d)
White solid in 67% yield. m.p. 160.1-161.4 °C. ESI-MS m/z: [M+H]+ 444.2. 1H NMR (400 MHz, CDCl3) δ 9.31 (s,
1H), 8.67 (s, 1H), 8.09 (s, 1H), 7.67 (d, J = 8.0 Hz, 1H), 7.27 (d, J = 1.4 Hz, 2H), 7.20 (t, J = 7.8 Hz, 1H), 7.05 – 6.96 (m,
2H), 6.20 (d, J = 15.3 Hz, 1H), 4.56 (t, J = 8.1 Hz, 2H), 4.07 (s, 3H), 3.24 (t, J = 8.0 Hz, 2H), 3.18 (dd, J = 6.0, 1.4 Hz,
2H), 2.46 (s, 4H), 1.63 (dt, J = 11.1, 5.5 Hz, 4H), 1.45 (d, J = 13.1 Hz, 2H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)-4-(4-methylpiperazin-1-yl)but-2-enamide (12e)
White solid in 62% yield. m.p. 139.3-141.1 °C. ESI-MS m/z: [M+H]+ 459.1. 1H NMR (400 MHz, CDCl3) δ 9.30 (s,
1H), 8.68 (s, 1H), 8.07 (s, 1H), 7.67 (d, J = 8.0 Hz, 1H), 7.28 (s, 2H), 7.20 (t, J = 7.7 Hz, 1H), 7.02 – 6.94 (m, 2H), 6.20
(d, J = 15.3 Hz, 1H), 4.56 (t, J = 8.0 Hz, 2H), 4.07 (s, 3H), 3.23 (dd, J = 14.6, 6.8 Hz, 4H), 2.54 (s, 8H), 2.33 (s, 3H).
(E)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)-4-morpholinobut-2-enamide (12f)
White solid in 65% yield. m.p. 157.9-169.3 °C. ESI-MS m/z: [M+H]+ 446.0. 1H NMR (400 MHz, CDCl3) δ 9.27 (s,
1H), 8.63 (s, 1H), 8.03 (s, 1H), 7.64 (d, J = 8.0 Hz, 1H), 7.23 (d, J = 4.7 Hz, 2H), 7.16 (t, J = 7.7 Hz, 1H), 6.99 – 6.90 (m,
2H), 6.18 (d, J = 15.2 Hz, 1H), 4.53 (t, J = 7.9 Hz, 2H), 4.03 (d, J = 4.5 Hz, 3H), 3.73 – 3.69 (m, 4H), 3.19 (dd, J = 17.3,
7.1 Hz, 4H), 2.47 (s, 4H).
(E)-4-(4-(hydroxymethyl)piperidin-1-yl)-N-(4-(indolin-1-yl)-7-methoxyquinazolin-6-yl)but-2-enamide (12g)
White solid in 68% yield. m.p. 130.5-131.8 °C. ESI-MS m/z: [M+H]+ 474.2. 1H NMR (400 MHz, DMSO-d6) δ 9.64
(s, 1H), 8.99 (s, 1H), 8.59 (s, 1H), 7.41 (d, J = 8.1 Hz, 1H), 7.31 (s, 1H), 7.28 (d, J = 7.0 Hz, 1H), 7.09 (t, J = 7.5 Hz, 1H),
6.93 (t, J = 7.3 Hz, 1H), 6.74 (d, J = 15.3 Hz, 1H), 6.61 (d, J = 15.3 Hz, 1H), 4.39 (t, J = 7.6 Hz, 2H), 4.03 (s, 3H), 3.24
(d, J = 5.7 Hz, 2H), 3.20 – 3.15 (m, 2H), 3.14 (s, 1H), 2.86 (d, J = 10.3 Hz, 2H), 1.96 (d, J = 12.9 Hz, 2H), 1.64 (d, J =
12.1 Hz, 2H), 1.34 (s, 1H), 1.19 (t, J = 15.3 Hz, 4H); 13C NMR (101 MHz, DMSO-d6) δ 163.84, 159.02, 154.20, 153.24,
150.30, 144.61, 142.12, 132.49, 126.91, 126.27, 125.82, 124.76, 122.21, 115.78, 115.40, 110.83, 106.19, 65.92(2C),
58.93, 56.27, 53.79, 53.27(2C), 38.17, 28.69(2C).
(R,E)-4-(dimethylamino)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)but-2-enamide (13a)
1
White solid in 66% yield. m.p. 144.2-146.0 °C. H NMR (400 MHz, CDCl3) δ 9.35 (s, 1H), 8.66 (s, 1H), 8.14 (s,
1H), 7.69 (d, J = 8.1 Hz, 1H), 7.27 (s, 1H), 7.19 (d, J = 10.7 Hz, 2H), 7.03 – 6.95 (m, 2H), 6.26 (d, J = 15.3 Hz, 1H), 5.20
(s, 1H), 4.56 (t, J = 8.0 Hz, 2H), 4.20 (d, J = 10.7 Hz, 1H), 4.13 – 4.02 (m, 2H), 3.95 (td, J = 8.5, 5.0 Hz, 1H), 3.23 (dd, J
= 14.2, 6.6 Hz, 4H), 2.44 (dd, J = 14.1, 6.3 Hz, 1H), 2.35 (s, 6H), 2.29 (dd, J = 13.1, 6.7 Hz, 1H); 13C NMR (101 MHz,

ACCEPTED MANUSCRIPT
DMSO-d6) δ 163.75, 159.01, 153.27, 152.02, 150.09, 144.59, 142.03, 132.50, 127.07, 126.26, 126.01, 124.75, 122.24,
115.88, 115.67, 110.82, 107.51, 78.95, 71.90, 66.60, 59.63, 53.79, 45.04(2C), 32.31, 28.68. HRMS (ESI): m/z: [M+H]+
calcd for C26H29N5O3, 460.2270; found 460.2265.
(R,E)-4-(diethylamino)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)but-2-enamide (13b)
White solid in 67% yield. m.p. 140.7-142.2 °C. ESI-MS m/z: [M+H]+ 488.0. 1H NMR (400 MHz, CDCl3) δ 9.35 (s,
1H), 8.66 (s, 1H), 8.01 (s, 1H), 7.70 (d, J = 8.1 Hz, 1H), 7.28 (s, 1H), 7.23 – 7.17 (m, 2H), 7.08 – 6.97 (m, 2H), 6.20 (d, J
= 15.3 Hz, 1H), 5.20 (s, 1H), 4.57 (t, J = 8.0 Hz, 2H), 4.19 (d, J = 10.7 Hz, 1H), 4.13 – 4.04 (m, 2H), 3.96 (td, J = 8.5,
5.1 Hz, 1H), 3.31 (d, J = 5.7 Hz, 2H), 3.24 (t, J = 7.9 Hz, 2H), 2.58 (q, J = 7.1 Hz, 4H), 2.44 (dt, J = 14.1, 7.6 Hz, 1H),
2.33 – 2.24 (m, 1H), 1.07 (t, J = 7.1 Hz, 6H).
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(pyrrolidin-1-yl)but-2-enamide (13c)
White solid in 64% yield. m.p. 137.4-139.8 °C. ESI-MS m/z: [M+H]+ 486.1. 1H NMR (400 MHz, CDCl3) δ 9.35 (s,
1H), 8.66 (s, 1H), 8.09 (s, 1H), 7.69 (d, J = 8.0 Hz, 1H), 7.28 (s, 1H), 7.23 – 7.17 (m, 2H), 7.10 – 6.97 (m, 2H), 6.23 (d, J
= 15.3 Hz, 1H), 5.20 (t, J = 5.2 Hz, 1H), 4.56 (t, J = 8.0 Hz, 2H), 4.19 (d, J = 10.7 Hz, 1H), 4.14 – 4.03 (m, 2H), 3.95 (td,
J = 8.5, 5.0 Hz, 1H), 3.33 (dd, J = 5.8, 1.5 Hz, 2H), 3.24 (t, J = 8.0 Hz, 2H), 2.60 (s, 4H), 2.44 (td, J = 14.1, 8.0 Hz, 1H),
2.29 (dd, J = 13.2, 6.7 Hz, 1H), 1.84 (dd, J = 6.6, 3.2 Hz, 4H); 13C NMR (101 MHz, DMSO-d6) δ 153.27, 150.09, 144.61,
142.70, 132.51, 127.11, 126.27, 125.18, 124.76, 122.46, 122.24, 115.82(2C), 107.54, 78.96, 78.28, 71.90, 66.60, 56.13,
53.78, 53.58(2C), 46.06, 32.31, 28.67, 28.41, 23.23(2C).
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(piperidin-1-yl)but-2-enamide (13d)
White solid in 66% yield. m.p. 149.9-152.8 °C. ESI-MS m/z: [M+H]+ 500.1. 1H NMR (400 MHz, CDCl3) δ 10.25 (s,
1H), 9.11 (s, 1H), 8.62 (s, 1H), 7.39 (d, J = 7.9 Hz, 2H), 7.31 (d, J = 7.5 Hz, 1H), 7.10 (d, J = 7.3 Hz, 1H), 6.98 – 6.92 (m,
1H), 5.45 (s, 1H), 4.39 (d, J = 7.8 Hz, 2H), 4.05 (d, J = 10.4 Hz, 1H), 3.96 – 3.85 (m, 4H), 3.19 (d, J = 7.8 Hz, 4H), 2.60
(s, 4H), 2.12 (s, 1H), 1.05 (d, J = 6.8 Hz, 6H).
(R,E)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(4-methylpiperazin-1-yl)but-2-enamide
(13e)
White solid in 69% yield. m.p. 148.1-149.2 °C. ESI-MS m/z: [M+H]+ 515.3. 1H NMR (400 MHz, CDCl3) δ 9.33 (s,
1H), 8.66 (s, 1H), 7.99 (s, 1H), 7.67 (d, J = 8.1 Hz, 1H), 7.26 (s, 1H), 7.20 (d, J = 3.3 Hz, 2H), 7.02 – 6.93 (m, 2H), 6.18
(d, J = 15.3 Hz, 1H), 5.20 (s, 1H), 4.55 (t, J = 8.0 Hz, 2H), 4.19 (d, J = 10.6 Hz, 1H), 4.13 – 4.03 (m, 2H), 3.95 (td, J =
8.5, 5.1 Hz, 1H), 3.23 (t, J = 6.5 Hz, 4H), 2.62 (s, 8H), 2.46 (dd, J = 14.0, 7.9 Hz, 1H), 2.39 (s, 3H), 2.29 (dd, J = 13.3,

ACCEPTED MANUSCRIPT
6.3 Hz, 1H).
(R,E)-4-(4-(hydroxymethyl)piperidin-1-yl)-N-(4-(indolin-1-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)but-2-ena
mide (13f)
White solid in 67% yield. m.p. 134.5-135.8 °C. ESI-MS m/z: [M+H]+ 530.1. 1H NMR (400 MHz, DMSO-d6) δ 9.36
(s, 1H), 9.03 (s, 1H), 8.57 (s, 1H), 7.43 (d, J = 8.0 Hz, 1H), 7.28 (d, J = 7.3 Hz, 1H), 7.24 (s, 1H), 7.10 (t, J = 7.6 Hz, 1H),
6.94 (t, J = 7.3 Hz, 1H), 6.81 – 6.72 (m, 1H), 6.60 (d, J = 15.3 Hz, 1H), 5.29 (s, 1H), 4.43 (s, 1H), 4.39 (t, J = 7.8 Hz,
2H), 4.07 (d, J = 10.0 Hz, 1H), 4.03 – 3.91 (m, 2H), 3.78 (dd, J = 13.0, 7.8 Hz, 1H), 3.24 (d, J = 6.0 Hz, 2H), 3.17 (t, J =
7.7 Hz, 2H), 3.11 (d, J = 5.4 Hz, 2H), 2.84 (d, J = 10.2 Hz, 2H), 2.35 (td, J = 14.1, 7.4 Hz, 1H), 2.24 – 2.14 (m, 1H), 1.92
(t, J = 10.9 Hz, 2H), 1.63 (d, J = 12.1 Hz, 2H), 1.33 (s, 1H), 1.21 – 1.10 (m, 2H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-4-(dimethylamino)but-2-enamidee (14a)
1
White solid in 60% yield. m.p. 144.4-147.6 °C. H NMR (400 MHz, CDCl3) δ 8.87 (s, 1H), 8.76 (s, 1H), 8.01 (s,
1H), 7.26 (s, 1H), 7.21 – 7.18 (m, 1H), 6.97 (dd, J = 6.6, 3.1 Hz, 2H), 6.89 (dt, J = 15.4, 5.9 Hz, 1H), 6.84 – 6.81 (m, 1H),
6.14 (d, J = 15.4 Hz, 1H), 4.05 (d, J = 8.8 Hz, 5H), 3.11 (d, J = 5.6 Hz, 2H), 2.94 (t, J = 6.6 Hz, 2H), 2.28 (s, 6H), 2.05 (s,
2H); 13C NMR (101 MHz, DMSO-d6) δ 163.52, 162.22, 154.40, 153.76, 150.36, 141.98, 141.74, 129.10, 128.04, 127.76,
125.97, 125.81, 122.02, 120.03, 115.39, 112.79, 106.27, 59.66, 56.32, 48.77, 45.04(2C), 26.37, 23.00. HRMS (ESI): m/z:
[M+H]+ calcd for C24H27N5O2, 418.0197; found 418.0201.
(E)-4-(diethylamino)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)but-2-enamide (14b)
White solid in 65% yield. m.p. 136.7-139.2 °C. ESI-MS m/z: [M+H]+ 446.3. 1H NMR (400 MHz, CDCl3) δ 8.86 (s,
1H), 8.77 (s, 1H), 8.07 (s, 1H), 7.27 (s, 1H), 7.21 (d, J = 5.7 Hz, 1H), 6.96 (dd, J = 7.8, 4.4 Hz, 2H), 6.91 (t, J = 7.7 Hz,
1H), 6.82 (d, J = 8.9 Hz, 1H), 6.23 (d, J = 15.2 Hz, 1H), 4.04 (d, J = 7.6 Hz, 5H), 3.39 (d, J = 6.1 Hz, 2H), 2.95 (t, J = 6.5
Hz, 2H), 2.68 (dd, J = 14.0, 6.9 Hz, 4H), 2.09 – 2.04 (m, 2H), 1.13 (t, J = 7.1 Hz, 6H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-4-(pyrrolidin-1-yl)but-2-enamide (14c)
White solid in 68% yield. m.p. 156.3-158.2 °C. ESI-MS m/z: [M+H]+ 444.1. 1H NMR (400 MHz, CDCl3) δ 8.86 (s,
1H), 8.75 (s, 1H), 8.02 (s, 1H), 7.25 (s, 1H), 7.21 – 7.17 (m, 1H), 6.98 – 6.89 (m, 3H), 6.82 (dd, J = 7.3, 2.0 Hz, 1H),
6.15 (d, J = 15.4 Hz, 1H), 4.04 (d, J = 9.4 Hz, 5H), 3.28 (dd, J = 5.8, 1.5 Hz, 2H), 2.93 (t, J = 6.6 Hz, 2H), 2.57 (s, 4H),
2.09 – 2.01 (m, 2H), 1.81 (dt, J = 6.5, 3.2 Hz, 4H); 13C NMR (101 MHz, DMSO-d6) δ 163.70, 162.20, 154.40, 153.74,
150.33, 142.36, 141.99, 129.08, 128.04, 127.81, 125.81, 125.11, 122.02, 120.02, 115.35, 112.80, 106.25, 56.31, 56.10,
53.56(2C), 48.76, 26.37, 23.20(2C), 23.00.

ACCEPTED MANUSCRIPT
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-4-(piperidin-1-yl)but-2-enamide (14d)
1
White solid in 61% yield. m.p. 125.9-127.6 °C. H NMR (400 MHz, CDCl3) δ 8.86 (s, 1H), 8.76 (s, 1H), 8.01 (s,
1H), 7.26 (s, 1H), 7.19 (d, J = 6.1 Hz, 1H), 6.98 – 6.94 (m, 2H), 6.90 (dd, J = 15.1, 6.3 Hz, 1H), 6.82 (d, J = 7.3 Hz, 1H),
6.11 (d, J = 15.5 Hz, 1H), 4.05 (d, J = 6.6 Hz, 5H), 3.13 (d, J = 5.9 Hz, 2H), 2.93 (t, J = 6.4 Hz, 2H), 2.41 (s, 4H), 2.09 –
2.02 (m, 2H), 1.63 – 1.56 (m, 4H), 1.44 (s, 2H); 13C NMR (101 MHz, DMSO-d6) δ 163.56, 162.20, 154.45, 153.76,
150.36, 142.00(2C), 129.09, 128.06, 127.77, 125.82(2C), 122.03, 120.00, 115.48, 112.81, 106.28, 59.23, 56.32,
54.05(2C), 48.75, 26.37, 25.49(2C), 23.80, 23.00. HRMS (ESI): m/z: [M+H]+ calcd for C24H27N5O2, 458.02477; found
458.0251.
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-4-(4-methylpiperazin-1-yl)but-2-enamide (14e)
White solid in 66% yield. m.p. 143.9-148.3 °C. ESI-MS m/z: [M+H]+ 473.1. 1H NMR (400 MHz, CDCl3) δ 8.86 (s,
1H), 8.76 (s, 1H), 7.98 (s, 1H), 7.27 (s, 1H), 7.21 – 7.18 (m, 1H), 6.99 – 6.95 (m, 2H), 6.93 – 6.81 (m, 2H), 6.12 (d, J =
15.4 Hz, 1H), 4.07 – 4.03 (m, 5H), 3.17 (d, J = 6.1 Hz, 2H), 2.94 (t, J = 6.6 Hz, 2H), 2.56 (s, 8H), 2.35 (s, 3H), 2.09 –
2.02 (m, 2H).
(E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-methoxyquinazolin-6-yl)-4-(4-(hydroxymethyl)piperidin-1-yl)but-2-enamid
e (14f)
White solid in 64% yield. m.p. 131.1-134.5 °C. ESI-MS m/z: [M+H]+ 488.4. 1H NMR (400 MHz, DMSO-d6) δ 9.58
(s, 1H), 8.72 (s, 1H), 8.57 (s, 1H), 7.35 (s, 1H), 7.19 (d, J = 4.7 Hz, 1H), 6.92 – 6.88 (m, 2H), 6.73 – 6.65 (m, 1H), 6.58
(d, J = 15.7 Hz, 2H), 4.48 (d, J = 24.6 Hz, 1H), 4.03 (s, 3H), 3.89 (t, J = 5.6 Hz, 2H), 3.24 (d, J = 5.9 Hz, 4H), 2.95 (d, J
= 10.8 Hz, 2H), 2.87 (t, J = 6.4 Hz, 2H), 2.15 (s, 2H), 1.97 (d, J = 5.9 Hz, 2H), 1.67 (d, J = 12.8 Hz, 2H), 1.39 (s, 1H),
1.22 (s, 2H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(dimethylamino)but-2-ena
mide (15a)
White solid in 67% yield. m.p. 151.2-152.5 °C. ESI-MS m/z: [M+H]+ 474.1. 1H NMR (400 MHz, CDCl3) δ 8.96 (s,
1H), 8.82 (s, 1H), 8.07 (s, 1H), 7.32 (s, 1H), 7.28 (s, 1H), 7.06 – 7.02 (m, 2H), 6.96 (dt, J = 15.3, 5.9 Hz, 1H), 6.91 – 6.88
(m, 1H), 6.23 (d, J = 15.4 Hz, 1H), 5.24 (s, 1H), 4.22 (d, J = 10.6 Hz, 1H), 4.15 – 4.08 (m, 4H), 4.00 (td, J = 8.5, 5.1 Hz,
1H), 3.22 (d, J = 5.8 Hz, 2H), 3.01 (t, J = 6.6 Hz, 2H), 2.47 (dt, J = 14.3, 7.0 Hz, 1H), 2.38 (s, 6H), 2.33 (dd, J = 13.5, 6.5
Hz, 1H), 2.12 (dt, J = 12.2, 6.0 Hz, 2H); 13C NMR (101 MHz, DMSO-d6) δ 163.50, 162.22, 153.78, 152.28, 150.13,
142.09(2C), 129.10, 128.11, 127.90, 125.82(2C), 122.09, 120.14, 115.76, 112.74, 107.60, 78.99, 71.87, 66.58, 59.63,

ACCEPTED MANUSCRIPT
48.80, 45.10(2C), 32.28, 26.36, 23.00.
(R,E)-4-(diethylamino)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)but-2-enam
ide (15b)
White solid in 62% yield. m.p. 143.2-145.6 °C. ESI-MS m/z: [M+H]+ 502.1. 1H NMR (400 MHz, CDCl3) δ 8.89 (s,
1H), 8.75 (s, 1H), 8.05 (s, 1H), 7.21 (d, J = 5.6 Hz, 2H), 6.99 – 6.96 (m, 2H), 6.95 – 6.89 (m, 1H), 6.85 – 6.81 (m, 1H),
6.20 (d, J = 15.2 Hz, 1H), 5.18 (s, 1H), 4.17 (d, J = 10.7 Hz, 1H), 4.05 (dt, J = 6.3, 5.0 Hz, 4H), 3.93 (td, J = 8.5, 5.1 Hz,
1H), 3.33 (d, J = 5.9 Hz, 2H), 2.94 (t, J = 6.6 Hz, 2H), 2.62 (q, J = 7.1 Hz, 4H), 2.42 (dd, J = 13.9, 6.2 Hz, 1H), 2.30 –
2.25 (m, 1H), 2.08 – 2.04 (m, 2H), 1.09 (t, J = 7.1 Hz, 6H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(pyrrolidin-1-yl)but-2-ena
mide (15c)
White solid in 66% yield. m.p. 150.1-152.0 °C. ESI-MS m/z: [M+H]+ 500.2. 1H NMR (400 MHz, CDCl3) δ 8.89 (s,
1H), 8.75 (s, 1H), 8.08 (s, 1H), 7.20 (d, J = 6.7 Hz, 2H), 6.99 – 6.89 (m, 3H), 6.86 – 6.81 (m, 1H), 6.20 (d, J = 15.4 Hz,
1H), 5.18 (s, 1H), 4.17 (d, J = 10.6 Hz, 1H), 4.11 – 4.01 (m, 4H), 3.93 (td, J = 8.5, 5.0 Hz, 1H), 3.34 (d, J = 5.8 Hz, 2H),
2.94 (t, J = 6.6 Hz, 2H), 2.64 (s, 4H), 2.45 – 2.37 (m, 1H), 2.25 (d, J = 7.0 Hz, 1H), 2.06 (dt, J = 12.6, 6.5 Hz, 2H), 1.84
(s, 4H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(piperidin-1-yl)but-2-ena
mide (15d)
White solid in 68% yield. m.p. 162.1-163.2 °C. ESI-MS m/z: [M+H]+ 514.1. 1H NMR (400 MHz, CDCl3) δ 8.88 (s,
1H), 8.75 (s, 1H), 8.01 (s, 1H), 7.20 (s, 2H), 6.96 (dd, J = 8.1, 4.2 Hz, 2H), 6.94 – 6.87 (m, 1H), 6.85 – 6.81 (m, 1H),
6.15 (d, J = 15.2 Hz, 1H), 5.17 (s, 1H), 4.16 (d, J = 10.6 Hz, 1H), 4.09 – 4.02 (m, 4H), 3.93 (dt, J = 13.4, 6.8 Hz, 1H),
3.19 (d, J = 6.1 Hz, 2H), 2.94 (t, J = 6.6 Hz, 2H), 2.48 (s, 2H), 2.40 (dd, J = 14.1, 7.7 Hz, 1H), 2.27 (dd, J = 13.2, 6.5 Hz,
1H), 2.09 – 2.01 (m, 2H), 1.67 – 1.60 (m, 4H), 1.47 (d, J = 13.5 Hz, 2H), 0.87 (dd, J = 9.5, 5.8 Hz, 2H).
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(4-methylpiperazin-1-yl)b
ut-2-enamide (15e)
White solid in 67% yield. m.p. 141.2-142.7 °C. ESI-MS m/z: [M+H]+ 529.1. 1H NMR (400 MHz, CDCl3) δ 8.89 (s,
1H), 8.76 (s, 1H), 7.92 (s, 1H), 7.20 (s, 2H), 7.01 – 6.95 (m, 2H), 6.93 – 6.81 (m, 2H), 6.12 (d, J = 15.3 Hz, 1H), 5.18 (s,
1H), 4.17 (d, J = 10.4 Hz, 1H), 4.06 (dd, J = 12.5, 7.1 Hz, 4H), 3.94 (dd, J = 13.6, 8.5 Hz, 1H), 3.20 (d, J = 6.0 Hz, 2H),
2.94 (t, J = 6.5 Hz, 2H), 2.64 (s, 8H), 2.42 (s, 3H), 2.29 – 2.24 (m, 1H), 2.10 – 2.02 (m, 3H).

ACCEPTED MANUSCRIPT
(R,E)-N-(4-(3,4-dihydroquinolin-1(2H)-yl)-7-((tetrahydrofuran-3-yl)oxy)quinazolin-6-yl)-4-(4-(hydroxymethyl)piperid
in-1-yl)but-2-enamide (15f)
White solid in 69% yield. m.p. 130.6-131.9 °C. ESI-MS m/z: [M+H]+ 544.2. 1H NMR (400 MHz, DMSO-d6) δ 9.28
(s, 1H), 8.71 (s, 1H), 8.60 (s, 1H), 7.29 (s, 1H), 7.18 (d, J = 4.8 Hz, 1H), 6.93 – 6.87 (m, 2H), 6.73 – 6.65 (m, 1H), 6.64 –
6.60 (m, 1H), 6.55 (d, J = 15.3 Hz, 1H), 5.30 (s, 1H), 4.48 (s, 1H), 4.01 (dt, J = 10.3, 7.4 Hz, 2H), 3.97 – 3.91 (m, 1H),
3.89 (t, J = 5.5 Hz, 2H), 3.77 (dd, J = 13.1, 7.8 Hz, 1H), 3.24 (d, J = 5.9 Hz, 2H), 3.17 (s, 2H), 2.87 (t, J = 6.2 Hz, 4H),
2.35 (td, J = 14.2, 7.2 Hz, 1H), 2.22 – 2.14 (m, 1H), 2.05 – 1.92 (m, 4H), 1.64 (d, J = 12.1 Hz, 2H), 1.36 (d, J = 10.8 Hz,
1H), 1.18 (dd, J = 15.1, 7.5 Hz, 2H).
4. 23 antiproliferative assay
The cytotoxic activities of target compounds (8a–g, 9a–g, 10a–e, 11a–e, 12a–g, 13a–f, 14a–f and 15a–f) were
evaluated with A549, MCF-7 and PC-3 cancer cell lines by the standard MTT assay, with compounds EGFR inhibitors
afatinib as positive control. The cancer cell lines were cultured in minimum essential medium (MEM) supplement with
10% fetal bovine serum (FBS). Approximately 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 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.
4. 24 EGFR kinase assay
The target compounds 8e, 9d, 12c, 12d, 12g, 13a and 14a are tested for their activity against EGFR kinase through
the mobility shift assay. All kinases assays were performed in 96-well plates in a 50 µL reaction volume. The kinase
buffer contains 50 mM HEPES, pH 7.5, 10 mM MgCl2, 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 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