Oriented synthesis and in vitro anticancer activity of biquinazoline-2,2'- diones

Article abstract of DOI:10.1021/cc9001247

The synthesis of a series of biquinazoline-2,2'-diones starting from o-nitrobenzaldehydes, anilines, and triphosgene is presented. This general approach features a novel and easy way for access to the target products. The mechanistic course of the reaction suggests the involvement of reduction, coupling, and cyclization by one-pot. These compounds were also investigated in vitro for anticancer activity, and some were found to have good anticancer activity.

Full text of DOI:10.1021/cc9001247

J. Comb. Chem. 2010, 12, 195–199
195
Oriented Synthesis and In Vitro Anticancer Activity of
Biquinazoline-2,2′-diones
Guolan Dou,† Daqing Shi,*,† and Yonghai Li‡
Key Laboratory of Organic Synthesis of Jiangsu ProVince, College of Chemistry, Chemical Engineering
and Materials Science, Soochow UniVersity, Suzhou 215123, P. R. China, and Life Science and
Technique Department, Xinxiang Medical UniVersity, Xinxiang 453003, P. R. China
ReceiVed August 14, 2009
The synthesis of a series of biquinazoline-2,2′-diones starting from o-nitrobenzaldehydes, anilines, and
triphosgene is presented. This general approach features a novel and easy way for access to the target products.
The mechanistic course of the reaction suggests the involvement of reduction, coupling, and cyclization by
one-pot. These compounds were also investigated in vitro for anticancer activity, and some were found to
have good anticancer activity.
Introduction
2-nitrobenzamide and triphosgene or isothiocyanates pro-
moted by low-valent titanium reagent.20,21 As our earlier
work goes, in this paper, we would like to report a novel
access to the construction of the biquinazoline-2,2′-dione
skeleton (1). For our target structure 1, the most efficient
route would require the reductive coupling of N-(2-nitroben-
zylidene)aniline derivatives 2 with triphosgene (Scheme 1).
N-(2-Nitrobenzylidene)aniline can be obtained from corre-
sponding o-nitrobenzaldehydes and anilines. Therefore, we
now report for the first time an efficient strategy for the
synthesis of biquinazoline-2,2′-diones from o-nitrobenzal-
dehydes, anilines, and triphosgene.
It is well-known that heterocycles are abundant in nature
and are of great significance to life because their structural
subunits exist in many natural products such as vitamins,
hormones, antibiotics, and alkaloids, as well as pharmaceu-
ticals, herbicides, dyes, and many other compounds. Quinazoli-
nones are an important class of heterocycles1 with potential
pharmacological activities including antibacterial,2 antihy-
pertensive,3 antitumor,4,5 anti-inflammatory,6 antidiabetic
agents7 and protein kinase inhibitors.8,9 Moreover, the
quinazolinone moiety has been extensively utilized as a
druglike scaffold in medicinal chemistry.10,11 The quinazoli-
none derivatives can generally be prepared by the reaction
of anthranilamides with aldehydes or ketones under either
acidic or basic conditions,12-15 and the typical preparation
of anthranilamides, in turn, proceeds by the reaction of
anilines with isatoic anhydride. However, this is not an
effective protocol if the anilines are substituted with an
electron-withdrawing group16 or present either two ortho
substituents or one bulky ortho substituent. In these cases,
the aniline does not react with isatoic anhydride even at
elevated temperatures.17 Yoo reported a novel one-step
synthesis of 2,3-dihydro-1H-quinazoline-4-ones from 2-ni-
trobenzamides;18 however, the reaction must reflux 2 days,
which is too long. Reaction of anthranilic acids with
carboxylic acids in the presence of P(OPh)3 in pyridine at
150 °C for 10 min under microwave irradiation yielded a
library of quinazolinones;19 the reaction temperature limited
the application of the method. Additionally, the synthesis of
biquinazoline-2,2′-diones (Figure 1) is rather scarce, there-
fore, a novel and general approach would be desirable and
of high value.
The synthesis of 1 is depicted in Scheme 2. N-(2-
Nitrobenzylidene)aniline 2 was prepared from o-nitroben-
zaldehyde and aniline in EtOH, which is simple, cheaply
priced, and readily available. With compound 2 in hand we
Figure 1. Biquinazoline-2,2′-diones.
Scheme 1. Retrosynthetic Pathway for the Synthesis of
Biquinazoline-2,2′-diones (1)
Previously, we described a facile synthesis of quinazoli-
none derivatives by the novel reductive cyclization of
* To whom correspondence should be addressed. E-mail: dqshi@
suda.edu.cn. Phone: +86-512-65880049. Fax: +86-512-65880089.
† Soochow University.
‡ Xinxiang Medical University.
10.1021/cc9001247  2010 American Chemical Society
Published on Web 12/22/2009

196 Journal of Combinatorial Chemistry, 2010 Vol. 12, No. 1
Dou et al.
Scheme 2. Reagents and Conditions: (a) EtOH, Reflux 2 h; (b) TiCl4/Sm, Triphosgene, THF, Reflux 2 h
Table 1. Optimization of Reaction Conditions for Compound
1a
under the optimized conditions, and the corresponding
products were obtained in good yields; therefore, no remark-
able electronic effects on the reaction were observed. We
were also pleased to find that no significant steric effects
were observed for the ortho-, meta-, and para-substituted
anilines, especially, the anilines presenting two ortho sub-
stituents can also give moderate to good yields (Table 2,
entry 19). Good yield was also obtained when N-(2-
nitrobenzylidene)naphthalene-1-amine reacted with triphos-
gene (Table 2, entry 20).
In our study, we ran the reaction of 2n with the absence
of triphosgene. However, tetraamine 6 can not be obtained
and the reductive coupling product 2-(4-methylphenyl)-2H-
indazole 7 was surprisingly obtained in 30% yield (Scheme
3). Therefore, we think that nitro group reduction precedes
reductive coupling.
entry
TiCl4/M
ratioa
yield/%
1
2
3
4
5
6
7
8
9
TiCl4/Zn
TiCl4/Mg
TiCl4/Al
TiCl4/Fe
TiCl4/Sm
TiCl4/Sm
TiCl4/Sm
TiCl4/Sm
TiCl4/Sm
1:4
1:4
1:4
1:4
1:4
1:1
1:2
1:3
1:5
85
69
62
57
94
72
78
89
93
Although the mechanism of the reaction is presently
unclear, a few possible sequences of events are outlined in
Scheme 4. First of all, TiCl4 is reduced by Sm dust to give
low-valent titanium species. In the initial step, because nitro
group reduction precedes reductive coupling, nitro compound
was reduced by low-valent titanium to amine 8, which was
then reacted with triphosgene to give compound 9. Then
compound 9 was reductive coupled and produce intermediate
10 by low-valent titanium reagent.22 Then 11 was obtained
by addition, and product 1 was obtained by hydrolysis.
a This ratio is the ratio of 2a and low-valent titanium reagent.
then looked at the reductive coupling of double linkage, nitro
group of the Schiff base 2 and triphosgene to give biquinazo-
line-2,2′-diones 1.
1
All the products were characterized by H NMR, 13C
NMR, IR, and HRMS spectra. The structure of 1e was further
confirmed by X-ray diffraction analysis. The molecular
structure of the product 1e is shown in Figure 2.
Finally, selected compounds were screened for anticancer
activity by the sulforhadamine B (SRB) method. The cells
used in this study were Human liver cancer Bel7402. The
results of the prescreening were listed in Figure 3.
Initially, the reaction of N-(5-chloro-2-nitrobenzylidene)-
4-fluoroaniline 2a and triphosgene at reflux was examined.
As a preliminary study, low-valent titanium system and the
ratio of 2a and low-valent titanium reagent were screened
in the model reaction. The results are shown in Table 1.
It was observed that among the screened low-valent
systems, TiCl4/Sm was superior to other low-valent titanium
systems (Table 1, entries 1-4). To further optimize reaction
conditions, a similar test was carried out at ratios of 2a and
low-valent titanium reagent ranging from 1:1 to 1:5. The
yield of product 1a was increased as the ratio was increased
from 1:1 to 1:4 (Table 1, entries 5, 6-8). However, a further
increase of the ratio to 1:5 failed to improve the yield of
product 1a (Table 1, entry 9). Therefore, 1:4 was chosen
for all further reactions.
Then we chose 1b, 1h, and 1k to determine the IC50
further. The results are shown in Table 3.
Among the compounds synthesized, the most active
compound was 1b. The IC50 values for 1b, 1h, and 1k were
0.33 µM, 2.13 µM, and 1.04 µM, respectively. Good to
improved anticancer activity was observed for them. How-
ever, the activity of compounds 1d and 1i were lower than
that of other compounds, and the reason is presently unclear.
With the optimized conditions in hand, we then performed
the reaction of a variety of N-(2-nitrobenzylidene)aniline 2
and triphosgene 3 via low-valent titanium reagent (TiCl4/
Sm). The results are summarized in Table 2.
As shown in Table 2, we were pleased to find that the
method was applicable to a broad substrate scope on N-(2-
nitrobenzylidene)anilines. Anilines containing various electron-
donating and electron-withdrawing substituents were reacted
Conclusions
In conclusion, we have developed an efficient and con-
venient method for the preparation of biquinazoline-2,2′-
diones. The process was carried out in tetrahydrofuran (THF)
via TiCl4/Sm to generate diversity on the biquinazolindiones.
A variety of substrates can participate in the process with
good yields. The procedure used commercially available

Synthesis of a Series of Biquinazoline-2,2′-diones
Journal of Combinatorial Chemistry, 2010 Vol. 12, No. 1 197
Table 2. Synthesis of Biquinazoline-2,2′-diones 1 Induced by TiCl4/Sm System
13C NMR (100 MHz) spectra were recorded on a Bruker
Scheme 3. Reaction with the Absence of Triphosgene
DPX-400 MHz spectrometer in DMSO-d6 solution. J values
are in hertz. Chemical shifts are expressed in parts per million
downfield from internal standard TMS. The exact mass
measurements were obtained by high resolution mass instru-
ment (GCT-TOF instrument).
General Procedure for the Synthesis of Biquinazoline-
2,2′-diones 1. TiCl4 (0.5 mL, 4 mmol) was added dropwise
using a syringe to a stirred suspension of samarium powder
(0.6 g, 4 mmol) in freshly distilled anhydrous THF (10 mL)
at room temperature (r.t.) under a dry N2 atmosphere. After
completion of the addition, the mixture was refluxed for 2 h.
The suspension of the low-valent titanium reagent formed
was cooled to r.t., and a solution of N-(2-nitrobenzylidene)
anilines (1 mmol) and triphosgene (1 mmol) in THF (5 mL)
was added dropwise. The reaction mixture was then refluxed
for 2 h under N2 atmosphere. After this period, the thin-
layer chromatography (TLC) analysis of the mixture showed
the completion of this reaction. The mixture was then
quenched with 5% HCl (30 mL) and extracted with
ClCH2CH2Cl (3 × 50 mL). The extracts were washed with
water (3 × 50 mL) and dried over anhydrous Na2SO4. After
evaporation of the solvent under reduced pressure, the crude
products were purified by recrystallization from 95% ethanol
and N,N-dimethylformamide (DMF).
o-nitrobenzaldehydes and anilines and is suitable for library
synthesis in drug discovery efforts. In particular, the product
could easily be collected by recrystallization. The short
reaction time and easily available materials render this
method particularly attractive for the efficient preparation
of biologically and medicinally interesting molecules. Im-
portantly, almost all of the compounds exhibited good
anticancer activity. It should be pointed out that this is the
first report of anticancer activity among these compounds.
Experimental Section
General Information. THF was distilled from sodium-
benzophenone just prior to use. All the reactions were
conducted under N2 atmosphere. Melting Points are uncor-
rected. IR spectra were recorded on a Tensor 27 spectrometer
1
in KBr with absorptions in cm-1. H NMR (400 MHz) and

198 Journal of Combinatorial Chemistry, 2010 Vol. 12, No. 1
Scheme 4. Supposed Reaction Mechanism
Dou et al.
Table 3. IC50 of Compounds 1b, 1h, and 1k
6,6′-Dichloro-3,3′-bis(4-fluorophenyl)-3,3′,4,4′-tetrahy-
1
dro-4,4′-biquinazoline-2,2′-dione (1a). H NMR (400 Hz,
DMSO-d6) (δ, ppm): 9.53 (br, s, 2H, 2 × NH), 7.40-7.36
(m, 4H, ArH), 7.23 (d, J ) 2.4 Hz, 2H, ArH), 7.16 (dd, J1
) 2.4 Hz, J2 ) 8.4 Hz, 2H, ArH), 7.06 (t, J ) 8.8 Hz, 4H,
ArH), 6.59 (d, J ) 8.8 Hz, 2H, ArH), 5.25 (s, 2H, 2 × CH).
13C NMR (100 Hz, DMSO-d6) (δ, ppm): 161.9, 158.7,
152.1, 137.9, 137.8, 137.6, 129.4, 129.4, 129.2, 127.2, 125.3,
119.4, 115.9, 115.4, 65.3 (4 C).
IR (KBr) ν: 3197, 3054, 2931, 1684, 1597, 1507, 1447,
1392, 1306, 1218, 1156, 1091, 941, 857, 831, 817, 743, 716,
663 cm-1.
HRMS (ESI): m/z calcd for: 573.0673 [M+Na]+, found:
573.0674.
Figure 2. Molecular structure of product 1e.
6,6′-Dichloro-3,3′-bis(4-methylphenyl)-3,3′,4,4′-tetrahy-
1
dro-4,4′-biquinazoline-2,2′-dione (1c). H NMR (400 Hz,
DMSO-d6) (δ, ppm): 9.35 (br, s, 2H, 2 × NH), 7.25 (d, J )
8.4 Hz, 4H, ArH), 7.22 (d, J ) 2.4 Hz, 2H, ArH), 7.19 (dd,
J1 ) 2.4 Hz, J2 ) 8.4 Hz, 2H, ArH), 7.07 (d, J ) 8.4 Hz,
4H, ArH), 6.58 (d, J ) 8.4 Hz, 2H, ArH), 5.15 (s, 2H, 2 ×
CH), 2.28 (s, 6H, 2 × CH3).
13C NMR (100 Hz, DMSO-d6) (δ, ppm): 152.2, 139.1,
137.7, 135.8, 129.8, 129.3, 127.3, 127.2, 125.1, 119.1, 115.2,
64.6 (4 C), 21.2.
IR (KBr) ν: 3197, 3052, 2925, 1679, 1598, 1501, 1447,
1389, 1312, 1212, 1165, 1132, 1111, 1091, 1018, 995, 935,
821, 741, 717, 669 cm-1.
HRMS (ESI): m/z calcd for: 565.1174 [M+Na]+, found:
565.1235.
Figure 3. Inhibitor ratio of the products by prescreening.

Synthesis of a Series of Biquinazoline-2,2′-diones
Journal of Combinatorial Chemistry, 2010 Vol. 12, No. 1 199
(2) Kung, P. P.; Casper, M. D.; Cook, K. L.; Wilson-Lingardo,
L.; Risen, L. M.; Vickers, T. A.; Ranken, R.; Blyn, L. B.;
Wyatt, J. R.; Cook, P. D.; Ecker, D. J. J. Med. Chem. 1999,
42, 4705–4713.
3,3′-Bis(4-chlorophenyl)-3,3′,4,4′-tetrahydro-4,4′-
1
biquinazoline-2,2′-dione (1l). H NMR (400 Hz, DMSO-
d6) (δ, ppm): 9.31 (br, s, 2H, 2 × NH), 7.40 (d, J ) 8.8 Hz,
4H, ArH), 7.29 (d, J ) 8.8 Hz, 4H, ArH), 7.15 (d, J ) 7.2
Hz, 2H, ArH), 7.11 (t, J ) 7.6 Hz, 2H, ArH), 6.85 (t, J )
7.6 Hz, 2H, ArH), 6.56 (d, J ) 7.6 Hz, 2H, ArH), 5.19 (s,
2H, 2 × CH).
(3) Hess, H. J.; Cronin, T. H.; Scriabine, A. J. Med. Chem. 1968,
11, 130–136.
(4) Baek, D.; Park, Y.; Heo, H. I.; Lee, M.; Yang, Z.; Choi, M.
Bioorg. Med. Chem. Lett. 1998, 8, 3287–3290.
13C NMR (100 Hz, DMSO-d6) (δ, ppm): 151.0, 137.1,
136.7, 136.7, 128.5, 128.4, 128.1, 126.1, 120.3, 116.0, 114.7,
114.5, 112.5, 108.8, 63.9 (4 C).
(5) Webber, S. E.; Bleckman, T. M.; Attard, J.; Deal, J. G.;
Kathardekar, V.; Welsh, K. M.; Webber, S.; Janson, C. A.;
Matthews, D. A.; Smith, W. W. J. Med. Chem. 1993, 36, 733–
746.
IR (KBr) ν: 3212, 3068, 2918, 1674, 1600, 1491, 1448,
1419, 1389, 1311, 1286, 1247, 1218, 1155, 1092, 1035, 1014,
872, 847, 828, 744, 659 cm-1.
(6) Chao, Q.; Deng, L.; Shih, H.; Leoni, L. M.; Genini, D.; Carson,
D. A.; Cottam, H. B. J. Med. Chem. 1999, 42, 3860–3873.
(7) Malamas, M. S.; Millen, J. J. Med. Chem. 1991, 34, 1492–
1503.
HRMS (ESI): m/z calcd for: 537.0861 [M+Na]+, found:
537.0877.
(8) Gibson, K. H.; Grundy, W.; Godfrey, A. A.; Woodburn, J. R.;
Ashton, S. E.; Curry, B. J.; Scarlett, L.; Barker, A. J.; Brown,
D. S. Bioorg. Med. Chem. Lett. 1997, 7, 2723–2728.
(9) Myers, M. R.; Setzer, N. N.; Spada, A. P.; Zulli, A. L.; Hsu,
C. J.; Zilberstein, A.; Johnson, S. E.; Hook, L. E.; Jacoski,
M. V. Bioorg. Med. Chem. Lett. 1997, 7, 417–420.
(10) Horton, D. A.; Bourne, G. T.; Smythe, M. L. Chem. ReV. 2003,
103, 893–930.
3,3′-Bis(3-methoxyphenyl)-3,3′,4,4′-tetrahydro-4,4′-
1
biquinazoline-2,2′-dione (1q). H NMR (400 Hz, DMSO-
d6) (δ, ppm): 9.14 (br, s, 2H, 2 × NH), 7.23-7.16 (m, 4H,
ArH), 7.11 (t, J ) 7.2 Hz, 2H, ArH), 7.00 (d, J ) 7.6 Hz,
2H, ArH), 6.91-6.85 (m, 4H, ArH), 6.75 (dd, J1 ) 2.0 Hz,
J2 ) 8.0 Hz, 2H, ArH), 6.53 (d, J ) 8.0 Hz, 2H, ArH), 5.20
(s, 2H, 2 × CH), 3.65 (s, 6H, 2 × CH3O).
(11) For example, Raltitrexed (Tomudex, marketed for colorectal
cancer), Ispinesib (phase II for solid tumors), and Temposatin
(phase II for bladder cancer).
13C NMR (100 Hz, DMSO-d6) (δ, ppm): 152.1, 151.9,
139.1, 138.9, 138.2, 130.3, 129.6, 129.6, 129.3, 129.3, 127.8,
127.7, 127.5, 127.3, 121.7, 121.5, 119.6, 119.3, 119.3, 117.9,
117.4, 116.8, 116.8, 116.5, 114.1, 113.9, 67.4 (4 C), 66.2
(4′ C).
(12) Ozaki, K.; Yamada, Y.; Oine, T.; Ishizuka, T.; Iwasawa, Y.
J. Med. Chem. 1985, 28, 568–576.
(13) Yale, H. L. J. Heterocycl. Chem. 1977, 14, 1357–1359.
(14) Kilroe Smith, T. A.; Stephen, H. Tetrahedron 1957, 1, 38–
44.
(15) Feldman, J. R.; Wagner, E. C. J. Org. Chem. 1942, 7, 31–47.
(16) Tani, J.; Yamada, Y.; Oine, T.; Ochiai, T.; Ishida, R.; Inoue,
I. J. Med. Chem. 1979, 22, 95–99.
IR (KBr) ν: 3184, 3061, 2994, 2937, 1679, 1602, 1496,
1443, 1421, 1302, 1265, 1226, 1204, 1169, 1045, 815, 773,
760, 743, 697 cm-1.
HRMS (ESI): m/z calcd for: 529.1852 [M+Na]+, found:
529.1883.
(17) Coyne, W. E.; Cusic, J. W. J. Med. Chem. 1968, 11, 1208–
1213.
(18) Yoo, C. L.; Fettinger, J. C.; Kurth, M. J. J. Org. Chem. 2005,
Acknowledgment. Financial support from the Foundation
of Key Laboratory of Organic Synthesis of Jiangsu Province
is gratefully acknowledged.
70, 6941–6943.
(19) Liu, J. F.; Kaselj, M.; Isome, Y.; Ye, P.; Sargent, K.; Sprague,
K.; Cherrak, D.; Wilson, C. J.; Si, Y.; Yohannes, D.; Ng, S. C.
J. Comb. Chem. 2006, 8, 7–10.
(20) Shi, D. Q.; Dou, G. L.; Li, Z. Y.; Ni, S. N.; Li, X. Y.; Wang,
X. S.; Wu, H.; Ji, S. J. Tetrahedron 2007, 63, 9764–9773.
(21) Dou, G. L.; Wang, M. M.; Shi, D. Q. J. Comb. Chem. 2009,
11, 151–154.
Supporting Information Available. Detailed descriptions
of experimental procedures and spectroscopic and analytical
data are available for compounds 1. This material is available
free of charge via the Internet at http://pubs.acs.org.
(22) Periasamy, M.; Srinivas, G.; Karunakar, G. V.; Bharathi, P.
References and Notes
Tetrahedron Lett. 1999, 40, 7577–7580.
(1) For a recent and general synthetic review, see: Connolly, D. J.;
Cusack, D.; O’Sullivan, T. P.; Guiry, P. J. Tetrahedron 2005,
61, 10153–10202.
CC9001247