Dihydropyrido[2,3-d]pyrimidines as a new class of antileishmanial agents

Article abstract of DOI:10.1016/j.bmc.2005.07.043

A series of dihydropyrido[2,3-d]pyrimidines have been synthesized and screened for its in vitro antileishmanial activity profile in promastigote and amastigote models. Compounds 2a-2l have shown 83-100% inhibition against promastigotes and 79-100% inhibition against amastigotes at a concentration of 50 μg/mL.

Full text of DOI:10.1016/j.bmc.2005.07.043

Bioorganic & Medicinal Chemistry 13 (2005) 6678–6684
Dihydropyrido[2,3-d]pyrimidines as a new class
of antileishmanial agents
Anu Agarwal,a Ramesh,b Ashutosh,c Neena Goyal,c
Prem M. S. Chauhana,* and Suman Guptab
aDivision of Medicinal Chemistry, Central Drug Research Institute, Lucknow 226001, India
bDivision of Parasitology, Central Drug Research Institute, Lucknow 226001, India
cDivision of Biochemistry, Central Drug Research Institute, Lucknow 226001, India
Received 14 June 2005; revised 18 July 2005; accepted 18 July 2005
Available online 26 August 2005
Abstract—A series of dihydropyrido[2,3-d]pyrimidines have been synthesized and screened for its in vitro antileishmanial activity
profile in promastigote and amastigote models. Compounds 2a–2l have shown 83–100% inhibition against promastigotes and
79–100% inhibition against amastigotes at a concentration of 50 lg/mL.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
There is still no effective vaccine for Kala-azar and chemo-
therapy remains the most effective control measure. The
drugs for the treatment of leishmaniasis are sodium stib-
ogluconate (Pentostam) and meglumine antimonate
(Glucantime), despite the fact that they exhibit renal
and cardiac toxicity. Although new drugs, i.e., amphoter-
icin B and its lipid complex, are quite effective, they are
expensive and out of reach of poor people.2,5 Newly intro-
duced first orally active drug miltefosine, a phosphocho-
line analogue, is quite effective in presenting severe
gastrointestinal problems and also shows teratogenic
effects and cannot be used in pregnant women.6,7 The
search for new drugs continues, with bisphosphonates,
for example, risedronate and pamidronate. It is also
known that these drugs also contribute to increased co-in-
fections leishmaniasis-AIDS. No treatment has proven to
be effective in achieving radical cure of visceral leishman-
iasis when it is associated with HIV infection. Therefore,
development of more effective and safer chemotherapeu-
tic agents for treating Leishmaniasis seems to be desir-
able.8 The pharmaceutical industry has shown little
interest in drug discovery for parasitic disease due to a
lack of financial incentive. There is also lack of interest
among researchers in the developed nations as it is a dis-
ease of the developing countries.
Leishmaniasis is caused by different species belonging to
the genus Leishmania, a protozoan which is transmitted
to humans by the bite of an insect vector, phlebotomine
sandfly. Leishmaniasis has an overwhelming impact on
global public health and is endemic in many tropical
and subtropical regions of the world. It affects around
12 million people of the world and 350 million are esti-
mated to be prone to the diseases, of which around 1.7
million people will be infected each year.1–3 Infection
by various strains of Leishmania causes a wide spectrum
of disease in humans, with many different clinical man-
ifestations, i.e., cutaneous, mucocutaneous and visceral.
The visceral form of Leishmaniasis, commonly known
as kala-azar, is caused by the parasite Leishmania dono-
vani, which affects 61 out of the 88 countries worldwide.
It attacks the phagocytic cells of the spleen, liver and
bone marrow, and is fatal in more than 90% of the
untreated cases. The parasitic protozoan is digenetic
and has two distinct stages in its life cycle. The motile
flagellated promastigote stage lives in the alimentary
canal of the sandfly vector, which, by inoculation,
transmits the promastigotes into the mammalian host,
where they enter macrophages differentiating and multi-
plying into non-motile amastigotes.4
Compounds of both synthetic and natural origin com-
prising a diverse group of chemical structures have
been reported as antileishmanial agents. These include
mostly the nitrogen heterocycles as quinolines,9
acridines,10 phenothiazines,11 pyrimidines,12 purines,13
Keywords: Lieshmania; Dihydrofolate reductase; Dihydropyrido[2,3-
d]pyrimidine; Uracil.
*
Corresponding author. Fax: +91-522-2623405; e-mail addresses:
prem_chauhan_2000@yahoo.com; premsc58@hotmail.com
0968-0896/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.07.043

A. Agarwal et al. / Bioorg. Med. Chem. 13 (2005) 6678–6684
6679
bis-benzamidines,14 pyrazolo[3,4-b]pyridine,15 benzothia-
zoles,16 and imidazolidine.17 Other classes of compounds
include buparvaquone-oxime,18 bisbenzamidines,19 chal-
cones,20 quinines,21 amino acid esters and amides,22 amino
alcohols,23 alkyl phospholipids,24 ether phospholipids,25
sulfanilamides,26 artemisinin27 and certain platinum
complexes.28
As part of our ongoing program devoted to the synthesis
of diverse heterocycles of biological interest,39 we con-
centrated our investigations on dihydropyrido[2,3-d]py-
rimidines40 due to their broad range of biological
activities. In this communication, we have reported the
antileishmanial activity of these synthesized compounds.
Uracil derivatives are versatile building blocks for the
synthesis of nitrogen-containing heteroaromatic species
of biological importance.29 Pyrazolopyridines,30 pyrimi-
dopyrimidines,31 pyridopurines,32 pyrazolo-pyrimi-
dines33 and xanthine derivatives34 have all been
prepared by the functionalization of these important
heterocyclic building blocks, the structures of which
are interesting in their own right, as well as being biolog-
ically active pyrimidine nucleosides. The diverse range
of biological activities of uracil derivatives in parasitic
chemotherapy has stimulated considerable interest in
their synthesis.
2. Chemistry
Dihydropyrido[2,3-d]pyrimidines (2a–r) are synthesized
in high yields by heating a mixture of 6-amino-1,3-di-
methyl uracil, different aldehydes and acetyl acetone in
acetic acid (Scheme 1), at 110 ꢁC for 8 h.
Initially after 8 h high yields of dihydropyrido[2,3-d]-
pyrimidines (2) are obtained and oxidized products (3
and 4) are either not formed or are formed in a very
negligible amount in some cases. As the reaction time is
increased, oxidation of the dihydropyrido[2,3-d]pyrimi-
dine (2) takes place with the formation of dearylated
product (4) and normal dehydrogenated product (3),
The amount of 3 and 4 goes on increasing with time
(Scheme 2). All the synthesized compounds are well-char-
acterized by spectroscopic method, such as IR, mass,
NMR, and elemental analysis.
Dihydrofolate reductase (DHFR) is an important target
site in most of the parasitic diseases. Most of the clini-
cally used DHFR inhibitors show less selectivity for
leishmanial enzymes.35 This is due to the fact that the
gene for pteridine reductase (PTR1) is amplified in some
leishmanial mutants. PTR1 can reduce pterins and
folates, and therefore act as a bypass for DHFR
inhibition. This implies that antifolate drugs must simul-
taneously target both DHFR and PTR1 to be successful
antileishmanials.36
2.1. Results and discussion
The in vitro biological activity of dihydropyrido[2,3-
d]pyrimidines has shown encouraging results (Table 1)
against L. donovani. Compounds showed a good corre-
lation of activity with the structure and logP values.
Four compounds (2d, 2g, 2h and 2i) showed 100% inhi-
bition against promastigotes, whereas two compounds
(2d, 2g) exhibited 100% inhibition in amastigotes at a
concentration of 50 lg/mL. When the R group was
phenyl, the compound 2a showed 89% and 72% inhibi-
tion of promastigotes, and 71% and 56% inhibition of
amastigotes at a concentration of 50 and 10 lg/mL,
respectively. Substitution of the phenyl ring with a
methyl group at the para position, compound 2b showed
an increase in activity exhibiting 97% and 74% inhibi-
tion for promastigotes, and 80% and 62% for amastig-
otes. Substitution of another methyl group at meta
position, compound 2c showed a further increase in
activity. On substitution of the phenyl ring with an
isopropyl group at the para position, compound 2d
exhibited 100% activity in both promastigotes and
amastigotes at a concentration of 50 lg/mL, and 84%
and 94% inhibition at a concentration of 10 lg/mL.
Activity of compounds gradually increased on substitut-
ing the phenyl ring with methyl, dimethyl and isopropyl
groups as with the increase in logP value. Substituting
A number of compounds having pteridine, quinazoline
and pyrimidine moieties are reported to be potent inhib-
itors of PTR1 in Leishmania.37 Based on these observa-
tions we hypothesized to synthesize pyrido[2,3-
d]pyrimidines in which one N atom of pteridine was re-
placed by carbon and in comparison to quinazoline it
has one carbon atom replaced by a nitrogen atom which
could also act as a PTR1 inhibitor. Earlier reports have
shown that pyrido[2,3-d]pyrimidine is a potent inhibitor
of dihydrofolate reductase.38 Due to the same reason,
the synthesized compounds can act as inhibitors of
PTR1, as well as inhibitors of DHFR, and thus could
be potential antileishmanial agents.
O
O
R
RCHO
Me
O
Me
O
COCH3
CH3
CH3C(O)CH2COCH3
N
N
AcOH, Heat, 8h
N
NH2
N
N
H
Me
Me
1
2
Scheme 1.
O
R
O
R
O
Me
O
COCH3
Me
N
COCH3 Me
+
COCH
CH3
N
N
AcOH
Heat
N
N
H
CH
3
O
N
N
CH3
O
N
N
Me
Me
Me
2
3
4
Scheme 2.

6680
A. Agarwal et al. / Bioorg. Med. Chem. 13 (2005) 6678–6684
Table 1. Antileishmanial in vitro activity against luciferase-promastigote and luciferase-amastigote systems
Compounds
R
logP
% inhibition
Promastigotes
concentration (lg/mL)
Amastigotes
concentration (lg/mL)
50
10
50
10
2a
2b
2c
2d
2e
2f
C6H5
0.25
0.74
89.2
72.4
74.2
78.4
84.2
86.3
89.2
92.4
99.1
93.8
92.6
64.1
68.1
56.8
42.4
48.4
45.2
32.4
45.7
38.4
28.6
ND
ND
—
71.4
55.6
61.9
72.8
94.2
66.2
68.9
80.3
76.1
68.5
75.7
46.6
52.4
42.5
NI
4-Me-C6H4
3,4-diMe-C6H3
4-CH(CH3)2-C6H4
4-F-C6H4
96.8
98.2
80.1
89.9
1.23
1.49
100
100
0.41
0.81
96.7
99.3
89.4
93.2
4-Cl-C6H4
2g
2h
2i
3,4-diCl-C6H3
4-CN-C6H4
3-NO2-C6H4
4-NO2-C6H4
3-OCH3-C6H4
4-OCH3-C6H4
3,4-diOCH3-C6H3
3,4,5-TriOCH3-C6H2
4-OH-C6H4
4-COOH-C6H4
2-Furan
1.37
0.29
100
100
100
100
100
97.4
81.6
88.9
81.2
79.6
54.6
38.6
53.6
1.91
1.91
2j
2k
2l
0.13
0.13
84.2
83.2
69.2
52.6
59.8
54.6
56.9
59.4
55.6
43.3
45.6
38.6
2m
2n
2o
2p
2q
2r
3f
0.0
À0.13
À0.14
À0.19
À1.20
À0.94
2.53
42.1
NI
NI
35
22
CH3
4-Cl-C6H4
42.4
32.4
12.4
28.5
22.1
21
26.4
ND
ND
ND
ND
ND
30
31
3h
4
4-OCH3-C6H4
4-CN-C6H4
—
1.84
2.00
0.29
SSGꢂ
Pentamidineꢂ
a
b
—
90–99
a: SSG (sodium stilbogluconate) shows 40–50% inhibition at 500 lg/mL.
b: Pentamidine shows 85–90% inhibition at 0.5 lg/mL.
ND, not determined; NI, no inhibition.
the phenyl ring with groups as fluoro, the compound 2e
showed ꢀ97% and ꢀ86% inhibition in promastigotes at
a concentration of 50 and 10 lg/mL, respectively. Com-
pound 2e showed ꢀ89% and ꢀ66% inhibition in
amastigotes at a concentration of 50 and 10 lg/mL,
respectively. Replacement of the fluoro group with
chloro group the compound 2f exhibited a further in-
crease in activity in both promastigotes and amastigotes.
On disubstituting the phenyl ring with chloro group at 3
and 4 positions, the compound 2g showed 100% inhibi-
tion in both promastigotes and amastigotes at a concen-
tration of 50 lg/mL and inhibited 92% and 80% growth
at a concentration of 10 lg/mL. In case of compounds
having groups as fluoro and chloro, the activity in-
creased on going from fluoro to chloro and increased
further on disubstitution with the increase in logP value.
On substituting the phenyl ring with a cyano group at
the para position the compound 2h showed 100% and
99% inhibition against promastigotes, and 88% and
76% inhibition against amastigotes at a concentration
of 50 and 10 lg/mL. Substituting the phenyl ring with
the nitro group at meta or para positions, the com-
pounds 2i and 2j have shown 100% inhibition in pro-
mastigotes, and 82% and 89% inhibition against
amastigotes at a concentration of 50 lg/mL. At a con-
centration of 10 lg/mL, the compounds 2i and 2j
showed 92% and 93% inhibition against promastigotes,
and 68% and 76% inhibition against amastigotes.
Substituting the phenyl ring with methoxy group at
the meta (2k) or para position (2l) reduced the activity
against both promastigotes and amastigotes. Di or tri
substitution of the methoxy group in the phenyl ring
(2m and 2n) reduced the activity further and the activity
decreased with an increase in substitution. Substitution
of the methoxy group on the phenyl ring reduced the
logP value of the compounds, thus reducing the activity.
Di or tri substitution of the methoxy group decreased
further the logP value, showing a decrease in activity.
Substitution of the phenyl ring with hydroxyl (2o) or
carboxyl group (2p) reduced the logP value of com-
pounds, reducing the activity. Replacement of the phen-
yl ring with heterocyclic ring as furan (2q) or with
methyl group (2r), the logP value of compounds de-
creased to a greater amount, showing a lower activity.
When the dihydropyridine rings of the compounds 2f,
2l and 2h were oxidized to the corresponding pyridine
rings the activity of compounds 3f, 3l and 3h was re-
duced to a great extent. The results emphasize that, in
general, the activity increased when chloro, cyano, nitro
and methyl are in the aromatic ring. Compounds exhib-
ited a lower activity when methoxy, hydroxy, or carboxy
groups are present in the aromatic ring. The activity
increased or decreased on di or tri substituting the
groups, which increase or decrease the activity. The
dihydro pyridine moiety plays an important role in activ-
ity as when the dihydro ring was oxidized to the corre-
sponding pyridine ring the activity was reduced to a
great extent.

A. Agarwal et al. / Bioorg. Med. Chem. 13 (2005) 6678–6684
6681
3. Conclusion
amastigotes. The test material in appropriate concentra-
tions (50 and 10 lg/mL) in complete medium was added
after replacing the previous medium and the plates were
incubated at 37 ꢁC in a CO2 incubator for 24 h or more.
After incubation, the drug containing medium was
decanted and 50 lL PBS was added to each well and
mixed with an equal volume of the steady Glo reagent.
After gentle shaking for 1–2 min, the reading was taken
in a luminometer.
Leishmania is a disease of developing countries. The
majority of antileishmanial chemotherapy relies on
antimonials and benzamidines, which are highly toxic.
Therefore, development of more effective and safer
chemotherapeutic agents for treating Leishmaniasis re-
mains desirable. There is also lack of interest among those
in the pharmaceutical industry in the discovery of a new
antileishmanial agent due to a lack of financial incentives.
The synthesized dihydropyrido[2,3-d]pyrimidines have
shown good in vitro activity. These compounds are new
leads in antileishmanial chemotherapy and are very useful
for further optimization work on the same.
4.2. General procedure for the synthesis of 6-acetyl-5-(aryl/
alkyl)-1,3,7-trimethyl-5,8-dihydro-1H-pyrido[2,3-d]pyrimi-
dine-2,4-dione (2a–2r)
6-Amino-1,3-dimethyl uracil (I) (0.3 mmol), acetyl ace-
tone (0.35 mmol), and aromatic or aliphatic aldehyde
(0.35 mmol) were heated in acetic acid (15 ml) for 8 h
to obtain dihydropyrido[2,3-d]pyrimidines (2a–r). The
mixture was allowed to cool and water (50 mL) was add-
ed. The precipitated solid was washed with water and
dried. Recrystallization from ethanol gave light yellow
or white crystals of the desired compound.
4. Experimental section
IR spectra were recorded on Beckman Aculab-10,
Perkin Elmer 881 and FTIR 8210 PC, Shimadzu spec-
trophotometers either on KBr discs or in neat. Nuclear
magnetic resonance (NMR) spectra were recorded on
either Bruker Avance DRX-300 MHz or Bruker DPX
200 FT spectrometers using TMS as an internal refer-
ence. FAB mass spectra were recorded on JEOL SX
102/DA 6000 mass spectrometer using argon/xenon
(6 kV, 10 mA) as the FAB gas. Chemical analysis was
carried out on a Carlo-Erba-1108 instrument. The
melting points were recorded on an electrically heated
melting point apparatus and are uncorrected.
4.2.1. 6-Acetyl-5-phenyl-1,3,7-trimethyl-5,8-dihydro-1H-
pyrido[2,3-d]pyrimidine-2,4-dione (2a). MS: 326 (M+1);
IR (KBr) 3005, 2953, 1708, 1676, 1619, 1514,
1
1388 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.26
(s, 1H), 7.24 (d, 2H, J = 6.8 Hz), 7.16 (d, 2H,
J = 6.8 Hz), 6.05 (s, 1H, NH), 5.02 (s, 1H), 3.44 (s, 3H,
NMe), 3.28 (s, 3H, NMe), 2.45 (s, 3H), 2.15 (s, 3H); 13C
(CDCl3, 50 MHz) 199.6, 161.4, 150.7, 142.1, 141.8,
137.2, 135.4, 127.8, 127.1, 124.6, 91.6, 37.5, 29.9, 28.7,
28.1, 19.8; Anal. Calcd for C18H19N3O3: C, 66.45; H,
5.89; N, 12.91. Found: C, 66.25; H, 6.12; N, 12.84.
4.1. Antileishmanial activity
Luciferase transfected L. donovani promastigotes
(MHOM/IN/80/Dd-8, obtained from Imperial College,
London), which are more stable under the influence of G
41818, were maintained at 25 1 ꢁC in Medium 199 (SIG-
MA Chemical, USA) supplemented with 10% Fetal Calf
Serum (Gibco). The in vitro effect of compounds on the
growth of promastigotes was assessed by monitoring the
luciferase activity of viable cells after treatment. The trans-
genic promastigotes of late log phase were seeded at
5 · 105/100 lL medium 199/well in 96-well flat-bottomed
microtitre (MT) plates (CELLSTAR) and incubated for
72 h in medium alone or in the presence of serial dilutions
of drugs (250 ng/mL to 10 lg/mL) in DMSO.18 Parallel
dilutions of DMSO were used as controls. After incuba-
tion, an aliquot (50 lL) of promastigote suspension was
aspirated from each well of a 96-well plate and mixed with
an equal volume of Steady Glo(R) reagent (Promega) and
luminescence was measured in luminometer. The values
were expressed as RLU (relative luminescence unit).
4.2.2. 6-Acetyl-1,3,7-trimethyl-5-p-tolyl-5,8-dihydro-1H-
pyrido[2,3-d]pyrimidine-2,4-dione (2b). MS: 340 (M+1);
IR (KBr) 3006, 2955, 1704, 1661, 1556, 1559, 1514,
1482 cmÀ1;1H NMR (CDCl3, 200 MHz) d (ppm) 7.27
(d, 2H, J = 7.9 Hz), 7.09 (d, 2H, J = 7.9 Hz), 6.01 (s,
1H, NH), 5.09 (s, 1H), 3.44 (s, 3H, NMe), 3.28 (s, 3H,
NMe), 2.44 (s, 3H), 2.28 (s, 3H), 2.14 (s, 3H); 13C (CDCl3,
50 MHz) 199.4, 161.4, 150.6, 141.9, 141.7, 137.1, 130.6,
128.1, 126.8, 91.5, 37.4, 34.5, 29.8, 28.7, 28.2, 20.7, 19.8;
Anal. Calcd for C19H21N3O3: C, 67.24; H, 6.24; N,
12.38. Found: C, 67.52; H, 6.34; N, 12.04.
4.2.3. 6-Acetyl-5-(3,4-dimethyl-phenyl)-1,3,7-trimethyl-
5,8-dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2c).
MS: 354 (M+1); IR (KBr) 3015, 2945, 1708, 1665, 1548,
1
1556, 1519, 1472 cmÀ1; H NMR (CDCl3, 200 MHz) d
(ppm) 7.09 (d, 1H, J = 7.7 Hz), 7.04 (s, 1H), 6.99 (d, 1H
J = 7.7 Hz), 6.01 (s, 1H, NH), 5.09 (s, 1H), 3.44 (s, 3H,
NMe), 3.28 (s, 3H, NMe), 2.45 (s, 3H), 2.14 (s, 3H), 2.08
(s, 6H, 2CH3); 13C (CDCl3, 50 MHz) 199.3, 161.4,
134.9, 150.6, 141.8, 141.5, 136.9, 136.2, 133.8, 130.2,
129.3, 125.6, 91.6, 38.3, 29.7, 28.6, 28.2, 20.6, 20.3, 19.8;
Anal. Calcd for C20H23N3O3: C, 67.97; H, 6.56; N,
11.89. Found: C, 67.72; H, 6.34; N, 12.04.
For assessing the activity of compounds against amasti-
gote stage of the parasite, mouse macrophage cell line
(J-774A.1) infected with promastigotes expressing lucifer-
ase firefly reporter gene was used. Cells were seeded in a
96-well plate (5 · 104 cells/100 lL/well) in RPMI-1640
containing 10% foetal calf serum and the plates were incu-
bated at 37 ꢁC in a CO2 incubator. After 24 h, the medium
was replaced with fresh medium containing stationary-
phase promastigotes (2.5 · 105/100 lL/well). Promasti-
gotes invade the macrophage and are transformed into
4.2.4. 6-Acetyl-5-(4-isopropyl-phenyl)-1,3,7-trimethyl-5,8-
dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2d). MS:
368 (M+1); IR (KBr) 3020, 2930, 1709, 1675, 1554, 1541,

6682
A. Agarwal et al. / Bioorg. Med. Chem. 13 (2005) 6678–6684
1
1517, 1460 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm)
7.38 (d, 2H, J = 7.6 Hz), 7.16 (d, 2H, J = 7.6 Hz), 5.98 (s,
1H, NH), 5.11 (s, 1H), 3.44 (s, 3H, NMe), 3.28 (s, 3H,
NMe), 3.18–3.23 (m, 1H), 2.43 (s, 3H, COCH3), 2.26 (s,
3H, CH3), 1.24 (s, 6H, 2CH3); 13C (CDCl3, 50 MHz)
199.5, 161.5, 150.8, 147.5, 142.1, 141.4, 137.1, 127.5,
126.5, 113.9, 91.4, 38.0, 33.7, 29.8, 28.8, 28.2, 23.9, 19.8;
Anal. Calcd for C21H25N3O3: C, 68.64; H, 6.86; N, 11.44.
Found: C, 68.52; H, 6.54; N, 11.04.
200 MHz) d (ppm) 8.09 (d, 1H, J = 8.3Hz), 8.03 (s,
1H), 7.87 (d, 1H J = 8.3 Hz), 7.49 (t, 1H, J = 4.8 Hz),
6.25 (s, 1H, NH), 5.26 (s, 1H), 3.51 (s, 3H, NMe), 3.28
(s, 3H, NMe), 2.49 (s, 3H), 2.16 (s, 3H); 13C (CDCl3,
50 MHz) 199.4, 161.5, 150.8, 146.2, 142.0, 141.8, 137.3,
134.8, 128.6, 124.2, 140.2, 119.6, 91.7, 37.8, 29.9, 28.7,
28.2, 19.8; Anal. Calcd for C18H18N4O5: C, 58.37; H,
4.90; N, 15.13. Found: C, 58.52; H, 4.64; N, 15.04.
4.2.10. 6-Acetyl-1,3,7-trimethyl-5-(4-nitro-phenyl-5,8-dihy-
dro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2j). MS: 371
(M+1); IR (KBr) 3028, 2960, 2150, 1709, 1685, 1610,
4.2.5. 6-Acetyl-5-(4-fluoro-phenyl)-1,3,7-trimethyl-5,8-dihy-
dro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2e). MS: 344
(M+1); IR (KBr) 3030, 2912, 1705, 1664, 1628, 1514,
1
1508, 1382, 1354 cmÀ1; H NMR (CDCl3, 200 MHz) d
1
1358 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.37
(ppm) 8.07 (d, 2H, J = 8.4Hz), 7.77 (d, 2H, J = 8.4 Hz),
6.15 (s, 1H, NH), 5.18 (s, 1H), 3.48 (s, 3H, NMe), 3.28
(s, 3H, NMe), 2.49 (s, 3H), 2.14 (s, 3H); 13C (CDCl3,
50 MHz) 199.4, 161.5, 150.7, 143.2, 142.1, 141.8, 137.6,
137.2, 119.8, 121.5, 91.4, 37.4, 29.8, 28.7, 28.3, 19.9; Anal.
Calcd for C18H18N4O5: C, 58.37; H, 4.90; N, 15.13.
Found: C, 58.62; H, 4.84; N, 15.04.
(d, 2H, J = 8.4 Hz), 6.99 (d, 2H, J = 8.4 Hz), 5.91 (s,
1H, NH), 5.12 (s, 1H), 3.48(s, 3H, NMe), 3.29 (s, 3H,
NMe), 2.46 (s, 3H), 2.14 (s, 3H); 13C (CDCl3, 50 MHz)
199.3, 161.4, 158.5, 150.7, 141.8, 141.6, 137.2, 133.1,
128.9, 115.6, 91.5, 37.6, 29.8, 28.7, 28.1, 19.8; Anal. Calcd
for C18H18FN3O3: C, 62.97; H, 5.28; N, 12.24. Found: C,
62.72; H, 5. 44; N, 12.46.
4.2.11. 6-Acetyl-5-(3-methoxy-phenyl)-1,3,7-trimethyl-5,8-
dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2k). MS:
356 (M+1); IR (KBr) 3014, 2928, 1710, 1667, 1615,
4.2.6. 6-Acetyl-5-(4-chloro-phenyl)-1,3,7-trimethyl-5,8-dihy-
dro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2f). MS: 360
(M+1); IR (KBr) 3015, 2946, 1708, 1678, 1623, 1508,
1
1515, 1379 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm)
1
1376 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.33
7.16 (d, 1H, J = 8.4 Hz), 7.00 (s, 1H), 6.93 (d, 1H,
J = 8.5 Hz), 6.72 (t, 1H, J = 5.2 Hz), 6.10 (s, 1H, NH),
5.12 (s, 1H), 3.81 (s, 3H, OMe), 3.45 (s, 3H, NMe),
3.29 (s, 3H, NMe), 2.45 (s, 3H), 2.15 (s, 3H). 13C (CDCl3,
50 MHz): 199.5, 161.5, 158.5, 150.8, 142.0, 141.8, 137.1,
128.6, 128.4, 122.2, 114.2, 111.9, 91.5, 55.1, 37.4, 29.8,
28.7, 28.2, 19.8; Anal. Calcd for C19H21N3O4: C, 64.21;
H, 5.96; N, 11.82. Found: C, 63.98; H, 5.74; N, 11.99.
(d, 2H, J = 8.3 Hz), 7.25 (d, 2H, J = 8.3 Hz), 6.12 (s,
1H, NH), 5.09 (s, 1H), 3.45 (s, 3H, NMe), 3.28 (s, 3H,
NMe), 2.43 (s, 3H), 2.14 (s, 3H); 13C (CDCl3, 50 MHz)
199.5, 161.5, 150.8, 141.8, 141.6, 137.2, 134.4, 130.5,
129.2, 127.6, 91.6, 37.5, 29.8, 28.7, 28.1, 19.8; Anal. Calcd
for C18H18ClN3O3: C, 60.09; H, 5.04; N, 11.68. Found: C,
60.32; H, 5.24; N, 11.84.
4.2.7. 6-Acetyl-5-(3,4-dichloro-phenyl)-1,3,7-trimethyl-5,8-
dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2g). MS:
394 (M+1); IR (KBr) 3015, 2946, 1708, 1678, 1623, 1508,
4.2.12. 6-Acetyl-5-(4-methoxy-phenyl)-1,3,7-trimethyl-5,8-
dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2l). MS:
356 (M+1); IR (KBr) 3025, 2953, 1705, 1682, 1619, 1502,
1
1
1376 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.38
1388 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.30
(d, 1H, J = 7.8 Hz), 7.30 (d, 1H, J = 7.8 Hz), 7.26 (s,
1H), 5.95 (s, 1H, NH), 5.12 (s, 1H), 3.44 (s, 3H, NMe),
3.28 (s, 3H, NMe), 2.44 (s, 3H), 2.14 (s, 3H); 13C (CDCl3,
50 MHz) 199.5, 161.6, 150.7, 141.9, 141.8, 137.2, 135.4,
132.8, 130.8, 127.2, 129.8, 128.7, 91.6, 37.4, 29.7, 28.6,
28.2, 19.8; Anal. Calcd for C18H17Cl2N3O3: C, 54.84; H,
4.35; N, 10.66. Found: C, 54.62; H, 4.44; N, 10.34.
(d, 2H, J = 8.6 Hz), 6.81 (d, 2H, J = 8.6 Hz), 6.10 (s, 1H,
NH), 5.07 (s, 1H), 3.74 (s, 3H, OMe), 3.44 (s, 3H, NMe),
3.28 (s, 3H, NMe), 2.43 (s, 3H), 2.14 (s, 3H). 13C (CDCl3,
50 MHz): 199.5, 161.5, 158.5, 150.8, 142.0, 141.8, 137.1,
128.8, 128.4, 113.9, 91.5, 55.1, 37.4, 29.8, 28.7, 28.2, 19.8;
Anal. Calcd for C19H21N3O4: C, 64.21; H, 5.96; N, 11.82.
Found: C, 64.05; H, 6.24; N, 11.94.
4.2.8. 6-Acetyl-1,3,7-trimethyl-2,4-dioxo-1,2,3,4,5,8-hexa-
hydro-pyrido[2,3-d]pyrimidine-5-yl-benzonitrile (2h). MS:
351 (M+1); IR (KBr) 3026, 2958, 2155, 1706, 1686,
4.2.13. 6-Acetyl-5-(3,4-dimethoxy-phenyl)-1,3,7-trimethyl-
5,8-dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2m). MS:
386 (M+1); IR (KBr) 3025, 2953, 1705, 1682, 1619, 1502,
1
1
1619, 1508, 1384 cmÀ1; H NMR (CDCl3, 200 MHz) d
1388 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.02
(ppm) 7.34 (d, 2H, J = 8.4 Hz), 7.12 (d, 2H,
J = 8.4 Hz), 6.10 (s, 1H, NH), 5.10 (s, 1H), 3.45 (s, 3H,
NMe), 3.28 (s, 3H, NMe), 2.45 (s, 3H), 2.16 (s, 3H);
13C (CDCl3, 50 MHz) 199.5, 161.5, 150.8, 142.1, 141.8,
140.6, 137.2, 129.8, 127.9, 115.5, 118.2, 91.5, 37.5,
29.8, 28.7, 28.2, 19.8; Anal. Calcd for C19H18N4O3: C,
65.13; H, 5.18; N, 15.99. Found: C, 65.42; H, 5.34; N,
16.14.
(d, 1H, J = 7.8 Hz), 6.92 (d, 1H, J = 7.8 Hz), 6.84 (s,
1H), 6.10 (s, 1H, NH), 5.06 (s, 1H), 3.74 (s, 6H, 2OMe),
3.44 (s, 3H, NMe), 3.28 (s, 3H, NMe), 2.43 (s, 3H), 2.15
(s, 3H); 13C (CDCl3, 50 MHz): 199.5, 161.4, 150.8, 146.3,
142.5, 141.8, 141.6, 137.2, 129.4, 121.8, 115.4, 114.7, 91.7,
54.9, 37.6, 29.8, 28.8, 28.2, 19.9; Anal. Calcd for
C20H23N3O5: C, 62.33; H, 6.01; N, 10.90. Found: C,
62.45; H, 6.23; N, 10.84.
4.2.9. 6-Acetyl-1,3,7-trimethyl-5-(3-nitro-phenyl)-5,8-dihy-
dro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2i). MS: 371
(M+1); IR (KBr) 3026, 2958, 2155, 1706, 1686, 1619,
4.2.14. 6-Acetyl-1,3,7-trimethyl-5-(3,4,5-trimethoxy-phen-
yl)-5,8-dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2n).
MS: 416 (M+1); IR (KBr) 3025, 2953, 1705, 1682, 1619,
1
1550, 1508, 1384, 1350 cmÀ1
;
1H NMR (CDCl3,
1502, 1388 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm)

A. Agarwal et al. / Bioorg. Med. Chem. 13 (2005) 6678–6684
6683
6.62 (s, 2H), 5.99 (s, 1H, NH), 5.10 (s, 1H), 3.82 (s, 6H,
2OMe), 3.79 (s, 3H, OMe), 3.45 (s, 3H, NMe), 3.31 (s,
3H, NMe), 2.46 (s, 3H), 2.17 (s, 3H); 13C (CDCl3,
50 MHz) 199.5, 161.5, 149.2, 150.8, 142.0, 141.8, 137.1,
130.2, 128.4, 105.4, 91.5, 61.4, 56.8, 37.4, 29.8, 28.7,
28.2, 19.8. Anal. Calcd for C21H25N3O6: C, 60.71; H,
6.07; N, 10.11. Found: C, 60.39; H, 5.84; N, 10.34.
and dried over sodium sulfate to give the crude product,
which was purified by column chromatography (hexane/
ethyl acetate; 7:3) to give 2, 3, and 4.
4.3.1. 6-Acetyl-5-(4-chloro-phenyl)-1,3,7-trimethyl-1H-pyri-
do[2,3-d]pyrimidine-2,4-dione (3f). MS: 358 (M+1); IR
(KBr) 3015, 2962, 1712, 1652, 1558, 1541, 1508,
1482 cmÀ1; 1H NMR (CDCl3, 200 MHz) d (ppm) 7.54 (d,
2H, J = 8.2 Hz), 7.43 (d, 2H, J = 8.2 Hz), 3.74 (s, 3H,
NMe), 3.32 (s, 3H, NMe), 2.55 (s, 3H, COCH3), 1.82 (s,
3H, CH3); 13C (CDCl3, 50 MHz) 204.4, 160.3, 159.6,
151.2, 150.4, 149.5, 139.7, 132.4, 130.2, 127.4, 120.9, 105.5,
31.7, 29.9, 28.3, 23.5; Anal. Calcd for C18H16N3O3: C,
60.42; H, 4.51; N, 11.74. Found: C, 60.52; H, 4.44; N, 12.04.
4.2.15. 6-Acetyl-5-(4-hydroxy-phenyl)-1,3,7-trimethyl-5,8-
dihydro-1H-pyrido[2,3-d]pyrimidine-2,4-dione (2o). MS:
342 (M+1); IR (KBr) 3420, 2953, 1709, 1682, 1502,
1
1388 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm) 7.19
(d, 2H, J = 8.5 Hz), 6.74 (d, 2H, J = 8.5 Hz), 5.00 (s,
1H), 3.48 (s, 3H), 3.21 (s, 3H), 2.43 (s, 3H), 2.15 (s, 3H);
13C (CDCl3, 50 MHz) 199.5, 161.5, 158.5, 150.8, 142.0,
141.8, 137.1, 128.6, 128.2, 113.7, 91.5, 37.4, 29.8, 28.7,
28.2, 19.8. Anal. Calcd for C18H19N3O4: C, 63.33; H,
5.61; N, 12.31. Found: C, 63.61; H, 5.82; N, 12.58.
4.3.2. 4-(6-Acetyl-1,3,7-trimethyl-2,4-dioxo-1,2,3,4-tetra-
hydro-pyrido[2,3-d]pyrimidin-5-yl)-benzonitrile (3h). MS:
349 (M+1); IR (KBr) 3008, 2948, 2150, 1708, 1675,
1546, 1553, 1514, 1484 cmÀ1
;
1H NMR (CDCl3,
4.2.16. 4-(6-Acetyl-1,3,7-trimethyl-2,4-dioxo-1,2,3,4,5,8-
hexa-hydro-pyrido[2,3-d]pyrimidin-5-yl)-benzoic acid (2p).
MS: 370 (M+1); IR (KBr) 3446, 2923, 1704, 1678, 1509,
1382 cmÀ1; 1H NMR (CDCl3, 200 MHz) d (ppm) 7.97 (d,
2H, J = 8.9 Hz), 7.46 (d, 2H, J = 8.9 Hz), 5.18 (s, 1H),
3.54 (s, 3H), 3.27 (s, 3H), 2.45 (s, 3H), 2.13 (s, 3H); 13C
(CDCl3, 50 MHz) 199.5, 161.5, 150.8, 144.5, 142.0,
141.8, 137.1, 134.5, 131.8, 126.4, 91.5, 37.4, 29.8, 28.7,
28.2, 19.8; Anal. Calcd for C19H19N3O5: C, 61.78; H,
5.18; N, 11.38. Found: C, 61.56; H, 5.34; N, 11.62.
200 MHz) d (ppm) 7.18 (d, 2H, J = 7.9 Hz), 7.01 (d,
2H, J = 7.9 Hz), 3.72 (s, 3H, NMe), 3.38 (s, 3H,
NMe), 2.52 (s, 3H), 1.86 (s, 3H); 13C (CDCl3,
50 MHz) 204.5, 160.4, 159.6, 151.2, 150.6, 149.6, 146.7,
130.2, 129.3, 120.4, 115.4, 111.5, 105.5, 31.8, 29.9,
28.4, 23.6; Anal. Calcd for C19H16N4O3: C, 65.51; H,
4.63; N, 16.08. Found: C, 65.72; H, 4.34; N, 16.24.
4.3.3. 6-Acetyl-5-(4-methoxy-phenyl)-1,3,7-trimethyl-1H-
pyrido[2,3-d]pyrimidine-2,4-dione (3l). MS: 354 (M+1);
IR (KBr) 3005, 2957, 1706, 1665, 1558, 1558, 1513,
1480 cmÀ1;1H NMR (CDCl3, 200 MHz) d (ppm) 7.14
(d, 2H, J = 8.6Hz), 6.97 (d, 2H, J = 8.6 Hz), 3.85 (s,
3H, OMe), 3.76 (s, 3H, NMe), 3.35 (s, 3H, NMe), 2.53
(s, 3H, COCH3), 1.85 (s, 3H, CH3); 13C (CDCl3,
50 MHz) 204.4, 160.3, 159.8, 158.4, 151.2, 150.8, 149.9,
136.7, 129.6, 120.7, 105.2, 113.4, 55.3, 31.7, 29.9, 28.3,
23.5; Anal. Calcd for C19H19N3O4: C, 64.58; H, 5.42;
N, 11.89. Found: C, 64.82; H, 5.34; N, 12.04.
4.2.17. 6-Acetyl-5-furan-2-yl-1,3,7-trimethyl-5,8-dihydro-
1H-pyrido[2,3-d]pyrimidine-2,4-dione (2q). MS: 316
(M+1); IR (KBr) 2948, 2936, 1704, 1656, 1532, 1512,
1
1432, 1373 cmÀ1; H NMR (CDCl3, 200 MHz) d (ppm)
7.24 (d, 1H, J = 4.8 Hz), 6.34 (t, 1H, J = 3.2Hz), 6.22 (d,
1H, J = 4.5 Hz), 6.07 (s, 1H, NH), 5.25 (s, 1H), 3.49 (s,
3H, NMe), 3.34 (s, 3H, NMe), 2.40 (s, 3H), 2.28 (s, 3H).
13C (CDCl3, 50 MHz): 199.2, 161.4, 152.5, 150.8, 142.0,
141.8, 137.1, 141.2, 110.2, 105.6, 91.5, 55.1, 37.4, 29.8,
28.7, 28.2, 19.8. Anal. Calcd for C16H17N3O4: C, 60.94;
H, 5.43; N, 13.33. Found: C, 60.67; H, 5.24; N, 13.11.
4.3.4. 6-Acetyl-1,3,7-trimethyl-1H-pyrido[2,3-d]-pyrimidine-
2,4-dione (4). MS: 248 (M+1); IR (KBr) 2960, 2928, 1702,
1
1663, 1564, 1509, 1440, 1371 cmÀ1; H NMR (CDCl3,
4.2.18. 6-Acetyl-1,3,5,7-tetramethyl-5,8-dihydro-1H-pyri-
do[2,3-d]pyrimidine-2,4-dione (2r). MS: 264 (M+1); IR
(KBr) 2963, 2924, 1708, 1665, 1544, 1502, 1428,
1368 cmÀ1; 1H NMR (CDCl3, 200 MHz) d (ppm) 5.97 (s,
1H, NH), 4.02 (dd, 1H, J = 6.8 Hz), 3.49 (s, 3H, NMe),
3.34 (s, 3H, NMe), 2.34 (s, 3H), 2.28 (s, 3H), 1.13 (s, 3H);
13C (CDCl3, 50 MHz) 199.7, 161.6, 150.9, 141.9, 141.1,
139.6, 95.5, 30.4, 29.9, 28.8, 28.2, 19.8, 15.2; Anal. Calcd
for C13H17N3O3: C, 59.30; H, 6.51; N, 15.96. Found: C,
59.53; H, 6.34; N, 15.74.
200 MHz) d (ppm) 6.81 (s, 1H), 3.68 (s, 3H), 3.43 (s,
3H), 2.76 (s, 3H), 2.52 (s, 3H); 13C (CDCl3, 50 MHz)
162.8, 162.3, 153.4, 151.8, 122.2, 107.2, 30.2, 28.5, 25.0,
22.6; Anal. Calcd for C12H13N3O3: C, 58.29; H, 5.30; N,
17.00. Found: C, 58.48; H, 5.12; N, 17.24.
Acknowledgments
A.A. thanks the Council of Scientific and Industrial
Research (India) for the award of Senior Research
Fellowship. We are also thankful to S.A.I.F. Division,
CDRI, Lucknow, for providing spectroscopic data.
CDRI Communication No. 6764.
4.3. General procedure for the synthesis of 6-acetyl-5-(aryl)-
1,3,7-trimethyl-1H-pyrido[2,3-d]pyrimidine-2,4-dione (3) and
6-acetyl-1,3,7-trimethyl-1H-pyrido[2,3-d]pyrimidine-2,4-dione
(4)
6-Amino-1,3-dimethyl uracil (I) (0.3 mmol), acetyl ace-
tone (0.35 mmol) and aryl aldehyde (0.35 mmol) were
heated in acetic acid (15 mL) for 48 h. The mixture
was allowed to cool and water (50 mL) was added.
The reaction mixture was extracted with chloroform
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