Synthesis and biological evaluation of a novel series of 6,8-dibromo-4(3H)quinazolinone derivatives as anticancer agents

Article abstract of DOI:10.1002/ardp.201300158

Three novel series of 6,8-dibromo-4(3H)quinazolinone derivatives were synthesized. Some of the novel quinazolinone derivatives were tested for their antitumor activity against the human breast carcinoma cell line MCF-7. Compounds XIIIb, IX, XIVd, XIVb, XI

Full text of DOI:10.1002/ardp.201300158

610
Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
Full Paper
Synthesis and Biological Evaluation of a Novel Series of
6,8-Dibromo-4(3H)quinazolinone Derivatives as Anticancer Agents
Marwa F. Ahmed1 and Mahmoud Youns2
1
Faculty of Pharmacy, Department of Pharmaceutical Chemistry, Helwan University, Cairo, Egypt
Faculty of Pharmacy, Department of Biochemistry and Molecular Biology, Helwan University, Cairo, Egypt
2
Three novel series of 6,8-dibromo-4(3H)quinazolinone derivatives were synthesized. Some of the novel
quinazolinone derivatives were tested for their antitumor activity against the human breast carcinoma
cell line MCF-7. Compounds XIIIb, IX, XIVd, XIVb, XIVe, XIIIa, XIVc, XVc, and XIVa exerted powerful
cytotoxic effects against the MCF7 cells, with very low IC50 values compared to doxorubicin (positive
control). The IC50 values were 1.7, 1.8, 1.83, 5.4, 6.84, 10.8, 13.9, 15.7, and 29.6 mg/mL, respectively.
Keywords: 6,8-Dibromo-4(3H)quinazolinone / Human breast carcinoma / Sulfa drugs
Received: April 29, 2013; Revised: May 24, 2013; Accepted: June 12, 2013
DOI 10.1002/ardp.201300158
Introduction
such a way to accommodate Br, sulfonamides, aminothiazol,
pyrazoline, N-phenylpyrazoline, tetrahydropyrimidinone, or
tetrahydropyrimidine-thione moiety with the aim to evaluate
their antitumor activities.
It is well-known that cancer is continuing to be a major health
problem in developing as well as undeveloped countries [1–6].
Although major advances have been made in the chemother-
apeutic management of some patients, the continued Results and discussion
commitment to the laborious task of discovering new
anticancer agents remains critically important.
Chemistry
In the last few years, the attention was oriented towards the
synthesis and biological evaluation of quinazoline derivatives as
they exhibit a broad spectrum of biological activities. Indeed,
several quinazoline derivatives have been reported to possess
anticonvulsant [7, 8], sedative, antihypertensive [9, 10], vaso-
dilatory [11], anti-inflammatory [12], antibiosis [13], phosphodi-
esterase inhibitory [14], fibrinogen receptor antagonistic [15],
and antitumor activity [16–18]. Anilinoquinazoline containing
compounds were recently approved for the treatment of HER2-
positive metastatic breast cancer [19–27]. On the other hand,
diverse chemotherapeutic agents contain pharmacophores like
Br [28], sulfonamides [29], aminothiazol [30], pyrazoline [31],
N-phenylpyrazoline [32], tetrahydropyrimidinone [33], and
tetrahydropyrimidine-thione [34] functions are known to
contribute to the enhancement of the antitumor activity. In
view of the aforementioned facts, it seemed most interesting to
synthesize a new series of anilinoquinazoline compounds in
The desired 6,8-dibromo-2-phenyl-3-(4-acetylphenyl)-4(3H)qui-
nazolinone (II) was obtained by condensation of 6,8-dibromo-
2-phenyl-4H-3,1-benzoxazin-4-one (I) with p-aminoacetophe-
none to give the corresponding key intermediate (II). Reaction
of (II) with bromine in presence of glacial acetic acid afforded
6,8-dibromo-3-(4-(2-bromoacetyl)phenyl)-2-phenylquinazolin-
4(3H)-one (III). Reaction of (III) with malononitrile in absolute
ethanol gave 2-(4-(6,8-dibromo-4-oxo-2-phenylquinazolin-
3(4H)-yl)benzoyl)malononitrile (IV). Cyclocondensation of
(III) by thiourea in absolute ethanol furnished the target
aminothiazol derivative (V) (Scheme 1).
Synthesis of 4-(6,8-dibromo-4-oxo-2-phenylquinazolin-3(4H)-
yl)-N-(substituted) benzenesulfonamides (VI–XI) was obtained
by aromatic aminolysis [35] of the known benzoxazine
derivative (I), upon fusion with the appropriate sulfa drugs,
namely: sulfamethoxazole, sulfaguanidine, sulfacetamide
sodium, sulfamerazine, sulfapyridine, and sulfanilamide
(Scheme 2).
Correspondence: Dr. Marwa F. Ahmed, Faculty of Pharmacy, Depart-
ment of Pharmaceutical Chemistry, Helwan University, Nasr City, Cairo
2122, Egypt.
E-mail: marwafarag80@yahoo.com
Fax: þ20 2 25541601
Additional correspondence: Mahmoud Youns,
E-mail: mahmoud_younis@pharm.helwan.edu.eg
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Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
Quinazoline Anticancer Agents
611
O
N
O
COCH3
H2N
COCH3
Br
Br
N
O
fusion
N
Br
Br
II
I
O
N
O
N
COCH
COCH2Br
3Br2
Br
Br
N
N
Br
Br
III
II
thiourea
NH2
malononitrile
CN
COCH
CN
O
N
N
O
Br
N
Br
N
S
N
Br
Br
IV
V
Scheme 1. Synthesis of 2-(4-(6,8-dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)benzoyl)malononitrile (IV) and aminothiazol derivative (V).
Claisen–Schmidt condensation of the acetyl derivative (II)
with different aldehydes namely, benzaldehyde, p-anisalde-
hyde, p-chlorobenzaldehyde, p-hydroxy benzaldehyde, and/or
furan-2-carboxaldehyde in ethanolic sodium hydroxide solu-
tion afforded the corresponding a,b-unsaturated ketones
(chalcones) (XIIa–e) [36], respectively. These chalcones are
considered to be useful intermediates in several cyclization
reactions to produce different types of heterocyclic com-
pounds of diverse biological importance, according to the
reactants used and the reactions conditions [37].
thiourea in presence of NaOH according to a reported method
[27] afforded the corresponding tetrahydropyrimidinone or
tetrahydropyrimidine-thione derivatives (Scheme 3).
Conclusion
We have synthesized a novel series of quinazolin-4(3H)-ones
compounds. Some of the synthesized compounds were tested
in order to determine the possible anticancer activity. Most
tested compounds have shown promising anticancer activi-
ties against the human breast cancer cell line MCF-7 at very
low concentrations.
Cyclocondensation of the unsaturated ketone (XIId) by
hydrazine hydrate in absolute ethanol afforded the corre-
sponding pyrazoline derivatives (XIIIa).
Experimental
Also, cyclocondensation of the key ketones (XIIa,b) by
phenylhydrazine in absolute ethanol furnished the target
N-phenylpyrazoline derivatives (XIIIb,c).
Chemistry
All melting points are uncorrected, elemental analyses were
carried out in the microanalytical unit of the National Research
Centre and Cairo University, Egypt. IR spectra were recorded on
FT-IR spectrophotometer-Nexus 670-Nicolet (USA) and Perkin
Elmer-9712 spectrophotometer. 1H NMR spectra were determined
Further, the a,b-unsaturated ketones (XIIa–e) were allowed
to react with hydroxylamine hydrochloride in ethanolic
sodium hydroxide solution, affording the corresponding
isoxazolines (XIVa–e), respectively. Cyclocondensation of the
chalcones (XIId,e) with urea in presence of HCl or with
on
a
Varian-Gemmini-300 MHz and Joel-Ex 270 MHz NMR
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M. F. Ahmed and M. Youns
Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
O
O
from ethanol to give IV. m.p. 220°C, yield 70%. Analysis for
N
O
N
O
S
C
24H12Br2N4O2, M.wt. (548.19) calcd.: % C, 52.58; H, 2.21; N, 10.22;
Br
N
N
Found: % C, 52.50; H, 2.18; N, 10.10. IR (KBr, cmꢀ1): 3032 (CH,
H
sulphamethoxazole
–
–
–
–
aromatic), 2200 (CN), 2220 (CN), 1700 (C O), 1672 (C O), and 1635
Br
Br
VI
1
–
–
(C N). H NMR (DMSO-d , d ppm): 5.1 (s, 1H, CH) and 7.30–8.4 (m,
6
NH2
O
O
11H, aromatic-H). MS (m/z, R.I.): Mþ. 546.02, 548.00, 549.00 (51%,
O
N
S
Br
N
NH2
N
100%, 54%).
sulphaguanidine
VII
O
6,8-Dibromo-3-(4-(2-aminothiazol-4-yl)phenyl)-2-
phenylquinazolin-4(3H)-one (V)
O
O
O
N
O
Br
S
O
Br
N
N
Na
N
sulphacetamide Na
A mixture of III (5.6 g; 10 mmol) and thiourea (0.76 g, 10 mmol)
in absolute ethanol (20 mL) was refluxed for 7 h. The precipitated
solid formed upon cooling was filtered and crystallized from
ethanol to give V. m.p. 190°C, yield 70%. Analysis for
C23H14Br2N4OS, M.wt. (554.26) calcd.: % C, 49.84; H, 2.55; N,
N
Br
Br
Br
VIII
O
N
O
O
I
S
S
Br
N
N
N
H
sulphamerazine
N
10.11; Found: % C, 49.74; H, 2.41; N, 10.00. IR (KBr, cmꢀ1): 3380
1
IX
–
–
(NH ), 3028 (CH, aromatic), 1672 (C O) and 1635 (C N). H NMR
–
–
2
O
O
(DMSO-d6, d ppm): 7.1 (s, NH2, exchangeable with D2O) and
7.50–8.28 (m, 11H, aromatic-H). MS (m/z, R.I.): Mþ. 551.92, 553.92,
555.90 (50%, 100%, 52%).
O
Br
N
H
N
N
sulfapyridine
Br
X
General method for the preparation of 4-(6,8-dibromo-4-
oxo-2-phenylquinazolin-3(4H)-yl)-N-(substituted)-
benzenesulfonamides (VI–XI)
A mixture of the benzoxazine I (3.8 g, 0.01 mol) and appropriate
sulfa drug, namely, sulfamethoxazole, sulfaguanidine, sulface-
tamide sodium, sulfamerazine, sulfapyridine, and sulfanilamide
(0.01 mol), was heated together upon fusion at 150°C on a sand
bath for 2 h. After cooling, the crude mass was crystallized from
the proper solvent.
O
O
O
N
S
Br
NH2
N
sulphanilamide
Br
XI
Scheme 2. Synthesis of 4-(6,8-dibromo-4-oxo-2-phenylquinazolin-
3(4H)-yl)-N-(substituted) benzenesulfonamides (VI–XI).
spectrometer using TMS as an internal standard. Mass spectra
were determined on Finnigan Mat SSQ 7000, mode EI 70 eV
(Thermo Inst. Sys., Inc., USA). Thin layer chromatography was
carried out on silica gel 60 F254 (Merck) plates.
4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-N-
(5-methylisoxazol-3-yl)benzenesulfonamide (VI)
Crystallized from ethanol to give VI. m.p. 210°C, yield 60%.
Analysis for C24H16Br2N4O4S, M.wt. (613.93) calcd.: % C, 46.77; H,
2.62; N, 9.09; Found: % C, 46.72; H, 2.58; N, 9.00. IR (KBr, cmꢀ1):
1
6,8-Dibromo-2-phenyl-3-(4-acetylphenyl)-4(3H)-
–
–
3380 (NH), 3040 (CH, aromatic), 1678 (C O), and 1635 (C N). H
–
–
quinazolinone (II)
It was prepared according to reported literatures [36].
NMR (DMSO-d6, d ppm): 2.3 (s, 1H, CH3), 6.50–8.5 (m, 12H,
aromatic-H) and 11.01 (s, NH, exchangeable with D2O). MS (m/z,
R.I.): Mþ. 613.93, 615.92, 617.93 (51%, 100%, 49%).
6,8-Dibromo-3-(4-(2-bromoacetyl)phenyl)-2-
phenylquinazolin-4(3H)-one (III)
4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-N-
(diaminomethylene)benzenesulfonamide (VII)
To a solution of the ketone II (0.99 g, 0.002 mol) in glacial acetic
acid was added 4 mL bromine in 5 mL glacial acetic acid dropwise
and left over night, then poured into crushed ice. The separated
material was filtered, air dried, and crystallized from ethanol to
give III. m.p. 160°C, yield 65%. Analysis for C22H13Br3N2O2, M.wt.
(577.06) calcd.: % C, 45.79; H, 2.27; N, 4.85; Found: % C, 45.72; H,
Crystallized from ethanol to give VII. m.p. 250°C, yield 65%.
Analysis for C21H15Br2N5O3S, M.wt. (577.25) calcd.: % C, 43.69; H,
2.62; N, 12.13; Found: % C, 43.61; H, 2.58; N, 12.01. IR (KBr, cmꢀ1):
–
3380 (NH ), 3370 (NH ), 3040 (CH, aromatic), 1678 (C O), and
–
2
2
1635 (C N). 1H NMR (DMSO-d6, d ppm): 6.61 (s, 4H, NH2,
–
–
2.21; N, 4.79. IR (KBr, cmꢀ1): 3028 (CH, aromatic), 1700 (C O) 1672
exchangeable with D2O) and 6.50–8.5 (m, 11H, aromatic-H). MS
–
–
1
(C O), and 1635 (C N). H NMR (DMSO-d , d ppm): 4.5 (s, 1H, CH )
(m/z, R.I.): Mþ. 574.91, 576.92, 578.93 (53%, 100%, 50%).
–
–
–
–
6
2
and 7.60–8.1 (m, 12H, aromatic-H). MS (m/z, R.I.): Mþ. 573.80,
575.82, 577.80 (34%, 100%, 97%).
4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-
sodiumacetyl(phenylsulfonyl)amide (VIII)
2-(4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-
benzoyl)malononitrile (IV)
A mixture of III (5.6 g; 10 mmol) and malononitrile (0.66 g,
10 mmol) in absolute ethanol (15 mL) was treated with NaOH (4 g
in 10 mL H2O) dropwise with stirring, then diluted with 30 mL
H2O; the precipitated solid formed was filtered and crystallized
Crystallized from ethanol to give VIII. m.p. 290°C, yield 75%.
Analysis for C22H14Br2N3NaO4S, M.wt. (599.23) calcd.: % C, 44.10; H,
2.35; N, 7.01; Found: % C, 43.99; H, 2.31; N, 6.98. IR (KBr, cmꢀ1): 3040
1
–
–
–
–
(CH, aromatic), 1700 (C O), 1678 (C O), and 1635 (C N). H NMR
–
–
(DMSO-d6, d ppm): 2.1 (s, 1H, CH3) and 6.50–8.5 (m, 11H, aromatic-H).
MS (m/z, R.I.): Mþ. 596.91, 598.92, 600.93 (50%, 100%, 49%).
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Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
Quinazoline Anticancer Agents
613
O
O
Ar
O
N
COCH3
ArCHO
Br
Br
N
N
N
Br
Br
NaOH
XII
II
HO
a, Ar =
b, Ar =
c, Ar =
d, Ar =
e, Ar =
OMe
O
Cl
thiourea
NaOH/EtOH
urea
HCl
NH2NH2
98%
NH2OH
EtOH
or
N
O
phNHNH2
O
R
X
N
N
Br
O
N
H
N
N
Br
O
N
Ar
Ar
Br
N
Ar
N
N
N
Br
_
XIVa e
Br
Br
_
XIIIa c
a, Ar =
_
XVa c
OCH3
b, Ar =
HO
R= H
a, Ar = sal
a, X=O , Ar =
Cl
c, Ar =
d, Ar =
e, Ar =
HO
b, Ar = benz
c, Ar = anis
R= ph
R= ph
b, X=O , Ar =
c, X=S , Ar =
O
HO
O
Scheme 3. Synthesis of pyrazoline derivative (XIIIa), N-phenylpyrazolines (XIIIb,c), isoxazolines (XIVa–e), tetrahydropyrimidinones
(XVa,b), and tetrahydropyrimidine-thione derivative (XVc).
4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-N-
4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-N-
(4-methylpyrimidin-2-yl)benzenesulfonamide (IX)
(pyridin-2-yl)benzenesulfonamide (X)
Crystallized from ethanol to give IX. m.p. 280°C, yield 60%.
Analysis for C25H17Br2N5O3S, M.wt. (627.31) calcd.: % C, 47.87; H,
Crystallized from ethanol to give X. m.p. 232°C, yield 65%.
Analysis for C25H16Br2N4O3S, M.wt. (612.29) calcd.: % C, 49.04; H,
2.73; N, 11.16; Found: % C, 47.85; H, 2.70; N, 11.10. IR (KBr, cmꢀ1):
2.63; N, 9.15; Found: % C, 49.00; H, 2.59; N, 9.05. IR (KBr, cmꢀ1):
1
1
–
–
–
–
–
–
3380 (NH), 3040 (CH, aromatic), 1678 (C O), and 1630 (C N). H
3385 (NH), 3045 (CH, aromatic), 1670 (C O), and 1630 (C N). H
–
–
NMR (DMSO-d6, d ppm): 2.3 (s, 1H, CH3), 7.50–8.5 (m, 13H,
aromatic-H) and 11.01 (s, NH, exchangeable with D2O). MS (m/z,
R.I.): Mþ. 625.01, 627.02, 629.03 (49%, 100%, 51%).
NMR (DMSO-d6, d ppm): 7.50–8.5 (m, 15H, aromatic-H) and 11.2 (s,
NH, exchangeable with D2O). MS (m/z, R.I.): Mþ. 609.93, 611.92,
613.93 (50%, 100%, 53%).
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M. F. Ahmed and M. Youns
Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
4-(6,8-Dibromo-4-oxo-2-phenylquinazolin-3(4H)-yl)-
benzenesulfonamide (XI)
General method for the synthesis of 6,8-dibromo-2-phenyl-
3-(4-(5-(substituted)-4,5-dihydroisoxazol-3-yl)phenyl)-
quinazolin-4(3H)-ones (XIVa–e)
A mixture of chalcones XIIa–e (3 mmol) and hydroxylamine
hydrochloride (5 mmol) in sodium hydroxide solution (0.5 g
NaOH in 0.5 mL water) in ethanol (60 mL) was refluxed for 3 h.
The product obtained upon cooling was filtered off, washed with
water, and recrystalized from the proper solvents to obtain the
desired compounds XIVa–e, respectively.
Crystallized from ethanol to give XI. m.p. 254°C, yield 70%.
Analysis for C20H13Br2N3O3S, M.wt. (535.21) calcd.: % C, 44.88; H,
2.45; N, 7.85; Found: % C, 44.81; H, 2.40; N, 7.80. IR (KBr, cmꢀ1):
1
–
–
3380 (NH ), 3045 (CH, aromatic), 1675 (C O) and 1632 (C N). H
–
–
2
NMR (DMSO-d6, d ppm): 7.2 (s, NH2, exchangeable with D2O) and
7.60–8.3 (m, 11H, aromatic-H). MS (m/z, R.I.): Mþ. 532.93, 534.90,
536.93 (51%, 100%, 54%).
6,8-Dibromo-2-phenyl-3-(4-(5-phenyl-4,5-dihydroisoxazol-
General method for the synthesis of XIIa–e
6,8-Dibromo-2-phenyl-3-{4-[(E)-3-substituted arylacryloyl]-phenyl}-
3H-quinazolin-4-ones (XIIa–e) (chalcones) were prepared accord-
ing to reported literatures [36].
3-yl)phenyl)quinazolin-4(3H)-one (XIVa)
Crystallized from ethanol to give XIVa. m.p. 160°C, yield 70%.
Analysis for C29H19Br2N3O2, M.wt. (601.29) calcd.: % C, 57.90; H,
3.18; N, 6.99; Found: % C, 57.83; H, 3.12; N, 6.88. IR (KBr, cmꢀ1):
1
–
–
3036 (CH, aromatic), 1673 (C O), and 1630 (C N). H NMR (DMSO-
–
–
6,8-Dibromo-2-phenyl-3-{4-[5-(4-hydroxy phenyl)-4,5-
dihydro-1H-pyrazol-3-yl]-phenyl}-3H-quinazolin-4-one
(XIIIa)
d6, d ppm): 3.7–3.82 (d, d, 2H, CH2, isoxazoline ring, J ¼ 5.4 Hz),
6.34 (t, 1H, CH, isoxazoline ring), and 7.80–8.54 (m, 16H, aromatic-
H). MS (m/z, R.I.): Mþ. 598.98, 600.98, 602.98 (50%, 100%, 48.7%).
A mixture of the chalcone XIId (0.005 mol) and hydrazine
hydrate (2.5 mL, 0.005 mol, 98%) in absolute ethanol (25 mL) was
heated under reflux for 10 h. After cooling, the separated
material was filtered, air dried, and crystallized from ethanol to
give XIIIa. m.p. 120°C, yield 60%. Analysis for C29H22Br2N4O2, M.
wt. (616.3) calcd.: % C, 56.52; H, 3.27; N, 9.09; Found: % C, 56.42; H,
3.10; N, 9.00. IR (KBr, cmꢀ1): 3380 (OH), 3385 (NH), 3030 (CH,
6,8-Dibromo-3-(4-(5-(4-methoxyphenyl)-4,5-
dihydroisoxazol-3-yl)phenyl)-2-phenylquinazolin-4(3H)-
one (XIVb)
Crystallized from ethanol to give XIVb. m.p. 200°C, yield 65%.
Analysis for C30H21Br2N3O3, M.wt. (631.31) calcd.: % C, 57.07; H,
3.35; N, 6.66; Found: % C, 57.00; H, 3.29; N, 6.61. IR (KBr, cmꢀ1):
aromatic), 1670 (C O), and 1635 (C N). 1H NMR (DMSO-d6, d
1
–
–
–
–
–
–
3040 (CH, aromatic), 1670 (C O), and 1637 (C N). H NMR (DMSO-
–
–
ppm):, 3.1 (d, d, 2H, CH2, pyrazoline ring, J ¼ 5.6 Hz), 3.90 (t, 1H,
CH of pyrazoline), 7.21 (s, NH, exchangeable with D2O), 7.50–8.28
(m, 16H, aromatic-H) and 9.80 (s, 1H, OH). MS (m/z, R.I.): Mþ.
614.00, 616.08, 618.00 (49%, 100%, 50%).
d6, d ppm): 3.48–3.52 (d, d, 2H, CH2, isoxazoline ring), 3.80 (s, 1H,
OCH3), 6.34 (t, 1H, CH, isoxazoline ring), and 7.50–8.34 (m, 15H,
aromatic-H). MS (m/z, R.I.): Mþ. 629.00, 631.01, 633.00 (51%, 100%,
49.0%).
6,8-Dibromo-3-(4-(5-(2-chlorophenyl)-4,5-
6,8-Dibromo-2-phenyl-3-{4-[1,5-diphenyl-4,5-dihydro-1H-
dihydroisoxazol-3-yl)phenyl)-2-phenylquinazolin-4(3H)-
pyrazol-3-yl]-phenyl}-3H-quinazolin-4-one (XIIIb)
A mixture of the chalcone XIIa (0.003 mol) and phenylhydrazine
(0.32 g, 0.003 mol, 98%) in absolute ethanol (25 mL) was heated
under reflux for 10 h. After cooling, the separated material was
filtered, air dried, and crystallized from ethanol to give XIIIb. m.p.
180°C, yield 75%. Analysis for C35H24Br2N4O, M.wt. (674.03) calcd.:
% C, 62.15; H, 3.58; N, 8.28; Found: % C, 62.00; H, 3.52; N, 8.19. IR
one (XIVc)
Crystallized from ethanol to give XIVc. m.p. 170°C, yield 75%.
Analysis for C29H18Br2ClN3O3, M.wt. (635.73) calcd.: % C, 54.79; H,
2.85; N, 6.61; Found: % C, 54.72; H, 2.79; N, 6.58. IR (KBr, cmꢀ1):
3040 (CH, aromatic), 1675 (C O), and 1635 (C N). H NMR (DMSO-
d6, d ppm): 3.58–3.62 (d, d, 2H, CH2, isoxazoline ring), 6.24 (t, 1H,
CH, isoxazoline ring), and 7.40–8.34 (m, 15H, aromatic-H). MS
(m/z, R.I.): Mþ. 633.00, 636.03, 638.00 (44%, 100%, 70.2%).
1
–
–
–
–
(KBr, cmꢀ1): 3388 (NH), 3038 (CH, aromatic), 1675 (C O), and 1635
–
–
1
–
–
(C N). H NMR (DMSO-d , d ppm): 3.6 (d, d, 2H, CH , pyrazoline
6
2
ring), 5.1 (t, 1H, CH of pyrazoline), and 7.50–8.5 (m, 21H, aromatic-
H). MS (m/z, R.I.): Mþ. 674.03, 676.00, 678.03 (51%, 100%, 49%).
6,8-Dibromo-3-(4-(5-(2-hydroxyphenyl)-4,5-
dihydroisoxazol-3-yl)phenyl)-2-phenylquinazolin-4(3H)-
one (XIVd)
6,8-Dibromo-2-phenyl-3-{4-[5-(4-methoxyphenyl)-1-
phenyl-4,5-dihydro-1H-pyrazol-3-yl]-phenyl}-3H-
quinazolin-4-one (XIIIc)
Crystallized from ethanol to give XIVd. m.p. 270°C, yield 60%.
Analysis for C29H19Br2N3O3, M.wt. (617.29) calcd.: % C, 56.43; H,
3.10; N, 6.81; Found: % C, 56.40; H, 3.02; N, 6.76. IR (KBr, cmꢀ1):
3334 (OH), 3045 (CH, aromatic), 1675 (C O), and 1630 (C N). H
NMR (DMSO-d6, d ppm): 3.58–3.62 (d, d, 2H, CH2, isoxazoline ring),
6.24 (t, 1H, CH, isoxazoline ring), 7.40–8.34 (m, 15H, aromatic-H),
and 9.60 (s, 1H, OH). MS (m/z, R.I.): Mþ. 614.98, 617.00, 618.98 (50%,
100%, 52%).
A mixture of the chalcone XIIb (0.003 mol) and phenylhydrazine
(0.32 g, 0.003 mol, 98%) in absolute ethanol (25 mL) was heated
under reflux for 10 h. After cooling, the separated material was
filtered, air dried, and crystallized from ethanol to give XIIIc. m.p.
215°C, yield 65%. Analysis for C36H26Br2N4O2, M.wt. (706.04)
calcd.: % C, 61.21; H, 3.71; N, 7.93; Found: % C, 61.18; H, 3.62; N,
1
–
–
–
–
7.89. IR (KBr, cmꢀ1): 3390 (NH), 3040 (CH, aromatic), 1673 (C O),
–
–
and 1638 (C N). 1H NMR (DMSO-d6, d ppm): 3.8 (d, d, 2H, CH2,
6,8-Dibromo-3-(4-(5-(furan-2-yl)-4,5-dihydroisoxazol-3-yl)-
–
–
pyrazoline ring), 3.80 (s, 1H, OCH3), 5.31 (t, 1H, CH of pyrazoline),
and 7.60–8.54 (m, 20H, aromatic-H). MS (m/z, R.I.): Mþ. 704.03,
706.00, 708.00 (52%, 100%, 49.7%).
phenyl)-2-phenylquinazolin-4(3H)-one (XIVe)
Crystallized from ethanol to give XIVe. m.p. 110°C, yield 65%.
Analysis for C27H17Br2N3O3, M.wt. (591.25) calcd.: % C, 54.85; H,
ß 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com

Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
Quinazoline Anticancer Agents
615
2.90; N, 7.11; Found: % C, 54.80; H, 2.79; N, 7.08. IR (KBr, cmꢀ1): Pharmacology
1
–
–
–
3042 (CH, aromatic), 1677 (C O) and 1637 (C N). H NMR (DMSO-
–
d6, d ppm): 3.60–3.68 (d, d, 2H, CH2, isoxazoline ring), 6.44 (t, 1H,
CH, isoxazoline ring), and 7.60–8.5 (m, 14H, aromatic-H). MS (m/z,
R.I.): Mþ. 589.00, 591.02, 593.03 (48%, 100%, 50%).
Materials and methods
Cell line and treatments
The human breast cancer cell line (MCF-7) was obtained from
the American Type Culture Collection and grown in DMEM
medium with 10% fetal bovine serum, 100 units/mL penicil-
lin, and 100 mg/mL streptomycin (Invitrogen). Cells were
maintained at 37° in a humidified atmosphere of 5% CO2.
Stock solutions of our compounds were prepared in DMSO
and stored at ꢀ20°C. All controls were exposed to DMSO
alone; DMSO concentration was always <0.1%.
General method for the synthesis of 6,8-dibromo-3-
(4-(6-(substituted)-2-oxo-1,2,5,6-tetrahydropyrimidin-4-yl)-
phenyl)-2-phenylquinazolin-4(3H)-one (XVa,b)
A mixture of the chalcones XIId,e (0.005 mol) and urea (0.5 g,
0.005 mol) in ethanol (20 mL), and conc. HCl (5 mL) was refluxed
for 7 h. The reaction mixture was concentrated to 1/2 its volume,
cooled, and neutralized with NH4OH solution. The precipitated
solid was filtered off, washed with water, air dried, and
crystallized from the proper solvent to give compounds XVa,b.
Cell viability assay (sulforhodamine B assay)
To assess cellular proliferation, sulforhodamine B (SRB) assay
was used according to the manufacturer’s instructions.
Briefly, cells were grown in tissue culture flasks, and then
harvested by treating the flasks with 0.025% trypsin and
0.25 mM EDTA for 5 min. Once detached, cells were washed,
counted and an aliquot (5 ꢁ 103 cells) was placed in each well
of a 96-well cell culture plate in a total volume of 100 mL. Cells
were allowed to attach overnight and then treated with or
without increasing concentrations of curcumin. After treat-
ment, cell medium was aspirated, and 100 mm3/well of 10%
trichloroacetic acid was added. After fixation at 4°C and
washing, 50 mm3 of 0.4% w/v sulforhodamine B (SRB; Sigma–
Aldrich) was added. Plates were incubated at room tempera-
ture for 30 min. Unbound SRB was removed with 1% acetic
acid. Bound SRB was solubilized with 100 mm3 of 10 mM Tris-
base solution. Absorbance was measured using a precision
microplate reader (Molecular Devices, Sunnyvale, CA) at 570
and 650 nm (background). Cell viability was expressed as the
percentage of cell survival relative to untreated controls.
6,8-Dibromo-3-(4-(6-(2-hydroxyphenyl)-2-oxo-1,2,5,6-
tetrahydropyrimidin-4-yl)phenyl)-2-phenylquinazolin-
4(3H)-one (XVa)
Crystallized from ethanol to give XVa. m.p. 115°C, yield 70%.
Analysis for C30H20Br2N4O3, M.wt. (644.31) calcd.: % C, 55.92; H,
3.13; N, 8.70; Found: % C, 55.88; H, 3.10; N, 8.65. IR (KBr, cmꢀ1):
3200–3600 (OH enolic of pyrimidine), 3042 (CH, aromatic), 1688
1
–
–
(C O), and 1618 (C N). H NMR (DMSO-d , d ppm): at 3.0 (2H, d,
–
–
6
CH2 of pyrimidinone), 5.1 (1H, t, CH of pyrimidinone), 7.2–8.5
(16H, m, aromatic protons including that of pyrimidone and
quinazolone rings), and 9.8 (1H, S, OH). MS (m/z, R.I.): Mþ. 642.00,
644.02, 646.03 (49%, 100%, 52%).
6,8-Dibromo-3-(4-(6-(furan-2-yl)-2-oxo-1,2,5,6-
tetrahydropyrimidin-4-yl)phenyl)-2-phenylquinazolin-
4(3H)-one (XVb)
Crystallized from ethanol to give XVb. m.p. 180°C, yield 75%.
Analysis for C28H18Br2N4O3, M.wt. (618.28) calcd.: % C, 54.39; H,
2.93; N, 9.06; Found: % C, 54.34; H, 2.89; N, 9.00. IR (KBr, cmꢀ1):
3220–3610 (OH enolic of pyrimidine), 3045 (CH, aromatic), 1690
1
–
–
(C O), and 1620 (C N). H NMR (DMSO-d , d ppm): at 3.1 (2H, d,
–
–
6
CH2 of pyrimidinone), 5.4 (1H, t, CH of pyrimidinone), and 7.5–8.3
(15H, m, aromatic protons including that of pyrimidone and
quinazolone rings). MS (m/z, R.I.): Mþ. 615.97, 617.97, 619.96 (52%,
100%, 54%).
Caspase-Glo 3/7 assay
Caspase-GloTM3/7 Assay (Promega) was used to detect caspase
3/7 activities of MCF-7 cancer cell lines triggered by our
compounds. This test provides a proluminescent caspase 3/7
substrate, which contains the caspase 3 specific tetrapeptide
sequence DEVD in a reagent optimized for cell lysis and
determination of caspases and luciferase activity. MCF-7 cells
cultured in DMEM were seeded in 96-well plates and treated
with our compounds. Six hours after treatment, cellular
caspase 3/7 activity was determined according to the
manufacturer’s protocol. Luminescence was measured using
Mithras LB 940 (Berthold Technologies, BadWildbad,
Germany). Cellular apoptosis was expressed as relative to
the untreated medium control.
6,8-Dibromo-3-(4-(6-(2-hydroxyphenyl)-2-thioxo-1,2,5,6-
tetrahydropyrimidin-4-yl)phenyl)-2-phenylquinazolin-
4(3H)-one (XVc)
A mixture of the chalcone XIId (0.005 mol) and thiourea
(0.005 mol) in presence of 0.5 g of NaOH in 5 mL of water. The
mixture was refluxed in (25 mL) of ethanol for 6 h, then
concentrated under vaccum and neutralized with dilute HCl.
The precipitated material was filtered off, washed with water,
dried, and crystallized from ethanol to give XVc. m.p. 210°C, yield
65%. Analysis for C30H20Br2N4O2S, M.wt. (660.38) calcd.: % C,
54.56; H, 3.05; N, 8.48; Found: % C, 54.50; H, 3.00; N, 8.38. IR (KBr,
cmꢀ1): 3220–3610 (OH enolic of pyrimidine), 3045 (CH, aromatic),
1690 (C O), 1620 (C N) and 1275 (C S). 1H NMR (DMSO-d6, d
–
–
–
–
–
–
In this present work a novel series of quinazolin-4(3H)-ones
compounds was synthesized. Synthetic Schemes 1–3 illustrate
the way used for the synthesis of target compounds. Some of
the synthesized compounds were tested in order to determine
ppm): at 3.3 (2H, d, CH2 0f pyrimidinone), 5.2 (1H, t, CH of
pyrimidinone), 7.7–8.5 (15H, m, aromatic protons including that
of pyrimidone and quinazolone rings), and 9.7 (1H, S, OH). MS
(m/z, R.I.): Mþ. 657.95, 659.94, 661.95 (50%, 100%, 54%).
ß 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com

616
M. F. Ahmed and M. Youns
Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
Table 1. Cytotoxity of tested compounds against the human
breast cancer cell line MCF-7 compared to doxorubicin.
IC50 values (mg/mL) with MCF-7 breast cancer cell line
Compounds
IC50 (mg/mL)
XIVd
XIVc
XIVe
XIIIc
VIII
XIVa
IV
XVb
1.83 ꢂ 0.09
13.9 ꢂ 2.41
6.84 ꢂ 0.92
38.5 ꢂ 1.8
68.12 ꢂ 3.42
21.4 ꢂ 1.71
68.4 ꢂ 3.83
53.1 ꢂ 2.54
15.7 ꢂ 0.92
>100
XVc
XVa
Figure 1. Caspase-Glo 3/7 activity of tested compounds relative to
untreated control. Enzymatic activity of caspase 3 after 6-h
treatment of MCF-7 cells. The activity is expressed as percentage
relative to untreated cells.
XIVb
XIIIb
XIIIa
IX
5.4 ꢂ 1.23
1.7 ꢂ 0.35
10.8 ꢂ 1.98
1.8 ꢂ 0.12
29.6 ꢂ 2.78
Doxorubicin
their IC50 concentrations. Six hours after treatment, the
activity of caspase 3/7 was measured using the Caspase-Glo 3/7
assay. Figure 1 shows that some of the tested compounds
caused significant increase in activation of caspase 3/7 (e.g.,
compounds XIVd, VIII, XIVb).
the possible anticancer activity. MCF7 human breast cancer
cells were cultured in a monolayer and treated with our
compounds for 48 h. The SRB assay was performed as
previously shown [38] to assess the rate of proliferation,
and the resulting growth curves showed that our molecules
exhibited cytotoxicity against the above-mentioned cell line
with very low IC50 values (Table 1). Compounds XIIIb, IX,
XIVd, XIVb, XIVe, XIIIa, XIVc, XVc, and XIVa exerted powerful
cytotoxic effects against MCF7 cells with very low IC50 values
compared to doxorubicin (positive control; IC50 values were
1.7, 1.8, 1.83, 5.4, 6.84, 10.8, 13.9, 15.7, and 29.6 mg/mL,
respectively). Compounds XIIIc, VIII, IV, and XVb exerted
moderate cytotoxic effect against MCF7 cells (IC50 values were
38.51, 68.12, 68.4, and 53.1 mg/mL, respectively). Compound
XVa exerted a weak cytotoxic effect against MCF7 cells (IC50
values >100 mg/mL). The most potent anticancer activity
exhibited by compounds XIIIa,b, IX, XIVa–e, and XVc might
be due to the presence of N-phenylpyrazoline, N-(4-methyl-
pyrimidin-2-yl)benzenesulfonamide, isoxazolines, and tetra-
hydropyrimidine-thione moiety of the 2-phenyl substituted
quinazolin-4(3H)-one, respectively. It is interesting to note that
a minor alteration in the molecular configuration of the
investigated compounds may have a pronounced effect on
anticancer activity, e.g., compound XVc having tetrahydro-
pyrimidine-thione moiety showed more anticancer activity
than XVa with tetrahydropyrimidinone.
The authors have declared no conflict of interest.
References
[1] S. Eckhardt, Curr. Med. Chem. Anticancer Agents 2002, 2, 419–
439.
[2] J. C. Medina, B. Shan, H. Beckmann, R. P. Farrell, D. L. Clark,
R. M. Learned, D. Roche, A. Li, V. Baichwal, C. Case, P. A.
Baeuerle, T. Rosen, J. C. Jaen, Bioorg. Med. Chem. Lett. 1998, 8,
2653–2656.
[3] A. A. M. Abdel-Aziz, Eur. J. Med. Chem. 2007, 42, 614–626.
[4] J. C. Medina, D. Roche, B. Shan, R. M. Learned, W. P.
Frankmoelle, D. L. Clark, T. Rosen, J. C. Jaen, Bioorg. Med.
Chem. Lett. 1999, 9, 1843–1846.
[5] H. S. Hwang, E. Y. Moon, S. K. Seong, C. H. Choi, C. H. Chung,
S. H. Jung, S. J. Yoon, Anticancer Res. 1999, 19, 5087–5093.
[6] H. Y. P. Choo, M. Kim, S. K. Lee, S. W. Kim, S. W. Chung, Bioorg.
Med. Chem. 2002, 10, 517–523.
[7] D. G. Wenzel, J. Am. Pharm. Assoc. 1955, 44, 550–553.
[8] M. Hori, R. Iemura, H. Hara, A. Ozaki, T. Sukamoto, H.
Ohtaka, Chem. Pharm. Bull. 1990, 38, 681–687.
[9] S. Hayao, H. J. Havera, H. E. W. G. Stryeker, T. G. Leipzig, R. A.
Kulp, H. E. Hartzler, J. Med. Chem. 1965, 8, 807–815.
[10] Y. Nishikawa, T. Shindo, K. Ishii, H. Nakamura, T. Kona, H.
Uno, J. Med. Chem. 1989, 32, 583–593.
It is well-established that the induction of the apoptotic
cascade is one of the main mechanisms of chemotherapy-
induced cell death [39], in order to determine whether the
chemosensitizing effect of our compounds demonstrated
above is secondary to its ability to activate the apoptotic
cascade, MCF-7 cells were treated with test compounds at
[11] H. J. Havera, H. J. Vidrio, J. Med. Chem. 1979, 22, 1548–1550.
[12] Q. Chao, L. Deng, H. Shih, L. M. Leoni, D. Genini, D. A. Carson,
H. B. Cottam, J. Med. Chem. 1999, 42, 3860–3873.
[13] P. P. Kung, M. D. Casper, K. L. Cook, L. Wilson-Lingardo, L. M.
Risen, T. A. Vickers, R. Ranken, L. B. Blyn, J. R. Wyatt, P. D.
Cook, D. J. Ecker, J. Med. Chem. 1999, 42, 4705–4713.
ß 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com

Arch. Pharm. Chem. Life Sci. 2013, 346, 610–617
Quinazoline Anticancer Agents
617
[14] T. Glaser, J. Traber, Agents Actions 1984, 15, 314–317.
[27] L. F. Hennequin, E. S. E. Stokes, A. P. Thomas, C. Johnstone,
P. A. Plé, D. J. Ogilvie, M. Dukes, S. R. Wedge, J. Kendrew, J. O.
Curwen, J. Med. Chem. 2002, 45, 1300–1312.
[15] N. J. Liverton, D. J. Armstrong, D. A. Claremon, D. C. Remy,
J. J. Bardwin, R. J. Lynch, G. Zhang, R. J. Gould, Bioorg. Med.
Chem. Lett. 1998, 8, 483–486.
[28] P. Mani Chandrika, T. Yakaiah, A. Raghu Ram Rao, B.
Narsaiah, N. Chakra Reddy, V. Sridhar, J. Venkateshwara
Rao, Eur. J. Med. Chem. 2008, 43, 846–852.
[16] S. T. Al-Rashood, I. A. Aboldahab, M. N. Nagi, L. A. Abou-Zeid,
A. A.-M. Abdel-Aziz, S. G. Abdel-Hamide, K. M. Youssef, A. M.
Al-Obaid, H. I. El-Subbagh, Bioorg. Med. Chem. 2006, 14, 8608–
8621.
[29] M. Abdulrahman Al-Obaid, G. Sami Abdel-Hamide, A. Hassan
El-Kashef, A. M. Alaa Abdel-Aziz, S. Adel El-Azab, A. Hamad
Al-Khamees, I. Hussein El-Subbagh, Eur. J. Med. Chem. 2009, 44,
2379–2391.
[17] A. M. Al-Obaid, S. G. Abdel-Hamide, H. A. El-Kashef, A. A.-M.
Abdel-Aziz, A. S. El-Azab, H. A. Al-Khamees, H. I. El-Subbagh,
Eur. J. Med. Chem. 2009, 44, 2379–2391.
[30] H. A. Abdel-Aziz, A. M. Gamal-Eldeen, N. A. Hamdy, I. M.
Fakhr, Arch. Pharm. 2009, 342, 230–237.
[18] F. A. M. Al-Omary, L. A. Abou-Zeid, M. N. Nagi, E. E. Habib,
A. A.-M. Abdel-Aziz, A. S. El-Azab, S. G. Abdel-Hamide, M. A.
Al-Omar, A. M. Al-Obaid, H. I. El-Subbagh, Bioorg. Med. Chem.
2010, 18, 2849–2863.
[31] T. Taj, R. R. Kamble, T. M. Gireesh, R. K. Hunnur, S. B.
Margankop, Eur. J. Med. Chem. 2011, 46, 4366–4373.
[32] R. Narayan Sharma, K. P. Sharma, S. N. Dikshit, Adv. Appl. Sci.
Res. 2011, 2, 178–192.
[19] F. Barlesi, C. Tchouhadjian, C. Doddoli, P. Villani, L. Greillier,
J.-P. Kleisbauer, P. Thomas, P. Astoul, Fundam. Clin. Pharmacol.
2005, 19, 385–393.
[33] A. Kamal, E. Vijaya Bharathi, J. Surendranadha Reddy, M.
Janaki Ramaiah, D. Dastagiria, M. Kashi Reddy, A. Viswa-
nath, T. Lakshminarayan Reddy, T. Basha Shaik, S. N. C. V. L.
Pushpavallib, M. Pal Bhadrab, Eur. J. Med. Chem. 2011, 46, 691–
703.
[20] C. L. Arteaga, D. H. Johnson, Curr. Opin. Oncol. 2001, 13, 491–498.
[21] A. J. Baker, K. H. Gibson, W. Grundy, A. A. Godfrey, J. J.
Barlow, M. P. Healy, J. R. Woodburn, S. E. Ashton, B. J. Curry,
L. Scarlett, L. Henthorn, L. Richards, Bioorg. Med. Chem. Lett.
2001, 11, 1911–1914.
[34] S. Cesarini, A. Spallarossa, A. Ranise, S. Schenone, C. Rosano,
P. La Colla, G. Sanna, B. Busonera, R. Loddo, Eur. J. Med. Chem.
2009, 46, 1106–1118.
[22] K. N. Ganjoo, H. Wakelee, Biologics: Targets Ther. 2007, 1, 335–
346.
[35] S. I. El-Nagady, M. A. El-Hashash, A. S. El-Badaway, Egypt. J.
Chem. 2006, 49, 721–728.
[23] L. Kopper, Pathol. Oncol. Res. 2008, 14, 1–8.
[36] M. S. Mohamed, M. M. Kamel, E. M. M. Kassem, N. Abotaleb, S.
M. Nofal, M. F. Ahmed, Acta Polon. Pharm. Drug Res. 2010, 67,
159–171.
[24] S. Dhillon, A. J. Wagstaff, Drugs 2007, 67, 2101–2108.
[25] H. A. Burris, H. I. Hurwitz, E. C. Dees, A. Dowlati, K. L.
Blackwell, B. O’Neil, P. K. Marcom, M. J. Ellis, B. Overmoyer, S.
F. Jones, J. L. Harris, D. A. Smith, K. M. Koch, A. Stead, S.
Mangum, N. L. Spector, J. Clin. Oncol. 2005, 23, 5305–5313.
[37] M. Tobe, Y. Isobe, H. Tomizawa, T. Nagazaki, H. Tahahashi, H.
Hayashi, Bioorg. Med. Chem. 2003, 11, 609–616.
[38] L. Pinto-Garcia, T. Efferth, A. Torres, J. D. Hoheisel, M. Youns,
[26] E. R. Wood, A. T. Truesdale, O. B. McDonald, D. Yuan, A.
Hassell, S. H. Dickerson, B. Ellis, C. Pennisi, E. Horne, K.
Lackey, K. J. Alligood, D. W. Rusnak, T. M. Gilmer, L.
Shewchuk, Cancer Res. 2004, 64, 6652–6659.
Planta Med. 2010, 76, 1155–1161.
[39] M. Youns, T. Efferth, J. Reichling, K. Fellenberg, A. Bauer, J. D.
Hoheisel, Biochem. Pharmacol. 2009, 78, 273–283.
ß 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com