Synthesis and evaluation of c-Src kinase inhibitory activity of pyridin-2(1H)-one derivatives

Article abstract of DOI:10.1016/j.bioorg.2014.02.001

Src kinase, a prototype member of the Src family of kinases (SFKs), is over-expressed in various human tumors, and has become a target for anticancer drug design. In this perspective, a series of eighteen 2-pyridone derivatives were synthesized and evaluated for their c-Src kinase inhibitory activity. Among them, eight compounds exhibited c-Src kinase inhibitory activity with IC ₅₀ value of less than 25 μM. Compound 1-[2-(dimethylamino)ethyl]- 5-(2-hydroxy-4-methoxybenzoyl)pyridin-2(1H)-one (36) exhibited the highest c-Src kinase inhibition with an IC₅₀ value of 12.5 μM. Furthermore, the kinase inhibitory activity of compound 36 was studied against EGFR, MAPK and PDK, however no significant activity was observed at the highest tested concentration (300 μM). These results provide insights for further optimization of this scaffold for designing the next generation of 2-pyridone derivatives as candidate Src kinase inhibitors.

Full text of DOI:10.1016/j.bioorg.2014.02.001

Bioorganic Chemistry 53 (2014) 75–82
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Synthesis and evaluation of c-Src kinase inhibitory activity
of pyridin-2(1H)-one derivatives
Karam Chand a, Suchita Prasad a, Rakesh K. Tiwari b,c, Amir N. Shirazi b,c, Sumit Kumar a,d
,
Keykavous Parang b,c, , Sunil K. Sharma a,
⇑
⇑
a Department of Chemistry, University of Delhi, Delhi 110007, India
b Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, The University of Rhode Island, Kingston, RI 02881, USA
c School of Pharmacy, Chapman University, One University Drive, Orange, CA 92866, USA
d Department of Chemistry, Deenbandhu Chhotu Ram University of Science & Technology, Murthal 131039, Haryana, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 23 September 2013
Available online 17 February 2014
Src kinase, a prototype member of the Src family of kinases (SFKs), is over-expressed in various human
tumors, and has become a target for anticancer drug design. In this perspective, a series of eighteen 2-pyri-
done derivatives were synthesized and evaluated for their c-Src kinase inhibitory activity. Among them,
eight compounds exhibited c-Src kinase inhibitory activity with IC50 value of less than 25
1-[2-(dimethylamino)ethyl]-5-(2-hydroxy-4-methoxybenzoyl)pyridin-2(1H)-one (36) exhibited the
highest c-Src kinase inhibition with an IC50 value of 12.5 M. Furthermore, the kinase inhibitory activity
of compound 36 was studied against EGFR, MAPK and PDK, however no significant activity was observed
at the highest tested concentration (300 M). These results provide insights for further optimization of
lM. Compound
Keywords:
Chromone
Pyridin-2(1H)-one
Synthesis
c-Src kinase inhibition
l
l
this scaffold for designing the next generation of 2-pyridone derivatives as candidate Src kinase inhibitors.
Ó 2014 Elsevier Inc. All rights reserved.
1. Introduction
Activation of PTKs has been shown to be critical in neoplasia pro-
gress [4]. Thus, inhibition of PTKs has become a major strategy in
Cancer is considered to be a multi-step process, occurring
through an accumulation of intrinsic or extrinsic errors in respon-
sible genes for the regulation of cell proliferation and survival. The
development of human cancer can be triggered with different
genes [1]. Limited abnormalities are sufficient for the development
of neoplasia that is an abnormal growth of cells. The alarming in-
crease in the cancer patients worldwide has led an unprecedented
pressure on researchers to explore novel active pharmacophores
with higher bioactivity, selectivity, and minimal toxicity.
drug design against cancer [3].
The Src family of kinases (SFKs) are non-receptor tyrosine ki-
nases that are involved in signal transduction in cancer cells. c-
Src is a member of SFKs which has been reported to induce STATs
involved in the tumorigenesis process [5]. STAT3 is a member of
signal transducer and activator of transcription protein family that
regulates cell growth, survival and differentiation and has been
associated with various human cancers. It has been observed that
the activity of c-Src kinase in human mammary carcinomas is 4
to 20-fold greater than that in normal cells [6]. Increased Src activ-
ity elevates the cell growth rate and reduces adhesion between
cells, leading to the development of metastatic potential of cells
[7–9]. As a result, c-Src kinase plays an important role in the gen-
esis and progression of human cancers, including carcinomas of the
breast, colon, prostate, lung, ovary, and in myeloproliferative disor-
ders [10–12]. Thus, the design and discovery of novel and potent c-
Src kinase inhibitors remains critically important.
PTKs are enzymes that catalyze the phosphorylation of the hy-
droxyl groups of tyrosine residues in various proteins by the trans-
fer of the c
-phosphate of the ATP–Mg2+ complex to the said amino
acid side chain [2]. PTKs are key regulators of various cell functions,
such as cellular growth, proliferation, migration, differentiation,
and apoptosis [3]. Due to their physiological relevance, variety
and ubiquity, PTKs have become a subject of extensive study.
We have previously designed and synthesized several novel
derivatives of benzopyran-2-one (coumarin) [13,14] and benzopy-
ran-4-one (chromone) [15] scaffold and evaluated their antiprolif-
erative and c-Src kinase inhibitory activity. Finding new Src kinase
inhibitors remains a challenging task. A more practical approach to
such challenges encompasses modification of the structure of
⇑
Corresponding authors. Address: Department of Chemistry, University of Delhi,
Delhi 110 007, India. Fax: +91 11 27666950 (S.K. Sharma). 7 Greenhouse Road,
Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy,
University of Rhode Island, Kingston, RI 02881, USA. Fax: +1 401 874 5787 (K. Parang).
E-mail addresses: kparang@uri.edu (K. Parang), sk.sharma90@gmail.com
(S.K. Sharma).
http://dx.doi.org/10.1016/j.bioorg.2014.02.001
0045-2068/Ó 2014 Elsevier Inc. All rights reserved.

76
K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
Fig. 1. Some of the 2- and 4-pyridone derivatives active against protein kinases.
existing active pharmacophores. Pyridone skeleton has been
known as an ideal frame for further exploration. Recently, myriad
pyridone derivatives (Fig. 1) have been tested for their potency
against various protein kinases including Pim-1 kinase (1) [16], Fo-
cal adhesion kinase (FAK) (2) [17], MEK1 (3 & 4) [18,19], Met ki-
nase (5, 6 & 7) [20–23] and Checkpoint kinase 1 (CHK1) (8) [24].
Furthermore, 2-pyridone scaffold has been screened against Src
kinases. Some 2-pyridone derivatives such as aryl aminoquinazo-
linepyridone (9) [25], pyrido[2,3-d]pyrimidine (10) [26,27] and
pyrido-propanamide (11) [28] have been reported as potent Src ki-
nase inhibitors. Thus, the wealth of information for Src kinases and
pyridone skeleton obtained from literature provided a strong ratio-
nale for considering inhibition of this target using pyridones to
treat cancer. In the light of the above literature reports and in con-
tinuation of our efforts to explore new scaffolds as c-Src kinase
inhibitors, herein, we report the synthesis and evaluation of c-Src
kinase inhibitory activity of a class of novel 2-pyridone derivatives.
(23–27) with various alkylamines, N,N-dialkylaminoalkanes, and
t-butyl (2-aminoethyl)carbamate in the presence of triethylamine
and using ethanol as solvent (Scheme 2).
The key intermediates (20–22), in turn were synthesized from
corresponding hydroxyacetophenones by following the method
reported earlier from our group [29]. In the case of dihydroxyaceto-
phenone (13/14), first mono-O-acetylation was carried out using ace-
tic anhydride and pyridine while o-hydroxyacetophenone (12) was
used as such. 4-Oxo-4H-1-chromen-3-yl-carbaldehydes (17–19) were
then synthesized using Vilsmeir–Haack formylation reaction. Since
o-hydroxyacetophenone (12) and its derivatives containing various
substituents takes a ring form due to H-bonding and thus prohibit
enolization, therefore these compounds can be doubly formylated
using Vilsmeir–Haack reagent to get 3-formyl substituted chromone
derivatives [30]. The formylation reaction was followed by the Knoe-
venagel condensation with malonic acid to yield the respective 4-oxo-
4H-chromen-3-yl)acrylic acid (20–22) (Scheme 1).
The desired pyridone precursors i.e. compound 23–25 were ob-
tained by esterification of acrylyl acid derivatives of 4-oxo-4H-1-
benzopyran (20–22) with ethanol under acidic condition
(Scheme 1). The methylation of phenolic group for compounds
24 and 25 with methyl iodide under basic conditions gave (E)-ethyl
3-(7/6-methoxy-4-oxo-4H-chromen-3-yl)acrylates (26/27). All of
the compounds were well characterized from their physical and
2. Results and discussion
2.1. Chemistry
A class of novel 2-pyridone derivatives (28–45) were synthe-
sized by reacting (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylates
Scheme 1. Synthesis of (E)-alkyl 3-(4-oxo-4H-chromen-3-yl)acrylate; Reagents and conditions: a) Ac2O, pyridine, 6 h; b) POCl3, DMF, 50 °C, 13 h; c) CH2(COOH)2, pyridine,
1.5 h; d) EtOH, conc. H2SO4 (3 or 4 drops), 12 h; e) CH3I, K2CO3, anhyd. acetone, reflux, 12 h.

K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
77
Scheme 2. Synthesis of pyridin-2(1H)-one derivatives; Reagents and conditions: a) R1NH2 (1.1 eq), NEt3, C2H5OH, reflux, 8–10 h; b) R1NH2 (2.4 eq), NEt3, C2H5OH, reflux,
16–17 h; c) NEt3, C2H5OH, morpholine/aniline/fluoroaniline, reflux, 12–13 h.
Fig. 2. c-Src Kinase inhibitory activity of 2-pyridone derivatives (28–45). IC50 is the concentration at which the enzyme activity is inhibited by 50% and is calculated from
Graph Prism software. All the experiments were carried out in triplicate.
spectral data and by comparing the data with literature value for
the known compounds.
2.2. Biology
The t-butyl (2-aminoethyl)carbamate used in the reaction was
synthesized according to the literature procedure and character-
ized by comparing its physical and spectral data with the literature
values [31].
When (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylates (23–27)
were reacted with 1.1 eq. of alkyl amines, then monoalkylated
products 28–43 were obtained (Scheme 2). However, by reacting
ester 23 with alkylamines in the molar ratio of 1:2, dialkyl prod-
ucts 44–45 were obtained. Furthermore, secondary and aromatic
amines e.g. morpholine, aniline, and fluoroaniline followed a dif-
ferent reaction pathway. For these secondary and aromatic amines
we observed the nucleophilic addition of the amine across the dou-
2.2.1. c-Src kinase inhibitory activity
Fig. 2 shows the c-Src inhibitory potency of all of the pyridin-
2(1H)-one derivatives (28–45) compared to a standard protein ki-
nase inhibitor, Staurosporine, and a Src kinase inhibitor, PP2. These
compounds exhibited modest c-Src kinase inhibitory activity.
Among eighteen compounds, eight were found to have IC50 values
below 25
hydroxy-4-methoxybenzoyl)pyridin-2(1H)-one (36) was found to
be the most potent with IC50 value of 12.5 M as shown in
Table S1 (Supporting information). The compounds 38 and 45
showed also significant activity with IC50 values 19.9 M and
20.1 M, respectively.
lM. The compound 1-[2-(dimethylamino)ethyl]-5-(2-
l
l
l
ble bond of
a
, b-unsaturated ester. The addition of amine occurred
In general, among all compounds containing pyridin-2(1H)-one
template, compound 36 having hydroxy and methoxy groups at
meta positions on the phenyl ring and the pyridone ring linked to
2-(dimethylamino)ethyl group was found to be the most potent.
The presence of the dimethylamine was found to be important
since other derivatives with similar structures including com-
pounds 34, 35, and 37 showed higher IC50 values of 34.1, 47.0,
at the b-carbon (C-3) due to electron withdrawing influence of the
adjacent carbonyl group of acrylate 23. Thus, nucleophilic addition
gave the compounds 46–48 (Scheme 2). The structure of these
compounds were confirmed by their 1H NMR and 13C NMR spec-
trum, and comparing the data with the theoretical NMR obtained
by Mestrenova version 5.3.

78
K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
Table 1
done derivatives, compound 36 was found to be the most potent c-
Src kinase inhibitor (IC50: 12.5 M), however it did not exhibit ki-
nase inhibition activity against three other kinases studied namely
Inhibitory activity of compound 36 against other
kinases.
l
Kinase
IC50 (lM)
EGFR, MAPK and PDK at the highest tested concentration of
300 lM. In the antiproliferative activity assay, a modest inhibition
EGFR(h)
MAPK1(h)
PDK1(h)
>300
>300
>300
potency was exhibited by compounds 35 and 41 with the prolifer-
ation of CCRF-CEM cells by 35% and 53% respectively. However,
none of the compounds synthesized have any significant antiprolif-
erative activity against SK-OV-3 and MCF-7 cells, thus establishing
a weak correlation between Src kinase inhibition and antiprolifer-
ative activity. Structure-activity relationship of 2-pyridone deriva-
tives for Src kinase inhibition has not been studied extensively,
hence these results can be used for further optimization of 2-pyri-
dones for designing and investigation of the potentiality of these
compounds as the lead potent and selective Src kinase inhibitors.
and 27.6 lM, respectively. The nature of substituent in the phenyl
ring was found to be critical i.e. the methoxy group on the phenyl
ring appeared to be involved in contributing to Src kinase inhibi-
tory activity. The compound 31 that lacked a methoxy group
showed higher IC50 value (28.2 lM) when compared with the cor-
responding methoxy analog 36. Similarly, while comparing the
compounds 33 and 37 which differed in terms of presence of hy-
droxyl and methoxy groups, respectively, at C-4 position of phenyl
ring, higher c-Src kinase inhibitory activity was observed for com-
4. Experimental section
pound 37 (IC50: 27.6
lM) in comparison to compound 33 (IC50:
4.1. Materials and methods
57.8 M). Also, by comparing the IC50 values of pyridones reported
l
herein with that of coumarins [13] and chromones [15] published
earlier from our group, it was observed that 2-pyridones have sig-
nificantly higher c-Src kinase inhibitory activities.
The organic solvents were dried and distilled prior to their use.
Reactions were monitored by precoated TLC plates (Merck silica
gel 60F254); the spots were visualized either by UV light, or by
spraying with 5% alcoholic FeCl3 solution. Silica gel (100–
200 mesh) was used for column chromatography. All of the chem-
icals and reagents were procured from Spectrochem Pvt. Ltd., India
and Sigma–Aldrich Chemicals Pvt. Ltd., USA. Melting points were
measured on a Buchi M-560 apparatus and are uncorrected.
Infrared spectra were recorded on Perkin-Elmer FT-IR model 9
spectrophotometer. The 1H and 13C NMR spectra were recorded
on Jeol-400 (400 MHz, 100.5 MHz) NMR spectrometer and
Avance-300 (300 MHz, 75.5 MHz) spectrometer using tetramethyl-
silane as internal standard. The chemical shift values are on a d
scale and the coupling constant values (J) are in Hertz. The UV data
were recorded on Analytik Jena SPECORD 250 and Perkin-Elmer
Lambda 35. The HRMS data were recorded on Agilent-6210 ES-
TOF, JEOL JMX-SX-102A and Waters LCT Micromass-KC455.
2.2.2. Inhibitory activity aganist EGFR, MAPK and PDK
In order to further explore the selectivity of compound 36 (most
active compound for c-Src kinase inhibition) against other kinases,
three different kinases namely Epidermal Growth Factor Receptor
(EGFR), Mitogen-Activated Protein Kinase (MAPK) and Phosphoin-
ositide-Dependent Kinase (PDK) were chosen as the target. It was
observed that the compound 36 did not show any kinase inhibition
at the highest tested concentration of 300 lM and thus it can be
established that this compound has selective activity against c-
Src kinase (1).
Also, in continuation of our efforts to get further insights about
the various 2-pyridone derivatives synthesized, the antiprolifera-
tive screening was carried out. However, the majority of com-
pounds did not show significant antiproliferative potency
compared to the positive control (Dox) at the concentration of
50 lM after 72 h incubation as shown in Fig. S1 (Supporting infor-
4.2. Chemistry
mation). Compounds 35 and 41 exhibited noticeable inhibition po-
tency with the proliferation of CCRF-CEM cells by 35% and 53%,
respectively. Furthermore, compounds 33, 38, and 43 exhibited
modest inhibitory activities in CCRF-CEM cells by 27%, 27%, and
26%, respectively, after 72 h incubation. However, these com-
pounds did not exhibit a noticeable inhibition of the proliferation
of SK-OV-3 and MCF-7 cells. These data indicate that there is a
weak correlation between Src kinase inhibition and antiprolifera-
tive activity, presumably because of limited cellular uptake and
contribution of other mechanisms in antiproliferative activity of
these compounds.
4.2.1. General procedure for the synthesis of N-substituted pyridone
derivatives (28–43)
To
a solution of (4-oxo-4H-chromen-3-yl)acrylate (23–27)
(4 mmol) and aminoalkane/diaminoalkane/t-butylaminoethylcar-
bamate (4.4 mmol) in ethanol (70 mL) was added triethylamine
(2 drops), and the reaction mixture was refluxed for 8–10 h. The
progress of reaction was monitored on TLC. On completion of reac-
tion, the mixture was cooled to room temperature, and the solvent
was evaporated under reduced pressure. The crude product was
purified by column chromatography over silica gel (100–200
mesh) in 20–40% ethyl acetate/petroleum ether to give 2-pyridone
derivatives (28–43) in 74–85% yield.
3. Conclusion
In summary, a total of twenty one compounds including eigh-
teen 2-pyridone derivatives and three of chromone derivatives
were synthesized and fully characterized by 1H NMR, 13C NMR,
UV, FT-IR, and high resolution mass spectroscopy (HRMS). Nine-
teen compounds i.e. 28–33, and 36–48 are novel. Although com-
pounds 34 and 35 were known in literature, their complete
spectral data were not reported. Herein, we have reported the
spectral data for all of the compounds in the experimental section.
All of the synthesized 2-pyridones were evaluated for c-Src kinase
inhibitory activity. Preliminary results showed that eight
compounds exhibited relatively modest c-Src kinase inhibitory
4.2.1.1. 1-Hexyl-5-(2-hydroxybenzoyl)pyridin-2(1H)-one (28). The
reaction of (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylate (23)
(0.98 g, 4 mmol) with hexylamine (0.45 g, 4.4 mmol) gave the title
compound 28 as a light yellow solid (0.97 g, 81%) by following the
general procedure: mp = 73 °C; 1H NMR (300 MHz, CDCl3): d = 0.89
(t, 3H, J = 6.6 Hz, H-600), 1.34 (brs, 6H, H-500, H-400 & H-300), 1.76–1.81
(m, 2H, H-200), 3.99 (t, 2H, J = 7.3 Hz, H-100), 6.61 (d, 1H, J = 9.6 Hz, H-
30), 6.94 (t, 1H, J = 7.5 Hz, H-40), 7.07 (d, 1H, J = 8.1 Hz, H-3), 7.50–
7.58 (m, 2H, H-50 & H-60), 7.76 (dd, 1H, J = 2.4 & 9.6 Hz, H-4),
7.93 (d, 1H, J = 2.4 Hz, H-6), 11.43 ppm (s, 1H, OH); 13C NMR
(75.5 MHz, CDCl3): d = 13.92, 22.42, 26.23, 29.63, 31.26, 50.71,
117.01, 118.67, 118.83, 119.91, 131.51, 136.09, 138.84, 143.41,
activities with IC50 values less than 25 lM. Among all of the 2-pyri-

K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
79
161.93, 162.33, 195.17 ppm; IR (KBr):
m
max = 3424 (OAH str), 3063,
(brs, 1H, OH); 13C NMR (100.5 MHz, DMSO-d6): d = 14.40, 22.52,
26.12, 29.12, 31.34, 49.70, 103.30, 108.40, 114.22, 117.27, 119.07,
134.25, 139.58, 144.97, 161.76, 162.59, 164.24, 192.93 ppm; IR
2956, 1669 (C@O), 1624, 1483, 1337, 1247, 1137, 838, 760,
638 cmꢀ1; UV (MeOH): kmax = 293 nm; HRMS: m/z [M + Na]+ calcd
for C18H21NO3: 322.1419, found: 322.1418.
(KBr):
mmax = 3200 (OAH str), 2929, 2854, 1663 (C@O), 1629,
1602, 1432, 1340, 1236, 1117, 846, 721, 597, 554, 467 cmꢀ1; UV
(CHCl3): kmax = 284, 298 and 333 nm; HRMS: m/z [M + Na]+ calcd
for C18H21NO4: 338.1368, found: 338.1378.
4.2.1.2. 5-(2-Hydroxybenzoyl)-1-isopropylpyridin-2(1H)-one (29). The
reaction of (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylate (23)
(0.98 g, 4 mmol) with isopropylamine (0.26 g, 4.4 mmol) gave the
title compound 29 as a light yellow solid (0.81 g, 79%) by following
the general procedure: mp = 109 °C; 1H NMR (300 MHz, CDCl3):
d = 1.38 (d, 6H, J = 6.8 Hz, H-200), 5.25–5.28 (m, 1H, H-100), 6.58 (d,
1H, J = 9.2 Hz, H-3’), 6.91 (t, 1H, J = 7.6 Hz, H-40), 7.05 (d, 1H,
J = 8.4 Hz, H-3), 7.47–7.53 (m, 2H, H-50 & H-60), 7.70 (d, 1H,
J = 9.6 Hz, H-4), 7.97 (s, 1H, H-6), 11.44 ppm (s, 1H, OH); 13C
NMR (75.5 MHz, CDCl3): d = 22.05, 47.50, 117.46, 118.85, 118.96,
118.97, 119.74, 131.58, 136.20, 138.30, 139.45, 161.78, 162.54,
4.2.1.6. t-Butyl [2-{5-(2,4-dihydroxybenzoyl)-2-oxopyridin-1(2H)-
yl}ethyl]carbamate (33). The reaction of (E)-ethyl 3-(7-hydroxy-4-
oxo-4H-chromen-3-yl)acrylate (24) (1.04 g, 4 mmol) with t-butyl
(2-aminoethyl)carbamate (0.70 g, 4.4 mmol) gave the title com-
pound 33 as a light yellow solid (1.23 g, 82%) by following the gen-
eral procedure: mp = 180–182 °C; 1H NMR (400 MHz, DMSO-d6):
d = 1.24 (s, 9H, H-2000), 3.24–3.25 (m, 2H, H-200), 3.96 (t, 2H,
J = 4.8 Hz, H-100), 6.34–6.40 (m, 2H, H-30 & H-50), 6.45 (d, 1H,
J = 9.5 Hz, H-3), 6.95 (t, 1H, J = 5.92 Hz, CONH, D2O exchanged),
7.49 (d, 1H, J = 8.8 Hz, H-60), 7.72 (dd, 1H, J = 2.2 & 9.5 Hz, H-4),
7.99 (d, 1H, J = 2.2 Hz, H-6) 10.52 (brs, 1H, OH, D2O exchanged),
11.54 ppm (brs, 1H, OH, D2O exchanged); 13C NMR (100.5 MHz,
DMSO-d6): d = 28.04, 38.25, 49.62, 77.93, 102.86, 107.95, 113.16,
116.09, 118.79, 133.95, 139.10, 145.20, 155.71, 161.39, 162.66,
195.28 ppm; IR (KBr):
mmax = 3336 (OAH str), 3053, 2981, 1658
(C@O), 1621, 1589, 1440, 1338, 1245, 1135, 763, 640 cmꢀ1; UV
(MeOH): kmax = 297 nm; HRMS: m/z [M + Na]+ calcd for
C15H15NO3: 280.0950, found: 280.0949.
4.2.1.3. 1-Cyclohexyl-5-(2-hydroxybenzoyl)pyridin-2(1H)-one (30). The
reaction of (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylate (23)
(0.98 g, 4 mmol) with cyclohexylamine (0.44 g, 4.4 mmol) gave
the title compound 30 as a light yellow solid (0.90 g, 76%) by fol-
lowing the general procedure: mp = 101 °C; 1H NMR (300 MHz;
CDCl3): d = 1.18–1.78 (m, 6H, H-300, H-400 & H-500), 1.90–2.00 (m,
4H, H-200 & H-600), 4.89 (m, 1H, H-100), 6.60 (d, 1H, J = 9.2 Hz, H-30),
6.94 (t, 1H, J = 7.6 Hz, H-40), 7.08 (d, 1H, J = 8.2 Hz, H-3), 7.50–
7.54 (m, 2H, H-50 & H-60), 7.72 (dd, 1H, J = 2.8 & 9.6 Hz, H-4),
7.99 (d, 1H, J = 2.3 Hz, H-6), 11.48 ppm (s, 1H, OH); 13C NMR
(75.5 MHz, CDCl3): d = 25.38, 25.88, 32.83, 54.98, 117.34, 118.96,
119.06, 119.09, 119.75, 131.71, 136.29, 138.34, 140.15, 161.93,
163.94, 192.49 ppm; IR (KBr): mmax = 3368 (OAH str), 2976, 2698,
1686 (NHCOOA), 1647 (C@O), 1586, 1521, 1335, 1271, 1173,
851, 619, 584, cmꢀ1; UV (CHCl3): kmax = 288 and 334 nm; HRMS:
m/z [M + H]+ and [M + K]+ calcd for C19H22N2O6: 375.1556 and
413.1115, found: 375.1482 and 413.1033 respectively.
4.2.1.7. 1-Hexyl-5-(2-hydroxy-4-methoxybenzoyl)pyridin-2(1H)-one
(34) [32]. The reaction of (E)-ethyl 3-(7-methoxy-4-oxo-4H-chro-
men-3-yl)acrylate (26) (1.1 g, 4 mmol) with hexylamine (0.45 g,
4.4 mmol) gave the title compound 34 as a light yellow solid
(1.11 g, 84%) by following the general procedure: mp = 72–74 °C;
1H NMR (400 MHz, CDCl3): d = 0.85 (t, 3H, J = 7.3 Hz, H-600), 1.28–
1.33 (m, 6H, H-300–H-500), 1.72–1.76 (m, 2H, H-200), 3.83 (s, 3H,
OCH3), 3.95 (t, 3H, J = 7.7 Hz, H-100), 6.43 (dd, 1H, J = 2.2 & 8.8 Hz,
H-50), 6.47 (d, 1H, J = 2.2 Hz, H-30), 6.55 (d, 1H, J = 9.5 Hz, H-60),
7.45 (d, 1H, J = 8.8 Hz, H-3), 7.66 (dd, 1H, J = 2.9 & 9.5 Hz, H-4),
7.83 (d, 1H, J = 2.2 Hz, H-6), 12.19 ppm (brs, 1H, OH); 13C NMR
(100.5 MHz, CDCl3): d = 13.89, 22.40, 26.21, 29.17, 31.23, 50.58,
55.59, 101.32, 107.48, 112.49, 117.28, 119.79, 133.24, 138.83,
162.69, 195.61 ppm; IR (KBr): mmax = 3430 (OAH str), 3047, 2921,
1629 (C@O), 1570, 1533, 1451, 1096, 974, 850, 750 cmꢀ1; UV
(MeOH): kmax = 265 and 305 nm; HRMS: m/z [M + H]+ calcd for
C18H19NO3: 298.1443, found: 298.1460.
4.2.1.4. 1-[2-(Dimethylamino)ethyl]-5-(2-hydroxybenzoyl)pyridin-
2(1H)-one (31). The reaction of (E)-ethyl 3-(4-oxo-4H-chromen-
3-yl)acrylate (23) (0.98 g, 4 mmol) with N1,N1-dimethylethane-1,
2-diamine (0.39 g, 4.4 mmol) gave the title compound 31 as a light
yellow solid (0.94 g, 82%) by following the general procedure:
142.50, 161.92, 165.67, 166.03, 193.87 ppm; IR (KBr): mmax = 3427
(OAH str), 2929, 2258, 1675 (C@O), 1628, 1583, 1347, 1285,
1159, 1026, 828, 784, 622 cmꢀ1; UV (CHCl3): kmax = 290 and
337 nm; HRMS: m/z [M + H]+ calcd for C19H23NO4: 330.1705,
found: 330.1628.
1
mp = 147 °C; H NMR (300 MHz, CDCl3): d = 2.30 (s, 6H, H-1000),
2.66 (t, 2H, J = 5.4 Hz, H-200), 4.05 (t, 2H, J = 5.4 Hz, H-100), 6.61 (d,
1H, J = 9.3 Hz, H-30), 6.91 (t, 1H, J = 7.5 Hz, H-40), 7.07 (d, 1H,
J = 8.4 Hz, H-3), 7.51 (t, 1H, J = 7.5 Hz, H-50), 7.70 (d, 1H,
J = 7.5 Hz, H-60), 7.82 (dd, 1H, J = 2.1 & 9.6 Hz, H-4), 7.99 (d, 1H,
J = 1.8 Hz, H-6), 11.53 ppm (s, 1H, OH); 13C NMR (75.5 MHz, CDCl3):
d = 45.50, 47.27, 57.79, 116.11, 118.64, 118.69, 119.03, 119.82,
131.81, 135.98, 138.98, 145.08, 161.98, 162.53, 195.24 ppm; IR
4.2.1.8. 5-(2-Hydroxy-4-methoxybenzoyl)-1-isopropylpyridin-2(1H)-
one (35) [33]. The reaction of (E)-ethyl 3-(7-methoxy-4-oxo-4H-
chromen-3-yl)acrylate (26) (1.1 g, 4 mmol) with isopropyl amine
(0.26 g, 4.4 mmol) gave the title compound 35 as a light yellow
solid (0.95 g, 83%) by following the general procedure: mp = 141–
143 °C; 1H NMR (400 MHz, CDCl3): d = 1.37 (d, 6H, J = 6.6 Hz, H-
200), 3.83 (s, 3H, OCH3), 5.21–5.28 (m, 1H, H-100), 6.42–6.45 (m,
1H, H-50), 6.48 (d, 1H, J = 2.2 Hz, H-30), 6.55 (d, 1H, J = 9.5 Hz, H-
60), 7.43 (d, 1H, J = 8.8 Hz, H-3), 7.64 (dd, 1H, J = 2.9 & 9.5 Hz, H-
4), 7.89 (d, 1H, J = 2.9 Hz, H-6), 11.57 ppm (brs, 1H, OH); 13C
NMR (100.5 MHz, CDCl3): d = 21.88, 47.17, 55.59, 101.36, 107.44,
112.48, 117.60, 119.48, 133.18, 138.15, 138.36, 161.63, 165.69,
(KBr):
mmax = 3430 (OAH str), 3048, 1666 (C@O), 1624, 1590,
1336, 1246, 1174, 1138, 1048, 855, 764, 699 cmꢀ1; UV (MeOH):
kmax = 268 & 310 nm; HRMS: m/z [M + H]+ calcd for C16H18N2O3:
287.1396, found: 287.1398.
4.2.1.5. 5-(2,4-Dihydroxybenzoyl)-1-hexylpyridin-2(1H)-one (32). The
reaction of (E)-ethyl 3-(7-hydroxy-4-oxo-4H-chromen-3-yl)acry-
late (24) (1.04 g, 4 mmol) with hexylamine (0.45 g, 4.4 mmol) gave
the title compound 32 as a light yellow solid (1.07 g, 85%) by fol-
lowing the general procedure: mp = 166–168 °C; 1H NMR
(400 MHz, DMSO-d6): d = 0.81 (t, 3H, J = 6.8 Hz, H-600), 1.23 (brm,
6H, H-300–H-500), 1.58–1.61 (m, 2H, H-200), 3.91 (t, 2H, J = 7.2 Hz,
H-100), 6.32–6.36 (m, 2H, H-50 & H-30), 6.41 (d, 1H, J = 9.2 Hz, H-
3), 7.38 (d, 1H, J = 8.4 Hz, H-60), 7.67 (dd, 1H, J = 2.4 & 9.6 Hz, H-
4), 8.18 (d, 1H, J = 2.4 Hz, H-6), 10.45 (brs, 1H, OH); 11.33 ppm
166.01, 194.03 ppm; IR (KBr): mmax = 3462 (OAH str), 2982, 2851,
1664 (C@O), 1617, 1588, 1438, 1348, 1262, 1211, 1112, 937, 836,
606 cmꢀ1; UV (CHCl3): kmax = 283, 294 and 336 nm; HRMS: m/z
[M + H]+ calcd for C16H17NO4: 288.1236, found: 288.1159.
4.2.1.9. 1-[2-(Dimethylamino)ethyl]-5-(2-hydroxy-4-methoxybenzoyl)
pyridin-2(1H)-one (36). The reaction of (E)-ethyl 3-(7-methoxy-

80
K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
4-oxo-4H-chromen-3-yl)acrylate (26) (1.1 g, 4 mmol) with N1,N1-
dimethylethane-1,2-diamine (0.39 g, 4.4 mmol) gave the title
compound 36 as a light yellow solid (1.03 g, 81%) by following
the general procedure: mp = 150–152 °C; 1H NMR (400 MHz,
CDCl3): d = 2.27 (s, 6H, H-1000), 2.63 (t, 2H, J = 5.5 Hz, H-200), 3.84
(s, 3H, OCH3), 4.02 (t, 2H, J = 5.5 Hz, H-100), 6.41 (dd, 1H, J = 2.9 &
8.8 Hz, H-50), 6.48 (d, 1H, J = 2.9 Hz, H-30), 6.58 (d, 1H, J = 9.5 Hz,
H-60), 7.61 (d, 1H, J = 8.8 Hz, H-3), 7.73 (dd, 1H, J = 3.0 & 9.5 Hz,
H-4), 7.88 (d, 1H, J = 2.2 Hz, H-6), 12.30 ppm (brs, 1H, OH); 13C
NMR (100.5 MHz, CDCl3): d = 45.43, 47.09, 55.58, 57.71, 101.12,
107.38, 112.57, 116.25, 119.66, 133.47, 138.98, 144.07, 161.91,
49.18, 115.26, 116.87, 117.59, 118.39, 119.94, 125.03, 138.68,
145.64, 148.33, 149.85, 161.40, 191.66 ppm; IR (KBr): max = 3285
(OAH str), 2921, 2854, 1664 (C@O), 1602, 1446, 1208, 1137, 994,
831, 790, 644, 551 cmꢀ1; UV (CHCl3): kmax = 281 and 328 nm;
HRMS: m/z [M + H]+ calcd for C18H21NO4: 316.1549, found:
316.1588.
m
4.2.1.13.
5-(2,5-Dihydroxybenzoyl)-1-isopropylpyridin-2(1H)-one
(40). The reaction of (E)-ethyl 3-(6-hydroxy-4-oxo-4H-chromen-
3-yl)acrylate (25) (1.04 g, 4 mmol) with isopropyl amine (0.26 g,
4.4 mmol) gave the title compound 40 as a light yellow solid
(0.87 g, 80%) by following the general procedure: mp = 158–
160 °C; 1H NMR (400 MHz, DMSO-d6): d = 1.28 (d, 6H, J = 6.6 Hz,
H-200), 4.96–5.03 (m, 1H, H-100), 6.45 (d, 1H, J = 9.5 Hz, H-3), 6.70
(d, 1H, J = 2.2 Hz, H-60), 6.80 (d, 1H, J = 8.8 Hz, H-30), 6.84 (dd, 1H,
J = 2.9 & 8.8 Hz, H-40), 7.71 (dd, 1H, J = 2.9 & 9.5 Hz, H-4), 8.10 (d,
1H, J = 2.9 Hz, H-6), 9.08 (brs, 1H, OH), 9.54 ppm (brs, 1H, OH);
13C NMR (100.5 MHz, DMSO-d6): d = 21.06, 21.20, 47.49, 115.44,
116.98, 117.62, 118.40, 120.23, 124.85, 138.09, 141.36, 148.37,
165.76, 165.92, 193.87 ppm; IR (KBr):
mmax = 3427 (OAH str),
2944, 2778, 1669 (C@O), 1616, 1440, 1344, 1261, 1116, 1027,
921, 818, 608, 588 cmꢀ1; UV (CHCl3): kmax = 286 and 331 nm;
HRMS: m/z [M + H]+ calcd for C17H20N2O4: 317.1501, found:
317.1425.
4.2.1.10. t-Butyl [2-{5-(2-hydroxy-4-methoxybenzoyl)-2-oxopyridin-
1(2H)-yl}ethyl]carbamate (37). The reaction of (E)-ethyl 3-(7-meth-
oxy-4-oxo-4H-chromen-3-yl)acrylate (26) (1.1 g, 4 mmol) with
t-butyl (2-aminoethyl)carbamate (0.70 g, 4.4 mmol) gave the title
compound 37 as a light yellow solid (1.29 g, 83%) by following the
general procedure: mp = 164–166 °C; 1H NMR (400 MHz, CDCl3):
d = 1.31 (s, 9H, H-2000), 3.44–3.46 (m, 2H, H-200), 3.81 (s, 3H, OCH3),
4.09 (brs, 2H, H-100), 5.17 (brs, 1H, CONH), 6.41 (brm, 1H, H-50),
6.43–6.45 (m, 1H, H-30), 6.53 (d, 1H, J = 8.8 Hz, H-60), 7.48 (d, 1H,
J = 8.8 Hz, H-3), 7.68 (d, 1H, J = 9.6 Hz, H-4), 7.80 (d, 1H, J = 2.2 Hz,
H-6), 12.14 ppm (brs, 1H, OH); 13C NMR (100.5 MHz, CDCl3):
d = 28.14, 39.38, 49.98, 55.57, 79.73, 101.36, 107.51, 112.45,
117.36, 119.73, 133.35, 139.31, 143.23,156.03,162.19, 165.59,
149.95,161.05, 191.54 ppm; IR (KBr):
mmax = 3417 (OAH str),
3057, 2986, 1654, (C@O), 1612 (C@O), 1571, 1438, 1350, 1236,
1140, 795, 643, 574 cmꢀ1; UV (CHCl3): kmax = 286 and 337 nm;
HRMS: m/z [M + H]+ calcd for C15H15NO4: 274.1079, found:
274.1009.
4.2.1.14. 1-Hexyl-5-(2-hydroxy-5-methoxybenzoyl)pyridin-2(1H)-one
(41). The reaction of (E)-ethyl 3-(6-methoxy-4-oxo-4H-chromen-
3-yl)acrylate (27) (1.1 g, 4 mmol) with hexylamine (0.45 g,
4.4 mmol) gave the title compound 41 as a light yellow solid
(1.12 g, 85%) by following the general procedure: mp = 60–62 °C;
1H NMR (400 MHz, CDCl3): d = 0.86 (t, 3H, J = 6.8 Hz, H-600), 1.28–
1.36 (m, 6H, H-300-H-500), 1.72–1.79 (m, 2H, H-200), 3.74 (s, 3H,
OCH3), 3.96 (t, 2H, J = 7.3 Hz, H-100), 6.57 (d, 1H, J = 9.2 Hz, H-3),
6.97–7.00 (m, 2H, H-30 & H-60), 7.12 (dd, 1H, J = 2.8 & 8.7 Hz, H-
40), 7.73 (dd, 1H, J = 2.9 & 9.5 Hz, H-4), 7.93 (d, 1H, J = 2.9 Hz, H-
6), 10.88 ppm (brs, 1H, OH); 13C NMR (100.5 MHz, CDCl3):
d = 14.05, 22.51, 26.37, 29.44, 31.43, 50.85, 56.04, 114.09, 117.17,
118.67, 119.63,120.06, 123.47, 138.86, 143.48, 151.75,
166.04, 193.62 ppm; IR (KBr): mmax = 3363 (OAH str), 2979, 2962,
1663 (NHCOOA), 1627 (C@O), 1508, 1436, 1342, 1291, 1160, 857,
776, 621, cmꢀ1; UV (CHCl3): kmax = 285 and 332 nm; HRMS: m/z
[M + H]+ and [M + Na]+ calcd for C20H24N2O6: 389.1713 and
411.1532, found: 389.1654 and 411.1486 respectively.
4.2.1.11. 5-(2-Hydroxy-4-methoxybenzoyl)-1-(2-hydroxyethyl)pyri-
din-2(1H)-one (38). The reaction of (E)-ethyl 3-(7-methoxy-
4-oxo-4H-chromen-3-yl)acrylate (26) (1.1 g, 4 mmol) with
ethanolamine (0.27 g, 4.4 mmol) gave the title compound 38 as a
light yellow solid (0.88 g, 76%) by following the general procedure:
mp = 176–178 °C; 1H NMR (400 MHz, DMSO-d6): d = 3.60 (brm, 2H,
H-200), 3.77 (s, 3H, OCH3), 3.99 (t, 2H, J = 5.5 Hz, H-100), 4.93 (brs, 1H,
OH, D2O exchanged), 6.43–6.47 (m, 2H, H-50 & H-30), 6.49 (brs, 1H,
H-60), 7.49 (d, 1H, J = 8.0 Hz, H-3), 7.71 (dd, 1H, J = 2.2 & 9.5 Hz, H-
4), 8.11 (d, 1H, J = 2.2 Hz, H-6), 11.32 ppm (brs, 1H, OH, D2O ex-
changed); 13C NMR (100.5 MHz, DMSO-d6): d = 51.79, 55.57,
58.48,101.42, 106.45, 115.17, 115.97, 118.43, 133.10, 139.16,
156.53,162.05, 194.89 ppm; IR (KBr):
mmax = 3235 (OAH str),
2930, 2858, 1654 (C@O), 1599, 1422, 1316, 1286, 1213, 1131,
1039, 837, 803, 646, 423 cmꢀ1
; UV (CHCl3): kmax = 289 and
330 nm; HRMS: m/z [M + H]+ calcd for C19H23NO4: 330.1705,
found: 330.1629.
4.2.1.15.
1-Cyclohexyl-5-(2-hydroxy-5-methoxybenzoyl)pyridin-2
(1H)-one (42). The reaction of (E)-ethyl 3-(6-methoxy-4-oxo-4H-
chromen-3-yl)acrylate (27) (1.1 g, 4 mmol) with cyclohexylamine
(0.44 g, 4.4 mmol) gave the title compound 42 as a light yellow
solid (1.09 g, 83%) by following the general procedure: mp = 105–
146.02,161.36, 161.45, 164.37, 192.33 ppm; IR (KBr): mmax = 3336
1
(OAH str), 2917, 1660 (C@O), 1623 (C@O), 1577, 1351, 1290,
1163, 1023, 832, 790, 625, cmꢀ1; UV (CHCl3): kmax = 287 and
333 nm; HRMS: m/z [M + H]+ calcd for C15H15NO5: 290.1028,
found: 290.0988.
107 °C; H NMR (400 MHz, CDCl3): d = 1.18–1.74 (m, 6H, H-300–H-
500), 1.86–1.96 (m, 4H, H-200 & H-600), 3.72 (s, 3H, OCH3), 4.82–4.88
(m, 1H, H-100), 6.56 (d, 1H, J = 9.5 Hz, H-3), 6.94 (d, 1H, J = 2.9 Hz,
H-60), 6.95–6.98 (m, 1H, H-30), 7.09 (dd, 1H, J = 2.2 & 8.8 Hz, H-
40), 7.70 (dd, 1H, J = 2.2 & 9.5 Hz, H-4), 7.98 (d, 1H, J = 2.2 Hz, H-
6), 10.93 ppm (brs, 1H, OH); 13C NMR (100.5 MHz, CDCl3):
d = 25.08, 25.55, 32.54, 54.65, 55.77, 114.09, 117.02, 118.43,
119.48, 123.65, 137.96, 139.79, 151.55, 156.42,161.58,
4.2.1.12. 5-(2,5-Dihydroxybenzoyl)-1-hexylpyridin-2(1H)-one (39).
The reaction of (E)-ethyl 3-(6-hydroxy-4-oxo-4H-chromen-3-
yl)acrylate (25) (1.04 g, 4 mmol) with hexylamine (0.45 g,
4.4 mmol) gave the title compound 39 as a light yellow solid
(1.02 g, 81%) by following the general procedure: mp = 87–89 °C;
1H NMR (400 MHz, DMSO-d6): d = 0.85 (t, 3H, J = 5.8 Hz, H-600),
1.26 (brm, 6H, H-500, H-400 & H-300), 1.58–1.62 (m, 2H, H-200), 3.94
(t, 2H, J = 7.3 Hz, H-100), 6.44 (d, 1H, J = 9.5 Hz, H-3), 6.67 (d, 1H,
J = 2.9 Hz, H-60), 6.79 (d, 1H, J = 8.0, Hz, H-30), 6.83 (dd, 1H, J = 2.9
& 8.8 Hz, H-40), 7.73 (dd, 1H, J = 2.4 & 9.6 Hz, H-4), 8.19 (d, 1H,
J = 2.9 Hz, H-6), 9.07 (brs, 1H, OH); 9.46 ppm (brs, 1H, OH); 13C
NMR (100.5 MHz, DMSO-d6): d = 13.90, 21.98, 25.54, 28.69, 30.81,
194.82 ppm; IR (KBr):
mmax = 3437 (OAH str), 3061, 2941, 1667
(C@O), 1626, 1587, 1442, 1276, 1167, 1123, 1026, 846, 789,
626 cmꢀ1
; UV (CHCl3): kmax = 283 and 337 nm; HRMS: m/z
[M + H]+ calcd for C19H21NO4: 328.1549, found: 328.1488.
4.2.1.16. t-Butyl [2-{5-(2-hydroxy-5-methoxybenzoyl)-2-oxopyridin-
1(2H)-yl}ethyl]carbamate (43). The reaction of (E)-ethyl 3-(6-meth-
oxy-4-oxo-4H-chromen-3-yl)acrylate (27) (1.1 g, 4 mmol) with
t-butyl (2-aminoethyl)carbamate (0.70 g, 4.4 mmol) gave the title

K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
81
compound 43 as a light yellow solid (1.26 g, 81%) by following the
general procedure: mp = 145–146 °C; 1H NMR (400 MHz, CDCl3):
d = 1.32 (s, 9H, H-2000), 3.44–3.49 (m, 2H, H-200), 3.74 (s, 3H,
OCH3), 4.13 (brs, 2H, H-100), 4.95 (brs, 1H, CONH), 6.58 (d, 1H,
J = 9.5, H-3), 6.97 (d, 1H, 8.8 Hz, H-30), 7.03 (brs, 1H, H-60), 7.10
(dd, 1H, J = 2.9 & 8.8 Hz, H-40), 7.77 (dd, 1H, J = 2.2 & 9.5 Hz, H-4),
7.91 (d, 1H, J = 2.2 Hz, H-6), 10.85 ppm (brs, 1H, OH); 13C NMR
(100.5 MHz, CDCl3): d = 28.17, 39.39, 49.92, 56.05, 79.74, 114.78,
117.19,119.06, 119.63, 119.71, 123.13, 139.22, 144.18, 151.67,
fluoroaniline) (16 mmol) and few drops of triethylamine were
added and stirred under reflux for 12–13 h. After the mixture
was cooled to room temperature, and the solvent removed, the
crude product was purified by column chromatography over silica
gel using petroleum ether–ethyl acetate (15–20%) as eluent to give
analogues of 2-pyridone (46–48) in 68–79% yield.
4.2.3.1. Ethyl 3-morpholin-3-(4-oxo-4H-chromen-3-yl)propanoate
(46). The reaction of (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylate
(23) (0.98 g, 4 mmol) with morpholine (1.39 g, 16 mmol) gave the
title compound 46 as a light yellow solid (1.05 g, 79%) by following
the general procedure: mp = 110 °C; 1H NMR (300 MHz, CDCl3):
d = 1.20 (t, 3H, J = 7.0, H-2000), 2.49–2.62 (m, 4H, H-200 & H-600),
2.81–2.89 (dd, 1H, J = 15.0 & J = 7.5 Hz, H-2a/H-2b), 2.95–3.02
(dd, 1H, J = 15.0 & J = 7.8 Hz, H-2a/H-2b), 3.67 (brs, 4H, H-300 & H-
500), 4.11 (q, 2H, J = 6.9 Hz, H-1000), 4.30 (t, 1H, J = 7.3 Hz, H-3),
7.39–7.47(m, 2H, H-60 & H-70), 7.68 (d, 1H, J = 7.8 Hz, H-80), 7.89
(s, 1H, H-20), 8.22 ppm (d, 1H, J = 7.8 Hz, H-50); 13C NMR
(75.5 MHz, CDCl3): d = 14.20, 36.33, 50.22, 57.58, 60.51, 67.23,
118.04, 120.47, 124.05, 125.26, 126.09, 133.70, 154.56, 156.02,
155.98, 156.15,162.21, 194.51 ppm; IR (KBr):
mmax = 3365 (OAH
str), 3061, 2926, 1702 (NHCOOA), 1667 (C@O), 1634, 1578, 1479,
1327, 1246, 1048, 946, 839 cmꢀ1; UV (CHCl3): kmax = 282 and
336 nm; HRMS: m/z [M]+ calcd for C20H24N2O6: 388.1634, found:
388.1649.
4.2.2. General procedure for the synthesis of N,N-dialkylimino pyridin-
2(1H)-one (44–45)
To a solution of (E)-ethyl 3-(4-oxo-4H-chromen-3-yl)acrylate
(23) (4 mmol) in ethanol (45 mL), primary amines (10 mmol) and
few drops of triethylamine were added and stirred under reflux
for about 16 h. The mixture was then cooled to room temperature,
and the solvent evaporated under reduced pressure. The crude
product was purified by column chromatography over silica gel
using petroleum ether-ethyl acetate (15–20%) as eluent to give
the desired analogues of 2-pyridone (44–45) in 75–80% yield.
171.46, 177.35 ppm; IR (KBr):
mmax = 3092, 2892 (CAH), 2756,
1733 (C@O ester), 1638 (C@O), 1570, 1464, 1356, 1030, 913, 853,
767 cmꢀ1; UV (MeOH): kmax = 297 and 307 nm; HRMS: m/z [M]+
calcd for C18H21NO5: 331.1420, found: 331.1218.
4.2.2.1. (E)-5-[(2-Hydroxyphenyl)(isopropylimino)methyl]-1-isopro-
pylpyridin-2(1H)-one (44). The reaction of (E)-ethyl 3-(4-oxo-4H-
chromen-3-yl)acrylate (23) (0.98 g, 4 mmol) with isopropyl amine
(0.59 g, 10 mmol) gave the title compound 44 as a light yellow solid
(0.94 g, 79%) by following the general procedure: mp = 183 °C; 1H
NMR (300 MHz, CDCl3): d = 1.25 (d, 6H, J = 6.3 Hz, H-200), 1.37 (d,
6H, J = 6.9 Hz, H-2000), 3.64–3.73 (m, 1H, H-100), 5.29–5.38 (m, 1H,
H-1000), 6.66–6.76 (m, 2H, H-30 & H-40), 6.96–6.99 (m, 2H, H-3 & H-
50), 7.15 (d, 1H, J = 9.3 Hz, H-60), 7.26–7.31 (m, 2H, H-4 & H-6),
15.48 ppm (s, 1H, OH); 13C NMR (75.5 MHz, CDCl3): d = 21.97,
24.26, 46.60, 52.04, 112.26, 117.65, 118.29, 119.54, 121.03,130.60,
131.97, 132.66, 137.94, 161.20, 163.28, 167.12 ppm; IR (KBr):
4.2.3.2. Ethyl 3-(4-oxo-4H-chromen-3-yl)-3-(phenylamino)propano-
ate (47). The reaction of (E)-ethyl 3-(4-oxo-4H-chromen-3-
yl)acrylate (23) (0.98 g, 4 mmol) with aniline (1.49 g, 16 mmol)
gave the title compound 47 as a light yellow low melting solid
(0.94 g, 70%) by following the general procedure: 1H NMR
(300 MHz, CDCl3): d = 1.23 (t, 3H, J = 7.0 Hz, H-2000), 2.71–2.79 (dd,
1H, J = 15.3 & J = 7.2 Hz, H-2a/H-2b), 2.93–3.00 (dd, 1H, J = 15.0 &
J = 6.9 Hz, H-2a/H-2b), 4.15 (q, 2H, J = 7.0 Hz, H-1000), 5.47 (t, 1H,
J = 7.0 Hz, H-3), 6.92 (d, 1H, J = 8.1 Hz, H-400), 7.03–7.10 (m, 4H,
H-200, H-300, H-500 & H-600), 7.26–7.46 (m, 4H, H-20, H-60, H-70, H-
80), 7.93 (d, 1H, J = 7.8 Hz, H-50), 11.83 ppm (d, 1H, J = 12.0 Hz,
NH); 13C NMR (75.5 MHz, CDCl3): d = 14.18, 41.34, 60.78, 75.71,
103.65, 116.40, 118.01, 121.78, 122.82, 123.93, 126.47, 129.82,
134.65, 139.97, 141.57, 157.56, 170.21, 182.02 ppm; IR (Nujol):
m
max = 3433 (OAH str), 3053, 2966, 1661 (C@O), 1595, 1522, 1438,
1304, 1257, 926, 754 cmꢀ1; UV (MeOH): kmax = 260 and 322 nm;
HRMS: m/z [M + H]+ calcd for C18H22N2O2: 299.1760, found:
299.1775.
m
max = 3235, 2970 (CAH), 1747 (C@O ester), 1647 (C@O), 1597,
1542, 1453, 1230, 1143, 956, 742 and 635 cmꢀ1; UV (MeOH):
kmax = 298 nm; HRMS: m/z [M + H]+ calcd for C20H19NO4:
338.1392, found: 338.1267.
4.2.2.2.
(E)-1-Cyclohexyl-5-[(cyclohexylimino)(2-hydroxyphenyl)
methyl]pyridin-2(1H)-one (45). The reaction of (E)-ethyl 3-(4-oxo-
4H-chromen-3-yl)acrylate (23) (0.98 g, 4 mmol) with cyclohexyl
amine (0.99 g, 10 mmol) gave the title compound 45 as a yellow
low melting solid (1.18 g, 78%) by following the general procedure:
1H NMR (400 MHz, CDCl3): d = 1.14–1.99 (m, 20H, H-200–H-600 & H-
2000–H-6000), 3.31–3.38 (m, 1H, H-100), 4.89–4.97 (m, 1H, H-1000),
6.66–6.73 (m, 2H, H-30 & H-40), 6.95–6.97 (m, 2H, H-3 & H-50),
7.13 (dd, 1H, J = 2.3 & 9.2 Hz, H-60), 7.23–7.24 (m, 1H, H-4), 7.25–
7.30 ppm (m, 1H, H-6); 13C NMR (100.5 MHz, CDCl3): d = 23.98,
25.24, 25.32, 25.65, 32.60, 54.08, 59.44, 111.83, 117.44, 118.38,
119.40, 120.82, 130.57, 132.64, 132.75, 137.93, 161.27, 163.68,
4.2.3.3. Ethyl 3-(4-fluorophenylamino)-3-(4-oxo-4H-chromen-3-yl)
propanoate (48). The reaction of (E)-ethyl 3-(4-oxo-4H-chromen-
3-yl)acrylate (23) (0.98 g, 4 mmol) with fluoroaniline (1.78 g,
16 mmol) gave the title compound 48 as a light yellow low melting
solid (0.97 g, 68%) by following the general procedure: 1H NMR
(300 MHz, CDCl3): d = 1.24 (brs, 3H, H-2000), 2.71–2.79 (dd, 1H,
J = 14.7 & J = 6.9 Hz, H-2a/H-2b), 2.92–2.99 (dd, 1H, J = 15.3 &
J = 7.2 Hz, H-2a/H-2b), 4.14 (brs, 2H, H-1000), 5.46 (t, 1H, J = 6.3 Hz,
H-3), 6.92 (brs, 1H, H-20), 7.04–7.06 (m, 5H, H-200, H-300, H-500, H-
600 & H-60), 7.32–7.42(m, 2H, H-70 & H-80), 7.92 (d, 1H, J = 7.2 Hz,
H-50), 11.83 ppm (s, 1H, J = 11.4 Hz, NH); 13C NMR (75.5 MHz,
CDCl3): d = 14.08, 41.26, 60.78, 75.59, 103.65, 116.43, 116.74,
117.89, 118.01, 121.80, 122.74, 126.46, 134.70, 136.36, 142.04,
157.57, 157.85, 161.08, 170.22, 182.05 ppm; IR (Nujol):
167.24 ppm; IR (Nujol): mmax = 3441 (OAH str), 2932, 2856, 1668,
1624, 1582, 1337, 1133, 836, 758 cmꢀ1; UV (MeOH): kmax = 265 &
305 nm; HRMS: m/z [M + H]+ calcd for C24H30N2O2: 379.2386,
found: 379.2407.
4.2.3. General procedure for the synthesis of ethyl 3-morpholin-3-(4-
oxo-4H-chromen-3-yl)propanoate (46), ethyl 3-(4-oxo-4H-chromen-
3-yl)-3(phenylamino)propanoate (47) and ethyl 3-(fluorophenylamino)-
3-(4-oxo-4H-chromen-3-yl)propanoate (48)
To a solution of chromone ester (23) (4 mmol) in ethanol
(45 mL), secondary or aromatic amines (morpholine, aniline and
m
max = 3078, 2982 (CAH), 1731 (C@O ester), 1651 (C@O), 1515,
1469, 1371, 1283, 1029, 945 and 759 cmꢀ1
; UV (MeOH):
kmax = 297 nm; HRMS: m/z [M]+ calcd for C20H18FNO4: 355.1220,
found: 355.1121.

82
K. Chand et al. / Bioorganic Chemistry 53 (2014) 75–82
4.3. Biology
Appendix A. Supplementary material
4.3.1. c-Src kinase inhibitory activity assay
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bioorg.2014.02.
001.
The effect of synthesized compounds on the activity of c-Src ki-
nase was assessed by Transcreener ADP2 FI assay (Bell Brook Labs,
Madison, Wisconsin; catalogue no. 3013-1K) according to manu-
facturer’s protocol. A 384-well low-volume black non binding sur-
face round-bottom microplate was purchased from Corning (No.
3676). In summary, the kinase reaction was started in 384-well
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4.3.2. EGFR, MAPK and PDK inhibitory activity assay
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Tyr) 4:1. MAPK1 (h) was incubated with 25 mM Tris pH 7.5,
0.02 mM EGTA, 250
Millipore, Dundee, UK), whereas PDK1 (h) was incubated with
50 mM Tris pH 7.5, 100 KTFCGTPEYLAPEVRREPRILSEEE-
QEMFRDFDYIADWC (PDKtide). The incubation was followed by
the addition of 10 mM magnesium acetate and [
33P-ATP] (spe-
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40 min of incubation with the addition of 3% phosphoric acid solu-
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The financial support from Defence Research Development
Organization (DRDO), Delhi and Council of Scientific and Industrial
Research (CSIR) is gratefully acknowledged. We also acknowledge
the financial support from the American Cancer Society Grant #
RSG-07-290-01-CDD and NIH Grant Number 8 P20 GM103430-12
for sponsoring the core facility. The author KC is thankful to CSIR
for the Senior Research Fellow award.