Discovery of novel quinazolinones and their acyclic analogues as multi-kinase inhibitors: design, synthesis, SAR analysis and biological evaluation

Article abstract of DOI:10.1039/C6RA19137A

This work deals with the design and synthesis of some novel 6-iodo-2-(pyridin-3/4-yl)-3-substituted quinazolin-4-one derivatives 8a-l, 10a-h, 13-18 in addition to certain acyclic analogues thereof viz.9a-n and 12a-h. The molecular design strategy was based on structural analogy between the new compounds and reported quinazolines and their acyclic analogues. This design scheme led to the synthesis of 8 new intermediates and 58 new final quinazolinones. The target compounds were evaluated for their antitumor activity against a panel of nine cancer cell lines viz. breast cancer (MCF-7, MDAMB-231, MDAMB-435 and HS-578T), colon cancer (HT-29 and HCC-2998) and leukemia (CCRF-CEM, K-562 and HL-60). The quinazolinones 10a-h displayed exceptional antitumor activity and compounds 12a-h showed superior potency against MCF-7. These compounds were further subjected to in vivo study. Kinase inhibitory assay was also carried out to investigate the mechanism of action of the target compounds and they displayed the highest activity against ABL, ALK and c-RAF kinases. The 3-substituted quinazolinones 10a-h showed the highest kinase activity inhibitory potency against ABL, ALK and c-RAF with the most active compound in this study being the fluoro-3-pyridyl derivative 10a. These results are in compliance with the observed antitumor activity. Finally, a molecular modeling study was performed to interpret the potential molecular interactions of these chemotypes with the most responsive biomolecular target ABL.

Full text of DOI:10.1039/C6RA19137A

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Discovery of novel quinazolinones and their acyclic
analogues as multi-kinase inhibitors: design,
synthesis, SAR analysis and biological evaluation
Cite this: RSC Adv., 2016, 6, 111767
Nehad A. El Sayed,a Amal A. Eissa,a Ghada F. El Masry,a Mohamed M. Abdullahb
c
*
and Reem K. Arafa
This work deals with the design and synthesis of some novel 6-iodo-2-(pyridin-3/4-yl)-3-substituted
quinazolin-4-one derivatives 8a–l, 10a–h, 13–18 in addition to certain acyclic analogues thereof viz. 9a–
n and 12a–h. The molecular design strategy was based on structural analogy between the new
compounds and reported quinazolines and their acyclic analogues. This design scheme led to the
synthesis of 8 new intermediates and 58 new final quinazolinones. The target compounds were
evaluated for their antitumor activity against a panel of nine cancer cell lines viz. breast cancer (MCF-7,
MDAMB-231, MDAMB-435 and HS-578T), colon cancer (HT-29 and HCC-2998) and leukemia (CCRF-
CEM, K-562 and HL-60). The quinazolinones 10a–h displayed exceptional antitumor activity and
compounds 12a–h showed superior potency against MCF-7. These compounds were further subjected
to in vivo study. Kinase inhibitory assay was also carried out to investigate the mechanism of action of
the target compounds and they displayed the highest activity against ABL, ALK and c-RAF kinases. The 3-
substituted quinazolinones 10a–h showed the highest kinase activity inhibitory potency against ABL, ALK
and c-RAF with the most active compound in this study being the fluoro-3-pyridyl derivative 10a. These
results are in compliance with the observed antitumor activity. Finally, a molecular modeling study was
performed to interpret the potential molecular interactions of these chemotypes with the most
responsive biomolecular target ABL.
Received 28th July 2016
Accepted 11th November 2016
DOI: 10.1039/c6ra19137a
www.rsc.org/advances
role in disruption of various cellular activities like growth,
proliferation and migration, and thus is a hallmark of some
types of cancer.12
Introduction
Quinazoline nucleus-based small molecules have aroused
recent attention from chemical and biological view-points. This
is attributed to the fact that members of this chemotype have
found their way to the pharmaceutical products market aer
being granted FDA approval through displaying remarkable
antineoplastic activity supported by a diverse array of mecha-
nisms of action.1–5 Among the disclosed modes of cytotoxicity
activities elicited by quinazolines is their effect as dihydrofolate
reductase antimetabolites,6,7 DNA intercalators3 or protein
kinase inhibitors.8–11
The human genome contains about 500 protein kinase genes
constituting about 2% of all human genes.12 Up to 30% of all
human proteins are modiable by kinase activity where kinases
are well-known regulators of many cellular pathways, especially
those involved in signal transduction.13 Hence, it was found that
protein kinases' mutation and/or over-expression play a central
Much attention has been given to exploiting small molecules
that can target protein kinases as inhibitors where such
chemical entities have proved effective as anti-proliferative and/
or cytotoxic agents.10,12,14–16 Quinazoline-based molecules are
one such class that act through interfering with the enzymatic
phosphorylation ability of kinase enzyme family members. The
antikinase activity of quinazolines is generally due to their
ability to competitively occupy the adenosine triphosphate
(ATP)-binding pocket with high affinity, thus acting as ATP-
mimic inhibitors.17–20
Despite extensive efforts within the oncology eld to develop
kinase inhibitors (KIs), uncertainty remains over the relative
merits of selective compounds versus less selective or “multi-
targeted” inhibitors.21 Targeted molecules offer the clearest
indication that in vivo effects result from the intended in vitro
activity. Moreover, toxicity derived from additional activity
against other kinases is likely to be reduced.22 However, inhi-
bition of a single kinase may not be sufficient to achieve
a clinical benet, either through the built-in redundancy of
signaling pathways, or the ability of tumors to acquire resis-
tance.23 Inhibitors with activity against multiple kinases may in
aDepartment of Pharmaceutical Chemistry, Faculty of Pharmacy, Cairo University,
11562, Egypt
bResearch Unit, Mapco Pharmaceutical Industries, Balteim, Egypt
cUniversity of Science and Technology, Zewail City of Science and Technology, Cairo
12588, Egypt. E-mail: rkhidr@zewailcity.edu.eg; Tel: +20 01002074028
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fact be more effective anticancer agents, and several multi-
Moreover, the 6-position of the quinazolinone was kept
targeted kinase inhibitors are now commercially available like occupied by the bulky hydrophobic iodo substituent aiming to
sorafenib.24
furnish the necessary bulkiness and hydrophobic character
Consequently, and motivated by the aforementioned facts, it required at this position besides acting as a metabolic blocker
deemed of interest to design, synthesize and evaluate novel to prolong the duration of action of these molecules.
quinazolinones as potential kinase inhibitors and cytotoxic
agents.
Finally, modications related to decreasing the rigidity of the
structure through quinazoline ring opening were performed to
assess the effect of increasing the backbone exibility on the
molecular binding interactions mode with the active site amino
acids of the target enzyme and on hence on the investigated
biological activity.
Molecular design strategy
Design of the target compounds (Fig. 1) focused on alternating
the physicochemical properties of the functional groups on the
2,3,6-trisubstituted quinazolinone backbone to target different
regions of the ATP-binding pocket of the protein kinase
domain. The ATP binding site has multiple regions viz. the
adenine binding region which contains two key hydrogen bonds
formed by the interaction of N-1 and N-6 amino group of the
adenine ring; the sugar binding region which is mostly hydro-
philic in nature; a hydrophobic pocket that is not used by ATP
but plays a crucial role in inhibitor selectivity; hydrophobic
channels that are not occupied by ATP yet may be exploited for
inhibitor specicity, and nally the phosphate binding region
which can be targeted for improving inhibitor selectivity.20,25
The newly synthesized compounds were initially screened
for their cytotoxic activities against a panel of cancer cell lines.
Compounds showing remarkable anticancer activity were sub-
jected to in vivo study. The kinase inhibitory activity was also
evaluated to elucidate the mechanism of action of the active
derivatives. Furthermore, a molecular docking study was
carried out to predict the plausible binding mode of these
chemotypes with the active site of the most responsive kinase
ABL.
Chemistry
Molecular manipulations on the quinazoline backbone were The key starting materials, 5-iodoanthranilic acid 3,26 nicotinoyl
performed according to the following rationale:
chloride 4a 27 and isonicotinoyl chloride 4b,28 were prepared as
First the 2-position of the quinazolinone ring, a heteroaryl reported. 5-Iodo-2-[(pyridinyl-3/4-carbonyl)amino]benzoic acid
viz. 3-pyridyl or 4-pyridyl was introduced to study the effect of derivatives 5a, b were prepared via the reaction of 5-iodoan-
this positional isomerism on the enzymatic and anticancer thranilic acid 3 and the freshly prepared nicotinoyl chloride 4a
ꢀ
activities of these derivatives.
or isonicotinoyl chloride 4b in dry DMF at 80 C. The benzox-
For the 3-position, fragment molecular manipulation was azinone derivatives 6a, b were synthesized through the nucle-
carried out with different groups of diverse lipophilicity and ophilic reaction of the amidated anthranilic acid with acetic
length viz. substituted/unsubstituted urea or thiourea, carbox- anhydride or via the reaction of the substituted anthranilic acid
ꢀ
amide, carbothioamide, carbothiohydrazide, phenyl hydrazine, and the freshly prepared acyl chloride in dry pyridine at 5 C.
hydroxylamine, para-aminobenzoic acid, benzylideneamino Moreover, the reaction of benzoxazinones 6a, b with hydrazine
and substituted acetylhydrazinyl to study the effect of these hydrate was carried out in various solvents or under various
changes on their occupation of the lipophilic pocket of the ATP reactions conditions viz. reux in ethanol or n-butanol or direct
binding site of kinases and hence their associated cytotoxic solvent-free fusion of the reactants to obtain the desired 3-
activity as compared to well-known KIs.
aminoquinazolinone. In all cases, the reaction afforded the
Fig. 1 Design strategy for the target 2,3,6-trisubstituted quinazolinones and their acyclic analogs.
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Scheme 1 Synthesis of derivatives 7a, b. Reagents and solvents: (i) KOH; (ii) I2 KOH, glacial acetic acid; (iii) DMF; (iv) acetic anhydride; (v) dry
pyridine; (vi) NH2NH2$H2O, absolute ethanol.
open ring structures; N-[2-(hydrazinylcarbonyl)-4-iodophenyl] presence of anhydrous sodium acetate furnished the carbox-
pyridine-3/4-carboxamide 7a, b which were isolated and iden- amide derivatives 14a, b carbothioamides 15a, b and carbo-
tied. Attempts to cyclize the diamides 7a, b to the corre- thiohydrazides 16a, b. Moreover, the phenylamino derivatives
sponding 4-(3H)-quinazolin-4-one under various reaction 17a–f were prepared through reuxing the various hydrazine
conditions, including sodium ethoxide or polyphosphoric acid derivatives with the benzoxazinone compounds 6a, b in abso-
were unsuccessful (Scheme 1).
lute ethanol. Finally, reaction of a mixture of benzoxazinones
Compounds 8a–l were prepared by reuxing the diamide 6a, b with p-aminobenzoic acid under fusion conditions at
ꢀ
derivatives 7a, b with the appropriate isocyanate or iso- 200 C yielded the benzoic acid derivatives 18a, b (Scheme 3).
thiocyanate in dioxane in the presence of catalytic amount of
triethylamine. Synthesis of the benzylideneamino derivatives
9a–n was carried out by treating the hydrazido derivatives with
the appropriate aldehyde in absolute ethanol in presence of few
drops of glacial acetic acid. Cyclodehydration reaction was then
Results and discussion
(1) Biological evaluation
carried out using sodium ethoxide to obtain the benzylidene
(a) In vitro cytotoxic activity. All the newly synthesized
aminoquinazolinones 10a–h from the diamide analogues. Also, compounds were tested in vitro for their cytotoxic activity
the desired 2-chloroacetyl hydrazinyl derivatives 11a, b were against a panel of 9 human cancer cell lines. This panel involved
successfully synthesized by reacting compounds 7a, b with four breast cancer cell lines (MCF-7, MDAMB-231, MDAMB-435
chloroacetyl chloride in dry dimethylformamide at room and HS-578T), two colon cancer cell lines (HT-29 and HCC-2998)
temperature. The chloroacetylhydrazinyl derivative11a and its and three leukemia cell lines (CCRF-CEM, K-562 and HL-60).
4-pyridyl analogue 11b were reuxed with morpholine, piperi- The obtained IC50 values are displayed in Table 1.
dine or substituted piperazines in absolute ethanol containing
anhydrous potassium carbonate to produce the desired h, 13a, b, 14a, b, 15a, b, 16a, b, 18a, b and 17a–f and N-(2,4-
compounds 12a–h (Scheme 2).
disubstituted phenyl)pyridine-carboxamides; 9a–n and 12a–h
As shown in Table 1, most of the novel quinazolinones; 10a–
The hydroxy derivatives 13a and 13b were synthesized via displayed single digit to sub-nanomolar level cytotoxic activity
reuxing 6a and 6b with hydroxylamine hydrochloride in dry against the breast cancer cell lines MCF-7, MDA-MB-231, HS-
pyridine. Reuxing the benzoxazinone derivatives 6a, b with 578T and MDA-MB-435; the colon cancer cell lines HT29 and
urea, thiourea or thiosemicarbazide in glacial acetic acid in the HCC-2998 and leukemia cell lines CCRF-CEM, K-562 and HL-60.
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Scheme 2 Synthesis of derivatives 8–12. Reagents and solvents: (i) appropriate isocyanate/isothiocyanate, TEA, dioxane; (ii) p-substituted
benzaldehyde, absolute ethanol, glacial acetic acid; (iii) sodium ethoxide, absolute ethanol; (iv) ClCH2COCl, DMF; (v) secondary amines, absolute
ethanol, anhydrous K2CO3.
SAR analysis into the ndings of the cytotoxicity of the pyridinecarboxamides with acetyl hydrazinyl-carrying side
synthesized compounds against breast cancer cell lines chain12a–h (IC50 ¼ 0.017–0.65 mM) displayed high cytotoxicity
revealed that most of the quinazolinyl urea/thiourea derivatives against the colon cancer cell lines. The pyridinecarboxamides
8a–l had weak activity on MCF-7 (>100 mM). However, most of bearing benzylidene moiety 9a–n exhibited the weakest anti-
the pyridinecarboxamides bearing benzylidene moiety 9a–n neoplastic activity against colon cancer cell lines (IC50 ¼ 0.05–
exhibited potent growth inhibitory activity. All the 3-pyridyl 0.96 mM). Concerning leukemia CCRF-CEM, K-562 and HL-60
analogues displayed enhanced activity against all breast cancer cell lines, all the tested compounds exhibited high growth
cell lines (IC50 ¼ 0.0032–0.009 mM). Looking at the 4-pyridyl inhibitory activity (IC50 ¼ 0.085–0.0013 mM).
series, some congeners exhibited weak activity (>100 mM),
(b) In vitro cytotoxicity against the non-tumorigenic MCF-
particularly; the uoro derivative 9h against MCF-7, the chloro 10A cell line and selectivity. Before progressing into the in
derivative 9i against HS-578T and MDA-MB-435, the alkylated vivo cytotoxic screening against MCF-7 mouse xenogra model,
and nitro analogues 9l, 9m and 9n against MCF-7 and MDA-MB- selected compounds viz. 10a–h, 12b and 12f were subjected to in
435. Interestingly, the target quinazolinones 10a–h, 13–16, vitro evaluation against the non-tumorigenic breast cell lines
18a,b & 17a–f and pyridinecarboxamides with acetyl hydrazinyl- MCF-10A. The cell line used was selected to represent healthy
carrying side chain12a–h displayed superior cytotoxic activity cells and to determine whether the cytotoxic properties of the
against all the breast cancer cell lines. Compounds 12a–h tested compounds were selective for malignant cells in contrast
showed high potency, particularly against MCF-7 (IC50 ¼ 0.12– to nonmalignant cells. Data obtained are enlisted in Table 2.
0.78 nM). Furthermore, the quinazolinones Schiff's base deriv-
atives 10a–h displayed exceptional antitumor activity (IC50
With regards to selective cytotoxicity of the tested
compounds relative to examined breast cancer cell lines,
¼
0.010–0.022 nM) against MCF-7 in the subnanomolar range. derivatives in the 10a–h series were >2164 to >33 701 fold more
Regarding the cytotoxicity against colon HT29 and HCC-2998 selective to cancer cell lines than nonmalignant cells with 10b
cancer cell lines the quinazolinones Schiff's base derivatives showing the highest selectivity. On the other hand, compounds
10a–h (IC50 ¼ 0.0031–0.049 mM) and most of the quinazolinyl tested from other series 12b and 12f exhibited >223 and >518
urea/thiourea derivatives 8a–l (IC50 ¼ 0.0034–0.65 mM) dis- fold selectivity towards the malignant breast cancer cell lines,
played superior anticancer activity. Moreover, the target qui- respectively. It can be inferred from these results that the che-
nazolinones 13–16, 18a, b & 17a–f (IC50 ¼ 0.014–0.098 mM) and motypes in hand show considerable safety margin towards
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Scheme 3 Synthesis of derivatives 13–18. Reagents and solvents: (i) NH2OH$HCl, pyridine; (ii) urea, anhydrous CH3COONa, glacial acetic acid;
(iii) thiourea, anhydrous CH3COONa, glacial acetic acid; (iv) thiosemicarbazide, anhydrous CH3COONa, glacial acetic acid; (v) appropriate
phenylhydrazine, absolute ethanol; (vi) p-aminobenzoic acid.
nonmalignant breast cells compared to malignant breast cancer were also 1.12–2.56 fold more potent than the positive control
cell lines when tested under the same experimental conditions. drug sorafenib (tumor volume 9.1 cm3).
(c) In vivo cytotoxic activity. Compounds showing higher
It is worth mentioning that the in vivo experiments were tried
cytotoxic activity against all breast cancer cell lines, particularly on two doses of the tested compounds (low dose of 5 mg kgꢁ1
MCF-7 (10a–h and 12a–h) were selected to perform in vivo and high dose of 10 mg kgꢁ1, every 48 h for 3 weeks) and data
cytotoxicity and data are given in Table 3.
reported herein are for the low dose. This is because rst tumor
Results showed that all compounds were 3.89–11.23 fold volume changes observed at the two tested doses were not
more potent than the vehicle control, showing tumor volume in signicantly different. Furthermore, there was no dose-
the range of 3.58–8.12 cm3 as compared to the vehicle control dependent toxicity in animal groups treated at the two dose
which displayed a tumor volume of 40.21 cm3. This is perfectly levels (data not shown).
in compliance with the in vitro cytotoxicity data where the most
(d) Kinases inhibition assay. To investigate the mechanism
active compound among the pyridinecarboxamide series of action and the kinase inhibitory prole of the target
bearing acetyl hydrazinyl side chain 10a (IC50 ¼ 0.012 nM) was compounds, they were tested at a single dose concentration of
also the most active in vivo. Interestingly, the tested compounds 10 mM over a panel of 30 kinases to determine their kinase
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Table 1 Cytotoxic activities of the target quinazolinones; 8a–l, 10a–h, 13a, b, 14a, b, 15a, b, 16a, b, 18a, b, 17a–f and N-(2,4-disubstituted phenyl)
pyridinecarboxamides; 9a–n and 12a–h against breast, colon and leukemia cell lines
IC50a (mM)
Breast cancer
MCF-7
Colon cancer
MDA-MB-435
Leukemia
Cpd no.
MDA-MB-231
HS-578T
HT-29
HCC-2998
CCRF-CEM
K-562
HL-60
8a
8b
8c
8d
8e
8f
8g
8h
8i
8j
8k
8l
9a
9b
9c
9d
9e
9f
9g
9h
9i
9j
9k
9l
>100
>100
>100
0.0009
>100
0.00098
0.0065
>100
>100
>100
0.0012
>100
>100
0.0042
>100
>100
>100
>100
0.087
0.049
>100
>100
0.021
0.035
0.034
0.023
>100
0.024
0.0027
0.0016
0.0068
0.0045
0.0054
0.0039
0.0056
0.043
0.0064
0.0045
0.65
0.75
0.05
0.0078
0.0055
0.0046
0.0057
0.0074
0.0079
0.0064
0.26
0.15
0.48
0.65
0.39
0.57
0.76
0.95
0.87
0.0043
0.0054
0.71
0.82
0.93
0.0065
0.0075
0.0070
0.0099
0.0088
0.0076
0.0034
0.35
0.24
0.58
0.36
0.47
0.46
0.55
0.86
0.64
0.0575
0.0484
0.0395
0.0486
0.05679
0.0466
0.0579
0.0457
0.0335
0.0344
0.0455
0.0468
0.0254
0.0135
0.0443
0.0244
0.0334
0.0344
0.0354
0.0546
0.0455
0.0435
0.0565
0.0456
0.0544
0.0478
0.041
0.0696
0.087
0.0765
0.065
0.0658
0.0899
0.0640
0.0273
0.0219
0.0367
0.0349
0.0655
0.0894
0.0673
0.0456
0.0258
0.3212
0.1243
0.0565
0.0893
0.0674
0.4566
0.0904
0.0566
0.0874
0.0435
0.0566
0.0575
0.0264
0.0348
0.031
>100
>100
0.0068
0.0046
>100
>100
>100
>100
0.0057
>100
0.0065
0.0012
>100
>100
>100
>100
0.01
>100
0.0032
0.0067
0.0067
0.0080
0.0089
0.0045
0.0090
0.0043
>100
0.0046
45
11
0.0052
67
0.00045
0.00057
0.00078
0.00085
0.00087
0.00095
0.00097
0.00099
0.0044
0.0055
0.0046
0.0055
0.0074
0.0083
0.0075
0.0066
0.0047
0.0058
0.0069
0.0078
0.0087
0.0075
0.0064
0.0054
0.0054
0.004
0.054
0.0505
0.0344
0.0235
0.0366
0.0478
0.0484
0.0455
0.0268
0.0156
0.0467
0.0255
0.0366
0.0378
0.0755
0.0953
0.0843
0.0755
0.0544
0.0654
0.0566
0.043
0.0095
>100
0.0054
0.0034
0.0056
0.0065
0.0078
0.0032
0.0089
>100
0.0065
0.0045
>100
>100
0.0034
>100
0.000012
0.000014
0.000016
0.000018
0.000019
0.000021
0.000022
0.000023
0.00012
0.00024
0.00035
0.00046
0.00054
0.00064
0.00076
0.00078
0.0023
0.0043
0.0054
0.0067
0.0065
0.0043
0.0023
0.0045
0.0045
0.0034
0.0056
0.0078
0.0075
0.0045
0.0034
0.0034
0.0047
0.0043
0.0078
0.0087
0.0090
0.0065
0.0053
0.0065
0.0034
0.0038
0.0077
0.0056
0.0074
0.0064
0.00010
0.00011
0.00012
0.00014
0.00015
0.00016
0.00017
0.00022
0.0035
0.0043
0.0056
0.0078
0.0098
0.0054
0.0026
0.0045
0.0032
0.0045
0.0076
0.0053
0.0045
0.0067
0.0089
0.0087
0.0033
0.0044
0.0053
0.0032
0.0045
0.0054
0.0065
0.0074
>100
0.034
>100
>100
0.0043
>100
0.06
0.07
0.06
0.05
0.97
0.94
0.03
0.96
9m
9n
0.06
0.85
10a
10b
10c
10d
10e
10f
10g
10h
12a
12b
12c
12d
12e
12f
12g
12h
13a
13b
14a
14b
15a
15b
16a
16b
17a
17b
17c
17d
17e
17f
18a
18b
0.00028
0.00029
0.00035
0.00036
0.00044
0.00045
0.00046
0.00048
0.0035
0.0046
0.0047
0.0058
0.0067
0.0056
0.0045
0.0036
0.0078
0.0063
0.0052
0.0083
0.0091
0.0082
0.0093
0.0087
0.0078
0.0089
0.0036
0.0045
0.0054
0.0065
0.0056
0.0045
0.0051
0.0056
0.0063
0.0072
0.0074
0.0086
0.0095
0.0099
0.025
0.035
0.026
0.017
0.036
0.045
0.054
0.043
0.034
0.075
0.086
0.075
0.098
0.087
0.058
0.036
0.054
0.085
0.076
0.057
0.068
0.076
0.024
0.045
0.0031
0.0032
0.0033
0.0044
0.0045
0.0046
0.0048
0.0049
0.035
0.034
0.057
0.065
0.034
0.036
0.055
0.046
0.054
0.023
0.014
0.023
0.054
0.067
0.056
0.043
0.067
0.067
0.089
0.087
0.053
0.045
0.045
0.034
0.065
0.064
0.076
0.085
0.087
0.091
0.096
0.099
0.049
0.058
0.064
0.075
0.076
0.087
0.098
0.042
0.053
0.064
0.065
0.066
0.067
0.068
0.0013
0.0022
0.0035
0.0024
0.0043
0.0056
0.0065
0.0057
0.0078
0.0069
0.0058
0.0046
0.0065
0.0054
0.0043
0.0054
0.0065
0.0074
0.0085
0.0095
0.0086
0.0077
0.0066
0.0055
0.0023
0.0034
0.0046
0.0035
0.0027
0.0038
0.0057
0.0066
0.0075
0.0084
0.0093
0.0084
0.0095
0.0076
0.0047
0.0056
0.0035
0.0043
0.0054
0.0065
0.0076
0.0047
0.0035
0.0023
0.0019
0.0028
0.0037
0.0046
0.0059
0.0048
0.0034
0.0023
0.0072
0.0064
0.0096
0.0087
0.0096
0.0085
0.0053
0.0044
0.0065
0.0076
0.0057
0.0038
0.0049
0.0059
0.0048
0.0038
0.003
0.004
0.005
0.004
0.005
0.004
a
The average IC50 values were calculated from three independent tests.
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observed in glioblastoma33 and neuroblastoma.34 Moreover,
dysregulated signaling through RAF kinase isoforms has been
detected in ꢂ30% of human cancers where wild type c-RAF is
hyperactivated in a wide range of human solid tumors.35
The kinase inhibitory activities (nM) of the targeted quina-
zolinones; 8a–l, 10a–h, (13–16, 18)a,b and 17a–f and N-(2,4-
disubstituted phenyl)pyridinecarboxamides; 9a–n and 12a–h
against ABL kinase (100% inhibited) are listed in Table 4.
Interestingly, the target quinazolinones; 8a–l, 10a–h and 13–
18 as well as the substituted N-(2,4-disubstituted phenyl)pyr-
idinecarboxamides; 9a–n and 12a–h exhibited a wide range of
ABL, ALK and c-RAF inhibitory activity with IC50 ranges 0.011–
76 nM and 0.24–55 nM, 0.05–323 nM and 0.09–93 nM, 0.042–
66 nM and 0.1–79 nM, respectively. The quinazolinyl-3-urea/
thiourea derivatives 8a–l and the pyridinecarboxamides with
benzylidene-carrying side chain 9a–n displayed moderate to
very weak inhibitory activity against ABL (IC50 ¼ 17–76 nM), ALK
(IC50 ¼ 19–323 nM) and c-RAF kinases (IC50 ¼ 24 nM-inactive).
The pyridinecarboxamides bearing acetyl hydrazinyl-carrying
side chain 12a–h and the 3-substituted quinazolin-4(3H)-one
13–18 had higher potency against ABL, ALK and c-RAF kinases
(IC50 ¼ 0.24–0.79 nM, 0.09–0.89 nM and 0.1–0.97, respectively).
Additionally, the 3-substituted benzylidene quinazolinones
10a–h showed exceptional ABL, ALK and c-RAF inhibitory
potency (IC50 ¼ 0.011–0.037 nM, 0.05–0.074 nM, 0.042–
0.057 nM, respectively).
Table 2 In vitro cytotoxicity of derivatives 10a–h, 12b and 12f against
MCF-10A
Cpd no. IC50 (mM) Selectivityb Cpd no. IC50 (mM) Selectivity
10a
10b
10c
10d
10e
10f
12.09
19.21
2.10
3.20
1.98
2.30
2.10
3.30
>26 866
>33 701
>2692
>3764
>2275
>2421
>2164
>3333
12a
12b
12c
12d
12e
12f
NTa
1.23
NT
NT
NT
4.30
NT
NT
NT
>223
NT
NT
NT
>518
NT
NT
10g
10h
12g
12h
a
b
NT: not tested. Selectivity in fold relative to the lowest observed
potency among tested breast cancer cell lines.
activity percent inhibition. The synthesized compounds exerted
multiple inhibitions over the tested oncogenic protein kinases
at the tested concentration. The synthesized compounds
completely inhibited the enzymatic activity of ABL, ALK and c-
RAF kinases. On the other hand, the inhibitions were of low
percent in the other tested 27 kinases.
For those kinases that were 100% inhibited at the initial high
dose, viz. ABL, ALK and c-RAF, IC50 values of the test
compounds were determined. Kinase activity was assessed
using HotSpotSM Technology, a miniaturized radio-isotope
based lter binding assay.29
Concerning the quinazolinone derivatives 8a–l: generally,
replacement of the pyridin-3-yl moiety (8a–f) by pyridin-4-yl
moiety (8g–l) caused a slight increase in ABL inhibitory
activity and it did not have a signicant effect on ALK inhibitory
activity. On the other hand, it nearly abolished c-RAF activity.
Surprisingly, side chain shortening of the phenylureido
Abelson kinase (ABL) is a fusion tyrosine kinase playing
a role in 90% of chronic myeloid leukemia (CML) cases.30
Anaplastic Lymphoma Kinase (ALK) is a member of the insulin
receptor tyrosine kinase family.31 It is constitutively active and
plays an oncogenic role in 70–80% of all anaplastic large cell
lymphomas.32 On the other hand, over-expression of ALK was
Table 3 MCF-7 mouse xenograft model of breast cancer for compounds 10a–h and 12a–h
Tumor growth Vt/Vo for compounds at timea (days)
Compound no.
0
2
4
6
8
10
12
14
16
18
20
Vehicle control 1.0 ꢃ 0.01b 1.3 ꢃ 0.07 1.8 ꢃ 0.07 4.7 ꢃ 0.2 9.8 ꢃ 0.5 12.6 ꢃ 1.4 24.7 ꢃ 2.7 27.6 ꢃ 1.2 29.0 ꢃ 1.7 38.9 ꢃ 1.2 40.2 ꢃ 1.8
Sorafenib
10a
10b
10c
10d
10e
10f
10g
10h
12a
12b
12c
12d
12e
12f
1.0 ꢃ 0.04 1.1 ꢃ 0.06 1.2 ꢃ 0.06 1.5 ꢃ 0.02 2.1 ꢃ 0.2 2.2 ꢃ 0.1 4.5 ꢃ 0.2 5.5 ꢃ 0.3 6.89 ꢃ 0.5 8.8 ꢃ 0.7 9.1 ꢃ 0.3
1.0 ꢃ 0.05 1.0 ꢃ 0.08 1.0 ꢃ 0.07 1.1 ꢃ 0.01 1.1 ꢃ 0.1 1.3 ꢃ 0.2 2.0 ꢃ 0.3 2.6 ꢃ 0.4 3.10 ꢃ 0.4 3.3 ꢃ 0.3 3.5 ꢃ 0.2
1.0 ꢃ 0.03 1.0 ꢃ 0.09 1.0 ꢃ 0.08 1.1 ꢃ 0.02 1.3 ꢃ 0.1 1.4 ꢃ 0.1 2.1 ꢃ 0.4 2.7 ꢃ 0.5 3.16 ꢃ 0.5 3.3 ꢃ 0.2 3.8 ꢃ 0.2
1.0 ꢃ 0.05 1.0 ꢃ 0.08 1.0 ꢃ 0.07 1.1 ꢃ 0.03 1.4 ꢃ 0.1 1.4 ꢃ 0.1 2.2 ꢃ 0.3 2.8 ꢃ 0.3 3.32 ꢃ 0.6 4.1 ꢃ 0.3 4.5 ꢃ 0.1
1.0 ꢃ 0.06 1.0 ꢃ 0.07 1.0 ꢃ 0.08 1.1 ꢃ 0.04 1.5 ꢃ 0.2 1.4 ꢃ 0.2 2.3 ꢃ 0.2 2.9 ꢃ 0.2 3.52 ꢃ 0.4 4.2 ꢃ 0.4 4.8 ꢃ 0.4
1.0 ꢃ 0.07 1.0 ꢃ 0.06 1.0 ꢃ 0.09 1.1 ꢃ 0.05 1.6 ꢃ 0.2 1.6 ꢃ 0.1 2.3 ꢃ 0.3 3.0 ꢃ 0.3 3.74 ꢃ 0.3 4.5 ꢃ 0.3 4.9 ꢃ 0.5
1.0 ꢃ 0.06 1.0 ꢃ 0.07 1.0 ꢃ 0.08 1.2 ꢃ 0.04 1.6 ꢃ 0.1 1.7 ꢃ 0.1 2.5 ꢃ 0.2 3.1 ꢃ 0.4 3.90 ꢃ 0.4 4.6 ꢃ 0.5 5.0 ꢃ 0.4
1.0 ꢃ 0.07 1.0 ꢃ 0.06 1.0 ꢃ 0.07 1.2 ꢃ 0.06 1.7 ꢃ 0.2 1.8 ꢃ 0.1 2.6 ꢃ 0.3 3.2 ꢃ 0.5 4.04 ꢃ 0.2 4.7 ꢃ 0.6 5.0 ꢃ 0.5
1.0 ꢃ 0.06 1.0 ꢃ 0.07 1.1 ꢃ 0.08 1.2 ꢃ 0.05 1.8 ꢃ 0.1 1.9 ꢃ 0.1 2.7 ꢃ 0.2 3.5 ꢃ 0.3 4.19 ꢃ 0.5 4.8 ꢃ 0.7 5.1 ꢃ 0.3
1.0 ꢃ 0.07 1.0 ꢃ 0.06 1.1 ꢃ 0.07 1.2 ꢃ 0.06 1.5 ꢃ 0.1 1.7 ꢃ 0.1 2.0 ꢃ 0.3 2.5 ꢃ 0.2 2.98 ꢃ 0.1 3.5 ꢃ 0.8 4.3 ꢃ 0.2
1.0 ꢃ 0.06 1.0 ꢃ 0.07 1.1 ꢃ 0.06 1.2 ꢃ 0.07 1.6 ꢃ 0.1 1.8 ꢃ 0.2 2.1 ꢃ 0.2 2.6 ꢃ 0.2 3.10 ꢃ 0.2 4.8 ꢃ 0.8 5.4 ꢃ 0.6
1.0 ꢃ 0.05 1.0 ꢃ 0.06 1.1 ꢃ 0.06 1.2 ꢃ 0.08 1.7 ꢃ 0.2 1.9 ꢃ 0.2 2.2 ꢃ 0.1 2.7 ꢃ 0.3 3.34 ꢃ 0.4 4.3 ꢃ 0.7 5.8 ꢃ 0.5
1.0 ꢃ 0.06 1.1 ꢃ 0.05 1.1 ꢃ 0.04 1.3 ꢃ 0.07 1.8 ꢃ 0.1 1.9 ꢃ 0.2 2.3 ꢃ 0.2 2.8 ꢃ 0.1 3.56 ꢃ 0.3 4.6 ꢃ 0.4 6.3 ꢃ 0.6
1.0 ꢃ 0.05 1.1 ꢃ 0.04 1.2 ꢃ 0.03 1.3 ꢃ 0.03 1.8 ꢃ 0.1 2.0 ꢃ 0.3 2.5 ꢃ 0.1 3.2 ꢃ 0.2 4.11 ꢃ 0.5 5.3 ꢃ 0.5 6.5 ꢃ 0.7
1.0 ꢃ 0.04 1.1 ꢃ 0.06 1.2 ꢃ 0.04 1.4 ꢃ 0.04 1.9 ꢃ 0.2 2.1 ꢃ 0.2 3.0 ꢃ 0.4 3.6 ꢃ 0.3 4.48 ꢃ 0.4 5.6 ꢃ 0.6 6.8 ꢃ 0.8
1.0 ꢃ 0.03 1.1 ꢃ 0.07 1.2 ꢃ 0.05 1.4 ꢃ 0.05 1.9 ꢃ 0.1 2.1 ꢃ 0.3 3.2 ꢃ 0.3 4.2 ꢃ 0.4 5.23 ꢃ 0.5 5.9 ꢃ 0.7 7.1 ꢃ 0.7
1.0 ꢃ 0.04 1.1 ꢃ 0.06 1.3 ꢃ 0.06 1.5 ꢃ 0.06 2.0 ꢃ 0.2 2.2 ꢃ 0.2 3.5 ꢃ 0.2 4.5 ꢃ 0.5 5.67 ꢃ 0.7 6.1 ꢃ 0.8 8.1 ꢃ 0.9
12g
12h
a
Tested compounds and the control drug sorafenib were administered by intraperitoneal (i.p.) injection at a dose of 5 mg kgꢁ1, every 48 h for 3
weeks. b Each data point is the mean ꢃ SEM of ve animals.
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Table 4 Kinase inhibitory activity
IC50 (nM)
ABL
IC50 (nM)
ABL
Compound number
ALK
c-RAF
Compound number
ALK
c-Raf
8a
8b
8c
8d
8e
8f
8g
8h
8i
8j
8k
8l
9a
9b
9c
9d
9e
9f
9g
9h
9i
9j
9k
9l
9m
9n
10a
10b
54
43
66
76
68
43
44
33
26
32
47
55
17
24
35
39
48
26
20
36
18
21
46
39
40
55
0.011
0.021
0.022
0.58
—
64
72
21
35
76
56
37
25
323
34
47
50
84
93
65
46
54
75
37
59
28
30
58
19
20
64
53
62
43
34
55
Inactive
Inactive
Inactive
Inactive
66
Inactive
36
43
24
56
35
45
67
40
58
79
36
47
68
25
0.042
0.044
0.045
0.99
—
10d
10e
10f
10g
10h
12a
12b
12c
12d
12e
12f
12g
12h
13a
13b
14a
14b
15a
15b
16a
16b
17a
17b
17c
17d
17e
17f
0.023
0.035
0.037
0.036
0.037
0.26
0.35
0.44
0.33
0.24
0.44
0.35
0.26
0.57
0.46
0.65
0.74
0.65
0.54
0.45
0.56
0.65
0.76
0.67
0.79
0.60
0.59
0.46
0.37
0.03
—
0.065
0.067
0.074
0.076
0.078
0.35
0.43
0.65
0.78
0.76
0.45
0.24
0.35
0.35
0.43
0.54
0.32
0.67
0.89
0.09
0.76
0.45
0.34
0.56
0.54
0.35
0.34
0.56
0.78
—
0.046
0.057
0.057
0.058
0.059
0.55
0.44
0.35
0.26
0.19
0.18
0.10
0.17
0.14
0.13
0.45
0.36
0.48
0.56
0.44
0.55
0.76
0.84
0.95
0.86
0.97
0.85
0.76
0.65
—
67
0.050
0.056
0.068
—
18a
10c
18b
Bosutinib
Sorafenib
Imatinib
Staurosporine
0.0022
—
6
derivatives 8d, j to phenyl amino ones 17a, d enhanced the case of ABL), (IC50 ¼ 0.050, 0.056, 0.068 nM, respectively in case
inhibitory potency by 50–100 fold. Looking at the effect of of ALK), (IC50 ¼ 0.042, 0.044, 0.045 nM, respectively in case of c-
removing the nitrogen spacer between the quinazolinone core RAF). Similar results were observed for the 4-pyridyl derivatives.
and the side chain linked to the quinazolinyl-N3, it was found It is worth to mention that the uoro-3-pyridyl derivative 10a
that this structural modication could be benecial for the ABL, exhibited the most promising kinase inhibitory activity in this
ALK and c-RAF kinases inhibitory activity (IC50 ¼ 0.37–0.79, study.
0.09–0.89 and 0.13–0.97 nM, respectively) in the quinazolinones
Finally, as evidenced from the experimental data showed,
13, 14, 15, 16, 18a, b. With respect to the pyridinecarboxamides the most active kinase activity inhibiting series in this study
with benzylidene-carrying side chain 9a–n, it was observed that 10a–h were equi- to many folds more potent than the reference
this series displayed slightly higher activity than the quinazo- kinase inhibitors against their target kinases viz. imatinib (ABL
linones 8a–l.
and c-RAF), bosutinib (ABL), sorafenib (c-RAF) and staur-
Rigidication of the Schiff's bases 9a–n through cyclo- osporine (ALK).
dehydration afforded the most potent 3-substituted quinazoli-
nones in this work 10a–h which showed exceptional enzyme
(2) Molecular modelling
inhibitory potency against ABL, ALK and c-RAF (IC50 ¼ 0.011–
0.037 nM, 0.05–0.074 nM, 0.042–0.057 nM, respectively) which
is consistent with their antitumor activity. Exploring the effect
of different substituents on the benzylidene moiety, revealed
that decreasing the size and increasing the electronegativity of
the halogenated substituent resulted in improving the potency
in case of 3-pyridyl analogues. Therefore, the uoro derivative
10a displayed higher activity than the chloro analogue 10b
which in turn exhibited slightly higher activity than the bromo
derivative 10c (IC50 ¼ 0.011, 0.021, 0.022 nM, respectively in
The docking study was initiated to predict the interaction of the
newly synthesized compounds against the kinase ABL towards
which they showed the highest enzymatic activity inhibition.
The two PDB les 2HYY and 3UE4 representing inactive DFG-
out conformations of ABL-co-crystallized with imatinib and
bosutinib, respectively.36,37 It is reported that the two confor-
mations are distinct in the sense that the in the latter crystal
structure of ABL with its inhibitor bosutinib, the activation loop
adopts a conformation like that observed in active kinases,
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except for the conformation of the DFG motif itself. This is in 3UE4, quinazoline N and Met318 amide NH hydrogen bond of
contrast with the commonly known conformational orientation 1.9 A, S ¼ ꢁ18.42 kcal molꢁ1) gave rise to complexes with
˚
of the activation loop seen in case of the type II inhibitor ima- comparable binding modes and energetics to that of the co-
tinib (PDB entry 2HYY) in which the P-loop folds over the ATP crystallized ligand bosutinib. Results displayed herein are
binding site, and the activation-loop blocks the substrate those obtained for attempted docking of the most active
binding site with a shi of the DFG motif nearer to the front of compound 10a (IC50 ¼ 0.011 nM) where Fig. 2 shows the
the active site.
respective 2D and 3D interaction contacts of the top scoring and
Docking of the most active compounds 10a–h and 12a–h was best tting pose obtained through this molecular modeling
performed on both crystal–inhibitor complexes to predict study. The highest ABL inhibitory activity of the Schiff's base
whether these derivatives elicit their activity by binding to the derivative 10a might be attributed to high ABL binding, energy
active pocket with a behavior compared to that of imitanib or score (S ¼ ꢁ21.22 kcal molꢁ1) where the 3D interactions (Fig. 2:
bosutinib. Results showed that while the type II inhibitor ima- bottom panel) showed that the pyridyl N atom formed
tinib displayed a network of hydrogen bonds to anchor itself a hydrogen bond with the amide NH of the gatekeeper Met318
˚
tightly with the ABL binding cle residues, all attempts to dock (2.4 A). It is noteworthy that this interaction is considered the
the new chemotypes in the active site of ABL DFG-out confor- key contributor to the overall strength of drug–ABL kinase
mation were unsuccessful due to multiple steric clashes with complexes.38–40 Additionally, the carbonyl O of the quinazolin-4-
the active site residues (data not shown). On the other hand, one ring interacted with the amidic NH of the Asn322 through
˚
docking calculations based on the ABL kinase domain confor- a hydrogen (2.1 A). These results along with the other obtained
mation induced by binding of bosutinib to ABL (PDB entry but not displayed ones strongly suggest that our compounds
probably act as type II kinase inhibitors by binding to the DFG
out conformation of the catalytic domain of ABL.
Conclusion
The present study reports the design and synthesis of ve novel
series of 6-iodo-2-(pyridin-3/4-yl)-3-substituted quinazolin-
4(3H)-one and acyclic analogues as multi-kinase inhibitors. The
design was based on structural analogy between the new
compounds, reported quinazoline derivatives and acyclic
analogues. Synthesis of the target compounds was carried out
employing techniques reported in the literature with some
modications when necessary. This led to the synthesis of 8 new
intermediates and 58 new nal compounds. The structures of
the synthesized compounds were conrmed by various spectral
analysis techniques as well as the elemental analyses.
The target compounds were evaluated for their antitumor
activity against a panel of nine cancer cell lines. This panel
involved breast cancers (MCF-7, MDAMB-231, MDAMB-435 and
HS-578T), colon cancers (HT-29 and HCC-2998) and leukemia
cell lines (CCRF-CEM, K-562 and HL-60). The biological results
revealed that target quinazolinones 8a–l, 10a–h, 13a, b, 14a, b,
15a, b, 16a, b, 17a–f and 18a, b, and N-(2,4-disubstituted phenyl)
pyridinecarboxamides 9a–n and 12a–h exhibited potent anti-
tumor activity. Aiming to investigate their mechanism of action,
the synthesized were tested over a panel of 30 kinases; ABL, ALK
and c-RAF were the most responsive kinases. Molecular docking
was carried out to predict the binding mode of these chemo-
types with the active site amino acids of the ABL, being the most
responsive kinase to the inhibitory effect of the tested
compounds. Molecular modeling results showed that these
derivatives recognize the active pocket of the target enzyme
selectively to a more relaxed conformation of the ABL active site
as induced by the known inhibitor bosutinib. In conclusion, the
synthesized compounds, especially 10a, can be viewed as
promising leads for further investigation and optimization as
multi-kinase inhibitors.
Fig. 2 Top panel: 2D interactions; bottom panel: 3D interactions of
10a and ABL (PDB entry 3UE4).
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6-Iodo-2-(pyridin-3-yl)-4H-benzo[d][1,3]oxazin-4-one (6a)
Method 1. A mixture of the amidated anthranilic acid 5a
Experimental section
Melting points were determined on Electrothermal Stuart SMP3 (1.84 g, 5 mmol) in acetic anhydride (15 mL) was heated under
digital melting point apparatus and were uncorrected. reux for 5 h. Then the reaction mixture was poured onto ice-
Elemental microanalyses were performed at the Regional water. The obtained beige solid product was collected by
Center for Mycology and Biotechnology, Al-Azhar University and ltration, dried to give 1.4 g (84%) and then recrystallized from
at the Microanalytical Centre, Faculty of Science, Cairo methanol. Mp: 167–169 C. Yield: 84%. I.R. (KBr, cmꢁ1): ~nmax
ꢀ
University. Infrared spectra were determined (KBr) using Shi- 3049 (CH aromatic), 1753 (C]O), 1626 (C]N), 1585 (C]C), 530
madzu Infrared spectrometer (IR-435) and FT-IR 1650 (Perkin (C–I). 1H-NMR 400 MHz (DMSO-d6, ppm): d 7.53 (d, 1H, J ¼
Elmer), Faculty of Pharmacy, Cairo University. 1H-NMR spectra 8.44 Hz, phenyl-H), 7.64 (dd, 1H, J ¼ 4.84, 8.04 Hz, pyridyl-H),
were performed in CDCl3 or DMSO-d6 using Joel, 300 MHz NMR 8.27 (dd, 1H, J ¼ 2.00, 8.44 Hz, phenyl-H), 8.42 (d, 1H, J ¼
spectrometer, Faculty of Science, Cairo University, and Bruker, 1.96 Hz, phenyl-H), 8.49 (t, 1H, J ¼ 8.04 Hz, pyridyl-H), 8.83 (dd,
400 MHz NMR spectrometer, Microanalytical unit, Faculty of 1H, J ¼ 1.60, 4.80 Hz, pyridyl-H), 9.30 (d, 1H, J ¼ 2.20 Hz,
Pharmacy, Cairo University. 13C-NMR spectra were recorded pyridyl-H). Mass (m/z, rel. abundance): 350 (M+, 100%). Anal.
using Bruker, 100 MHz NMR spectrometer, Microanalytical calcd for C13H7IN2O2 (350.11): C, 44.60; H, 2.02; N, 8.00. Found:
unit, Faculty of Pharmacy, Cairo University. Mass spectra were C, 44.92; H, 1.85; N, 8.08.
carried out using Finnigan SSQ 7000 Gas chromatograph-Mass,
Method 2. A solution of 5-iodoanthranilic acid (2.63 g, 10
at the Microanalytical Centre, Faculty of Science, Cairo mmol) in dry pyridine (30 mL) was added to the freshly prepared
University. Thin layer chromatography was carried out on silica isonicotinoyl chloride hydrochloride (2.13 g, 12 mmol) and the
gel TLC plates with uorescence indicator (F254). 5-Iodoan- mixture was stirred at 5 ꢀC for 2 h, and then allowed to warm to
thranilic acid (3),26 nicotinoyl chloride (4a),27 isonicotinoyl room temperature. The reaction mixture was poured onto ice-
chloride (4b)28 were prepared according to the reported method. water, the solid obtained was ltered, washed with diluted
acetic acid, dried and recrystallized from methanol. Mp: 167–
169 C. Yield: 65%. I.R. (KBr, cmꢁ1): ~nmax 3049 (CH aromatic),
ꢀ
(1) Chemistry
1753 (C]O), 1626 (C]N), 1585 (C]C), 530 (C–I).
5-Iodo-2-[(pyridin-3-ylcarbonyl)amino]benzoic acid (5a). A
solution of 5-iodoanthranilic acid 3 (2.63 g, 10 mmol) in dry
dimethylformamide (20 mL) was added to the freshly prepared
nicotinoyl chloride hydrochloride (2.13 g, 12 mmol) and the
6-Iodo-2-(pyridin-4-yl)-4H-benzo[d][1,3]oxazin-4-one (6b)
Method 1. This compound was prepared according to method
1 used for preparation of 6a but using 5b as a starting material
and was also recrystallized from methanol.
Mp: 185–187 ꢀC. Yield: 75%. I.R. (KBr, cmꢁ1): ~nmax 3057 (CH
aromatic), 1764 (C]O), 1616 (C]N), 1556 (C]C), 534 (C–I). 1H-
NMR 400 MHz (DMSO-d6, ppm): d 7.50 (d, 1H, J ¼ 8.40 Hz,
phenyl-H), 7.99 (d, 2H, J ¼ 6.04 Hz, pyridyl-H), 8.23 (dd, 1H, J ¼
2.00, 8.40 Hz, phenyl-H), 8.37 (d, 1H, J ¼ 1.88 Hz, phenyl-H),
8.78 (d, 2H, J ¼ 6.04 Hz, pyridyl-H). Mass (m/z, rel. abun-
dance): 350 (M+, 100%). Anal. calcd for C13H7IN2O2 (350.11): C,
44.60; H, 2.02; N, 8.00. Found: C, 44.54; H, 1.81; N, 8.09.
Method 2. Method 2 used for preparation of 6a was also used
for synthesis of 6b using 5b as a starting material.
ꢀ
mixture was heated at 80 C for 2 h, and then allowed to cool.
The reaction mixture was poured onto ice-water. The solid ob-
tained was ltered, washed several times with water and dried
to give 3.2 g (71%) which was recrystallized from acetic acid.
Mp: 314–316 ꢀC. I.R. (KBr, cmꢁ1): ~nmax 3435–3414 (br.) (OH),
3338 (NH), 1676, 1664 (2C]O), 1608 (C]N), 1519 (C]C), 530
(C–I). 1H-NMR 300 MHz (DMSO-d6, D2O ppm): d 7.66 (dd, 1H, J
¼ 4.80, 7.80 Hz, pyridyl-H), 8.00 (dd, 1H, J ¼ 2.10, 8.70 Hz,
phenyl-H), 8.29 (t, 1H, J ¼ 2.40 Hz, pyridyl-H), 8.32 (dd, 1H, J ¼
1.80, 6.30 Hz, phenyl-H), 8.40 (d, 1H, J ¼ 8.70 Hz, phenyl-H),
8.83 (dd, 1H, J ¼ 1.50, 4.80 Hz, pyridyl-H), 9.11 (d, 1H, J ¼
1.80 Hz, pyridyl-H), 9.30 (s, 1H, NH exchanged by D2O), 12.02 (s,
1H, OH exchanged by D2O). Mass (m/z, rel. abundance): 368
(M+, 44.38%), 106 (100%). Anal. calcd for C13H9IN2O3 (368.13):
C, 42.41; H, 2.46; N, 7.61. Found: C, 42.64; H, 2.45; N, 7.70.
5-Iodo-2-[(pyridin-4-ylcarbonyl)amino]benzoic acid (5b).
Compound 5b was prepared and puried by adopting the same
procedure used for 5a using isonicotinoyl chloride. Mp: 290–
292 ꢀC. I.R. (KBr, cmꢁ1): ~nmax 3232 (OH), 3145 (NH), 1683, 1653
(2C]O), 1595 (C]N), 1573 (C]C), 526 (C–I). 1H-NMR 300 MHz
(DMSO-d6, D2O ppm): d 7.96 (d, 2H, J ¼ 4.80 Hz, pyridyl-H), 7.99
(dd, 1H, J ¼ 6.00, 8.10 Hz, phenyl-H), 8.29 (d, 1H, J ¼ 1.80 Hz,
phenyl-H), 8.38 (dd, 1H, J ¼ 3.00, 8.70 Hz, phenyl-H), 8.92 (d,
2H, J ¼ 4.80 Hz, pyridyl-H), 10.82 (s, 1H, NH exchanged by D2O),
12.11 (s, 1H, OH exchanged by D2O). Mass (m/z, rel. abundance):
368 (M+, 34.7%), 106 (100%). Anal. calcd for C13H9IN2O3
(368.13): C, 42.41; H, 2.46; N, 7.61. Found: C, 42.57; H, 2.48; N,
7.73.
Mp: 185–187 ꢀC. Yield: 55%. I.R. (KBr, cmꢁ1): ~nmax 3057 (CH
aromatic), 1764 (C]O), 1616 (C]N), 1556 (C]C), 534 (C–I).
N-[2-(Hydrazinylcarbonyl)-4-iodophenyl]pyridine-3-
carboxamide (7a). A mixture of benzoxazinone 6a (1.74 g, 5
mmol) and hydrazine hydrate 99% (15 mmol, 0.8 mL) in abso-
lute ethanol (20 mL) was reuxed for 4 h, le to cool then
ltered. The separated white solid was collected, washed with
ꢀ
ethanol and recrystallized from ethyl acetate. Mp: 221–223 C.
Yield: 75%. I.R. (KBr, cmꢁ1): ~nmax 3309 (2NH), 3213, 3151 (NH2),
3061 (CH aromatic), 1697 (2C]O), 1631 (C]N), 1516 (C]C),
530 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 4.70 (s, 2H,
NH2 exchanged by D2O), 7.62 (dd, 1H, J ¼ 4.88, 7.96 Hz, pyridyl-
H), 7.90 (dd, 1H, J ¼ 1.32, 8.80 Hz, phenyl-H), 8.11 (s, 1H,
phenyl-H), 8.26 (t, 1H, J ¼ 8.04 Hz, pyridyl-H), 8.39 (d, 1H, J ¼
8.8 Hz, phenyl-H), 8.80 (dd, 1H, J ¼ 1.52, 4.80 Hz, pyridyl-H),
9.10 (d, 1H, J ¼ 1.84 Hz, pyridyl-H), 10.29 (s, 1H, NH
exchanged by D2O), 12.49 (s, 1H, NH exchanged by D2O). Mass
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(m/z, rel. abundance): 382 (M+, 1.63%), 216 (100%). Anal. calcd 5CH2), 4.06 (m, 1H, cyclohexyl-CH), 7.52 (t, 1H, J ¼ 6.3 Hz,
for C13H11IN4O2 (382.16): C, 40.86; H, 2.90; N, 14.66. Found: C, pyridyl-H), 7.59 (dd, 1H, J ¼ 4.8, 8.1 Hz, quinazolinyl-H), 7.95 (s,
40.98; H, 2.96; N, 14.89.
1H, quinazolinyl-H), 8.14 (d, 1H, J ¼ 6.9 Hz, pyridyl-H), 8.21 (d,
N-[2-(Hydrazinylcarbonyl)-4-iodophenyl]pyridine-4-carbox- 1H, J ¼ 8.4 Hz, quinazolinyl-H), 8.79 (d, 1H, J ¼ 3.3 Hz, pyridyl-
amide (7b). Compound 7b was prepared and puried by using H), 9.04 (s, 1H, pyridyl-H), 10.59 (s, 1H, NH exchanged by D2O),
the same procedure adopted for the preparation of 7a but using 11.5 (s, 1H, NH exchanged by D2O). Mass (m/z, rel. abundance):
6b as a starting material. Mp: 238–240 ꢀC. Yield: 75%. I.R. (KBr, 505.3 (M+, 3.49%), 106.05 (100%). Anal. calcd for C20H20IN5OS
cmꢁ1): ~nmax 3250 (2NH), 3165, 3130 (NH2), 3074 (CH aromatic), (505.38): C, 47.53; H, 3.99; N, 13.86. Found: C, 47.49; H, 4.17; N,
1681 (C]O), 1638 (C]N), 1517–1434 (C]C), 530 (C–I). 1H-NMR 14.11.
400 MHz (DMSO-d6, D2O, ppm): d 4.72 (s, 2H, NH2 exchanged by
1-(6-Iodo-4-oxo-2-(pyridin-3-yl)quinazolin-3(4H)-yl)-3-phenylurea
D2O), 7.82 (d, 2H, J ¼ 5.96 Hz, pyridyl-H), 7.91 (dd, 1H, J ¼ 1.56, (8d). Mp: 230–232 ꢀC. Yield: 89% I.R. (KBr, cmꢁ1): ~nmax 3296
8.76 Hz, phenyl-H), 8.13 (s, 1H, phenyl-H), 8.40 (d, 1H, J ¼ (2NH), 3101 (CH aromatic), 1683, 1668 (2C]O), 1639 (C]N),
8.76 Hz, phenyl-H), 8.85 (d, 2H, J ¼ 5.96 Hz, pyridyl-H), 10.32 (s, 1506–1446 (C]C), 530 (C–I). 1H-NMR 300 MHz (DMSO-d6, D2O,
1H, NH exchanged by D2O), 12.60 (s, 1H, NH exchanged by ppm): d 6.99 (t, 1H, J ¼ 7.5 Hz, phenyl-H), 7.25 (t, 2H, J ¼ 7.5 Hz,
D2O). Anal. calcd for C13H11IN4O2 (382.16): C, 40.86; H, 2.90; N, phenyl-H), 7.45 (d, 2H, J ¼ 8.4 Hz, phenyl-H), 7.57 (dd, 1H, J ¼
14.66. Found: C, 40.98; H, 2.93; N, 14.82.
5.1 Hz, 7.8 Hz, pyridyl-H), 7.97 (dd, 1H, J ¼ 2.1 Hz, 9.00 Hz,
1-(6-Iodo-4-oxo-2-(pyridin-3-yl)
quinazolin-3(4H)-yl)-3- quinazolinyl-H), 8.18–8.24 (m, 2H, pyridyl-H, quinazolinyl-H),
substituted urea/thiourea derivatives (8a–f)
8.78 (d, 1H, J ¼ 4.8 Hz, pyridyl-H), 8.88 (s, 1H, pyridyl-H), 9.06
General method. A mixture of the hydrazido derivative 7a (d, 1H, J ¼ 2.1 Hz, quinazolinyl-H), 10.66 (s, 1H, NH exchanged by
(0.72 g, 2 mmol) and the appropriate alkyl or aryl isocyanate (2.2 D2O), 11.73 (s, 1H, NH exchanged by D2O). 13C-NMR (100 MHz
mmol) or isothiocyanate (3 mmol) in dioxane (20 mL) was DMSO-d6, ppm): d 88.05, 119.22, 122.60, 123.67, 123.88, 124.30
reuxed for 6–10 h. Then allowed to cool. The precipitated solid (phenyl C-2, C-6), 129.09, 130.35, 135.40, 137.07, 138.40, 139.88,
product was collected by ltration, washed with aqueous 141.27, 148.77, 153.16, 155.80, 163.98 (C]O), 167.71 (C]O).
ethanol then dried and recrystallized from absolute ethanol to Mass (m/z, rel. abundance): 483 (M+, 3.49%), 106.05 (100%). Anal.
give 8a–f.
calcd for C20H14IN5O2 (483.26): C, 49.71; H, 2.92; N, 14.49. Found:
1-Ethyl-3-(6-iodo-4-oxo-2-(pyridin-3-yl)quinazolin-3(4H)-yl)thio- C, 49.61; H, 2.66; N, 14.70.
urea (8a). Mp: 210–211 ꢀC. Yield: 85%. I.R. (KBr, cmꢁ1): ~nmax
1-(6-Iodo-4-oxo-2-(pyridin-3-yl)quinazolin-3(4H)-yl)-3-phenylthiourea
3300, 3251 (2NH), 3215 (CH aromatic), 2972, 2926 (CH2, CH3), (8e). Mp: 270 C. Yield: 85%. I.R. (KBr, cmꢁ1): ~nmax 3246 (NH),
1681 (C]O), 1654 (C]N), 1508 (C]C), 1215 (C]S), 530 (C–I). 3062 (CH aromatic), 1681 (C]O), 1635 (C]N), 1521–1483 (C]
ꢀ
1H-NMR 300 MHz (DMSO-d6, D2O, ppm): d 1.02 (t, 3H, J ¼ C), 1180 (C]S), 530 (C–I). H-NMR 300 MHz (DMSO-d6, D2O,
1
6.90 Hz, CH3CH2), 3.45 (q, 2H, J ¼ 6.90 Hz, CH3CH2), 7.62 (dd, ppm): d 7.05 (t, 1H, J ¼ 6.9 Hz, phenyl-H), 7.38 (t, 2H, J ¼ 6.9 Hz,
1H, J ¼ 4.80, 8.10 Hz, pyridyl-H), 7.98 (d, J ¼ 2.10 Hz, quina- phenyl-H), 7.61–7.70 (m, 3H, phenyl-2H, pyridyl-H), 7.95 (d, 1H, J
zolinyl), 8.13–8.24 (m, 3H, quinazolinyl-H, pyridyl-H), 8.81 (dd, ¼ 8.70 Hz, quinazolinyl-H), 8.11 (s, 1H, quinazolinyl-H), 8.36 (d,
1H, J ¼ 1.8, 4.80 Hz, 1H, pyridyl-H), 9.06 (d, 1H, J ¼ 2.40 Hz, 1H, J ¼ 7.80 Hz, quinazolinyl-H), 8.46 (d, 1H, J ¼ 8.70 Hz, pyridyl-
pyridyl-H), 10.7 (s, 1H, NH exchanged by D2O), 11.68 (s, 1H, NH H), 8.85 (d, 1H, J ¼ 4.2 Hz, pyridyl-H), 9.22 (s, 1H, pyridyl-H),
exchanged by D2O). Mass (m/z, rel. abundance): 451 (M+, 10.83 (s, 1H, NH exchanged by D2O), 11.48 (s, 1H, NH
53.61%), 171 (100%). Anal. calcd for C16H14IN5OS (451.28): C, exchanged by D2O). Mass (m/z, rel. abundance): 499 (M+, 3.49%),
42.58; H, 3.13; N, 15.52. Found: C, 42.33; H, 3.45; N, 15.26.
106.05 (100%). Anal. calcd for C20H14IN5OS (499.33): C, 48.11; H,
1-Cyclohexyl-3-(6-iodo-4ꢀ-oxo-2-(pyridin-3-yl)quinazolin-3(4H)- 2.83; N, 14.03. Found: C, 47.99; H, 2.49; N, 14.33.
yl)urea (8b). Mp: 235–237 C. Yield: 80%. I.R. (KBr, cmꢁ1): ~nmax
1-(4-Chlorophenyl)-3-(6-iodo-4-oxo-2-(pyridin-3-yl)quinazolin-
3327, 3296 (2NH), 2935 (CH2 aliphatic), 1687, 1660 (2C]O), 3(4H)-yl)urea (8f). Mp: 251–253 ꢀC. Yield%: 90%. I.R. (KBr,
1
1647 (C]N), 1506–1440 (C]C), 530 (C–I). H-NMR 300 MHz cmꢁ1): ~nmax 3296, 3226 (2NH), 3099 (CH aromatic), 1683, 1670
(DMSO-d6, D2O, ppm): d 1.07–1.59 (m, 10H, cyclohexyl-5CH2), (2C]O), 1645 (C]N), 1552–1448 (C]C), 723 (C–Cl), 530 (C–I).
3.35 (m, 1H, cyclohexyl-CH), 7.58 (dd, 1H, J ¼ 4.8 Hz, 7.8 Hz, 1H-NMR 300 MHz (DMSO-d6, D2O, ppm): d 7.28 (d, J ¼ 5.1 Hz,
quinazolinyl-H), 7.86 (s, 1H, pyridyl-H), 7.96 (d, 1H, J ¼ 2.1 Hz, 2H, phenyl-H), 7.48–7.58 (m, 3H, phenyl-H, pyridyl-H), 7.98 (d,
quinazolinyl-H), 8.10 (s, 1H, quinazolinyl-H), 8.19–8.22 (m, 1H,
J
¼
2.10 Hz, quinazolinyl-H), 8.18–8.23 (m, 2H,
1H, pyridyl-H), 8.80 (dd, 1H, J ¼ 1.8, 5.1 Hz, pyridyl-H), 9.05 (s, quinazolinyl-H, pyridyl-H), 8.43 (s, 1H, quinazolinyl-H), 8.78
1H, pyridyl-H), 10.44 (s, 1H, NH exchanged by D2O), 11.78 (s, (dd, 1H, J ¼ 3.00, 6.30 Hz, pyridyl-H), 9.06 (s, 1H, pyridyl-H),
1H, NH exchanged by D2O). Mass (m/z, rel. abundance): 489 10.69 (s, 1H, NH exchanged by D2O), 11.74 (s, 1H, NH
(M+, 5.6%), 106.05 (100%). Anal. calcd for C20H20IN5O2 exchanged by D2O). Mass (m/z, rel. abundance): 517 (M+,
(489.31): C, 49.09; H, 4.12; N, 14.31. Found: C, 49.16; H, 4.18; 19.22%), 519 (M+ + 2, 13.7%), 78 (100%). Anal. calcd for C20
-
N, 14.58.
H13ClIN5O2 (517.71): C, 46.4; H, 2.53; N, 13.53. Found: C, 46.55;
1-Cyclohexyl-3-(6-iodo-4-oxo-2-(pyridin-3-yl)quinazolin-3(4H)- H, 2.80; N, 13.24.
yl)thiourea (8c). Mp: 232–234 C. Yield: 79%. I.R. (KBr, cmꢁ1):
1-(6-Iodo-4-oxo-2-(pyridin-4-yl)quinazolin-3(4H)-yl)-3-sub-
ꢀ
~nmax 3404, 3309 (2NH), 2924 (CH2 aliphatic), 1676 (C]O), 1649 stituted urea/thiourea derivatives (8g–l). Compounds 8g–l were
(C]N), 1516–1448 (C]C), 1201 (C]S), 530 (C–I). 1H-NMR 300 prepared and puried by applying the same method described
MHz (DMSO-d6, D2O, ppm): d 1.01–1.75 (m, 10H, cyclohexyl-
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for 8a–f starting with 7b and using the appropriate isocyanate or 1531–1481 (C]C), 1213 (C]S), 530 (C–I). 1H-NMR 400 MHz
isothiocyanate.
(DMSO-d6, D2O, ppm): d 7.20 (t, 1H, J ¼ 7.20 Hz, phenyl-H),
1-Ethyl-3-(6-iodo-4-oxo-2-(pyridin-4-yl)quinazolin-3(4H)-yl)thio- 7.32–7.36 (m, 4H, phenyl-H), 7.40–7.55 (m, 1H, quinazolinyl-
urea (8g). Mp: 229–231 ꢀC. Yield: 83%. I.R. (KBr, cmꢁ1): ~nmax H), 7.75–7.77 (m, 2H, pyridyl-H), 7.98 (dd, 1H, J ¼ 8.70 Hz,
3321 (2NH), 3134 (CH aromatic), 2968, 2931 (CH2, CH3), 1683 quinazolinyl-H), 8.23 (s, 1H, quinazolinyl-H), 8.80 (d, 2H, J ¼
(C]O), 1598 (C]N), 1512–1483 (C]C), 1213 (C]S), 530 (C–I). 6.30 Hz, pyridyl-H), 9.72 (s, 1H, NH exchanged by D2O), 11.82 (s,
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 1.04 (t, 3H, J ¼ 7.04, 1H, NH exchanged by D2O). Mass (m/z, rel. abundance): 499
CH3CH2), 3.43–3.5 (q, 2H, J ¼ 7.00, CH3CH2), 7.79 (d, 2H, J ¼ (M+, 1.55%), 364 (100%). Anal. calcd for C20H14IN5OS (499.33):
6.00 Hz, pyridyl-H), 7.98 (dd, 1H, J ¼ 1.88, 8.68 Hz, quinazolinyl- C, 48.11; H, 2.83; N, 14.03. Found: C, 48.32; H, 2.81; N, 14.25.
H), 8.21 (d, 2H, J ¼ 8.68 Hz, quinazolinyl-H), 8.86 (d, 2H, J ¼
1-(4-Chlorophenyl)-3-(6-iodo-4-oxo-2-(pyridin-4-yl)quinazolin-
6.00 Hz, pyridyl-H), 10.74 (s, 1H, NH exchanged by D2O), 11.85 3(4H)-yl)urea (8l). Mp: 238–240 ꢀC. Yield%: 92%. I.R. (KBr,
(s, 1H, NH exchanged by D2O). 13C-NMR (100 MHz DMSO-d6, cmꢁ1): ~nmax 3280 (2NH), 3107 (CH aromatic), 1666, 1649 (2CO),
1
ppm): d 19.03 (CH3), 39.05 (CH2), 88.34, 121.32, 123.51, 123.86, 1595 (C]N), 1558–1444 (C]C), 688 (C–Cl), 530 (C–I). H-NMR
137.4, 138.26, 141.44, 141.57, 151.28, 163.76, 167.48 (C]O), 400 MHz (DMSO-d6, D2O, ppm): d 7.31 (d, 2H, J ¼ 8.52 Hz,
181.63 (C]S). Mass (m/z, rel. abundance): 451 (M+, 0.12%), 75 phenyl-H), 7.50 (d, 2H, J ¼ 8.52 Hz, phenyl-H), 7.78 (d, 2H, J ¼
(100%). Anal. calcd for C16H14IN5OS (451.28): C, 42.58; H, 3.13; 4.84 Hz, pyridyl-H), 7.99 (d, H, J ¼ 8.56 Hz, quinazolinyl-H), 8.22
N, 15.52. Found: C, 42.67; H, 3.33; N, 15.26.
(s, 1H, quinazolinyl-H), 8.26 (d, 1H, J ¼ 8.56 Hz, quinazolinyl-
1-Cyclohexyl-3-(6-iodo-4-oxo-2-(pyridin-4-yl)quinazolin-3(4H)- H), 8.80 (d, 2H, J ¼ 4.84 Hz, pyridyl-H), 10.72 (s, 1H, NH
yl)urea (8h). Mp: 225–227 C. Yield: 85%. I.R. (KBr, cmꢁ1): ~nmax exchanged by D2O), 11.87 (s, 1H, NH exchanged by D2O). Mass
ꢀ
3315 (br.) (2NH), 2924, 2850 (CH2 aliphatic), 1687, 1660 (2C] (m/z, rel. abundance): 517 (M+, 2.16%), 519 (M+ + 2, 1.48%), 115
O), 1647 (C]N), 1506 (C]C), 530 (C–I). 1H-NMR 400 MHz (100%). Anal. calcd for C20H13ClIN5O2 (517.71): C, 46.4; H, 2.53;
(DMSO-d6, D2O, ppm): d 1.25–1.72 (m, 10H, cyclohexyl-5CH2), N, 13.53. Found: C, 46.36; H, 2.82; N, 13.37.
3.39–3.46 (m, 1H, cyclohexyl-CH), 7.79 (d, 2H, J ¼ 6.00 Hz,
N-(2-{[2-(4-Substituted
benzylidene)hydrazinyl]carbonyl}-4-
pyridyl-H), 7.95 (dd, 1H, J ¼ 1.92, 8.72 Hz, quinazolinyl-H), 8.14 iodophenyl)pyridine-3-carboxamides (9a–g)
(d, 1H, J ¼ 1.92 Hz, quinazolinyl-H), 8.21 (d, 1H, J ¼ 8.68 Hz,
General method. The appropriate p-substituted benzaldehyde
quinazolinyl-H), 8.82 (d, 2H, J ¼ 6.00 Hz, pyridyl-H), 10.48 (s, (2.40 mmol), the hydrazido derivative 7a (0.72 g, 2 mmol) in
1H, NH exchanged by D2O), 11.86 (s, 1H, NH exchanged by absolute ethanol (10 mL) and in the presence of glacial acetic
D2O). Mass (m/z, rel. abundance): 489 (M+, 0.24%), 56 (100%). acid (1 mL) were reuxed for 5–8 h. The reaction mixture was
Anal. calcd for C20H20IN5O2 (489.31): C, 49.09; H, 4.12; N, 14.31. le to cool; the obtained solid was ltered, washed with diethyl
Found: C, 49.16; H, 3.92; N, 14.56.
ether, dried and recrystallized from absolute ethanol to produce
1-Cyclohexyl-3-(6-iodo-4-oxo-2-(pyridin-3-yl)quinazolin-3(4H)- the pure compounds 9a–g.
yl)thiourea (8i). Mp: 254–256 ꢀC. Yield: 70%. I.R. (KBr, cmꢁ1):
N-(2-{[2-(4-Fluorobenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)-
~nmax 3325, 3230 (2NH), 2929, 2850 (CH2 aliphatic), 1680 (C]O), pyridine-3-carboxamide (9a). Mp: 237–239 ꢀC. Yield: 65%. I.R.
1
1640 (C]N), 1508 (C]C), 1116 (C]S), 530 (C–I). H-NMR 400 (KBr, cmꢁ1): ~nmax 3309 (2NH), 3100 (CH aromatic), 2920 (C–H),
MHz (DMSO-d6, D2O, ppm): d 0.98–1.22 (m, 5H, cyclohexyl), 1687 (C]O), 1654 (C]O), 1597 (C]N), 1506 (C]C), 1234 (C–F),
1.53–1.75 (m, 5H, cyclohexyl), 4.07 (m, 1H, cyclohexyl-CH), 7.77 530 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.32 (t, 2H, J
(d, J ¼ 5.40 Hz, 2H, pyridyl-H), 7.95–8.09 (m, 3H, quinazolinyl- ¼ 8.76 Hz, uorophenyl-H), 7.63 (dd, 1H, J ¼ 4.84, 7.88 Hz,
H), 8.82 (d, 2H, J ¼ 5.40 Hz, pyridyl-H), 10.64 (s, 1H, NH pyridyl-H), 7.82 (dd, 2H, J ¼ 5.64, 8.76 Hz, uorophenyl-H), 7.97
exchanged by D2O), 11.67 (s, 1H, NH exchanged by D2O). Anal. (dd, 1H, J ¼ 1.8 Hz, 8.64 Hz, iodophenyl-H), 8.19–8.27 (m, 3H,
calcd for C20H20IN5OS (505.38): C, 47.53; H, 3.99; N, 13.86. iodophenyl-H, pyridyl-H), 8.44 (s, 1H, C–H), 8.81 (dd, 1H, J ¼ 1.36,
Found: C, 47.77; H, 4.08; N, 13.98.
1-(6-Iodo-4-oxo-2-(pyridin-4-yl)quinazolin-3(4H)-yl)-3-phenylurea 1H, NH exchanged by D2O), 12.56 (s, 1H, NH exchanged by D2O).
(8j). Mp: 263–265 ꢀC. Yield: 81% I.R. (KBr, cmꢁ1): ~nmax 3304, Anal. calcd for C20H14FIN4O2 (488.25): C, 49.20; H, 2.89; N, 11.47.
4.76 Hz, pyridyl-H), 9.1 (d, 1H, J ¼ 1.84 Hz, pyridyl-H), 11.78 (s,
3211 (2NH), 3042 (CH aromatic), 1678, 1668 (2C]O), 1639 (C] Found: C, 49.29; H, 2.91; N, 11.62.
1
N), 1558 (C]C), 530 (C–I). H-NMR 400 MHz (DMSO-d6, D2O,
N-(2-{[2-(4-Chlorobenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)-
ppm): d 6.97 (t, 1H, J ¼ 7.32 Hz, phenyl-H), 7.26 (t, 2H, J ¼ pyridine-3-carboxamide (9b). Mp: 242–244 ꢀC. Yield: 66%. I.R.
7.92 Hz, phenyl-H), 7.46 (d, 2H, J ¼ 7.92 Hz, phenyl-H), 7.95 (d, (KBr, cmꢁ1): ~nmax 3199 (NH), 3103 (CH aromatic), 2924 (C–H),
2H, J ¼ 5.76 Hz, pyridyl-H), 8.00 (dd, 1H, J ¼ 1.6, 8.72 Hz 1672, 1660 (2C]O), 1595 (C]N), 1508 (C]C), 705 (C–Cl), 514
quinazolinyl-H), 8.22 (d, 1H, J ¼ 1.64 Hz, quinazolinyl-H), 8.26 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.54 (d, 2H, J ¼
(s, 1H, J ¼ 8.72 Hz, quinazolinyl-H), 8.79 (d, 2H, J ¼ 5.76 Hz, 8.44 Hz, chloro phenyl-H), 7.63 (dd, 1H, J ¼ 4.84 Hz, 7.88 Hz,
pyridyl-H), 10.7 (s, 1H, NH exchanged by D2O), 11.88 (s, 1H, NH pyridyl-H), 7.78 (d, 2H, J ¼ 8.44 Hz, chlorophenyl-H), 7.97 (dd,
exchanged by D2O). Mass (m/z, rel. abundance): 483 (M+, 1H, J ¼ 1.80, 8.64 Hz, iodophenyl-H), 8.19–8.27 (m, 3H,
0.22%), 91 (100%). Anal. calcd for C20H14IN5O2 (483.26): C, iodophenyl-H, pyridyl-H), 8.43 (s, 1H, C–H), 8.81 (dd, 1H, J ¼
49.71; H, 2.92; N, 14.49. Found: C, 49.65; H, 2.98; N, 14.36.
1.24, 4.76 Hz, pyridyl-H), 9.1 (d, 1H, J ¼ 1.8 Hz, pyridyl-H), 11.75
1-(6-Iodo-4-oxo-2-(pyridin-4-yl)quinazolin-3(4H)-yl)-3-phenylthiourea (s, 1H, NH exchanged by D2O), 12.20 (s, 1H, NH exchanged by
(8k). Mp: 227–229 C. Yield: 78%. I.R. (KBr, cmꢁ1): ~nmax 3340, D2O). 13C-NMR 100 MHz (DMSO-d6, D2O, ppm): d 88.16, 124.21,
ꢀ
3300 (NH), 3062 (CH aromatic), 1693 (C]O), 1614 (C]N), 124.37, 129.23, 129.40, 129.47, 130.36, 133.38, 135.36, 135.42,
111778 | RSC Adv., 2016, 6, 111767–111786
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137.09, 138.76, 141.32, 148.33, 148.79, 153.14 (C–H), 163.77 C20H15IN4O3 (486.26): C, 49.40; H, 3.11; N, 11.52. Found: C, 49.56;
(C]O), 163.82 (C]O). Mass (m/z, rel. abundance): 504 (M+, H, 3.16; N, 11.63.
1.06%), 506 (M+ + 2, 0.37%), 78 (100%). Anal. calcd for C20-
N-(4-Iodo-2-{[2-(4-nitrobenzylidene)hydrazinyl]carbonyl}phenyl)
H14ClIN4O2 (504.71): C, 47.59; H, 2.80; N, 11.10. Found: C, pyridine-3-carboxamide (9g). Mp: 298–300 ꢀC. Yield: 88%. I.R.
47.72; H, 2.78; N, 11.24.
(KBr, cmꢁ1): ~nmax 3329 (2NH), 3103 (CH aromatic), 2949 (C–H),
N-(2-{[2-(4-Bromobenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)- 1683, 1660 (2C]O), 1598 (C]N), 1577 (C]C), 1517 (C–NO2),
1
pyridine-3-carboxamide (9c). Mp: 243–245 ꢀC. Yield: 75%. I.R. 510 (C–I). H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.63 (dd,
(KBr, cmꢁ1): ~nmax 3197 (2NH), 3059 (CH aromatic), 2922 (C–H), 1H, J ¼ 4.88, 7.08 Hz, pyridyl-H), 7.97–8.07 (m, 3H, phenyl-H),
1672, 1660 (2C]O), 1593 (C]N), 1508 (C]C), 610 (C–Br), 520 8.16–8.32 (m, 5H, phenyl-H, pyridyl), 8.53 (s, 1H, C–H), 8.80
(C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.63 (dd, 1H, J (d, 1H, J ¼ 3.56 Hz, pyridyl-H), 9.1 (s, 1H, pyridyl-H), 11.65 (s,
¼ 4.92, 7.96 Hz, pyridyl-H), 7.69 (m, 4H, bromophenyl-H), 7.97 1H, NH exchanged by D2O), 12.41 (s, 1H, NH exchanged by
(d, 1H, J ¼ 1.88, 6.84 Hz, iodophenyl-H), 8.19–8.28 (m, 3H, D2O). Anal. calcd for C20H14IN5O4 (515.26): C, 46.62; H, 2.74; N,
iodophenyl-H, pyridyl-H), 8.41 (s, 1H, C–H), 8.81 (dd, 1H, J ¼ 13.59. Found: C, 46.80; H, 2.71; N, 13.68.
1.28, 3.48 Hz, pyridyl-H), 9.1 (d, 1H, J ¼ 1.84 Hz, pyridyl-H),
N-(2-{[2-(4-Substituted benzylidene)hydrazinyl]carbonyl}-4-
11.75 (s, 1H, NH exchanged by D2O), 12.20 (s, 1H, NH iodophenyl)pyridine-4-carboxamides (9h–n). Compounds 9h–n
exchanged by D2O). Anal. calcd for C20H14BrIN4O2 (549.16): C, were prepared according to the last mentioned procedure using
43.74; H, 2.57; N, 10.20. Found: C, 43.79; H, 2.53; N, 10.28.
7b as a starting material and were recrystallized from absolute
N-[2-({2-[4-(Dimethylamino)benzylidene]hydrazinyl}carbonyl)-4- ethanol.
ꢀ
iodophenyl]pyridine-3-carboxamide (9d). Mp: 255–257 C. Yield:
N-(2-{[2-(4-Fluorobenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)
80%. I.R. (KBr, cmꢁ1): ~nmax 3253 (2NH), 3194 (CH aromatic), pyridine-4-carboxamide (9h). Mp: 288–290 ꢀC. Yield: 59%. I.R.
2987 (CH, CH3), 1668, 1653 (2C]O), 1589 (C]N), 1510 (C]C), (KBr, cmꢁ1): ~nmax 3296 (NH), 3122 (CH aromatic), 2820 (C–H),
510 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 2.99 (s, 6H, 1683 (C]O), 1630 (C]N), 1508 (C]C), 1232 (C–F), 532 (C–I). 1H-
2 ꢄ CH3), 6.67 (d, 2H, J ¼ 8.80 Hz, N,N-dimethylaminophenyl- NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.37 (t, 2H, J ¼ 8.68 Hz,
H), 7.56 (d, 2H, J ¼ 8.80 Hz, dimethylaminophenyl-H), 7.64 uorophenyl-H), 7.61 (d, 1H, J ¼ 8.56 Hz, iodophenyl-H), 7.67 (d,
(dd, 1H, J ¼ 4.88, 7.88 Hz, pyridyl-H), 7.95 (dd, 1H, J ¼ 1.92, 2H, J ¼ 5.92 Hz, pyridyl-H), 7.79 (d, 2H, J ¼ 3.12 Hz, uorophenyl-
8.76 Hz, iodophenyl-H), 8.2 (d, 1H, J ¼ 1.92 Hz, iodophenyl-H), H), 8.22 (dd, 1H, J ¼ 8.72 Hz, iodophenyl-H), 8.52 (d, 1H, J ¼
8.26–8.31 (m, 2H, iodophenyl-H, pyridyl-H), 8.31 (s, 1H, C–H), 1.92 Hz, iodophenyl-H), 8.69 (d, 2H, J ¼ 5.92 Hz, pyridyl-H), 9.08
8.82 (dd, 1H, J ¼ 1.44, 4.8 Hz, pyridyl-H), 9.11 (d, 1H, J ¼ 1.88 Hz, (s, 1H, C–H), 11.70 (s, 1H, NH exchanged by D2O), 12.20 (s, 1H,
pyridyl-H), 11.87 (s, 1H, NH exchanged by D2O), 12.01 (s, 1H, NH exchanged by D2O). Anal. calcd for C20H14FIN4O2 (488.25): C,
NH exchanged by D2O). Mass (m/z, rel. abundance): 513 (M+, 49.20; H, 2.89; N, 11.47. Found: C, 49.29; H, 2.92; N, 11.59.
1.44%), 163 (100%). Anal. calcd for C22H20IN5O2 (513.33): C,
51.47; H, 3.93; N, 13.64. Found: C, 51.53; H, 3.98; N, 13.81.
N-(2-{[2-(4-Chlorobenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)
pyridine-3-carboxamide (9i). Mp: 262–264 ꢀC. Yield: 83%. I.R. (KBr,
N-[4-Iodo-2-({2-[4-(triuoromethyl)benzylidene]hydrazinyl}carbonyl) cmꢁ1): ~nmax 3446 (NH), 3113 (CH aromatic), 2843 (C–H), 1680
phenyl]-pyridine-3-carboxamide (9e). Mp: 245–247 ꢀC. Yield: 72%. (2C]O), 1587 (C]N), 1506–1442 (C]C), 750 (C–Cl), 520 (C–I).
I.R. (KBr, cmꢁ1): ~nmax 3427 (2NH), 3192 (CH aromatic), 2980 (C–H), 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.54 (d, 2H, J ¼
1
1681 (2C]O), 1595 (C]N), 1508 (C]C), 510 (C–I). H-NMR 400 8.40 Hz, chloro phenyl-H), 7.78 (d, 2H, J ¼ 8.40 Hz, chloro phenyl-
MHz (DMSO-d6, D2O, ppm): d 7.62 (dd, 1H, J ¼ 3.00, 4.92 Hz, H), 7.82 (d, 2H, J ¼ 5.88 Hz, pyridyl-H), 7.99 (dd, 1H, J ¼ 1.64,
pyridyl-H), 7.82 (d, 2H, J ¼ 8.28 Hz, triuoromethyl phenyl-H), 8.72 Hz, iodophenyl-H), 8.21 (d, 1H, J ¼ 1.60 Hz, iodophenyl-H),
7.95–7.99 (m, 3H, phenyl-H), 8.18–8.27 (m, 3H, phenyl-H, pyridyl- 8.24 (d, 1H, J ¼ 8.72 Hz, iodophenyl-H), 8.44 (s, 1H, C–H), 8.84 (d,
H), 8.50 (s, 1H, C–H), 8.81 (dd, 1H, J ¼ 3.36, 4.76 Hz, pyridyl-H), 2H, J ¼ 5.88 Hz, pyridyl-H), 11.88 (s, 1H, NH exchanged by D2O),
9.1 (s, J ¼ 1.8 Hz, 1H, pyridyl-H), 11.69 (s, 1H, NH exchanged by 12.22 (s, 1H, NH exchanged by D2O). Mass (m/z, rel. abundance):
D2O), 12.31 (s, 1H, NH exchanged by D2O). Anal. calcd for 504 (M+, 0.23%), 506 (M+ + 2, 0.13%), 78 (100%). Anal. calcd for
C21H14F3IN4O2 (538.26): C, 46.86; H, 2.62; N, 10.41. Found: C, 46.94; C20H14ClIN4O2 (504.71): C, 47.59; H, 2.80; N, 11.10. Found: C,
H, 2.68; N, 10.54.
47.67; H, 2.82; N, 11.23.
N-(2-{[2-(4-Hydroxybenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)
N-(2-{[2-(4-Bromobenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)
pyridine-3-carboxamide (9f). Mp: 273–275 ꢀC. Yield: 79%. I.R. (KBr, pyridine-4-carboxamide (9j). Mp: 283–285 ꢀC. Yield: 70%. I.R.
cmꢁ1): ~nmax 3238 (OH), 3161 (2 NH), 3080 (CH aromatic), 2924 (C– (KBr, cmꢁ1): ~nmax 3423 (NH), 3076 (CH aromatic), 2843 (C–H),
1
H), 1718, 1680 (2C]O), 1593 (C]N), 1496 (C]C), 510 (C–I). H- 1685 (2C]O), 1577 (C]N), 1506 (C]C), 526 (C–Br), 514 (C–
1
NMR 400 MHz (DMSO-d6, D2O, ppm): d 6.85 (d, 2H, J ¼ 8.44 Hz, I). H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.67–7.72 (m,
hydroxyphenyl-H), 7.59 (d, 2H, J ¼ 8.44 Hz, hydroxyphenyl-H), 7.63 4H, bromophenyl-H), 7.82 (d, 2H, J ¼ 5.80 Hz, pyridyl-H),
(dd, 1H, J ¼ 2.92, 4.96 Hz, pyridyl-H), 7.95 (dd, 1H, J ¼ 1.44, 7.98 (dd, 1H, J ¼ 1.60, 8.72 Hz, iodophenyl-H), 8.21 (d, 1H,
8.76 Hz, iodophenyl-H), 8.20 (d, 1H, J ¼ 1.56 Hz, iodophenyl-H), J ¼ 1.60 Hz, iodophenyl-H), 8.24 (d, 1H, J ¼ 8.76 Hz,
8.26–8.28 (m, 2H, iodophenyl-H, pyridyl-H), 8.34 (s, 1H, C–H), iodophenyl-H), 8.42 (s, 1H, C–H), 8.84 (d, 2H, J ¼ 5.80 Hz,
8.82 (dd, 1H, J ¼ 3.72, 4.72 Hz, pyridyl-H), 9.10 (d, 1H, J ¼ 1.84 Hz, pyridyl-H), 11.87 (s, 1H, NH exchanged by D2O), 12.22 (s, 1H,
pyridyl-H), 10.00 (s, 1H, OH exchanged by D2O), 11.93 (s, 1H, NH NH exchanged by D2O). Anal. calcd for C20H14BrIN4O2
exchanged by D2O), 11.96 (s, 1H, NH exchanged by D2O). Mass (m/ (549.16): C, 43.74; H, 2.57; N, 10.20. Found: C, 43.85; H, 2.53;
z, rel. abundance): 486 (M+, 1.8%), 67 (100%). Anal. calcd for N, 10.34.
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mmol) was heated under reux for 3 h. The reaction mixture
was cooled to room temperature. The separated solid was
ltered, washed with water and recrystallized from ethanol.
3-[(4-Fluorobenzylidene)amino]-6-iodo-2-(pyridin-3-yl)quinazo-
lin-4(3H)-one (10a). Mp: 337–339 ꢀC. Yield: 80%. I.R. (KBr,
cmꢁ1): ~nmax 3190 (CH aromatic), 2922 (C–H), 1681 (C]O), 1593
N-[2-({2-[4-(Dimethylamino)benzylidene]hydrazinyl}carbonyl)-4-
iodophenyl]-pyridine-4-carboxamide (9k). Mp: 228–230 C. Yield:
ꢀ
80%. I.R. (KBr, cmꢁ1): ~nmax 3408 (NH), 3159 (CH aromatic), 2880
(CH, CH3(s)), 1672 (2C]O), 1610 (C]N), 1508 (C]C), 514 (C–I).
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 3.01 (s, 6H, 2 ꢄ
CH3), 6.77 (d, 2H, J ¼ 8.92 Hz, aminophenyl-H), 7.56 (d, 2H, J ¼
8.92 Hz, aminophenyl-H), 7.83 (d, 2H, J ¼ 5.96 Hz, pyridyl-H),
7.96 (dd, 1H, J ¼ 1.80, 8.76 Hz, iodophenyl-H), 8.22 (d, 1H, J
¼ 1.84 Hz, iodophenyl-H), 8.31 (d, 1H, J ¼ 1.6 Hz, iodophenyl-
H), 8.33 (s, 1H, C–H), 8.85 (d, 2H, J ¼ 5.96 Hz, pyridyl-H),
11.89 (s, 1H, NH exchanged by D2O), 12.43 (s, 1H, NH
exchanged by D2O). 13C-NMR 100 MHz (DMSO-d6, D2O, ppm):
d 56.49 (2 ꢄ CH3 carbon), 88.25, 112.24, 121.36, 121.48, 123.69,
124.23, 129.24, 136.95, 138.78, 141.18, 141.70, 150.88 (C–H),
151.28, 152.25, 163.25 (C]O), 163.48 (C]O). Anal. calcd for
1
(C]N), 1506 (C]C), 1226 (C–F), 530 (C–I). H-NMR 400 MHz
(DMSO-d6, ppm): d 7.23 (t, 2H, J ¼ 8.64 Hz, uorophenyl-H), 7.59
(d, 2H, J ¼ 7.44 Hz, uorophenyl-H), 7.64 (d, 1H, J ¼ 8.68 Hz,
quinazolinyl-H), 7.70 (t, 1H, J ¼ 7.00 Hz, pyridyl-H), 8.18 (s, 1H,
C–H), 8.32 (d, 1H, J ¼ 7.76 Hz, quinazolinyl-H), 8.47 (d, 1H, J ¼
8.64 Hz, pyridyl-H), 8.51 (s, 1H, quinazolinyl-H), 8.77 (d, 1H, J ¼
3.76 Hz, pyridyl), 9.17 (s, 1H, pyridyl). Mass (m/z, rel. abun-
dance): 470 (M+, 9.28%), 78 (100%). Anal. calcd for C20H12FIN4O
(470.24): C, 51.08; H, 2.57; N, 11.91. Found: C, 51.19; H, 2.55; N,
12.04.
C
22H20IN5O2 (513.33): C, 51.47; H, 3.93; N, 13.64. Found: C,
3-[(4-Chlorobenzylidene)amino]-6-iodo-2-(pyridin-3-yl)quinazo-
lin-4(3H)-one (10b). Mp: 238–340 ꢀC. Yield: 77%. I.R. (KBr,
cmꢁ1): ~nmax 3290 (CH aromatic), 2922 (C–H), 1645 (C]O), 1577
51.53; H, 3.98; N, 13.81.
N-[4-Iodo-2-({2-[4-(triuoromethyl)benzylidene]hydrazinyl}carbonyl)
phenyl]-pyridine-4-carboxamide (9l). Mp: 238–240 ꢀC. Yield: 67%. I.R.
(KBr, cmꢁ1): ~nmax 3412 (NH), 3211 (CH aromatic), 2840 (C–H), 1678
(C]O), 1651 (C]O), 1589 (C]N), 1508–1444 (C]C), 540 (C–I). 1H-
NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.82–7.84 (m, 4H,
triuorophenyl-H), 7.97 (d, 2H, J ¼ 7.88 Hz, pyridyl-H), 8.00 (d, 1H, J
¼ 1.64, iodophenyl-H), 8.21–8.23 (m, 2H, iodophenyl-H), 8.51 (s,
1H, C–H), 8.84 (d, 2H, J ¼ 7.88 Hz, pyridyl-H), 11.82 (s, 1H, NH
exchanged by D2O), 12.33 (s, 1H, NH exchanged by D2O). Mass (m/
z, rel. abundance): 538 (M+, 0.44%), 106 (100%). Anal. calcd for
C21H14F3IN4O2 (538.26): C, 46.86; H, 2.62; N, 10.41. Found: C, 46.95;
H, 2.61; N, 10.50.
N-(2-{[2-(4-Hydroxybenzylidene)hydrazinyl]carbonyl}-4-iodophenyl)
pyridine-4-carboxamide (9m). Mp: 297–299 ꢀC. Yield: 90%. I.R. (KBr,
cmꢁ1): ~nmax 3176 (OH), 3111 (NH), 3057 (CH aromatic), 2868 (C–H),
1685 (C]O), 1643 (C]O), 1610 (C]N), 1512 (C]C), 536 (C–I). 1H-
NMR 400 MHz (DMSO-d6, D2O, ppm): d 6.86 (d, 2H, J ¼ 8.44 Hz,
hydroxyphenyl-H), 7.59 (d, 2H, J ¼ 8.44 Hz, hydroxyphenyl-H), 7.82
(d, 2H, J ¼ 5.68 Hz, pyridyl-H), 7.96 (d, 1H, J ¼ 8.64 Hz, iodophenyl-
H), 8.21 (d, 1H, J ¼ 1.40 Hz, iodophenyl-H), 8.30 (d, 1H, J ¼ 8.72 Hz,
iodophenyl-H), 8.35 (s, 1H, C–H), 8.84 (d, 2H, J ¼ 5.68 Hz, pyridyl-
H), 10.01 (s, 1H, OH exchanged by D2O), 11.98 (s, 1H, NH
exchanged by D2O), 12.07 (s, 1H, NH exchanged by D2O). Anal.
calcd for C20H15IN4O3 (486.26): C, 49.40; H, 3.11; N, 11.52. Found:
C, 49.48; H, 3.14; N, 11.58.
1
(C]N), 1504 (C]C), 750 (C–Cl), 520 (C–I). H-NMR 400 MHz
(DMSO-d6, ppm): d 7.44 (d, 2H, J ¼ 7.76 Hz, chloro phenyl-H),
7.82 (d, 2H, J ¼ 8.54 Hz, chloro phenyl-H), 7.89 (d, 1H, J ¼
5.88 Hz, pyridyl-H), 8.20 (dd, 1H, J ¼ 1.58, 8.70 Hz, quinazlinyl-
H), 8.24–8.35 (m, 3H, quinazolinyl-H, pyridyl-H), 8.44 (d, 1H, J ¼
8.72 Hz, quinazolinyl-H), 8.65 (s, 1H, C–H), 8.84 (d, 1H, J ¼
6.00 Hz, pyridyl-H). Anal. calcd for C20H12ClIN4O (486.69): C,
49.36; H, 2.49; N, 11.51. Found: C, 49.08; H, 2.60; N, 11.77.
3-[(4-Bromobenzylidene)amino]-6-iodo-2-(pyridin-3-yl)quinazo-
lin-4(3H)-one (10c). Mp: 335–337 ꢀC. Yield: 81%. I.R. (KBr,
cmꢁ1): ~nmax 3284 (CH aromatic), 2951 (C–H), 1678 (C]O), 1587
1
(C]N), 1506 (C]C), 526 (C–Br), 514 (C–I). H-NMR 400 MHz
(DMSO-d6, ppm): d 7.56–7.62 (m, 4H, bromophenyl-H), 7.65 (d,
2H, J ¼ 2.76 Hz, pyridyl-H, quinazolinyl-H), 8.13 (s, 1H, C–H),
8.32 (d, 1H, J ¼ 7.84 Hz, quinazolinyl-H), 8.47 (d, 2H, J ¼
8.72 Hz, quinazolinyl-H, pyridyl-H), 8.50 (s, 1H, pyridyl-H), 8.77
(d, 1H, J ¼ 4.44 Hz, pyridyl-H). Mass (m/z, rel. abundance): 531
(M+, 19.59%), 532 (M+ + 1, 13.21%), 78 (100%). Anal. calcd for
C
20H12BrIN4O (531.14): C, 45.23; H, 2.28; N, 10.55. Found: C,
45.31; H, 2.26; N, 10.63.
6-Iodo-3-((4-nitrobenzylidene)amino)-2-(pyridin-3-yl)quinazolin-
4(3H)-one (10d). Mp: 315–317 C. Yield: 70%. I.R. (KBr, cmꢁ1):
ꢀ
~nmax 3064 (CH aromatic), 2954 (CH), 1654 (C]O), 1629 (C]N),
1512 (C–NO2), 1506 (C]C), 514 (C–I). 1H-NMR 400 MHz (DMSO-
d6, ppm): d 7.60–7.73 (m, 4H, nitrophenyl-H), 8.22 (d, 2H, J ¼
8.28 Hz, pyridyl-H, quinazolinyl-H), 8.30–8.34 (m, 2H, pyridyl-H,
quinazolinyl-H), 8.46 (d, 1H, J ¼ 7.36 Hz, quinazolinyl-H), 8.53
(s, 1H, C–H), 8.80 (d, 1H, J ¼ 4.64 Hz, pyridyl-H), 9.17 (s, 1H,
pyridyl-H). Mass (m/z, rel. abundance): 497 (M+, 37.31%), 78
(100%). Anal. calcd for C20H12IN5O3 (497.25): C, 48.31; H, 2.43;
N, 14.08. Found: C, 48.50; H, 2.46; N, 14.17.
3-[(4-Substituted benzylidene)amino]-6-iodo-2-(pyridin-4-yl)
quinazolin-4(3H)-one (10e–h). Compounds 10e–h were simi-
larly prepared and puried adopting the method described for
compounds 10a–f.
N-(4-Iodo-2-{[2-(4-nitrobenzylidene)hydrazinyl]carbphenyl)pyridine-
4-carboxamide (9n). Mp: 298–300 ꢀC. Yield: 88%. I.R. (KBr, cmꢁ1):
~nmax 3487 (NH), 3109 (CH aromatic), 2837 (C–H), 1689 (C]O), 1645
1
(C]O), 1635 (C]N), 1555 (C–NO2), 1514 (C]C), 530 (C–I). H-
NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.82 (d, 2H, J ¼ 5.8 Hz,
pyridyl-H), 8.00–8.02 (m, 3H, phenyl-H), 8.19 (d, 1H, J ¼ 8.7 Hz,
iodophenyl-H), 8.21 (d, 1H, J ¼ 1.64 Hz, iodophenyl-H), 8.32 (d, 2H,
J ¼ 8.44 Hz, nitrophenyl-H), 8.53 (s, 1H, C–H), 8.84 (d, 2H, J ¼
5.8 Hz, pyridyl-H), 11.76 (s, 1H, NH exchanged by D2O), 12.41 (s,
1H, NH exchanged by D2O). Anal. calcd for C20H14IN5O4 (515.26):
C, 46.62; H, 2.74; N, 13.59. Found: C, 46.80; H, 2.71; N, 13.68.
3-[(4-Substituted benzylidene)amino]-6-iodo-2-(pyridin-3-yl)
quinazolin-4(3H)-one (10a–d). A mixture containing equimolar
amounts of sodium ethoxide and the diamide 9a, 9b, 9c or 9g (1
3-[(4-Fluorobenzylidene)amino]-6-iodo-2-(pyridin-4-yl)quinazo-
lin-4(3H)-one (10e). Mp: 304–306 ꢀC. Yield: 59%. I.R. (KBr,
cmꢁ1): ~nmax 3111 (CH aromatic), 2921 (C–H), 1645 (C]O), 1598
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(C]N), 1506 (C]C), 1230 (C–F), 536 (C–I). H-NMR 400 MHz calcd for C15H12ClIN4O3 (458.64): C, 39.28; H, 2.64; N, 12.22.
(DMSO-d6, ppm): d 7.32 (t, 2H, J ¼ 7.60 Hz, phenyl-H), 7.82 (d, Found: C, 39.41; H, 2.68; N, 12.38.
2H, J ¼ 7.60 Hz, phenyl-H), 7.98 (d, 2H, J ¼ 8.76 Hz,
N-(2-{[2-(Chloroacetyl)hydrazinyl]carbonyl}-4-iodophenyl)
quinazolinyl-H), 8.21 (d, 2H, J ¼ 5.44 Hz, pyridyl-H), 8.21–8.26 pyridine-4-carboxamide (11b). This compound was prepared
(m, 2H, quinazolinyl-H), 8.45 (s, 1H, C–H), 8.84 (d, 2H, J ¼ and puried as per the previously mentioned procedure, using
5.44 Hz, pyridyl-H). Mass (m/z, rel. abundance): 470 (M+, 0.46%), the hydrazido derivative 7b. Mp: 289–291 ꢀC. Yield: 80%. I.R.
75 (100%). Anal. calcd for C20H12FIN4O (470.24): C, 51.08; H, (KBr, cmꢁ1): ~nmax 3468, 3448 (NHs), 3178 (CH aromatic), 2974
2.57; N, 11.91. Found: C, 51.22; H, 2.62; N, 12.07.
(CH aliphatic), 1695, 1685 (3C]O), 1631 (C]N), 1508–1436
3-[(4-Chlorobenzylidene)amino]-6-iodo-2-(pyridin-4-yl)quinazo- (C]C), 524 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm):
lin-4(3H)-one (10f). Mp: 331–333 ꢀC. Yield: 65%. I.R. (KBr, cmꢁ1): d 4.24 (s, 2H, CH2), 7.79 (d, 2H, J ¼ 5.48 Hz, pyridyl-H), 7.99 (d,
~nmax 3111 (CH aromatic), 2920 (C–H), 1654 (C]O), 1597 (C]N), 1H, J ¼ 8.68 Hz, phenyl), 8.20 (s, 1H, phenyl), 8.30 (d, 1H, J ¼
1508 (C]C), 750 (C–Cl), 520 (C–I). 1H-NMR 400 MHz (DMSO-d6, 8.76 Hz, phenyl-H), 8.83 (dd, 2H, J ¼ 5.48 Hz, pyridyl-H), 10.58
ppm): d 7.44 (d, 2H, J ¼ 8.28 Hz, chloro phenyl-H), 7.64 (dd, 1H, (s, 1H, NH exchanged by D2O), 11.00 (s, 1H, NH exchanged by
J ¼ 1.76, 8.76 Hz, quinazolinyl-H), 7.67 (d, 2H, J ¼ 8.40 Hz, D2O), 11.89 (s, 1H, NH exchanged by D2O). Mass (m/z, rel.
chloro phenyl-H), 7.88 (d, 2H, J ¼ 5.44 Hz, pyridyl-H), 8.16 (s, abundance): 460 (M+ + 2, 1.10%), 458 (M+, 2.37%), 56 (100%).
1H, C–H), 8.45 (d, 1H, J ¼ 1.60 Hz, quinazolinyl-H), 8.50 (d, 1H, J Anal. calcd for C15H12ClIN4O3 (458.64): C, 39.28; H, 2.64; N,
¼ 8.72 Hz, quinazolinyl-H), 8.78 (d, 2H, J ¼ 5.44 Hz, pyridyl-H). 12.22. Found: C, 39.38; H, 2.68; N, 12.28.
Anal. calcd for C20H12ClIN4O (486.69): C, 49.36; H, 2.49; N,
11.51. Found: C, 49.43; H, 2.54; N, 11.67.
N-(4-Iodo-2-{[2-(substituted
phenyl)pyridine-3-carboxamide (12a–d). Equimolar amounts
acetyl)hydrazinyl]carbonyl}
3-[(4-Bromobenzylidene)amino]-6-iodo-2-(pyridin-4-yl)quinazo- of chloroacetyl hydrazido derivative 11a (0.45 g, 1 mmol) and
lin-4(3H)-one (10g). Mp: 327–329 ꢀC. Yield: 72%. I.R. (KBr, the appropriate secondary amine (1 mmol) in absolute ethanol
cmꢁ1): ~nmax 3203 (CH aromatic), 2968 (C–H), 1649 (2C]O), 1577 (15 mL) containing anhydrous potassium carbonate (1.5 mmol,
1
(C]N), 1506 (C]C), 526 (C–Br), 512 (C–I). H-NMR 400 MHz 0.2 g), were reuxed for 12 h. The reaction mixture was ltered
(DMSO-d6, ppm):
d 7.57–7.65 (m, 5H, bromophenyl-H, while hot and the clear solution was evaporated under reduced
quinazolinyl-H), 7.88 (d, 2H, J ¼ 5.84 Hz, pyridyl-H), 8.14 (d, pressure. The remaining residue was collected, washed with
1H, J ¼ 3.60 Hz, quinazolinyl-H), 8.46 (d, 1H, J ¼ 8.60 Hz, water and recrystallized from ethanol affording the corre-
quinazolinyl-H), 8.51 (s, 1H, C–H), 8.79 (d, 2H, J ¼ 5.84 Hz, sponding derivatives 12a–d.
pyridyl-H). Anal. calcd for C20H12BrIN4O (531.14): C, 45.23; H,
2.28; N, 10.55. Found: C, 45.34; H, 2.31; N, 10.73.
N-(4-Iodo-2-{[2-(piperidin-1-ylacetyl)hydrazinyl]carbonyl}phenyl)
pyridine-3-carboxamide (12a). Mp: 200–202 ꢀC. Yield: 55%. I.R.
6-Iodo-3-((4-nitrobenzylidene)amino)-2-(pyridin-4-yl)quinazolin- (KBr, cmꢁ1): ~nmax 3419 (NHs), 3132 (CH aromatic), 2941 (CH2),
4(3H)-one (10h). Mp: 333 C. Yield: 84%. I.R. (KBr, cmꢁ1): ~nmax 1670 (C]O), 1653 (C]N), 1541 (C]C), 516 (C–I). H-NMR 400
1
ꢀ
3070 (CH aromatic), 2960 (CH), 1657 (C]O), 1610 (C]N), 1518 MHz (DMSO-d6, D2O, ppm): d 1.39 (d, J ¼ 4.44 Hz, 2H,
1
(C–NO2), 1508 (C]C), 514 (C–I). H-NMR 400 MHz (DMSO-d6, piperidinyl-H), 1.55 (t, J ¼ 5.36 Hz, 4H, piperidinyl-H), 3.09 (s,
ppm): d 7.68 (d, 1H, J ¼ 8.36 Hz, quinazolinyl-H), 7.88–7.90 (m, 2H, CH2), 3.34 (s, 4H, piperidinyl-H), 7.60 (dd, 1H, J ¼ 4.84,
4H, nitrophenyl-H), 8.20 (d, 2H, J ¼ 5.28 Hz, pyridyl-H), 8.29 (s, 7.92 Hz, pyridyl-H), 7.95 (dd, 1H, J ¼ 1.48, 8.72 Hz, phenyl-H),
1H, C–H), 8.47 (d, 1H, J ¼ 8.40 Hz, quinazolinyl-H), 8.53 (s, 1H, 8.20 (d, 1H, J ¼ 1.68 Hz, phenyl-H), 8.30 (d, 1H, J ¼ 7.60 Hz,
quinazolinyl-H), 8.82 (d, 2H, J ¼ 5.28 Hz, pyridyl-H). Mass (m/z, pyridyl-H), 8.36 (d, 1H, J ¼ 8.76 Hz, phenyl-H), 8.79 (dd, 1H, J ¼
rel. abundance): 497 (M+, 1.4%), 59 (100%). Calcd for 1.04, 4.72 Hz, pyridyl-H), 9.07 (d, 1H, J ¼ 1.64 Hz, pyridyl-H), 9.93
C
20H12IN5O3 (497.25): C, 48.31; H, 2.43; N, 14.08. Found: C, (s, 1H, NH exchanged by D2O), 10.74 (s, 1H, NH exchanged by
48.39; H, 2.41; N, 14.19.
D2O), 12.07 (s, 1H, NH exchanged by D2O). Anal. calcd for
N-(2-{[2-(Chloroacetyl)hydrazinyl]carbonyl}-4-iodophenyl) C20H22IN5O3 (507.32): C, 47.35; H, 4.37; N, 13.80. Found: C,
pyridine-3-carboxamide (11a). The hydrazido derivative 7a 47.54; H, 4.44; N, 13.96.
(10 mmol, 3.64 g) and chloroacetyl chloride (15 mmol, 1.69 g)
N-(4-Iodo-2-{[2-(morpholin-4-ylacetyl)hydrazinyl]carbonyl}phenyl)
were stirred overnight at room temperature in dimethylforma- pyridine-3-carboxamide (12b). Mp: 268–270 ꢀC. Yield: 53%. I.R.
mide (10 mL). The reaction mixture was poured onto ice-cold (KBr, cmꢁ1): ~nmax 3251 (NHs), 3182 (CH aromatic), 2856 (CH2),
water and the separated solid was collected, dried and recrys- 1700, 1681 (3C]O), 1641 (C]N), 1556 (C]C), 514 (C–I). 1H-NMR
ꢀ
tallized from methanol. Mp: 276–278 C. Yield: 83% I.R. (KBr, 400 MHz (DMSO-d6, D2O, ppm): d 2.52 (t, 4H, J ¼ 4.04 Hz,
cmꢁ1): ~nmax 3479 (NH), 3176 (CH aromatic), 2953 (CH2), 1707, morpholinyl-H), 3.11 (s, 2H, CH2), 3.63 (t, 4H, J ¼ 4.52 Hz,
1687 (3C]O), 1651 (C]N), 1516 (C]C), 785 (C–Cl), 524 (C–I). morpholinyl-H), 7.61 (dd, 1H, J ¼ 4.80, 7.88 Hz, pyridyl-H), 7.97
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 4.24 (s, 2H, CH2), (dd, 1H, J ¼ 1.73, 8.80 Hz, phenyl-H), 8.18 (d, 1H, J ¼ 1.44 Hz,
7.62 (dd, 1H, J ¼ 3.64, 7.88 Hz, pyridyl-H), 7.98 (dd, 1H, J ¼ 1.72, phenyl-H), 8.25 (d, 1H, J ¼ 7.88 Hz, pyridyl-H), 8.35 (d, 1H, J ¼
8.76 Hz, phenyl-H), 8.19 (d, 1H, J ¼ 1.76 Hz, phenyl-H), 8.24 (d, 8.76 Hz, phenyl-H), 8.81 (dd, 1H, J ¼ 1.12, 4.68 Hz, pyridyl-H), 9.06
1H, J ¼ 7.96 Hz, pyridyl-H), 8.29 (d, 1H, J ¼ 8.76 Hz, phenyl-H), (d, 1H, J ¼ 1.76 Hz, pyridyl-H), 9.98 (s, 1H, NH exchanged by D2O),
8.80 (dd, 1H, J ¼ 1.04, 4.68 Hz, pyridyl-H), 9.1 (d, 1H, J ¼ 1.72 Hz, 10.74 (s, 1H, NH exchanged by D2O), 11.90 (s, 1H, NH exchanged
pyridyl-H), 10.56 (s, 1H, NH exchanged by D2O), 10.98 (s, 1H, NH by D2O). Mass (m/z, rel. abundance): 509 (M+, 1.04%), 60 (100%).
exchanged by D2O), 11.81 (s, 1H, NH exchanged by D2O). Anal. Anal. calcd for C19H20IN5O4 (509.30): C, 44.81; H, 3.96; N, 13.75.
Found: C, 44.89; H, 3.99; N, 13.89.
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Ethyl 4-({N0[(5-iodo-2-pyridine-3-amido)benzoyl]hydrazinecarbonyl}
methyl)-piperazine-carboxylate (12c). Mp: 237–239 C. Yield: 78%.
(dd, 1H, J ¼ 1.92, 8.76 Hz, phenyl-H), 8.20 (d, 1H, J ¼ 1.96 Hz,
phenyl-H), 8.35 (d, 1H, J ¼ 8.76 Hz, phenyl-H), 8.82 (dd, 2H, J ¼
1.40, 4.52 Hz, pyridyl-H), 9.99 (s, 1H, NH exchanged by D2O),
10.80 (s, 1H, NH exchanged by D2O), 11.96 (s, 1H, NH exchanged
by D2O). Anal. calcd for C19H20IN5O4 (509.30): C, 44.81; H, 3.96; N,
13.75. Found: C, 44.89; H, 3.98; N, 13.79.
ꢀ
I.R. (KBr, cmꢁ1): ~nmax 3244 (NHs), 3111 (CH aromatic), 2927 (CH2
aliphatic), 1701, 1681, 1654 (3C]O), 1597 (C]N), 1508 (C]C),
516 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 1.18 (t, 3H, J
¼ 7.08 Hz, CH3CH2), 3.15 (s, 2H, CH2), 3.41 (s, 8H, piperazinyl-H),
4.04 (q, 2H, J ¼ 7.04 Hz, CH3CH2), 7.61 (dd, 1H, J ¼ 4.84, 7.84 Hz,
pyridyl-H), 7.97 (dd, 1H, J ¼ 1.68, 8.76 Hz, phenyl-H), 8.18 (d, 1H, J
¼ 1.76 Hz, phenyl-H), 8.24 (d, 1H, J ¼ 7.96 Hz, pyridyl-H), 8.35 (d,
1H, J ¼ 8.80 Hz, phenyl-H), 8.80 (dd, 1H, J ¼ 3.56, 4.76 Hz, pyridyl-
H), 9.06 (d, 1H, J ¼ 1.68 Hz, pyridyl-H), 10.01 (s, 1H, NH exchanged
by D2O), 10.80 (s, 1H, NH exchanged by D2O), 11.89 (s, 1H, NH
exchanged by D2O). Mass (m/z, rel. abundance): 580 (M+, 0.17%),
165 (100%). Anal. calcd for C22H25IN6O5 (580.38): C, 45.53; H, 4.34;
N, 14.48. Found: C, 45.58; H, 4.36; N, 15.55.
Ethyl 4-({N0[(5-iodo-2-pyridine-4-amido)benzoyl]hydrazinecarbonyl}
methyl)-piperazinecarboxylate (12g). Mp: 176–178 ꢀC. Yield: 60%.
I.R. (KBr, cmꢁ1): ~nmax 3479, 3419 (NHs), 3228 (CH aromatic), 2980
(CH2 aliphatic), 1683, 1653 (3C]O), 1591 (C]N), 1508 (C]C), 538
1
(C–I). H-NMR 400 MHz (CDCl3, D2O, ppm): d 1.30 (t, 3H, J ¼
7.08 Hz, CH3CH2), 2.69 (s, 4H, piperazinyl-H), 3.32 (s, 2H, CH2),
3.62 (s, 4H, piperazinyl-H), 4.18 (q, 2H, J ¼ 7.08 Hz, CH3CH2), 7.60
(d, 1H, J ¼ 8.72 Hz, phenyl-H), 7.84 (d, 2H, J ¼ 5.36 Hz, pyridyl-H),
8.05 (s, 1H, phenyl-H), 8.53 (d, 1H, J ¼ 8.72 Hz, phenyl-H), 8.85 (d,
2H, J ¼ 5.36 Hz, pyridyl-H), 9.38 (s, 1H, NH exchanged by D2O),
10.53 (s, 1H, NH exchanged by D2O), 12.14 (s, 1H, NH exchanged
by D2O). Anal. calcd for C22H25IN6O5 (580.38): C, 45.53; H, 4.34; N,
14.48. Found: C, 45.70; H, 4.19; N, 14.50.
N-(4-Iodo-2-{[2-(4-phenylpiperazin-1-ylacetyl)hydrazinyl]carbonyl}
phenyl)pyridine-3-carboxamide (12d). Mp: 213–215 ꢀC. Yield: 62%.
I.R. (KBr, cmꢁ1): ~nmax 3219 (NHs), 3188 (CH aromatic), 2823 (CH
aliphatic), 1683, 1658 (3C]O), 1597 (C]N), 1506–1452 (C]C),
514 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 2.51 (t, 4H, J
¼ 1.68 Hz, piperazinyl), 2.69 (s, 4H, piperazinyl), 3.19 (s, 2H, CH2),
6.78 (t, 1H, J ¼ 7.2 Hz, phenyl-H), 6.95 (d, 2H, J ¼ 7.44 Hz, phenyl-
H), 7.22 (t, 2H, J ¼ 7.44 Hz, phenyl-H), 7.60 (dd, 1H, J ¼ 4.84,
7.88 Hz, pyridyl-H), 7.97 (dd, 1H, J ¼ 1.80, 8.72 Hz, iodophenyl-H),
8.19 (d, 1H, J ¼ 1.88 Hz, iodophenyl-H), 8.25 (d, 1H, J ¼ 7.96 Hz,
pyridyl-H), 8.35 (d, 1H, J ¼ 8.76 Hz, iodophenyl-H), 8.80 (dd, 1H, J
¼ 1.32, 4.76 Hz, pyridyl-H), 9.07 (d, 1H, J ¼ 1.80 Hz, pyridyl-H),
10.01 (s, 1H, NH exchanged by D2O), 10.82 (s, 1H, NH
exchanged by D2O), 11.90 (s, 1H, NH exchanged by D2O). Anal.
calcd for C25H25IN6O3 (584.41): C, 51.38; H, 4.31; N, 14.38. Found:
C, 51.54; N, 4.36; N, 14.48.
N-(4-Iodo-2-{[2-(4-phenylpiperazin-1-ylacetyl)hydrazinyl]carbonyl}
phenyl)pyridine-3-carboxamide (12h). Mp: 240–242 ꢀC. Yield: 68%.
I.R. (KBr, cmꢁ1): ~nmax 3277 (NHs), 3103 (CH aromatic), 2891 (CH2
aliphatic), 1695, 1678, 1647 (2C]O), 1593 (C]N), 1556–1452
(C]C), 511 (C–I). 1H-NMR 400 MHz (CDCl3-d6, D2O, ppm): d 2.88
(t, 4H, J ¼ 4.56 Hz, piperazinyl-H), 3.33 (t, 4H, J ¼ 4.6 Hz,
piperazinyl-H), 3.36 (s, 2H, CH2), 6.93 (t, 1H, J ¼ 7.4 Hz, phenyl-
H), 6.97 (d, 2H, J ¼ 8.32 Hz, phenyl-H), 7.32 (t, 2H, J ¼ 8.32 Hz,
phenyl-H), 7.77 (dd, 1H, J ¼ 1.80, 8.88 Hz, iodophenyl-H), 7.84
(dd, 2H, J ¼ 1.48, 4.48 Hz, pyridyl-H), 8.04 (d, 1H, J ¼ 1.72 Hz,
iodophenyl-H), 8.61 (d, 1H, J ¼ 8.88 Hz, iodophenyl-H), 8.83 (d,
2H, J ¼ 6.00 Hz, pyridyl-H), 9.58 (s, 2H, NH exchanged by D2O),
11.99 (s, 1H, NH exchanged by D2O). Anal. calcd for C25H25IN6O3
(584.41): C, 51.38; H, 4.31; N, 14.38. Found: C, 51.46; N, 4.16; N,
14.46.
3-Hydroxy-6-iodo-2-(pyridin-3-yl)quinazolin-4(3H)-one (13a).
A mixture of (pyridin-3-yl)-benzoxazin-4-one 6a (3.5 g, 10 mmol)
and hydroxylamine hydrochloride (0.69 g, 10 mmol) in dry
pyridine (10 mL) was heated under reux for 8 h. The product
obtained upon cooling was ltered off, washed with diluted
acetic acid and recrystallized from ethanol. Mp: 317–319 ꢀC.
Yield: 48%. I.R. (KBr, cmꢁ1): ~nmax 3151 (OH), 3082 (CH
aromatic), 1666 (C]O), 1608 (C]N), 1517 (C]C), 530 (C–I). 1H-
NMR 300 MHz (DMSO-d6, D2O, ppm): d 7.64 (t, J ¼ 5.10 Hz, 1H,
pyridyl-H), 7.99 (dd, J ¼ 1.80, 8.70 Hz, 1H, quinazolinyl-H), 8.26–
8.29 (m, 2H, pyridyl-H), 8.40 (d, J ¼ 8.7 Hz, 1H, quinazolinyl-H),
8.83 (s, 1H, quinazolinyl-H), 9.12 (s, 1H, pyridyl-H), 12.04 (s, 1H,
OH exchanged by D2O). Mass (m/z, rel. abundance): 365 (M+,
0.15%), 350 (100%). Anal. calcd for C13H8IN3O2 (365.13): C,
42.76; H, 2.21; N, 11.51. Found: C, 42.90; H, 2.29; N, 11.66.
3-Hydroxy-6-iodo-2-(pyridin-4-yl)quinazolin-4(3H)-one (13b).
Compound 13b was similarly prepared and puried as
compound 13a but using (pyridin-4-yl)-benzoxazin-4-one (6b) as
a starting material. Mp: 297–299 ꢀC. Yield: 42%. I.R. (KBr,
cmꢁ1): ~nmax 3431 (OH), 3068 (CH aromatic), 1681 (C]O), 1593
(C]N), 1496 (C]C), 534 (C–I). 1H-NMR 400 MHz (DMSO-d6,
D2O, ppm): d 7.82 (d, 2H, J ¼ 5.32 Hz, pyridyl-H), 8.01 (d, 1H, J ¼
8.64 Hz, quinazolinyl-H), 8.30 (d, 1H, J ¼ 1.20 Hz, quinazolinyl-
N-(4-Iodo-2-{[2-(substituted
acetyl)hydrazinyl]carbonyl}
phenyl)pyridine-4-carboxamide (12e–h). Derivatives 12e–h
were prepared by applying the procedure used for the prepara-
tion of the nicotinamido analogues 12a–d and were recrystal-
lized from ethanol.
N-(4-Iodo-2-{[2-(piperidin-1-ylacetyl)hydrazinyl]carbonyl}phenyl)
pyridine-4-carboxamide (12e). Mp: 204–206 ꢀC. Yield: 45%. I.R.
(KBr, cmꢁ1): ~nmax 3412 (NHs), 3118 (CH aromatic), 2926 (CH2
aliphatic), 1668 (3C]O), 1629 (C]N), 1577 (C]C), 518 (C–I).
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 1.40 (s, 2H,
piperidinyl-H), 1.57 (s, 4H, piperidinyl-H), 3.08 (s, 2H, CH2), 3.32
(s, 4H, piperidinyl-H), 7.83 (d, 2H, J ¼ 5.68 Hz, pyridyl-H), 7.96 (d,
1H, J ¼ 8.92 Hz, phenyl-H), 8.22 (s, 1H, phenyl-H), 7.36 (d, 1H, J
¼ 8.84 Hz, phenyl-H), 8.82 (d, 2H, J ¼ 5.68 Hz, pyridyl-H), 9.93 (s,
1H, NH exchanged by D2O), 10.78 (s, 1H, NH exchanged by D2O),
12.02 (s, 1H, NH exchanged by D2O). Mass (m/z, rel. abundance):
507 (M+, 1.32%), 85 (100%). Anal. calcd for C20H22IN5O3 (507.32):
C, 47.35; H, 4.37; N, 13.80. Found: C, 47.50; H, 4.39; N, 13.89.
N-(4-Iodo-2-{[2-(morpholin-4-ylacetyl)hydrazinyl]carbonyl}phenyl)
yridine-4-carboxamide (12f). Mp: 229–231 ꢀC. Yield: 70%. I.R. (KBr,
cmꢁ1): ~nmax 3444 (NHs), 3120 (CH aromatic), 2927 (CH2 aliphatic),
1697, 1681, 1651 (3C]O), 1598 (C]N), 1510–1440 (C]C), 520
(C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 2.51 (t, 4H, J ¼
1.72 Hz, morpholinyl-H), 3.12 (s, 2H, CH2), 3.63 (t, 4H, J ¼
4.52 Hz, morpholinyl-H), 7.79 (d, 2H, J ¼ 5.96 Hz, pyridyl-H), 7.98
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H), 8.42 (d, 1H, J ¼ 8.80 Hz, quinazolinyl-H), 8.85 (d, 2H, J ¼
Mp: 326–328 ꢀC. Yield: 66%. I.R. (KBr, cmꢁ1): ~nmax 3400
5.32 Hz, pyridyl-H), 12.19 (s, 1H, OH exchanged by D2O). Mass (NH2), 3043 (CH aromatic), 1680 (C]O), 1597 (C]N), 1496 (C]
(m/z, rel. abundance): 365 (M+, 9.23%), 350 (100%). Anal. calcd C), 1085 (C]S), 534 (C–I). H-NMR 400 MHz (DMSO-d6, D2O,
1
for C13H8IN3O2 (365.13): C, 42.76; H, 2.21; N, 11.51. Found: C, ppm): d 7.83 (d, 2H, J ¼ 5.84 Hz, pyridyl-H), 8.00 (dd, 1H, J ¼
42.87; H, 2.24; N, 11.67.
1.76, 8.76 Hz, quinazolinyl-H), 8.30 (d, 1H, J ¼ 1.84 Hz,
6-Iodo-4-oxo-2-(pyridin-3-yl)quinazoline-3(4H)-carboxamide quinazolinyl-H), 8.42 (d, 1H, J ¼ 8.80 Hz, quinazolinyl-H), 8.84
(14a). To a suspension of the benzoxazinone 6a (3.5 g, 10 mmol) (d, 2H, J ¼ 5.84 Hz, pyridyl-H), 12.24 (s, 2H, NH2 exchanged by
in glacial acetic acid (10 mL), anhydrous sodium acetate (1.64 g, D2O). Mass (m/z, rel. abundance): 408 (M+, 0.97%), 350 (100%).
20 mmol) and urea (0.6 g, 10 mmol) were added. The mixture Anal. calcd for C14H9IN4OS (408.22): C, 41.19; H, 2.22; N, 13.72.
was heated under reux for 2 h, cooled to room temperature Found: C, 41.34; H, 2.24; N, 13.80.
and poured on ice/water. The separated solid was ltered,
6-Iodo-4-oxo-2-(pyridin-3-yl)quinazoline-3(4H)-carbothio-
washed with water and recrystallized from methanol. Mp: 307– hydrazide (16a). A solution of the benzoxazine derivative 6a
309 ꢀC. Yield: 71%. I.R. (KBr, cmꢁ1): ~nmax 3153, 3107 (NH2), 3080 (3.5 g, 10 mmol) and thiosemicarbazide (0.75 g, 10 mmol) in
(CH aromatic), 1660 (2C]O), 1606 (C]N), 1516 (C]C), 530 (C– glacial acetic acid (15 mL) in the presence of sodium acetate
I). 1H-NMR 300 MHz (DMSO-d6, D2O, ppm): d 7.63 (dd, 1H, J ¼ (1.64 g, 20 mmol) was reuxed for 6 h. The reaction was poured
4.8, 8.1 Hz, pyridyl-H), 7.99 (dd, 1H, J ¼ 2.10, 8.10 Hz, onto ice/water and the obtained product was ltered off, washed
quinazolinyl-H), 8.26–8.29 (m, 2H, pyridyl-H, quinazolinyl-H), with water and recrystallized from ethanol to give white crystals.
8.42 (d, 1H, J ¼ 8.7 Hz, quinazolinyl-H), 8.81 (d, 1H, J ¼ Mp: 230–232 C. Yield: 71%. I.R. (KBr, cmꢁ1): ~nmax 3290 (NH2),
ꢀ
4.8 Hz, pyridyl-H), 9.11 (s, 1H, pyridyl-H), 12.08 (s, 2H, NH2 3176 (NH), 3088 (CH aromatic), 1685 (C]O), 1591 (C]N), 1498
exchanged by D2O). Mass (m/z, rel. abundance): 394 (M+ + 2, (C]C), 540 (C–I). 1H-NMR 300 MHz (DMSO-d6, D2O, ppm):
85%), 393 (M+ + 1, 1.68%), 392 (M+, 93.42%), 76 (100%). Anal. d 7.55 (d, 1H, J ¼ 8.40 Hz, quinazolinyl-H), 8.05 (d, 1H, J ¼
calcd for C14H9IN4O2 (392.15): C, 42.88; H, 2.31; N, 14.29. 7.80 Hz, pyridyl-H), 8.12–8.20 (m, 1H, pyridyl-H), 8.22 (d, 1H, J ¼
Found: C, 43.01; H, 2.28; N, 14.54.
2.10 Hz, pyridyl-H), 8.46 (d, 1H, J ¼ 4.00 Hz, quinazolinyl-H),
6-Iodo-4-oxo-2-(pyridin-4-yl)quinazoline-3(4H)-carboxamide 8.69 (dd, 1H, J ¼ 4.8, 12.90 Hz, quinazolinyl-H), 8.85 (d, 1H, J
(14b). Compound 14b was synthesized according to the adopted ¼ 20.40, pyridyl-H), 10.17 (s, 2H, NH2 exchanged by D2O), 10.43
procedure for 14a using (pyridin-4-yl)-benzoxazin-4-one (6b) (s, 1H, NH exchanged by D2O). Mass (m/z, rel. abundance): 423
and was recrystallized from methanol. Mp: 330–332 C. Yield: (M+, 0.67%), 349 (100%). Anal. calcd. for C14H10IN5OS (423.23):
ꢀ
70%. I.R. (KBr, cmꢁ1): ~nmax 3099, 3066 (NH2), 3043 (CH C, 39.73; H, 2.38; N, 16.55. Found: C, 39.89; H, 2.41; N, 16.79.
aromatic), 1681 (2C]O), 1597 (C]N), 1496 (C]C), 534 (C–I).
6-Iodo-4-oxo-2-(pyridin-4-yl)quinazoline-3(4H)-carbothio-
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.83 (d, 2H, J ¼ hydrazide (16b). Compound 16b was prepared by applying the
5.80 Hz, pyridyl-H), 8.01 (dd, 1H, J ¼ 1.72, 8.72 Hz, quinazolinyl- same procedure used for preparation of 16a and was recrystal-
H), 8.30 (d, 1H, J ¼ 1.88 Hz, quinazolinyl-H), 8.42 (d, 1H, J ¼ lized from ethanol to give beige crystals. Mp: 210–212 ꢀC. Yield:
8.80 Hz, quinazolinyl-H), 8.76 (d, 2H, J ¼ 5.80 Hz, pyridyl-H), 60%. I.R. (KBr, cmꢁ1): ~nmax 3290 (NH2), 3242 (NH), 3062 (CH
12.17 (s, 1H, NH2 exchanged by D2O). Mass (m/z, rel. abun- aromatic), 1681 (C]O), 1591 (C]N), 1504 (C]C), 540 (C–I). 1H-
dance): 392 (M+, 0.14%), 78 (100%). Anal. calcd for C14H9IN4O2 NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.55–7.60 (m, 1H,
(392.15): C, 42.88; H, 2.31; N, 14.29. Found: C, 43.07; H, 2.32; N, quinazolinyl-H), 7.70 (d, 2H, J ¼ 5.12 Hz, pyridyl-H), 8.22 (d, 1H,
14.51.
J ¼ 8.52 Hz, quinazolinyl-H), 8.48 (s, 1H, quinazolinyl-H), 8.75
6-Iodo-4-oxo-2-(pyridin-3-yl)quinazoline-3(4H)-carbothioa- (d, 2H, J ¼ 5.12 Hz, pyridyl-H), 10.16 (s, 1H, NH2 exchanged by
mide (15a). A mixture of benzoxazinone 6a (3.5 g, 10 mmol) and D2O), 10.43 (s, 1H, NH exchanged by D2O). Mass (m/z, rel.
thiourea (0.76 g, 10 mmol) in glacial acetic acid (15 mL) con- abundance): 423 (M+, 0.61%), 64 (100%). Anal. calcd. for
taining sodium acetate anhydrous (1.64 g, 20 mmol), was
C14H10IN5OS (423.23): C, 39.73; H, 2.38; N, 16.55. Found: C,
reuxed for 4 h. Upon pouring on crushed ice/water, beige 39.85; H, 2.37; N, 16.68.
powder was obtained, ltered, washed with water and recrys-
6-Iodo-3-(4-un/substituted phenylamino)-2-(pyridin-3-yl)qui-
tallized from ethanol. Mp: 303–305 ꢀC. Yield: 70%. I.R. (KBr, nazolin-4(3H)-one (17a–c). To a suspension of benzoxazinone
cmꢁ1): ~nmax 3365 (NH2), 1666 (C]O), 1606 (C]N), 1516 (C]C), 6a (3.5 g, 10 mmol) in absolute ethanol (20 mL), the required
1159 (C]S), 530 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, hydrazine derivative (10 mmol) was added and the reaction
ppm): d 7.54 (d, 1H, J ¼ 8.56 Hz, quinazolinyl-H), 7.58 (dd, 1H, J mixture was heated under reux for 8–12 h. Then concentrated.
¼ 4.84, 7.88 Hz, pyridyl-H), 8.12 (dd, 1H, J ¼ 1.36, 8.48 Hz, Upon cooling the separated solid mass was ltered, washed
quinazolinyl-H), 8.42–8.48 (m, 2H, quinazolinyl-H, pyridyl-H), with water and recrystallized from absolute ethanol to afford
8.76 (d, 1H, J ¼ 4.28 Hz, pyridyl-H), 9.28 (d, 1H, J ¼ 1.44 Hz, the titled derivatives 17a–c.
pyridyl-H), 12.86 (s, 2H, NH2 exchanged by D2O). Mass (m/z, rel.
6-Iodo-3-(phenylamino)-2-(pyridin-3-yl)quinazolin-4(3H)-one (17a).
abundance): 408 (M+, 44.59%), 78 (100%). Anal. calcd for Mp: 176–178 ꢀC. Yield: 55%. I.R. (KBr, cmꢁ1): ~nmax 3325 (NH), 2922
C
14H9IN4OS (408.22): C, 41.19; H, 2.22; N, 13.72. Found: C, (CH aromatic), 1681 (C]O), 1645 (C]N), 1598 (C]C), 520 (C–I).
41.30; H, 2.18; N, 13.85.
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 6.74 (t, 1H, J ¼ 7.28 Hz,
phenyl-H), 6.84 (d, 2H, J ¼ 7.72 Hz, phenyl-H), 7.16 (t, 2H, J ¼
6-Iodo-4-oxo-2-(pyridin-4-yl)quinazoline-3(4H)-carbothioa-
mide (15b). This compound was prepared and puried as per 7.72 Hz, phenyl-H), 7.59 (dd, 1H, J ¼ 4.84, 7.92 Hz, pyridyl-H), 7.98
the previously mentioned procedure.
(d, 1H, J ¼ 8.80 Hz, quinazolinyl-H), 8.20–8.23 (m, 1H, pyridyl-H),
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8.30–8.32 (m, 2H, quinazolinyl-H), 8.78 (dd, 1H, J ¼ 1.16, 4.68 Hz, Anal. calcd for C19H12FIN4O (458.23): C, 49.80; H, 2.64; N, 12.23.
pyridyl-H), 9.05 (d, 1H J ¼ 1.76 Hz, pyridyl-H), 12.03 (s, 1H, NH Found: C, 49.98; H, 2.67; N, 12.39.
exchanged by D2O). Mass (m/z, rel. abundance): 440 (M+, 100%).
3-[(4-Chlorophenyl)amino]-6-iodo-2-(pyridin-4-yl)quinazolin-
Anal. calcd for C19H13IN4O (440.24): C, 51.84; H, 2.98; N, 12.73. 4(3H)-one (17f). Mp: 259–261. Yield: 51%. I.R. (KBr, cmꢁ1): ~nmax
Found: C, 51.99; H, 3.03; N, 12.94. 3290 (NH), 3180 (CH aromatic), 1656 (C]O), 1643 (C]N), 1483
3-[(4-Fluorophenyl)amino]-6-iodo-2-(pyridin-3-yl)quinazolin-4(3H)- (C]C), 755 (C–Cl), 520 (C–I). 1H-NMR 300 MHz (DMSO-d6,
one (17b). Mp: 310–312 ꢀC. Yield: 52%. I.R. (KBr, cmꢁ1): ~nmax 3325 ppm): d 7.35 (d, 2H, J ¼ 7.80 Hz, phenyl-H), 7.45 (d, 2H, J ¼
(NH), 2924 (CH aromatic), 1681 (C]O), 1645 (C]N), 1598 (C]C), 7.80 Hz, phenyl-H), 7.55 (d, 1H, J ¼ 8.70 Hz, quinazolinyl-H),
501 (C–I). 1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 7.00 (dd, 2H, 8.01 (d, 2H, J ¼ 6.00 Hz, pyridyl-H), 8.20 (s, 1H, quinazolinyl-
J ¼ 4.52, 8.88 Hz, phenyl-H), 7.17 (t, 2H, J ¼ 4.72, 7.76 Hz, phenyl- H), 8.69 (d, 2H, J ¼ 6.00 Hz, pyridyl-H), 9.27 (s, 1H,
H), 7.65 (dd, 1H, J ¼ 4.72, 7.76 Hz, pyridyl-H), 8.01 (dd, 1H, J ¼ quinazolinyl-H), 10.84 (s, 1H, NH exchanged by D2O). Mass (m/
1.88, 8.68 Hz, quinazolinyl-H), 8.27–8.30 (m, 2H, pyridyl-H, z, rel. abundance): 474 (M+, 35.35%), 476 (M+ + 2, 11.89%), 126
quinazolinyl-H), 8.42 (d, J ¼ 8.76 Hz, 1H, quinazolinyl-H), 8.82 (100%). Anal. calcd for C19H12ClIN4O (474.68): C, 48.07; H, 2.55;
(d, J ¼ 3.80 Hz, 1H, pyridyl-H), 9.11 (s, 1H, pyridyl), 12.06 (s, 1H, N, 11.80. Found: C, 48.31; H, 2.60; N, 11.94.
NH exchanged by D2O). 13C-NMR (100 MHz DMSO-d6, ppm):
4-(6-Iodo-4-oxo-2-(pyridin-3-yl)quinazolin-3(4H)-yl)benzoic
d 79.46, 87.38, 116.10, 116.33, 117.07, 117.15, 120.06, 123.14, acid (18a). Equimolar amounts of benzoxazin-4-one 6a (0.70 g, 2
124.72, 135.95, 139.56, 140.22, 142.95, 148.18, 152.71, 162.21 (C– mmol) and p-aminobenzoic acid (0.28 g, 2 mmol) were fused
F), 168.70 (C]O). Mass (m/z, rel. abundance): 458 (M+, 8.24%), 78 together at 200 ꢀC in an oil bath for 1 h. The mixture was cooled,
(100%). Anal. calcd for C19H12FIN4O (458.23): C, 49.80; H, 2.64; N, dissolved in boiling glacial acetic acid (30 mL) and ltered; the
12.23. Found: C, 49.95; H, 2.68; N, 12.40.
ltrate was concentrated up to (10 mL) in vacuo and cooled. The
resulting solid was ltered, washed with water, dried and
3-[(4-Chlorophenyl)amino]-6-iodo-2-(pyridin-3-yl)quinazolin-
4(3H)-one (17c). Mp: 254–256 C. Yield: 65%. I.R. (KBr, cmꢁ1): recrystallized from glacial acetic acid. Mp: 307–309 ꢀC. Yield:
~nmax 3265 (NH), 3082 (CH aromatic), 1681 (C]O), 1583 (C]N), 50%. I.R. (KBr, cmꢁ1): ~nmax 3292 (OH), 1685 (2C]O), 1635 (C]
1487 (C]C), 520 (C–I). 1H-NMR 300 MHz (DMSO-d6, D2O, ppm): N), 1508 (C]C), 540 (C–I). 1H-NMR 300 MHz (DMSO-d6, ppm):
d 6.67 (d, 2H, J ¼ 8.70 Hz, phenyl-H), 7.15 (d, 2H, J ¼ 8.70 Hz, d 7.58 (dd, 1H, J ¼ 4.80, 7.80 Hz, pyridyl-H), 7.82 (d, 2H, J ¼
phenyl-H), 7.47 (d, 1H, J ¼ 6.78 Hz, pyridyl-H), 7.68 (d, 1H, J ¼ 8.70 Hz, phenyl-H), 7.92 (d, 2H, J ¼ 8.70 Hz, phenyl-H), 7.99 (d,
8.40 Hz, quinazolinyl-H), 8.11 (d, 1H, J ¼ 7.80 Hz, pyridyl-H), 1H, J ¼ 2.10 Hz, quinazolinyl-H), 8.20–8.23 (m, 1H, pyridyl-H),
8.22 (d, 1H, J ¼ 8.70 Hz, quinazolinyl-H), 8.24 (s, 1H, 8.29 (d, 1H, J ¼ 1.80 Hz, quinazolinyl-H), 8.40 (s, 1H,
quinazolinyl-H), 8.65 (s, 1H, pyridyl-H), 8.92 (s, 1H, pyridyl-H), quinazolinyl-H), 8.76 (d, 1H, J ¼ 4.20 Hz, pyridyl-H), 9.04 (s, 1H,
9.30 (s, 1H, NH exchanged by D2O). Mass (m/z, rel. abun- J ¼ 1.76 Hz, pyridyl-H), 11.23 (s, 1H, OH exchanged by D2O).
dance): 474 (M+, 49.72%), 476 (M+ + 2, 17.35%), 349 (100%). Mass (m/z, rel. abundance): 469 (M+, 1.01%), 78 (100%). Anal.
Anal. calcd for C19H12ClIN4O (474.68): C, 48.07; H, 2.55; N, calcd for C20H12IN3O3 (469.23): C, 51.19; H, 2.58; N, 8.96.
ꢀ
11.80. Found: C, 48.28; H, 2.61; N, 11.94.
Found: C, 51.34; H, 2.60; N, 9.08.
6-Iodo-3-(4-un/substituted phenylamino)-2-(pyridin-4-yl)qui-
4-(6-Iodo-4-oxo-2-(pyridin-4-yl)quinazolin-3(4H)-yl)benzoic
nazolin-4(3H)-one (17d–f). The quinazolinones 17d–f were acid (18b). This compound was prepared and puried as per the
prepared and puried according to the previously mentioned previously mentioned procedure, using the benzoxazinone
procedure starting from the benzoxazinone 6b.
derivative 6b. Mp: 244–246 ꢀC. Yield: 55%. I.R. (KBr, cmꢁ1): ~nmax
6-Iodo-3-(phenylamino)-2-(pyridin-4-yl)quinazolin-4(3H)-one (17d). 3334 (OH), 1681 (2C]O), 1589 (C]N), 1506 (C]C), 519 (C–I).
Mp: 239–241 ꢀC. Yield: 40%. I.R. (KBr, cmꢁ1): ~nmax 3304 (NH), 3057 1H-NMR 300 MHz (DMSO-d6, ppm): d 7.35 (d, 2H, J ¼ 7.80 Hz,
(CH aromatic), 1672 (C]O), 1647 (C]N), 1519 (C]C), 534 (C–I). phenyl-H), 7.45 (d, 2H, J ¼ 7.80 Hz, phenyl-H), 7.55 (d, 1H, J ¼
1H-NMR 400 MHz (DMSO-d6, D2O, ppm): d 6.74 (t, 1H, J ¼ 7.28 Hz, 8.70 Hz, quinazolinyl-H), 8.01 (d, 2H, J ¼ 6.00 Hz, pyridyl-H),
phenyl-H), 6.84 (d, 2H, J ¼ 7.88 Hz, phenyl-H), 7.16 (t, 2H, J ¼ 8.20 (d, 1H, J ¼ 8.64 Hz, quinazolinyl-H), 8.69 (d, 2H, J ¼
7.64 Hz, phenyl-H), 7.77 (d, 2H, J ¼ 5.68 Hz, pyridyl-H), 7.99 (s, 1H, 6.00 Hz, pyridyl-H), 9.27 (s, 1H, quinazolinyl-H), 12.12 (s, 1H,
quinazolinyl-H), 8.31–8.33 (m, 2H, quinazolinyl-H), 8.80 (d, 2H, J ¼ OH exchanged by D2O). Mass (m/z, rel. abundance): 469 (M+,
5.68 Hz, pyridyl-H), 10.77 (s, 1H, NH exchanged by D2O). Anal. 42.74%), 78 (100%). Anal. calcd for C20H12IN3O3 (469.23): C,
calcd for C19H13IN4O (440.24): C, 51.84; H, 2.98; N, 12.73. Found: C, 51.19; H, 2.58; N, 8.96. Found: C, 51.31; H, 2.62; N, 9.03.
51.97; H, 2.96; N, 12.91.
3-[(4-Fluorophenyl)amino]-6-iodo-2-(pyridin-4-yl)quinazolin-
4(3H)-one (17e). Mp: 234–236 C. Yield: 42%. I.R. (KBr, cmꢁ1):
(2) Biology
ꢀ
~nmax 3240 (NH), 2922 (CH aromatic), 1678 (C]O), 1645 (C]N),
(a) In vitro cytotoxicity. The in vitro cytotoxicity of the newly
1598 (C]C), 1230 (C–F), 501 (C–I). 1H-NMR 400 MHz (DMSO-d6, synthesized compounds against a panel of nine cancer cell lines
ppm): d 6.05 (d, 1H, J ¼ 8.76 Hz, quinazolinyl-H), 7.09 (t, 2H, J ¼ and one non-tumorigenic cell line was assessed using the MTT
8.80 Hz, phenyl-H), 7.30 (dd, 2H, J ¼ 4.72, 8.84 Hz, phenyl-H), assay according to Mosmann's method.41 The MTT assay which
7.52 (d, 2H, J ¼ 5.92 Hz, pyridyl-H), 7.56 (d, 1H, J ¼ 1.72 Hz, was developed in 1983, is based on the reduction of the soluble
quinazolinyl-H), 8.19 (d, 1H, J ¼ 1.76 Hz, quinazolinyl-H), 8.52 3-(4,5-methyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide
(d, 2H, J ¼ 5.92 Hz, pyridyl-H), 10.06 (s, 1H, NH exchanged by (MTT) into a blue-purple formazan product, mainly by mito-
D2O). Mass (m/z, rel. abundance): 458 (M+, 30.88%), 110 (100%). chondrial reductase activity inside living cells. Mosmann
111784 | RSC Adv., 2016, 6, 111767–111786
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showed that MTT assay has very wide applicability for ltermat and washed three times for 5 minutes in 75 mM
measuring survival and/or proliferation of various cells and can phosphoric acid and once in methanol prior to drying and
potentially be applied to any assay in which living cells must be scintillation counting.
distinguished from dead cells or a lack of cells. The assay also
has potential value for quantitative and rapid measurement of
(3) Molecular docking
cell death.
The molecular modeling study was carried out using Molecular
The cells used in cytotoxicity assay were cultured in RPMI
Operating Environment (MOE 2009.10) soware provided by
chemical computing group, Canada. The energy minimization
for the compounds was performed with MOE with MMFF94X
force eld and the partial charges were automatically
calculated.43,44
1640 medium supplemented with 10% fetal calf serum. Cells
suspended in the medium (2 ꢄ 104/mL) were plated in 96-well
ꢀ
culture plates and incubated at 37 C in a 5% CO2 incubator.
Aer 12 h, the test sample (2 mL) was added to the cells in 96-
ꢀ
well plates and cultured at 37 C for 3 days.
Crystallographic complexes of ABL with its respective
inhibitors imatinib and bosutinib (PDB IDs: 2HYY and 3UE4)
were downloaded from the protein data bank available at http://
www.rcsb.org/pdb. The protein-ligand complex obtained from
the protein data bank was prepared for docking as follows: the
enzyme was 3D protonated, where hydrogen atoms were added
at their standard geometry, the partial charges were computed
and the system was optimized.
Deletion of water of crystallization together with extra chains
of the protein was then done. Finally, isolation of the active site
and recognition of the involved amino acids was carried out
then the backbone was hidden.
Docking was performed using Triangle Matcher placement
method, poses were prioritized based on London dG scoring
and renement of the results was done using forceeld. The
most stable docking model was selected per the best scored
conformation predicted by the MOE scoring function.
The cultured cells were mixed with 20 mL of MTT solution
and incubated for 4 h. at 37 C. The supernatant was carefully
ꢀ
removed from each well and 100 mL of DMSO were added to
each well to dissolve the formazan crystals which were formed
by the cellular reduction of MTT. Aer mixing with a mechan-
ical plate mixer, the absorbance of each well was measured by
a microplate reader using a test wavelength of 570 nm.
(b) In vivo cytotoxicity. Female athymic pathogen-free nude
mice (nu/nu, 4–6 weeks) were purchased and maintained in
accordance with the guidelines of the so adopted Guide for the
Care and Use of Laboratory Animals 8th Edition 2011, and as
approved by the Research Ethics Committee (REC) of Faculty of
Pharmacy, Cairo University. To establish MCF-7 human breast
cancer xenogras, each of the female nude mice was rst
implanted with a 60 day subcutaneously (s.c.) slow release
estrogen pellet (SE-121, 1.7 mg 17b-estradiol/pellet). The next
day, cultured MCF-7 cells were harvested from conuent
monolayer cultures, washed twice with serum-free medium,
resuspended and injected s.c. (5 ꢄ 106 cells, total volume 0.2
mL) into the le inguinal area of the mice. All animals were
monitored for activity, physical condition, body weight, and
tumor growth. Tumor size was determined by caliper
measurement in two perpendicular diameters of the implant
every other day. Tumor volume was calculated by the formula, 1/
2a ꢄ b2 where “a” is the long diameter and “b” is the short
diameter (in cm).
Conflict of interest
The authors declare no conict of interest.
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afenib
were
dissolved
in
PEG400 : ethanol : saline
(57.1 : 14.3 : 28.6, v/v/v), and were administered by intraperito-
neal (i.p.) injection at doses equivalent of 5 or 10 mg kgꢁ1, every
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based on previous reports of sorafenib.42
(c) Kinase inhibitory assay. In a nal reaction volume of 25
mL, kinase (5–10 mU) was incubated with 25 mM Tris pH 7.5,
0.02 mM EGTA, 0.66 mg mLꢁ1 myelin basic protein, 10 mM
magnesium acetate and [g33P-ATP] (specic activity approxi-
mately 500 cpm pmolꢁ1, concentration as required). The reac-
tion was initiated by the addition of the Mg–ATP mixture. Aer
incubation for 40 minutes at room temperature, the reaction
was stopped by the addition of 5 mL of a 3% phosphoric acid
solution. 10 mL of the reaction was then spotted onto a P30
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Paper
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