Regioselective addition reactions at C-2 of 3,4-dihydropyrimidinones. Synthesis and evaluation of multifunctionalized tetrahydropyrimidines

Article abstract of DOI:10.1016/j.tet.2010.08.034

Multifunctionalized tetrahydropyrimidines derivatives have been synthesized from Biginelli 3,4-dihydropyrimidin-2-(1H)-thiones (DHPMs) efficiently. The transformation includes desulfurization of DHPMs with Raney-Ni and subsequent regioselective C-2 functi

Full text of DOI:10.1016/j.tet.2010.08.034

Tetrahedron 66 (2010) 8175e8180
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Regioselective addition reactions at C-2 of 3,4-dihydropyrimidinones. Synthesis
and evaluation of multifunctionalized tetrahydropyrimidines
,
a
b
Kamaljit Singh a , Divya Arora , Jan Balzarini
*
a Organic Synthesis Laboratory, Department of Applied Chemical Sciences & Technology, Guru Nanak Dev University, Amritsar, Punjab 143 005, India
b Rega Institute for Medical Research, Katholieke Universiteit Leuven, 10 Minderbroedersstraat, B-3000 Leuven, Belgium
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 29 June 2010
Received in revised form 11 August 2010
Accepted 12 August 2010
Available online 19 August 2010
Multifunctionalized tetrahydropyrimidines derivatives have been synthesized from Biginelli 3,4-dihy-
dropyrimidin-2-(1H)-thiones (DHPMs) efficiently. The transformation includes desulfurization of DHPMs
with Raney-Ni and subsequent regioselective C-2 functionalization using a variety of C-nucleophiles with
simultaneous activation with ethyl chloroformate. Functionalized pyrimidine derivatives containing
bulky substituents at C-2 of the pyrimidine ring are cytostatic.
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Biginelli compounds
Carbon nucleophiles
Pyrimidines
Regioselectivity
Desulfurization
1. Introduction
These reactions proceeded with high regio- and chemoselectivity
and without any chemical activation of pyrimidinone.15e17
The elaboration of heterocycles represents one of the most
vibrant research areas in organic chemistry and has a rich history
within the realm of fragment-based drug design.1 3,4-Dihy-
dropyrimidin-2-(1H)-ones (DHPMs) and their appropriately func-
tionalized derivatives have interesting pharmacological profiles.
These are potent antihypertensive agents,2 mitotic kinesin inhib-
itors,3e6 a1a-adrenergic receptor antagonists,7 or hepatitis B virus
replication inhibitors8 and depict a variety of other biological
effects.9,10 Consequently, synthetic investigations on DHPMs have
received extensive attention by both synthetic as well as medicinal
chemists. Although a large number of DHPM derivatives have been
prepared in a single-pot Biginelli multi component reaction (MCR)
and its variants, very useful and convincing structural variability of
these interesting heterocycles have been achieved by chemical
functionalization of the six positions around the DHPM core.11 We
have also recently reported highly regio- as well as chemoselective
functionalization of Biginelli DHPMs at the C-6 methyl12 position
alone or in combination with N-1 (leading to bicyclic products), N-1
and N-3 positions with a wide variety of substituents.13,14 Likewise,
elaboration of the C-4 position of DHPMs through addition of
C-nucleophiles on the precursor pyrimidinones has been achieved.
Multifunctionalized pyrimidine derivatives form basic skeleton
of a wide range of biologically active molecules and are often pro-
cured through multistep routes. Many important pyrimidine
derivatives such as Bay 41e4109 1, Bay 39-5493 2 (non-nucleosidic
inhibitor of hepatitis B virus),8 HAP-1 3,18 N-cyano-iminopyr-
imidine 4 (potent antimycotic agent),19 the hypocholesterolemic
agent Rosuvastatin (Ca salt) 5 (HMG-CoA reductase inhibitor)20 and
the potent anticancer drug Gleevec 6 (a tyrosine kinease in-
hibitor)21 contain a C-2 functionalized pyrimidine core (Fig. 1).
Biginelli DHPMs could be visualized as ideal precursors for
obtaining pyrimidine derivatives through simple transformations.
A survey of recent literature revealed different approaches to
access pyrimidines from pyrimidinone derivatives.22,23 For exam-
ple, pyrimidines bearing a sulfone moiety at C-2, obtained through
treatment of 2-thioxopyrimidines with oxone can react with
nucleophiles23 to obtain 2-substituted pyrimidines. Alternatively,
a palladium(0)-catalyzed copper(I)-mediated coupling of boronic
acids with cyclic thioamides has been used.24 Kang et al. have
reported a two-step procedure for efficient transformation of
Biginelli DHPMs to pyrimidines via PyBroP-mediated coupling.25
Similarly, pyrimidines have been reported via palladium cata-
lyzed Suzuki Sonogashira cross-coupling and Eschenmoser sulfide
contraction reactions.26,27 However, these useful approaches are
multistep as well as involve expensive reagents and lack generality
in many instances. Regioselective addition reactions at C-2 position
* Corresponding author. Tel.: þ91 183 2258853; fax: þ91 183 2258819/20; e-mail
address: kamaljit19in@yahoo.co.in (K. Singh).
0040-4020/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2010.08.034

8176
K. Singh et al. / Tetrahedron 66 (2010) 8175e8180
F
F
F
Cl
CN
NH
Cl
Cl
N
N
MeOOC
Me
EtOOC
Me
N
F
MeOOC
N
N
N
N
H
S
N
H
N
H
N
Me
F
N
H
1
4
2
3
F
O
N
HN
OH OH
ROOC
Me
OOC
N
N
N
N
Me
SO2Me
N
N
N
N
N
N
H
H
Me
N
Me
Me
Me
7 (R = Me)
8 (R = Et)
5
6
Figure 1. Medicinally potent C-2 functionalized pyrimidine derivatives.
of DHPMs appeared to be an attractive tool for introducing diversity
at the C-2 position in order to probe the effect of substitution on the
cytostatic activity. Herein, we report a general and practicable
protocol for preparing a number of C-2 substituted pyrimidine
derivatives and results of their cytostatic activity.
a synthetically useful manner (Table 1). The overall reaction thus
depicts a new carbonecarbon bond formation at the C-2 position.
Representative compounds have also been screened for cytostatic
activity. 3,4-Dihydropyrimidin-2-(1H)-thione 9 was obtained in
quantitative yield through the reaction of methyl acetoacetate,
benzaldehyde, and thiourea under acid catalysed conditions of
Biginelli condensation reaction.29 Desulfurisation28 of 9 was ach-
ieved using Raney-Ni, under atmosphere of hydrogen gas and C-2
2. Results and discussion
unsubstituted
5-methoxycarbonyl-6-methyl-4-phenyl-1,4-dihy-
We envisaged that electronegativity of the two nitrogen atoms
could invoke considerable electrophilic character to the methine
carbon (C-2) flanked by two nitrogen atoms in the 1,4-dihy-
dropyrimidine derivative 10 (Scheme 1), and consequently, under
dropyrimidine 10 was obtained in 60% yield. Reaction of 10 with
various nucleophilic species, such as carbanion of acetone, thio-
phene, Grignard reagents of alkyl groups (short, medium, and long
MeOOC
Me
MeOOC
Me
MeOOC
ClCOOEt
NCOOEt
NH
N
R1
N
H
S
(R1Li/R1MgX)
11
N
H
H
Me
N
H
H
10
12
9
Scheme 1. Synthesis of C-2 elaborated tetrahydropyrimidines through regioselective C-2 addition reactions on 1,4-dihydropyrimidine derivative.
Table 1
appropriate reaction conditions, addition of nucleophiles 11 may be
facilitated, exclusively at C-2 position, with or without any chemical
activation of the nitrogen centers. This would allow synthesis of C-2
functionalized pyrimidine derivatives 12 with retention of chemical
diversity at other positions. Such addition reactions at C-2 of 10
shall open a novel general route for accessing multifunctionalized
pyrimidine derivatives, which are otherwise inaccessible through
the conventional approaches. In view of facile desulfurisation28 of
9e10, using Raney-Ni, synthesis of 12 may be visualized as a single
step approach from 10. Indeed, it has been found that when 10 was
treated with a number of carbon nucleophiles, with simultaneous
activation of N-3 with ethyl chloroformate (Scheme 1), synthesis of
a number of multifunctionalized pyrimidine derivatives 12, bearing
a range of substituents, such as alkyl, aryl, and heterocyclyl at its
C-2 position was achieved with high regioselectivity and in
Addition of carbon nucleophiles to 5-methoxycarbonyl-6-methyl-4-phenyl-1,4-di-
hydropyrimidine 10. Formation of C-2 substituted DHPM derivatives 12 (Scheme 1)
Entrya
C-Nucleophile
11 R1Li/MgX
Product
12 R1
Yieldb (%)
Mp (ꢀC)
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11a LiCH2COMe
11b 2-Thienyllithium
11c MeMgI
12a CH2COMe
12b Thien-2yl
12c Me
12d Et
12e n-Pr
60
68
65
64
67
70
72
62
70
68
120e121
155e156
135e136
128e129
120e121
96e97
98e99
Liquid
95e96
140e141
11d EtMgBr
11e n-PrMgBr
11f n-BuMgBr
11g n-C5H11MgBr
11h n-C11H23MgBr
11i AllylMgBr
11j PhMgBr
12f n-Bu
12g n-C5H11
12h n-C11H23
12i allyl
12j Ph
a
Compound 15 (Scheme 2) was also detected in 10e15% in each case.
Isolated purified yields.
b

K. Singh et al. / Tetrahedron 66 (2010) 8175e8180
8177
aliphatic chains), allyl and phenyl groups proceeded smoothly
and exclusively at the C-2 position in the presence of ethyl
chloroformate.
CeC bonds. Thus, treatment of 10 with freshly prepared methyl
magnesium iodide at ꢁ5 ꢀC, under an atmosphere of dry nitrogen
gas furnished 3-ethoxycarbonyl-5-methoxycarbonyl-2,6-dimethyl-
4-phenyl-1,2,3,4-tetrahydropyrimidine 12c in 65% yield (Table 1).
Other Grignard reagents32 bearing aliphatic carbon chains of length
varying from C1e5 and C11 reacted with 10, analogously at ꢁ5 ꢀC
under an atmosphere of nitrogen to furnish the corresponding
pyrimidine derivatives 12deh (Table 1, entries 4e8) in good yields.
Similar reaction with allylmagnesium bromide furnished C-2
addition product 12i in 70% yield after column chromatography.
The reaction of phenylmagnesium bromide with 10 was also very
facile and furnished the pyrimidine derivative 12j in 68% yield.
While the deprotection of carbamate from N-3 positions of prod-
ucts 12aej using LiAlH4 in methanol17 (stirring at 0 ꢀC to room
temperature and then reflux), NaOMe in refluxing methanol33 or
tetrabutyl ammonium fluoride (TBAF) in THF34 could not be ach-
ieved, the current methodology served the purpose of appending
a variety of substituents at the C-2 position, regioselectively and in
a synthetically useful manner.
When a solution of carbanion of acetone30 11a (LDA, THF,
ꢁ78 ꢀC) and ethyl chloroformate were slowly added, simulta-
neously, to a stirring solution of 10 in anhydrous THF maintained at
ꢁ5 ꢀC, 12a was obtained in 60% isolated yield. The structure of 12a
was confirmed through the spectral and physical data (vide ex-
perimental). To provide an unambiguous characterization of 12a,
the structure was additionally confirmed from single crystal X-ray
analysis (Fig. 2). Interestingly, in this reaction a side product 15
(Scheme 2) was also obtained.
The formation of the products 12 and 15 can be visualized to be
proceeding through the mechanism shown in Scheme 2. Thus,
during the simultaneous addition of ethyl chloroformate as well as
pre-generated anion (R1Li/MgX) to a solution of 10, the in situ
generated intermediate species 13 can undergo reaction with 11 to
furnish 12, as well as with chloride ion, formed as a consequence of
the N-3 activation, leading to 14, which upon dehydrohalogenation
may give 15, which may also arise from 13 through deprotonation.
In support of this mode of reaction, when under similar reaction
conditions, a solution of 10 was stirred with ethyl chloroformate, 15
was formed in 45% yield.
Representative members22 of the newly synthesized com-
pounds 12 were evaluated for their inhibitory effect against the
proliferation of murine leukemia (L1210), murine mammary car-
cinoma (FM3A), human T-lymphocyte (CEM), and human cervix
carcinoma (HeLa) cells (Table 2). Whereas, pyrimidine derivatives
Figure 2. Ortep diagram of 12a (CCDC-735335) showing stereo view of the molecule
and the numbering scheme used in the structure analysis. (N2, N1, C1, C3 positions
may be read as N1, N3, C4, and C6, respectively, following IUPAC nomenclature
scheme).
COOEt
H
MeOOC
MeOOC
Me
COOEt
R1
(R1Li/R1MgX)
N
N
Me
N
H
N
H
H
11
12
13
ClCOOEt
H
MeOOC
Me
N
Cl
N
H
10
MeOOC
Me
COOEt
Cl
MeOOC
Me
COOEt
H
N
-HCl
N
N
H
N
H
15
14
Scheme 2. Proposed mode of addition reactions of R1Li/R1MgX to 10.
Likewise, red colored anion 11b formed as a result of lithiation of
thiophene31 using n-BuLi at ꢁ40 ꢀC upon quenching with 10, in the
presence of ethyl chloroformate resulted in the formation of the
corresponding product 12b (Table 1).
The nucleophilic addition reaction of Grignard reagents to the
activated carbonecarbon and carboneheteroatom double bonds
constitutes one of the most versatile methods for the formation of
bearing acetylmethyl, methyl, ethyl and n-propyl substituents at
the C-2 position (i.e., 12a, 12c, 12d, and 12e, Table 2) depicted
a
marginal cytostatic activity of the test compounds (IC50
56e434 M), more bulky alkyl groups such as butyl (12f) or pentyl
(12g) rendered a 5- to 10-fold higher antiproliferative activity (IC50
23e45 M). The C2-phenyl substituted pyrimidine derivative 12j
showed a higher cytostatic activity against HeLa cells.
:
m
:
m

8178
K. Singh et al. / Tetrahedron 66 (2010) 8175e8180
Table 2
using flash chromatography using silica gel (60e120 mesh) and
mixtures of ethyl acetate/hexane as eluent.
Inhibitory effect against the proliferation of murine leukemia (L1210), murine
mammary carcinoma (FM3A), human T-lymphocyte (CEM) and human cervix car-
cinoma (HeLa) cells
4.2. Synthesis of 5-methoxycarbonyl-6-methyl-4-phenyl-3,4-
dihydropyrimidin-2-(1H)-thione 9
a
Compound
IC50
(m
M)
L1210
FM3A
CEM
HeLa
A solution of methyl acetoacetate (0.35 mol), benzaldehyde
(0.35 mol), and thiourea (0.52 mol) in methanol (30e40 mL) con-
taining dilute hydrochloric acid (0.5 mL) was refluxed till the
reaction was completed (TLC). The product got directly precipitated
in the reaction flask as off-white solid, which was filtered and
washed with water (2ꢂ50 mL). Recrystallization from ethanol gave
the pure 5-methoxycarbonyl-6-methyl-4-phenyl-3,4-dihydropyr-
imidin-2-(1H)-thione 9 in 90% yield as an off-white solid; [found: C,
59.51; H, 5.11; N, 10.45; S, 12.11. C13H14N2O2S requires C, 59.54; H,
5.34; N, 10.68; S, 12.21%]; Rf (45% EtOAc/hexane) 0.5; mp:
205e206 ꢀC (methanol); dH (300 MHz, CDCl3þDMSO-d6) 9.46 (1H,
s, D2O exchangeable, NH), 8.78 (1H, s, D2O exchangeable, NH),
7.24e7.41 (5H, m, ArH), 5.35 (1H, d, J 2.7 Hz, C4eH), 3.62 (3H, s,
estereCH3), 2.36 (3H, s, CH3); dC (75 MHz, CDCl3) 174.2, 165.6, 144.1,
142.7, 128.0, 127.2, 126.2, 101.1, 54.6, 50.5, 17.3; m/z 285 (Mþ23).
12a
12c
12d
12e
12f
330ꢃ52
146ꢃ130
206ꢃ16
128ꢃ9
37ꢃ4
434ꢃ93
385ꢃ163
168ꢃ15
142ꢃ11
34ꢃ7
412ꢃ124
169ꢃ13
134ꢃ59
56ꢃ12
45ꢃ8
210ꢃ23
114ꢃ30
65ꢃ3
98ꢃ72
28ꢃ2
12 g
12j
37ꢃ3
35ꢃ3
42ꢃ10
76ꢃ20
23ꢃ2
75ꢃ7
77ꢃ8
10ꢃ18
a
Inhibitory concentration (50%).
It is also worth noticing that the cytostatic activity of the in-
dividual test compounds was independent of the nature of the
tumor cell line (either murine vs human, or carcinoma vs leukemia
(lymphoma)). It would now be imperative to further explore the
C-2 position on the pyrimidine to potentiate the cytostatic activity
of the test compounds.
3. Conclusion
4.3. Desulfurisation of 5-methoxycarbonyl-6-methyl-4-
phenyl-3,4-dihydropyrimidin-2-(1H)-thione 9. Synthesis of
C-2 unsubstituted DHPM derivative 10
In summary, a simple and efficient method for the synthesis of
novel multifunctionalized tetrahydropyrimidines from Biginelli
dihydropyrimidin-2-(1H)-thiones has been devised. The method-
ology holds potential for the introduction of a number of tailor-
made nucleophilic fragments at the C-2 position of DHPMs in
a synthetically useful manner. The nature of the C-2 substituent
modulates the cytostatic activity in cell culture.
A solution of compound 9 (5 mmol) in methanol (50 mL) was
treated with excess of Raney-Ni (2 g) and refluxed for approxi-
mately 6 h under hydrogen atmosphere. After complete desulfur-
isation (TLC), reaction was filtered through a Celite bed and the
solvent was removed under reduced pressure. Crystallization of the
crude product from methanol gave the pure C-2 unsubstituted
product, i.e., 5-methoxycarbonyl-6-methyl-4-phenyl-1,4-dihy-
dropyrimidine 10 in 60% yield as an off-white solid; [found: C,
67.78; H, 6.01; N, 12.05. C13H14N2O2 requires C, 67.82; H, 6.08; N,
12.17]; Rf (20% EtOAc/hexane) 0.5; mp: 175e176 ꢀC (DCM); nmax
4. Experimental
4.1. General
The solvents: acetone (K2CO3), diethyl ether, hexane, and tet-
rahydrofuran (THF) (Naebenzophenone ketyl), alkyl halides
(CaCl2), thiophene (refluxing with Naemetal and then dried over
KOH pellets), diisopropylamine (KOH pellets) were adequately
dried and drawn under N2 atmosphere using hypodermic glass
(KBr): 2877, 1687, 1626 cmꢁ1
; dH (300 MHz, CDCl3) 7.12e7.33 (5H,
m, ArH), 6.91 (1H, s, C2eH), 5.53 (1H, s, C4eH), 3.60 (3H, s,
estereCH3), 2.25 (3H, s, CH3); dC (75 MHz, CDCl3) 167.2, 144.8, 142.5,
128.4, 127.3, 126.9, 100.0, 57.4, 50.9, 18.9; m/z 231 (Mþ1).
ꢀ
syringes. Low boiling reagents were distilled and stored over 4 A
molecular sieves. Freshly prepared n-BuLi (2.0e2.3 N in hexanes)
standardized using diphenylacetic acid was used. Lithium diiso-
propyl amide (LDA) was generated from equimolar quantities of
diisopropylamine and n-BuLi at 0 ꢀC. Grignard reagents were pre-
pared using activated magnesium metal and appropriate alkyl
halides in anhydrous diethyl ether. Reactions were run under
a blanket of dry nitrogen gas in a sealed (rubber septum, Aldrich)
round-bottomed flasks. Organometallic reagents were added using
cannula. The low temperature was achieved by using slush of liquid
nitrogen with appropriate solvent.
4.4. Synthesis of 3-ethoxycarbonyl-5-methoxycarbonyl-6-
methyl-2-(2-oxopropyl)-4-phenyl-1,2,3,4-tetrahydropyrimidine
12a and 3-ethoxycarbonyl-5-methoxycarbonyl-6-methyl-4-
phenyl-3,4-dihydropyrimidine 15
To a solution of diisopropylamine (17.39 mmol) in THF (2 mL),
n-BuLi (17.39 mmol) was added dropwise at ꢁ78 ꢀC, under nitrogen
atmosphere. The solution was allowed to warm to 0 ꢀC and stirred
for an additional 10 min. The solution was cooled to ꢁ78 ꢀC and
pre-cooled THF (25 mL at ꢁ78 ꢀC) was added with stirring. Acetone
(17.39 mmol) dissolved in THF (10 mL) was then introduced via
hypodermic syringe and the solution stirred for additional 10 min
to ensure complete deprotonation to monoanion 11a. The mono-
anion 11a was then introduced with the help of a cannula, to
a round-bottomed flask containing a solution of 10 (4.34 mmol) in
anhydrous THF (20 mL) with simultaneous addition of ethyl
chloroformate (4.34 mmol) at ꢁ5 ꢀC. Reaction was warmed to room
temperature and stirring was continued to complete the reaction
(TLC), after which a saturated aqueous solution of NH4Cl was
introduced. The reaction was extracted with ethyl acetate
(3ꢂ25 mL) and the extract washed once with brine (100 mL). The
extract was dried over anhydrous Na2SO4 and the solvent removed
under reduced pressure. Column chromatography of the residue
over silica gel (60e120 mesh), using mixtures of hexane and ethyl
acetate furnished two products. The product 12a was isolated in
Melting points were determined in open capillaries and are
uncorrected. 1H NMR (300 and 400 MHz) spectra were recorded on
Jeol FT-AL-300 and Bruker-400 MHz multinuclear spectrometers,
using deuterated solvents. All chemical shifts are reported in part
per million (d) relative to tetramethylsilane (TMS, d 0.0) used as
internal reference standard. 13C NMR spectra were recorded at
75 MHz on Jeol FT-AL-300 and 100 MHz on Bruker-400 in-
struments. EI mass spectra (MS) were recorded on Bruker Daltonics
esquire 3000 spectrometer. Elemental analyses were performed on
FLASH EA 112 (Thermo Electron Corporation) analyzer and the
results are quoted in %. For monitoring the progress of a reaction
and for comparison purpose, thin layer chromatography (TLC) was
performed on pre-coated aluminium sheets Merck (60 F254
,
0.2 mm) using an appropriate solvent system. The chromatograms
were visualized under UV light. The compounds were purified

K. Singh et al. / Tetrahedron 66 (2010) 8175e8180
8179
60% yield as a white solid; [found: C, 63.07; H, 6.83; N, 7.70.
C19H24N2O5 requires C, 63.33; H, 6.66; N, 7.77]; Rf (15% EtOAc/
hexane) 0.65; mp: 120e121 ꢀC (DCM); nmax (KBr): 3280, 2971, 1720,
anhydrous ether, was transferred with the help of cannula to
a solution of 10 (4.34 mmol) in anhydrous THF maintained at
ꢁ5 ꢀC, under nitrogen atmosphere with simultaneous addition of
ethyl chloroformate (4.34 mmol). Reaction was warmed to room
temperature and stirring was continued to complete the reaction
(TLC), after which a saturated aqueous solution of NH4Cl was
introduced. The reaction was extracted with ethyl acetate
(3ꢂ25 mL) treated with brine. The extract was dried over anhy-
drous Na2SO4 and the mixture was concentrated under reduced
pressure. The following products have been isolated after column
chromatography.
1621 cmꢁ1
; dH (400 MHz, CDCl3) 7.23e7.36 (5H m, ArH), 6.42 (1H,
br, NH), 5.78 (1H, br, C2eH), 5.29 (1H, br, C4eH), 4.30 (2H, br,
estereCH2), 3.56 (3H, s, estereCH3), 2.40 (3H, s, CH3), 2.01e2.31
(2H, m, eCH2), 1.85 (3H, s, eCOCH3), 1.29 (3H, br, CH3); dC (100 MHz,
CDCl3) 207.67, 167.4, 128.4, 127.5, 127.3, 62.3, 59.4, 50.6, 30.0, 14.7;
m/z 383 (Mþ23). The product 3-ethoxycarbonyl-5-methoxy-
carbonyl-6-methyl-4-phenyl-3,4-dihydropyrimidine 15 was obta-
ined in 15% yield also as white solid; [found C, 63.55; H, 6.35; N,
8.88. C16H18N2O4 requires C, 63.57; H, 5.96; N, 9.27]; Rf (15% EtOAc/
hexane) 0.65; mp: 70e71 ꢀC (DCM); nmax (KBr): 3310, 2956, 1702,
4.6.1. 3-Ethoxycarbonyl-5-methoxycarbonyl-2,6-dimethyl-4-phenyl-
1,2,3,4-tetrahydropyrimidine (12c). White solid; [found: C, 64.07; H,
6.83; N, 8.70. C17H22N2O4 requires C, 64.15; H, 6.91; N, 8.80]; Rf (15%
EtOAC/hexane) 0.6; mp: 135e136 ꢀC (DCM); nmax (KBr): 3320, 2975,
1621 cmꢁ1
; dH (400 MHz, CDCl3) 8.03 (1H, s, C2eH), 7.26e7.35 (5H,
m, ArH), 6.05 (1H, s, C4eH), 4.25 (2H, m, estereCH2), 3.67 (3H, s,
estereCH3), 2.44 (3H, s, CH3), 1.30 (3H, t, J 7.2 Hz, CH3); dC (100 MHz,
CDCl3) 166.2, 151.4, 144.6, 140.5, 128.6, 128.5, 127.6, 111.6, 63.7, 53.3,
51.6, 22.1, 14.2; m/z 303 (Mþ1).
1705, 1620 cmꢁ1
; dH (300 MHz, CDCl3) 7.17e7.60 (5H, m, ArH), 6.40
(1H, s, NH), 5.60 (1H, br, C2eH), 4.54 (1H, br, C4eH), 4.24 (2H, m,
estereCH2), 3.51 (3H, s, estereCH3), 2.41 (3H, s, CH3), 1.36 (3H, t, J
7.2, CH3), 0.83 (3H, d, J 6.6 Hz, CH3); dC (75 MHz, CDCl3) 171.4, 160.1,
154.9, 138.3, 128.0, 127.8, 127.4, 126.8, 73.6, 67.2, 60.4, 51.4, 22.7,
14.7,14.3d; m/z 341 (Mþ23).
4.4.1. Alternate procedure for 15. To a solution of 10 (4.33 mmol) in
THF, ethyl chloroformate (4.76 mmol) was added slowly at ꢁ5 ꢀC
under nitrogen atmosphere. Reaction was warmed to room tem-
perature and stirring was continued to complete the reaction (TLC),
after which a saturated aqueous solution of NH4Cl was introduced.
The reaction was extracted with ethyl acetate (3ꢂ25 mL) treated in
sequence with brine. The extract was dried over anhydrous Na2SO4
and the mixture was concentrated under reduced pressure. The
corresponding product 15 was isolated after column chromato-
graphy using silica gel (60e120 mesh) and mixtures of ethyl acetate
and hexane as eluent.
4.6.2. 3-Ethoxycarbonyl-5-methoxycarbonyl-2-ethyl-6-methyl-4-
phenyl-1,2,3,4-tetrahydropyrimidine (12d). White solid; [found: C,
65.07; H, 7.13; N, 8.39. C18H24N2O4 requires C, 65.06; H, 7.22; N,
8.43]; Rf (15% EtOAC/hexane) 0.6; mp: 128e129 ꢀC (DCM); nmax
(KBr): 3318, 2976, 1702, 1621 cmꢁ1
; dH (300 MHz, CDCl3)
7.16e7.37 (5H, m, ArH), 6.39 (1H, s, NH), 5.35 (1H, s, C2eH), 4.69
(1H, s, C4eH), 4.26 (2H, q, J 7.2 Hz, estereCH2), 3.54 (3H, s,
estereCH3), 2.40 (3H, s, CH3), 1.36 (3H, br, CH3), 1.14 (2H, m, CH2),
0.52 (3H, t, J 7.5 Hz, CH3); dC (75 MHz, CDCl3) 167.6, 155.6, 127.9,
127.5, 126.8, 103.3, 92.9, 62.2, 51.5, 50.5, 32.3, 14.7, 9.66, 6.79; m/z
355 (Mþ23).
4.5. Synthesis of 3-ethoxycarbonyl-5-methoxycarbonyl-6-
methyl-2-(thiophen-2-yl)-4-phenyl-1,2,3,4-
tetrahydropyrimidine 12b
To a solution of thiophene (17.39 mmol) in THF, n-BuLi
(17.39 mmol) was added slowly at ꢁ40 ꢀC under nitrogen atmo-
sphere. The temperature of the mixture was held between ꢁ30 ꢀC
and ꢁ20 ꢀC for an hour, and then lowered to ꢁ78 ꢀC. Dark red
colored anion of thiophene 11b was added to solution of the elec-
trophile 10 (4.34 mmol) in anhydrous THF (25 mL) with simulta-
neous addition of ethyl chloroformate (4.34 mmol) at ꢁ5 ꢀC.
Reaction was warmed to room temperature and stirring was con-
tinued to complete the reaction (TLC), after which a saturated
aqueous solution of NH4Cl was introduced. The reaction was
extracted with ethyl acetate (3ꢂ25 mL) and the extract washed
once with brine (100 mL). The extract was dried over anhydrous
Na2SO4 and the solvent removed under reduced pressure. The
corresponding products 12b (68%) and 15 (14%) were isolated after
column chromatography using mixtures of ethyl acetate and hex-
ane as eluent. Compound 12b was obtained as a white solid;
[found: C, 62.07; H, 5.83; N, 7.17. C20H22N2O4S requires C, 62.17; H,
5.69; N, 7.25]; Rf (15% EtOAc/hexane) 0.60; mp: 155e156 ꢀC (DCM);
4.6.3. 3-Ethoxycarbonyl-5-methoxycarbonyl-6-methyl-4-phenyl-2-
propyl-1,2,3,4-tetrahydropyrimidine (12e). Off-white solid; [found:
C, 65.81; H, 7.49; N, 8.01. C19H26N2O4 requires C, 65.89; H, 7.51; N,
8.09]; Rf (15% EtOAC/hexane) 0.6; mp: 120e121 ꢀC (DCM); nmax
(KBr): 3315, 2970, 1702, 1628 cmꢁ1
; dH (400 MHz, CDCl3)
7.17e7.35 (5H, m, ArH), 6.37 (1H, br, NH), 5.37 (1H, br, C2eH),
4.70 (1H, br, C4eH), 4.26 (2H, m, estereCH2), 3.58 (3H, s,
estereCH3), 2.40 (3H, s, CH3), 0.78e1.41 (4H, m, 2ꢂCH2), 0.49
(3H, t, J 6.4 Hz, CH3); dC (75 MHz, CDCl3) 167.7, 155.7, 128.0, 127.6,
126.9, 63.4, 62.3, 51.7, 50.6, 38.4, 29.7, 21.5, 18.4, 14.8, 13.3; m/z
369 (Mþ23).
4.6.4. 3-Ethoxycarbonyl-5-methoxycarbonyl-2-butyl-6-methyl-4-
phenyl-1,2,3,4-tetrahydropyrimidine (12f). White solid; [found: C,
66.47; H, 7.77; N, 7.69. C20H28N2O4 requires C, 66.66; H, 7.77; N,
7.77]; Rf (15% EtOAC/hexane) 0.6; mp: 96e97 ꢀC (DCM); nmax (KBr):
3320, 2966, 1700, 1620 cmꢁ1
; dH (300 MHz, CDCl3) 7.16e7.37 (5H,
m, ArH), 6.37 (1H, s, NH), 5.43 (1H, s, C2eH), 4.72 (1H, s, C4eH),
4.30 (2H, q, J 7.5 Hz, estereCH2), 3.53 (3H, s, estereCH3), 2.40 (3H, s,
CH3), 0.74e1.42 (9H, m, CH3, and 3ꢂCH2), 0.59 (3H, t, J 6.9 Hz, CH3);
dC (75 MHz, CDCl3) 167.7, 155.6, 127.9, 127.5, 126.8, 62.2, 50.5, 36.0,
29.6, 27.1, 21.9, 14.7, 13.6; m/z 383 (Mþ23).
nmax (KBr): 3388, 1666, 1607 cmꢁ1
; dH (300 MHz, CDCl3) 6.89e7.00
(5H, m, ArH), 6.82 (1H, br, NH), 6.43e6.53 (3H, m, ArH), 6.37 (1H, br,
C2eH), 4.99 (1H, br, C4eH), 4.34 (2H, q, J 6.9 Hz, estereCH2), 3.46
(3H, s, estereCH3), 2.51 (3H, s, CH3), 1.29 (3H, t, J 7.2 Hz, CH3); dC
(75 MHz, CDCl3) 155.6, 144.1, 128.1, 127.7, 127.6, 127.4, 127.2, 126.4,
126.1, 125.3, 124.6, 96.3, 68.9, 63.3, 62.5, 62.2, 50.6, 14.7, 14.3; m/z
409 (Mþ23).
4.6.5. 3-Ethoxycarbonyl-5-methoxycarbonyl-6-methyl-2-pentyl-4-
phenyl-1,2,3,4-tetrahydropyrimidine (12g). White solid; [found: C,
67.21; H, 7.97; N, 7.39. C21H30N2O4 requires C, 67.37; H, 8.02; N,
7.48]; Rf (15% EtOAC/hexane) 0.6; mp: 98e99 ꢀC (DCM); nmax (KBr):
4.6. General procedure for the synthesis of pyrimidine
derivatives 12cej
3316, 2976, 1709,1621 cmꢁ1
; dH (300 MHz, CDCl3) 7.16e7.60 (5H, m,
ArH), 6.38 (1H, s, NH), 5.44 (1H, s, C2eH), 4.65 (1H, s, C4eH), 4.27
(2H, q, J 6.9 Hz, estereCH2), 3.53 (1H, s, CH3), 2.40 (3H, s, CH3),
0.78e1.37 (11H, m, CH3, and 4ꢂCH2), 0.72 (3H, t, J 7.2 Hz, CH3); dC
Appropriate Grignard reagent 11cej (17.39 mmol) prepared
using activated magnesium metal and alkyl/aryl/allyl halide in

8180
K. Singh et al. / Tetrahedron 66 (2010) 8175e8180
(75 MHz, CDCl3) 167.7, 155.6, 127.9, 127.5, 126.8, 63.6, 62.1, 50.5,
36.3, 31.0, 24.6, 22.2, 21.3, 14.6, 13.8; m/z 397 (Mþ23).
for financial assistance, and National Single Crystal X-ray Diffrac-
tion Facility, IIT Bombay for X-ray analysis. DA thanks the CSIR,
New Delhi for senior research fellowship.
4.6.6. 3-Ethoxycarbonyl-5-methoxycarbonyl-6-methyl-4-phenyl-2-
undecyl-1,2,3,4-tetrahydropyrimidine (12h). Yellow liquid; [found:
C, 70.58; H, 9.16; N, 6.06. C27H42N2O4 requires C, 70.74; H, 9.17; N,
6.11]; Rf (15% EtOAC/hexane) 0.6; nmax (KBr): 3320, 2986, 1710,
References and notes
1. Hajduk, P. J.; Greer, J. Nat. Rev. Drug Discovery 2007, 6, 211e219.
2. Atwal, K. S.; Swanson, B. N.; Unger, S. E.; Floyd, D. M.; Moreland, S.; Hedberg, A.;
O’Reilly, B. C. J. Med. Chem. 1991, 34, 806e811.
3. Mayer, T. U.; Kapoor, T. M.; Haggarty, S. J.; King, R. W.; Schreiber, S. L.; Mitchison, T.
J. Science 1999, 286, 971e974.
1621 cmꢁ1
; dH (300 MHz, CDCl3) 7.15e7.37 (5H, m, ArH), 6.36 (1H, s,
NH), 5.44 (1H, s, C2eH), 4.71 (1H, s, C4eH), 4.30 (2H, m,
estereCH2), 3.53 (3H, s, estereCH3), 2.40 (3H, s, CH3), 0.69e1.42
(26H, t, J 7.2 Hz, 10ꢂCH2, and 2ꢂCH3); dC (75 MHz, CDCl3) 167.7,
155.6, 127.9, 127.5, 126.8, 63.5, 62.1, 50.5, 31.8, 29.5, 29.4, 29.3, 29.2,
28.8, 25.0, 22.6, 14.6, 14.0; m/z 481 (Mþ23).
4. Haggarty, S. J.; Mayer, T. U.; Miyamoto, D. T.; Fathi, R.; King, R. W.; Mitchison, T.
J.; Schreiber, S. L. Chem. Biol. 2000, 7, 275e286.
5. Maliga, Z.; Kapoor, T. M.; Mitchison, T. J. Chem. Biol. 2002, 9, 989e996.
6. DeBonis, S.; Simorre, J. P.; Crevel, I.; Lebeau, L.; Skoufias, D. A.; Blangy, A.; Ebel,
C.; Gans, P.; Cross, R.; Hackney, D. D.; Wade, R. H.; Kozielski, F. Biochemistry
2003, 42, 338e349.
7. Barrow, J. C.; Nantermet, P. G.; Selnick, H. G.; Glass, K. L.; Rittle, K. E.; Gilbert, K.
F.; Steele, T. G.; Homnick, C. F.; Freidinger, R. M.; Ransom, R. W.; Kling, P.; Reiss,
D.; Broten, T. P.; Schorn, T. W.; Chang, R. S. L.; O’Malley, S.; Olah, T. V.; Ellis, J. D.;
Barrish, A.; Kassahun, K.; Leppert, P.; Nagarathnam, D.; Forray, C. J. Med. Chem.
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Kramer, T.; Niewoehner, U.; Pleiss, U.; Stoltefuss, J.; Graef, E.; Koletzki, D.;
Masantschek, R. N. A.; Reimann, A.; Jaeger, R.; Grob, R.; Beckermann, B.;
Schlemmer, K.-H.; Haebich, D.; Ruebsamen-Waigmann, H. Science 2003, 299,
893e896.
4.6.7. 3-Ethoxycarbonyl-5-methoxycarbonyl-2-allyl-6-methyl-4-
phenyl-1,2,3,4-tetrahydropyrimidine (12i). Yellow solid; [found: C,
66.23; H, 6.98; N, 8.06. C19H24N2O4 requires C, 66.27; H, 6.97; N,
8.13]; Rf (15% EtOAC/hexane) 0.6; mp 95e96 ꢀC (DCM); nmax (KBr):
3310, 2988, 1711, 1640 cmꢁ1
; dH (300 MHz, CDCl3) 7.18e7.35 (5H, m,
ArH), 5.93 (1H, s, C4eH), 5.68 (1H, m, CH), 5.10 (2H, m, CH2), 4.68 (2H,
m, C4eH, and NH), 4.08 (2H, m, estereCH2), 3.61 (3H, s, estereCH3),
2.71 (2H, m, CH2), 2.29 (3H, s, CH3), 1.14 (3H, t, J 7.2 Hz, CH3); dC
(75 MHz, CDCl3) 167.1, 155.0, 153.1, 144.3, 133.6, 127.8, 127.5, 126.6,
119.0, 99.4, 63.6, 61.4, 56.0, 50.5, 37.9, 21.1, 14.3; m/z 367 (Mþ23).
9. Kappe, C. O. Eur. J. Med. Chem. 2000, 35, 1043e1052.
10. Singh, K.; Arora, D.; Singh, K.; Singh, S. Mini-Rev. Med. Chem. 2009, 99, 5e106.
11. Dallinger, D.; Kappe, C. O. Pure Appl. Chem. 2005, 77, 155e161.
12. Singh, K.; Singh, S.; Mahajan, A. J. Org. Chem. 2005, 70, 6114e6117.
13. Singh, K.; Singh, S. Tetrahedron Lett. 2006, 47, 8143e8146.
14. Singh, K.; Singh, K. Tetrahedron 2009, 65, 10395e10399.
15. Singh, K.; Arora, D.; Falkowaski, D.; Liu, Q.; Moreland, R. S. Eur. J. Org. Chem.
2009, 3258e3264.
4.6.8. 3-Ethoxycarbonyl-5-methoxycarbonyl-6-methyl-2,4-diphenyl-
1,2,3,4-tetrahydropyrimidine (12j). White solid; [found: C, 69.23; H,
6.28; N, 7.21. C22H24N2O4 requires C, 69.47; H, 6.31; N, 7.36]; Rf (15%
EtOAC/hexane) 0.6; mp 140e141 ꢀC (DCM); nmax (KBr): 3341,
16. Singh, K.; Arora, D.; Singh, S. Tetrahedron Lett. 2007, 48, 1349e1352.
17. Singh, K.; Singh, S. Tetrahedron 2009, 65, 4106e4112.
1680 cmꢁ1
; dH (300 MHz, CDCl3) 7.18e7.47 (10H, m, ArH), 6.90 (1H,
s, NH), 5.83 (1H, m, C2eH), 4.64 (1H, m, C4eH), 3.86e3.99 (2H, m,
estereCH2), 3.65 (3H, s, estereCH3), 2.55 (3H, s, CH3), 0.93 (3H, t, J
7.2 Hz, CH3); dC (75 MHz, CDCl3) 152.2, 141.8, 128.6, 128.1, 128.0,
127.6, 127.3, 126.8, 125.3, 66.8, 61.4, 55.7, 50.7, 21.3, 14.0; m/z 403
(Mþ23).
18. Stray, S. J.; Bourne, C. R.; Punna, S.; Lewis, W. G.; Finn, M. G.; Zlotnick, A. Z. PNAS
2005, 10, 8138e8143.
19. Kreutzberger, A.; Sellheim, M. J. Heterocycl. Chem. 1985, 22, 721e723.
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Chem. 1997, 5, 437e444.
21. Capdeville, R.; Buchdunger, E.; Zimmermann, J.; Matter, A. Nat. Rev. Drug
Discovery 2002, 1, 493e502.
22. While screening pyrimidine derivatives for their activity against a broad variety
of DNA and RNA viruses (including HIV) in the appropriate cell culture models,
we observed that while none of the dihydropyrimidinone derivatives showed
appreciable activity against any of the investigated viruses at subtoxic con-
centrations, C-2 substituted pyrimidine derivatives 7 and 8 proved markedly
4.7. Cytostatic activity assays
Murine leukemia L1210, murine mammary carcinoma FM3A,
human T-lymphocyte CEM and human cervix carcinoma HeLa cells
were suspended at 300,000e500,000 cells/mL in RPMI-1640 cul-
ture medium supplemented with 10% foetal bovine serum and
cytostatic against MDCK cell cultures (IC50: 0.9
spectively). Compound 7 was more cytotoxic (MCC: 4e10
cell cultures (i.e., human embryonic lung cells, feline Crandell kidney cells)
g/mL) in these cell cultures. Singh, K.; Arora, D.; Balzarini,
m
g/mL and 1.2
mg/mL, re-
m
g/mL) to confluent
than 8 (MCC: ꢄ100
m
2 mM
L-glutamine, and 100 ml of the cell suspensions were added to
J. unpublished results.
100 l of an appropriate dilution of the test compounds in 96-well-
m
23. Matloobi, M.; Kappe, C. O. J. Comb. Chem. 2007, 9, 275e284.
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1957e1960.
microtiter plates. After incubation at 37 ꢀC for two (L1210 and
FM3A) or three (CEM and HeLa) days, the cell number was
determined using a Particle Counter ZI (Coulter, Analis, Ghent,
Belgium). The number of the suspension cells could be counted
directly; the number of the monolayer HeLa cells was counted after
detachment of the cells upon trypsinization. The IC50 was defined
as the compound concentration required for inhibiting cell pro-
liferation by 50%.
26. Gholap, A. R.; Toti, K. S.; Shirazi, F.; Deshpande, M. V.; Srinivasan, K. V. Tetra-
hedron 2008, 64, 10214e10223.
27. Singh, S.; Schober, A.; Gebinoga, M.; Gross, A. G. Tetrahedron Lett. 2009, 50,
1838e1843.
28. Kashima, C.; Shimizu, M.; Katoh, A.; Omote, Y. J. Chem. Soc., Perkin Trans. 1 1983,
1799e1802.
29. Biginelli, P. Gazz. Chim. Ital. 1893, 23, 360e416.
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31. Jones, E.; Moodie, I. M. Org. Synth. 1988, Coll. Vol. 6, 979e980.
32. Hiers, G. S. Org. Synth. 1941, Coll. Vol. 1, 550e552.
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4719e4728.
Acknowledgements
We are thankful to UGC (F. No. 37-188/2009 (SR)) and CSIR 01
(2364)/10/EMR-II, New Delhi and the K.U. Leuven (GOA no. 05/19)
34. Jacquemard, U.; Beneteau, V.; Lefoix, M.; Routeir, S.; Merour, J.-Y.; Coudert, G.
Tetrahedron 2004, 60, 10039e10047.