Discovery of 2-methoxy-3-phenylsulfonamino-5-(quinazolin-6-yl or quinolin-6-yl)benzamides as novel PI3K inhibitors and anticancer agents by bioisostere

Article abstract of DOI:10.1016/j.ejmech.2014.01.053

2-Substituted-3-sulfonamino-5-(quinazolin-6-yl or quinolin-6-yl)benzamides have been proposed as novel structures of PI3K inhibitors and anticancer agents based on bioisostere. In the present study, 2-substituted-3-sulfonamino-5-(4- morpholinoquinazolin-6-yl)benzamides and 2-methoxy-3-sulfonamino-5-(4- morpholinoquinolin-6-yl)benzamides were synthesized. Their antiproliferative activities in vitro were evaluated via MTT assay against four human cancer cell lines, including A549, HCT-116, U-87 MG and KB. The SAR of the title compounds was preliminarily discussed. Compound 1a with potent antiproliferative activity was tested for its inhibitory activity against PI3K and mTOR and its effect on the AKT and p-AKT⁴⁷³. The anticancer effect of 1a was evaluated in established nude mice U-87 MG xenograft model. The results suggest that compound 1a can significantly inhibit PI3K/AKT/mTOR pathway and tumor growth. These findings strongly support the assumption that title compounds are potent PI3K inhibitors and anticancer agents.

Full text of DOI:10.1016/j.ejmech.2014.01.053

European Journal of Medicinal Chemistry 75 (2014) 96e105
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Discovery of 2-methoxy-3-phenylsulfonamino-5-(quinazolin-6-yl or
quinolin-6-yl)benzamides as novel PI3K inhibitors and anticancer
agents by bioisostere
,
,
Teng Shao a 1, Juan Wang b 1, Jian-Gang Chen a, Xiao-Meng Wang a, Huan Li a, Yi-Ping Li a,
Yan Li a, Guang-De Yang a, Qi-Bing Mei b , San-Qi Zhang
,
a,
*
*
a Department of Medicinal Chemistry, School of Pharmacy, Health Science Center, Xi’an Jiaotong University, Xi0an 710061, PR China
b Center for Pharmacological Evaluation and Research, Shanghai Institute of Pharmaceutical Industry, Shanghai 200437, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
2-Substituted-3-sulfonamino-5-(quinazolin-6-yl or quinolin-6-yl)benzamides have been proposed as
novel structures of PI3K inhibitors and anticancer agents based on bioisostere. In the present study, 2-
substituted-3-sulfonamino-5-(4-morpholinoquinazolin-6-yl)benzamides and 2-methoxy-3-sulfona
mino-5-(4-morpholinoquinolin-6-yl)benzamides were synthesized. Their antiproliferative activities
in vitro were evaluated via MTT assay against four human cancer cell lines, including A549, HCT-116, U-
87 MG and KB. The SAR of the title compounds was preliminarily discussed. Compound 1a with potent
antiproliferative activity was tested for its inhibitory activity against PI3K and mTOR and its effect on the
AKT and p-AKT473. The anticancer effect of 1a was evaluated in established nude mice U-87 MG xenograft
model. The results suggest that compound 1a can significantly inhibit PI3K/AKT/mTOR pathway and
tumor growth. These findings strongly support the assumption that title compounds are potent PI3K
inhibitors and anticancer agents.
Received 3 August 2013
Received in revised form
17 December 2013
Accepted 25 January 2014
Available online 29 January 2014
Keywords:
Benzamide
Quinazoline
PI3K inhibitor
Antiproliferative activity
Anticancer effect
Bioisostere
Ó 2014 Elsevier Masson SAS. All rights reserved.
1. Introduction
remarkable progress has been made on the design, synthesis and
evaluation of PI3Ka and mTOR dual inhibitors [4e11], and there-
Cancer is characterized by uncontrolled cell growth and prolif-
eration, which usually results from the abnormal activation of some
enzymes. In many cancers, the PI3K/AKT/mTOR signal transduction
pathway is dysregulated, contributing to cellular transformation
and tumor growth [1]. Thus, the phosphatidylinositol 3-kinase
(PI3K, a family of lipid kinases) signaling pathway plays a crucial
role in cell growth, proliferation and survival. Genomic aberrations
upon the pharmacophore of the dual inhibitors has been put for-
ward [12]. Among the reported PI3K and mTOR dual inhibitors, N-
(5-(quinolin-6-yl)pyridin-3-yl)phenylsulfonamide is
a class of
important active compounds. In these structures, the two ring ni-
trogen atoms in pyridine and quinoline are the main components of
pharmacophore.
N-(2-Methoxy-5-(4-pyridazin-4-yl)quinolin-6-
ylpyridin-3-yl)-2,4-difluorophenylsulfonamide, GSK2126458, was
identified by GlaxoSmithKline as a potent, orally bioavailable dual
in the PI3K pathway such as mutational activation of PI3K
a or
dysfunction of tumor suppressor PTEN are closely linked to the
development and progression of a wide range of cancers, including
cancers of colon, breast, head and neck, and non-small cell lung.
Hence, inhibition of the key targets in the pathway, e.g. PI3K, AKT or
mTOR (mammalian target of rapamycin), has great potential for the
inhibitor of PI3Ka and mTOR [13]. Having designed, synthesized
and evaluated several classes of N-(2,5-disubstituted-pyridin-3-yl)
phenylsulfonamides, Amgen Inc discovered that N-(2-chloro-5-(4-
morpholinoquinolin-6-yl)pyridine-3-yl)-4-
fluorophenylsulfonamide [14], N-(2-chloro-5-(2-acetylaminobenzo
[d]thiazol-6-yl)pyridine-3-yl)-4-fluoro phenylsulfonamide [15] and
N-(2-chloro-5-(2-acetylaminoimidazo[1,2-b]pyridazin-6-yl)pyri-
treatment of cancer. Therefore, PI3Ka and mTOR have become the
targets of quest research for anticancer drug [2,3]. In recent years, a
dine-3-yl)-4-fluorophenylsulfonamide [16] were effective PI3K
and mTOR dual inhibitors. Recently, they have reported that AMG
511 is a potent PI3K selective inhibitor [17]. Our group synthesized
a series of [1,2,4]triazolo[1,5- ]pyridinylpyridines and evaluated
their anticancer activity [18]. Meanwhile, other group reported 3D-
a
* Corresponding authors.
a
E-mail addresses: qbmei@hotmail.com (Q.-B. Mei), sqzhang@xjtu.edu.cn,
sqzhang@mail.xjtu.edu.cn (S.-Q. Zhang).
a
1
T. Shao and J. Wang contributed equally to this work.
0223-5234/$ e see front matter Ó 2014 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2014.01.053

T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
97
QSAR and docking studies of 3-pyridinyl heterocyclic derivatives as
potent PI3K/mTOR inhibitors [19]. The first PI3K /mTOR dual in-
hibitor, BEZ235 developed by Novartis, is being in clinical trials [20].
The pharmacophore of PF-04691502 and PF-04979064 is similar to
that of GSK2126458. They are also PI3Ka/mTOR dual inhibitors,
developed by Pfizer in phase I/II clinical trials for treating solid
tumors [21e23]. (Fig. 1).
bromo-4-chloroquinozoline or 6-bromo-4-chloroquinoline by reac-
tion with thionyl chloride and phosphorus oxychloride. Without
further purification, the reaction of 4-chloroquinozoline or 6-bromo-
4-chloroquinoline with morpholine produced intermediate 6a or 6b.
Catalyzed by PdCl2(dppf), 6a or 6b was reacted with bis(pinacolato)
diboron to produce arylboronic esters. Without isolation of arylbor-
onic esters, intermediate 5, PdCl2(dppf), water and sodium carbonate
were added to the above reaction mixture. The resulted mixture was
refluxed for 1e2 h to produce the title compounds 1 or 2. The prep-
aration of arylboronic esters and Suzuki coupling were completed in
one pot. 1,2-dimethoxyethane was used as solvent to improve the
yield.
To investigate the different activity of substituted groups at 2-
position in the benzamide ring, 5-bromo-2-chloro-3-nitrobenzoic
acid and 5-bromo-3-nitrobenzoic acid were used as starting ma-
terials to synthesize the compounds 1iek. The synthetic route for
the compounds is shown in Scheme 2. The synthetic procedure is
similar to that of 1a.
a
The co-crystal of GSK2126458 with PI3Kg protein has clarified
the interaction of GSK2126458 with PI3K [13]. In the co-crystal
structure, we find that the nitrogen atom in the pyridine ring can
form two hydrogen bonds at the affinity pocket in PI3K kinase with
the aid of a water molecule. One hydrogen atom in water molecule
forms a hydrogen bond with the nitrogen atom in pyridine ring, the
other hydrogen atom forms a hydrogen bond with the carboxyl in
Asp841 and the oxygen atom in water molecule forms a hydrogen
bond with the phenolic hydroxyl in Tyr867. Meanwhile, the nitro-
gen atom at 1-position in the quinoline ring can form a hydrogen
bond with the Val882 at the hinge region in PI3K kinase. The
interaction is outlined in Fig. 2.
In order to obtain a novel scaffold of anticancer agents, we
propose that the complex of pyridine with a water molecule be
replaced by the structure of benzamide on the basis of bioisostere
(Fig. 2). In this way, benzamide can replace the pyridine ring in
GSK2126458 to produce a novel scaffold of PI3K inhibitors. In this
novel scaffold, we suspect that the oxygen atom in benzamide can
form a hydrogen bond with the phenolic hydroxyl in Tyr867, and
the hydrogen atom in benzamide can form a hydrogen bond with
the carboxyl in Asp841 as well, which might indicate that 2-
methoxy-3-phenylsulfonylamino-5-(quinolin-6-yl)benzamide
shares the same pharmacophore (denoted as red in Fig. 2) with
GSK2126458. In other words, 2-methoxy-3-phenylsulfonylamino-
5-(quinazolin-6-yl or quinolin-6-yl)benzamides might interact
with PI3K without the aid of water molecule, serve as a novel
scaffold of PI3K inhibitors and exhibit potential anticancer effect.
Thus, we further assume that this design may help us discover the
3. Pharmacology
3.1. Antiproliferative assays in vitro
We evaluated antiproliferative activities of compounds 1aek, 2a
and 2b against four human cancer cell lines, including human lung
adenocarcinoma epithelial cell line (A549), human colon carcinoma
cell line (HCT-116), human glioblastoma-astrocytoma epithelial-
like cell line (U-87 MG) and human epidermis mouth carcinoma
cell line (KB) by applying the MTT colorimetric assay. The PI3K and
mTOR dual inhibitor BEZ235 was used as the positive control.
3.2. PI3K and mTOR enzymatic activity assay
Selected compound 1a was evaluated for its PI3K and mTOR
enzymatic activity using a ATP depletion assay [24]. BEZ235 was
used as the positive drug. Compounds 1a and BEZ235 were dis-
solved in DMSO and tested at final concentrations of 1.0, 3.3, 10, 33,
100, 333, 1000, 3333, 10,000 nM. IC50s of compounds were calcu-
lated according to inhibitory rates using GraphPad Prism5.
novel PI3K inhibitors with
a scaffold of 2-substituted-3-
phenylsulfonylamino-5-(quinazolin-6-yl) benzamide by using
bioisostere principle. Hereafter in this paper, we describe the syn-
thesis of 2-substituted-3-phenylsulfonylamino-5-(quinazolin-6-yl
or quinolin-6-yl)benzamides, and the evaluation of their bioac-
tivity, anticancer effect in vivo as well.
3.3. Western blot assay
To further determine whether these compounds affect the PI3K/
AKT/mTOR signaling pathway, we evaluated the suppressive effects
2. Chemistry
of compound 1a at 10 mM and positive compound BEZ235 at 10 mM
The synthetic route for the title compounds 1aeh and 2aeb is
outlined in Scheme 1.
on AKT and p-AKT473 in HCT-116 cells through Western blot.
The commercially available 5-bromo-3-nitro-2-methoxybenzoic
acid 3a was converted into corresponding intermediate 4aed with
the conventional steps. Reduction of 4 with stannous chloride in
concentrated hydrochloric acid produced corresponding amines.
Without further purification, the amines were converted into sulfa-
mides 5aeh. The commercially available 6-bromo-4-hydroxyquin
ozoline or 6-bromo-4-hydroxyquinoline was converted into 6-
3.4. Anticancer effects in established nude mouse U-87 MG
xenograft model in vivo
To preliminarily test the anticancer effects of the synthesized
compounds in vivo, a test using U-87 MG xenograft model was
performed. Compound 1a displayed evident activities against the
F
F
S
HO
O
O
N
NC
O
HN
O
O
N
N
O
OH
MeO
O
N
O
N
N
N
N
N
N
N
N
N
N
N
N
NH2
N
N
GSK2126458
NVP-BEZ235
PF-04691502
PF-04979064
Fig. 1. Some structures of PI3Ka and mTOR dual inhibitors developing in clinical trials.

98
T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
H
H
N
F
F
S
N
R'
O
H
H
R'
O
S
Lys 833
O
Lys 833
O
S
HN
N
O
HN
N
O
HN
MeO
MeO
R
MeO
O
H
N
O
R
N
N
O
H
O
O
X
H
O
N
NH2
N
O
Asp 841
H
O
H
H
O
H
N
O
Asp 841
O
H
N
N
N
H
H
Tyr 867
Tyr 867
Val 882
Val 882
proposed structures
proposed interaction
Co-crystal structure of GSK2126458 with PI3Kr
Fig. 2. The design idea of the research subject. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
O
O
NO2
S
NO2
R'
HN
MeO
R
MeO
HO
MeO
R
b
a
R = OMe, NH2, NHMe, NMe2
R' = cPr, 4-FPh, 4-ClPh,
4-MePh, 2,4-diFPh
Br
Br
Br
O
O
O
3a
4a-4d
5a-5h
O
O
O
N
S
R'
HN
O
N
OH
Br
MeO
R
c
d
Br
X
X
N
X
N
N
O
6a: X = N; 6b: X = CH
1a-1h: X = N; 2a-2b: X = CH
Scheme 1. a) i: Me2SO4, K2CO3, acetone; or ii: SOCl2, DMF, toluene, reflux, 2 h; then amine, THF, r.t, 1 h. b) i: SnCl2$2H2O, conc. HCl, r.t, overnight; ii: sulfonyl chloride, pyridine, 0 ꢀC-
r.t, overnight. c) i: POCl3, SOCl2, toluene, DMF, reflux, 4 h; ii: morpholine, isopropanol, reflux, 1 h. d) i: bis(pinacolato)diboron, KOAc, PdCl2(dppf), 1,2-dimethoxyethane, reflux, 2 h; ii:
5, H2O, Na2CO3, PdCl2(dppf), reflux, 2 h.
four carcinoma cell lines and was thus chosen to evaluate its anti-
cancer effect in vivo. BEZ235 (20 mg/kg) was used as positive drug.
1a was dosed orally at 20 mg/kg or 50 mg/kg once a day for 12 days.
The control group was administrated orally the solvent only.
3.5. Docking studies
We performed docking analysis by utilizing the C-DOCKER
program within Discovery Studio 2.5 software package to further
O
O
NO2
NO2
S
HN
R
O
R'
R
O
a
b
R
O
HO
H2N
Br
Br
H2N
Br
3b, 3c
4e, 4f
5i-5k
O
O
O
N
S
R'
HN
O
N
R
c
Br
R = Cl, H
N
H2N
N
R' = 4-FPh, 4-ClPh
N
O
N
6a
1i-1k
Scheme 2. a) i: SOCl2, DMF, toluene, reflux, 1 h; ii: aqueous ammonia, THF, r.t, 1 h. b) i: SnCl2$2H2O, conc. HCl, r.t, overnight; ii: sulfonyl chloride, pyridine, 0 ꢀC-r.t, overnight. c) i:
bis(pinacolato)diboron, KOAc, PdCl2(dppf), 1,2-dimethoxyethane, reflux, 2 h; ii: 5, H2O, Na2CO3, PdCl2(dppf), reflux, 2 h.

T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
99
Table 2
look into the binding mode of the compound 1a. Docking simula-
Enzymatic activities of compound 1a against PI3K and mTORC1 (IC50, nM).
tions were performed on human PI3Kg (PDB code 3L08) [10].
Kinase
1a
65
14
190
56
BEZ235
4. Results and discussion
mTORC1
18
58
15
65
97
PI3K
PI3K
PI3K
PI3K
a
b
g
d
4.1. Chemistry
74
All the newly synthesized compounds show satisfactory ana-
lyses for the proposed structures, which were characterized on the
basis of their 1H NMR, 13C NMR and HRMS. The structures of re-
ported compounds were characterized by 1H NMR and HRMS.
(IC50 ¼ 4.71
mM against HCT-116), which suggests that a fragment of
quinazoline at 5-position of benzamide is favorable to maintain the
activity. From the above mentioned results we could detect a SAR in
which amino at R1, methoxy at R2 and aryl at R3 are favorable for
antiproliferative activity. In its antiproliferative activity, the IC50 of
compound 1a is comparable to that of PI3K and mTOR dual in-
hibitor BEZ235. Therefore, we further investigated compound 1a.
4.2. Antiproliferative assays in vitro
The results of the antiproliferative activities are summarized in
Table 1. Firstly, compounds 1aee exhibited significant activities
against the four cancer cell lines, especially against HCT-116, which
4.3. PI3K and mTOR enzymatic activity assay
might be related to PI3K
a
mutant in HCT-116. The fact that com-
M against HCT-116) was more potent than
31.60 against HCT-116) and 1j
pound 1a (IC50 ¼ 0.54
m
¼
The results of the PI3K and mTOR enzymatic activity are sum-
marized in Table 2.
compound 1i (IC50
mM
(IC50 ¼ 20.26
mM against HCT-116) indicates that the title com-
Compound 1a exhibited significant activities against PI3K and
pound with a methoxy at 2-position (R2) of benzamide ring may
improve the antiproliferative activity. To further investigate the
structureeactivity relationship (SAR) of compound 1, we synthe-
sized compounds 1f, 1g and 1h by changing substituent R1. As we
had expected, almost all the activities of compounds 1f and 1g
mTORC1. The activity against PI3K
other kinases tested. These results suggest that compound 1a is a
selective PI3K inhibitor.
a is four-fold higher than the
a
4.4. Western blot assay
(IC50 > 50 mM) were weaken or disappeared. The activity of 1h
distinctly decreased compared with 1a against the four cancer call
lines. These results strongly support our design idea that amino
group at R1 position is essential to maintain the interactions of
As shown in Fig. 3, the suppressive effect of compound 1a on p-
AKT473 was a slightly stronger than that of BEZ235. The results
suggest compound 1a inhibit the PI3K/AKT/mTOR pathway and
thus is a potential PI3K inhibitor.
inhibitor with PI3K protein. Compound 1e (IC50 ¼ 3.74
mM against
HCT-116) was less potent than 1a, 1b and 1c (IC50 ¼ 0.54e0.85
mM
against HCT-116), indicating that aromatic group at position of R3 is
more beneficial to activity than cyclopropyl group. When 4-
fluorophenyl (1a and 2a) was replaced by 2,4-difluorophenyl (1d
and 2b) at position of R3, the antiproliferative activities of com-
4.5. Anticancer effects in established nude mouse U-87 MG
xenograft model in vivo
pounds did not improve. The activities of 1a (IC50 ¼ 0.54
m
M against
The change in the tumor volumes was observed inFig. 4 (A). In this
model, treatment groups with compound 1a at 20 mg/kg or 50 mg/kg
significantly inhibit tumor growth by 30.05%and 57.20%, compared to
HCT-116) and 1d (IC50 ¼ 1.92
m
M against HCT-116) were more
potent than those of 2a (IC50 ¼ 3.84
mM against HCT-116) and 2b
Table 1
Antiproliferative activities of compounds 1aek and 2a, 2b against four human cancer cell lines (x ꢁ s, n ¼ 3).
O
O
S
R3
HN
O
N
R2
O
R1
X
N
Compds
R1
R2
R3
X
IC50
(
m
M)
A549
HCT-116
U87 MG
KB
1a
1b
1c
1d
1e
1f
1g
1h
1i
1j
1k
2a
NH2
NH2
NH2
NH2
NH2
NHMe
NMe2
OMe
NH2
NH2
NH2
NH2
NH2
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
Cl
H
H
OMe
OMe
4-FPh
N
N
N
N
N
N
N
N
N
N
N
H
H
4.32 ꢁ 1.52
3.85 ꢁ 1.52
2.18 ꢁ 1.10
5.52 ꢁ 2.14
9.80 ꢁ 1.36
>50
0.54 ꢁ 0.28
0.96 ꢁ 0.02
0.85 ꢁ 0.16
1.92 ꢁ 1.08
3.74 ꢁ 1.03
>50
1.37 ꢁ 0.40
1.12 ꢁ 0.07
1.06 ꢁ 0.57
2.88 ꢁ 1.66
5.21 ꢁ 1.87
>50
4.45 ꢁ 2.82
14.61 ꢁ 3.68
3.59 ꢁ 0.75
2.69 ꢁ 1.41
4.02 ꢁ 0.98
>50
4-ClPh
4-MePh
2,4-diFPh
cPr
4-FPh
4-FPh
4-FPh
4-FPh
4-FPh
4-ClPh
4-FPh
>50
>50
>50
>50
35.19 ꢁ 7.13
26.60 ꢁ 4.38
31.60 ꢁ 5.12
20.26 ꢁ 4.09
12.31 ꢁ 3.13
3.84 ꢁ 0.88
4.71 ꢁ 2.49
1.16 ꢁ 0.15
7.80 ꢁ 1.53
5.96 ꢁ 2.02
33.62 ꢁ 4.16
16.76 ꢁ 3.88
10.74 ꢁ 3.01
4.00 ꢁ 0.69
4.47 ꢁ 2.98
3.34 ꢁ 2.43
>50
>50
>50
>50
4.85 ꢁ 1.87
6.49 ꢁ 1.77
0.54 ꢁ 0.36
20.80 ꢁ 4.83
11.85 ꢁ 3.65
3.28 ꢁ 1.61
5.09 ꢁ ꢁ 2.77
1.32 ꢁ 0.41
2b
BEZ235
2,4-diFPh

100
T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
the control group, respectively. BEZ235 group displayed a delay in
tumor growth by 42.44% with 20 mg/kg. These results suggest that 1a
display significant inhibitory effect on the tumor growth.
The change in mouse body weights in the four groups is outlined
in Fig 4 (B). The loss of weight in the tested animals was observed in
two dosages groups of 1a and BEZ235 group, indicating that com-
pound 1a and BEZ235 display some toxicity at the dosage admin-
istered. After the medicative was stopped, mouse body weights
recovered gradually.
by thin-layer chromatography on silica gel plates (GF-254) and
visualized with UV light. All the melting points were determined on
a Beijing micromelting-point apparatus and thermometer was
uncorrected. 1H NMR spectra and 13C NMR spectra were recorded
on a 400 Bruker NMR spectrometer with tetramethylsilane (TMS)
as an internal reference. All chemical shifts are reported in parts per
million (ppm). High-resolution exact mass measurements were
performed using electrospray ionization (positive mode) on a
quadrupole time-of-flight (QTOF) mass spectrometer (Maxis Q-TOF,
Bruker Inc.).
4.6. Docking studies
6.1.1. Methyl 5-bromo-2-methoxy-3-nitrobenzate (4a)
The docking result of compound 1a with PI3Kg (Fig. 5) has the
To a solution of compound 3a (8.0 g, 29.0 mmol) in acetone
(100 ml) were added K2CO3 (6.5 g, 47.0 mmol) and Me2SO4 (4.5 ml,
45.0 mmol). The reaction mixture was refluxed with stirring for 6 h,
cooled to room temperature and filtrated. The filtrate was
concentrated in vacuum and the residue was suspended in water1
(100 ml). The mixture was extracted with ethyl acetate (40 ml ꢂ3).
The combined organic phase was washed with saturated solution of
Na2CO3 and brine, dried over anhydrous Na2SO4 and concentrated
to afford title compound (7.7 g, 87.0%) as an oil, which was solidified
following three indications: (a) 1-position of quinazoline group can
form a hydrogen bond with Val882; (b) The oxygen atom of benza-
midecan form one hydrogen bond with Tyr867. The hydrogen atom of
benzamide can form a hydrogen bond with Asp841. These interactions
indicate that benzamide can replace the complex of pyridine ring in
the structure of GSK2126458 with water molecule; (c) The oxygen of
sulfonylamide and methoxy group at 2-position of benzamide can
form two hydrogen bonds with Lys833. As for compound 1i or 1j,
chloride or hydrogen atom at 2-position of benzamide cannot form
hydrogen bond with Lys833, which suggest that the methoxygroup is
a more suitable substitute group than chloride in compound 1.
in one day keeping. mp 60.0e60.5 ꢀC. 1H NMR (CDCl3)
d: 8.17 (1H, d,
J ¼ 2.4 Hz, AreH), 8.05 (1H, d, J ¼ 2.4 Hz, AreH), 4.02 (3H, s, OCH3),
3.99 (3H, s, OCH3). ESI-HRMS m/z: calcd for C9H8BrNO5 [M þ H]þ:
288.9586; found 288.9590, 290.9568.
5. Conclusion
6.1.2. 5-Bromo-2-methoxy-3-nitrobenzamide (4b)
The present study proves that 2-substituted-3-sulfonamino-5-
(quinazolin-6-yl or quinolin-6-yl)benzamide can serve as a novel
structure of PI3K inhibitors and anticancer agents. Thirteen com-
pounds were synthesized, whose structures were characterized by
1H NMR, 13C NMR and HRMS. The results of antiproliferative ac-
tivities indicate that title compounds exhibit significant activity
against the four cancer cell lines, especially against HCT-116. The
SAR of these compounds indicates that a methoxy at 2-position of
benzamide ring can improve the inhibitory activity of title com-
pounds. The PI3K and mTOR enzymatic activity assay and Western
blot assay results of compound 1a suggest that 1a can block the
PI3K/AKT/mTOR pathway. Furthermore, compound 1a significantly
exhibited inhibitory effect on tumor growth in established nude
mice U-87 MG xenograft model. These findings strongly support
our hypothesis that the structure of benzamide can replace the
complex of pyridine with a water molecule to design novel PI3K
inhibitors and anticancer agents.
The mixture of 5-bromo-2-methoxy-3-nitrobenzoic acid 3a
(7.2 g, 26.0 mmol), thionyl chloride (10 ml, 137.0 mmol), toluene
(20 ml) and DMF (3 drops) was refluxed for 2 h. The volatile was
removed in vacuum to afford a brown oil which was diluted with
dry THF (20 ml). The obtained solution was added dropwise to the
stirred ammonia water cooled by an ice-water bath. The reaction
mixture was stirred at room temperature for 30 min. THF was
removed in vacuum and the resultant solid was collected by
filtration, washed with water and dried to yield 4b (6.9 g, 96.2%) as
a yellow solid. mp: 150.0e153.0 ꢀC. 1H NMR (DMSO-d6)
d, J ¼ 2.4 Hz, AreH), 8.04 (1H, s, CONH2), 7.91 (1H, d, J ¼ 2.4 Hz, Are
H), 7.87 (1H, s, CONH2), 3.87 (3H, s, OCH3). 13C NMR (DMSO-d6)
165.60, 149.26, 145.26, 136.14, 134.97, 128.59, 115.42, 63.78. ESI-
HRMS m/z: calcd for C8H8BrN2O4 [M þ H]þ: 274.9667; found
274.9665, 276.9644.
d: 8.25 (1H,
d
:
Compounds 4b and 4f were synthesized according to the pro-
cedure described in 4a.
6. Experimental protocols
6.1.3. N-Methyl-5-bromo-2-methoxy-3-nitrobenzamide (4c)
6.1. Chemistry and chemical methods
Yield 84.4%, mp 142.0e142.5 ꢀC. 1H NMR (CDCl3)
d: 8.41 (1H, d,
J ¼ 2.4 Hz, AreH), 8.05 (1H, d, J ¼ 2.4 Hz, AreH), 7.35 (1H, s, NH),
Unless specified otherwise, all starting materials, reagents and
solvents were commercially available. All reactions were monitored
3.98 (3H, s, OCH3), 3.08 (3H, d, J ¼ 5.2 Hz, NCH3). 13C NMR (DMSO-
d6) d: 164.23, 149.27, 145.20, 136.23, 134.82, 128.57, 115.41, 63.72,
Fig. 3. Effects of BEZ235 and compound 1a on AKT in HCT-116 cells. (A) Effect of BEZ235 and compound 1a on the AKT and p-AKT473. (B) The quantified effect of BEZ235 and
compound 1a on the AKT and p-AKT473. Results are x ꢁ s for three independent experiments each performed in duplicate. ***P < 0.001vs DMSO.

T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
101
Fig. 4. In vivo efficacy of 1a in U-87 MG xenograft tumor model on a once daily dosing schedule. (A) the change of tumor volume; (B) the change of mouse body weight. ***,
p < 0.001 compared with the control group.
26.74. ESI-HRMS m/z: calcd for C9H10BrN2O4 [M þ H]þ: 288.9824;
6.1.7. 5-Bromo-2-methoxy-3-(4-fluorophenylsulfonamino)
benzamide (5a)
found 288.9820, 290.9801.
To a suspension of compound 4a (6.9 g, 25.0 mmol) in
concentrated HCl (30 ml) was added SnCl2$2H2O (20.0 g,
88.0 mmol) in portions. After addition, the mixture was stirred at
room temperature overnight, basified to pH 11 with saturated
NaOH solution and extracted with ethyl acetate (70 ml ꢂ4). The
combined organic phase was washed with brine (100 ml ꢂ3), dried
over Na2SO4 and concentrated in vacuum to give reduced com-
pound (6.1 g, quantitative) as light-yellow solid. To a solution of
reduced product in pyridine (70 ml) was added 4-
fluorophenylsulfonyl chloride (5.4 g, 27.5 mmol) in portions at
0 ꢀC. Then the mixture was stirred at room temperature overnight.
Pyridine was removed under reduced pressure to give a residue,
which was solidified by adding H2O (70 ml). The solid was filtered,
washed with H2O and dried to give a brown solid, which was
recrystallized in ethanol to afford 5a (8.6 g, 85.3%) as white solid.
6.1.4. N,N-Dimethyl-5-bromo-2-methoxy-3-nitrobenzamide (4d)
Yield 87.3%, mp 96.0e98.0 ꢀC. 1H NMR (CDCl3)
d
: 8.00 (1H, d,
J ¼ 2.4 Hz, AreH), 7.65 (1H, d, J ¼ 2.4 Hz, AreH), 3.95 (3H, s, OCH3),
3.17 (3H, s, NCH3), 2.91 (3H, s, NCH3). 13C NMR (DMSO-d6)
: 164.93,
d
148.26, 144.65, 135.33, 134.84, 128.24, 115.64, 63.11, 38.50, 34.87.
ESI-HRMS m/z: calcd for C10H12BrN2O4 [M þ H]þ: 302.9980; found
302.9976; 302.9956.
6.1.5. 5-Bromo-2-chloro-3-nitrobenzamide (4e)
Yield 91.6%, mp 191.0e192.0 ꢀC. 1H NMR (DMSO-d6)
d
: 8.41 (1H,
s, AreH), 8.17 (1H, s, CONH2), 8.01 (1H, s, AreH), 7.95 (1H, s,
CONH2). 13C NMR (DMSO-d6)
: 165.55, 149.61, 141.38, 134.58,
d
128.10, 121.14, 120.85. ESI-HRMS m/z: calcd for C7H5BrClN2O3
[M þ H]þ: 278.9172; found 278.9170, 280.9148.
mp 186.0e188.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.18 (1H, s, SO2NH), 7.89
(2H, dd, J ¼ 8.8, 4.2 Hz, AreH), 7.79 (1H, s, CONH2), 7.63 (1H, s,
CONH2), 7.48 (1H, d, J ¼ 1.6 Hz, AreH), 7.46 (2H, t, J ¼ 9.0 Hz, AreH),
7.34 (1H, d, J ¼ 1.6 Hz, AreH), 3.45 (3H, s, OCH3). 13C NMR (DMSO-
6.1.6. 5-Bromo-3-nitrobenzamide (4f)
Yield 84.2%, mp 188.0e190.5 ꢀC. 1H NMR (DMSO-d6): 8.63 (1H, s,
AreH), 8.51 (1H, s, AreH), 8.44 (1H, s, AreH), 8.10 (2H, s, CONH2).
d6)
d
: 166.64, 164.94 (d, JCeF ¼ 250 Hz), 149.09, 136.74, 132.75,
13C NMR (DMSO-d6)
d: 164.76, 149.10, 137.72, 136.71, 128.98, 122.61,
132.44, 130.25, 130.15, 128.14, 126.73, 117.19, 116.96, 115.23, 61.99.
ESI-HRMS m/z: calcd for C14H13BrFN2O4S [M þ H]þ: 402.9763;
found 402.9760, 402.9740.
121.98. ESI-HRMS m/z: calcd for C7H6BrN2O3 [M þ H]þ: 244.9562;
found 244.9560, 246.9538.
Compounds 5bek were synthesized according to the procedure
described in 5a.
6.1.8. 5-Bromo-2-methoxy-3-(4-chlorophenylsulfonamino)
benzamide (5b)
Yield 84.3%, mp 177.5e178.5 ꢀC. 1H NMR (DMSO-d6)
d: 10.26 (1H,
s, SO2NH), 7.82 (2H, d, J ¼ 8.8 Hz, AreH), 7.81 (1H, s, CONH2), 7.70
(2H, d, J ¼ 8.8 Hz, AreH), 7.64 (1H, s, CONH2), 7.48 (1H, d, J ¼ 2.4 Hz,
AreH), 7.34 (1H, d, J ¼ 2.4 Hz, AreH), 3.45 (3H, s, OCH3). 13C NMR
(DMSO-d6) d: 166.64, 149.15, 139.24, 138.54, 132.77, 132.31, 130.02,
129.02, 128.25, 126.83, 115.22, 61.98. ESI-HRMS m/z: calcd for
C
14H13BrClN2O4S [M þ H]þ: 418.9468; found 418.9466, 420.9443.
6.1.9. 5-Bromo-2-methoxy-3-(4-methylphenylsulfonamino)
benzamide (5c)
Yield 87.4%, mp 169.0e170.5 ꢀC. 1H NMR (DMSO-d6)
d: 10.11 (1H,
s, SO2NH), 7.81 (1H, s, CONH2), 7.72 (2H, d, J ¼ 8.0 Hz, AreH), 7.64
(1H, s, CONH2), 7.50 (1H, d, J ¼ 2.4 Hz, AreH), 7.41 (2H, d, J ¼ 8.4 Hz,
AreH), 7.29 (1H, d, J ¼ 2.4 Hz, AreH), 3.44 (3H, s, OCH3), 2.36 (3H, s,
AreCH3). 13C NMR (DMSO-d6)
d: 166.65, 148.58, 144.16, 137.42,
132.88, 132.76, 130.28, 127.55, 127.14, 125.82, 115.29, 62.01, 21.46.
ESI-HRMS m/z: calcd for C15H16BrN2O4S [M þ H]þ: 399.0014; found
399.0010, 400.9990.
Fig. 5. Docking mode of compound 1a with PI3Kg. Selected residues Val882, Tyr867,
Lys833, Asp841 and Asp964 are shown. Green dashed lines indicate hydrogen bond.
(For interpretation of the references to color in this figure legend, the reader is referred
to the web version of this article.)

102
T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
6.1.10. 5-Bromo-2-methoxy-3-(2,4-difluorophenylsulfonamino)
benzamide (5d)
6.1.16. 5-Bromo-3-(4-fluorophenylsulfonamino)benzamide (5j)
Yield 83.1%, mp 238.0e240.0 ꢀC. 1H NMR (DMSO-d6)
: 10.77
d
Yield 75.4%, mp 152.0e153.0 ꢀC. 1H NMR (CDCl3)
d: 8.05 (1H, m,
(1H, s, SO2NH), 8.06 (1H, s, AreH), 7.84 (2H, dd, J ¼ 8.8, 5.2 Hz, Are
AreH), 7.81 (1H, d, J ¼ 1.6 Hz, AreH), 7.73 (1H, d, J ¼ 1.6 Hz, AreH),
7.47 (1H, s, CONH), 7.09 (1H, m, AreH), 7.05 (1H, s, CONH), 7.00 (1H,
m, AreH), 5.84 (1H, s, SO2NH), 3.82 (3H, s, OCH3). 13C NMR (DMSO-
H), 7.74 (1H, s, CONH2), 7.60 (1H, s, AreH), 7.56 (1H, s, CONH2), 7.44
(2H, t, J ¼ 8.8 Hz, AreH), 7.38 (1H, s, AreH). 13C NMR (DMSO-d6)
d:
166.24, 164.86 (d, JCeF ¼ 250 Hz), 139.69, 137.81, 135.79, 130.26,
130.16, 125.95, 124.96, 122.12, 118.79, 117.32, 117.10. ESI-HRMS m/z:
calcd for C13H11BrFN2O3S [M þ H]þ: 372.9658; found 372.9655,
374.9635.
d6)
d
: 166.90, 159.58 (d, JCeF ¼ 256 Hz), 155.51 (d, JCeF ¼ 227 Hz),
150.13, 132.69, 132.47, 131.82, 128.94, 128.42, 125.21, 125.07, 115.00,
112.88, 112.69, 106.90, 106.65, 106.38, 61.89. ESI-HRMS m/z: calcd
for
422.9646.
C
14H12BrF2N2O4S [M
þ
H]þ: 420.9669; found 420.9667,
6.1.17. 5-Bromo-3-(4-chlorophenylsulfonamino)benzamide (5k)
Yield 95.0%, mp 206.5e208.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.82
(1H, s, SO2NH), 8.06 (1H, s, AreH), 7.78 (2H, d, J ¼ 8.8 Hz, AreH),
6.1.11. 5-Bromo-2-methoxy-3-cyclopropylsulfonaminobenzamide
7.76 (1H, s, CONH2), 7.68 (2H, d, J ¼ 8.8 Hz, AreH), 7.61 (1H, s, Are
(5e)
H), 7.56 (1H, s, CONH2), 7.39 (1H, s, AreH). 13C NMR (DMSO-d6)
d:
Yield 57.5%, mp 175.5e177.0 ꢀC. 1H NMR (DMSO-d6)
d: 9.55 (1H,
166.20, 139.57, 138.69, 138.30, 137.83, 130.16, 129.03, 126.04, 125.01,
122.14, 118.82. ESI-HRMS m/z: calcd for C13H11BrClN2O3S [M þ H]þ:
388.9362; found 388.9360, 390.9340.
s, SO2NH), 7.85 (1H, s, CONH2), 7.69 (1H, s, CONH2), 7.64 (1H, d,
J ¼ 2.4 Hz, AreH), 7.39 (1H, d, J ¼ 2.4 Hz, AreH), 3.77 (3H, s, OCH3),
2.82 (1H, m, CH), 0.99 (4H, m, 2CH2). 13C NMR (DMSO-d6)
d: 166.66,
149.26, 133.44, 132.76, 127.78, 127.15, 115.42, 62.28, 30.96, 5.73
H]þ:
6.1.18. 6-Bromo-4-morpholinoquinazoline (6a)
(2CH2). ESI-HRMS m/z: calcd for C11H14BrN2O4S [M
þ
To
a mixture of 6-bromo-4-hydroxyquinazoline (7.9 g,
348.9858; found 348.9860, 350.9834.
35.1 mmol) in POCl3 (60 ml) was added DMF (1 ml). Then the
mixture was heated to reflux under N2 for 6 h. The dark clear so-
lution was cooled to room temperature and concentrated in vac-
uum to produce a brown solid. To the mixture of above brown solid
in 2-propanol (150 ml) was added morpholine (12.2 ml,
140.5 mmol). Then the mixture was heated to reflux under N2 for
1 h, concentrated in vacuum to give a residue which was dissolved
in ethyl acetate (200 ml). The ethyl acetate solution was washed
with brine (80 ml ꢂ5), dried over Na2SO4 and concentrated to give a
brown oil, which was solidified on keeping to produce 6a (8.9 g).
6.1.12. Methyl 5-bromo-2-methoxy-3-(4-fluorophenylsulfonamino)
benzoate (5f)
Yield 77.8%, mp 147.0e147.5 ꢀC. 1H NMR (CDCl3)
d: 7.93 (1H, d,
J ¼ 2.8 Hz, AreH), 7.89 (2H, dd, J ¼ 4.8, 8.8 Hz, AreH), 7.69 (1H, d,
J ¼ 2.4 Hz, AreH), 7.31 (1H, s, SO2NH), 7.19 (2H, t, J ¼ 8.8 Hz, AreH),
3.91 (3H, s, OCH3), 3.63 (3H, s, OCH3). 13C NMR (DMSO-d6)
d: 164.71,
164.94 (d, JCeF ¼ 251 Hz), 151.18, 136.70, 133.41, 130.22, 130.13,
129.47, 129.09, 127.04, 117.20, 116.99, 115.33, 62.49, 53.13. ESI-HRMS
m/z: calcd for C15H14BrFNO5S [M þ H]þ: 417.9760; found 417.9764,
419.9748.
Yield 85.7%, mp 112.0e114.0 ꢀC. 1H NMR (CDCl3)
d: 8.75 (1H, s, Are
H), 8.00 (1H, d, J ¼ 1.6 Hz, AreH), 7.82 (1H, d, J ¼ 8.8 Hz, AreH), 7.80
(1H, dd, J ¼ 9.2, 2.0 Hz, AreH), 3.90 (4H, t, J ¼ 4.4 Hz, NCH2), 3.79
(4H, t, J ¼ 4.4 Hz, OCH2). ESI-HRMS m/z: calcd for C12H12BrClN3O
[M þ H]þ: 293.0164; found 293.0166, 295.0140.
6.1.13. N-Methyl-5-bromo-2-methoxy-3-(4-
fluorophenylsulfonamino)benzamide (5g)
Yield 83.6%, mp 170.5e171.5 ꢀC. 1H NMR (CDCl3)
d: 7.88 (2H, dd,
J ¼ 5.2, 8.8 Hz, AreH), 7.82 (1H, s, AreH), 7.75 (1H, s, SO2NH), 3.07
6.1.19. 6-Bromo-4-morpholinoquinoline (6b)
(3H, s, NCH3), 7.19 (3H, t, J ¼ 8.4 Hz, AreH), 6.94 (1H, s, NH), 3.51
Yield 88.1%, mp: 72.0e73.0 ꢀC. 1H NMR (CDCl3)
d: 8.76 (1H, d,
(3H, s, OCH3). 13C NMR (DMSO-d6)
d: 165.33, 164.93 (d, JCe
J ¼ 4.8 Hz, AreH), 8.17 (1H, d, J ¼ 1.6 Hz, AreH), 8.00 (1H, d,
J ¼ 8.8 Hz, AreH), 7.76 (1H, dd, J ¼ 1.6, 8.8 Hz, AreH), 6.91 (1H, d,
J ¼ 4.8 Hz, AreH), 4.02 (4H, t, J ¼ 4.4 Hz, OCH2), 3.26 (4H, t,
J ¼ 4.4 Hz, NCH2). ESI-HRMS m/z: calcd for C13H11BrClN2O3S
[M þ H]þ: 388.9362; found 388.9360, 390.9340.
¼ 251 Hz), 149.01, 136.73, 132.78, 132.36, 130.24, 130.14, 128.11,
F
126.61, 117.19, 116.97, 115.20, 61.90, 26.65. ESI-HRMS m/z: calcd for
C
15H15BrFN2O4S [M þ H]þ: 416.9920; found 416.9918, 418.9896.
6.1.14. N,N-Dimethyl-5-bromo-2-methoxy-3-(4-
fluorophenylsulfonamino)benzamide (5h)
6.1.20. 3-(4-Fluorophenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinazolin-6-yl)benzamide (1a)
Yield 89.1%, mp 169.0e170.0 ꢀC. 1H NMR (CDCl3)
d: 7.86 (2H, dd,
J ¼ 4.8, 8.8 Hz, AreH), 7.79 (1H, d, J ¼ 2.4 Hz, AreH), 7.17 (2H, t,
J ¼ 8.8 Hz, AreH), 7.15 (1H, s, SO2NH), 7.13 (1H, d, J ¼ 2.4 Hz, AreH),
3.54 (3H, s, OCH3), 3.09 (3H, s, NCH3), 2.75 (3H, s, NCH3). 13C NMR
The mixture of 6a (1.0 g, 3.4 mmol), bis(pinacolato)diboron
(1.0 g, 4.1 mmol), potassium acetate (5.0 g, 10.2 mmol), PdCl2(dppf)
(126 mg, 0.16 mmol) and 1,2-dimethoxyethane (25 ml) was reflux
for 2 h under nitrogen atmosphere, cooled to room temperature. To
the resulted mixture were added 5a (1.4 g, 3.4 mmol), Na2CO3 (1.1 g,
10.4 mmol), PdCl2(dppf) (126 mg, 0.16 mmol) and water (12 ml).
The reaction mixture was refluxed under nitrogen atmosphere for
2 h. The volatiles were removed in vacuum and the residue was
purified through a column chromatography on silica with chloro-
form/methanol (V:V ¼ 30:1) as eluent to produce 1a (1.49 g, 81.4%).
(DMSO-d6)
d
: 166.17, 165.08 (d, JCeF ¼ 246 Hz), 148.17, 136.53,
132.60, 131.77, 130.28, 130.18, 127.51, 127.31, 117.03, 116.81, 115.39,
61.61, 38.10, 34.68. ESI-HRMS m/z: calcd for C16H17BrFN2O4S
[M þ H]þ: 431.0076; found 431.0074, 433.0053.
6.1.15. 5-Bromo-2-chloro-3-(4-fluorophenylsulfonamino)
benzamide (5i)
Yield 75.1%, mp 228.0e229.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.47
mp 168.5e170.5 ꢀC. 1H NMR (DMSO-d6)
d: 10.13 (1H, s, SO2NH), 8.67
(1H, s, SO2NH), 7.96 (1H, s, CONH2), 7.82 (2H, dd, J ¼ 5.2, 8.8 Hz, Are
(1H, s, AreH), 8.04 (1H, d, J ¼ 8.8 Hz, AreH), 8.03 (1H, s, AreH), 7.92
(1H, d, J ¼ 8.8 Hz, AreH), 7.91 (2H, dd, J ¼ 8.8, 5.2 Hz, AreH), 7.85
(1H, s, CONH2), 7.74 (1H, d, J ¼ 2.0 Hz, AreH), 7.63 (1H, s, CONH2),
7.60 (1H, d, J ¼ 2.0 Hz, AreH), 7.45 (2H, t, J ¼ 8.8 Hz, AreH), 3.80 (8H,
H), 7.70 (1H, s, CONH2), 7.47 (1H, s, AreH), 7.45 (2H, t, J ¼ 8.8 Hz,
AreH), 7.41 (1H, d, J ¼ 2.0 Hz, AreH). 13C NMR (DMSO-d6)
d: 166.68,
165.00 (d, JCeF ¼ 251 Hz), 140.59, 136.90, 135.88, 130.29, 130.19,
129.86, 128.68, 125.41, 119.74, 117.21, 116.98. ESI-HRMS m/z: calcd
for C13H10BrClFN2O3S [M þ H]þ: 406.9268; found 406.9264,
408.9244.
s, OCH2, NCH2), 3.49 (3H, s, OCH3). 13C NMR (DMSO-d6)
d: 167.96,
164.83 (d, JCeF ¼ 250 Hz), 164.25, 154.24, 151.22, 149.99, 137.20,
136.31, 134.65, 131.83, 131.27, 130.23, 130.13, 129.41, 124.71, 123.55,

T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
103
122.83, 117.10, 116.88, 116.42, 66.45 (2CH2), 61.91, 50.23 (2CH2). ESI-
HRMS m/z: calcd for C26H24FN5O5S [M þ H]þ: 538.1560; found
538.1557.
Compounds 1bek, 2a and 2b were synthesized according to the
procedure described in 1a.
J ¼ 8.6, 2.4 Hz, AreH), 7.90 (1H, s, CONH), 7.75 (1H, d, J ¼ 1.6 Hz, Are
H), 7.56 (1H, d, J ¼ 2.4 Hz, AreH), 7.49 (2H, t, J ¼ 8.8 Hz, AreH), 3.80
(8H, s, OCH2, NCH2), 3.45 (3H, s, OCH3), 2.77 (3H, d, J ¼ 4.8 Hz,
NCH3). 13C NMR (DMSO-d6)
d
: 166.64, 164.82 (d, JCeF ¼ 250 Hz),
164.23, 154.24, 151.22, 149.84, 137.20, 136.26, 134.67, 131.83, 131.40,
131.20, 130.22, 130.12, 129.39, 124.60, 123.35, 122.81, 117.11, 116.89,
116.40, 66.45 (2CH2), 61.83, 50.23 (2CH2), 26.59. ESI-HRMS m/z:
calcd for C27H26FN5O5S [M þ H]þ: 552.1717; found 552.1725.
6.1.21. 3-(4-Chlorophenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinazolin-6-yl)benzamide (1b)
Yield 68.5%, mp 134.0e137.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.19 (1H,
s, SO2NH), 8.67 (1H, s, AreH), 8.04 (2H, m, AreH), 7.92 (1H, d,
J ¼ 8.8 Hz, AreH), 7.85 (3H, m, AreH), 7.73 (1H, d, J ¼ 2.4 Hz, AreH),
7.70 (1H, s, CONH2), 7.68 (1H, s, AreH), 7.63 (1H, s, CONH2), 7.61
(1H, d, J ¼ 2.4 Hz, AreH), 3.81 (8H, s, OCH2, NCH2), 3.51 (3H, s,
6.1.26. N,N-Dimethyl-3-(4-fluorophenylsulfonamino)-2-methoxy-
5-(4-morpholinoquinazolin-6-yl)benzamide (1g)
Yield 39.7%, mp 103.0e103.5 ꢀC. 1H NMR (DMSO-d6)
d: 10.07
(1H, s, SO2NH), 8.67 (1H, s, AreH), 8.06 (1H, m, AreH), 8.04 (1H, s,
AreH), 7.90 (1H, d, J ¼ 5.6 Hz, AreH), 7.85 (2H, dd, J ¼ 8.8, 5.2 Hz,
AreH), 7.73 (1H, d, J ¼ 2.0 Hz, AreH), 7.43 (2H, t, J ¼ 8.0 Hz, AreH),
7.41 (1H, d, J ¼ 9.2 Hz, AreH), 3.81 (8H, s, OCH2, NCH2), 3.43 (3H, s,
OCH3). 13C NMR (DMSO-d6)
d: 167.96, 164.24, 154.24, 151.23, 150.09,
139.70, 138.33, 136.28, 134.64, 131.84, 131.29, 131.15, 129.95, 129.40,
129.04, 124.82, 123.68, 122.84, 116.41, 66.45 (2CH2), 61.90, 50.23
(2CH2). ESI-HRMS m/z: calcd for C26H24ClN5O5S [M
554.1265; found 554.1260.
þ
H]þ:
OCH3), 2.99 (3H, s, NCH3), 2.67 (3H, s, NCH3). 13C NMR (CDCl3)
d:
168.10, 165.38, 164.68 (d, JCeF ¼ 251 Hz), 154.23, 151.30, 145.91,
136.53, 136.39, 135.17, 131.54, 130.16, 129.86, 129.77 (2CH), 129.46,
123.16, 122.63, 120.56, 116.63 (2CH), 116.41, 66.71 (2CH2), 61.80,
50.37 (2CH2), 38.23, 35.06. ESI-HRMS m/z: calcd for C28H28FN5O5S
[M þ H]þ: 566.1873; found 566.1879.
6.1.22. 3-(4-Methylphenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinazolin-6-yl)benzamide (1c)
Yield 72.1%, mp 146.0e147.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.00 (1H,
s, SO2NH), 8.67 (1H, s, AreH), 8.02 (1H, s, AreH), 8.01 (1H, d,
J ¼ 8.0 Hz, AreH), 7.92 (1H, d, J ¼ 9.2 Hz, AreH), 7.85 (1H, s, CONH2),
7.75 (2H, d, J ¼ 8.4 Hz, AreH), 7.74 (1H, d, J ¼ 2.4 Hz, AreH), 7.61
(1H, s, CONH2), 7.57 (1H, d, J ¼ 2.4 Hz, AreH), 7.45 (2H, d, J ¼ 8.0 Hz,
AreH), 3.80 (8H, s, OCH2, NCH2), 3.51 (3H, s, OCH3), 2.35 (3H, s, Are
6.1.27. Methyl 3-(4-fluorophenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinazolin-6-yl)benzoate (1h)
Yield 47.7%, mp > 250.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.20 (1H, s,
SO2NH), 8.68 (1H, s, AreH), 8.04 (2H, s, AreH), 7.89e7.93 (4H, m,
AreH), 7.82 (1H, s, AreH), 7.45 (2H, t, J ¼ 8.2 Hz, AreH), 3.86 (3H, s,
OCH3), 3.81 (8H, s, OCH2, NCH2), 3.46 (3H, s, OCH3). 13C NMR
CH3). 13C NMR (DMSO-d6)
d: 167.95, 164.25, 154.25, 151.23, 149.58,
143.91, 137.91, 136.39, 134.60, 131.81, 131.75, 131.27, 130.21, 129.43,
127.16, 124.15, 122.74, 122.64, 116.43, 66.47 (2CH2), 61.94, 50.23
(2CH2), 21.43. ESI-HRMS m/z: calcd for C27H27N5O5S [M þ H]þ:
534.1811; found 534.1809.
(DMSO-d6)
d
: 166.04, 164.83 (d, JCeF ¼ 251 Hz), 164.20, 154.30,
151.73, 151.25, 137.12, 135.82, 134.97, 132.23, 131.79, 130.20, 130.10,
129.38, 125.94, 125.80, 125.69, 123.01, 117.11, 116.88, 116.34, 66.44
(2CH2), 62.41, 53.00, 50.20 (2CH2). ESI-HRMS m/z: calcd for
6.1.23. 3-(2,4-Difluorophenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinazolin-6-yl)benzamide (1d)
C
27H25FN4O6S [M þ H]þ : 553.1557; found 553.1552.
Yield 61.5%, mp 112.0e116.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.44 (1H,
6.1.28. 2-Chloro-3-(4-fluorophenylsulfonamino)-5-(4-
morpholinoquinazolin-6-yl)benzamide (1i)
b, SO2NH) 8.73 (1H, d, J ¼ 5.2 Hz, AreH), 8.14 (1H, d, J ¼ 2.0 Hz, Are
H), 8.05 (1H, d, J ¼ 8.8 Hz, AreH), 7.96 (1H, m, AreH), 7.89 (1H, s,
CONH), 7.85 (1H, m, AreH), 7.76 (1H, d, J ¼ 2.4 Hz, AreH), 7.66 (1H,
d, J ¼ 2.4 Hz, AreH), 7.62 (1H, s, CONH), 7.57 (1H, m, AreH), 7.28
(1H, m, AreH), 7.07 (1H, d, J ¼ 4.8 Hz, AreH), 3.91 (4H, t, J ¼ 4.4 Hz,
NCH2), 3.58 (3H, s, OCH3), 3.23 (4H, d, J ¼ 3.6 Hz, OCH2). 13C NMR
Yield 62.7%, mp 223.0e225.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.43
(1H, s, SO2NH), 8.67 (1H, s, AreH), 8.08 (1H, s, AreH), 8.06 (1H, d,
J ¼ 8.8 Hz, AreH), 7.97 (1H, s, CONH2), 7.92 (1H, d, J ¼ 8.8 Hz, AreH),
7.85 (2H, dd, J ¼ 8.4, 5.2 Hz, AreH), 7.68 (3H, d, J ¼ 2.8 Hz, 2AreH,
CONH2), 7.45 (2H, t, J ¼ 8.8 Hz, AreH), 3.81 (8H, d, J ¼ 4.8 Hz, OCH2,
(DMSO-d6)
d
: 167.97, 165.44 (d, JCeF ¼ 250 Hz), 164.24, 164.18,
NCH2). 13C NMR (DMSO-d6)
d
: 167.97, 164.90 (d, JCeF ¼ 251 Hz),
159.42 (d, JCeF ¼ 249 Hz), 154.24, 151.22, 150.96, 136.23, 134.56,
132.42, 131.86, 131.20, 130.77, 129.40, 125.41, 125.13, 122.84, 121.50,
116.42, 112.70, 106.64, 66.44 (2CH2), 61.81, 50.22 (2CH2). ESI-HRMS
m/z: calcd for C27H24F2N4O5S [M þ H]þ: 555.1514; found 555.1519.
164.16, 154.42, 151.48, 139.45, 138.52, 137.41, 135.39, 135.20, 131.65,
130.28, 130.18, 129.41, 126.17, 125.76, 124.77, 123.43, 117.11, 116.38,
116.32, 66.43 (2CH2), 50.17 (2CH2). ESI-HRMS m/z: calcd for
C
25H21ClFN5O4S [M þ H]þ: 542.1065; found 542.1071.
6.1.24. 3-Cyclopropylsulfonamino-2-methoxy-5-(4-
6.1.29. 3-(4-Fluorophenylsulfonamino)-5-(4-morpholinoquinazolin-
morpholinoquinazolin-6-yl)benzamide (1e)
6-yl)benzamide (1j)
Yield 72.2%, mp 189.0e192.0 ꢀC. 1H NMR (DMSO-d6)
d
: 9.44 (1H,
Yield47.2%, mp142.0e144.5 ꢀC.1HNMR (DMSO-d6)
d: 10.68(1H, s,
s, SO2NH), 8.67 (1H, s, AreH), 8.12 (1H, d, J ¼ 9.2 Hz, AreH), 8.09
(1H, s, AreH), 7.93 (1H, d, J ¼ 8.8 Hz, AreH), 7.88 (2H, s, AreH,
CONH2), 7.67 (1H, s, AreH), 7.66 (1H, s, CONH2), 3.86 (3H, s, OCH3),
3.80 (8H, s, OCH2, NCH2), 1.99 (1H, s, CH), 1.01 (4H, m, 2CH2). 13C
SO2NH), 8.68 (1H, s, AreH), 8.14 (1H, s, AreH), 8.10 (1H, s, AreH), 8.08
(1H, d, J ¼ 2.4 Hz, AreH), 7.97 (1H, s, CONH2), 7.94 (1H, d, J ¼ 9.2 Hz,
AreH), 7.88 (2H, dd, J ¼ 8.8, 5.2 Hz, AreH), 7.65 (1H, s, CONH2), 7.63
(1H, s, AreH), 7.52 (1H, s, AreH), 7.43 (2H, t, J ¼ 8.8 Hz, AreH), 3.81
NMR (DMSO-d6)
d: 167.98, 164.28, 154.23, 151.21, 150.12, 136.52,
(8H, s, OCH2, NCH2). 13C NMR (DMSO-d6)
d: 167.53, 164.86 (d, JCe
134.85, 132.26, 131.97, 131.31, 129.37, 124.28, 123.86, 122.91, 116.42,
66.46 (2CH2), 62.26, 50.25 (2CH2), 30.95, 5.75 (2CH2). ESI-HRMS m/
z: calcd for C23H25N5O5S [M þ H]þ: 484.1655; found 484.1652.
¼ 251 Hz), 164.24, 154.37, 151.50, 140.54, 139.03, 136.80, 136.40,
F
136.18, 132.05, 130.25, 130.15, 129.39, 123.23, 122.03, 121.37, 119.35,
117.24,117.02,116.35, 66.54(2CH2), 50.21(2CH2). ESI-HRMS m/z: calcd
for C25H22FN5O4S [M þ H]þ: 508.1455; found 508.1452.
6.1.25. N-Methyl-3-(4-fluorophenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinazolin-6-yl)benzamide (1f)
6.1.30. 3-(4-Chlorophenylsulfonamino)-5-(4-morpholinoquinazolin-
Yield 43.5%, mp 199.5e201.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.12
6-yl)benzamide (1k)
(1H, s, SO2NH), 8.67 (1H, s, AreH), 8.34 (1H, d, J ¼ 4.4 Hz, AreH),
Yield 43.1%, mp 146.0e148.5 ꢀC. 1H NMR (DMSO-d6)
d: 10.75
8.04 (1H, d, J ¼ 6.4 Hz, AreH), 8.03 (1H, s, AreH), 7.92 (2H, dd,
(1H, s, SO2NH), 8.68 (1H, s, AreH), 8.16 (1H, s, AreH), 8.10 (1H, m,

104
T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
AreH), 8.08 (1H, s, AreH), 7.98 (1H, s, CONH2), 7.94 (1H, d,
J ¼ 9.2 Hz, AreH), 7.81 (2H, d, J ¼ 8.8 Hz, AreH), 7.67 (2H, d,
J ¼ 8.8 Hz, AreH), 7.66 (1H, s, CONH2), 7.63 (1H, s, AreH), 7.54 (1H, s,
viability. The IC50 value was calculated according to the inhibition
ratios.
AreH), 3.81 (8H, s, OCH2, NCH2). 13C NMR (DMSO-d6)
d
: 167.50,
6.2.3. PI3K and mTOR enzymatic activity assay
164.23, 154.37, 151.50, 140.57, 138.89, 138.65, 136.82, 136.38, 132.04,
132.07, 129.38, 129.11, 129.05, 123.24, 122.14, 121.42, 119.39, 116.35,
66.45 (2CH2), 50.21 (2CH2). ESI-HRMS m/z: calcd for C25H22ClN5O4S
[M þ H]þ: calcd 524.1159; found 524.1151.
PI3K and mTOR enzymatic activity assay was performed ac-
cording to process described in supporting information of Ref. [24].
Briefly, different enzyme phosphorylate different substrates, the
concentration of kinases and corresponding substrates was
different. The enzyme reaction buffer contained 40 mM TriseHCl,
6.1.31. 3-(4-Fluorophenylsulfonamino)-2-methoxy-5-(4-
pH 7.4, 10 mM MgCl2, 0.1 mg/ml BSA, 1 mM DTT, 10 mM ATP, enzyme
morpholinoquinolin-6-yl)benzamide (2a)
and substrate. Various concentration of compounds was added to
Yield 69.8%, mp 141.0e142.0 ꢀC. 1H NMR (DMSO-d6)
d
: 10.17 (1H,
this enzyme reaction buffer, and the final reaction volume was
s, SO2NH), 8.73 (1H, d, J ¼ 5.2 Hz, AreH), 8.15 (1H, s, AreH), 8.05
(1H, d, J ¼ 8.8 Hz, AreH), 7.96 (1H, s, CONH2), 7.91 (2H, dd, J ¼ 8.6,
5.6 Hz, AreH), 7.89 (1H, s, AreH), 7.81 (1H, s, AreH), 7.64 (1H, s,
CONH2), 7.62 (1H, d, J ¼ 1.6 Hz, AreH), 7.45 (2H, t, J ¼ 8.8 Hz, AreH),
7.07(1H, d, J ¼ 4.8 Hz, AreH), 3.92 (4H, s, OCH2), 3.50 (3H, s, OCH3),
50
then stopped by the addition of 25
m
l. The reaction mixture was incubated at 30 ꢀC for 40 min, and
mL of KinaseGlo solution. The
stopped reaction was incubated for 5 min and the remaining ATP
was then detected via luminescence.
3.23 (4H, s, NCH2). 13C NMR (DMSO-d6)
d: 168.00, 164.80 (d, JCe
6.2.4. Western blot assay
¼ 250 Hz), 156.63, 151.63, 149.79, 148.95, 137.29, 135.95, 135.01,
The suppressive activity of AKTand p-AKT473 in HCT-116 cells was
determined by western blot. HCT-116 cells were seeded into six-well
plates at 1 ꢂ 106 cells per well respectively and cultured in DMEM
10% FBS, incubated at 37 ꢀC for 16 h prior to drug exposure. Cells
F
131.51, 131.19, 131.00, 130.14, 130.04, 128.14, 124.38, 123.24, 123.21,
121.25, 117.11, 116.88, 110.05, 66.56 (2CH2), 61.91, 52.77 (2CH2). ESI-
HRMS m/z: calcd for C27H25FN4O5S [M þ H]þ: 537.1608; found
537.1616.
were treated with final concentrations of 10 mM of compounds 1a,
BEZ235 and DMSO and incubated at 37 ꢀC for 1 h. Cells were washed
twice with ice-cold PBS and scraped into ice-cold cell lysis buffer.
Then the cell lysates were clarified by centrifugation at 12,000 rpm
for 20 min at 4 ꢀC and the supernatant was collected. Protein content
was measured by BCA protein Assay Kit and the proteins were
separated on SDS-PAGE and then transferred onto nitrocellulose
membranes. The membranes were incubated with antibodies
against AKT (Cell Signaling Technology), p-Ser473 AKT (Cell
6.1.32. 3-(2,4-Difluorophenylsulfonamino)-2-methoxy-5-(4-
morpholinoquinolin-6-yl) benzamide (2b)
Yield 52.8%, mp 111.0e116.0 ꢀC. 1H NMR (DMSO-d6)
d: 10.44 (1H,
b, SO2NH), 8.73 (1H, d, J ¼ 5.2 Hz, AreH), 8.14 (1H, d, J ¼ 2.0 Hz, Are
H), 8.05 (1H, d, J ¼ 8.8 Hz, AreH), 7.96 (1H, m, AreH), 7.89 (1H, s,
CONH), 7.85 (1H, m, AreH), 7.76 (1H, d, J ¼ 2.4 Hz, AreH), 7.66 (1H,
d, J ¼ 2.4 Hz, AreH), 7.62 (1H, s, CONH), 7.57 (1H, m, AreH), 7.28
(1H, m, AreH), 7.07 (1H, d, J ¼ 4.8 Hz, AreH), 3.91 (4H, t, J ¼ 4.4 Hz,
OCH2), 3.58 (3H, s, OCH3), 3.23 (4H, d, J ¼ 3.6 Hz, NCH2). 13C NMR
Signaling Technology) and b-Acin (Santa Cruz), washed by TBST and
then incubated with Mouse and rabbit horseradish peroxidase-
conjugated secondary antibodies. The proteineantibody complexes
were detected by chemiluminescence with a GeneGnome5 system
(Syngene, UK). Protein bands were quantified using ImageJ software
(National Institutes of Health, Bethesda, MD, USA).
(DMSO-d6) d: 168.02, 165.43, 160.66, 158.17, 156.67, 151.56, 150.72,
148.86, 135.88, 134.94, 132.30, 131.85, 131.14, 130.92, 128.19, 125.10,
124.78, 124.0, 123.21, 121.29, 112.70, 110.02, 106.70, 66.54 (2CH2),
61.82, 52.76 (2CH2). ESI-HRMS m/z: calcd for C27H24F2N4O5S
[M þ H]þ: 555.1514; found 555.1519.
6.2.5. Anticancer effects in established nude mouse U-87 MG
xenograft model in vivo
6.2. Biological assay methods
Mice (BALB/C, SPF grade, male, 18e20 g) were purchased from
Shanghai Slakey Laboratory Animal Co., LTD and fed at Shanghai
Institute of Pharmaceutical Industry. The experimental protocol
was approved by Ethic Committee of Shanghai Institute of Phar-
maceutical Industry.
6.2.1. Cell culture
The human cell lines A549, HCT-116, U-87 MG or KB were
maintained as a monolayer culture in DMEM, supplemented with
10% FBS in a humidified atmosphere (5% CO2) at 37 ꢀC.
U-87 MG cells at 3 ꢂ 106 were injected subcutaneously into the
flank. Once the tumor xenografts reached 100 mm3, all tumor-
bearing nude mice were randomly divided into four groups, with
8 mice in control group and 5 mice in each drug group. In the
control group, the same volume of solvent was administered orally.
BEZ235 and 1a were dissolved in NMP/PEG300 (1: 9). BEZ235 was
dosed orally at 20 mg/kg to positive group. 1a was dosed orally at
20 mg/kg and 50 mg/kg for the low and high dosage groups once a
day for 12 days, respectively. Tumor volumes and body weights
were recorded at intervals of 3 or 4 days. Tumor volume was
calculated as length ꢂ width ꢂ width/2 and is reported in mm3.
Results are expressed as the mean ꢁ standard error. The mice were
anesthetized and sacrificed on day 21. The weights of the body, the
neoplasm, and the volumes of neoplasm, were measured and
inhibitory ratios for tumor volume and weight was calculated.
6.2.2. Antiproliferative activity
3-[4,5-Dimethyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bro-
mide (MTT) was purchased from Sigma (St. Louis, MO, USA).
Cellular chemosensitivity was determined by using a modified MTT
method assay in vitro. Cell numbers were titrated to keep cells
growing in the exponential phase throughout the 48 h incubation
period. In brief, HCT-116, U-87 MG, A549 or KB cells in 100 ml cul-
ture medium were seeded into 96-well microplates at 3000e5000
cells per well respectively and cultured in DMEM 10% FBS, incu-
bated at 37 ꢀC for 24 h prior to drug exposure. Compounds were
prepared in DMSO and then diluted with medium to the desired
concentrations. Cells were treated with final concentrations of
100.0, 10.0, 1.0, 0.1 and 0.01
neously and incubated for 72 h and then 20
(5 mg/ml in medium) was added to each well and incubated for 4 h.
The formed blue formazan crystals were dissolved in 200 l of
m
M of tested compounds simulta-
ml of MTT solution
m
6.3. Molecular modeling
DMSO. The optical density at 490 nm was determined by Varioskan
Flash Multimode Reader (Thermo scientific). Three separate ex-
periments with triplicate data were performed to obtain mean cell
The proteineligand complex crystal structure of GSK2126458
bound to PI3Kg was chosen as the template to compare the docking

T. Shao et al. / European Journal of Medicinal Chemistry 75 (2014) 96e105
105
[9] A.L. Smith, N.D. D’Angelo, Y.Y. Bo, S.K. Booker, V.J. Cee, B. Herberich, F. Hong,
C.L.M. Jackson, B.A. Lanman, L. Liu, N. Nishimura, L.H. Pettus, A.B. Reed,
S. Tadesse, N.A. Tamayo, R.P. Wurz, K. Yang, K.L. Andrews, D.A. Whittington,
J.D. McCarter, T.S. Miguel, L. Zalameda, J. Jiang, R. Subramanian, E.L. Mullady,
S. Caenepeel, D.J. Freeman, L. Wang, N. Zhang, T. Wu, P.E. Hughes,
M.H. Norman, Journal of Medicinal Chemistry 55 (2012) 5188e5219.
[10] H. Cheng, J.E. Hoffman, P.T. Le, M. Pairish, R. Kania, W. Farrell, S. Bagrodia,
J. Yuan, S. Sun, E. Zhang, C. Xiang, D. Dalvie, S.V. Rahavendran, Bioorganic &
Medicinal Chemistry Letters 23 (2013) 2787e2792.
[11] G.A. Morales, J.R. Garlich, J. Su, X. Peng, J. Newblom, K. Weber, D.L. Durden,
Journal of Medicinal Chemistry 56 (2013) 1922e1939.
[12] P. Wu, Y.Z. Hu, Medicinal Chemistry Communications 3 (2012) 1337e1355.
[13] S.D. Knight, N.D. Adams, J.L. Burgess, ACS Medicinal Chemistry Letters 1
(2010) 39e43.
[14] N. Nishimura, A. Siegmund, L. Liu, K. Yang, M.C. Bryan, K.L. Andrews, Y. Bo,
S.K. Booker, S. Caenepeel, D. Freeman, H. Liao, J. McCarter, E.L. Mullady,
T.S. Miguel, R. Subramanian, N. Tamayo, L. Wang, D.A. Whittington,
L. Zalameda, N. Zhang, P.E. Hughes, M.H. Norman, Journal of Medicinal
Chemistry 54 (2011) 4735e4751.
[15] N.D. D’Angelo, T. Kim, K. Andrews, S.K. Booker, S. Caenepeel, K. Chen,
D. D’Amico, D. Freeman, J. Jiang, L. Liu, J.D. McCarter, T.S. Miguel, E.L. Mullady,
M. Schrag, R. Subramanian, J. Tang, R.C. Wahl, L. Wang, D.A. Whittington,
T. Wu, N. Xi, Y. Xu, P. Yakowec, K. Yang, L.P. Zalameda, N. Zhang, P. Hughes,
M.H. Norman, Journal of Medicinal Chemistry 54 (2011) 1789e1811.
[16] M.M. Stec, K.L. Andrews, S.K. Booker, S. Caenepeel, D.J. Freeman, J. Jiang,
H. Liao, J. McCarter, E.L. Mullady, T. San Miguel, R. Subramanian, N. Tamayo,
L. Wang, K. Yang, L.P. Zalameda, N. Zhang, P.E. Hughes, M.H. Norman, Journal
of Medicinal Chemistry 54 (2011) 5174e5184.
mode between compound 1a bound to PI3K
PI3K . The molecular docking procedure was performed by using
C-DOCKER protocol within Discovery Studio 2.5. For enzyme
preparation, the hydrogen atoms were added. The whole PI3K
enzyme was defined as a receptor and the site sphere was selected
on the basis of the ligand binding location of GSK2126458.
GSK2126458 and water molecule were removed and compound 1a
was placed. After end of molecular docking, ten docking poses were
scored and selected based on calculated C-DOCKER energy.
g and 1a bound to
g
g
Acknowledgments
Financial support from National Natural Science Foundation of
China (Grant No. 21072156 and 81272448) is gratefully
acknowledged.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.01.053.
[17] M.H. Norman, K.L. Andrews, Y.Y. Bo, S.K. Booker, S. Caenepeel, V.J. Cee,
N.D. D’Angelo, D.J. Freeman, B.J. Herberich, F. Hong, C.L.M. Jackson, J. Jiang,
B.A. Lanman, L. Liu, J.D. McCarter, E.L. Mullady, N. Nishimura, L.H. Pettus,
A.B. Reed, T.S. Miguel, A.L. Smith, M.M. Stec, S. Tadesse, A. Tasker, D. Aidasani,
X. Zhu, R. Subramanian, N.A. Tamayo, L. Wang, D.A. Whittington, B. Wu, T. Wu,
R.P. Wurz, K. Yang, L. Zalameda, N. Zhang, P.E. Hughes, Journal of Medicinal
Chemistry 55 (2012) 7796e7816.
[18] X.M. Wang, J. Xu, Y.P. Li, H. Li, C.S. Jiang, G.D. Yang, S.M. Lu, S.Q. Zhang, Eu-
ropean Journal of Medicinal Chemistry 67 (2013) 243e251.
[19] W. Yang, M. Shu, Y. Wang, R. Wang, Y. Hu, L. Meng, Z. Lin, Journal of Molecular
Structure 1054e1055 (2013) 107e116.
[20] S. Maira, F. Stauffer, J. Brueggen, P. Furet, C. Schnell, C. Fritsch, S. Brachmann,
P. Chene, A.D. Pover, K. Schoemaker, D. Fabbro, D. Gabriel, M. Simonen,
L. Murphy, P. Finan, W. Sellers, C. Garcia-Echeverria, Molecular Cancer Ther-
apeutics 7 (2008) 1851e1863.
[21] H. Cheng, S. Bagrodia, S. Bailey, M. Edwards, J. Hoffman, Q. Hu, R. Kania,
D.R. Knighton, M.A. Marx, S. Ninkovic, S. Sun, E. Zhang, Medicinal Chemistry
Communications 1 (2010) 139e144.
[22] J. Yuan, P.P. Mehta, M. Yin, S. Sun, A. Zou, J. Chen, K. Rafidi, Z. Feng, J. Nickel,
J. Engebetsen, J. Hallin, A. Blasina, E. Zheng, L. Nguyen, M. Sun, P.K. Vogt,
A. McHarg, H. Cheng, J.G. Christensen, J.L.C. Kan, S. Bagrodia, Molecular Cancer
Therapeutics 10 (2011) 2189e2199.
[23] H. Cheng, C. Li, S. Bailey, S.M. Baxi, L. Goulet, L. Guo, J. Hoffman, Y. Jiang,
T.O. Johnson, T.W. Johnson, D.R. Knighton, J. Li, K.K.-C. Liu, Z. Liu,
A.M. Matthew, M. Walls, A.W. Peter, M. Yin, J. Zhu, M. Zientek, ACS Medicinal
Chemistry Letters 4 (2013) 91e97.
References
[1] L. Salmena, A. Carracedo, P.P. Pandolfi, Cell 133 (2008) 403e414.
[2] P. Liu, H. Cheng, T.M. Roberts, J. Zhao, Nature Reviews Drug Discovery 8 (2009)
627e644.
[3] Q. Liu, C. Thoreen, J. Wang, D. Sabatini, N.S. Gray, Drug Discovery Today:
Therapeutic Strategies 6 (2009) 47e55.
[4] M. Dehnhardt, A.M. Venkatesan, E.D. Santos, Z. Chen, O. Santos, S. Ayral-
Kaloustian, N. Brooijmans, R. Mallon, I. Hollander, L. Feldberg, J. Lucas,
I. Chaudhary, K. Yu, J. Gibbons, R. Abraham, T.S. Mansour, Journal of Medicinal
Chemistry 53 (2010) 798e810.
[5] M. Venkatesan, C.M. Dehnhardt, E.D. Santos, Z. Chen, O.D. Santos, S. Ayral-
Kaloustian, G. Khafizova, N. Brooijmans, R. Mallon, I. Hollander, L. Feldberg,
J. Lucas, K. Yu, J. Gibbons, R.T. Abraham, I. Chaudhary, T.S. Mansour, Journal of
Medicinal Chemistry 53 (2010) 2636e2645.
[6] M. Venkatesan, Z. Chen, O.D. Santos, C. Dehnhardt, E.D. Santos, S. Ayral-
Kaloustian, R. Mallon, I. Hollander, L. Feldberg, J. Lucas, K. Yu, I. Chaudhary,
T.S. Mansour, Bioorganic & Medicinal Chemistry Letters 20 (2010) 5869e5873.
[7] J. Folkes, K. Ahmadi, W.K. Alderton, S. Alix, S.J. Baker, G. Box, I.S. Chuckowree,
P.A. Clarke, P. Depledge, S.A. Eccles, L.S. Friedman, A. Hayes, T.C. Hancox,
A. Kugendradas, L. Lensun, P. Moore, A.G. Olivero, J. Pang, S. Patel, G.H. Pergl-
Wilson, F.I. Raynaud, A. Robson, N. Saghir, L. Salphati, S. Sohal, M.H. Ultsch,
M. Valenti, H.J.A. Wallweber, N.C. Wan, C. Wiesmann, P. Workman,
Al. Zhyvoloup, M.J. Zvelebil, S.J. Shuttleworth, Journal of Medicinal Chemistry
51 (2008) 5522e5532.
[8] D.P. Sutherlin, L. Bao, M. Berry, G. Castanedo, I. Chuckowree, J. Dotson, A. Folks,
L. Friedman, R. Goldsmith, J. Gunzner, T. Heffron, J. Lesnick, C. Lewis,
S. Mathieu, J. Murray, J. Nonomiya, J. Pang, N. Pegg, W.W. Prior, L. Rouge,
L. Salphati, D. Sampath, Q. Tian, V. Tsui, N.C. Wan, S. Wang, B. Wei,
C. Wiesmann, P. Wu, B. Zhu, A. Olivero, Journal of Medicinal Chemistry 54
(2011) 7579e7587.
[24] M.T. Burger, S. Pecchi, A. Wagman, Z. Ni, M. Knapp, T. Hendrickson, G. Atallah,
K. Pfister, Y. Zhang, S. Bartulis, K. Frazier, S. Ng, A. Smith, J. Verhagen,
J. Haznedar, K. Huh, E. Iwanowicz, X. Xin, D. Menezes, H. Merritt, I. Lee,
M. Wiesmann, S. Kaufman, K. Crawford, M. Chin, D. Bussiere, K. Shoemaker,
I. Zaror, S. Maira, C.F. Voliva, ACS Medicinal Chemistry Letters 2 (2011) 774e779.