Synthesis and pharmacological evaluation of novel 1- and 8-substituted-3-furfuryl xanthines as adenosine receptor antagonists

Article abstract of DOI:10.1016/j.bmc.2009.07.034

The synthesis of an important set of 3-furfurylxanthine derivatives is described. Binding affinities were determined for rat A₁ and human A_2A, A_2B and A₃ receptors. Several of the 3-furfuryl-7-methylxanthine der

Full text of DOI:10.1016/j.bmc.2009.07.034

Bioorganic & Medicinal Chemistry 17 (2009) 6755–6760
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and pharmacological evaluation of novel 1- and 8-substituted-
3-furfuryl xanthines as adenosine receptor antagonists
a,c,
a,c
María Carmen Balo a, José Brea b,c, Olga Caamaño a,c, , Franco Fernández
, Xerardo García-Mera
,
*
*
Carmen López a,c, María Isabel Loza b,c, María Isabel Nieto d, José Enrique Rodríguez-Borges e
a Departamento de Química Orgánica, Facultade de Farmacia, Universidade de Santiago de Compostela, E-15782 Santiago de Compostela, Spain
b Departamento de Farmacología, Facultade de Farmacia, Universidade de Santiago de Compostela, E-15782 Santiago de Compostela, Spain
c Instituto de Farmacia Industrial, Facultade de Farmacia, Universidade de Santiago de Compostela, E-15782 Santiago de Compostela, Spain
d Departamento de Química Fundamental, Facultade de Química, Universidade de A Coruña, Campus da Zapateira, E-15071, A Coruña, Spain
e CIQ—Departamento de Química, Facultade de Ciencias, Universidade de Porto, Rua do Campo Alegre, 687, 4169-007 Porto, Portugal
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis of an important set of 3-furfurylxanthine derivatives is described. Binding affinities were
determined for rat A1 and human A2A, A2B and A3 receptors. Several of the 3-furfuryl-7-methylxanthine
derivatives showed moderate-to-high affinity at human A2B receptors, the most active compound (10d)
having a Ki of 7.4 nM for hA2B receptors, with selectivities over rA1 and hA2A receptors up to 14-fold and
11-fold, respectively. Affinities for hA3 receptors were very low for all members of the set.
Ó 2009 Elsevier Ltd. All rights reserved.
Received 28 April 2009
Revised 10 July 2009
Accepted 16 July 2009
Available online 23 July 2009
Keywords:
3-Furfuryl xanthines
Synthesis
Adenosine receptor antagonist
A2B, A2A and A3 Binding affinities
1. Introduction
(AMP) cause bronchoconstriction in atopic and asthmatic patients,
but not in normal control subjects. Furthermore, dipyridamole,
Adenosine modulates many important physiological functions,
that affect the cardiovascular, renal, immune and central nervous
systems. In recent years the search for adenosine analogues that
show agonistic or antagonistic properties against adenosine recep-
tors has intensified. These receptors belong to a superfamily of rho-
dopsin-like G protein-coupled receptors (GPCRs), of which four
subtypes are known and designated as A1, A2A, A2B and A3. The
receptors modulate the activity of adenylate cyclase either by
stimulation (A2A and A2B) or inhibition (A1 and A3) of the activity.1
All of the subtypes have been cloned, and each receptor can there-
fore be studied individually by use of recombinant systems.
In the past decade a large body of experimental evidence has
been obtained that supports the idea that adenosine plays an
important role in allergic asthma.2–5 Patients with asthma and
bronchitis show higher concentrations of adenosine in bronchoal-
veolar fluid than control subjects, suggesting that adenosine may
be a marker of pulmonary inflammation. In in vivo assays, both in-
haled adenosine and its precursor 5-adenosine monophosphate
which is an inhibitor of the cellular adenosine reuptake, increases
bronchospasms induced by adenosine in asthmatic patients, an ef-
fect that may be inhibited by theophylline, an antagonist of aden-
osine receptors.6
The presence of A2B receptors has been demonstrated in bron-
choalveolar mastocytes, a finding that supports the hypothesis that
adenosine participates in the physiopathology of asthma through
activation of these receptors.4,7 As a result A2B receptor antagonists
can be proposed as potential antiasthmatic drugs.
Several groups have designed and tested many xanthine deriv-
atives with the aim of discovering new, potent and A2B-selective li-
gands.8–11 Good initial results have been obtained by Jacobson and
coworkers in the series of 8-(4-substitutedphenyl)xanthines, with
terms such as XAC (1),12 a potent adenosine receptors (AR) antag-
onist, and MRS 1754 (2), that was shown to be selective for human
A2B (hA2B) AR versus hA1, hA2A, and hA3 subtypes of AR.13 Excellent
results in terms of both hA2B AR affinity and selectivity have also
been obtained by another group for the xanthine derivative 3.14
Even though a number of A2B AR antagonists have been re-
ported, only a few have shown high affinity and selectivity for
the A2B AR relative to the A1, A2A, and A3 subtypes of AR. In a first
foray into this field, we studied a short series of new [1,2,4]triazol-
o[1,5-c]quinazoline15 analogues of triazoloquinazoline 4, a strong
* Corresponding authors. Tel.: +34 981563100x15047; fax: +34 981594912
(O.C.); tel.: +34 981563100x14942; fax: +34 981594912 (F.F.).
E-mail addresses: molga.caamano@usc.es (O. Caamaño), franco.fernandez@
usc.es (F. Fernández).
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.07.034

6756
M. C. Balo et al. / Bioorg. Med. Chem. 17 (2009) 6755–6760
antagonist for A2A and A2B AR,4 but unfortunately our compounds
showed poor activities. More promising results were found in a
complete study over the design, synthesis, structure–activity rela-
tionships (SAR) and structure–selectivity relationships (SSR) of a
large series of 9-deaza- and 9-OH-9-deazaxanthines.16 Recently,
we have reported a series of new 1-alkyl-8-substituted-3-(3-
methoxypropyl)xanthines,17 bearing the same 8-(N-substituted-
phenoxyacetamido) moiety proposed by Jacobson,18 and a series
of 1,3-dialkyl-8-(N-substituted-benzyloxycarbonylamino)-9-deaz-
axanthines,19 bearing an isosteric group of the Jacobson motif. In
both series, a number of compounds are endowed with nanomolar
affinities for hA2B, hA1 and/or hA2A AR subtypes.
O
N
O
H
N
i
ii
NH2
N
NH2
H
NH2
O
O
O
O
5
iii
O
N
O
N
O
N
R1
NO
R1
NH2
R1
iv
N
N
v
N
NH2
O
NH2
O
NH2
O
O
O
O
8a-8l
6a-6l
7a-7l
R1 = Me, Et, n-Pr, i-Bu, n-Pen, c-PrMe, prop-2-ynyl, allyl, 2-MeOEt, 2-EtOEt, 2-(MeS)Et, 2-(EtS)Et
O
H
vi
R1
N
N
O
OCH2COR
O
H
N
R1
O
N
N
R1
N
N
N
N
R2
R3
R2
vii
N
N
N
O
O
1, XAC: R1 = R3 = CH2CH2CH3; R = NH(CH2)2NH2
2
, MRS 1754: R1 = R3 = CH2CH2CH3; R = NHC6H4-4-CN
O
O
10a-10o
9a-9aa
Scheme 1. Reagents and conditions: (i) KOCN, H2SO4; (ii) NCCH2CO2H; (iii) (a)
Method A: RX, NaOH, EtOH; (b) Method B: (NH4)2SO4, HMDS, I2, RX, to, Na2S2O3,
H2O; (iv) NaNO2, AcOH; (v) Na2S2O4, NH4OH; (vi) a) R2CO2H, DIC, MeOH, rt, 0.5 h;
(b) 2.5 N NaOH, MeOH, reflux, 10 min–1 h; (vii) CH3I, NaH, DMF, 60 °C, 2 h.
O
O
N
N
CF3
H
N
N
Cl
N
N
N
N
ated derivatives 10(a–o) were obtained by methylation of 9(a–g),
9(l,m), 9(p–t) and 9y with excess methyl iodide in DMF in the pres-
ence of NaH.
N
O
NH2
4, CGS 15943
N
3
Oxidation of the xanthines 9(r–aa) (Scheme 2) to the corre-
sponding sulfoxides 11(a–c) and sulfone 12 was achieved by fol-
lowing previously reported literature procedures.17,19,25
The study reported here was carried out as part of a line of
investigation directed at finding a potentially selective, high affin-
ity A2B-AR antagonist through the preparation of a series of xanth-
ines bearing appropriate and/or more simple substituents in
several positions of the xanthine nucleus, while exploring their
SAR and SSR. The new adenosine analogues of xanthine type pres-
ent structural variations at position 1 (alkyl or functionalized alkyl
substituents), position 7 (unsubstituted or methyl substituent) and
position 8 (aryl or heteroaryl substituents) of the xanthine nucleus,
with a furfuryl substituent at position 3 (Schemes 1 and 2). The
rationale behind this election was the strong adenosine antagonist
activity reported for xanthines such as 1–3 combined with earlier
reports that 3-furfurylxanthines have shown interesting bronchod-
ilating and vasodilatadors effects,20 as well as the presence of the
2-furyl motif at the triazoloquinazoline CGS 15943 (4).
3. Test results and discussion
The affinity (pKi or displacement percentage) values of the 3-
furfurylxanthine derivatives 9(a–aa), 10(a–o), 11(a–c) and 12, at
cloned human adenosine receptors expressed in HeLa cells (hA2A
and hA3) and HEK-293 cells (hA2B) and at rat A1 receptors in mem-
branes from rat cortex,26 are given in Tables 1 and 2. The radioli-
gand [3H]DPCPX was used for competition binding assays on A1
and A2B receptors whereas [3H]ZM241385 was used for A2A and
[3H]NECA for A3 receptors.15,17 The affinity values of compounds
that did not fully displace specific radioligand binding at 1 lM
are given only in terms of displacement percentage. The biological
methods employed are fully described into the Supplementary
data including a representative binding curve for compound 10d
at hA2B receptors.
2. Chemistry
The 7-unsubstituted xanthine derivatives (Table 1) compounds
showed moderate-to-low affinity for hA2B and rA1 receptors, a les-
ser affinity for hA2A receptors, and an almost complete lack of affin-
ity for hA3 receptors. Results shown in Table 1 enable certain
trends to be deduced concerning the SAR and SSR for this group
of xanthines. For example, in as much as these compounds showed
some level of affinity for the receptors in question, an increase in
the size of the 1-alkyl chain from two to three or more carbon
atoms (compounds 9(f–m)) yielded analogues with decreased
A2B and A2A receptors binding affinity relative to the 1-methyl or
1-ethyl analogues 9(a–d), while most of them retained or even in-
creased their affinity for A1 receptors, giving rise to members with
the selectivity A2B/A1 reversed (9(f–h,j–l)). However, the presence
of an unsaturation in the 1-alkyl chain, see compounds 9(n–p), or
the replacement of a methylene group by a sulfur atom in that al-
kyl chain, see compounds 9(w–aa), lead to an improvement of the
The target compounds 9(a–aa) and 10(a–o) were synthesized
as illustrated in Scheme 1. The 5,6-diamino-1-furfuryl-3-substi-
tuted uracils 8 were synthesized according to previously described
methods. Thus, 1-furfurylurea was condensed with cyanoacetic
acid21,22 to give uracil 5 (79%). Direct alkylation of this uracil was
performed with 15% aqueous NaOH and the appropriate alkyl ha-
lide21,23 or by heating under reflux with (NH4)2SO4 in HMDS and
the subsequent addition of I2 and the appropriate halide17,24 to give
the corresponding 1,3-disubstituted-6-aminouracil 6. Standard
nitrosation of 6(a–l) with sodium nitrite in acetic acid was fol-
lowed by reduction with sodium dithionite to give diaminouracils
8(a–l). Finally, condensation of diaminouracils 8 with an appropri-
ate carboxylic acid in the presence of diisopropylcarbodiimide
(DIC) in MeOH, and subsequent cyclization by heating under reflux
with 2.5 N NaOH in MeOH afforded the xanthines 9. The 7-methyl-

M. C. Balo et al. / Bioorg. Med. Chem. 17 (2009) 6755–6760
6757
O
S
O
O
N
O
O
O
N
H
N
H
N
S
H
N
S
R
R
N
N
N
i)
ii)
N
O
N
O
N
N
O
O
O
O
11a: R = phenyl
9x: R = phenyl
9y: R = furan-2-yl
9z
12
11b: R = furan-2-y
11c
: R = thiophen-2-yl
: R = thiophen-2-yl
Scheme 2. Reagents and conditions: (i) oxone, aliquat, CH3CN, H2O, CH2Cl2, rt; (ii) AcOH, 30%, H2O2, rt.
Table 1
Chemical structures and binding affinitiesa at hA2B, hA2A, rA1 and hA3 ARs of 1,8-disubstituted 3-furfurylxanthine derivatives
O
H
R1
N
R2
N
N
N
O
O
Compound
R1
R2
R2
hA2B
hA2A
rA1
9a
9b
9c
9d
9e
9f
9g
9h
9i
9j
9k
9l
9m
9n
9o
9p
9q
9r
Metil
Ethyl
Ethyl
Ethyl
Propyl
Propyl
Propyl
Isobutyl
Isobutyl
Pentyl
Pentyl
Cyclopropylmethyl
Cyclopropylmethyl
Prop-2-ynyl
Prop-2-ynyl
Allyl
Furan-2-yl
Phenyl
Furan-2-yl
Thiophen-2-yl
Phenyl
Furan-2-yl
Thiophen-2-yl
Phenyl
Thiophen-2-yl
Phenyl
Thiophen-2-yl
Thiophen-2-yl
2,6-Difluorophenyl
Thiophen-2-yl
2,6-Difluorophenyl
Thiophen-2-yl
2,6-Difluorophenyl
Phenyl
Thiophen-2-yl
2,6-Difluorophenyl
Phenyl
Thiophen-2-yl
Thiophen-2-yl
2,6-Difluorophenyl
Phenyl
Furan-2-yl
Thiophen-2-yl
Phenyl
Furan-2-yl
Thiophen-2-yl
Phenyl
6.50
6.79
6.90
6.92
6.93
66%
68%
33%
44%
47%
17%
42%
59%
6.57
7.05
6.26
24%
65%
70%
70%
31%
27%
7.03
6.70
6.08
6.19
6.56
9%
6.37
5.98
6.28
6.44
6.38
33%
19%
13%
11%
7%
14%
41%
24%
6.33
6.52
6.38
6.47
6.31
n.d.b
6.06
21%
6.46
14%
6.66
6.36
23%
3%
27%
17%
28%
25%
n.d.
6.82
6.70
6.40
42%
32%
1%
6%
5%
18%
3%
9%
3%
2%
8%
n.d.
24%
11%
21%
7%
11%
5%
4%
1%
5%
2%
4%
6%
n.d.
8%
2%
14%
4%
13%
26%
n.d.
n.d.
1%
10%
12%
6%
6.35
6.34
5.84
11%
32%
34%
23%
2%
Allyl
2-Methoxyethyl
2-Methoxyethyl
2-Methoxyethyl
2-Ethoxyethyl
2-Ethoxyethyl
2-(Methylthio)ethyl
2-(Methylthio)ethyl
2-(Ethylthio)ethyl
2-(Ethylthio)ethyl
2-(Ethylthio)ethyl
2-(Ethylsulfinyl)ethyl
2-(Ethylsulfinyl)ethyl
2-(Ethylsulfinyl)ethyl
2-(Ethylsulfonyl)ethyl
9s
9t
9u
9v
9w
9x
9y
9z
9aa
11a
11b
11c
12
1%
5.68
5.91
11%
17%
4%
11%
5%.
10%
1%
13%
33%
20%
n.d.
n.d.
2%
a
Binding affinity is expressed as pKi or displacement percentage at 1 lM where indicated. The SEM was always lower than 10%.
Not determined.
b
affinity values for A2B receptors, while selectivities with regard to
both A2A and A1 receptors remained low. Finally, oxidation of the
sulfur atom of the 1-(alkylthioalkyl) chain of xanthines 9(y–aa), re-
sulted in the loss of binding affinity at all the AR assayed [com-
pounds 11(a–c) and 12].
On the other hand, the 7-methylxanthine derivatives (Table 2)
showed increased values of affinity for hA2B, hA2A and rA1 recep-
tors, while keeping an almost complete lack of affinity for hA3
receptors. Generally speaking, A2B/A2A and A2B/A1 selectivities im-
proved for this series, though not spectacularly.
was reached (compound 10i) at the cost of a decrease in A2B affin-
ity (Ki = 41 nM) and in A2B/A1 selectivity (2.6-fold). A more bal-
anced situation can be seen for compound 10m, with
intermediate affinity at A2B receptors (Ki = 28 nM) and 25-fold
and 14-fold selectivities over A2A and A1 receptors, respectively.
Still, in some cases affinity for A1 receptors was equal (10o) or even
slightly greater (10e) than for A2B receptors.
4. Conclusions
Thus, compound 10d showed high affinity at A2B receptors
(Ki = 7.4 nM), with 11-fold and 14-fold selectivities over A2A and
A1 receptors, respectively. Maximum (39-fold) A2B/A2A selectivity
In summary, an important set of xanthine derivatives, com-
pounds 9(a–aa), 10(a–o), 11(a–c) and 12, have been synthesized

6758
M. C. Balo et al. / Bioorg. Med. Chem. 17 (2009) 6755–6760
Table 2
Chemical structures and binding affinitiesa at hA2B, hA2A, rA1 and hA3 ARs of 1,8-disubstituted 3-furfuryl-7-methylxanthine derivatives
O
CH3
R1
R2
N
N
N
N
O
O
Compound
R1
R2
hA2B
hA2A
rA1
hA3 (%)
A2B/A2A ratiob
A2B/A1 ratio
10a
10b
10c
10d
10e
10f
10g
10h
10i
10j
10k
10l
10m
10n
10o
Methyl
Ethyl
Ethyl
Ethyl
Propyl
Propyl
Propyl
Cyclopropylmethyl
Cyclopropylmethyl
Allyl
Furan-2-yl
Phenyl
Furan-2-yl
Thiophen-2-yl
Phenyl
7.49
7.89
7.86
8.13
7.36
7.72
7.83
7.45
7.33
7.67
7.67
7.21
7.56
7.41
7.05
6.83
7.05
6.98
7.08
6.31
6.51
6.62
6.31
5.74
6.51
6.42
6.04
6.16
6.31
6.05
6.28
7.54
6.97
6.99
7.57
6.88
6.75
7.0
6.92
6.88
6.88
6.76
6.40
6.71
7.05
45
16
11
5
41
41
65
65
1
12
23
5
4
1
4.57
6.90
7.60
11.2
11.2
16.2
16.2
13.8
38.9
14.5
17.8
14.8
25.1
12.6
10
16.2
2.24
7.76
13.8
0.62
6.92
12.0
2.82
2.75
6.17
6.17
2.82
14.4
5.02
1.0
Furan-2-yl
Thiophen-2-yl
Thiophen-2-yl
2,6-Difluorophenyl
Thiophen-2-yl
2,6-Difluorophenyl
Phenyl
Thiophen-2-yl
2,6-Difluorophenyl
Thiophen-2-yl
Allyl
2-Methoxyethyl
2-Methoxyethyl
2-Methoxyethyl
2-(Ethylthio)ethyl
52
a
Binding affinity is expressed as pKi or displacement percentage at 1
Affinity ratios were calculated on the basis of Ki values.
lM where indicated. The SEM was always lower than 10%.
b
and their affinities for four subtypes of AR have been evaluated.
The pharmacological activity data (Tables 1 and 2) show unequiv-
ocally that the presence of an additional methyl group at position 7
proved to be beneficial for A2B, A2A and A1 affinities. This is a rather
surprising finding, at the light of a short but illustrative precedent
found in a related series of xanthines.17 Also important is the fact
that affinities for A2B receptors at the one digit nM level can be
found for a combination of appropriate substituents at several
positions of the xanthine scaffold, without adhering to the 8-(N-
substituted-phenoxyacetamido) motif proposed by Jacobson.18 De-
spite the low A2B/A2A and A2B/A1 selectivity ratios found for these
compounds, this work gives a deeper insight into the universe of
structural variability which significantly affects the biological
properties here studied.
recorded on Hewlett–Packard HP5988A or Micromass Autospec
spectrometers. Elemental analyses were performed in a FISONS
EA 1108 Elemental Analyser at the University of Santiago Micro-
analysis Service; all results shown are within 0.4% of the theoret-
ical values (C, S, N, H).
5.1. General procedure for the preparation of 3-substituted 6-
amino-1-furfuryluracils 6(a–l)
5.1.1. Method A
A mixture of 6-amino-1-furfuryluracil (1 mmol), 15% NaOH
(0.3 mL) and 95% EtOH (0.62 mL) was heated under reflux for
15 min and the corresponding alkylating agent RX (2 mmol) was
added dropwise. The resulting solution was heated under reflux
for 0.25–48 h. The solvents were removed under reduced pressure
and the residue was partitioned between CHCl3/H2O (2/1, 6.3 mL).
The organic layer was washed with H2O, dried (Na2SO4) and evap-
orated to dryness to give the corresponding 3-substituted 6-ami-
no-1-furfuryluracils, which in most cases were used in
subsequent steps without further purification.
5. Experimental
All chemicals were of reagent grade and were obtained from Al-
drich Chemical Co. and used without further purification. When
necessary, solvents were dried by standard techniques and dis-
tilled. All air-sensitive reactions were carried out under argon.
Flash chromatography was performed on silica gel (Merck 60,
230–240 mesh) and analytical TLC was carried out on pre-coated
silica gel plates (Merck 60 F254, 0.25 mm) type E. Chromatographic
spots were visualized by UV light or with Hanessian reagent.27
Melting points (uncorrected) were measured in glass capillary
tubes on a Stuart Scientific electro thermal apparatus SMP3. Infra-
red spectra were recorded on a Perkin–Elmer 1640 FTIR spectro-
photometer. 1H NMR and 13C NMR spectra were recorded on a
Bruker AMX 300 spectrometer at 300 and 75.47 MHz, respectively,
using TMS as internal standard (chemical shifts in d values, J in
hertz). All of the observed signals are consistent with the proposed
structures. Chemical shifts (d scale) are reported in parts per mil-
lion (ppm) relative to the centre of the solvent peak. Coupling con-
stants (J values) are given in hertz (Hz). Spin multiplicities are
given as s (singlet), d (doublet), dd (double doublet), t (triplet), m
(multiplet), br s (broad singlet), v s (virtual singlet), dt (double trip-
let), q (quadruplet), qt (quintuplet), sex (sextet). Mass spectra were
5.1.1.1. 6-Amino-1-furfuryl-3-propyluracil (6c). Method A,
alkylating agent: propyl bromide, reaction time 6 h, foamy solid,
yield 54%. 1H NMR (DMSO-d6) d: 7.58 (d, J = 0.9 Hz, 1H, 5-H
C4H3O), 6.85 (s, 2H D2O exchange, NH2), 6.38 (dd, J = 3.1, 1.9 Hz,
1H, 4-H C4H3O), 6.30 (d, J = 3.1 Hz, 1H, 3-H C4H3O), 5.05 (s, 2H,
CH2–C4H3O), 4.66 (s, 1H, 5-H), 3.66 (t, J = 7.4 Hz, 2H, NCH2), 1.47
(sex, J = 7.4 Hz, 2H, CH2CH3), 0.79 (t, J = 7.4 Hz, 3H, CH3). HRMS
m/z calcd for C12H15N3O3, 249.1113; found, 227.1130.
5.1.2. Method B
A suspension of 6-amino-1-furfuryluracil (1.0 g, 7.8 mmol) and
(NH4)2SO4 (0.05 g) in hexamethyldisilazane (HMDS, 10 mL) was
heated under reflux for 2 h and during this time the mixture be-
came homogeneous. Excess HMDS was distilled off, at first under
atmospheric pressure and then under vacuum. The product was al-
lowed to cool to 70 °C, a temperature above the melting point of
the compound as it had to be liquid for the subsequent step. A cat-

M. C. Balo et al. / Bioorg. Med. Chem. 17 (2009) 6755–6760
6759
alytic amount of I2 (ca. 8 mg) was dissolved in the product and an
80% toluene solution of the corresponding organic bromide
(1.0 mL, 9.0 mmol) was added. The mixture was heated in an oil
bath under reflux for (1.5–3 h) and then was allowed to cool to
room temperature and a solution of Na2S2O3 (1.5 g) in H2O
(5 mL) was added. The flask was cooled in an ice bath and a satu-
rated aqueous NaHCO3 (ca. 40 mL), was added in small portions
over a period of 15 min with vigorous stirring until effervescence
ceased, and the gelatinous mixture had turned into a suspension.
The precipitate was filtered off, washed with cold water (15 mL),
toluene (10 mL), and ether (10 mL).
5.4. General procedure for the preparation of xanthines 9(a–aa)
Diisopropylcarbodiimide (1 mmol) was added to a solution or
suspension of the corresponding carboxylic acid (1 mmol) in anhy-
drous MeOH (2 mL) and this was followed by the addition of the
appropriate diaminouracil 8(a–l) (1 mmol). The reaction mixture
was stirred at room temperature for 30 min. The solvent was re-
moved under reduced pressure and the sticky residue was tritu-
rated with H2O. The resulting solid was filtered off and mixed
with MeOH (3.5 mL) and 2.5 N NaOH (5 mL). The mixture was
heated under reflux for the appropriate time in each case and al-
lowed to cool down to room temperature. The solid was filtered
off and the filtrate was adjusted to pH 6 by the addition of 2 N
HCl. The corresponding 1H-purine-2,6(3H,7H)-dione precipitated
and was filtered off, washed with H2O and purified by crystalliza-
tion or washing with the appropriate solvent.
5.1.2.1. 6-Amino-1-furfuryl-3-(prop-2-ynyl)uracil (6g). Method B,
alkylating agent: prop-2-ynyl bromide, reaction time 1 h, pink solid,
yield 38%. 1H NMR (CDCl3) d: 7.38 (v s, 1H, 5-H C4H3O), 6.41 (v s,
1H, 3-H C4H3O), 6.30 (v s, 1H, 4-H C4H3O), 5.01 (s, 2H, CH2–
C4H3O), 4.70 (s, 1H, 5-H), 3.46–3.45 (m, 2H, CHCCH2), 2.21 (s, 1H,
CHCCH2), 2.16 (br s, 2H, D2O exchange, NH2). Anal. Calcd for
C12H11N3O3, (245.23): C, 58.77; H, 4.52; N, 17.13. Found: C, 59.04;
H, 4.76; N, 17.40.
5.4.1. 3,8-Difurfuryl-1-methyl-1H-purine-2,6(3H,7H)-dione (9a)
Reaction time 30 min, white solid, mp 204–206 °C (MeOH),
yield 18%. IR (KBr)
m
(cmÀ1): 3144, 3092, 3033, 1708, 1656, 1555,
1504, 1411, 1398, 1288, 1222, 1151, 1004, 939, 825, 760, 507. 1H
NMR (CDCl3) d: 12.52 (s, 1H, D2O exchange, NH), 7.32–7.29 (m,
2H, 2 Â (5-H C4H3O), 6.43 (d, J = 3.0 Hz, 1H, 3-H C4H3O), 6.31–
6.28 (m, 2H, 2 C4H3O), 6.21 (d, J = 2.6 Hz, 1H, C4H3O), 5.31 (s, 2H,
NCH2–C4H3O), 4.27 (s, 2H, CH2–C4H3O), 3.42 (s, 3H, CH3). 13C
NMR and DEPT (CDCl3) d: 156.05 (C8), 151.39 (C6), 151.32 (C2),
149.82 (C4), 149.37 (C2 C4H3O), 148.96 (C2 C4H3O), 142.83 (C5
C4H3O) 142.64 (C5 C4H3O), 111.05, 110.85, 109.98 and 108.07
(2C3 and 2C4 C4H3O), 107.55 (C5), 40.27 (CH2–C4H3O), 28.86
(CH2–C4H3O), 28.74 (CH3). MS (EI) m/z (%): 326 (M, 6), 148 (1),
269 (3), 84 (13), 82 (6), 81 (100), 78 (1), 68 (2), 54 (1), 53 (12),
52 (2), 51 (2). Anal. Calcd for C16H14N4O4 (326.30): C, 58.89; H,
4.32; N, 17.17. Found: C, 59.12; H, 4.61; N, 17.42.
5.2. General procedure for the preparation of 3-substituted 6-
amino-1-furfuryl-5-nitrosouracils 7(a–l)
A solution of NaNO2 (18 mmol) in H2O (7 mL) was added slowly
over 15 min to a solution of the corresponding 3-substituted 6-
amino-1-furfuryluracil 6 (6 mmol) in 50% aqueous AcOH (30 mL)
at 80 °C. The reaction mixture was stirred at room temperature.
In cases where a precipitate was formed the solid was filtered
off, washed with water and dried under vacuum. In cases where
precipitation was only slight the aqueous solution was extracted
with EtOAc. The organic phase was dried (Na2SO4), the solvent
was removed under reduced pressure and the residue was crystal-
lized from the appropriate solvent.
5.5. General procedure for the methylation of compounds 9(a–
g), 9(l,m), 9(p–t) and (9y)
5.2.1. 6-Amino-3-ethyl-1-furfuryl-5-nitrosouracil (7b)
Reaction time 24 h, violet solid, mp 192–194 °C (EtOAc), yield
54%. 1H NMR (DMSO-d6) d: 3.24 (br s, 1H, D2O exchange, NHH),
9.27 (br s, 1H, D2O exchange, NHH), 7.61 (d, J = 0.8 Hz, 1H, 5-H
C4H3O), 6.45 (d, J = 3.0 Hz, 1H, 3-H C4H3O), 6.40 (dd, J = 3.0,
1.8 Hz, 1H, 4-H C4H3O), 5.13 (s, 2H, CH2–C4H3O), 3.93 (q,
J = 7.0 Hz, 2H, CH2CH3), 1.06 (t, J = 7.0 Hz, 3H, CH3). Anal. Calcd
for C11H12N4O4, (264.23): C, 50.00; H, 4.58; N, 21.20. Found: C,
50.42; H, 4.69; N, 21.09.
To a suspension of NaH (1.2 mmol) in dry DMF (10 mL) was
added the corresponding 1H-purine-2,6(3H,7H)-dione 9 (1 mmol)
and the mixture was shaken at room temperature for 15 min and
at 60 °C for a further 15 min. Once the mixture had reached room
temperature CH3I (1 mmol) was added. The reaction was heat at
100 °C for 2 h and then allowed to cool down to room temperature.
Water was added and the resulting precipitate was filtered off,
washed with H2O and dried under vacuum.
5.3. General procedure for the preparation of 3-substituted 5,6-
diamino-1-furfuryluracils 8(a–l)
5.5.1. 3,8-Difurfuryl-1,7-dimethyl-1H-purine-2,6(3H,7H)-dione
(10a)
Na2S2O4 (16 mmol) was added in small portions to a well-stir-
red suspension of the corresponding 3-substituted 6-amino-1-fur-
furyl-5-nitrosouracil 6 (8 mmol) in 30% NH4OH (25 mL) at 60 °C.
On completion of the addition the reaction mixture was heated
for 1 to 4 h and then cooled to 4–5 °C for 18 h. The resulting precip-
itate was filtered off, washed with H2O and dried under vacuum.
The volume of the filtrate was reduced to give a second crop of
the corresponding diaminouracil.
White solid, mp 121–123 °C, yield 58%. IR (KBr)
m
(cmÀ1): 3433,
3146, 2956, 1709, 1660, 1542, 1502, 1446, 1411, 1336, 1171, 1124,
1012, 971, 760, 737. 1H NMR (CDCl3) d: 7.41 (v s, 1H, 5-H C4H3O),
7.33 (v s, 1H, 5-H C4H3O), 6.44–6.41 (m, 1H, C4H3O), 6.37–6.29 (m,
2H, 2C4H3O), 6.16 (d, J = 3.1 Hz, 1H, C4H3O), 5.27 (s, 2H, NCH2–
C4H3O), 4.20 (s, 2H, CH2–C4H3O), 3.92 (s, 3H, CH3), 3.39 (s, 3H,
CH3). 13C NMR and DEPT (CDCl3) d: 155.85 (C8), 151.49 (C6),
150.02 (C2), 149.82 (C4), 149.37 (C2 C4H3O), 147.61 (C2 C4H3O),
142.74 (C5 C4H3O), 142.65 (C5 C4H3O), 111.11 (C4H3O), 110.81
(C4H3O), 109.79 (2C C4H3O), 107.96 (C5), 39.97 (CH2–C4H3O),
30.10 (CH2–C4H3O), 28.74 (CH3), 27.29 (CH3). MS (EI) m/z (%):
340 (M 9), 326 (23), 316 (39), 256 (20), 153 (20), 141 (17), 127
(21), 125 (21), 113 (24), 111 (25), 99 (29), 85 (51), 83 (24), 80
65), 70 (67), 68 (49), 56 (100), 54 (19). Anal. Calcd for
C17H16N4O4 (340.34): C, 60.00; H, 4.74; N, 16.46. Found: C, 59.72;
H, 4.44; N, 16.32.
5.3.1. 5,6-Diamino-3-ethyl-1-furfuryluracil (8b)
Reaction time 2.5 h, green solid, yield 91%. 1H NMR (DMSO-d6)
d: 7.31 (d, J = 1.7 Hz, 1H, 5-H C4H3O), 6.39 (d, J = 3.1 Hz, 1H, 3-H
C4H3O), 6.31 (dd, J = 3.1, 1.9 Hz, 1H, 4-H C4H3O), 5.11 (br s, 2H,
D2O exchange, NH2), 5.03 (s, 2H, CH2–C4H3O), 3.90 (q, J = 7.1 Hz,
2H, CH2CH3), 2.24 (br s, 2H, D2O exchange, NH2), 1.17 (t,
J = 7.1 Hz, 3H, CH3). Anal. Calcd for C11H14N4O3 (250.25): C,
52.79; H, 5.64; N, 22.39. Found: C, 53.04; H, 5.86; N, 22.53.

6760
M. C. Balo et al. / Bioorg. Med. Chem. 17 (2009) 6755–6760
5.6. General procedure for the oxidation of sulfides 9(y–aa) to
sulfoxides 11(a–c), respectively
(CH2–C6H5), 35.18 (NCH2), 6.52 (CH3). MS (EI) m/z (%): 442 (M,
13), 279 (2), 278 (4), 250 (3), 226 (2), 91 (9), 82 (6), 81 (100), 53
(15). Anal. Calcd for C22H24N4O5S (456.51): C, 57.88; H, 5.30; N,
12.27; S, 7.02. Found: C, 57.35; H, 5.62; N, 12.44; S, 7.48.
OXONEÒ (0.67 mmol) and aliquat (three drops) in a mixture of
H2O (5.8 mL) and CH2Cl2 (3.8 mL) were added to a solution of the
corresponding 1-[2-(ethylthio)ethyl]-1H-purine-2,6(3H,7H)-dione
9(y–aa) (1 mmol) in CH3CN (0.4 mL) cooled to 0 °C, and the mix-
ture was stirred at room temperature for the time quoted below.
The reaction mixture was then poured into EtOAc (100 mL) and
the organic layer was washed twice with H2O, dried (Na2SO4)
and the solvents evaporated under reduced pressure to leave a res-
idue that was purified by recrystallization or column chromatogra-
phy on silica gel.
Acknowledgments
The authors thank Almirall Prodesfarma S.A. (Barcelona-Spain)
for promoting and financially supporting this work. I.N. thanks
the Xunta de Galicia for financial support under ‘Programa Isidro
Parga Pondal’. J.B. thanks the Xunta de Galicia for financial support
under ‘Isabel Barreto Contract’.
5.6.1. 8-Benzyl-1-[2-(ethylsulfinyl)ethyl]-3-furfuryl-1H-purine-
2,6(3H,7H)-dione (11a)
Supplementary data
White solid, mp 165–167 °C (CH3CN), yield 65%. IR (KBr) m
(cmÀ1):
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmc.2009.07.034.
3140, 3088, 2969, 1704, 1658, 1602, 1556, 1503, 1453,1409, 1268,
1215, 1155, 1105, 1018, 980, 770, 754, 729, 697. 1H NMR (CDCl3) d:
12.36 (s, 1H, D2O exchange, NH), 7.34–7.19 (m, 6H, 2-H, 3-H, 4-H, 5-
H, 6-H C6H5 and 5-H C4H3O), 6.41 (d, J = 3.2 Hz, 1H, 3-H C4H3O),
6.29 (dd, J = 3.1, 1.9 Hz, 1H, 4-H C4H3O), 5.28 (s, 2H, CH2–C4H3O),
4.45–4.22 (m, 2H, CH2N), 4.18 (s, 2H, CH2–C6H5), 3.31–3.22 (m, 1H,
SOCHH), 3.02–2.73 (m, 3H, CH2SOCHH), 1.32 (t, J = 7.5 Hz, 3H, CH3).
13C NMR and DEPT (CDCl3) d: 154.85 (C8), 154.35 (C6), 151.32 (C2),
149.86 (C4), 149.08 (C2 C4H3O), 142.80 (C5 C4H3O), 136.57 (C1
C6H5), 129.26 and 129.15 (C2, C3, C5, C6 C6H5), 127.46 (C4 C6H5),
110.86 (C4 C4H3O), 109.94 (C3 C4H3O), 107.46 (C5), 50.19 (SOCH2),
45.92 (CH2–C4H3O), 40.24 (CH2SO) 36.33 (CH2–C6H5), 35.70 (NCH2),
7.17 (CH3). Anal. Calcd for C21H22N4O4S (426.49): C, 59.14; H, 5.20;
N, 13.14; S, 7.52. Found: C, 59.55; H, 5.47; N, 13.45; S, 7.12.
References and notes
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5.7. 8-Benzyl-1-[2-(ethylsulfonyl)ethyl]-3-furfuryl-1H-purine-
2,6(3H,7H)-dione (12)
11. Volpini, R.; Costanzi, S.; Vittori, S.; Cristalli, G.; Klotz, K. N. Curr. Top. Med. Chem.
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12. Kim, Y.-C.; Karton, Y.; Ji, X.-d.; Melman, N.; Linden, J.; Jacobson, K. A. Drug Dev.
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A mixture of the 1-[2-(ethylthio)ethyl]-1H-purine-2,6(3H,7H)-
dione 9y (0.14 g, 0.35 mmol), HOAc (0.18 mL) and 30% H2O2
(0.24 mL) was stirred at room temperature for 24 h, H2O (30 mL)
was then added and the mixture was extracted with CH2Cl2
(3 Â 30 mL). The combined organic layers were washed with
H2O, dried (Na2SO4) and the solvent was evaporated under reduced
pressure to leave a solid residue (0.15 g) that was purified by
recrystallization.
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14. Elzein, E.; Kalla, R. V.; Li, X.; Perry, T.; Gimbel, A.; Zeng, D.; Lustig, D.; Leung, K.;
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Compound 12: White solid, mp 210–212 °C (MeOH), yield 97%.
IR (KBr)
m
(cmÀ1): 3176, 3110, 3036, 1705, 1667, 1553, 1495,
1347, 1289, 1121, 1013, 782, 696. 1H NMR (CDCl3) d: 11.84 (s,
1H, D2O exchange, NH), 7.35–7.22 (m, 6H, 2-H, 3-H, 4-H, 5-H, 6-
H C6H5 and 5-H C4H3O), 6.43 (d, J = 3.2 Hz, 1H, 3-H C4H3O), 6.31
(dd, J = 3.2, 1.9 Hz, 1H, 4-H C4H3O), 5.31 (s, 2H, CH2–C4H3O), 4.48
(t, J = 3.2 Hz, 2H, CH2N), 4.24 (s, 2H, CH2–C6H5), 3.33 (t, J = 7.3 Hz,
2H, SO2CH2), 3.08 (q, J = 7.5 Hz, 2H, CH2SOCH2), 1.38 (t, J = 7.4 Hz,
3H, CH3). 13C NMR and DEPT (CDCl3) d: 154.70 (C8), 154.49 (C6),
150.54 (C2), 149.16 (C4), 149.12 (C2 C4H3O), 142.49 (C5 C4H3O),
135.64 (C1 C6H5), 128.91 and 128.76 (C2, C3, C5, C6 C6H5),
127.33 (C4 C6H5), 110.49 (C4 C4H3O), 109.70 (C3 C4H3O), 106.70
(C5), 48.99 (CH2–C4H3O), 47.25 (SO2CH2), 39.90 (CH2SO2), 35.31
27. Touchstone, J. In Advances in Thin-Layer Chromatography; Wiley: New York,
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