Regioselective synthesis of 6-substituted-2-amino-5-bromo-4(3H)- pyrimidinones and evaluation of their antiviral activity

Article abstract of DOI:10.1016/j.ejmech.2013.06.036

A series of 2-amino-5-bromo-4(3 H)-pyrimidinone derivatives bearing different substituents at the C-6 position were synthesized using a highly regioselective lithiation-substitution protocol, and the effect of structural variation at the C-6 position on their antiviral activity in cell culture was evaluated. Although some of the derivatives were found to be active against various virus strains, they were effective only close to their toxicity threshold.

Full text of DOI:10.1016/j.ejmech.2013.06.036

European Journal of Medicinal Chemistry 67 (2013) 428e433
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Regioselective synthesis of 6-substituted-2-amino-5-bromo-4(3H)-
pyrimidinones and evaluation of their antiviral activity
a
,
a
b
*
Kamaljit Singh , Kawaljit Singh , Jan Balzarini
a
Department of Chemistry, UGC-Centre of Advance Study-I, Guru Nanak Dev University, Amritsar 143005, India
Rega Institute for Medical Research, Katholieke Universiteit Leuven, 10 Minderbroedersstraat, B-3000 Leuven, Belgium
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 11 April 2013
Received in revised form
A series of 2-amino-5-bromo-4(3H)-pyrimidinone derivatives bearing different substituents at the C-6
position were synthesized using a highly regioselective lithiationesubstitution protocol, and the effect of
structural variation at the C-6 position on their antiviral activity in cell culture was evaluated. Although
some of the derivatives were found to be active against various virus strains, they were effective only
close to their toxicity threshold.
21 May 2013
Accepted 15 June 2013
Available online 5 July 2013
Ó 2013 Elsevier Masson SAS. All rights reserved.
Keywords:
Interferon inducers
Antiviral
Pyrimidinone
Cytotoxicity
Regioselectivity
Lithiation
1. Introduction
ABMP, 6-phenylpyrimidinones does not possess this toxicity and
exhibits enhanced interferon-inducing antiviral potency and ac-
The efficacy of interferons in mediating an antiviral response in
the virus-infected host has been well established [1]. One of the
most important approaches adopted in antiviral therapy is the use
of interferon inducers [2,3]. Agents capable of stimulating inter-
feron production range from viruses and bacterial cell walls to
synthetic compounds or biopolymers [4] along with a small group
of low-molecular-weight compounds (Fig. 1). Some of the com-
pounds which have been reported to induce interferon include N,N-
tivity [12]. Further biological evaluation of an initial lead candidate
in this second-generation pyrimidinone series, 2-amino-5-bromo-
6-phenyl-4(3H)-pyrimidinone 7, revealed an intriguing spectrum of
immunomodulatory activity which may be related to its antiviral
[13] and antitumour activity [14]. In the present study, a highly
regioselective and efficacious protocol to prepare C-6 elaborated 2-
amino-5-bromo-4(3H)-pyrimidinones required for the elucidation
of the structureeactivity relationship (SAR) in this pyrimidinone
series has been reported. In addition, a rather elaborated antiviral
assay of this series of new compounds along with examination of
their cytotoxicity has been carried out.
0
0
0
dioctadecyl-N ,N -bis(2-hydroxyethyl)propanediamine 1 [5], N,N -
dihexadecyl-m-xylylenediamine [6], bis(diethy1amino)fluo-
2
renone (3, tilorone) [7], 1,5-bis[[(diethylamino)ethyl]amino]-9,10-
anthraquinone 4 [8] and nucleobase derived 9-benzyl-8-hydroxy
adenines 5 [9].
Although the compound 2-amino-5-bromo-6-methyl-4(3H)-
pyrimidinone 6 (Fig. 1) has been reported to induce interferon in
rodents and cats when administered orally or intraperitoneally
2. Chemistry
The compound 2-amino-6-methyl-4(3H)-pyrimidinone
8
[10], it has been observed to exhibit toxicity-limiting crystal
was readily synthesized by condensation of the -ketoester, eth-
b
deposition in the renal papillae of rats upon chronic administration
ylacetoacetate with guanidine carbonate in ethanoletoluene
mixture (Scheme 1), obtaining 65% yield following a reported
procedure [13].
[11]. Subsequent studies have demonstrated that the analogue of
In order to append different substituents at the C-6 position of
compound 8, our synthesis scheme was commenced by protecting
the NH group of compound 8 with a tert-butyloxycarbonyl (Boc)
2
*
Corresponding author. Tel.: þ91 183 2258853; fax: þ91 183 2258819x20.
E-mail addresses: kamaljit19in@yahoo.co.in, kamaljitgndu@gmail.com (K. Singh).
0223-5234/$ e see front matter Ó 2013 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2013.06.036

K. Singh et al. / European Journal of Medicinal Chemistry 67 (2013) 428e433
429
O
NH(CH2)15CH3
O
O
H C(H C)
3
2
17
CH CH OH
2 2
N
N
N
N
H C(H C)
3
2
17
CH CH OH
2
2
NH(CH2)15CH3
1
3
N
2
NH2
N
O
N
OH
O
HN
Br
R
N
NH
NH2
OH
N
R
N
CH Ph
2
NH
O
OH
4
R = S-alkyl, OC H9
6 R = CH3
N
4
5
7 R = C H5
6
Fig. 1. Selected examples of low molecular weight interferon inducers.
ꢀ
group using Et
3
N as the base in THF at 40 C to obtain the desired
and subsequent deprotection with TFA produced good yields of
compounds 13h and 13i (Table 1, entries 8 and 9). Evidently, the
formation of 13h and 13i products suggests in situ elimination of
water from the corresponding carbinols 12h and 12i, under the acid
catalyzed conditions used in the deprotection reaction.
The 5-bromo substituted analogues of compound 13aei,
namely, compounds 14aei, were obtained in good to excellent
yields (Table 1, entries 10e18) through the reaction of the respec-
tive compounds 13aei, with bromine in acetic acid at room tem-
perature (Scheme 2).
compound 9 with a yield of 47% (Scheme 2) [15]. A metallatione
alkylation sequence was used for the synthesis of 6-substituted
derivatives of compound 8. In order to fully exploit the synthesis
potential of this procedure, the reaction of the lithium derivative of
compound 8 with several electrophiles was investigated (Scheme
2
). All the compounds exhibited satisfactory characteristic data
(vide experimental).
The anion intermediate 10 was generated in situ by treating
ꢀ
compound 9 with n-BuLi (3.1 equiv) in anhydrous THF at 0 C in an
atmosphere of dry nitrogen gas. The treatment of 10 with an
appropriate electrophile 11 such as ethyl bromide, n-propyl bro-
3. Biology
ꢀ
mide and n-butyl bromide (1.2 equiv, Table 1) at 0 C led to the
substitution at the more nucleophilic C-6 (vinylogous) methyl.
Thus, the formation of the corresponding O/N-alkylated product
The antiviral assays were based on the inhibition of virus-
induced cytopathicity in CRFK [Feline corona virus (FIPV) and Fe-
line herpes virus], HEL [herpes simplex virus (HSV) type-1 (KOS),
HSV-2 (G), vaccinia virus, vesicular stomatitis virus (VSV), and
was not observed. Subsequent quenching with a saturated aqueous
ꢀ
solution of NH
4
Cl at 0 C, afforded 6-substituted derivatives 12aec
-
R
(Scheme 2). These derivatives were then treated with trifluoro-
HSV-1 (TK KOS ACV )], Vero (parainfluenza-3, reovirus-1, Sindbis
virus B4, Coxsackie B4, and Punta Toro virus), HeLa (VSV, Coxsackie
virus B4, and respiratory syncytial virus) and MDCK [(influenza A
(H1N1, H3N2) and B virus)] cell cultures. Confluent cell cultures in
96-well microtiter plates were inoculated with 100 cell culture
inhibitory dose-50 (CCID50) of virus (1 CCID50 is the virus dose to
infect 50% of the cell cultures), along with varying concentrations of
the test compounds. Viral cytopathicity was recorded as soon as it
reached completion in the control virus-infected cell cultures that
were not treated with the test compounds. Table S1 (see Supporting
information) shows the activity and cytotoxicity data of com-
pounds 14aei against various virus strains. The anti-VSV results
observed in HeLa cell cultures (Table S1, entry 5) could not be
observed in HEL cell cultures (Table S1, entry 4).
acetic acid: methylene chloride (2:8 v/v) mixture at room tem-
perature without further purification to obtain compounds 13aec
in good yields (Table 1, entries 1e3).
Similarly, compound 9 was subjected to the above-mentioned
metallation reaction, followed by quenching with 2-bromoethyl
benzene to obtain compound 12d, and subsequent deprotection of
12d produced the desired compound 13d in 70% yield (Table 1).
Likewise, reaction of the anionic intermediate 10 with acetone fol-
lowed by deprotection of the corresponding 12e produced a good
yield of compound 13e. To achieve further diversification at the
C6-position of compound 8, reactions of the anionic intermediate
10 were performed with a few aldehydes. Thus, the reaction
of compound with n-BuLi followed by reaction with 4-
9
chlorobenzaldehyde and 2-trifluoromethylbenzaldehyde and sub-
sequent deprotection of the corresponding intermediates 12f and
Compound 14h (R ¼ 2,4,6-(CH ) C H CH]CH) showed activity
3
3 6 2
against the AD-169 (EC ¼ 11
5
0
m
M) and Davis strains (EC ¼ 8.9
50
mM)
12g with TFA produced good yields of compounds 13f and 13g
of cytomegalovirus in HEL cell cultures, but its activity was close to
its toxicity threshold (Table S1, entry 1). Similarly, compound 14h
also exhibited activity against varicella-zoster virus (VZV) strains
(Table 1, entries 6 and 7). Likewise, reaction of the anionic species 10
with 2,4,6-trimethylbenzaldehyde and 3,4-dimethoxybenzaldehyde
þ
ꢁ
[
TK VZV OKA (EC50 ¼ 2.5
m
M) and TK VZV 07-1 (EC50 ¼ 5.3
mM)] in
HEL cell cultures, which was close to its toxicity threshold (Table S1,
entry 2). In CRFK cell cultures, compound 14h presented activity
against Feline Herpes Virus at a concentration well below its cellular
toxicity threshold (Table S1, entry 3). However, none of the com-
pounds presented any activity against HSV-1, HSV-2, vaccinia virus
and VSV in HEL cell cultures (Table S1, entry 4). Compounds 14b, 14c,
O
O
NH
NH2
EtOH/PhMe
Heat/3h
NH
NH2
EtO
Me
+
H N
2
Me
N
O
8
1
4f and 14i possessing, respectively,
a n-butyl, n-pentyl, 4-
Scheme 1. Synthesis of 2-amino-6-methyl-4(3H)-pyrimidinone.
ClC CH(OH)CH and 3,4-(OCH CH]CH substituents at C-6
6
H
4
2
3 2 6 3
) C H

430
K. Singh et al. / European Journal of Medicinal Chemistry 67 (2013) 428e433
O
O
O
NH
NH2 DMAP/THF
0 oC
(
Boc) O/Et N
NH
NHBoc
NLi
2
3
n-BuLi
THF/0 C
o
Me
N
Me
N
LiHC
N
NBoc
Li
4
8
9
10
+
o
(
i) 11 (E )/0 C
o
(
ii) aq. NH Cl/0 C
4
O
O
N
O
Br
R
Br /AcOH
NH
NH
2
TFA/DCM
RT
NH
RT
N
NH2
R
NH2
N
NHBoc
E
14
13
12
(
(
(
(
(
(
(
a) E = C H5 (b) E = n-C H7
2 3
c) E = n-C H (d) E = -CH CH Ph
4
9
2
2
e) E = -C(CH ) OH
3
2
f) E = 4-ClC H CH(OH)
6
4
g) E = 2-CF C H CH(OH)
3
6 4
h) E = 2,4,6-(CH ) C H CH(OH)
3
3 6 2
i) E = 3,4-(OCH ) C H CH(OH)
3
2 6 3
Scheme 2. Synthesis of C6-substituted 2-amino-6-methyl-4(3H)-pyrimidinone 13 and 5-bromo derivatives 14.
positions showed activity against VSV in HeLa cell cultures with EC50
values of 10, 45, 45 and 45 M, respectively. Although, compound 14c
with an n-pentyl substituent at the C-6 position had presented
M), it was close to its
toxicity threshold (Table S1, entry 5). Furthermore, the anti-VSV ac-
tivity was not observed in HEL cell cultures, in which the toxicity of
the compound was also lower. In Vero cell cultures, none of the
compounds showed any activity against parainfluenza virus,
reovirus-1, Sindbis virus, Coxsackie virus B4 and Punta Toro virus
pyrimidinones in a synthetically useful manner. The obtained de-
rivatives were evaluated for antiviral activity in a series of cell
cultures. Although some of the derivatives were found to be active
against various virus strains in different cell cultures, they were
effective only close to their toxicity threshold.
m
pronounced antiviral activity (EC50 ¼ 2
m
5. Experimental section
5.1. General
(Table S1, entry 6). In MDCK cell cultures, all the compounds were
found to be inactive against influenza virus (Table S1, entry 7).
All liquid reagents were dried/purified using recommended
ꢀ
1
drying agents and/or distilled over 4 A molecular sieves. H NMR
13
4
. Conclusions
(300 MHz) and C NMR (75 MHz) spectra were recorded in DMSO-
and CDCl on a multinuclear Jeol FT-AL-300 instruments with
chemical shifts being reported in parts per million ( ) relative to
d
6
3
A new series of 2-amino-5-bromo-4(3H)-pyrimidinone de-
rivatives bearing different substituents at the C-6 position were
synthesized from the corresponding 2-amino-6-methyl-4(3H)-
d
1
13
internal tetramethylsilane (TMS,
d
0.0, H NMR or CDCl ,
3
d
77.0,
C
NMR). Mass spectra were recorded at The Indian Institute of Inte-
grative Medicine (CSIR), Jammu, India, under electron impact at
7
0 eV on a Bruker Daltonics Esquire 3000 spectrometer. Elemental
Table 1
analysis was performed on FLASH EA 112 (Thermoelectron Corpo-
ration) analyzer at the Department of Chemistry, Guru Nanak Dev
University, Amritsar, India and the results were presented in per-
centage (%). IR was recorded on FTIR Shimadzu 8400 Fourier-
Synthesis of C6-substituted derivatives of 2-amino-4(3H)-pyrimidinone 13 and 2-
amino-5-bromo-4(3H)-pyrimidinone 14.
Entry Electrophile 11
Compound
R
Yield (%)
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
.
.
.
.
.
.
.
.
C
n-C
n-C
BrCH
(CH
4-ClC
2-CF
2,4,6-(CH
2
H
5
Br
13a
13b
13c
13d
13e
13f
n-C
n-C
n-C
n-C
CH
4-ClC
2-CF
2,4,6-(CH
3,4-(OCH
n-C
n-C
n-C
n-C
CH
4-ClC
2-CF
2,4,6-(CH
3,4-(OCH
3
4
5
3
H
H
H
H
7
65
69
62
70
75
58
61
ꢁ1
transform spectrophotometer in the range of 400e4000 cm us-
3
H
7
Br
Br
9
ing KBr. The melting points were determined in open capillaries
and were not corrected. For monitoring the progress of a reaction
and for comparison purpose, thin layer chromatography was per-
formed on pre-coated aluminium sheets (Merck; 60F254, 0.2 mm)
using an appropriate solvent system. The chromatograms were
visualized under UV light. For column chromatography silica gel
(60e120 mesh) was employed and the eluents were ethyl acetate/
hexane mixtures.
4
H
9
11
2
CH
CO
CHO
2
Ph
6
Ph
3
)
2
2
C(CH
3
)
2
OH
6
H
4
6
H
4
CH(OH)CH
2
3
C
6
H
4
CHO
13g
3
C
6
H
4
3
CH(OH)CH
2
3
)
3
C
6
H
2
CHO 13h
CHO 13i
14a
)
3
6
C H
2
CH]CH 71
CH]CH 69
.
3,4,-(OCH
3
)
2
C
6
H
3
3
)
2
6
C H
3
0.
1.
2.
3.
4.
5.
6.
7.
8.
e
e
e
e
e
e
e
e
e
3
H
4
H
5
H
3
H
7
88
82
90
83
92
14b
14c
14d
14e
14f
14g
14h
14i
9
11
6
Ph
5.2. General procedure for the synthesis of tert-butyl (4-methyl-6-
2
C(CH
3
)
2
OH
oxo-1,6-dihydropyrimidin-2-yl) carbamate 9
6
H
4
CH(OH)CH
2
78
73
3
C
6
H
4
3
CH(OH)CH
2
)
3
6
C H
2
CH]CH 87
CH]CH 76
A mixture of compound 8 (3.99 mmol), Boc
2
O (4.79 mmol),
3
)
2
6
C H
3
triethylamine (5.98 mmol), and DMAP (catalytic amount) in THF

K. Singh et al. / European Journal of Medicinal Chemistry 67 (2013) 428e433
431
ꢀ
(
20 mL) was stirred at 40 C for 2 days. The solvent was evaporated
20.1, 25.0, 28.9, 32.2, 99.1, 152.8, 154.9 and 161.3. Anal. Calcd. for
and the residue was purified by column chromatography on silica
gel using ethyl acetate and hexane as the eluent to obtain the pure
compound. The characteristic data of the compound are as follows.
C
9
H
15
N
3
O: C, 59.64; H, 8.34; N, 23.19; Found: C, 59.23; H, 8.11; N,
þ
22.94. MS: m/z 182 (M þ1).
Viscous oil. Rf: 0.4 (Ethyl acetate). Yield: 47%. IR (KBr):
n
max 1090,
1.48 (s,
5.3.4. 2-Amino-6-phenethylpyrimidin-4(3H)-one (13d)
ꢁ1 1
ꢀ
1
1
D
2
C
1
615, 1680, 3320 cm . H NMR (300 MHz, CDCl
3
, 25 C):
d
Brown solid. Rf: 0.4 (methanol:ethyl acetate/20:80). Yield: 70%.
ꢀ
2H, 3ꢂ CH
3
), 2.12 (s, 3H, C6eCH
O exchangeable, 2ꢂ NH). C NMR (75 MHz, CDCl
3
), 5.85 (s, 1H, C5eH), 7.55 (br, 2H,
M.p. 85e87 C (DCM). IR (KBr):
n
max 1255, 1465, 1625, 1715,
13
ꢀ
ꢁ1 1
ꢀ
2
3
, 25 C)
d
22.3,
3450 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
d
2.71 (t, J ¼ 7.2 Hz,
7.5, 84.0, 101.2, 147.4, 153.9, 154.6, and 165.4. Anal. Calcd. for
2H, CH
2
), 2.89 (t, J ¼ 7.2 Hz, 2H, CH
2
), 5.73 (s, 1H, CH), 7.18e7.33 (m,
13
10
H
15
N
3
O
3
: C, 53.32; H, 6.71; N, 18.66; Found: C, 53.21; H, 6.32; N,
5H, ArH), 8.51 (br, 2H, D
2
2
O exchangeable, NH ). C NMR (75 MHz,
þ
ꢀ
8.44. MS: m/z 248 (M þ23).
DMSO-d
6
, 25 C):
d
32.8, 34.1, 102.0, 126.5, 128.5, 140.0, 153.3 and
1
61.5. Anal. Calcd. for C12
H
13
N
3
O: C, 66.96; H, 6.09; N, 19.52; Found:
þ
5
13
.3. General procedure for the synthesis of C6-substituted derivatives
C, 66.75; H, 5.84; N, 19.19. MS: m/z 216 (M þ1).
5.3.5. 2-Amino-6-(2-hydroxy-2-methylpropyl)pyrimidin-4(3H)-
To a suspension of compound 9 (5.0 mmol) in dry THF (50 mL)
one (13e)
under a blanket of dry N
wise at 0 C. After the addition, reaction mixture was stirred at
room temperature for 0.5 h and quenched at 0 C with electrophile
2
, 2.1 N n-BuLi (15.5 mmol) was added drop
Brown solid. Rf: 0.5 (methanol:ethyl acetate/10:90). Yield: 75%.
ꢀ
ꢀ
M.p. 165e167 C (methanol/DCM). IR (KBr):
n
max 1110, 1245, 1345,
ꢀ
ꢁ1 1
ꢀ
1655, 1702, 3253 cm . H NMR (300 MHz, CDCl
3
, 25 C):
d
1.28 (s,
1
1 (7.5 mmol), dissolved in dry THF (10 mL). The reaction was
6H, 2ꢂ CH
3
), 2.59 (s, 2H, CH
exchangeable, NH). C NMR (75 MHz, CDCl
68.4, 103.3, 151.6 and 152.0. Anal. Calcd. for C
2
), 5.67 (s, 1H, CH), 8.99 (br, 1H, D
2
O
13
ꢀ
stirred at room temperature for 0.5 h. After completion of reaction,
a cold saturated aqueous solution of NH
at 0 C. The reaction contents were extracted with ethyl acetate
3
, 25 C):
d
28.0, 44.1,
4
Cl (30 mL) was introduced
H
8 13
N
3
O
2
: C, 52.45; H,
ꢀ
7.15; N, 22.94; Found: C, 52.11; H, 6.91; N, 22.60. MS: m/z 184
þ
(
3 ꢂ 25 mL), treated with brine, washed with water (2 ꢂ 25 mL),
(M þ1).
dried over anhydrous Na SO and concentrated under reduced
2
4
pressure to obtain crude compound 12 (60e75%). This compound
was further treated with TFA:DCM (2:8) at room temperature
for 0.5 h. After completion of reaction, solvent was removed
under reduced pressure and reaction residue was neutralized with
base. The reaction contents were extracted with ethyl acetate
5.3.6. 2-Amino-6-(2-(4-chlorophenyl)-2-hydroxyethyl)pyrimidin-
4(3H)-one (13f)
White solid. Rf: 0.2 (methanol:ethyl acetate/10:90). Yield: 58%.
ꢀ
M.p. 206e207 C (methanol/DCM). IR (KBr):
n
max 1067, 1287, 1344,
ꢁ
1
1
ꢀ
1615, 1709, 3140, 3644 cm
.
H NMR (300 MHz, CDCl
3
, 25 C):
(
3 ꢂ 20 mL), dried over anhydrous Na
2
SO
4
and concentrated under
d
2.56 (dd, J ¼ 3.6 Hz, 1.8 Hz, 2H, CH
2
), 4.94e4.98 (m, 1H, CH), 5.56
reduced pressure. Required compounds 13 were purified by column
chromatography using silica gel-G (230e400 mesh) and mixtures
of ethyl acetate/hexane as the eluent. The characteristic data of the
compounds are as follows.
(s, 1H, CH), 7.29 (d, J ¼ 8.2 Hz, 2H, ArH), 7.40 (d, J ¼ 8.2 Hz, 2H, ArH),
13
7.89 (br, 2H, D
2
O exchangeable, NH
d 42.1, 69.5, 103.1, 127.3, 127.8, 131.4, 143.0, 152.9 and 161.0.
2
). C NMR (75 MHz, DMSO-d
6
,
ꢀ
25 C):
Anal. Calcd. for C12
5
12
H N
3
O
2
Cl: C, 54.25; H, 4.55; N, 15.82; Found: C,
þ
4.10; H, 4.21; N, 15.43. MS: m/z 288 (M þ23).
5
.3.1. 2-Amino-6-propylpyrimidin-4(3H)-one (13a)
Brown solid. Rf: 0.4 (methanol:ethyl acetate/20:80). Yield: 65%.
5.3.7. 2-Amino-6-(2-hydroxy-2-(2-(trifluoromethyl)phenyl)ethyl)
pyrimidin-4(3H)-one (13g)
ꢀ
M.p. 192e195 C (methanol/DCM). IR (KBr):
n
max 1010, 1313, 1635,
ꢁ1
1
ꢀ
1
699, 3353 cm . H NMR (300 MHz, DMSO-d
), 1.62e1.75 (m, 2H, CH
), 4.11 (br, 2H, D O exchangeable, NH
H, D
6
, 25 C):
d
0.99 (t,
Brown solid. Rf: 0.5 (methanol:ethyl acetate/20:80). Yield: 61%.
ꢀ
J ¼ 7.2 Hz, 3H, CH
3
2
), 2.47 (t, J ¼ 7.5 Hz, 2H,
M.p. 186e187 C (methanol/DCM). IR (KBr):
n
max 1142, 1224, 1633,
ꢁ1
1
ꢀ
CH
1
d
2
2
2
), 5.62 (s, 1H, CH), 5.84 (br,
1688, 3332 cm
J ¼ 8.7 Hz, 2H, CH
(br, 1H, D
.
H NMR (300 MHz, CDCl
3
, 25 C):
d
2.59 (d,
13
ꢀ
2
O exchangeable, NH). C NMR (75 MHz, DMSO-d
6
, 25 C):
2
), 5.31 (t, J ¼ 4.8 Hz, 1H, CH), 5.47 (s, 1H, CH), 6.57
12.7, 19.8, 33.7, 101.5, 153.2, 156.2 and 161.0. Anal. Calcd. for
2
O exchangeable, NH), 7.39 (t, J ¼ 7.5 Hz, 1H, ArH), 7.58e
13
C
2
7
H
11
N
3
O: C, 54.89; H, 7.24; N, 27.43; Found: C, 54.55; H, 7.00; N,
7.63 (m, 2H, ArH), 7.88 (d, J ¼ 8.1 Hz, 1H, ArH). C NMR (75 MHz,
þ
ꢀ
7.23. MS: m/z 154 (M þ1).
CDCl
3
, 25 C):
d
44.4, 66.0, 99.8, 123.5, 123.9, 125.7, 126.6, 130.8,
1
42.4 and 154.0. Anal. Calcd. for C13
H
12
F
3
N
3
O
2
: C, 52.18; H, 4.04; N,
þ
5
.3.2. 2-Amino-6-butylpyrimidin-4(3H)-one (13b)
14.04; Found: C, 51.88; H, 3.90; N, 13.76. MS: m/z 322 (M þ23).
White crystalline solid. Rf: 0.6 (methanol:ethyl acetate/20:80).
ꢀ
Yield: 69%. M.p. 205e207 C (methanol/DCM). IR (KBr):
n
max 1112,
, 25 C):
5.3.8. 2-Amino-6-(2,4,6-trimethylstyryl)pyrimidin-4(3H)-one
(13h)
ꢁ1 1
ꢀ
1
330, 1655, 1705, 3220 cm . H NMR (300 MHz, DMSO-d
6
d
0.92 (t, J ¼ 6.9 Hz, 3H, CH
H, CH
), 2.40 (t, J ¼ 7.2 Hz, 2H, CH
O exchangeable, NH
3
), 1.31e1.39 (m, 2H, CH
2
), 1.52e1.59 (m,
Light yellow solid. Rf: 0.7 (methanol:ethyl acetate/10:90). Yield:
ꢀ
2
D
2
2
), 5.62 (s, 1H, CH), 8.25 (br, 2H,
71%. M.p. 158e160 C (methanol/DCM). IR (KBr):
n
max 1030, 1210,
13
ꢀ
ꢁ1 1
ꢀ
2
2
). C NMR (75 MHz, DMSO-d
6
, 25 C):
d
11.7,
1299, 1629, 1710, 3455 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
1
C
2
9.7, 27.1, 30.2, 99.7, 151.6, 155.4 and 159.5. Anal. Calcd. for
d
2.33 (s, 9H, 3ꢂ CH ), 5.76 (s, 1H, CH), 6.29 (d, J ¼ 16.5 Hz, 1H, CH),
3
13
8
H
13
N
3
O: C, 57.46; H, 7.84; N, 25.13; Found: C, 57.32; H, 7.51; N,
6.88 (s, 2H, ArH), 7.68 (d, J ¼ 16.5 Hz, 1H, CH). C NMR (75 MHz,
þ
ꢀ
4.90. MS: m/z 168 (M þ1).
DMSO-d
1
6
, 25 C):
d
20.0, 100.6, 126.2, 128.1, 130.9, 134.4, 135.4,
36.6, 153.8 and 163.2. Anal. Calcd. for C15
H
17
N
3
O: C, 70.56; H, 6.71;
þ
5.3.3. 2-Amino-6-pentylpyrimidin-4(3H)-one (13c)
N, 16.46; Found: C, 70.23; H, 6.51; N, 16.40. MS: m/z 256 (M þ1).
White crystalline solid. Rf: 0.2 (methanol:ethyl acetate/10:90).
ꢀ
Yield: 62%. M.p. 150e152 C (methanol). IR (KBr):
n
max 1250, 1630,
5.3.9. 2-Amino-6-(3,4-dimethoxystyryl)pyrimidin-4(3H)-one (13i)
ꢁ
1 1
ꢀ
1635, 1699, 3244 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
d
0.89
Yellow solid. Rf: 0.4 (methanol:ethyl acetate/10:90). Yield: 69%.
ꢀ
ꢁ
(
t, J ¼ 6.9 Hz, 3H, CH
CH
), 2.40 (t, J ¼ 7.2 Hz, 2H, CH
exchangeable, NH
3
), 1.28e1.33 (m, 4H, 2ꢂ CH
2
), 1.53e1.65 (m, 2H,
M.p. 220e222 C (methanol). IR (KBr):
1315, 3467 cm . H NMR (300 MHz, CDCl
OCH ), 3.92 (s, 3H, OCH
n
max 1122, 1267, 1356, 1678,
1
1
ꢀ
2
2
), 5.59 (s, 1H, CH), 7.71 (br, 2H, D
2
O
3
, 25 C):
d
3.90 (s, 3H,
13
ꢀ
2
). C NMR (75 MHz, DMSO-d
6
, 25 C):
d
12.0,
3
3
), 5.79 (s, 1H, CH), 6.62 (d, J ¼ 16.2 Hz, 1H,

432
K. Singh et al. / European Journal of Medicinal Chemistry 67 (2013) 428e433
CH), 6.90 (d, J ¼ 8.1 Hz, 1H, ArH), 7.11 (s, 1H, ArH), 7.13 (d, J ¼ 8.7 Hz,
H, ArH), 7.63 (d, J ¼ 16.2 Hz, 1H, CH), 8.07 (br, 1H, D
exchangeable, OH), 6.75 (br, 2H, D
2
O exchangeable, NH
2
), 11.35 (br,
13
ꢀ
1
2
O
55.5,
1H, D
2
O exchangeable, NH). C NMR (75 MHz, DMSO-d
6
, 25 C):
13
ꢀ
exchangeable, NH). C NMR (75 MHz, DMSO-d
6
, 25 C):
d
d
29.4, 45.4, 100.9, 151.4, 153.2 and 157.4. Anal. Calcd. for
5
5.6, 100.9, 109.5, 111.8, 121.8, 128.2, 136.1, 149.0, 150.2, 154.3 and
C
8
H
12BrN
3
O
2
: C, 36.66; H, 4.61; N, 16.03; Found: C, 36.45; H, 4.33;
þ
162.3. Anal. Calcd. for C14
H
15
N
3
O
3
: C, 61.53; H, 5.53; N, 15.38;
N, 15.78. MS: m/z 262 and 264 (M þ1).
þ
Found: C, 61.20; H, 5.32; N, 15.10. MS: m/z 296 (M þ23).
5.4.6. 2-Amino-5-bromo-6-(2-(4-chlorophenyl)-2-hydroxyethyl)
5.4. General procedure for the synthesis of C5-bromo substituted
pyrimidin-4(3H)-one (14f)
derivatives 14
Light yellowish solid. Rf: 0.3 (methanol:ethyl acetate/10:90).
ꢀ
Yield: 78%. M.p. 255e257 C (methanol). IR (KBr):
n
max 598, 710,
ꢁ1 1
To a suspension of compound 13 (1.0 equiv) in glacial acetic acid
1066, 1343, 1685, 1729, 3183 cm . H NMR (300 MHz, DMSO-d
6
,
ꢀ
(
10 mL), bromine (1.1 equiv) was added drop wise and reaction
25 C):
d
2.78e2.93 (m, 2H, CH
2
), 3.87 (br, 1H, D
2
O exchangeable,
mixture was allowed to stir at room temperature for 0.5 h. After
completion of reaction (TLC), solvent was removed under reduced
pressure. Solid residue was washed with water to obtain required
compound. The characteristic data of the compounds are as follows.
OH), 5.43e5.44 (m, 1H, CH), 6.72 (br, 2H, D
7.35e7.52 (m, 4H, ArH). C NMR (75 MHz, DMSO-d
67.5, 103.7, 126.3,127.8, 132.4, 144.0, 156.9 and 161.9. Anal. Calcd. for
2
O exchangeable, NH
2
),
13
ꢀ
6
, 25 C):
d
40.1,
C
12
H
11BrClN
3
O
2
: C, 41.83; H, 3.22; N, 12.19; Found: C, 41.54; H, 3.11;
þ
N, 11.90. MS: m/z 344 and 346 (M þ1).
5.4.1. 2-Amino-5-bromo-6-propylpyrimidin-4(3H)-one (14a)
Light brown solid. Rf: 0.6 (methanol:ethyl acetate/20:80). Yield:
5.4.7. 2-Amino-5-bromo-6-(2-hydroxy-2-(2-(trifluoromethyl)
phenyl)ethyl)pyrimidin-4(3H)-one (14g)
ꢀ
8
1
2
8%. M.p. 200e202 C (methanol/DCM). IR (KBr):
212, 1373, 1695, 1745, 3166 cm . H NMR (300 MHz, DMSO-d
n
max 550, 1120,
ꢁ
1 1
6
,
White solid. Rf: 0.7 (methanol:ethyl acetate/20:80). Yield: 73%.
ꢀ
ꢀ
5 C):
d
0.93 (t, J ¼ 7.2 Hz, 3H, CH
3
), 1.54e1.66 (m, 2H, CH
2
), 2.54 (t,
). C NMR
M.p. 177e179 C (methanol/DCM). IR (KBr):
n
max 615, 1210, 1342,
1
3
ꢁ1 1
ꢀ
J ¼ 7.2 Hz, 2H, CH
2
), 7.29 (br, 2H, D
2
O exchangeable, NH
2
1513, 1735, 3443 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
d
2.75
ꢀ
(
75 MHz, DMSO-d
6
, 25 C):
10BrN
d
13.3, 20.1, 35.4, 98.9, 152.0 and 157.3.
(dd, J ¼ 13.8 Hz, 9.3 Hz, 1H, CH), 2.96 (dd, J ¼ 13.8 Hz, 9.3 Hz, 1H,
CH), 3.90 (br,1H, D
O exchangeable, OH), 5.40 (d, J ¼ 7.2 Hz,1H, CH),
6.76 (br, 2H, D O exchangeable, NH
), 7.48 (t, J ¼ 7.2 Hz, 1H, ArH),
.64e7.74 (m, 2H, ArH), 7.89 (d, J ¼ 7.5 Hz, 1H, ArH). C NMR
Anal. Calcd. For C
6.02; H, 4.22; N, 17.90. MS: m/z 232 and 234 (M þ1).
7
H
3
O: C, 36.23; H, 4.34; N, 18.11; Found: C,
2
þ
3
5
8
1
2
1
2
2
13
7
ꢀ
.4.2. 2-Amino-5-bromo-6-butylpyrimidin-4(3H)-one (14b)
Light brown solid. Rf: 0.6 (methanol:ethyl acetate/10:90). Yield:
(75 MHz, DMSO-d
6
, 25 C):
d
39.7, 66.9, 99.7, 123.6, 124.5, 126.5,
3 3 2
11BrF N O :
130.0,134.6,154.0,155.6 and 162.4. Anal. Calcd. for C13
H
ꢀ
2%. M.p. 210e212 C (methanol/DCM). IR (KBr):
n
max 523, 1016,
C, 41.29; H, 2.93; N, 11.11; Found: C, 40.99; H, 2.56; N, 10.87. MS: m/z
ꢁ1
1
þ
249, 1346, 1679, 1721, 3264 cm . H NMR (300 MHz, DMSO-d
6
,
378 and 380 (M þ1).
ꢀ
5 C):
.59 (m, 2H, CH
exchangeable, NH
75 MHz, DMSO-d
d
0.89 (t, J ¼ 7.2 Hz, 3H, CH
3
), 1.26e1.38 (m, 2H, CH
2
), 1.49e
2
), 2.50 (t, J ¼ 7.2 Hz, 2H, CH
2
), 6.62 (br, 2H, D
2
O
5.4.8. 2-Amino-5-bromo-6-(2,4,6-trimethylstyryl)pyrimidin-
4(3H)-one (14h)
13
2
), 11.18 (br, 1H, D
2
O exchangeable, NH). C NMR
14.1, 22.2, 29.2, 34.0, 99.2, 152.6 and
ꢀ
(
1
6
, 25 C):
d
Light yellow solid. Rf: 0.2 (ethyl acetate). Yield: 87%. M.p. 178e
ꢀ
58.0. Anal. Calcd. for C
Found: C, 38.82; H, 4.70; N, 16.90. MS: m/z 246 and 247 (M þ1).
8
H
12BrN
3
O: C, 39.04; H, 4.91; N, 17.07;
180 C (methanol/DCM). IR (KBr):
n
max 611, 1030, 1262, 1364, 1653,
þ
ꢁ1 1
ꢀ
1679, 3433 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
d
2.23 (s, 3H,
CH
3
), 2.31 (s, 6H, 2ꢂ CH
3
), 6.85e6.93 (m, 3H, CH & ArH), 7.76 (d,
13
ꢀ
5
.4.3. 2-Amino-5-bromo-6-pentylpyrimidin-4(3H)-one (14c)
White solid. Rf: 0.5 (methanol:ethyl acetate/10:90). Yield: 90%.
J ¼ 16.2 Hz,1H, CH). C NMR (75 MHz, DMSO-d
6
, 25 C):
d
20.7, 21.0,
98.3,128.2,129.2,131.8,136.2,137.3,153.3 and 158.9. Anal. Calcd. for
ꢀ
M.p. 195e197 C (methanol). IR (KBr):
n
max 610, 1120, 1255, 1344,
C
15
H
16BrN
3
O: C, 53.91; H, 4.83; N,12.57; Found: C, 53.67; H, 4.45; N,
ꢁ
1 1
ꢀ
þ
1
(
677, 1712, 3453 cm . H NMR (300 MHz, DMSO-d
t, J ¼ 6.0 Hz, 3H, CH ),1.26e1.30 (m, 4H, 2ꢂ CH
CH ), 6.81 (br, 1H, D O exchangeable, NH), 11.05 (br, 2H, D
exchangeable, NH
1.8, 26.6, 30.8, 35.9, 97.3, 153.6 and 158.5. Anal. Calcd. for
6
, 25 C):
d
0.85
12.19. MS: m/z 334 and 336 (M þ1).
3
2
), 2.04e2.15 (m, 2H,
2
2
2
O
13.8,
5.4.9. 2-Amino-5-bromo-6-(3,4-dimethoxystyryl)pyrimidin-4(3H)-
one (14i)
13
ꢀ
2
). C NMR (75 MHz, DMSO-d
6
, 25 C):
d
2
C
1
Yellow solid. Rf: 0.7 (methanol:ethyl acetate/10:90). Yield: 76%.
ꢀ
9
H
14BrN
3
O: C, 41.55; H, 5.42; N, 16.15; Found: C, 41.34; H, 5.33; N,
M.p. 211e213 C (methanol/DCM). IR (KBr):
n
max 613, 1120, 1255,
þ
ꢁ1 1
ꢀ
5.90. MS: m/z 260 and 262 (M þ1).
1353, 1655, 1711, 3329 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
d
3.86 (s, 3H, OCH
3
), 3.90 (s, 3H, OCH
3
), 5.88 (d, J ¼ 11.4 Hz, 1H, CH),
5
.4.4. 2-Amino-5-bromo-6-phenethylpyrimidin-4(3H)-one (14d)
White solid. Rf: 0.7 (methanol:ethyl acetate/20:80). Yield: 83%.
6.18 (d, J ¼ 11.4 Hz, 1H, CH), 6.77 (br, 2H, D
2
O exchangeable, NH ),
2
13
7.11 (d, J ¼ 8.1 Hz, 2H, ArH), 8.06 (s, 1H, ArH). C NMR (75 MHz,
ꢀ
ꢀ
M.p. 220e222 C (methanol/DCM). IR (KBr):
n
max 615, 1130, 1278,
DMSO-d
6
, 25 C):
d
51.5, 56.0, 56.2, 104.5, 121.8, 128.2, 129.2, 136.2,
ꢁ1 1
ꢀ
1
d
369, 1699, 1755, 3283 cm . H NMR (300 MHz, DMSO-d
), 6.71 (br, 2H, D
), 7.21e7.31 (m, 5H, ArH), 11.25 (br, 1H, D O exchangeable, NH).
32.8,126.0, 128.2, 128.4, 141.0
6
, 25 C):
3 3
148.6,151.2,155.3 and 164.7. Anal. Calcd. for C14H14BrN O : C, 47.74;
2.82 (d, J ¼ 8.4 Hz, 4H, 2ꢂ CH
2
2
O exchangeable,
H, 4.01; N, 11.93; Found: C, 47.54; H, 3.91; N, 11.70. MS: m/z 350 and
ꢁ
NH
2
2
352 (M ꢁ1).
13
ꢀ
C NMR (75 MHz, DMSO-d
6
, 25 C):
d
and 154.2. Anal. Calcd. for C12
Found: C, 48.78; H, 3.90; N, 14.10. MS: m/z 316 and 318 (M þ23).
H
12BrN
3
O: C, 49.00; H, 4.11; N, 14.29;
6. Antiviral activity assays
þ
The compounds were evaluated against the following viruses:
herpes simplex virus type 1 (HSV-1) strain KOS, thymidine kinase-
5
4
.4.5. 2-Amino-5-bromo-6-(2-hydroxy-2-methylpropyl)pyrimidin-
(3H)-one (14e)
ꢁ
r
deficient (TK ) HSV-1 KOS strain resistant to ACV (ACV ), herpes
simplex virus type 2 (HSV-2) strains Lyons and G, cytomegalovirus
(CMV) strains AD-169 and Davis, varicella-zoster virus (VZV) strains
OKA and 07-1, vaccinia virus Lederle strain, respiratory syncytial
virus (RSV) strain Long, vesicular stomatitis virus (VSV), Coxsackie
White solid. Rf: 0.7 (methanol:ethyl acetate/10:90). Yield: 92%.
ꢀ
M.p. 180e182 C (methanol/DCM). IR (KBr):
1
n
max 597, 1123, 1345,
ꢁ1 1
ꢀ
395, 1695, 1725, 3353 cm . H NMR (300 MHz, DMSO-d
6
, 25 C):
d
1.14 (s, 6H, 2ꢂ CH ), 2.70 (s, 2H, CH
3
2 2
), 5.29 (br, 1H, D O

K. Singh et al. / European Journal of Medicinal Chemistry 67 (2013) 428e433
433
B4, Parainfluenza 3, Influenza virus A (subtypes H1N1, H3N2),
influenza virus B, Reovirus-1, Sindbis and Punta Toro virus. The
antiviral assays were based on inhibition of virus-induced cytopa-
thicity or plaque formation in human embryonic lung (HEL) fibro-
blasts, African green monkey cells (Vero), human cervix carcinoma
epithelial cells (HeLa) or MadineDarby canine kidney cells (MDCK).
Confluent cell cultures in microtiter 96-well plates were inoculated
with 100 CCID50 of virus (1 CCID50 being the virus dose to infect 50%
of the cell cultures) or 100 or 20 plaque forming units (PFU) (for
CMV or VZV, respectively) in the presence of varying concentra-
tions of the test compounds. Viral cytopathicity or plaque forma-
tion was recorded as soon as it reached completion in the control
virus-infected cell cultures that were not treated with the test
compounds. Antiviral activity was expressed as the EC50 or com-
pound concentration required to reduce virus-induced cytopatho-
genicity or viral plaque formation by 50%.
[2] D.A. Stringfellow (Ed.), Interferon and Interferon Inducers, Marcel Dekker,
New York, 1980.
[3] W. Wierenga, in: H.-J. Hess (Ed.), Annual Reports in Medicinal Chemistry, vol.
17, Academic Press, New York, 1981. (Chapter 16), and references therein.
[4] I. Musmuca, S. Simeoni, A. Caroli, R. Ragno, Small molecule interferon in-
ducers. Toward the comprehension of the molecular determinants through
ligand-based approaches, J. Chem. Inf. Model. 49 (2009) 1777e1786.
[
5] (a) B.G. Gatmaitan, E.D. Stanley, G.G. Jackson, The limited effect of nasal,
interferon induced by rhinovirus and a topical chemical inducer on the course
of, infection, J. Infect. Dis. 127 (1973) 401e407;
0 0
(b) W.W. Hoffman, J.J. Korst, J.F. Niblack, T.H. Cronin, N,N-Dioctadecyl-N ,N -
bis(2-hydroxyethyl) propanediamine:, Antiviral activity and interferon stim-
ulation in mice, Antimicrob. Agents Chemother. 3 (1973) 498e502.
[
6] R.G. Douglas Jr., R.H. Waldman, R.F. Bette, R. Ganguly, Lack of effect of an,
interferon inducer, N,N-dihexadecyl-m-xylylenediamine, on rhinovirus chal-
lenge in, humans, Antimicrob. Agents Chemother. 15 (1979) 269e272.
[7] (a) R.F. Krueger, G.D. Mayer, Tilorone hydrochloride: an orally active antiviral,
agent, Science 169 (1970) 1213e1214;
(b) E.R. Andrews, R.W. Fleming, J.M. Grisar, J.C. Kihm, D.L. Wenstrup,
G.D. Mayer, Bis basic-substituted polycyclic, aromatic compounds. New class
of antiviral agents. 2. Tilorone and related bis-basic, ethers of fluorenone,
fluorenol, and fluorene, J. Med. Chem. 17 (1974) 882e886.
[
8] D.A. Stringfellow, S.D. Weed, G.E. Underwood, Antiviral and interferon-
inducing, properties of 1,5-diamino anthraquinones, Antimicrob. Agents
Chemother. 15 (1979) 111e118.
Acknowledgements
We thank CSIR [01/(2364)/10/EMR-II], UGC [F. No. 37-188/
009(SR)] and SAP (DRS-1) UGC, New Delhi for providing financial
assistance and KU Leuven for financial support of the antiviral ex-
periments (GOA 10/14). We thank Anita Camps, Steven Carmans,
Lies Van der Heurck, Leentje Persoons and Frieda De Meyer for
excellent technical assistance.
[9] K. Hirota, K. Kazaoka, I. Niimoto, H. Kumihara, H. Sajiki, Y. Isobe, H. Takaku,
M. Tobe, H. Ogita, T. Ogino, S. Ichii, A. Kurimoto, H. Kawakami, Discovery of 8-
hydroxyadenines as a novel type of interferon inducer, J. Med. Chem. 45
2
(2002) 5419e5422.
[10] F.R. Nichol, S.D. Weed, G.E. Underwood, Stimulation of murine interferon by a,
substituted pyrimidine, Antimicrob. Agents Chemother. 9 (1976) 433e439.
[
[
11] E.R. Larsen, R.D. Hamilton, J.E. Gray, J.J. Clark, in: J.D. Nelson, C. Grassi (Eds.),
Current Chemotherapy and Infectious Diseases, American Society for Micro-
biology, Washington, DC, 1980, p. 1413.
12] W. Wierenga, H.I. Skulnick, D.A. Stringfellow, S.D. Weed, H.E. Renis,
E.E. Eidson, 5-Substituted 2-amino-6-phenyl-4(3H)-pyrimidinones. Antiviral
and, interferon-inducing agent, J. Med. Chem. 23 (1980) 237e239.
[13] H.I. Skulnick, S.D. Weed, E.E. Eidson, H.E. Renis, W. Wierenga,
D.A. Stringfellow, Pyrimidinones. 1. 2-Amino-5-halo-6-aryl-4(3H)-pyr-
imidinones., interferon-inducing antiviral agent, J. Med. Chem. 28 (1985)
Appendix A. Supporting information
Supplementary data related to this article can be found at
http://dx.doi.org/10.1016/j.ejmech.2013.06.036.
1
864e1869. and, references therein.
References
[
14] (a) D.A. Stringfellow, 6-Arylpyrimidinols: interferon inducers-immuno
modulators-, antiviral and antineoplastic agents, Prog. Cancer Res. Ther. 16
(1981) 215e228;
[1] (a) E.De. Clercq, A 40-year journey in search of selective antiviral chemo-
therapy, Annu. Rev. Pharmacol. Toxicol. 51 (2011) 1e24;
(b) L. Milas, E.M. Hersch, D.A. Stringfellow, N. Hunter, Studies on the antitumor,
activities of pyrimidinone-interferon inducers. I. Effect against artificial and,
spontaneous lung metastases of murine tumors, J. Natl. Cancer Inst. 68 (1982)
139e145.
(
b) D.S. Silin, O.V. Lyubomska, F.I. Ershov, V.M. Frolov, G.A. Kutsuna, Synthetic
and natural immunomodulators acting as interferon inducers, Curr. Pharm.
Des. 15 (2009) 1238e1247;
(
“
c) E. De Clercq, Interferon and its inducers. A never ending story: “Old” and
New” data in a new perspective, J. Infect. Dis. 194 (2006) S19eS26;
d) S. Baron, F. Dianzani, The interferons: a biological system with therapeutic
potential in viral infections, Antivir. Res. 24 (1994) 97e110.
[15] U. Hahn, J.J. González, E. Huerta, M. Segura, J.-F. Eckert, F. Cardinali,
J. Mendoza, J.-F. Nierengarten, A highly directional fourfold hydrogen-bonding
motif for, supramolecular structures through self-assembly of full-
erodendrimers, Chem. Eur. J. 11 (2005) 6666e6672.
(