Synthesis and evaluation of quinazolinone derivatives as inhibitors of NF-κB, AP-1 mediated transcription and eIF-4E mediated translational activation: Inhibitors of multi-pathways involve in cancer

Article abstract of DOI:10.1016/j.ejmech.2010.04.038

In our effort to discover and develop small molecule multi-pathway inhibitors which may be useful as tools for treating cancerous conditions, we have synthesized a small library of 2-thiazole-5-yl-3H-quinazolin-4-one derivatives. Synthesized compounds were evaluated as inhibitors of NF-κB and AP-1 mediated transcriptional and eIF-4E mediated translational activation as these transcription and translation factors are known to play a pivotal role in initiation and progression of cancer. The results from the study suggest the utility of the 2-thiazole-5-yl-3H-quinazolin-4-one scaffold as a promising scaffold for the design of novel multi-pathway inhibitors, which can be explored as anti-cancer agents. This study describes the synthesis and evaluation of 2-thiazole-5-yl-3H-quinazolin-4-one derivatives as inhibitors of multiple pathways involved in cancerous conditions.

Full text of DOI:10.1016/j.ejmech.2010.04.038

European Journal of Medicinal Chemistry 45 (2010) 3558e3563
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and evaluation of quinazolinone derivatives as inhibitors of NF-kB, AP-1
mediated transcription and eIF-4E mediated translational activation: Inhibitors of
multi-pathways involve in cancer
,
,
Rajan S. Giri a 1, Hardik M. Thaker a, Tony Giordano b, Bing Chen b, Sam Nuthalapaty b, Kamala K. Vasu a
,
*
Vasudevan Sudarsanam a
a B. V. Patel Pharmaceutical Education and Research Development (PERD) Centre, S.G. Road, Thaltej, Ahmedabad 54, India
b Feist-Weiller Cancer Center, LSUHSCs, 1501 Kings Highway, Shreveport, LA 71130, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
In our effort to discover and develop small molecule multi-pathway inhibitors which may be useful as
tools for treating cancerous conditions, we have synthesized a small library of 2-thiazole-5-yl-3H-qui-
nazolin-4-one derivatives. Synthesized compounds were evaluated as inhibitors of NF-kB and AP-1
mediated transcriptional and eIF-4E mediated translational activation as these transcription and trans-
lation factors are known to play a pivotal role in initiation and progression of cancer. The results from the
study suggest the utility of the 2-thiazole-5-yl-3H-quinazolin-4-one scaffold as a promising scaffold for
the design of novel multi-pathway inhibitors, which can be explored as anti-cancer agents.
Ó 2010 Elsevier Masson SAS. All rights reserved.
Received 26 January 2010
Received in revised form
19 March 2010
Accepted 30 April 2010
Available online 7 May 2010
Keywords:
Transcription factors
NF-kB
AP-1
Translational factor
eIF-4E
Quinazolinone derivatives
Molecular modeling
Thiazole derivatives
1. Introduction
combined action of NF-
kB and AP-1. Furthermore, activated AP-1
and NF- B are found in transformed keratinocytes, pancreatic
k
Cancer is a complex disease that is the result of perturbations in
multiple intracellular regulatory systems [1] leading to an overall
increase in cell number and thus tumor formation. As cancer is
a multifactor process involving the expression of a large number of
proteins, a single molecule that can simultaneously modulate more
than one pathway or target involved in cancer would be a signifi-
cant advantage over the current combination therapy strategies.
cancers, and head and neck squamous cell carcinoma cell lines
[8e10]. Similarly, the translation initiation factors, eIF-4E, eIF-4F,
and eIF-4G, as well as mTOR have been shown to play a significant
role in tumor progression [11e13]. The level of eIF-4E is found to be
elevated in a majority of human breast, head and neck cancers. The
over expression of eIF-4E is also correlated with elevated angio-
genic growth factors and induced by hypoxia [14e16]. These
studies provide a clear physiological role for these transcription
NF-kB and AP-1 are two important transcription factors for many
cell types and are thought to be promising targets for the devel-
opment of anti-cancer therapeutics. It has been well documented
that the NF-kB and AP-1 pathways play an important role in
promoting metastases [2e4], tumor progression [5], angiogenesis
[6] and chemoresistance [7]. A variety of genes involved in cancer
(IL-6, IL-8, MMP-9, COX-2, and MCP-1) are regulated by the
(NF-kB and AP-1) and translation (eIF-4E) factors in cancer. These
studies also suggest that the modulation of these transcription and
translation factors represents a sound strategy for the treatment of
cancer. Previously, we had developed an in vitro screening assay for
the analysis of changes in transcriptional (AP-1 and NF-kB) and
translational (eIF-4E) activity [17,18], which was used in the present
study for screening of the newly synthesized molecules.
Quinazoline scaffolds and their bioisosteres are well known as
a privileged scaffold in drug discovery. Quinazolinones and their
analogs have shown to have utility in a number of therapeutic
* Corresponding author. Tel.: þ91 079 27439375; fax: þ91 07927450449.
E-mail address: kamkva@gmail.com (K.K. Vasu).
Present address: Institute for Tuberculosis Research, UIC, Chicago, IL 60612, USA.
1
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.04.038

R.S. Giri et al. / European Journal of Medicinal Chemistry 45 (2010) 3558e3563
3559
areas, including allergies [19,20], Parkinson’s and Alzheimer’s
diseases, epilepsy and pain [21,22], cytotoxic agents [23], inflam-
mation [24], malaria [25], tuberculosis [26], and cancer [27]. In our
effort to discover and develop inhibitors of NF-kB and AP-1 medi-
ated transcriptional activation as potential anti-inflammatory and
anti-cancer agents, we have reported a series of 2-(2-alkylami-
no,4-substituted-thiazole-5-yl)-3-aryl-3H-quinazolin-4-one deriv-
atives (Fig. 1) [28]. The results of the previous study encouraged us
to synthesize additional analogues of 2-thiazole-5-yl-3H-quinazo-
line-4-one derivatives with more diverse substitutions and to
analyze their activities against the transcription and translation
factors (eIF-4E, NF-kB and AP-1) described above to identify multi-
pathway inhibitors that may be useful in treating cancer. In the
previous study, only alkyl amino and aryl amino substitutions were
studied at the R1 position, therefore carboxyethyl amino and
benzoyl amino groups at this position were included in this study to
expand the scope of the SAR. Similarly, a methyl substitution on the
R2 position was explored in the previous study, whereas in the
current study phenyl substitutions at the R2 position were explored.
Para position in the phenyl ring at the 3rd position of quinazolinone
was found to be the key for the activity of this scaffold [28]
therefore, in present study effects of an electron withdrawing
group (eCl) and an electron donating group (eOCH3) were inves-
tigated at the R3 position in combination with the substitutions at
the R1 and R2 positions. To get some insights into the potential
mechanism of action of the compounds molecular docking study
was performed and results of the same is also discussed herein with
the results of in vitro experiments.
Scheme 1. Probable mechanism and the condition for the synthesis of desired
compounds. (a) Acetonitrile, 75e80 ꢀC, 4e5 h [28].
Table 1
2. Chemistry
Evaluation of synthesized compounds for their inhibitory activity towards NF-
AP-1 and eIF-4e mediated transcriptional activation in vitro.
kB,
The synthesis of designed compounds is shown in Scheme 1. The
thiourea derivatives 3 and 2-chloromethyl-3-aryl-3H-quinazolin-4-
one derivatives 2 were prepared according to a previously reported
procedure [28]. An appropriately substituted thiourea derivative 3
was S-alkylated by active methylene group of 2-chloromethyl-3-
aryl-3H-quinazolin-4-one derivatives 2 to give intermediate 4
which in the same reaction follows the intramolecular cyclization
followed by aromatic ring formation to give the desired 2-thiazole-
5-yl-3H-quinazolin-4-one derivatives 5 (Scheme 1). The elaborated
discussion of reaction mechanism for the synthesis of these
compounds is previously discussed [28]. Table 1 shows the list of
compounds that were prepared with modifications at R1, R2 and R3.
R3
O
N
R1
S
N
NH
N
R2
Entry
R1
R2
R3
% Inhibition in vitro at 1 mM
NF-kB
AP-1
eIFe4E
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
eCOOC2H5
eCH3
ePh
ePh
ePh
ePh
ePh
ePh
ePh
ePh
ePh
ePh
eCH3
eCH3
eCH3
eCH3
ePh
ePh
ePh
ePh
ePh
eH
eH
eH
eH
32
71
71
43
71
29
63
12
46
54
80
66
79
24
72
67
73
62
7
58
51
26
57
34
19
8
59
30
83
29
75
20
67
39
36
47
ꢁ63
33
37
30
ꢁ66
15
16
31
32
50
61
67
26
18
40
38
39
34
3. Biology
eCOPh
Development of the screens for NF-
previously discussed in detail [18]. In brief for the NF-
screens, two HEK293 cell lines were purchased from Panomics, one
was stably transfected with a plasmid in which luciferase expres-
k
B, AP-1 and eIF-4E was
B and AP-1
4-Cl Phenyl
eCOOC2H5
ePh
eH
k
eOCH3
eOCH3
eOCH3
eOCH3
eCl
eCl
eCl
eCl
eCl
eCl
eCl
eCl
eCl
eCOPh
4-Cl Phenyl
eCOOC2H5
eCH3
sion is regulated by six copies of the NF-kB transcriptional element
(50AGTTGAGGGGACTTTCCCAGGC-30) and the other in which lucif-
erase expression is regulated by three copies of the AP-1 tran-
scriptional element (TGACTAA). Activity of the compounds were
ePh
4-Cl Phenyl
eCOOC2H5
eCH3
ePh
eCOPh
4-Cl Phenyl
All the samples were analyzed in triplicate in each of two or three individual
experiments and data presented here is the average of two to three individual
experiments. Rapamycin a well-established inhibitor of eIF-4E was found to inhibit
50% luciferase activity at 10 nM. A pyrimidine carboxymide compound that had
e
previously been shown [29] to be an inhibitor of NF-
k
B and AP-1 mediated tran-
scriptional activation, reduced luciferase activity by 40% in the NF-
kB and AP-1 cell
e
Fig. 1. General structure of compounds.
lines @ 1 mM.

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R.S. Giri et al. / European Journal of Medicinal Chemistry 45 (2010) 3558e3563
measured both following induction of transcription by a transcrip-
tional stimulator and without induction. To test the biological
similar to their activity in inflammation. To analyze the effects of the
compounds on NF- B induction of transcriptional activity, the stably
transfected NF- B cell line was plated in a 96-well plate then 24 h
later, compounds 5i, 5j and 5r, which demonstrated robust NF-
inhibition in the un-induced cells, were added at six different
concentrations for 2 h followed by addition of 20 ng/ml of TNF- to
induce NF- B dependent gene transcription. Twenty-four hours
k
activity following induction, the promoters the HEK/NF-
were stimulated with TNF- (20 ng/ml) for 24 h and the HEK/AP-1
cell line with PMA (10 ng/ml) for 24 h. Both cell lines produced the
desired response with NF- driven luciferase expression
k
B cell line
k
a
kB
k
B
a
increasing by 100-fold and AP-1 driven expression by 30-fold
following the appropriate stimulation. Screening for eIF-4E was
carried out by cloning the highly structured 50 UTR of FGF (which
requires high levels of eIF-4E before protein synthesis can initiate)
into the pMS110 plasmid (pGL3-control plasmid from Promega
containing a neo selectable marker) and both the UTR and parental
plasmids were stably transfected into a cell line that has high levels
of eIF-4E (FaDu cell line), then analyzing luciferase activity in the
presence and absence of compound [18].
k
later, the cells were lysed, luciferase activity recorded and the values
normalized. Compounds 5i, 5j and 5r stronglyinhibited (>70%) TNF-
a
induced NF-
Similarly compounds 5k and 5l were also found to be giving IC50 of
3.3 and 5.5 M respectively in this assay [28].
kB gene expression at the 3.3 mM concentration.
m
To further analyze the binding conformations and binding affinity
of the active compounds from the above series, 5r was chosen as
a representative compound and analyzed for its potential binding
interaction with NF-kB using the docking protocol Flexidock. The
binding conformations and binding affinity of 5r to the NF-
binding site (the site at which the transcription factor binds to the
DNA sequence) was obtained. The Sybyl 6.9 package was used to
k
B’s DNA
4. Results and discussion
All the compounds synthesized were analyzed for activity in the
three screens discussed above and the results are shown in Table 1.
A number of the compounds inhibited luciferase expression under
the control of the FGF 50 UTR, while a few compounds actually
increased luciferase expression. These former molecules may
represent general inhibitors of translation, may reduce luciferase
activity by reducing cell number through inhibition of growth or
cell toxicity, or may actually act specifically to inhibit highly cap-
dependent translation. All the compounds were then tested for
analyze if and how the active compound interacts with NF-kB. The
thermodynamically stable conformations were first calculated and
the automated docking procedure (FlexiDock) with manual adjust-
ment was used to determine the most energetically favorable binding
location and orientation of 5r. The complex model with the lowest
energy among several docking models was selected for binding
observations. As an outcome of this study we have observed strong
charged interactions of 5r and NF-
k
B with a binding energy of ꢁ26.70
kcal/mol, where >C]O at 4th position of quinazoline ring presents
two [H] bond acceptor sites for Ser 72 and Ser 78 residues, N-1 of
quinazoline ring presents one [H] bond acceptor site for Gly66 and
eNHe of p-Cl phenylamino group at second position of thiazole ring
their ability to modulate NF-kB and AP-1 activity. A number of these
compounds exhibited mild to good inhibition in these assays
(Table 1). Seven representative compounds (5a, 5i, 5j, 5k, 5m, 5o
and 5r) were selected and retested in a six-point dose response
beginning at 10 nM and increasing in ½ log units. Screening was
carried out as described above monitoring the compound effects on
presents one [H] bond acceptor sites for Gly-52 residue of NF-
(Fig. 2). These results suggest that a direct interaction of 5r withNF-
may lead to the reduced transcriptional activity. This potential inter-
action of the compound in the DNA binding pocket of NF- B may
inhibit its binding to DNA and thus inhibit transcriptional activation.
k
k
B
B
basal NF-
Six of these compounds produced a very good dose dependent
inhibition of NF- B and AP-1 mediated transcriptional activity
k
B, AP-1 and FGF 50 UTR dependent luciferase expression.
k
k
(Table 2). Conversely, none of the compounds tested for the eIF-4E
elicited a good dose response inspite of the fact that during the
primary screening for eIF-4E, a few of these compounds did
demonstrate a good inhibition at 1 uM concentration. To analyze
general effects of compounds on cell viability, the seven
compounds were tested for their effects on cell number. An MTT
assay was used to analyze this effect on FaDu cells since it would
measure both inhibition of cell growth and cell toxicity. The
compounds were tested in six point dose-response starting at 10
nM and increasing to 3.3
mM. Of these, only compound 5k was
found to be inhibiting cell growth at 3.3
mM by more than 20%.
The above screening for transcriptional inhibition of luciferase
expression was carried out in non-induced cell lines to determine
the effects of compound on basal gene expression. However, it is also
possible that in cancer these transcriptional factors are induced,
Table 2
IC50 values of the selected representative compounds.
Entry
IC50 (mM) in vitro
NF-lB
AP-1
5.7
14
10
3.0
1.2
>500
4.3
5a
5i
>10
20
16
12
15
5j
Fig. 2. Compound 5r bound to NF-
(The protein model of NF- B (p50 homodimer) was taken from the Protein Data Bank
entry 1NFK [31]). Where -helix is purple colored, -stand is yellow and loops are in
blue. Selected protein residues are rendered in ball stick representation. The ligand (5r)
is rendered I capped stick representation. Color code for atoms: Carbon (white),
Hydrogen (cyan), Nitrogen (blue), Oxygen (red), Sulfur (yellow), Chlorine (green).
H-bonds are depicted by yellow dotted lines.
kB’s DNA binding site in ribbon tube representation
5k
5m
5o
5r
k
a
b
>250
3.3
IC50 values were calculated on the basis of dose response shown by the compounds
in dose dependent screening by using graph pad software.

R.S. Giri et al. / European Journal of Medicinal Chemistry 45 (2010) 3558e3563
3561
Though a more detailed study is necessary before concluding this is
5.2.2. 2-(2-Methylamino-4-phenyl-thiazol-5-yl)-3-phenyl-3H-
quinazolin-4-one (5b)
the primary mechanism of action of the compound, these docking
results partially explain a probable mechanism for transcriptional
inhibition elicited by these compounds.
As discussed in our previous communication [28] 2-(2,4-disub-
stituted-thiazole-5-yl)-3-aryl-3H-quinazolin-4-one has again proven
Yield: 48%, mp 210-2 ꢀC, Rf: 0.48, M.F.: C24H18N4OS, LC-MS (m/e):
411 (Mþ1), 1H NMR (400 MHz, CDCl3)
d 2.85 (s, 3H), 5.92 (d, 1H),
6.42e7.79 (m, 12H), 8.27 (d, 1H), 9.61 (s,1H)
to be a reasonable molecular scaffold to develop as inhibitor of NF-kB
5.2.3. 3-Phenyl-2-(4-phenyl-2-phenylamino-thiazol-5-yl)-3H-
quinazolin-4-one (5c)
and AP-1 mediated transcriptional activation. Although not as robust,
this scaffold may also show some promise in modulating eIF-4E
mediated translational activity. Although the limited number of
compounds and mixed biological response makes it very difficult to
develop a good SAR from this study the data does suggest a more
elaborated study is warranted to explore the full potential of this
Yield: 78%, mp 210-2 ꢀC, Rf: 0.70, M.F.: C29H20N4OS, LC-MS
(m/e): 473 (Mþ1), 1H NMR (400 MHz, CDCl3)
d 6.47e6.49
(m, 2H), 6.89e7.12 (m, 5H), 7.24e7.83 (m, 11H), 8.29 (d, 1H), 9.5
(s, 1H).
scaffold. As the transcription factors, NF-kB and AP-1, and the trans-
5.2.4. N-[5-(4-Oxo-3-phenyl-3,4-dihydro-quinazolin-2-yl)-4-
phenyl-thiazol-2-yl]-benzamide (5d)
lation factor, eIF-4E, have been demonstrated to play a pivotal role in
cancer, these inhibitors may represent novel multi targeted anti-
cancer agents. Additional characterization of these compounds and
related analogs will be reported in due course.
In conclusion we have synthesized and evaluated 2-thiazole-5-yl-
3H-quinazolin-4-one derivatives as inhibitors of NF-kB and AP-1
Yield: 75%, mp >275 ꢀC, Rf: 0.67, M.F.: C30H20N4O2S, LC-MS (m/e):
501 (Mþ1), 1H NMR (300 MHz, DMSO-d6)
d 6.65e7.12 (m, 5H),
7.18e7.77 (m, 9H), 7.84 (d,1H), 7.91 (dd,1H), 8.08e8.18 (m, 3H),11.91
(s, 1H)
mediated transcriptional and eIF-4E mediated translational activa-
tion. The outcome of this study suggest that the 2-thiazole-5-yl-3H-
quinazolin-4-one derivative is a promising scaffold for developing
multi-pathwayinhibitorsandcanbeexploredtoitsfullpotentialasan
anti-cancer scaffold.
5.2.5. 2-[2-(4-Chloro-phenylamino)-4-phenyl-thiazol-5-yl]-3-
phenyl-3H-quinazolin-4-one (5e)
Yield:71%, mp >275 ꢀC,Rf:0.68, M.F.:C29H19N4OSCl, LC-MS(m/e):
507 (Mþ1), 1H NMR (400 MHz, CDCl3)
d 6.60 (dd, 2H), 7.07e7.84
(m, 15H), 8.3 (dd, 1H), 9.38 (s, 1H)
5.2.6. {5-[3-(4-Methoxy-phenyl)-4-oxo-3,4-dihydro-quinazolin-2-yl]-
4-phenyl-thiazol-2-yl}-carbamic acid ethyl ester (5f)
5. Experimental
Yield: 46%, mp 256-7 ꢀC, M.F.: C27H22N4O4S, LC-MS (m/e):
499,521 (Mþ1, Mþ23), 1H NMR (300 MHz, DMSO-d6)
d 1.23 (t, 3H),
Unlessmentionedotherwiseallthestartingmaterialsandsolvents
were purchased from commercially available sources. Melting points
were determined on a Toshniwal melting point apparatus using open
glass capillary tubes and represent the uncorrected values. Proton
NMR spectra were measured on a Bruker spectrometer using the
specified solvents at RSIC Chandigarh. LC-MS analysis was done on
a Perkin Elmer Applied Bio-Sciences API-165 in the analytical labo-
ratory of B. V. Patel PERD Centre, Ahmedabad. Elemental analysis was
carried out at RSIC, IIT Powai on Perkin Elmer 2400 CHN elemental
analyzer. All the reactions were monitored using thin layer chroma-
tography (TLC) using glass plate coated with silica gel G or GF254. TLC
plates were developed in iodine and toluene: acetonitrile (7:3) was
taken as mobile phase, unless mentioned otherwise.
2.47 (s, 3H), 4.20 (q, 2H), 6.85e7.95 (m, 12H), 8.16 (d, 1H), 11.9
(s, 1H).
5.2.7. 3-(4-Methoxy-phenyl)-2-(4-phenyl-2-phenylamino-thiazol-
5-yl)-3H-quinazolin-4-one (5g)
Yield:49%, mp135-7ꢀC, Rf:0.58, M.F.:C30H22N4O2S, LC-MS(m/e):
503 (Mþ1), 1H NMR (300 MHz, DMSO-d6)
d 3.43 (s, 3H), 6.76e7,75
(m, 17H), 8.2 (d, 1H), 10.42 (s, 1H).
5.2.8. N-{5-[3-(4-Methoxy-phenyl)-4-oxo-3,4-dihydro-quinazolin-
2-yl]-4-phenyl-thiazol-2-yl}-benzamide (5h)
Yield: 57%, mp >275 ꢀC, Rf: 0.61, M.F.: C31H22N4O3S, LC-MS (m/e):
531, 553, 569 (Mþ1, Mþ23, Mþ39), 1H NMR (300 MHz, DMSO-d6)
d
3.64 (s, 3H,), 6.62e7.65 (m, 13H), 7.81 (d, 1H), 7.90 (dd, 1H),
8.08e8.18 (m, 3H), 12.88 (s, 1H).
5.1. General synthesis of 2-(2,4-disubstituted-thiazole-5-yl)-3-aryl-
3H-quinazolin-4-one derivatives
5.2.9. 2-[2-(4-Chloro-phenylamino)-4-phenyl-thiazol-5-yl]-3-(4-
methoxy-phenyl)-3H quinazolin-4-one (5i)
The general procedure for synthesis of compounds with general
structure 5 is as follows. 0.01 mol of thiourea derivatives 3 in 5 ml of
acetonitrile was added to a stirred solution of 0.01 mol of the
respective 2-chloromethyl-3-aryl-3H-quinazolin-4-one 2 in 5 ml of
acetonitrile at 75e80 ꢀC. This reaction mixture was stirred at this
temperature for 2e4 h and then cooled to room temperature. The
compound that precipitated was filtered and recrystallized in
methanol to obtain solid crystals.
Yield: 52%, mp 220-21 ꢀC, Rf: 0.62. M.F.: C30H21N4O2SCl, LC-MS
(m/e): 537 (Mþ1), 1H NMR (400 MHz, CDCl3)
6.50e7.82 (m, 16H), 8.32 (dd, 1H), 9.63 (s, 1H).
d 3.73 (s, 3H),
5.2.10. {5-[3-(4-Chloro-phenyl)-4-oxo-3,4-dihydro-quinazolin-2-
yl]-4-methyl-thiazol-2-yl}-carbamic acid ethyl ester (5j)
Yield: 46%, mp >275 ꢀC, Rf: 0.46, M.F.: C21H17N4O3SCl, LC-MS
(m/e): 441, 463 (Mþ1, Mþ23), 1H NMR (300 MHz, CDCl3)
d 1.33
(t, 3H), 2.3 (s, 3H), 4.24 (q, 2H), 7.18e7.84 (m, 7H), 8.28 (dd,1H), 9.85
(s, 1H).
5.2. Characterization of synthesized compounds
5.2.11. 3-(4-Chloro-phenyl)-2-(4-methyl-2-methylamino-thiazol-
5-yl)-3H-quinazolin-4-one (5k)
5.2.1. [5-(4-Oxo-3-phenyl-3,4-dihydro-quinazolin-2-yl)-4-phenyl-
thiazol-2-yl]-carbamic acid ethyl ester (5a)
Yield:41%, mp195-7ꢀC, Rf:0.61, M.F.:C19H15N4OSCl, LC-MS(m/e):
Yield: 79%, mp 245-7 ꢀC, Rf: 0.41, M.F.: C26H20N4O3S, LC-MS (m/e):
384 (Mþ1), 1H NMR (400 MHz, CDCl3)
d 2.33 (s, 3H), 2.82 (s, 3H),
469,491 (Mþ1, Mþ23), 1H NMR (300 MHz, DMSO-d6)
d 1.90 (t, 3H),
7.17e7.82 (m, 7H), 8.28e8.30 (m, 1H), 9.37 (s, 1H).
4.24 (q, 2H), 6.81e7.92 (m, 13H), 8.1 (d, 1H), 10.5 (s, 1H).

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R.S. Giri et al. / European Journal of Medicinal Chemistry 45 (2010) 3558e3563
5.2.12. 3-(4-Chloro-phenyl)-2-(4-methyl-2-phenylamino-thiazol-
5-yl)-3H-quinazolin-4-one (5l)
distance-dependent dielectric constants and the conjugate
ꢀꢁ1
gradient method until the gradient reached 0.001 kcal molꢁ1
A .
Yield: 51%, M.P.: 277e80 ꢀC, Rf: 0.84, M.F.: C24H17N4O2SCl, LC-MS
Representative conformers selected from clusters of low-energy
conformers obtained in the conformational search were re-optimized
by semi empirical molecular orbital calculations with the PM3
(m/e): 446 (Mþ1), 1H NMR (400 MHz, CDCl3)
d 2.46 (s, 3H),
6.94e7.85 (m, 12H), 8.23e8.25 (dd, 1H), 9.91 (s, 1H).
method in the MOPAC 6.0 package. The protein model of NF-kB (p50
5.2.13. 3-(4-Chloro-phenyl)-2-[2-(4-chloro-phenylamino)-4-
methyl-thiazol-5-yl]-3H-quinazolin-4-one(5m)
homodimer) was taken from the Protein Data Bank entry 1NFK [31].
DNA-bindingregion,aminoacidresidues59e71(59:Arg, Tyr, Val, Cys,
Glu, Gly, Pro, Ser, His, Gly, Gly, Leu, Pro: 71) of NFkBwere identified the
as active site. This site was used for the docking of ligand [32,33].
Flexible docking was facilitated through the FlexiDock utility in
the Biopolymer module of SYBYL 6.9. During flexible docking, the
ligand and the side chains of hydrophilic amino acids in the puta-
tive binding site were defined as rotatable bonds. After the
hydrogen atoms were added to the receptor, atomic charges were
recalculated by using Kollman All-atom for the protein and Gas-
teigereHuckel for the ligand. H-bonding sites were marked for all
residues in the active site and ligands with H-bond donor or
acceptor. Ligands were pre-positioned in the putative binding
cavity guided by several superimposition results. Default FlexiDock
parameters were set at 3000-generations for genetic algorithms. To
increase the binding interaction, the torsion angles of the side
M.F.: C24H16N4OSCl2, LC-MS (m/e): 479, 480, 502 (Mþ, Mþ2,
Mþ23), 1H NMR (300 MHz, CDCl3)
d 2.29 (s, 3H), 7.29 (d, 2H),
7.44e7.58 (m, 7H), 7.72 (d, 1H), 7.86 (dd, 1H), 8.14 (d, 1H), 10.33 (s,
1H), Elemental analysis results were: calculated for C24H16N4OSCl2;
C 60.25, H 4.18, N 7.27, obtained; C 60.26, H 4.09, N 7.26.
5.2.14. {5-[3-(4-Chloro-phenyl)-4-oxo-3,4-dihydro-quinazolin-2-
yl]-4-phenyl-thiazol- 2-yl}-carbamic acid ethyl ester (5n)
Yield:50%, mp >275 ꢀC, Rf:0.62,M.F.:C26H19N4O3SCl, LC-MS (m/e):
502, 505, 525 (Mþ, Mþ3, Mþ23), 1H NMR (400 MHz, CDCl3)
d 1.67
(t, 3H), 4.22e4.30 (q, 2H), 6.47e7.88 (m,12H), 8.25e8.32 (m,1H),11.40
(s,1H), Elemental analysis results were calculated for C26H19N4O3SCl; C
62.09, H 3.81, N 11.14, obtained; C 62.12, H 3.77, N 11.15.
ꢀ
5.2.15. 3-(4-Chloro-phenyl)-2-(2-methylamino-4-phenyl-thiazol-
5-yl)-3H-quinazolin-4-one (5o)
chains within 5 A of the ligands were manually adjusted from the
results of FlexiDock. Finally, the complexes were minimized by
Yield: 45%, mp >275 ꢀC, Rf: 0.61, M.F.: C24H17N4OSCl, LC-MS (m/e):
using the powell method with a fixed dielectric constant (4.0), until
446 (Mþ1), 1H NMR (400 MHz, CDCl3)
d
2.87 (s, 3H), 5.96 (d, 1H),
the conjugate gradient reached 0.001 kcal molꢁ1
A .
ꢀꢁ1
6.46e7.84(m,11H),8.29(d,1H),9.47(s,1H), Elementalanalysisresults
were: calculated for C24H17N4OSCl; C 64.79, H 3.85, N 12.59, obtained;
C 64.78, H 3.84, N 12.58.
Acknowledgements
R.S.G. thanks to RSIC Mumbai and Chandigarh for recording
NMR and elemental analysis. Support from the Feist-Weiller Cancer
Center and an IC grant provided by Industrial Commissionerate of
Gujarat for carrying out this work is acknowledged. We thank
Professor Harish Padh and Professor C. J. Shisoo, Directors, B.V. Patel
PERD centre, for their support towards this project.
5.2.16. 3-(4-Chloro-phenyl)-2-(4-phenyl-2-phenylamino-thiazol-5-
yl)-3H-quinazolin-4-one (5p)
Yield: 53%, mp 218-20 ꢀC, Rf: 0.64, M.F.: C29H19N4OSCl, LC-MS
(m/e): 508 (Mþ1), 1H NMR (400 MHz, CDCl3)
7.01e7.83 (m, 15H), 8.3 (dd, 1H), 8.5 (s, 1H).
d 6.47e6.49 (m, 2H),
5.2.17. N-{5-[3-(4-Chloro-phenyl)-4-oxo-3,4-dihydro-quinazolin-
2-yl]-4-phenyl-thiazol-2-yl}-benzamide (5q)
References
Yield: 53%, mp >275 ꢀC, Rf: 0.54, M.F.: C30H19N4O2SCl, LC-MS
(m/e): 535 (Mþ1), 1H NMR (400 MHz, CDCl3)
8.27e8.29 (dd, 1H), 12.2 (s, 1H)
d 6.50e8.18 (m, 17H),
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