Synthesis and adenosine receptor binding studies of some novel triazolothienopyrimidines

Article abstract of DOI:10.1016/j.ejmech.2007.05.001

A new series of 5-alkyl/aryl-8,9-dimethyl/8,9,10,11-tetrahydro[1]benzothieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thiones (4a-k) have been synthesized through a facile cyclization reaction of 4-hydrazino-2-alkyl/aryl-5,6-dimethyl/5,6,7,8-tetrahydr

Full text of DOI:10.1016/j.ejmech.2007.05.001

Available online at www.sciencedirect.com
European Journal of Medicinal Chemistry 43 (2008) 614e620
http://www.elsevier.com/locate/ejmech
Original article
Synthesis and adenosine receptor binding studies of some
novel triazolothienopyrimidines
b,2
Mailavaram Raghu Prasad a, , Akkinepally Raghuram Rao , Pamulaparthy Shanthan Rao
a,1
,
*
Kombu Subramanian Rajan a,3, Shanmugam Meena a,4, Kuchana Madhavi a,5
a Medicinal Chemistry Research Division, University College of Pharmaceutical Sciences, Kakatiya University, Warangal 506 009, AP, India
b Indian Institute of Chemical Technology, Tarnaka, Hyderabad, AP, India
Received 7 October 2006; received in revised form 30 April 2007; accepted 1 May 2007
Available online 21 May 2007
Abstract
A new series of 5-alkyl/aryl-8,9-dimethyl/8,9,10,11-tetrahydro[1]benzothieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine-3(2H )-thiones (4aek)
have been synthesized through a facile cyclization reaction of 4-hydrazino-2-alkyl/aryl-5,6-dimethyl/5,6,7,8-tetrahydro[1]benzothieno
[2,3-d]pyrimidines (3aek) using carbon disulphide under basic conditions. 4-Hydrazino-2-alkyl/aryl-5,6-dimethyl/5,6,7,8-tetrahydro[1]benzo-
thieno[2,3-d]pyrimidines (3aek) were prepared by replacing the chloro group of 4-chloro-2-substituted-5,6-dimethyl/5,6,7,8-tetrahydro[1]ben-
zothieno[2,3-d]pyrimidines (2aek) with hydrazine hydrate which were obtained by a known one-pot synthesis. The affinities of these
compounds for adenosine A1/A2A receptors were determined at 1 mM concentration. The test compounds which exhibited more than 20%
inhibition were selected and further screened at six different concentration levels to estimate their EC50/Ki values. The most potent compounds
in the series were 4c and 4d having an ethyl side chain at C5 position with dimethyl and cyclohexyl substitution at the C8eC9 positions, ex-
hibiting Ki values of 2.1 and 1.1 mM, respectively, at A1ARs. The SAR indicates that by increasing or decreasing the alkyl chain length at
C5 led to reduced affinity. The remaining aryl/arylalkyl derivatives of the series were inactive showing that a simple alkyl side chain at C5
is necessary for these ligands to bind at A1ARs. However, none of the compounds showed inhibition on A2A receptors at 1 mM concentration
indicating their selectivity. This communication describes the design, synthesis and evaluation of these new molecules.
Ó 2007 Elsevier Masson SAS. All rights reserved.
Keywords: Synthesis; Triazolopyrimidines; Adenosine A1 receptor antagonists; Triazolothienopyrimidines; Radioligand binding assays
1. Introduction
* Corresponding author. P.E.S. College of Pharmacy, Hanumanthnagar, Ban-
Adenosine is an endogenous neuromodulator which medi-
galore 560 050, Karnataka, India. Tel.: þ91 80 26507428 (O)/þ91 80
26691255 (R); fax: þ91 80 26507428; mobile: þ91 9448435886.
E-mail addresses: raghumrp@mailcity.com (M. Raghu Prasad),
raghumed@gmail.com (A. Raghuram Rao).
ates its biological effects by interacting with four adenosine
receptor (AR) subtypes namely A1, A2A, A2B and A3 identified
and cloned from various mammalian tissues [1e3]. Adenosine
A1 receptors (A1AR) play significant role in a number of var-
ied biological effects. These receptors are found both pre- and
post-synaptically and are involved in the sedative, anxiolytic
and locomotor-depressant effects of adenosine in the central
nervous system [4e7]. Abundant evidence reveals an impor-
tant role for A1ARs in protection of heart [8e10], lung and
brain [11] from ischemia reperfusion injury. Like A1 receptors,
the A2A receptors have been implicated in a wide range of
1
Present address: Medicinal Chemistry Division, University Institute of
Pharmaceutical Sciences, Panjab University, Chandigarh 160 014, India. Tel.:
þ91 172 253 4111 (O); fax: þ91 172 254 1142; mobile: þ91 9815846064.
2
Present address: BASR Chemicals Limited, Hyderabad, Andhra Pradesh,
India.
3
Present address: Case Western Reserve University, 2109 Adelbert Road,
School of Medicine, BRB 901 Cleveland, OH, USA.
4
Present address: K.M. College of Pharmacy, Madurai, Tamil Nadu, India.
Present address: Institute of Pharmacy, Sri Padmavati Mahila University,
Tirupathi, Andhra Pradesh, India.
5
0223-5234/$ - see front matter Ó 2007 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2007.05.001

M. Raghu Prasad et al. / European Journal of Medicinal Chemistry 43 (2008) 614e620
615
biological events. Because of the interaction between dopamine
and A2A receptors and as these receptors are primarily ex-
pressed in striatum, antagonists of A2A receptors have potential
therapeutic utility in alleviating Parkinson’s disease [12].
Despite a large number of potential therapeutic applica-
tions, relatively few compounds entered clinical trials
[13,14]. This could be due to a poor pharmacokinetic profile
of some of the known AR antagonists, including low water
solubility and/or low central nervous system penetration. As
a consequence, a lot of interest has been generated in recent
years to find ligands of ARs with high degree of selectivity.
A large number of studies are aimed at designing new ligands
which are free from undesirable side effects.
proposed structures have three lipophilic pockets (one of
which can be filled better with cycloalkyl moiety) along
with hydrogen donoreacceptor motif (Fig. 4) as per proposed
pharmacophore model for A1AR binding [26]. Hence it was
felt worthwhile to synthesize a novel series of 5-alkyl/aryl-
8,9-dimethyl/8,9,10,11-tetrahydro[1]benzothieno[3,2-e][1,2,4]
triazolo[4,3-c]pyrimidine-3(2H)-thiones (4aek) as potential
A1AR antagonists.
2. Results and discussion
2.1. Chemistry
The naturally occurring xanthines, caffeine and theophyl-
line are the prototypic AR antagonists. Several attempts
have been made to improve the potency and selectivity of xan-
thine-based compounds which resulted in the synthesis and
screening of a large number of new analogues. Of them,
DPCPX and KW3802 (Fig. 1) are the standard xanthine
A1AR antagonists. However, these compounds failed in clini-
cal trials as they lack acceptable pharmacokinetic profiles.
This gave an impetus for the design of new xanthine-based,
non-xanthine heterocyclic analogues and test their efficacy
and selectivity towards ARs. Literature reveals that a wide va-
riety of nitrogen containing heterocycles have been synthesized
as possibleA1/A2A selectiveantagonists (A1AR) [15,16]. Partic-
ularly some tricyclic heteroaromatic (six-six-five ring systems)
compounds (Fig. 2) like triazolo[1,5-c]quinazoline, tria-
zolo[4,3-a]quinoxaline and imidazo[4,5-c]quinoline [17e20]
derivativeswere shown to be selectiveligands with potent antag-
onistic activity at A1ARs. They were designed as agents with
potential therapeutic utility as cognitive enhancers, agents
with antidementia properties, as psychostimulants, antide-
pressant drugs and ameliorants of cerebral function [21].
Several of these A1AR antagonists were demonstrated with
promising therapeutic potential for renal and cardiac failure
[22,23]. Sarges et al. suggested that CP-68247 [24], a tria-
zolo[4,3-a]quinoxaline derivative binds to ARs, by mimick-
ing adenosine derivatives (Fig. 3).
The synthetic route to the target compounds is outlined in the
Scheme 1. 2-Amino-3-carbethoxy-4,5-dimethylthiophene, 2-
amino-3-carbethoxy-4,5,6,7-tetrahydro[1]benzothiophene [27,
28], 4-chloro-2-substituted-5,6-dimethylthieno[2,3-d]pyrimi-
dines and 4-chloro-2-substituted-5,6,7,8-tetrahydro[1]benzo-
thieno[2,3-d]pyrimidines 2aek [29] were prepared following
reported procedures. The chlorine atom in 2aek was replaced
by hydrazine by treating 2aek with hydrazine hydrate (99%)
to yield 4-hydrazino-2-alkyl/aryl-5,6-dimethylthieno[2,3-d]
pyrimidines and 4-hydrazino-2-alkyl/aryl-5,6,7,8-tetrahy-
dro[1]benzothieno[2,3-d]pyrimidines 3aek in good yields
[30]. The prominent peaks (3400e3200 cmꢀ1) for primary
and secondary amino groups in the IR spectra indicated the for-
mation of the expected product. The 1H NMR spectrum showed
a triplet at d 6.4e6.6 ppm and a doublet at d 3.8 ppm for NH and
NH2 groups and the presence of characteristic signals for di-
methyl and cyclohexyl groups confirmed the product formation.
Furthermore, the mass spectrum showed a prominent molecular
ion peak [Mþ] as the base peak and fragmentation pattern char-
acteristic to its structure. The absence of an isotopic peak
(Mþ þ 2) also confirmed the replacement of the chloro group
in the product.
Compounds 3aek were condensed with carbon disulphide
under basic conditions using ethanol as solvent to yield the
title compounds. The products thus obtained were purified
and characterized as 5-alkyl/aryl-8,9-dimethylthieno[3,2-
e][1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thiones and 5-alkyl/
aryl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e][1,2,4]triazolo
[4,3-c]pyrimidine-3(2H)-thiones 4aek on the basis of their
analytical and spectral data. The IR spectra showed character-
istic peaks (C]S) at 1200 cmꢀ1 [weak] and 3090 cmꢀ1 (sec-
amino group). The disappearance of the primary amino proton
signal and the appearance of the secondary amino signal at
d 14.0 in PMR spectrum indicated the product formation. Fur-
thermore, the mass spectrum showed molecular ion peak [Mþ]
as the base peak and fragmentation pattern characteristic to its
structure. The elemental analyses of all the newly synthesized
compounds confirmed their formation.
Further various triazolopyrazolopyrimidines (five-six-five
ring systems) [25] have been explored for affinity towards
ARs, which showed better affinity/selectivity. These observa-
tions led us to design these novel molecules by replacing pyr-
azole ring from its bioisosteric thiophene which resulted in
triazolothienopyrimidines. These were also found to be bioi-
sosteres of triazoloquinoxalines in a fused system. The
O
H3C
H
N
O
N
H3C
H
N
N
N
N
N
O
N
O
2.2. Pharmacology
C3H
C3H
DPCPX
2.2.1. Radioligand binding studies
KW3902
Thesynthesizedcompounds,4aekweretestedfortheirability
to displace [3H]2-chloro-N6-cyclohexyladenosine ([3H]CCPA)
Fig. 1. Standard xanthine-based adenosine A1 receptor antagonists.

616
M. Raghu Prasad et al. / European Journal of Medicinal Chemistry 43 (2008) 614e620
R2
R2
R2
H
N
N
N
N
R3
N
N
N
N
R3
R3
N
NHR1
NHR1
NHR1
N
N
Triazolo[1,5-c]quinazolines
Triazolo[4,3-a]quinoxalines
Triazolo[ 4,5-c]quinolines
Fig. 2. Basic structures of representative tricyclic xanthine-based non-xanthine adenosine antagonists.
from A1ARs and [3H]MSX-2 from A2AAR in rat cortical and
striatal membranes, respectively. The A1 and A2A receptor bind-
ing affinitiesfor compounds 4aek are expressedasKi values orin
percent inhibition of radioligand binding (at 1.0 mM concentra-
tion). The binding affinities data of compounds 4aek are pre-
sented in Table 1.
this study reveals that the compounds presently synthesized,
although weakly active at A1ARs, are fairly selective. Further
optimization may potentially result in better affinity.
3. Conclusion
None of the compounds showed affinity towards A2AARs at
1.0 mM concentration. This gives a clue that the compounds fit
fairly well into the proposed pharmacophoric model for
A1AR’s.
Alkyl derivatives of the series (4b, 4c, 4d) showed affinity
towards A1ARs i.e., the percentage of inhibition at 1.0 mM
concentration was found to be 26, 21 and 56%, respectively,
whereas the aryl and arylalkyl derivatives did not bind at the
receptor site. This may be due to the increase in bulkiness
of the lipophilic site which may result in lesser interaction.
The results are in agreement with the fact that DPCPX, the ar-
chetypical A1AR antagonist also has alkyl side chains in its
structure (Fig. 1).
Those compounds which showed 20% inhibition were fur-
ther screened at six different concentrations ranging from
0.01 mMe10 mM to determine their EC50 and Ki values. The
most potent compounds in the series were 4c and 4d having
an ethyl side chain at the C5 position with dimethyl and cyclo-
hexyl substitution at the C8eC9 positions, exhibiting Ki values
of 2.1 and 1.1 mM, respectively, for A1ARs.
Of the alkyl derivatives (methyl, ethyl and propyl deriva-
tives), ethyl derivatives, 4c and 4d showed inhibition at micro-
molar ranges. Increasing or decreasing the length of the side
chain (replacement of ethyl group by methyl or propyl) at
C5 led to reduced affinity indicating ethyl group at C5 position
as optimal structural requirement for the compounds to bind to
A1ARs.
In conclusion, we designed a set of novel triazolothienopyr-
imidines based on the adenosine A1 receptor pharmacophoric
model. The designed compounds were successfully synthe-
sized in quantitative yields. The compounds were screened
for their affinity towards A1 and A2A receptors and showed
no affinity towards A2AARs. This indicates the validity of
our design as they fit well in the proposed pharmacophoric
model. Alkyl derivatives showed affinity at micromolar ranges
towards A1ARs out of which, ethyl derivative with a fused cy-
clohexyl ring system at C8e9 showed highest affinity with se-
lectivity towards A1ARs. Hence it is worthwhile to attempt at
fusing triazolothienopyrimidines with bulky cycloalkyl rings
and to evaluate them for affinity. Further efforts are currently
being taken up to optimize the lead structure and the results of
which will be the basis of our future research endeavor.
4. Experimental
Analytical TLC was performed on Silica Gel F254 plates
(Merck) with visualization by UV (254 nm) chamber with pro-
tective filters (Wiswo Instruments, Mumbai). Melting points
were determined in open capillaries on a Gallenkemp Melting
Point Apparatus and are uncorrected. The IR spectra (KBr,
ymax, cmꢀ1) were run on Perkin Elmer FTIR Spectrophotom-
eter. Proton (1H) NMR spectra were recorded on a AMX-
400 and chemical shifts are expressed as d values (ppm)
downfield from tetramethylsilane (TMS) using either CDCl3
Out of the 8,9,10,11-terahydrobenzo series (4b, 4d, 4f, 4h,
4j, 4k) and 8,9-dimethyl series (4a, 4c, 4e, 4g, 4i) the former
showed more affinity towards A1ARs which is in agreement
with the fact that one of the lipophilic pockets when filled
with cycloalkyl moiety increases the affinity. As a whole
R1
O
H3C
H
S
H
N
N
N
N
R2
F3C
Ribose
N
N
N
N
S
N
N
N
O
N
N
N
N
Cl
R3
CH3
N
N
N
H
N6-cyclopentyladenosine
Proposed Structure
DPCPX
H
CP-68247
Fig. 4. Comparison of proposed structure with published pharmacophore
model (DPCPX) for A1 receptor antagonists. Dashed arrows indicate hydrogen
bond acceptoredonor motifs; dashed squares indicate lipophilic pockets.
Fig. 3. Putative binding mode of CP-68247: overlap with N6-
cyclopentyladenosine.

M. Raghu Prasad et al. / European Journal of Medicinal Chemistry 43 (2008) 614e620
Cl
617
O
R1
R2
COOEt
R1
R1
N-H
N
POCl3
RCN/HCl
R3
R3
N
N
NH2
S
R2
N
S
S
R2
[1]
[2a-k]
H
N
H2NHN
S
S
R1
R1
R2
N
N
CS2/KOH/EtOH
N2H4H2O
R3
R3
N
N
S
R2
[4a-k]
[3a-k]
R
R
R
R
R
3
R
1
2
3
2
1
Pr
4a Me
4b -(CH2)4-
4c Me
4d -(CH2)4-
Me
Me
Me
Et
Et
Pr
4f -(CH2)4-
4g Me Me
4h -(CH2)4-
4i Me Me
4j -(CH2)4-
Ph
Ph
Bz
Bz
Me
4e Me
Me
4k -(CH2)4- 4-Pyridyl
Scheme 1. Synthesis of some novel 5-alkyl/aryl-8,9-dimethyl/8,9,10,11-tetrahydro[1]benzothieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine-3(2H )-thiones.
or DMSO-d6 as solvents. Mass [EI-MS (70 eV)] spectra were
recorded on Autospec Mass spectrometer. Elemental analyses
were performed on Carlo Erba 1108 elemental analyzer. All
the chemicals were obtained from E. Merck (Mumbai) of an-
alytical grade, and were used without further purification.
Graph Pad PRISM [Version 3.0 from San Diego, CA) was
used for biological data analyses.
poured onto crushed ice. The solid precipitate thus obtained
was filtered, dried and recrystallized from ethanol (95%).
4.1.1.1. 4-Hydrazino-2-ethyl-5,6-dimethylthieno[2,3-d]pyrimi-
dine (3c). M.p. 109e110 ꢁC; yield 71%; IR (KBr) ymax
:
3190, 3320 (NH2), 2910 (CH3CH2) cmꢀ1; H NMR (CDCl3)
d: 1.2e1.4 (t, 3H, J ¼ 8.7 Hz, CH2CH3), 2.42 (s, 6H, CH3),
3.35e3.40 (q, 2H, J ¼ 8.7 Hz, CH2CH3), 3.65e3.85 (d, 2H,
J ¼ 5.6 Hz, NHNH2, D2O exchangeable), 6.5e6.7 (t, 1H,
J ¼ 5.7 Hz, NHNH2, D2O exchangeable) ppm. EI-MS [m/z,
%]: 222 [Mþ 100]. Anal. Calcd for C10H14N4S: C, 54.05; H,
6.30; N, 25.22. Found: C, 54.25; H, 6.55; N, 25.52.
1
4.1. Chemistry
4.1.1. General procedure for the preparation of
4-hydrazino-2-alkyl/aryl-5,6-dimethyl/5,6,7,8-
tetrahydro[1]benzothieno[2,3-d]pyrimidines (3aek)
To a warm solution of 2-alkyl/aryl-4-chloro-5,6-dimethyl/
5,6,7,8-tetrahydrobenzo[b]thieno[2,3-d]pyrimidines [10 mmol]
(2aek)] in ethanol (95%, 20 ml) was added a solution of hydra-
zine hydrate [99%, 4.3 g, 100 mmol] dropwise and heated un-
der reflux for 2 h. Then the reaction mixture was cooled and
4.1.1.2. 4-Hydrazino-2-ethyl-5,6,7,8-tetrahydro[1]benzothieno
[2,3-d]pyrimidine (3d). M.p. 159e160 ꢁC; yield 75%; IR
(KBr) ymax: 3190, 3320 (NH2), 2910 (CH3CH2) cmꢀ1 1H
;
NMR (CDCl3) d: 1.2e1.4 (t, 3H, J ¼ 7.9 Hz, CH3), 1.8e2.0
(m, 4H, CH2 at 6 and 7), 2.7e3.0 (m, 6H, eCH2 at 2, 5 and
8), 3.6e3.8 (d. 2H, J ¼ 5.79 Hz, eNH2, D2O exchangeable),
6.4e6.6 (t, 1H, J ¼ 6.0 Hz, NH, D2O exchangeable) ppm; EI-
MS [m/z, %]: 248 [Mþ 100]. Anal. Calcd for C12H16N4S: C,
58.06; H, 6.45; N, 22.58. Found: C, 58.35; H, 6.85; N, 22.88.
Table 1
Binding affinity of compounds 4aek at rat A1 and A2A adenosine receptors
Compd. no.
R1
R2
R3
A1ARa %
Ki ꢂ SEM
mM
inhibition at
1.0 mM conc.
4.1.1.3. 4-Hydrazino-2-propyl-5,6-dimethylthieno[2,3-d]pyrimi-
dine (3e). M.p. 102e103 ꢁC; yield 77%; IR (KBr) ymax
:
4a
4b
4c
Me
e(CH2)4e
Me
Me
Me
Et
4
26
21
56
13
0
e
3190, 3320 (NH2), 2910 (alk) cmꢀ1; H NMR (CDCl3,) d:
1.10e1.12 (t, 3H, J ¼ 11.1 Hz, CH3), 2.25 (s, 6H, CH3),
2.42e2.52 (m, 2H, eCH2CH2CH3), 3.01e3.05 (m, 2H,
eCH2CH2CH3), 3.62e3.82 (d. 2H, J ¼ 5.8 Hz, eNHNH2,
D2O exchangeable), 6.6e6.8 (t, 1H, J ¼ 6.0 Hz, NHNH2, D2O
exchangeable) ppm; EI-MS [m/z, %]: 236 [Mþ 100]. Anal.
Calcd for C11H16N4S: C, 55.93; H, 6.77; N, 23.72. Found: C,
56.13; H, 7.13; N, 24.10.
1
4.40 ꢂ 1.1
Me Me
e(CH2)4e
2.10 ꢂ 0.4
4d
4e
Et
1.17 ꢂ 0.4
Me Me
e(CH2)4e
Pr
e
e
4f
Pr
Ph
4g
4h
4i
Me Me
e(CH2)4e
0
e
Ph
Bz
0
e
Me
Me
0
e
4j
e(CH2)4e
e(CH2)4e
e
Bz
4-Pyridyl
e
0
e
4k
DPCPX
a
0
e
e
0.5 ꢂ 0.2 (nM)
4.1.1.4. 4-Hydrazino-2-propyl-5,6,7,8-tetrahydro[1]benzothieno
[2,3-d]pyrimidine (3f). M.p. 128e130 ꢁC; yield 82%; IR (KBr)
All the test compounds failed to show any A2AAR inhibition up to 1 mM
and DPCPX showed Ki values at 157 ꢂ 6.0 nM on rat A2A receptors.

618
M. Raghu Prasad et al. / European Journal of Medicinal Chemistry 43 (2008) 614e620
ymax: 3190, 3320 (NH2), 2910 (alk) cmꢀ1; 1H NMR (CDCl3) d:
1.0e1.1 (t, 3H, J ¼ 10.9 Hz, CH3CH2CH2e), 1.7e2.0 (m, 4H,
CH2 at 6 and 7), 2.45e2.55 (m, 2H, eCH2CH2CH3),
2.75e2.80 (m, 2H, CH2 at 5 and 8), 3.85e4.05 (m, 2H, e
CH2CH2CH3), 4.20e4.30 (d. 2H, J ¼ 5.5 Hz, eNHNH2, D2O
exchangeable), 6.6e6.8 (t, 1H, J ¼ 5.6 Hz, NHNH2, D2O ex-
changeable) ppm; EI-MS [m/z, %]: 262 [Mþ 100]. Anal. Calcd
for C13H18N4S: C, 59.54; H, 6.87; N, 21.37. Found: C, 59.85; H,
7.05; N, 21.60.
C13H14N4S2: C, 53.79; H, 7.3; N, 19.3. Found: C, 54.04; H,
7.26; N, 19.12.
4.1.2.5. 5-Propyl-8,9-dimethylthieno[3,2-e][1,2,4]triazolo[4,3-c]
pyrimidine-3(2H)-thione (4e). M.p. 198e200 ꢁC; yield 65%; IR
(KBr) ymax: 3190 (NH), 2910 (CH3), 1200 (C]S), 1290 (CeN)
1
cmꢀ1; H NMR (DMSO-d6) d: 1.10e1.13 (t, 3H, J ¼ 10.9 Hz,
CH3CH2CH2e), 2.28 (s, 6H, CH3), 2.45e2.55 (m, 2H,
CH3CH2CH2e), 3.05e3.15 (m, 2H, CH3CH2CH2e), 11.2 (s,
1H, NH, D2O exchangeable) ppm; EI-MS [m/z, %]: 278 [Mþ
100]. Anal. Calcd for C12H14N4S2: C, 51.79; H, 5.03; N, 20.14.
Found: C, 52.05; H, 5.25; N, 20.35.
4.1.2. General procedure for the preparation of 5-alkyl/aryl-
8,9-dimethyl/8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thiones (4aek)
2-Alkyl/aryl-4-hydrazino-5,6-dimethyl/5,6,7,8-tetrahydro-
benzo[b]thieno[2,3-d]pyrimidines (3aek) [10 mmol] and car-
bon disulphide [7.6 g, 10 ml, 100 mmol] in 10% alcoholic
potassium hydroxide (20 ml) solution was heated under reflux
for 5 h. The reaction mixture was cooled and added onto
crushed ice. The solid product thus obtained was filtered, dried
and recrystallized from chloroformeethanol (1:1) mixture.
4.1.2.6. 5-Propyl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thione (4f). M.p. > 300 ꢁC;
yield 74%; IR (KBr) ymax: 3190 (NH), 2910 (CH3), 1200
1
(C]S), 1290 (CeN) cmꢀ1; H NMR (DMSO-d6) d: 1.0e
1.1 (t, 3H, J ¼ 11.1 Hz, CH3CH2CH2), 1.7e2.0 (m, 4H, CH2
at 9 and 10), 2.45e2.55 (m, 2H, CH3CH2CH2), 2.75e2.80
(t, 2H, J ¼ 8.3 Hz, CH2 at 5), 2.85e3.05 (t, 2H, J ¼ 8.3 Hz,
CH2 at 8), 3.85e4.0 (m, 2H, CH3CH2CH2), 14.0 (s, 1H,
NH, D2O exchangeable) ppm; EI-MS [m/z, %]: 304 [Mþ
50]. Anal. Calcd for C14H16N4S2: C, 55.26; H, 5.26; N,
18.42. Found: C, 55.50; H, 5.55; N, 18.65.
4.1.2.1. 5-Methyl-8,9-dimethylthieno[3,2-e][1,2,4]triazolo[4,3-
c]pyrimidine-3(2H)-thione (4a). M.p. 275e276 ꢁC; yield 69%;
IR (KBr) ymax: 3190 (NH), 2910 (CH3), 1200 (C]S), 1290
1
(CeN) cmꢀ1; H NMR (CDCl3) d: 1.2 (s, 3H, CH3 at 5), 2.45
(s, 6H CH3 at 8 and 9), 11.4 (s, 1H, NH, D2O exchangeable)
ppm; EI-MS [m/z, %]: 250 [Mþ 100]. Anal. Calcd for
C10H10N4S2: C, 48.00; H, 4.00; N, 22.40. Found: C, 48.25; H,
4.30; N, 22.75.
4.1.2.7. 5-Phenyl-8,9-dimethylthieno[3,2-e][1,2,4]triazolo[4,3-c]
pyrimidine-3(2H)-thione (4g). M.p. > 300 ꢁC; yield 80%; IR
(KBr) ymax: 3190 (NH), 2910 (CH3), 1200 (C]S), 1290 (Ce
1
N) cmꢀ1; H NMR (DMSO-d6) d: 2.4 (s, 6H, CH3 at 8 and
9), 7.2e7.6 (m, 5H, C6H5), 14.2 (s, 1H, NH, D2O exchangeable)
ppm; EI-MS [m/z, %]: 312 [Mþ 100]. Anal. Calcd for
C15H12N4S2: C, 57.69; H, 3.84; N, 17.9. Found: C, 57.84; H,
4.12; N, 18.15.
4.1.2.2.
5-Methyl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thione (4b). M.p. 280e
281 ꢁC; yield 76%; IR (KBr) ymax: 3190 (NH), 2910 (CH3),
1200 (C]S), 1290 (CeN) cmꢀ1; H NMR (CDCl3) d: 1.9
1
(m, 4H, CH2 at 9 and 10), 2.35 (s, 1H CH3), 2.7e3.0 (m, 4H,
CH2 at 8 and 11), 11.1 (s, 1H, NH, D2O exchangeable) ppm;
EI-MS [m/z, %]: 276 [Mþ 100]. Anal. Calcd for C12H12N4S2:
C, 52.17; H, 4.34; N, 20.28. Found: C, 52.45; H, 4.66; N, 20.50.
4.1.2.8. 5-Phenyl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thione (4h). M.p. >
300 ꢁC; yield 80%; IR (KBr) ymax: 3190 (NH), 2910 (CH3),
1
1200 (C]S), 1290 (CeN) cmꢀ1; H NMR (DMSO-d6) d:
1.5e1.7 (m, 4H, CH2 at 9 and 10), 2.5e2.7 (m, 4H, CH2 at
8 and 11), 7.2e7.6 (m, 5H, ArH ), 14.3 (s, 1H, NH, D2O ex-
changeable) ppm; EI-MS [m/z, %]: 339 [Mþ100]. Anal. Calcd
for C15H12N4S2: C, 60.35; H, 4.14; N, 16.56. Found: C, 60.60;
H, 4.45; N, 16.95.
4.1.2.3. 5-Ethyl-8,9-dimethylthieno[3,2-e][1,2,4]triazolo[4,3-c]
pyrimidine-3(2H)-thione (4c). M.p. 180e181 ꢁC; yield 68%; IR
(KBr) ymax: 3190 (NH), 2910 (CH3), 1200 (C]S), 1290 (CeN)
1
cmꢀ1; H NMR (DMSO-d6) d: 1.2e1.4 (t, 3H, J ¼ 8.69 Hz,
CH2CH3), 2.35 (s, 6H CH3 at 8 and 9), 3.4 (t, 2H,
J ¼ 8.69 Hz, CH2CH3), 11.1 (s, 1H, NH, D2O exchangeable)
ppm; EI-MS [m/z, %]: 264 [Mþ 100]. Anal. Calcd for
C11H12N4S2: C, 50.76; H, 4.61; N, 21.53. Found: C, 50.95;
H, 4.75; N, 21.65.
4.1.2.9. 5-Benzyl-8,9-dimethylthieno[3,2-e][1,2,4]triazolo[4,3-
c]pyrimidine-3(2H)-thione (4i). M.p. 165e167 ꢁC; yield 73%;
IR (KBr) ymax: 3190 (NH), 2910 (CH3), 1200 (C]S), 1290 (Ce
N) cmꢀ1; 1H NMR (DMSO-d6) d: 2.5 (s, 6H, CH3), 3.2 (S, 2H,
CH2Ph), 7.28e7.67 (m, 5H, C6H5), 14.1 (s, 1H, NH, D2O ex-
changeable) ppm; EI-MS [m/z, %] (70 eV): 326 [Mþ 25], 294
[MþꢀSH, 100]. Anal. Calcd for C16H14N4S2: C, 58.89; H,
4.29; N, 17.17. Found: C, 59.16; H, 4.55; N, 17.35.
4.1.2.4. 5-Ethyl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thione (4d). M.p. 244e
245 ꢁC; yield 72%; IR (KBr) ymax: 3190 (NH), 2910 (CH3),
;
1200 (C]S), 1290 (CeN) cmꢀ1 1H NMR (DMSO-d6) d:
1.1 (t, 3H J ¼ 4.4 Hz, CH3), 1.7e2.0 (m, 4H, CH2 at 9 and
10), 2.8e3.0 (m, 4H, CH2 at 8 and 11), 3.9e4.0 (q, 2H,
J ¼ 4.24 Hz, eCH2CH3), 14.0 (s, 1H, NH, D2O exchangeable)
ppm; EI-MS [m/z, %]: 290 [Mþ 100]. Anal. Calcd for
4.1.2.10. 5-Benzyl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thione (4j). M.p. 232e
234 ꢁC; yield 75%; IR (KBr) ymax: 3190 (NH), 2910 (CH3),
;
1200 (C]S), 1290 (CeN) cmꢀ1 1H NMR (DMSO-d6) d:

M. Raghu Prasad et al. / European Journal of Medicinal Chemistry 43 (2008) 614e620
619
1.3e1.5 (m, 4H, CH2 at 9 and 10), 2.2e2.5 (m, 4H, CH2 at 8
Acknowledgements
and 11), 3.35 (s, 2H, CH2Ph), 7.25e7.8 (m, 5H, ArH ), 14.0 (s,
1H, NH, D2O exchangeable) ppm; EI-MS [m/z, %] (70 eV):
352 [Mþ 100]. Anal. Calcd for C18H16N4S2: C, 61.36; H,
4.54; N, 15.90. Found: C, 61.50; H, 4.85; N, 16.27.
One of the authors (MRP) is thankful to All India Council
for Technical Education (AICTE) for granting a Career Award
for Young Teachers (CAYT no. 1-51/FD/CA(11)/2004e05).
The authors wish to thank the Principal, University College
of Pharmaceutical Sciences, Kakatiya University for providing
facilities and IISC, Bangalore for spectral data. We thankfully
acknowledge the cooperation extended by Prof. Dr. Christa
4.1.2.11. 5-Pyridyl-8,9,10,11-tetrahydro[1]benzothieno[3,2-e]
[1,2,4]triazolo[4,3-c]pyrimidine-3(2H)-thione (4k). M.p. 238e
240 ꢁC; yield 67%; IR (KBr) ymax: 3190 (NH), 2910 (CH3),
1200 (C]S), 1290 (CeN) cmꢀ1
;
1H NMR (DMSO-d6) d:
E. Muller, University of Bonn, Germany by providing facili-
¨
1.6e1.9 (m, 4H, CH2 at 9 and 10), 2.4e2.8 (m, 4H, CH2 at
8 and 11), 7.65e8.25 (m, 4H, ArH ), 14.4 (s, 1H, NH, D2O
exchangeable) ppm; EI-MS [m/z, %]: 339 [Mþ 100]. Anal.
Calcd for C16H13N5S2: C, 56.63; H, 3.83; N, 20.64. Found:
C, 56.95; H, 4.10; N, 20.89.
ties, equipment and biochemicals for performing radioligand
binding studies and guidance.
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