Novel thiazole-thiophene conjugates as adenosine receptor antagonists: Synthesis, biological evaluation and docking studies

Article abstract of DOI:10.1016/j.bmcl.2015.01.040

Here we report novel thiazole-thiophene conjugates as adenosine receptor antagonists. All the molecules were evaluated for their binding affinity for adenosine receptors. Most of the molecules were found to interact with the A₁, A_2A and A₃ adenosine receptor subtypes with good affinity values. The most potent and selective compound 8n showed an A₃ K_i value of 0.33 μM with selectivity ratios of >90 versus the A₁ and >30 versus the A₂ subtypes. For compound 8n docking studies into the binding site of the A₃ adenosine receptor are provided to visualize its binding mode.

Full text of DOI:10.1016/j.bmcl.2015.01.040

Accepted Manuscript
Novel thiazole-thiophene conjugates as adenosine receptor antagonists: Syn-
thesis, biological evaluation and docking studies
Dhaivat H. Pandya, Jayesh A. Sharma, Hitesh B. Jalani, Amit N. Pandya, V.
Sudarsanam, Sonja Kachler, Karl Norbert Klotz, Kamala K. Vasu
PII:
S0960-894X(15)00052-9
DOI:
http://dx.doi.org/10.1016/j.bmcl.2015.01.040
BMCL 22380
Reference:
To appear in:
Bioorganic & Medicinal Chemistry Letters
Received Date:
Revised Date:
Accepted Date:
28 October 2014
16 January 2015
20 January 2015
Please cite this article as: Pandya, D.H., Sharma, J.A., Jalani, H.B., Pandya, A.N., Sudarsanam, V., Kachler, S.,
Klotz, K.N., Vasu, K.K., Novel thiazole-thiophene conjugates as adenosine receptor antagonists: Synthesis,
biological evaluation and docking studies, Bioorganic & Medicinal Chemistry Letters (2015), doi: http://dx.doi.org/
1
0.1016/j.bmcl.2015.01.040
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Novel thiazole-thiophene conjugates as adenosine receptor antagonists:
Synthesis, biological evaluation and docking studies†
a¶
Dhaivat H. Pandya , Jayesh A. Sharma , Hitesh B. Jalani , Amit N. Pandya , V. Sudarsanam ,
a
a
a
a
b
b
a
Sonja Kachler , Karl Norbert Klotz , Kamala K. Vasu *
Department of Medicinal Chemistry, B. V. Patel Pharmaceutical Education and Research
Development (PERD) Centre, Sarkhej-Gandhinagar Highway, Thaltej, Ahmedabad-380 054,
Gujarat, India.
b
Institut für Pharmakologie und Toxikologie, Julius-Maximilians-Universität Würzburg,
Germany.
†
Communication Ref. No.: PERD250714
*
Corresponding author. Tel.: + 91 79 27439375; Fax: + 91 79 27450449
E-mail: kamalav@perdcentre.com
¶
Dhaivat H. Pandya is registered research scholar in Institute of Science, Nirma University.
Abstract: Here we report novel thiazole-thiophene conjugates as adenosine receptor antagonists.
All the molecules were evaluated for their binding affinity for adenosine receptors. Most of the
molecules were found to interact with the A
affinity values. The most potent and selective compound 8n showed an A
with selectivity ratios of > 90 versus the A and > 30 versus the A subtypes. For compound 8n
docking studies into the binding site of the A adenosine receptor are provided to visualize its
binding mode.
1
, A2A and A
3
adenosine receptor subtypes with good
3
K
i
value of 0.33 µM
1
2
3
Key Words: adenosine receptors, thiazoles, thiophenes, molecular docking
1

Adenosine regulates a variety of cellular functions through interaction with the four G-protein
coupled receptor (GPCR) subtypes A . The A and A adenosine receptors
couple to G and thereby inhibit adenylyl cyclase (AC) with a consequent decrease of cellular
cAMP levels. The A2A and A2B receptor couple to G and mediate a stimulation of AC resulting
1
, A2A, A2B and A
3
1
3
i
s
1
,2
in enhanced intracellular cAMP levels. Adenosine receptor subtypes are a target of great
interest because of their pathological involvement in numerous diseases. Activation of adenosine
receptors is beneficial in many conditions like epilepsy, pain, cancer, etc. whereas inhibition of
adenosine receptors is helpful in Parkinson’s disease, Alzheimer’s disease, asthma, diabetes and
1
,2,3
cancer.
Development of adenosine receptor antagonists is a major focus of medicinal chemistry since
several decades. Many selective antagonists have been developed so far and are helpful in
pathological conditions like renal failure (A
1
), Parkinson’s disease (A2A), diabetes (A2B), asthma
), glaucoma and cancer (A ). The
and chronic obstructive pulmonary disease COPD (A2B/A
3
3
traditional xanthine derivatives like caffeine and theophylline are naturally occurring
nonselective antagonists. Selective antagonists have been developed by modifying the xanthine
4
or adenine scaffolds, or heterocyclic moieties with mono-, bi- or tricyclic ring systems. Several
of them are in preclinical or clinical trials and the selective A2A receptor antagonist Istradefylline
got approval in Japan for the treatment of Parkinson’s disease (Figure 1). The major problem
associated with xanthine derivatives is related to pharmacokinetic issues. Consequently, there is
a need for new non-xanthine molecules. Structures incorporating thiazole, thiophene and
5
–7
benzothiazinones have been developed as selective adenosine receptor antagonists.
INSERT FIGURE-1 HERE
As published in many reports, aminothiazoles and aminothiophenes are two important structural
elements in adenosine receptor antagonists. Such 2-aminothiophene substituents are also found
8
in modulation of adenosine receptors. Based on similar observations, Aurelio et al published 2-
9
aminothiophene derivatives as adenosine receptor modulators . Aminothiazole compounds as
adenosine receptor antagonists have been reported from our group to show high affinity and
5
selectivity. So keeping in mind the importance of both moieties, i.e. thiophene and thiazole,
,7
here we have designed novel thiazole-thiophene conjugates with an amide spacer (Figure-2).
2

INSERT FIGURE-2 HERE
The synthesis of thiazole-thiophenes (8a-8r, Table-1) was carried out by a four-step reaction.
The respective starting derivatives of 2-chloroacetamidothiophene and amidinothiourea were
prepared separately. The reaction between 2-chloroacetamidothiophenes (4) and
amidinothioureas (7) was carried out to get the final thiazole-thiophene conjugate compounds
(
Scheme-1).
For the preparation of 2-chloroacetamidothiophene, 2-aminothiophenes (3) were prepared by the
1
Gewald reaction utilizing various carbonyl compounds (1), active methylene nitriles (2) and
0
elemental sulphur in presence of morpholine as a base. This 2-aminothiophene (3) was further
0
reacted with chloroacetyl chloride in THF with triethyl amine as a base at 0-5 C to furnish 2-
chloroacetamidothiophene derivatives (4a-d, Scheme-1).
Our group generated many novel biologically important ligands from the intermediate
1
1–13
amidinothiourea (7).
Various isothiocyanates were reacted with different amidine derivatives
in THF to give the amidinothiourea (7a-h, Scheme-1).
The amidinothiourea was reacted with 2-chloroacetamidothiophenes in DMF to give the final
thiazolyl-thiophenes (8a-r, Table-1). The reaction takes place by the S-alkylation of
amidinothiourea followed by the 5-exo-trig cyclization and removal of secondary amine to
furnish the desired thiazolyl-thiophenes (Scheme-1).
INSERT SCHEME HERE
All the molecules were screened for their binding affinity for human adenosine receptor subtypes
1
4,15
by radioligand binding at hA
Interestingly, 15 out of 18 synthesized thiazole-thiophene conjugates showed micromolar or sub
micromolar affinity towards hA adenosine receptors. None of the compounds showed
measurable interaction with the hA2B subtype (K > 10 µM, not shown). Compounds 8c, 8d and
2.2, 0.52, 0.33 µM, respectively) and high selectivity (≥ 14-
adenosine receptor (Table 1). Compound 8k also showed high A
1 3
, hA2A, hA as previously described by Klotz et al.
3
i
8
n showed the highest affinity (K
i
9
0fold) towards the hA
3
3
3

affinity but was not very selective (≤ tenfold) due to significant affinity for the A
subtypes (K 2.3 and 3.4 µM, respectively) (Table-1). The three compounds 8c, 8d and 8n with
the highest A affinity and selectivity were characterized by the presence of a 4-methylphenyl or
-methoxyphenyl group at R and R position. These substituents might be responsible for good
1
and A2A
i
3
4
5
4
4
3
A affinity and selectivity. Replacement of 4-methylphenyl in R position for an unsubstituted
phenyl resulted invariably in nonselective compounds (e.g. 8a, 8e, 8k) (Table-1). Also, a
5
dimethylamino group in R position produced nonselective ligands (Table-1).
INSERT TABLE-1 HERE
To see the possible interactions of the newly synthesized ligands with the A
flexible docking studies into an A homology model (PDB id: 1OEA) were carried out in
Surflex-Dock, SYBYL X-2.0. The three A -selective compounds 8c, 8d and 8n, and the
3
adenosine receptor
3
3
nonselective ligand 8k, showing the highest affinity, were investigated. All four molecules were
found to interact with the receptor (Supplymentary figure S1) with the amino acids ASN250,
PHE168, GLN167 and SER170 through hydrogen bonding. The common interaction site of the
four active molecules with the receptor was the amide spacer present in the molecule. All of
these four molecules were found to interact through hydrogen bonding to the amino acid
1
ASN250 which is reported to be responsible for antagonist interaction. In the most potent
6
molecule 8n, the carbonyl oxygen of –CONH present on the 3-position of the thiophene ring
2
interacted with a hydrogen of the ASN250 amido group. The carbonyl oxygen of the amide
linker present between the thiazole and thiophene has shown three interactions, namely with the
second amido hydrogen of ASN250 and with the two amido hydrogens of GLN167. The amine
nd
spacer hydrogen present of the 2 position of thiazole ring was found to interact with the
hydroxyl oxygen of the SER170. Figure-3 shows the highest consensus score (CScore = 4) pose
of the molecule 8n in the cavity of the ligand binding site. The study documents interaction of
3
the new compounds with ligand binding domain of the A receptor and also shows the important
role of the amide spacer between the two ring systems which may be responsible for good
antagonist affinity.
INSERT FIGURE 3 HERE
4

In conclusion, novel thiazole-thiophene conjugates were designed, synthesized, characterized
and evaluated for their binding to adenosine receptors. Most of the novel compounds synthesized
were found to interact with adenosine receptors showing that the chosen thiazole-thiophene
conjugate structure is favorable for the interaction with adenosine recpetors. In particular, some
were found to bind with high affinity and selectivity to the A
potent and selective molecule 8n bound to the A subtype with a K
compounds revealing the highest A affinity were docked into a homology model of the A
3
adenosine receptor. The most
3
i
-value of 0.33 µM. The four
3
3
adenosine receptor confirming their similar interaction with the ligand binding domain.
Acknowledgement
We would like to acknowledge B.V. Patel PERD Centre, for providing research facilities. We
also thank Industrial Commissioner (IC) of Gujarat and Indian Council of Medical Research
(
ICMR) for their financial assistance. We thank Prof. C. J. Shishoo and Dr. Manish Nivsarkar,
Directors, B.V. Patel PERD Centre, for their constant encouragement and support.
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2
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Fredholm, B. B.; IJzerman, A. P.; Jacobson, K. A.; Linden, J.; Müller, C. E. Pharmacol.
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3
4
5
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Chen, J.-F.; Eltzschig, H. K.; Fredholm, B. B. Nat. Rev. Drug Discov. 2013, 12, 265–286.
Müller, C. E.; Jacobson, K. A. Biochim. Biophys. Acta 2011, 1808, 1290–1308.
Scheiff, A. B.; Yerande, S. G.; El-Tayeb, A.; Li, W.; Inamdar, G. S.; Vasu, K. K.;
Sudarsanam, V.; Müller, C. E. Bioorg. Med. Chem. 2010, 18, 2195–2203.
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8
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Inamdar, G. S.; Pandya, A. N.; Thakar, H. M.; Sudarsanam, V.; Kachler, S.; Sabbadin, D.;
Moro, S.; Klotz, K.-N.; Vasu, K. K. Eur. J. Med. Chem. 2013, 63, 924–934.
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5

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1
1
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7
220–7222.
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1
1
1
1
2. Giri, R. S.; Thaker, H. M.; Giordano, T.; Chen, B.; Nuthalapaty, S.; Vasu, K. K.;
Sudarsanam, V. Eur. J. Med. Chem. 2010, 45, 3558–3563.
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6

Figure 3: The most active molecule 8n in the active site of adenosine A receptor showing
3
hydrogen interactions (PDB Id: 1OEA)

i
Table 1: Binding affinity (K ) of synthesized compounds at adenosine receptors with selectivity
K
i
(µM)
Selectivity
/hA hA2A/hA
3
1
R
2
R
3
R
4
R
5
R
No
a
b
c
hA
1
hA2A
hA
3
hA
1
3
8
a
b
c
d
e
f
40.8
36.2 – 45.9) (3.91 – 10.6) (3.94 – 4.32)
6.45
4.12
-
-
-
-
-
-
-
-
-
-
-
(CH
(CH
(CH
(CH
(CH
(CH
(CH
(CH
(CH
(CH
(CH
2
2
2
2
2
2
2
2
2
2
2
)
4-
)
4-
)
4-
)
4-
)
4-
)
4-
)
4-
)
4-
)
4-
)
4-
)
4-
-COOEt
-COOEt
-COOEt
C
6
H
4
-N(Me)
Me
2
9.9
> 3.7
> 14
> 19
1.9
1.6
> 3.7
> 14
> 19
6.9
(
8
8
8
8
8
8
8
8
8
8
8.22
6.96 – 9.70)
4-MeC
6
6
H
4
> 30
> 30
> 30
> 30
(
2.16
1.25 – 3.73)
4-MeC
H
4
4-MeC
4-MeC
4-MeC
6
6
6
H
4
(
0.520
347 – 778)
-COOEt 4-OMe C
-COOEt
-COOEt 4-Me C
6
H
4
H
4
> 10
7.75
> 10
28.4
(
4.09
C
6
H
4
H
4
(
6.42 – 9.35) (17.1 – 47.2) (2.28 – 7.33)
23.5 18.0 4.71
18.8 – 29.3) (13.6 – 12.8) (3.91 – 5.67)
6
H
4
-N(Me)
-N(Me)
-N(Me)
-N(Me)
-N(Me)
2
2
2
2
2
5.0
3.8
(
g
h
i
-COOEt
-CN
CH
3
OCO
> 100
> 30
> 100
> 30
> 30
> 3.3
> 0.84
> 2.9
4.5
> 3.3
> 0.84
> 0.29
3.1
35.6
25.3 – 50.2)
6 4
C H
(
34.0
30.4 – 38.0)
-CN
4-MeC
6
6
H
4
> 100
32.4
> 10
22.3
(
j
7.13
-CONH
-CONH
2
2
4-MeC
H
4
(
21.4 – 49.3) (15.8 – 31.5) (6.40 – 7.95)
2.38 3.44 0.334
1.68 – 3.38) (2.10 – 5.62) (236 – 473)
k
C
6
H
4
4-MeC
6
H
4
7.1
10
(

8
8
8
8
8
8
8
l
-
-
-
-
(CH
(CH
(CH
(CH
2
2
2
2
)
)
)
)
4-
4-
4-
4-
-CN
-CN
4-MeC
6
H
4
4-MeC
4-MeC
4-MeC
Me
6
6
6
H
4
> 10
> 30
> 30
> 10
> 30
> 10
-
-
m
n
o
p
q
r
9.56
6.05 – 15.1)
4-OMeC
6
H
4
H
4
> 3.1
> 90
> 7.8
5.1
> 3.1
> 30
1.5
(
0.33
239 – 456)
-CONH
-CN
2
4-MeC
6
6
H
4
H
4
> 10
5.67
(
3.87
4-MeC
H
4
> 30
14.0
(
5.01 – 6.41) (3.01 – 4.99)
9.66 2.73
-
-
-
COOMe Me -COOEt 4-OMeC
6
H
4
-N(Me)
Me
2
2
3.5
(
12.0 – 16.5) (7.30 – 12.8) (2.11 – 3.55)
COOMe Me -COOEt
COOMe Me -COOEt
4-MeC
6
6
H
4
> 30
19.0
> 30
26.6
>30
-
0.33
6.8
3.91
4-MeC
H
4
-N(Me)
4.9
(
15.4 – 23.6) (19.8 – 35.9) (2.98 – 5.11)
Data are expressed as geometric means with 95% confidence intervals in parentheses.
a
3
Displacement of specific [ H]CCPA binding at human A
1
receptors expressed in CHO cells.
Displacement of specific [ H]NECA binding at human A2A receptors expressed in CHO cells.
b
3
c
3
3
Displacement of specific [ H]HEMADO binding at human A receptors expressed in CHO cells.