Design, synthesis and bioactivities evaluation of novel quinazoline analogs containing oxazole units

Article abstract of DOI:10.1002/cjoc.201400271

A novel type of quinazoline derivatives, which were designed by the combination of quinazoline as the backbone and oxazole scaffold as the substituent, have been synthesized and their biological activities were evaluated for anti-proliferative activities and EGFR inhibitory potency. Compound 12b demonstrated the most potent inhibitory activity (IC ₅₀=0.95 μmol/L for EGFR), which could be optimized as a potential EGFR inhibitor in the further study. The structures of the synthesized quinazoline analogs and all intermediates were comfirmed by ¹H and ¹³C NMR, 2D NMR spectra, IR spectra and MS spectra. Copyright

Full text of DOI:10.1002/cjoc.201400271

FULL PAPER
DOI: 10.1002/cjoc.201400271
Design Synthesis and Bioactivities Evaluation of Novel
Quinazoline Analogs Containing Oxazole Units
Xuehui Hou,a,† Jingyu Zhang,b,† Xuan Zhao,b Liming Chang,b Ping Hu,*,a and Hongmin Liu*,c
a Department of Quality Detection and Management, Henan University of Animal Husbandry and
Economy, Zhengzhou, Henan 450011, China
b School of Pharmaceutical Science, Henan University of TCM, Zhengzhou, Henan 450008, China
c School of Pharmaceutical Science, Zhengzhou University, Zhengzhou, Henan 450001, China
A novel type of quinazoline derivatives, which were designed by the combination of quinazoline as the back-
bone and oxazole scaffold as the substituent, have been synthesized and their biological activities were evaluated
for anti-proliferative activities and EGFR inhibitory potency. Compound 12b demonstrated the most potent inhibi-
tory activity (IC50＝0.95 μmol/L for EGFR), which could be optimized as a potential EGFR inhibitor in the further
1
study. The structures of the synthesized quinazoline analogs and all intermediates were comfirmed by H and 13C
NMR, 2D NMR spectra, IR spectra and MS spectra.
Keywords quinazoline, synthesis, anticancer activity
Introduction
breast and prostate cancer and nonsmall cell lung carci-
noma. In addition, coexpression of EGFR and Her-2 has
been found in various cancers such as breast, ovarian,
colon and prostate cancers, and is associated with poor
prognosis of the patients. Therefore, the EGFR has be-
come an attractive target for the design and develop-
ment of potential anticancer drugs.
Over the past decades, there has been a growing in-
terest in the use of EGFR-TK inhibitors, such as quina-
zoline derivatives. In our research for the development
of novel quinazoline inhibitors of tyrosine kinase activ-
ity, we have prepared novel quinazoline derivatives and
investigated their bioactivity. In our effort to develop
new quinazoline derivatives as effective anticancer
agents or EGFR kinase inhibitors, we have focused on
the molecules possessing oxazole scaffold. Thus, we
herein report the syntheses characterization and pre-
liminary biological evaluation of the novel quinazoline
analogues 12 (a－d) prepared under mild conditions,
which is a new and practical synthetic route of prepar-
ing quinazoline analogues containing oxazole unit
(Scheme 1).
Cancer is advancing to be a leading health problem
in developed as well as developing countries.[1-9] Sur-
passing heart diseases; it is taking the position of num-
ber one killer due to various worldwide factors. The
enormous cancer incidence increases the search for new,
safer and efficient anticancer agents, aiming at the pre-
vention or the cure of this illness. Quinazoline analogs
have been identified as new classes of cancer chemo-
therapeutic agents with significant therapeutic efficacy
against solid tumors.
As an important pharmacophore, quinazoline has a
variety of biological activities,[10-13] such as anticancer,
antibacterial, anti-inflammatory and so on. Anticancer
agents have been widely used in clinical therapy. FDA
has approved several quinazoline derivatives as anti-
cancer drugs, such as Gefitinib, Erlotinib,[14] Lapatinib[15]
and others (Figure 1). Based on the good performances
of quinazoline derivatives in anticancer application, de-
velopment of novel quinazoline derivatives as antican-
cer drugs is a promising field. Several irreversible in-
hibitors, such as afatinib and dacomitinib (Figure 1), are
currently in clinical trials. In the last few years, a large
structural variety of compounds,[16-18] were reported as
epidermal growth factors receptor (EGFR) tyrosine
kinase (TK) inhibitors. The EGFR belongs to the ErbB
family of receptor tyrosine kinases (TK) involved in the
proliferation of normal and malignant cells. EGFR is
often overexpressed in many tumour cells, namely in
Experimental
Apparatus and materials
The starting materials used in the present study were
of AR grade and purchased and used without further
purification. Melting point was measured on an XT5A
*
E-mail: liuhmwork2014@163.com; Tel.: 0086-0371-67781739; Fax: 0086-0371-67781739.
Received April 28, 2014; accepted June 3, 2014; published online XXXX, 2014.
Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/cjoc.201400271 or from the author.
† The authors contributed equally to this work and should be considered co-first authors.
Chin. J. Chem. 2014, XX, 1—7
© 2014 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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Hou et al.
FULL PAPER
F
F
O
HN
N
O
HN
N
Cl
N
HN
N
Cl
O
N
O
O
HN
O
O
O
N
N
N
HCl
O
Gefitinib
Erlotinib
Dacomitinib
F
O
O
N
HN
N
Cl
F
HN
O
O
S
N
HN
N
Cl
H
N
N
O
O
O
Lapatinib
Afatinib
Figure 1 Chemical structures of several EGFR-TK inhibitors.
Scheme 1 Synthetic pathway of quinazoline analogs 12a－12d
Cl
N
O
N
OH
O2N
160 oC, 2 h
NH2
O2N
COOH
NH2
O
O2N
O2N
SOCl2
N
NH
N
reflux, 3 h
78%
H
82%
N
2
4
1
3
F
S
S
N
N
O2N
Cl
OH
O
N
O
N
S
7
CH3OH
Zn/NH4Cl
CHO
NaBH4
CH3OH, r.t.
2 h, 99%
NaH/DMF
0 oC to r.t., 3 h
86%
S
0 oC to r.t., 1 h
95%
H2N
Cl
O2N
Cl
6
5
8
9
S
S
S
N
N
N
O
ThioCDI, Amino alcohol
THF, NaOH/TsCl
Zn, NH4Cl
O
THF,
O
4 + 9
CH3OH, r.t,
2 h, 99%
reflux, 3 h
78%
HN
N
Cl
H
HN
Cl
HN
N
Cl
N
N
O2N
R
H2N
N
N
N
12
10
11
O
O
O
O
N
N
O
N
O
N
R =
12c
12a
61%
12b
65%
12d
60%
52%
1
apparatus and was uncorrected. H NMR (500 MHz)
and 13C NMR (125 MHz) spectra were acquired on a
Bruker AVANCE DPX-500 spectrometer with chemical
shifts (δ) relative to tetramethylsilane as the internal
standard (δ＝0.00). Data were reported as follows: s,
singlet; d, doublet; t, triplet; q, quartet; m, multiplet; dd,
double doublet; br s, broad singlet. Infrared spectra were
recorded on Nicolet IR200 instrument using KBr disks
in the 400－4000 cm−1 region. High resolution mass
spectra (HRMS) were obtained on a Waters Micromass
Q-Tof MicroTM instrument using the ESI technique.
Optical rotations were determined on a Perkin Elmer
341 polarimeter at 20 ℃ in MeCN. Reaction progress
was monitored using analytical thin-layer chromatogra-
2
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© 2014 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Chin. J. Chem. 2014, XX, 1—7

Design Synthesis and Bioactivities Evaluation of Novel Quinazoline Analogs
phy (TLC) on 0.25 mm F-254 silica gel glass plates.
Visualization was achieved by UV light (254 nm).
Room temperatures are within 20－30 ℃. The purity of
the target compounds (10, 11, 12a－12d) was deter-
mined by analytical HPLC and was more than 95%.
6-Nitro-4-hydroxyquinazoline (3) 2-Amino-5-
nitrobenzoic acid (5.46 g, 30 mmol) was added to a 250
mL flask equipped with a reflux condenser. Then 50 mL
formamide was added. The mixture was heated with
vigorous stirring at 160 ℃ for 3 h. After cooling the
solution was poured in ice-water to give 3 in almost
pure form (Yellow solid 4.70 g, yield 82.0%). m.p.
12.16; found C 41.74, H 4.33, N 12.18; MS (ESI) m/z:
116.11 (M＋H).
2-((2-Chloro-4-nitrophenoxy)methyl)thiazole (8)
2-(2-Chloro-4-nitro-phenoxymethy1)-thiazole was pre-
pared by adding thiazol-2-yl-methanol (5.48 g, 47.65
mmol) to a slurry of sodium hydride (2.42 g of a 60%
dispersion in oil, 60.5 mmol) in THF (50 mL) at 0 ℃.
After several minutes, 2-chloro-1-fluoro-4-nitrobenzene
(7.58 g, 43.60 mmol) was added and the reaction mix-
ture was warmed to room temperature. The reaction
mixture was stirred at room temperature for 3 h, and at
60 ℃ for 16 h. After cooling to room temperature, the
reaction mixture was poured into 300 mL water. The
resulting precipitate was collected by filtration, washed
with water, and dried in vacuo to give 2-(2-chloro-4-
nitrophenoxymethy1)-thiazole (11.06 g, 86%) which
was used in the next step without further purification.
m.p. 170－171 ℃; 1H NMR (DMSO-d6) δ: 8.35 (d, J＝
2.8 Hz, 1H), 8.25 (dd, J＝2.8, 9.15 Hz, 1H), 7.87 (d,
J＝3.3 Hz, 1H), 7.83 (d, J＝3.3 Hz, 1H), 7.54 (d, J＝
9.2 Hz, 1H), 5.73 (s, 2H); 13C NMR (DMSO-d6) δ:
164.2, 158.5, 143.2, 141.7, 125.9, 124.9, 122.4, 122.2,
114.6, 68.4; IR (KBr) ν: 3112, 3009, 1587, 1509, 1500,
1354, 1319, 1284, 1255, 1154, 1125, 1054, 1006, 894,
780, 746, 728 cm−1. Anal. calcd for C10H7N2O3SCl: C
44.37, H 2.61, N 10.35; found C 44.31, H 2.67, N 10.29;
MS (ESI) m/z: 268.89 (M−H).
3-Chloro-4-(thiazol-2-ylmethoxy)aniline (9) In a
flask equipped with a reflux condenser, the compound 8
(15.00 g, 55.6 mmol), reduced zinc powder (14.44 g,
222.0 mmo1, 4 equiv.), saturated ammonia chloride (5
mL) and methanol (100 mL) were mixed. The mixture
was stirred at a temperature of 40 ℃ for 1.5 h. Then
the zinc powder was filtered off, the filtrate was con-
centrated to obtain yellow solid 13.21 g, yield 99%. m.p.
60－61 ℃; 1H NMR (DMSO-d6) δ: 7.80 (d, J＝3.3 Hz,
1H), 7.75 (d, J＝3.3 Hz, 1H), 6.96 (d, J＝8.8 Hz, 1H),
6.64 (d, J＝2.7 Hz, 1H), 6.46 (dd, J＝2.7, 8.7 Hz, 1H),
5.30 (s, 2H), 5.04 (s, 2H, NH2, exchangeable); 13C NMR
(DMSO-d6) δ: 166.8, 145.1, 144.1, 142.80, 123.1, 121.5,
117.7, 115.2, 113.6, 69.1; IR (KBr) ν: 3322, 3192, 3112,
1607, 1499, 1457, 1436, 1291, 1274, 1221, 1191, 1144,
1057, 1027, 857, 797, 767, 733, 584 cm−1. Anal. calcd
for C10H9N2OSCl: C 49.90, H 3.77, N 11.64; found C
49.95, H 3.76, N 11.66; MS (ESI) m/z: 239.01 (M−H).
N-(3-Chloro-4-(thiazol-2-ylmethoxy)phenyl)-6-
nitroquinazolin-4-amine (10) In a flask equipped
with a reflux condenser, 6-nitro-4-chloroquinazoline
(8.0 g, 38.3 mmol) and 3-chloro-4-(thiazol-2-ylmeth-
oxy)aniline (8.9 g, 37.0 mmol) were dissolved into 150
mL of THF, and the solution was refluxed for 3 h. Then
a lot of yellow solid was deposited. Then it was filtered
affording yellow solid 12.8 g, yield 81%. m.p. 183－
184 ℃ (decompose); 1H NMR (DMSO-d6) δ: 11.97 (s,
1H, exchangeable), 9.84 (s, 1H), 9.00 (s, 1H), 8.76 (d,
J＝9.1 Hz, 1H), 8.12－8.14 (m, 1H), 7.94 (d, J＝2.3 Hz,
1H), 7.87 (d, J＝3.2 Hz, 1H), 7.81 (d, J＝3.2 Hz, 1H),
7.44 (d, J＝9.0 Hz, 1H), 7.69 (dd, J＝2.5, 8.9 Hz, 1H),
1
317－318 ℃; H NMR (DMSO-d6) δ: 12.74 (s, 1H,
OH, exchangeable), 8.78 (d, J＝2.4 Hz, 1H), 8.53 (dd,
J＝2.6, 9.0 Hz, 1H), 8.30 (s, 1H), 7.84 (d, J＝9.0 Hz,
1H); 13C NMR (DMSO-d6) δ: 160.1, 152.9, 148.9, 145.0,
129.1, 128.3, 122.7, 121.9; IR (KBr) ν: 3172, 3046,
2879, 1674, 1615, 1577, 1514, 1491, 1469, 1343, 1289,
1242, 1167, 1112, 928, 920, 901, 803, 753, 630, 574,
531 cm−1. Anal. calcd for C8H5N3O3: C 50.27, H 2.64,
N 21.98; found C 50.30, H 2.65, N 21.96; MS (ESI) m/z:
189.97 (M−H).
4-Chloro-6-nitroquinazoline (4) In a 100 mL
flask equipped with a reflux condenser, 6-nitroquinazo-
lin-4-one (2.86 g, 15 mmol) and thionyl chloride (SOCl2,
25 mL) were added. The mixture was heated under re-
flux with vigorous stirring for 2 h. After the solution
was clear, the reaction mixture was heated for another 2
h. Then, 150 mL of ice MeOH was dropped into it
carefully; the mixture was extracted with CH2Cl2. The
organic layer was dried under MgSO4, filtered and the
solvent was removed to give 4-chloro-6-nitroquinazo-
line (4). Yellow solid 2.45 g, yield 78%. m.p. 134－135
1
℃; H NMR (DMSO-d6) δ: 8.80 (d, J＝3.0 Hz, 1H),
8.54 (dd, J＝2.7, 9.0 Hz, 1H), 8.35 (s, 1H), 7.87 (d, J＝
9.0 Hz, 1H); 13C NMR (DMSO-d6) δ: 160.0, 152.5,
149.1, 145.1, 128.7, 128.4, 122.7, 122.0; IR (KBr) ν:
3431, 3082, 3038, 2664, 2613, 2567, 1724, 1685, 1676,
1646, 1617, 1578, 1526, 1468, 1359, 1346, 1269 cm−1.
Anal. calcd for C8H4N3O2Cl: C 45.84, H 1.92, N 20.05;
found C 45.81, H 1.97, N 20.02; MS (ESI) m/z: 207.96
(M−H).
Thiazol-2-yl-methano1 (6) Sodium borohydride
(16.0 g, 140 mmol) was added to a stirred solution of
thiazole-2-carbaldehyde (24.2 g, 214 mmol) in MeOH
(400 mL) at 0 ℃. The reaction mixture was warmed to
room temperature. After 1 h, the reaction mixture was
quenched by the addition of water and the organics were
removed by concentration. The resulting aqueous mix-
ture was extracted with EtOAc. The combined organic
extracts were dried under Na2SO4 and concentrated to
give thiazol-2-yl-methano1 (23.39 g, 95%). b.p. 75－76
℃ (0.2 mmHg) [lit.[19] b.p. 70－80 ℃ (0.2 mmHg)];
1
m.p. 63－64 ℃. H NMR (CDCl3) δ: 4.91 (s, 2H), 5.1
(br, lH), 7.28 (d, J＝3.2 Hz, 1H), 7.68 (d, J＝2.9 Hz,
1H); IR (KBr) ν: 3135, 3099, 3082, 2814, 1509, 1446,
1351, 1189, 1149, 1073, 1050, 977, 775, 745, 613, 603
cm−1. Anal. calcd for C4H5NOS: C 41.72, H 4.38, N
Chin. J. Chem. 2014, XX, 1—7
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3

Hou et al.
FULL PAPER
5.61 (s, 2H); 13C NMR (DMSO-d6) δ: 166.8, 145.1,
144.1, 142.8, 123.1, 121.5, 117.7, 115.2, 113.7, 69.1; IR
(KBr) ν: 3442, 3100, 1636, 1618, 1570, 1552, 1523,
1492, 1442, 1400, 1377, 1344, 1301, 1267, 1069, 805
cm−1. Anal. calcd for C18H12N5O3SCl: C 52.24, H 2.92,
N 16.92; found C 52.26, H 2.93, N 16.96; MS (ESI) m/z:
412.84 (M−H).
1H), 7.36 (d, J＝3.2 Hz, 1H), 7.36 (s, 1H), 7.24－7.28
(m, 3H), 6.95－6.99 (m, 2H), 6.90 (d, J＝8.0 Hz, 2H),
5.40 (s, 2H), 5.00－5.05 (m, 1H), 4.14 (s, 1H), 4.13 (s,
1H), 4.07 (dd, J＝9.6, 11.4 Hz, 1H), 3.85 (dd, J＝6.6,
11.7 Hz, 1H), 3.64－3.68 (m, 1H); 13C NMR (CDCl3) δ:
166.6, 158.1, 157.3, 156.6 153.5, 152.8, 150.1, 146.5,
146.2, 146.0, 142.6, 133.4, 129.7, 129.3, 128.6, 126.1,
124.5, 123.5, 123.1, 121.7, 121.7, 120.2, 115.6, 114.6,
68.7, 68.5, 64.2, 29.7; IR (KBr) ν: 3423, 3280, 3115,
2922, 1664, 1630, 1617, 1598, 1560, 1529, 1496, 1426,
1401, 1240, 1056 cm−1. Anal. calcd for C28H23N6O3SCl:
C 60.16, H 4.15, N 15.03; found C 60.18, H 4.11, N
15.08; MS (ESI) calcd for m/z: 559.14 (M＋H).
N4-(3-Chloro-4-(thiazol-2-ylmethoxy)phenyl)-
quinazoline-4,6-diamine (11) In a flask equipped
with a reflux condenser, the compound 10 (5.00 g, 12.1
mmol), reduced zinc powder (3.2 g, 48.5 mmo1, 4
equiv.), saturated ammonia chloride (3 mL) and
methanol (60 mL) were mixed. The mixture was stirred
at room temperature for 30 min. Then the zinc powder
was filtered off, the filtrate was concentrated to obtain
yellow solid 4.58 g, yield 98%. m.p. 197－198 ℃
(decompose); 1H NMR (DMSO-d6) δ: 9.33 (s, 1H,
exchangeable), 8.31 (s, 1H), 8.05 (d, J＝2.6 Hz, 1H),
7.85 (d, J＝3.3 Hz, 1H), 7.79 (d, J＝3.3 Hz, 1H), 7.73
(dd, J＝2.5, 9.0 Hz, 1H), 7.51 (d, J＝8.9 Hz, 1H), 7.30
(d, J＝2.4 Hz, 1H), 7.29 (d, J＝4.7 Hz, 1H), 7.23 (dd,
J＝2.3, 8.9 Hz, 1H), 5.57 (s, 2H, exchangeable), 5.52 (s,
2H); 13C NMR (DMSO-d6) δ: 165.9, 155.8, 149.7, 148.5,
147.3, 142.6, 142.5, 134.6, 128.7, 123.6, 123.2, 121.4,
121.3, 121.1, 116.5, 114.7, 100.9, 67.8; IR (KBr) ν:
3443, 3358, 3211, 3100, 1631, 1596, 1577, 1560, 1530,
1494, 1431, 1383, 1217, 910 cm−1. Anal. calcd for
C18H14N5OSCl: C 56.32, H 3.68, N 18.24; found C
56.34, H 3.70, N 18.22; MS (ESI) m/z: 382.66 (M−H).
12b: Employing the same method as above, com-
pound 12b was prepared and the amino alcohol was
(R)-1-amino-3-phenoxypropan-2-ol. Yellow solid, yield
65% (calculated on compound 11). m.p. 89－90 ℃;
1
[α]2D0 ＝−26.6° (c 1.0, CHCl3); H NMR (CDCl3) δ:
8.60 (s, 1H), 8.22 (s, 1H), 7.84 (d, J＝2.2 Hz, 1H), 7.78
(d, J＝3.3 Hz, 1H), 7.73 (d, J＝8.9 Hz, 1H), 7.53 (dd,
J＝3.5, 8.8 Hz, 1H), 7.37 (d, J＝3.2 Hz, 1H), 7.36 (s,
1H), 7.24－7.28 (m, 3H), 6.95－6.99 (m, 2H), 6.90 (d,
J＝8.0 Hz, 2H), 5.40 (s, 2H), 5.00－5.05 (m, 1H), 4.14
(s, 1H), 4.13 (s, 1H), 4.07 (dd, J＝9.6, 11.4 Hz, 1H),
3.85 (dd, J＝6.6, 11.7 Hz, 1H), 3.64－3.68 (m, 1H); 13C
NMR (CDCl3) δ: 166.6, 158.1, 157.3, 153.5, 150.1,
142.6, 133.4, 129.7, 129.3, 124.5, 123.5, 121.7, 120.2,
115.6, 114.6, 68.7, 68.5, 29.7; IR (KBr) ν: 3423, 3280,
3115, 2922, 1664, 1630, 1617, 1598, 1560, 1529, 1496,
1426, 1401, 1240, 1056 cm−1. Anal. calcd for
C28H23N6O3SCl: C 60.16, H 4.15, N 15.03; found C
60.18, H 4.11, N 15.08; MS (ESI) calcd for m/z: 559.14
(M＋H).
Method for the synthesis of quinazoline analogs 12a
to 12d
General procedure for 12 (one pot) ThioCDI
was added to a stirred solution of N4-(3-chloro-4-
(thiazol-2-ylmethoxy)phenyl)quinazoline-4,6-diamine
(11) in THF. After stirring for 3 h, amino alcohol was
added and the reaction mixture was stirred for another 4
h. Then NaOH solution (2.0 mol/L) was added followed
by addition of a TsCl solution (2.0 mol/L) in THF. After
2 h, the reaction mixture was diluted and the organic
phase was dried and concentrated providing the desired
product 12.
12c: Employing the same method as above, com-
pound 12c was prepared and the amino alcohol was
(S)-2-amino-propan-1-ol. Yellow solid, yield 52%. m.p.
1
243－244 ℃; [α]2D0 ＝＋22.5° (c 1.0, CH3CN); H
NMR (DMSO-d6) δ: 9.54 (s, 1H), 8.46 (s, 1H), 8.06 (s,
2H), 7.85 (d, J＝3.3 Hz, 2H), 7.79 (d, J＝3.3 Hz, 2H),
7.75 (d, J＝8.9 Hz, 1H), 7.64 (d, J＝8.3 Hz, 1H), 7.30
(d, J＝9.0 Hz, 1H), 5.54 (s, 2H), 4.73－4.77 (m, 1H),
3.72 (s, 1H), 3.19 (s, 1H), 1.34 (d, J＝6.15 Hz, 3H); 13C
NMR (DMSO-d6) δ: 165.8, 156.9, 152.0, 148.8, 145.3,
142.6, 134.3, 128.7, 128.0, 123.5, 121.7, 121.3, 121.0,
115.6, 114.6, 72.5, 67.7, 63.0, 29.8, 29.0, 20.0, 13.9; IR
(KBr) ν: 3439, 3278, 3101, 2925, 1660, 1631, 1601,
1557, 1500, 1428, 1404, 1384, 1329, 1291, 1257, 1225,
1052 cm−1. Anal. calcd for C22H19N6O2SCl: C 55.59, H
4.10, N 18.00; found C 55.55, H 4.13, N 18.02; MS
(ESI) m/z: 467.2 (M＋H).
12d: Employing the same method as above, com-
pound 12d was prepared and the amino alcohol was
(R)-2-amino-propan-1-ol. Yellow solid, yield 60%. m.p.
242－243 ℃; [α]2D0 ＝−22.3° (c 1.0, CH3CN); 1H
NMR (DMSO-d6) δ: 9.52 (s, 1H), 8.80 (s, 1H), 8.52 (dd,
J＝2.7, 8.9 Hz, 1H), 8.45 (s, 1H), 8.30 (s, 1H), 8.07 (s,
1H), 7.85 (d, J＝3.2 Hz, 1H), 7.79 (d, J＝3.2 Hz, 1H),
7.75 (s, 1H), 7.63 (d, J＝8.2 Hz, 1H), 7.31 (d, J＝9.0
12a: ThioCDI (4.65 g, 26.3 mmol) was added to a
stirred solution of 11 (9.57 g, 25 mmol) in THF (100
mL). After 3 h, (S)-1-amino-3-phenoxypropan-2-ol[20]
(4.18 g, 25 mmol) was added and the reaction mixture
was stirred for 4 h. Solution of NaOH (2 mol/L, 25 mL)
in water was added followed by addition of a 2 mol/L
TsCl solution in THF (14 mL). After 30 min, the reac-
tion mixture was diluted with EtOAc and water, and the
phases separated. The aqueous phase was extracted with
EtOAc and the combined organic extracts were dried
(Na2SO4) and concentrated. Purification by recrystal-
lization provided the desired product 12a (8.50 g,
yellow solid, 61%). m.p. 88－89 ℃; [α]2D0 ＝＋26.4°
1
(c 1.0, CHCl3); H NMR (CDCl3) δ: 8.60 (s, 1H), 8.22
(s, 1H), 7.84 (d, J＝2.2 Hz, 1H), 7.78 (d, J＝3.3 Hz,
1H), 7.73 (d, J＝8.9 Hz, 1H), 7.53 (dd, J＝3.5, 8.8 Hz,
4
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Chin. J. Chem. 2014, XX, 1—7

Design Synthesis and Bioactivities Evaluation of Novel Quinazoline Analogs
Hz, 1H), 5.53 (s, 2H), 4.73－4.77 (m, 1H), 3.81 (s, 1H),
3.19 (s, 1H), 1.34 (d, J＝6.2 Hz, 3H); 13C NMR
(DMSO-d6) δ: 165.8, 156.9, 152.0, 148.8, 145.3, 142.6,
134.3, 128.7, 128.0, 123.5, 121.7, 121.3, 121.0, 115.6,
114.6, 72.5, 67.7, 63.0, 29.8, 29.0, 20.0, 13.9; IR (KBr)
ν: 3439, 3278, 3101, 2925, 1660, 1631, 1601, 1557,
1500, 1428, 1404, 1384, 1329, 1291, 1257, 1225, 1052
cm−1. Anal. calcd for C22H19N6O2SCl: C 55.59, H 4.10,
N 18.00; found C 55.55, H 4.13, N 18.02; MS (ESI) m/z:
467.20 (M＋H).
using the following equation: 100%−[(negative control)/
(positive control−negative control)]. The IC50 was ob-
tained from curves of percentage inhibition with eight
concentrations of compound.
Results and Discussion
Chemistry
Quinazoline derivative 11 was prepared using a
practical synthetic route as depicted in Scheme l. More
specifically, 4-hydroxy-6-nitroquinazoline (3), prepared
by cyclization reaction of 2-amino-5-nitrobenzoic acid 1,
was used as the starting material for the preparation of
4-chloro-6-nitroquinazoline (4). Hence the synthesis of
4 was accomplished according to a slightly modified
reported procedure by treatment of the 4-hydroxy-6-
nitroquinazoline with SOCl2 at 79 ℃.[21] On the other
hand, thiazol-2-ylmethanol (6), was obtained by reduc-
ing thiazole-2-carbaldehyde (5) with sodium boro-
hydride at room temperature. In the next step,
2-((2-chloro-4-nitrophenoxy)methyl)thiazole (8), was
prepared by coupling thiazol-2-ylmethanol to 2-chloro-
1-fluoro-4-nitrobenzene (7). The nitro group of the
compound 8 was reduced to amine by hydrogenation
with Zn/NH4Cl at room temperature affording 3-chloro-
4-(thiazol-ylmethoxy)aniline (9). The reduction of this
nitro group was confirmed in the 1H NMR spectrum by
the presence of a singlet (δ 5.04) corresponding to the
amine protons, which were exchangeable with D2O. The
amine 9 was subsequently reacted with 4-chloro-6-nitro-
quinazoline to give N-(3-chloro-4-(thiazol-2-ylmethoxy)-
phenyl)-6-nitroquinazolin-4-amine (10). Then the nitro
group at the 6-position of the quinazoline ring in com-
pound 10 was reduced to amine employing the same
strategy affording N4-(3-chloro-4-(thiazol-2-ylmethoxy)-
phenyl)quinazoline-4,6-diamine (11). Analogously, the
General procedure for cell proliferation assay
The anti-proliferative activities against A431 cells of
the six quinazoline derivatives were determined using a
standard (MTT)-based colorimetric assay (Sigma).
Briefly, exponentially growing A431 cells were seeded
into 96-well flat-bottomed plates at a density of 5×103
cells pre well. After 24 h at 37 ℃, exponentially grow-
ing cells were exposed to the indicated compounds at
final concentrations ranging from 0.01 to 100 μg/mL.
The cells were incubated for another 48 h, and cell sur-
vival was determined by the addition of an MTT solu-
tion (20 μL of 5 mg/mL MTT in PBS). After 6 h, 100
μL of 10% SDS in 0.01 equiv. HCl was added, and the
plates were incubated for a further 4 h at 37 ℃. The
optical absorbance was measured at 570 nm using a
spectrophotometric microplate reader. Each concentra-
tion was analyzed in triplicate and the experiment was
repeated three times. The average 50% inhibitory con-
centration (IC50) was determined from the dose-
response curves according to the inhibition ratio for
each concentration using curve-fitting software (Graph-
Pad Prism version 3.00 for Windows, GraphPad Soft-
ware, San Diego, CA).
General procedure of EGFR inhibitory assay
1
The EGFR kinase assay was set up to assess the
level of autophosphorylation based on DELFIA/Time-
Resolved Fluorometry. Compounds 10－12 were dis-
solved in DMSO and diluted to the appropriate concen-
trations with 25 mmol/L HEPES at pH 7.4. In each well,
10 μL compound was incubated with 10 μL (5 ng for
EGFR) recombinant enzyme (1∶85 dilution in 100
mmol/L HEPES) for 10 min at room temperature. Then,
10 μL of 5×buffer (containing 20 mmol/L HEPES, 2
mmol/L MnCl2, 100 μmol/L Na3VO4, and 1 mmol/L
DTT) and 20 μL of 0.1 mmol/L ATP-50 mmol/L MgCl2
were added for 1 h. Positive and negative controls were
included in each plate by incubation of enzyme with or
without ATP-MgCl2. At the end of incubation, liquid
was aspirated, and plates were washed three times with
wash buffer. A 75 μL (400 ng) sample of europiumla-
beled anti-phosphotyrosine antibody was added to each
well for another 1 h of incubation. After washing, en-
hancement solution was added and the signal was de-
tected by Victor (Wallac Inc.) with excitation at 340 nm
and emission at 615 nm. The percentage of auto-phos-
phorylation inhibition by the compounds was calculated
reduction of this nitro group was confirmed in the H
NMR spectrum by the presence of a singlet (δ 5.57)
corresponding to the amine protons, which were ex-
changeable with D2O.
Referring to the preparation methodology of quina-
zoline analogs 12 (a－d), two strategies were employed
outlined in Scheme 2. One method was step by step re-
action starting from amino alcohol and thioCDI. De-
tailedly, shown as method A, amino alcohols were con-
densed with thioCDI in a suitable organic solvent such
as THF or DCM to afford oxazolidine-2-thione (13).
Then the oxazolidine-2-thione was transformed to
bromo-oxazolidine derivative (14) via bromination re-
action at room temperature or at slightly elevated tem-
peratures (30－80 ℃). Then the compound 14 was cou-
pled with N4-(3-chloro-4-(thiazol-2-ylmethoxy)phenyl)quina-
zoline-4,6-diamine (11) to afford the objective 12.
Alternatively, compound 12 can also be prepared
using a facile ‘one pot’ reaction, that is, method B, de-
scribed in Scheme 2. Specifically, the quinazoline 12
was prepared by adding thioCDI to a stirred solution of
N4-(3-chloro-4-(thiazol-2-ylmethoxy)phenyl) quinazo-
Chin. J. Chem. 2014, XX, 1—7
© 2014 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.cjc.wiley-vch.de
5

Hou et al.
FULL PAPER
Scheme 2 Strategies for the preparetion of quinazoline analogs 12 (a－d)
S
NH2
N
+
N
OH
R
N
N
H
N
S
Method A
O
R
Step by step reaction
13
S
S
PBr3
N
N
N
O
Cl
O
Br
O
HN
R
HN
N
Cl
H
14
H2N
N
O
N
N
R
N
N
11
12
Method B
one pot reaction
Condensation agent
S
S
N
N
S
N
N
N
N
O
OH
O
H2N
R
HN
Cl
H
H
N
HN
Cl
N
N
H
N
N
N
S
N
R
OH
N
S
N
16
15
line-4,6-diamine (11) in THF. After several hours,
amino alcohol (1-amino-3-phenoxypropan-2-ol or
2-aminopropan-1-ol) was added and the reaction mix-
ture was stirred for another certain time at room tem-
perature. Some solvent was removed and the residue
was treated with a variety of carbodiinidates (Table 1)
such as in EDCI, DCC, DCI, DIC, DMTT, PFPOH, or
TsCl/NaOH in suitable organic solvents such as THF,
DMF, DCM, DCE at room temperature or slightly ele-
vated temperature. After appropriate time, the reaction
mixture was diluted with EtOAc and water. The aque-
ous phase was extracted and organic phase was dried
followed by purifying to provide the desired product
12a－12d. This synthetic process was more suitable for
industrialized production due to the simplicity and good
yield and purity. The structures of the synthesized
quinazoline analogs and all intermediates were com-
Table 1 Studies on the effect of condensation agent on the
yields of compound 12
Yield/%
Entry
Condensation agent/Solvent
12a 12b 12c 12d
44 34 39 59
48 46 50 42
32 39 35 32
54 56 51 57
46 41 41 59
55 58 45 56
56 59 48 43
61 65 52 60
1
2
3
4
5
6
7
8
DCC/THF
DCI/DCM
DIC/DCM
EDCI/DMF
EDCI/DCE
DMTT/THF
PFPOH/THF
TsCl, NaOH/THF,H2O
cause 50% growth inhibition or EGFR kinase inhibition
with respect to the control culture, were reported. The
results were presented in Table 2. Obviously, com-
pounds 12a and 12b, possessing substituent of oxazole
scaffold at the 7-positions, demonstrated more potent
inhibitory activities for EGFR (IC50＝1.21 and 0.95
μmol/L) than those of compounds 10 and 11 only own-
ing nitro or amino group at the 7-positions. Their activi-
ties positively correlated with antiproliferative activities
and had the same trends. Although the results were less
comparable to the positive control Erlotinib (IC50＝0.03
μmol/L for EGFR), it is possible and necessary to
1
firmed by H and 13C NMR, 2D NMR HSQC and
HMBC spectra, IR spectra and HRMS.
Bioactivities of synthetic compounds
The in vitro anticancer activity of new quinazoline
analogs of A431 cells and EGFR inhibitory potency
were preliminary evaluated by applying the MTT
colorimetric assay and the EGFR kinase assay. Com-
pounds were tested and the calculated IC50 values, that
is, the concentration of a compound that was able to
6
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© 2014 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Chin. J. Chem. 2014, XX, 1—7

Design Synthesis and Bioactivities Evaluation of Novel Quinazoline Analogs
Abdel-Aziz, A. A. M.; Abdel-Hamide, S. G.; Youssef, K. M.;
Al-Obaid, A. M.; El-Subbagh, H. I. Bioorg. Med. Chem. 2006, 14,
8608.
proceed with the investigation of modification and bio-
activity of quinazoline analogs substitued by oxazole
scaffold for discovery of new EGFR inhibitors. Further
research work of them about bioactivity and structure-
function relationship is in progress.
[8] Al-Obaid, A. M.; Abdel-Hamide, S. G.; El-Kashef, H. A.;
Abdel-Aziz, A. A. M.; El-Azab, A. S.; Al-Khamees, H. A.;
El-Subbagh, H. I. Eur. J. Med. Chem. 2009, 44, 2379.
[9] Al-Omary, F. A. M.; Abouzeid, L. A.; Nagi, M. N.; Habib, E. E.;
Abdel-Aziz, A. A. M.; El-Azab, A. S.; Abdel-Hamide, S. G.;
Al-Omar, M. A.; Al-Obaid, A. M.; El-Subbagh, H. I. Bioorg. Med.
Chem. 2010, 18, 2849.
[10] (a) Mendelson, J.; Baselga, J. Oncogene 2000, 19, 6550; (b) Liu, J.;
Liu, Y.; Jian, J.; Bao, X. Chin. J. Org. Chem. 2013, 33, 370 (in
Chinese); (c) Zhang, Y.; Wang, P.; Hu, D.; Jin, L.; Yang, S. Chin. J.
Org. Chem. 2012, 32, 444 (in Chinese).
Table 2 EGFR tyrosine kinase (TK) inhibition and antipro-
liferative data for combinatorial molecules containing nitro group
IC50/(μmol•L−1)
Compound
EGFR
Inhibition of A431 basal growth
inhibition
12a
12b
2.61
2.53
5.31
4.04
7.54
3.32
0.05
1.21
0.95
2.37
2.36
25
[11] (a) Sirisoma, N.; Kasibhatla, S.; Pervin, A.; Zhang, H.; Jiang, S.;
Willardsen, J. A.; Anderson, M. B.; Baichwal, V.; Mather, G. G.;
Jessing, K.; Hussain, R.; Hoang, K.; Pleiman, C. M.; Tseng, B.;
Drewe, J.; Cai, S. X. J. Med. Chem. 2008, 51, 4771; (b) Foote, K. M.;
Mortlock, A. A.; Heron, N. M.; Jung, F. H.; Hill, G. B.; Pasquet, G.;
Brady, M. C.; Green, S.; Heaton, S. P.; Kearney, S.; Keen, N. J.;
Odedra, R.; Wedgea, S. R.; Wilkinsona, R. W. Bioorg. Med. Chem.
Lett. 2008, 18, 1904; (c) Wissner, A.; Fraser, H. L.; Ingalls, C. L.;
Dushin, R. G.; Floyd, M. B.; Cheung, K.; Nittoli, T.; Ravi, M. R.;
Tan, X.; Loganzo, F. Bioorg. Med. Chem. 2007, 15, 3635; (d) Liu,
G.; Hu, D.-Y.; Jin, L.-H.; Song, B.-A.; Yang, S.; Liu, P.-S.; Bhadury,
P. S.; Ma, Y.; Luo, H.; Zhou, X. Bioorg. Med. Chem. 2007, 15, 6608;
(e) Barlaam, B.; Acton, D. G.; Ballard, P.; Bradbury, R. H.; Cross,
D.; Ducray, R.; Germain, H.; Hudson, K.; Klinowska, T.; Magnien,
F.; Ogilvie, D. J.; Olivier, A.; Ross, H. S.; Smith, R.; Trigwell, C. B.;
Vautier, M.; Wright, L. Bioorg. Med. Chem. Lett. 2008, 18, 1799; (f)
Chandrika, P. M.; Yakaiah, T.; Ram Rao, A. R.; Narsaiah, B.; Reddy,
N. C.; Sridhar, V.; Rao, J. V. Eur. J. Med. Chem. 2008, 43, 846.
[12] Nesterova, I. N.; Radkevich, T. P.; Granik, V. G. J. Pharm. Chem.
1991, 25, 786.
12c
12d
11
10
8.32
0.03
Erlotinib
Conclusions
In summary, a series of quinazoline derivatives
possessing oxazole scaffold have been designed and
synthesized, and their biological activities were also
evaluated as potent EGFR inhibitory firstly and then
anticancer activity. Compound 12b demonstrated the
most potent inhibitory activity (IC50＝0.95 μmol/L for
EGFR and 2.53 μmol/L for A431 cells proliferation),
which could be optimized as a potential EGFR inhibitor
and anticancer drug in the further study. More
quinazoline analogs are being prepared according to this
concise strategy and further research work about bioac-
tivity and structure-function relationship is in progress.
[13] Chao, Q.; Deng, L.; Shih, H.; Leoni, L. M.; Genini, D.; Carson, D.
A.; Cottam, H. B. J. Med. Chem. 1999, 42, 3860.
[14] (a) Khalil, M. Y.; Grandis, J. R.; Shin, D. M. Exp. Rev. Anticancer
Ther. 2003, 3, 367; (b) Baker, A. J.; Gibson, K. H.; Grudy, W.; God-
frey, A. A.; Barlow, J. J.; Healy, M. P.; Woodburn, J. R.; Ashton, S.
E.; Curry, B. J.; Scarlett, L.; Henthorn, L.; Richards, L. Bioorg. Med.
Chem. Lett. 2001, 11, 1191; (c) Moyer, J. D.; Barbacci, E. G.; Iwata,
K. K.; Arnold, L.; Boman, B.; Cunningham, A.; DiOrio, C.; Doty, J.;
Morin, M. J.; Moyer, M. P.; Neveu, M.; Pollack, V. A.; Pustilnik, L.
R.; Reynolds, M. M.; Sloan, D.; Theleman, A.; Miller, P. Cancer Res.
1997, 57, 4838.
[15] Rusnak, D. W.; Lackey, K. E.; Wood, E. R.; Alligood, K. J.; Rhodes,
N.; Knight, W. B.; Gilmer, T. M. Mol. Cancer Ther. 2001, 1, 85.
[16] (a) Wissner, A.; Overbeek, E.; Reich, M. F.; Floyd, M. B.; Johnson,
B. D.; Mamuya, N.; Rosfjord, E. C.; Discafani, C.; Davis, R.; Shi, X.
Q.; Rabindran, S. K.; Gruber, B. C.; Ye, F.; Hallett, W. A.; Nilakan-
tan, R.; Shen, R.; Wang, Y. F.; Greenberger, L. M.; Tsou, H. R. J.
Med. Chem. 2003, 46, 49; (b) Tsou, H. R.; Overbeek-Klumpers, E.
G.; Hallett, W. A.; Reich, M. F.; Floyd, M. B.; Johnson, B. D.;
Michalak, R. S.; Shen, R.; Shi, X.; Wang, Y. F.; Upeslacis, J.; Wiss-
ner, A. J. Med. Chem. 2005, 48, 1107.
[17] Traxeler, P.; Bold, G.; Frei, J.; Lang, M.; Lydon, N.; Mett, H.;
Buchdunger, E.; Meyer, T.; Mueller, M.; Furet, P. J. J. Med. Chem.
1997, 40, 3601.
[18] Palmer, B. D.; Trumpp-Kallmeyer, S.; Fry, D. W.; Nelson, J. M.;
Showalter, H. D. H.; Denny, W. A. J. Med. Chem. 1997, 40, 1519.
[19] Alessandro, D.; Giancarlo, F.; Marco, F.; Alessandro, M.; Paola, P.
Tetrahedron 1988, 44, 2021.
[20] Zhang, J. Y.; Liu, H. M.; Xu, H. W.; Shan, L. H. Tetrahedron:
Asymmetry 2008, 19, 512.
[21] Celia, F.; Cristina, O.; Lurdes, G.; Athanasia, B.; Ioannis, P.; Isabel,
S. Bioorg. Med. Chem. 2007, 15, 3974.
Acknowledgement
We are grateful for the financial support from the
National Natural Science Foundation of China (Project
No. 81172937), as well as surpport from the Education
Department of Henan Province Science and Technology
Research Projects (No. 14A150052) and the Scientific
Research Plan of Zhenzhou (No. 121PPTGG509-2).
References
[1] Eckhardt, S. Curr. Med. Chem. Anti-Cancer Agents 2002, 2, 419.
[2] Medina, J. C.; Shan, B.; Beckmann, H.; Farrell, R. P.; Clark, D. L.;
Learned, R. M.; Roche, D.; Baichwal, A.; Li, V.; Case, C.; Baeuerle,
P. A.; Rosen, T.; Jaen, J. C. Bioorg. Med. Chem. Lett. 1998, 8, 2653.
[3] Abdel-Aziz, A. A. M. Eur. J. Med. Chem. 2007, 42, 614.
[4] Medina, J. C.; Roche, D.; Shan, B.; Learned, R. M.; Frankmoelle, W.
P.; Clark, D. L.; Rosen, T.; Jaen, J. C. Bioorg. Med. Chem. Lett. 1999,
9, 1843.
[5] Hwang, H. S.; Moon, E. Y.; Seong, S. K.; Choi, C. H.; Chung, C. H.;
Jung, S. H.; Yoon, S. J. Anticancer Res. 1999, 19, 5087.
[6] Choo, H. Y. P.; Kim, M.; Lee, S. K.; Kim, S. W.; Chung, S. W.
Bioorg. Med. Chem. 2002, 10, 517.
[7] Al-Rashood, S. T.; Aboldahab, I. A.; Nagi, M. N.; Abouzeid, L. A.;
(Pan, B.; Qin, X.)
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© 2014 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.cjc.wiley-vch.de
7