Chalcone-based derivatives as new scaffolds for hA3 adenosine receptor antagonists

Article abstract of DOI:10.1111/jphp.12028

Objectives With the aim of finding new adenosine receptor (AR) ligands based on the chalcone scaffold, we report the synthesis of a new series of coumarin-chalcone hybrids and the pharmacological characterization of their actions at four subtypes of AR. Methods The synthesized compounds 5-10 were characterized in radioligand binding (A₁, A_2A and A ₃) and adenylyl cyclase activity assays (A_2B) to determine the affinity of the compounds for the four human AR (hAR) subtypes. Key findings Coumarin-chalcone hybrids were found to be ligands with a novel structure, not reported thus far, that showed varying affinity and selectivity for AR subtypes. Conclusions The coumarin-chalcone hybrids in which ring B of the chalcone scaffold was a thiophene (compounds 5 and 9) were found to be the most potent compounds of the series. Compound 9, in which ring A of the chalcone moiety was the phenyl ring of the coumarin, showed similar activity against hA₁, hA_2A and hA₃ ARs, while compound 5, in which ring A of the chalcone was substituted by the benzopyrone ring of the coumarin moiety, showed similar activity only at the hA₃ AR and, therefore, was deemed to be selective (K_i (dissociation constant) = 5160 nm). A new series of chalcone-based derivatives, including a coumarin moiety in their structures, have been synthesized and binding assays for the hA1, hA2A, and hA3 adenosine receptors were employed to assess the affinity for receptor subtypes. Isosterical substitutions of one or both phenyl rings of the chalcone moiety reveal that thiophene-containing compounds 5 and 9 have good binding affinity for ARs. Compound 5, in which both phenyl rings of the original chalcone scaffold (rings A and B) were substituted for a thiophene and pyrone rings, resulted selective for the hA3 adenosine receptor (Ki = 5,160 nM). Based on the obtained results, a preliminary structure-activity relationship (SAR) study showed a high selectivity and good potency of these thiophene-containing compounds. 2013 The Authors. JPP

Full text of DOI:10.1111/jphp.12028

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Research Paper
Journal of Pharmacy
And Pharmacology
Chalcone-based derivatives as new scaffolds for hA3
adenosine receptor antagonists
Saleta Vazquez-Rodrigueza, Maria João Matosa,b, Lourdes Santanaa, Eugenio Uriartea,
Fernanda Borgesb, Sonja Kachlerc and Karl-Norbert Klotzc
aDepartamento de Química Orgánica, Facultad de Farmacia, Universidad de Santiago de Compostela, Santiago de Compostela, Spain,
c
bCIQUP/Departmento de Química e Bioquímica, Faculdade de Ciências, Universidade do Porto, Porto, Portugal and Institut für Pharmakologie und
Toxikologie, Universität Würzburg, Würzburg, Germany
Keywords
adenosine receptor; chalcone;
Abstract
Objectives With the aim of finding new adenosine receptor (AR) ligands based
on the chalcone scaffold, we report the synthesis of a new series of coumarin–
chalcone hybrids and the pharmacological characterization of their actions at four
subtypes of AR.
Methods The synthesized compounds 5–10 were characterized in radioligand
binding (A1, A2A and A3) and adenylyl cyclase activity assays (A2B) to determine the
affinity of the compounds for the four human AR (hAR) subtypes.
Key findings Coumarin–chalcone hybrids were found to be ligands with a novel
structure, not reported thus far, that showed varying affinity and selectivity for AR
subtypes.
coumarin-chalcone hybrids; structure–activity
relationship
Correspondence
Saleta Vazquez-Rodriguez, Departamento de
Química Orgánica, Facultad de Farmacia,
Universidad de Santiago de Compostela,
15782 Santiago de Compostela, Spain.
E-mail: svre77@gmail.com
Received November 9, 2012
Accepted December 19, 2012
Conclusions The coumarin–chalcone hybrids in which ring B of the chalcone
scaffold was a thiophene (compounds 5 and 9) were found to be the most potent
compounds of the series. Compound 9, in which ring A of the chalcone moiety
was the phenyl ring of the coumarin, showed similar activity against hA1, hA2A and
hA3 ARs, while compound 5, in which ring A of the chalcone was substituted
by the benzopyrone ring of the coumarin moiety, showed similar activity only
at the hA3 AR and, therefore, was deemed to be selective (Ki (dissociation
constant) = 5160 nm).
doi: 10.1111/jphp.12028
Introduction
Adenosine is a purine nucleoside produced by all meta-
bolically active cells. It acts as an endogenous modulator
controlling a wide range of physiological processes due to
its interaction with four specific cell membrane G-protein-
coupled receptors (GPCRs) classified as A1, A2A, A2B and A3
adenosine receptors (ARs).[1–3] Because of their ubiquitous
presence in cells, ARs have been seen as promising targets
in the field of medicinal chemistry. ARs are distributed
along different tissues in mammalian systems and regulate
diverse physiological functions by modulating cell signal-
ling, being activated by endogenous adenosine and blocked
by antagonists.[4,5] In the last two decades, a large number
of ligands have been synthesized in the search for potent
and selective agonists and antagonists for each AR subtype;
selective A2BAR agonists are among the most recently
reported.[6] Targeting ARs has opened a new window for
potential drug treatment of a variety of pathologies such
as asthma, neurodegenerative disorders, cancer and inflam-
matory and ischaemic conditions.[7–12] More specifically,
AR antagonists are involved in several pathological proc-
esses such as inflammation (A2A),[13,14] heart and renal
failure (A1)[15] or neurological disorders like Parkinson’s[16]
and Alzheimer’s disease (A2A and/or A1).[17]
AR antagonists developed recently present a chemical
diversity, while the 1,3-dialkylxanthines are considered as
derivatives of the classical scaffold.[18,19] Among the non-
classical antagonists, the flavone and isoflavone derivatives
have played a remarkable role, namely genistein, which has
been described as a competitive antagonist at the A1 AR in
FRTL (thyroid) cells[20] or galangin, which was found to
bind three subtypes of ARs, and it has been shown that this
type of compound presents micromolar affinity for the A3
AR.[21] (Figure 1)
Coumarins (chromone isosteres) and chalcones (flavone
with an opened pyrone ring) are another class of
benzopyran-related compounds of natural origin that
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
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Chalcone derivatives as hA3 receptors
Saleta Vazquez-Rodriguez et al.
O
O
OH
O
OH
HO
O
Galangin
Flavone
O
O
A
Phenyl/Thiophenyl
O
O
O
B
O
Chalcone
Coumarin
Coumarin–chalcone hybrid
Figure 1 Rational design of the synthesized compounds based on flavonoid scaffolds.
present a wide range of pharmacological actions.[22–24]
Coumarins have health value mainly due to their antimi-
crobial,[25] enzyme-inhibitory,[26–28] anticancer[29] and anti-
oxidant[30] activity.
75.47 MHz, respectively, using tetramethylsilane as internal
standard (chemical shifts in d values, J in Hz). Mass spectra
were obtained using a Hewlett-Packard 5988A spectrom-
eter. Elemental analyses were performed using a Perkin-
Elmer 240B microanalyser and were within Ϯ 0.4% of
calculated values in all cases. Silica gel (Merck 60, 230–00
mesh) was used for flash chromatography. Analytical thin-
layer chromatography (TLC) was performed on plates pre-
coated with silica gel (Merck 60 F254, 0.25 mm). The purity
of compounds was assessed by high-performance liquid
chromatography (HPLC) coupled at diode array detector
(DAD) on a Thermo Quest Spectrasystem (Thermo Fisher
Scientific, Waltham, MA, USA) equipped with a P4000
pump, a UV6000 UV-Vis diode array detector and an
SN4000 interface for operation via a personal computer.
Instrument software ChromQuest 5.0 (Thermo Fisher Sci-
entific) was used for data acquisition. Different analytical
columns and mobile phases (all solvents were HPLC grade)
were tested. The mobile phase was H2O–CH3CN (70:30)
and an Eclipse xdb C18 column (5 mm particle size,
0.46 mm i.d., 25 cm length; Agilent Technologies) was used.
The purity of the compounds was found to be higher than
95%.
On the other hand, chalcones also present related bio-
logical activity as antimicrobial and anti-inflammatory,[24]
anti-tumour,[29] antioxidant[31] or enzyme-inhibitory
effects.[32] Thus, due to structural similarities between fla-
vones, coumarins and chalcones, we decided to synthesize
hybrid compounds bearing both scaffolds in the same mol-
ecule, and to evaluate their activity towards the four sub-
types of human AR expressed in Chinese hamster ovary
(CHO) cells. The designed compounds have the chalcone
scaffold as the main structural core, and the different
derivatives are mainly based on the substitution of one or
both of the chalcone phenyl rings (A and/or B) for a cou-
marin and/or thiophene with different substituents
(Figure 1). In addition, the drug-like properties of the
hybrids were also evaluated.
Materials and Methods
Synthetic methodologies
Melting points were determined using a Reichert Kofler
thermopan or in capillary tubes on a Büchi 510 apparatus
and are uncorrected. 1H and 13C NMR spectra were
recorded on a Bruker AMX spectrometer at 300 and
Chemistry
The chalcone-based derivatives 5–10 were efficiently
synthesized according to the protocol outlined in Figure 2.
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
698

Saleta Vazquez-Rodriguez et al.
Chalcone derivatives as hA3 receptors
R1
O
O
R1
O
O
O
O
CHO
(a)
(b)
R2
(c)
EtO2C
O
OH
1 R1 = H
5 R1 = H, R2 = 3-thiophenyl
OH
3 R1 = OH
6 R1 = H, R2 = p-methoxyphenyl
7 R1 = OH, R2 = 3-thiophenyl
8 R1 = OH, R2 = p-methoxyphenyl
O
O
2
OH
O
O
OH
O
R2
(c)
O
O
O
4
9
R2 = 3-thiophenyl
10 R2 = p-methoxyphenyl
Figure 2 Protocol for synthesis of chalcone-based derivatives 5–10. Reagents and conditions: (a) solvent-free, piperidine, rt, 10 min; (b) POCl3,
glacial acetic acid, reflux, 30 min; (c) appropriate aldehyde, EtOH, piperidine, reflux, 2–6 h.
Different starting materials were used in accordance with
the substitution pattern of the final compounds. To
synthesize compounds 5 and 6,[33] we first prepared the
3-acetylcoumarin (1) by a Knoevenagel reaction using
salicylaldehyde and ethyl acetoacetate without solvent
and employing piperidine in a catalytic amount at room
Biological assays
The binding affinity of the compounds for the human AR
subtypes hA1, hA2A and hA3 was determined using radiolig-
and competition experiments in CHO cells that had been
stably transfected with the individual receptor subtypes.[36,37]
The radioligands used were 1 nm [3H]2-chloro-N6-
cyclopentyladenosine (CCPA) for hA1, 10 nm [3H]
N-ethylcarboxamidoadenosine (NECA) for hA2A and 1 nm
[3H]2-(1-Hexynyl)-N-methyladenosine (HEMADO) for
hA3 receptors. Due to the lack of a suitable radioligand for
hA2B receptors the potency of antagonists at hA2B receptor
(expressed on CHO cells) was determined by inhibition
of NECA-stimulated adenylyl cyclase activity.[36] The 50%
inhibitory concentration (IC50) for inhibition of cAMP
(cyclic adenosine monophosphate) production was deter-
mined and converted to a Ki value (dissociation constants)
using the Cheng and Prusoff equation.[38] For all the tested
compounds no measurable activity for the hA2B AR
(Ki > 10 000 nm) was detected.
temperature.[34] To prepare compounds
7
and 8,[35]
4-hydroxycoumarin (2) was used as the starting material.
Acylation at the 3 position of compound 2, using POCl3 in
glacial acetic acid under reflux, afforded precursor 3
(93%).[35] Compounds 9 and 10 were prepared from the
commercially available 8-acetyl-7-hydroxycoumarin (4).
Starting from precursors 1, 3 and 4, a Claisen-Schmidt
condensation in EtOH, using piperidine as base, and under
reflux mixed with the corresponding aromatic aldehydes,
afforded the desired final compounds 5–10 in good yields
(47–87%).
Synthesis of 3-(3-aryl)acryloylcoumarin (5–8) and
8-(3-aryl)acryloylcoumarin (9, 10) derivatives
The corresponding 3-acetylcoumarin (1 or 3, 1 mmol) or
8-acetylcoumarin (4) and the conveniently substituted
aromatic aldehyde (1.1 mmol) were dissolved in EtOH
(3 ml) and a catalytic amount of piperidine (0.05 ml) was
added. The reaction mixtures were stirred for 2–6 h under
reflux. After completion of reaction (followed by TLC), the
solvent was evaporated under vacuum and the dry residue
was purified by flash chromatography (hexane–ethyl
acetate, 85:15) to give the desired products 5–10.
Statistical methods
Ki values (dissociation constants) were determined in radio-
ligand competition experiment with seven or eight different
concentrations of test compound and each concentration
was tested in duplicate. Ki values are given as geometric
means of three independent experiments with 95% confi-
dence intervals. For analysis of the competition curves the
programme SCTFIT was used.[39]
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
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Chalcone derivatives as hA3 receptors
Saleta Vazquez-Rodriguez et al.
(E)-4-Hydroxy-3-(3-(thiophen-3-yl)acryloyl)coumarin
(7): Yield 87%. Mp 192–194°C 1H NMR (300 MHz, CDCl3)
dppm 8.25 (d, J = 15.6 Hz, 1H, Hb), 8.15–7.99 (m, 2H, Ha,
H-5), 7.77–7.61 (m, 2H, H-2’, H-5’), 7.52 (dd, J = 5.2,
1.3 Hz, 1H, H-4’), 7.43–7.23 (m, 3H, H-6, H-7, H-8). 13C
NMR (75 MHz, CDCl3) dppm 192.5, 181.5, 160.1, 154.6,
140.5, 138.3, 135.9, 131.1, 127.1, 125.9, 125.6, 124.2, 122.1,
116.9, 116.2. MS m/z (%) (ESI): 299 ([M + 1]+, 13), 298
([M]+, 68), 280 (38), 270 (46), 137 (100), 121 (99), 109 (86).
Anal. Calcd. for C16H10O4S: C, 64.42; H, 3.38. Found: C,
64.39; H, 3.32
Theoretical evaluation of absorption,
distribution, metabolism and
excretion properties
The absorption, distribution, metabolism and excretion
(ADME) properties of the studied compounds were calcu-
lated using the Molinspiration property programme.[40]
LogP was calculated using the methodology developed by
Molinspiration as a sum of fragment-based contributions
and correction factors. Topological polar surface area
(TPSA) was calculated based on the methodology published
by Ertl et al. as a sum of fragment contributions.[42] Oxygen-
and nitrogen-centred polar fragments were considered.
Polar surface area (PSA) has been shown to be a very good
descriptor characterizing drug absorption, including intesti-
nal absorption, bioavailability, Caco-2 permeability and
blood–brain barrier penetration. The method for calcula-
tion of molecule volume developed at Molinspiration is
based on group contributions. These have been obtained by
fitting the sum of fragment contributions to ‘real’ three-
dimensional (3D) volume for a training set of about 12 000,
mostly drug-like molecules. 3D molecular geometries for
a training set were fully optimized by the semi-empirical
AM1 method.
(E)-7-Hydroxy-8-(3-(thiophen-3-yl)acryloyl)coumarin (9):
1
Yield 47%. Mp 169–171°C. H NMR (300 MHz, CDCl3)
dppm 14.10 (bs, 1H, OH), 8.16 (d, J = 15.3 Hz, 1H, Hb),
7.98 (d, J = 15.1 Hz, 1H, Ha), 7.80–7.59 (m, 2H, H-4, H-5),
7.59–7.44 (m, 2H, H-6, H-2’), 7.39 (dd, J = 5.0, 2.8 Hz, 1H,
H-4’), 6.93 (d, J = 8.7 Hz, 1H, H-5’), 6.30 (d, J = 9.6 Hz, 1H,
H-3). 13C NMR (75 MHz, CDCl3) dppm 193.2, 167.8, 159.4,
144.3, 139.6, 138.3, 134.4, 130.4, 127.3, 125.9, 125.7, 115.8,
111.9, 111.0, 109.6. MS m/z (%) (ESI): 299 ([M + 1]+, 11),
298 ([M]+, 28), 203 (25), 189 (100), 84 (89). Anal. Calcd. for
C16H10O4S: C, 64.42; H, 3.38. Found: C, 64.44; H, 3.40.
(E)-7-Hydroxy-8-(3-(4-methoxyphenyl)acryloyl)coumarin
(10): Yield: 68%. Mp 187–188°C. 1H NMR (300 MHz,
CDCl3) dppm 14.00 (s, 1H, OH), 8.03 (d, J = 15.4 Hz, 1H,
Hb), 7.82 (d, J = 15.5 Hz, 1H, Ha), 7.62–7.42 (m, 3H, H-4,
H2’, H6’), 7.34 (d, J = 8.8 Hz, 1H, H-5), 6.86–6.66 (m, 3H,
H-6, H3’, H-5’), 6.12 (d, J = 9.5 Hz, 1H, H-3), 3.70 (s, 3H,
-OMe). 13C NMR (75 MHz, CDCl3) dppm 192.85, 167.84,
162.22, 159.47, 155.44, 146.39, 144.28, 134.17, 131.05,
127.55, 123.42, 115.71, 114.59, 111.88, 111.01, 109.70, 55.41.
MS m/z (%) (ESI): 323 ([M + 1]+, 24), 322 ([M]+, 47), 321
(100), 293 (10), 134 (44). Anal. Calcd. for C19H14O5: C,
70.80; H, 4.38. Found: C, 70.77; H, 4.35.
Results
Structural identification
(E)-3-(3-(Thiophen-3-yl)acryloyl)coumarin (5): Yield 61%.
1
Mp 146–148°C H NMR (300 MHz, CDCl3) dppm 8.57 (s,
1H, H-4), 7.87 (d, J = 15.6 Hz, 1H, Hb), 7.74 (d, J = 15.7 Hz,
1H, Ha), 7.69–7.60 (m, 3H, H-2’, H-4’, H-5’), 7.43 -7.29
(m, 4H, H-5, H-6, H-7, H-8). 13C NMR (75 MHz, CDCl3)
dppm 186.5, 155.1, 148.1, 141.8, 138.4, 138.3, 134.2, 132.6,
130.7, 129.9, 126.9, 125.7, 124.9, 123.6, 118.6, 116.5. MS m/z
(%) (ESI): 283 ([M + 1]+, 8), 282 ([M]+, 35), 254 (100), 226
(73), 137 (88), 109 (93). Anal. Calcd. for C16H10O3S: C,
68.07; H, 3.57. Found: C, 68.11; H, 3.59
Binding affinity assays
The data obtained for the binding affinity assays from
radioligand binding experiments for compounds 5–10 are
Table 1 The binding affinity of compounds 5–10 for the human adenosine receptor subtypes hA1, hA2A and hA3
Ki (nM)
Compound
hA1
hA2A
hA3
5 160 (3 000–8 900)
5
6
7
>30 000
>10 000
>30 000
>10 000
>30 000
>10 000
>30 000
>10 000
>10 000
31 500 (21 600–46 000)
8
>10 000
9
10
8 330 (7 510–9 230)
11 900 (7 790–18 300)
5 020 (3 260–7 730)
20 200 (14 600–28 100)
>10 000
>10 000
The binding affinity (Ki) for the human AR subtypes hA1, hA2A and hA3 of compounds 5–10 was determined using radioligand competition experi-
ments in Chinese hamster ovary (CHO) cells. Highest concentrations tested were different depending on the solubility of compounds under the
respective assay conditions. Values are geometric means of three experiments and given with 95% confidence intervals in parentheses.
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
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Saleta Vazquez-Rodriguez et al.
Chalcone derivatives as hA3 receptors
Table 2 Theoretical structural properties of the coumarin–chalcone hybrids
Compound
logP
Molecular weight
TPSA (Å2)
n-OH acceptors
n-OHNH donors
Volume (Å3)
5
6
7
8
9
3.22
3.87
2.93
3.58
3.19
3.84
282.32
306.32
298.32
322.32
298.32
322.32
47.28
56.52
67.51
76.54
67.51
76.74
3
4
4
5
4
5
0
0
1
1
1
1
237.11
271.94
245.12
279.96
245.12
279.96
10
n-OH, number of hydrogen acceptors; n-OHNH, number of hydrogen bond donors; TPSA, topological polar surface area. The data was determined
with Molinspiration calculation software.
summarized in Table 1. Compound 9 showed binding affin-
ity to hA1 and hA2A ARs in the micromolar range, compara-
ble with the prototypical antagonist theophylline.[4,36]
Compounds 5 and 9 bound with affinity in the low micro-
molar range to hA3AR, with compound 5 being the one that
showed selectivity for this subtype.
exhibits similar affinity for A1 and A2A ARs whereas com-
pound 5 is devoid of measurable affinity for these subtypes.
The configuration with both A and B rings replaced seems
to be favourable for high A3 affinity and selectivity. Compar-
ing compounds 5, 7 and 9 suggests that the 4-hydroxyl
group in compound 7 and the corresponding 7-hydroxyl
group in compound 9, respectively, might abolish subtype
selectivity. Compound 9 presents a binding affinity for the
hA3 AR (Ki = 5020 nm) and also for hA1 AR (Ki = 8330 nm)
and hA2A AR (Ki = 11 900 nm). The A1 and A2A affinities are
thus comparable to the affinity of the classical naturally
occurring antagonist theophylline. [36]
Comparison of the structurally related compounds 5 and
7, reveals that the only difference between them is the
absence or presence of a hydroxyl group at position 4,
respectively. One can conclude that the presence of the
hydroxyl group seems to cause a marked decrease in the
binding affinity for the hA3 AR (Ki = 5160 and 31 500 nm,
respectively).
Comparing another pair of 8-substituted derivatives,
compounds 9 and 10, that differ in the substitution of ring
B of the chalcone (compound 10 has the original phenyl
ring of the chalcone while compound 9 presents a thiophe-
nyl ring as a result of an isosteric change), one can observe
that compound 9 shows affinity with no selectivity and
compound 10 is less potent at all three receptor subtypes.
Due to the limited affinity close to the detection limit the
degree of selectivity cannot be determined. From these pre-
liminary structure–activity relationships, one can conclude
that substitution of ring B of the chalcone (Figure 1) by a
thiophenyl ring (compounds 5, 7 and 9) favoured binding
affinity towards the AR. However, hA3 AR selectivity is
attained when ring B of the chalcone is substituted by a thi-
ophene ring and when ring A is substituted for the pyrone
ring contained in the coumarin moiety (compound 5).
On the other hand, looking at the theoretical evaluation
of ADME properties (Table 2), it can be observed that no
violations of Lipinski’s rule of five (molecular weight, logP,
number of hydrogen donors and acceptors) were found,
making these hybrid compounds promising leads for
drug candidates.[41] TPSA, described as being a predictive
Theoretical evaluation of ADME properties
To better correlate the drug-like properties of the
coumarin–chalcone hybrid compounds, the lipophilicity,
expressed as the octanol/water partition coefficient and
herein called logP, as well as other theoretical calculations
such as the TPSA, the number of hydrogen acceptors and
the number of hydrogen bond donors were calculated using
the Molinspiration property programme.[40] From the data
obtained, it was noticed that all the hybrid compounds,
5–10, not only had logP values compatible with those
required to cross membranes but also they did not break
any point of the Lipinski’s rule of five. Theoretical predic-
tion of ADME properties of all compounds is summarized
in Table 2.
Discussion
With the aim of finding novel and selective AR ligands, we
have synthesized chalcone- and coumarin-containing
hybrids in which one or both phenyl groups of the chalcone
scaffold either retains the two phenyl groups corresponding
to an original chalcone structure (rings A and B, Figure 1),
or is isosterically substituted by heteroaromatic rings
(pyrone ring of the coumarin and/or thiophene).
The data obtained in the binding affinity assays are
shown in Table 1. From all the synthesized derivatives, com-
pounds 5, 7 and 9 exhibit significant binding affinity in the
low micromolar range for one or more AR. The common
feature of compounds 5, 7 and 9 is the presentation of a thi-
ophenyl substituent as the key isosterical change in ring B of
the chalcone. However, only compound 5 displays a note-
worthy selectivity for the hA3 AR (Ki = 5160 nm). It is inter-
esting to note that compound 9, when compared with
compound 5, shows similar A3 affinity; however, it also
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
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Chalcone derivatives as hA3 receptors
Saleta Vazquez-Rodriguez et al.
indicator of the drug capacity of membrane penetration,
was also found to be positive.[42]
The remarkable results found for compounds 5 and 9
encourage us to continue our research looking for the opti-
mization of these lead compounds with the aim of obtain-
ing potent and selective chalcone-based hA3 AR ligands.
These findings encourage us to continue the efforts
towards the optimization of the pharmacological profile for
this type of hybrid as potent and selective ligands for the
hA3 ARs.
Declarations
Conclusions
Conflict of interest
Isosteric substitution of the phenyl rings of the chalcone
moiety for one or two heteroaromatic rings (benzopyrone
and/or thiophenyl) results in compounds (5 and 9) with
binding affinity towards particular subtypes of ARs. A pre-
liminary structure–activity relationship study of the synthe-
sized derivatives allowed verification that hA3 AR selectivity
was achieved only when ring B of the chalcone was substi-
tuted for a thiophene and ring A was substituted for the
pyrone ring included in the coumarin scaffold (compound
5). Replacing ring B by a thiophenyl group but keeping a
phenyl group in place of ring A, namely the benzene ring of
coumarin (compound 9), resulted in an increase of hA1 and
hA2A AR affinity. As binding affinity remained similar for
hA3 AR, selectivity was, therefore, lost.
The Author(s) declare(s) that they have no conflicts of
interest to disclose.
Funding
This work was partially supported by the Ministerio de
Sanidad y Consumo (PS09/00501) and Xunta de Galicia
(PGIDIT09CSA030203PR) (Spain) and by the Foundation
for Science and Technology (FCT) project PTDC/QUI-
QUI/113687/2009 (Portugal). S. Vazquez-Rodriguez thanks
the Ministerio de Educación y Ciencia for the PhD FPU
grant (AP2008-04263). M. J. Matos thanks Fundação para a
Ciência e Tecnologia for the PhD grant (SFRH/BD/61262/
2009).
receptor agonists. WO 9502604 A1 14. Lappas CM et al. Adenosine A2A ago-
References
19950126. Patent 1995.
8. Feoktistov et al. Adenosine A2B
nists in development for the treatment
of inflammation. Expert Opin Investig
Drugs 2005; 14: 797–806.
1. Poulsen SA, Quinn RJ. Adenosine
receptors: new opportunities for
future drugs. Bioorg Med Chem 1998;
6: 619–641.
I
receptors as therapeutic targets. Drug
Dev Res 1998; 45: 198–206.
15. Shah R, Frishman WH. Adenosine1
receptor antagonism: a new therapeu-
tic approach for the treatment of
decompensated heart failure. Cardiol
Rev 2009; 17: 125–131.
9. Jacobson KA. Adenosine A3 receptors:
novel ligands and paradoxical effects.
Trends Pharmacol Sci 1998; 19: 184–
195.
2. Fredholm BB et al. International
union of pharmacology. XXV. Nomen-
clature and classification of adenosine
receptors. J Pharmacol Rev 2001; 53: 10. Gessi S et al. Elevated expression of A3 16. Shook BC et al. Design and characteri-
527–552.
adenosine receptors in human color-
ectal cancer is reflected in peripheral
blood cells. Clin Cancer Res 2004; 10:
5895–5901.
zation of optimized adenosine
A2A/A1 receptor antagonists for
the treatment of Parkinson’s disease.
J Med Chem 2012; 55: 1402–1417.
3. Fredholm BB et al. Structure and
function of adenosine receptors and
their genes. Arch Pharmacol 2000; 362:
364–374.
11. Jacobson KA, Gao ZG. Adenosine 17. Stone TW et al. Adenosine receptors
4. Klotz K-N. Adenosine receptors and
their ligands. Naunyn Schmiedebergs
Arch Pharmacol 2000; 362: 382–391.
5. Baraldi PG et al. A3 adenosine recep-
tor ligands: history and perspectives.
Med Res Rev 2000; 20: 103–128.
6. Baraldi PG. Recent improvements in
the development of A2B adenosine
receptors as therapeutic targets. Nat
Rev Drug Discov 2006; 49: 247–264.
and neurological disease: neuroprotec-
tion and neurodegeneration. Handb
Exp Pharmacol 2009; 193: 535–597.
12. Kim
Y
et al. Novel 2- and
4-substituted
1H-imidazo[4,5- 18. Baraldi PG et al. Adenosine receptor
c]quinolin-4-amine derivatives as
allosteric modulators of the A3 adeno-
sine receptor. J Med Chem 2009; 52:
2098–2108.
antagonists: translating medicinal
chemistry and pharmacology into
clinical utility. Chem Rev 2008; 108:
238–263.
receptor agonists. Purinergic Signal 13. Sitkovsky MV et al. Physiological 19. Moro S et al. Progress in the pursuit of
2009; 5: 3–19.
control of immune response and
inflammatory tissue damage by
hypoxia-inducible factors and adenos-
therapeutic
adenosine
receptor
7. Jacobson KA et al. Preparation of
N6-benzyladenosine-5’-uronamides,
modified xanthine ribosides, and
related compounds as adenosine A3
antagonists. Med Res Rev 2006; 26:
131–159.
ine A2A receptors. Annu Rev Immunol 20. Okajima F et al. Genistein, an inhibi-
2004; 22: 657–682.
tor of protein tyrosine kinase, is
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
702

Saleta Vazquez-Rodriguez et al.
Chalcone derivatives as hA3 receptors
also
a
competitive antagonist for
Bioorg Med Chem Lett 2011; 21: 3342–
transfected receptors in CHO cells.
Naunyn Schmiedebergs Arch Pharma-
col 1998; 357: 1–9.
P1-purinergic (adenosine) receptor in
3345.
FRTL-5 thyroid cells. Biochem Biophys 29. Sashidhara KV et al. Synthesis and
Res Commun 1994; 203: 1488–1495.
21. Jacobson KA et al. Interactions of fla-
vones and other phytochemicals with
adenosine receptors. Adv Exp Med Biol
2002; 505: 163–171.
in vitro evaluation of novel coumarin- 37. Klotz KN et al. [3H]HEMADO-
a
novel tritiated agonist selective for the
human adenosine A3 receptor. Eur J
Pharmacol 2007; 556: 14–18.
chalcone hybrids as potential antican-
cer agents. Bioorg Med Chem Lett
2010; 20: 7205–7211.
30. Vukovic N et al. An efficient synthesis 38. Cheng YC et al. Relation between the
22. Borges F et al. Simple coumarins and
analogues in medicinal chemistry:
occurrence, synthesis and biological
activity. Curr Med Chem 2005; 12:
887–916.
and antioxidant properties of novel
imino and amino derivatives of
4-hydroxycoumarins. Arch Pharm Res
2010; 33: 5–15.
inhibition constant (Ki) and the con-
centration of inhibitor which causes
fifty per cent inhibition (I50) of an
enzymic reaction. Biochem Pharmacol
1973; 22: 3099–3108.
31. Vogel S et al. Natural and non-narural
23. Go ML et al. Chalcones: an update on
cytotoxic and chemoprotective prop-
erties. Curr Med Chem 2005; 12: 483–
495.
prenylated chalcones: synthesis, cyto- 39. De Lean
A
et al. Validation and
toxicity and anti-oxidative activity.
Bioorg Med Chem 2008; 16: 4286–
4293.
statistical analysis of computer
a
modeling method for quantitative
analysis of radioligand binding data
for mixtures of pharmacological
receptor subtypes. Mol Pharmacol
1982; 21: 5–16.
24. Nowakowska Z. A review of anti- 32. Chimenti F et al. Chalcones: a valid
infective and anti-inflammatory chal-
cones. Eur J Med Chem 2007; 42: 125–
137.
scaffold for monoamine oxidases
inhibitors. J Med Chem 2009; 52:
28218–22824.
40. M Cheminformatics,Bratislava, Slovak
Republic. 2009. http://www.molin
spiration.com/services/properties.
html
25. Bryskier A, Klich M. Coumarin antibi- 33. Liu XH et al. Synthesis and molecular
otics: novobiocin, coumermycin, and
clorobiocin. In: Bryskier A, ed. Antimi-
crob Agents Washington, DC, USA:
American Society for Microbiology Press
2005: 816–825.
docking study of novel coumarin
derivatives containing 4,5-
dihydropyrazole moiety as potential 41. Lipinski CA et al. Experimental and
antitumor agents. Biorg Med Chem
Lett 2010; 20: 5705–5708.
computational approaches to estimate
solubility and permeability in drug
discovery and development settings.
Adv Drug Deliv Rev 1997; 23: 3–25.
26. Viña D et al. 3-Substituted coumarins 34. Sugino T, Tanaka K. Solvent-free cou-
as dual inhibitors of AChE and MAO
for the treatment of Alzheimer’s
marin synthesis. Chem Lett 2001; 30:
110–111.
42. Ertl P et al. Fast calculation of
disease. Med Chem Comm 2012; 3: 35. Hamdi N et al. A rapid access to new
molecular polar surface area as a sum
of fragment-based contributions and
its application to the prediction of
213–218.
coumarinyl chalcone and substituted
chromeno[4,3-c]pyrazol-4(1H)-ones
and their antibacterial and DPPH
radical scavenging activities. Med
Chem Res 2011; 20: 522–530.
27. Matos MJ et al. Synthesis and study of
a SEries of 3-arylcoumarins as Potent
and selective monoamine oxidase B
inhibitors. J Med Chem 2011; 54:
7127–7137.
drug transport properties.
J
Med
Chem 2000; 43: 3714–3717.
36. Klotz KN et al. Comparative pharma-
cology of human adenosine receptor
subtypes – characterization of stably
28. Matos MJ et al. New halogenated phe-
nylcoumarins as tyrosinase inhibitors.
© 2013 The Authors. JPP © 2013
Royal Pharmaceutical Society 2013 Journal of Pharmacy and Pharmacology, 65, pp. 697–703
703