Development of antimycobacterial tetrahydrothieno[2,3-c]pyridine-3- carboxamides and hexahydrocycloocta[b]thiophene-3-carboxamides: Molecular modification from known antimycobacterial lead

Article abstract of DOI:10.1016/j.ejmech.2014.02.028

Twenty derivatives of 2,6-disubstituted 4,5,6,7-tetrahydrothieno[2,3-c] pyridine-3-carboxamide and ten of 2-substituted 4,5,6,7,8,9- hexahydrocycloocta[b]thiophene-3-carboxamide were synthesized by molecular modification of a known antimycobacterial molecule. Compounds were evaluated in vitro against Mycobacterium tuberculosis (MTB), and cytotoxicity against RAW 264.7 cell line. Among the compounds, 2-(4-phenoxybenzamido)-4,5,6,7,8,9- hexahydrocycloocta[b]thiophene-3-carboxamide (12f) was found to be the most active compound against MTB with MIC of 3.70 μM and was more potent than Ethambutol (MIC of 7.64 μM), Ciprofloxacin (MIC of 9.41 μM) and standard lead compound SID 92097880 (MIC of 9.15 μM). Compound 12f also showed MTB MIC of 1.23 μM in the presence of an efflux pump inhibitor verapamil, and showed no cytotoxicity at 50 μM.

Full text of DOI:10.1016/j.ejmech.2014.02.028

European Journal of Medicinal Chemistry 76 (2014) 110e117
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Development of antimycobacterial tetrahydrothieno[2,3-c]pyridine-3-
carboxamides and hexahydrocycloocta[b]thiophene-3-carboxamides:
Molecular modification from known antimycobacterial lead
Radhika Nallangi, Ganesh Samala, Jonnalagadda Padma Sridevi, Perumal Yogeeswari,
*
Dharmarajan Sriram
Department of Pharmacy, Birla Institute of Technology & Science-Pilani, Hyderabad Campus, Shameerpet, R.R. District, Hyderabad 500078, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Twenty derivatives of 2,6-disubstituted 4,5,6,7-tetrahydrothieno[2,3-c]pyridine-3-carboxamide and ten
of 2-substituted 4,5,6,7,8,9-hexahydrocycloocta[b]thiophene-3-carboxamide were synthesized by mo-
lecular modification of a known antimycobacterial molecule. Compounds were evaluated in vitro against
Mycobacterium tuberculosis (MTB), and cytotoxicity against RAW 264.7 cell line. Among the compounds,
2-(4-phenoxybenzamido)-4,5,6,7,8,9-hexahydrocycloocta[b]thiophene-3-carboxamide (12f) was found
Received 9 December 2013
Received in revised form
29 January 2014
Accepted 8 February 2014
Available online 11 February 2014
to be the most active compound against MTB with MIC of 3.70
(MIC of 7.64 M), Ciprofloxacin (MIC of 9.41 M) and standard lead compound SID 92097880 (MIC of
9.15 M). Compound 12f also showed MTB MIC of 1.23 M in the presence of an efflux pump inhibitor
verapamil, and showed no cytotoxicity at 50 M.
mM and was more potent than Ethambutol
m
m
Keywords:
Tuberculosis
Cytotoxicity
Tetrahydrothieno[2,3-c]pyridine-3-
carboxamides
m
m
m
Ó 2014 Elsevier Masson SAS. All rights reserved.
Hexahydrocycloocta[b]thiophene-3-
carboxamides
Efflux pump
1. Introduction
and the lack of drugs that effectively target persistent bacteria, there
seem to be an urgent need for identification of new antimicrobial
targets and inhibitors [5]. Target-based approaches are widely used
in drug discovery; questions have been raised about the efficiency of
this approach given the very high attrition rates that these projects
have historically shown in the anti-infective field [6]. Compounds
identified in whole-cell screens provide suitable chemical and bio-
logical starting points. The Tuberculosis Antimicrobial Acquisition
and Coordinating Facility (TAACF) was established by the National
Institute of Allergy and Infectious Diseases (NIAID) in 1994 to allow
researchers access to high quality screening services in order to
encourage antituberculosis drug discovery research. Recently
TAACF reported anti-TB high-throughput results of large libraries of
drug-like small molecules [7e9]. One among them was the mole-
cule SID 92097880 6-acetyl-2-(thiophene-2-carboxamido)-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide that showed ac-
tivity against M. tuberculosis H37Rv (MTB) with minimum inhibitory
Mycobacterium tuberculosis (MTB), the causative agent of
tuberculosis (TB), is one of the most lethal infectious agents
affecting mankind [1]. The disease infects almost two billion people
or one-third of the world’s population, and accounts for an esti-
mated 2 million deaths per year [2]. MTB infection is difficult to
treat, requiring 6e9 months of chemotherapy with a cocktail of four
antibiotics isoniazid, rifampin, pyrazinamide and ethambutol. In
addition to toxic side effects, the lengthy treatment regime results in
poor patient compliance and thus drug resistant strains are begin-
ning to emerge [3]. The World Health Organization estimates that
up to 50 million people worldwide are infected with multidrug
resistant strains of MTB (MDR-TB) [4]. This number continues to
grow as 300,000 new MDR-TB cases are diagnosed each year with
79 percent of individuals showing resistance to three or more
frontline drugs [4]. Taken together, the growing problem of MDR-TB
concentration (MIC) of 9.15 mM and selectivity index of 13 (Fig. 1).
We took SID 92097880 as the starting point to design and synthesize
various 2,6-disubstituted 4,5,6,7-tetrahydrothieno[2,3-c]pyridine-
3-carboxamides and 2-substituted 4,5,6,7,8,9-hexahydrocycloocta
[b]thiophene-3-carboxamides to study extensively their structure
* Corresponding author.
E-mail addresses: dsriram@hyderabad.bits-pilani.ac.in, drdsriram@yahoo.com
(D. Sriram).
http://dx.doi.org/10.1016/j.ejmech.2014.02.028
0223-5234/Ó 2014 Elsevier Masson SAS. All rights reserved.

R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
111
at 70 ꢀC, we noticed good yields (nearly 20% more) when the re-
action was carried out at room temperature, and in addition also
observed that there was a good amount of solids formed in the
reaction mixture which eased the purification process. The primary
aromatic amine group of compound 4 was coupled with various
alkyl, cycloalkyl, (sub)aryl carboxylic acids by employing peptide
coupling reagents 1-ethyl-3-(3-dimethylaminopropyl)carbodii-
mide (EDCI) and HOBt to produce compounds 5aej. Due to the poor
solubility of reaction products, we observed formation of desired
product as solid in reaction mixture. Direct filtration of reaction
mixture washing with distilled water, hexane produced pure
products without any further purification. In next step deprotection
of Boc-protection was carried out by using trifluoroacetic acid to get
compounds 6aej. The conversion of compounds 6aej to target
compounds 7aej was achieved by treating with acetyl chloride in
dichloromethane using Et3N as base.
In order to achieve the synthesis of compound 8aej, we tried
direct methylation using CH3I and sodium hydride conditions, but
the reaction was unsuccessful. Then we employed reductive amina-
tion conditions using formaldehyde and Na(OAc)3BH in MeOH with
catalytic amount of acetic acid to produce compounds 8aej. We also
tried to synthesize the final compounds 7aej and 8aej using 1-
acetylpiperidin-4-one and 1-methylpiperidin-4-one respectively
as the starting materials in order to achieve them in two steps.
We succeeded in first step i.e. synthesis of 6-acetyl-2-amino-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide and 2-amino-6-
methyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine-3-carboxamide. But
in next step, due to very poor solubility, the coupling reaction was
unsuccessful with various carboxylic acids.
O
R
NH
S
H2N
O
O
R1= Acetyl,
Methyl:
R = Alkyl,
Cycloalkyl,
(sub)Aryl:
S
NH
S
7a-j,
8a-j
H2N
N
R1
O
Structural
modification
CONH2
NH
12a-j
N
R
O
CH3
SID 92097880
MTB MIC: 9.15uM
S
O
Fig. 1. Structural modification of lead compound.
activity relationship (SAR) (Fig. 1) and to develop a more potent
compound.
2. Results and discussion
2.1. Chemistry
In Scheme 1, we synthesised 2,6-disubstituted 4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamides by following a
five step synthetic protocol starting from the commercially avail-
able, less expensive 4-piperidone monohydrate hydrochloride (1).
In first step we protected the secondary amine group by Boc-
protection to get ‘tert-butyl 4-oxopiperidine-1-carboxylate’ (2). In
the next step, by following Gewald reaction conditions using
morpholine, cyanoacetamide (3) and elemental sulphur achieved
In Scheme 2 (2-substituted 4,5,6,7,8,9-hexahydrocycloocta[b]
thiophene-3-carboxamides), for the preparation of intermediate
compound 11, we used the same reaction conditions which are fol-
lowed for the conversion 2 to 4 (morpholine, cyanoacetamide and
elemental sulphur), but the reactionwas unsuccessful. Hence instead
of cyanoacetamide we used malanonitrile under Gewald reaction
conditions to get 2-amino-4,5,6,7,8,9-hexahydrocycloocta[b]thio-
phene-3-carbonitrile (10). Compound 10 was then treated with conc.
‘tert-butyl
2-amino-3-carbamoyl-4,5-dihydrothieno[2,3-c]pyri-
dine-6(7H)-carboxylate’ (4). This conversion (2e4) was successful
in both the conditions i.e. at room temperature and at 70 ꢀC. But
they differ in reaction time, purification process and yield. When
reaction carried out at 70 ꢀC we observed the complete conversion
of starting material in less than 2 h and it has taken 6e9 h to
complete the reaction at room temperature. Compared to reaction
H2SO4
to
afford
2-amino-4,5,6,7,8,9-hexahydrocycloocta[b]
R
O
O
NH2
S
HN
O
N
O
H2N
H2N
O
NC
O
NH2
(Boc)2O
a
R-COOH
c
3
S
b
N
H
N
N
O
O
HCl. H2O
O
O
O
O
4
5a-j
1
2
R
O
O
R
O
O
HN
S
HN
H2N
H2N
TFA
d
S
e
N
N
R1
H CF3COOH
R1 = Ac:
7a-j,
R1 = CH3: 8a-j
6a-j
Reagents & Conditions: a: (Boc)2O, Et3N, DCM, MeOH, rt; b: S8, Morpholine, EtOH, rt, 6h;
c: EDCI, HOBt, Et3N, DCM, rt, 9h; d: TFA, DCM, rt, 2h; e: R1 = Acetyl; Acetylchloride, Et3N,
CH2Cl2 and R2 = Methyl; HCHO, Na(OAc)3BH, MeOH, H2O, rt, 2h:
Scheme 1. Synthetic protocol of tetrahydrothieno[2,3-c]pyridine-3-carboxamides.

112
R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
O
O
NH2
CN
NH2
NH2
O
c
a
b
H
N
NH2
O
S
S
S
R
12a-j
9
10
11
Reagents & Conditions: a: Malanonitrile, S8, Et3N, DMF, rt; b: Concd H2SO4, rt, 6 h; c: RCOOH,
EDCI, HOBt, Et3N, DCM, rt, 9 h:
Scheme 2. Synthetic protocol of hexahydrocycloocta[b]thiophene-3-carboxamides.
thiophene-3-carboxamide (11). The free amino group of compound
11 on reaction with various carboxylic acids produced the target
molecules 12aej. The purity of the synthesized compounds was
checked by elemental analysis and TLC and the structures were
identified by spectral analysis. In the nuclear magnetic resonance
spectra (1H NMR and 13C NMR), the signals of the respective protons
of the prepared derivatives were verified on the basis of their
chemical shifts, multiplicities, and coupling constants. The elemental
analysis results were within ꢁ0.4% of the theoretical values.
with MIC of 3.70 mM against log-phase culture of MTB. All the
synthesized compounds were less potent than standard first line
antitubercular compound Isoniazid. Three compounds were found
to be more potent than Ethambutol (MIC 7.64
were found to be more potent than lead compound SID 92097880
(MIC 9.15 M).
mM). Five compounds
m
With respect to SAR of 2,6-disubstituted 4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamides (7aej and 8aej);
we have prepared various alkyl, cycloalkyl and (sub)aryl amide at
2nd position and at N-6 position we have prepared acetyl (7aej)
and methyl (8aej) derivatives. At N-6 position, compounds with N-
methyl derivatives showed better activity (MIC ranging 5.06e
The structures of all final compounds were established from 1H
and 13C NMR spectroscopic data as illustrated for a representative
example 12f (Fig. 2). The 1H NMR spectrum of 12f has a doublet at
7.89 ppm (J ¼ 8.7 Hz) corresponds to aromatic H-10, a doublet at
7.44 ppm (J ¼ 7.8 Hz) is due to H-40, a triplet at 7.24 ppm (J ¼ 7.5 Hz)
corresponds to H-50 and the other doublet at 7.13 ppm (J ¼ 8.4 Hz)
corresponds to H-20 and H-30 protons. The two triplets at 2.84,
2.75 ppm corresponds to C-4 and C-9 protons respectively, the
protons of C-5, C-6, C-7 and C-8 gave a multiplet (1.56e1.19) ppm.
39.43 mM) than N-acetyl derivatives (MIC ranging 57.30e88.60 mM).
These results indicated that alkyl substituent favour bioactivity
than the acyl derivative. Among compounds 8aej; hydrophobic
substituent at C-2 position increased the activity. Cycloalkyl de-
rivatives (8he8j) showed promising activity with MIC less than
10 mM; followed by naphthyl derivative (8g) with MIC of 17.07 mM.
When compared to phenyl derivative (8b), naphthyl derivative
showed two times more potency. Among (sub)phenyl derivatives
(8be8f), not much difference were found between electron with-
drawing and donating groups at phenyl ring. With respect to SAR of
4,5,6,7,8,9-hexahydrocycloocta[b]thiophene-3-carboxamides
(12aej), where we replaced N-acetyl tetrahydropyridine moiety of
lead compound SID 92097880 with highly hydrophobic cyclooctyl
moiety showed better activity with MIC ranging from 3.70 to
2.2. In vitro MTB screening
All the synthesized compounds were screened for their in vitro
anti-tubercular activity against M. tuberculosis H37Rv (ATCC27294)
using agar dilution method and drug concentrations from 50
mg/mL
to 0.78 g/mL in duplicates. The minimum inhibitory concentration
m
(MIC) was determined for each compound. The MIC was taken as
the minimum concentration of compound required to completely
inhibit the bacterial growth. Isoniazid, Ethambutol, Ciprofloxacin
and SID 92097880 were used as reference compounds for com-
parison. The MIC values of the synthesized compounds along with
the standard drug for comparison are presented in Table 1. All the
synthesized compounds showed activity against MTB with MIC
18.72 mM. In this series some of substituted phenyl derivatives at C-
2 position (12e and 12f) showed better activity (less than 5
than cycloalkyl (12he12j) and naphthyl derivative (12g).
mM)
When compared to lead compound SID 92097880, many of the
synthesized compounds showed less activity. It could be due to the
involvement of wide array of efflux mechanisms mediated by
several ABC (ATP-binding cassette) transporters and major facili-
tator superfamily (MFS) proteins, or antibiotic-modifying and-
degrading enzymes, to name a few possibilities [10]. Multiple drugs
like verapamil, reserpine, phenothiazines such as thioridazine, and
piperine have been shown to inhibit bacterial efflux pumps in vitro
[11]. In general, the mechanisms by which these agents act were
poorly understood. Several models have been proposed [12], such
as: (1) direct binding and inhibition of pump assembly or function;
(2) disruption of the transmembrane gradients utilized by secondary
transporters; (3) inhibitor binding to the antimicrobial compound;
and (4) competition for efflux. We have tested some selected com-
ranging from 3.70 to 88.6
m
M. Seven compounds (8he8j, 12e, 12f,
M. Compound 12f (2-(4-
12i and 12j) inhibited MTB with MIC <10
phenoxybenzamido)-4,5,6,7,8,9-hexahydrocycloocta[b]thiophene-
3-carboxamide) was found to be the most active compound in vitro
m
7.24, t,
J = 7.5 Hz
2.84, t
J = 5.7 Hz
124.6
H
H
O
5'
4'
NH2
4
5
7.44, d,
J = 7.8 Hz
129.4
3
3'
m
H
,
6
7
9
2'
1'
pounds [MIC of <20 mM], in presence of reported efflux pump in-
hibitor verapamil; and in most of the cases MIC was decreased 2e3
fold when compared to absence of efflux pump inhibitor. Most active
1
.
NH
O
1
2
-
6
5
S
O
.
1
1
9
8
7.13, d,
J = 8.4 Hz
117.9
2.75, t,
compound 12f showed MIC of 1.23 mM in this study.
H
H
7.13, d,
J = 8.4 Hz
119.8
7.89, d,
J = 8.7 Hz
130.3
2.3. Cytotoxicity study
Compounds with MIC less than 20
mM were further examined
Fig. 2. 1H and 13C chemical shifts in compound 12f.
for cytotoxicity in a RAW 264.7 cell line (mouse leukaemic

R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
113
Table 1
Biological activities of synthesized compounds.
Compound
R
Yield (%)
MP (ꢀC)
MTB MIC in
m
M
MTB MIC in
mM
Cytotoxicity at 50 mM
[in presence of verapamil]
(RAW 264.7 cells) % inhibition
7a
7b
7c
7d
7e
7f
7g
7h
7i
7j
8a
8b
8c
8d
8e
8f
8g
8h
8i
8j
12a
12b
12c
12d
12e
12f
12g
12h
12i
12j
Methyl
Phenyl
81
63
70
84
79
72
61
74
64
78
58
66
62
81
73
73
56
72
62
72
79
88
60
69
63
69
70
66
54
79
223e224
231e232
202e204
251e252
204e205
190e191
270e271
204e205
280e281
208e209
222e223
182e183
205e206
224e225
133e135
207e208
180e181
231e232
260e261
206e207
186e187
213e214
243e244
228e229
216e217
210e211
190e191
145e146
175e176
223e224
88.6
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
8.53
2.78
5.06
2.58
9.36
5.06
6.96
16.71
2.27
1.23
3.29
8.53
9.73
3.11
0.72
3.82
9.41
4.57
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
28.42
22.16
18.16
18.28
24.56
26.62
24.80
36.14
20.52
22.16
28.32
16.86
18.34
16.64
NT
72.65
66.84
64.26
69.80
57.30
63.45
81.15
74.40
71.40
39.43
37.87
36.12
34.62
37.87
30.60
17.07
5.57
2-Methoxyphenyl
3-Nitrophenyl
4-Tolyl
4-Phenoxyphenyl
1-Napthyl
Cyclopropyl
Cyclopentyl
Cyclohexyl
Methyl
Phenyl
2-Methoxyphenyl
3-Nitrophenyl
4-Tolyl
4-Phenoxyphenyl
1-Napthyl
Cyclopropyl
Cyclopentyl
Cyclohexyl
Methyl
5.06
7.76
18.72
15.19
13.92
16.71
4.54
Phenyl
2-Methoxyphenyl
3-Nitrophenyl
4-Tolyl
4-Phenoxyphenyl
1-Napthyl
Cyclopropyl
Cyclopentyl
Cyclohexyl
3.70
13.19
17.06
9.73
9.33
0.72
7.64
9.41
9.15
Isoniazid
Ethambutol
Ciproflaxacin
SID 92097880
NT
NT
NT
NT indicates not tested.
monocyte macrophage) at single concentration of 50
m
M. We
compounds, 2-(4-phenoxybenzamido)-4,5,6,7,8,9-hexahydrocy-
have selected this macrophage cell line to test the toxicity as
generally MTB reside inside the macrophages and we expected
the new molecules not to show any toxicity against macrophages.
After 72 h of exposure, viability was assessed on the basis of
cellular conversion of MTT into a formazan product using the
Promega Cell Titer 96 non-radioactive cell proliferation assay.
Most of the tested compounds were not cytotoxic to RAW
264.7 cells based on their percentage growth inhibitions (less
than 30%) as reported in Table 1. The most active anti-TB com-
cloocta[b]thiophene-3-carboxamide (12f) was found to be the
most active compound in vitro with MIC of 3.70
phase culture of MTB and also non-toxic up to 50
pound 12f was w3 times more potent than lead compound SID:
92097880. Compound 12f showed MIC of 1.23 M in the pres-
m
M against log-
m
M. Com-
m
ence of efflux pump inhibitor. Further structural optimization
can be performed to get compounds with better potency than
the lead compound and also to explore the possible mechanism
of action against various MTB essential enzymes. In addition,
efflux could be the rate-limiting step in the discovery of novel
anti-TB compounds, as has already been recognized in the dis-
covery of drugs for Gram-negative bacterial infections. The
discovery of new pumps with multiple specificities in MTB and
the impact of these pumps on antitubercular therapy by
conferring resistance to many of the new molecules discovered
necessitate the study of efflux mechanism as an important
therapeutic target.
pound 12f showed 22.6% cytotoxicity at 50
index of >10 for MTB.
mM with selectivity
3. Conclusion
In this study we have designed, synthesized and studied
SAR of various inhibitors of MTB based on the lead compound
SID 92097880 earlier reported by TAACF. Among the

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R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
4. Experimental
4.1.7. 6-Acetyl-2-benzamido-4,5,6,7-tetrahydrothieno[2,3-c]
pyridine-3-carboxamide (7b)
4.1. Chemistry
MS (ESI) 344 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 10.17
(s, 2H), 7.81e7.63 (m, 6H), 4.50 (s, 2H), 3.81 (t, J ¼ 7.6 Hz, 2H), 3.10
4.1.1. Preparation of tert-butyl 4-oxopiperidine-1-carboxylate (2)
Et3N (26.45 mL, 185.17 mmol) was added drop-wise to a stirred
solution of compound (1) (4.0 g, 29.60 mmol) in CH2Cl2 (20 mL) and
MeOH (4 mL) at 0 ꢀC, then (Boc)2O (7.74 mL, 35.52 mmol) was
added drop wise to the reaction mixture at same temperature and
allowed to stir at room temperature for 16 h. The reaction mixture
was diluted with H2O (30 mL) and extracted with CH2Cl2
(3 ꢂ 60 mL). The separated organic layer was concentrated under
reduced pressure and the crude residue was washed with hexanes
to get compound 2 (4.30 g, 72%) as an off-white solid. MS (ESI) 200
[M þ H]þ. Anal. calcd for C10H17NO3: C, 60.28; H, 8.60; N, 7.03%
Found C, 60.34; H, 8.64; N, 7.11%.
(t, J ¼ 7.6 Hz, 2H), 2.20 (s, 3H); 13C NMR (100 MHz, DMSO-d6)
d
178.3, 172.4, 167.9, 139.4, 133.8, 131.4(2C), 130.8, 128.4(2C), 126.3,
123.8, 118.9, 43.8, 39.7, 25.0, 22.5. Anal. calcd for C17H17N3O3S: C,
59.46; H, 4.99; N, 12.24% Found C, 59.52; H, 5.12; N, 12.32%.
4.1.8. 6-Acetyl-2-(2-methoxybenzamido)-4,5,6,7-tetrahydrothieno
[2,3-c]pyridine-3-carboxamide (7c)
MS (ESI) 374 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 12.45
(s, 1H), 9.97 (s, 2H), 7.74e7.63 (m, 2H), 7.56 (d, J ¼ 7.6 Hz, 1H), 7.49
(t, J ¼ 8.0 Hz, 1H), 4.58 (s, 2H), 3.94 (s, 3H), 3.72 (t, J ¼ 7.2 Hz, 2H),
3.15 (t, J ¼ 7.6 Hz, 2H), 2.16 (s, 3H); 13C NMR (75 MHz, DMSO-d6)
d
181.3, 176.3, 171.9, 148.9, 141.8, 139.4, 133.8, 132.9, 130.3, 128.7,
127.4, 122.4, 114.8, 55.8, 41.6, 38.9, 22.5, 21.6. Anal. calcd for
C18H19N3O4S: C, 57.89; H, 5.13; N, 11.25% Found C, 57.92; H, 5.22; N,
4.1.2. Preparation of tert-butyl 2-amino-3-carbamoyl-4,5-
dihydrothieno[2,3-c]pyridine-6(7H)-carboxylate (4)
11.32%.
To the stirred solution of compound 2 (6.0 g, 30.14 mmol),
cyanoacetamide (3.01 g, 37.18 mmol), sulphur powder (2.13 g,
30.14 mmol) in ethanol (60 mL) was added morpholine (6.01 mL,
60.30 mmol) and stirred the reaction mixture at room temperature
for 12 h the reaction mixture was concentrated, and the crude solid
was filtered, washed with H2O and dried in vacuum oven for 2 h.
The obtained dried compound was purified by column chroma-
tography to yield (3.67 g, 82%) as a light yellow solid. MS (ESI) 298
4.1.9. 6-Acetyl-2-(3-nitrobenzamido)-4,5,6,7-tetrahydrothieno[2,3-
c]pyridine-3-carboxamide (7d)
MS (ESI) 389 [M þ H]þ. 1H NMR (400 MHz, CDCl3):
d 9.21 (s, 1H),
8.31 (s, 1H), 8.10 (s, 2H), 7.99e7.90 (m, 2H), 7.72 (t, J ¼ 8.4 Hz, 1H),
4.59 (s, 2H), 3.79 (t, J ¼ 7.6 Hz, 2H), 3.09 (t, J ¼ 8.0 Hz, 2H), 2.23 (s,
3H); 13C NMR (100 MHz, DMSO-d6)
d 177.9,175.3,168.4,141.9,139.8,
132.5, 130.5, 128.4, 126.9, 124.8, 122.4, 120.4, 116.3, 45.2, 38.4, 24.9,
22.5. Anal. calcd for C17H16N4O5S: C, 52.57; H, 4.15; N, 14.43% Found
C, 52.62; H, 4.22; N, 14.52%.
[M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 11.25 (s, 2H), 7.90 (s, 2H),
4.32 (s, 2H), 3.51 (t, J ¼ 6.8 Hz, 2H), 3.04 (t, J ¼ 6.8 Hz, 2H); Anal.
calcd for C13H19N3O3S: C, 52.51; H, 6.44; N, 14.13% Found C, 52.64;
H, 6.55; N, 14.11%.
4.1.10. 6-Acetyl-2-(4-methylbenzamido)-4,5,6,7-tetrahydrothieno
[2,3-c]pyridine-3-carboxamide (7e)
4.1.3. General procedure for the preparation of 5aej
MS (ESI) 358 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 10.27 (s,
To the stirred solution of ReCOOH (1.20 equiv) in CH2Cl2 was
added EDCI (1.30 equiv), HOBt (1.30 equiv) and Et3N (2.50 equiv)
allowed the reaction mixture to stir for few minutes, then added
compound 4 (1.00 equiv). The reaction mixture was stirred at room
temperature for 3 h. Concentrated and triturated with H2O to get
solid compound. The solids were given cold ethanol, diethyl ether
and hexane washings to get pure products.
2H), 7.81 (s, 1H), 7.90 (d, J ¼ 8.0 Hz, 2H), 7.63 (d, J ¼ 8.0 Hz, 2H), 4.60
(s, 2H), 3.81 (t, J ¼ 7.6 Hz, 2H), 3.18 (t, J ¼ 7.2 Hz, 2H), 2.41 (s, 3H),
2.18 (s, 3H); 13C NMR (75 MHz, DMSO-d6)
d 178.7, 169.3, 166.2,
134.0, 132.9, 130.3(2C), 129.3(2C), 127.4, 123.4, 121.4, 116.3, 51.8,
42.6, 26.2, 24.5, 23.7. Anal. calcd for C18H19N3O3S: C, 60.49; H, 5.36;
N, 11.76% Found C, 60.52; H, 5.39; N, 11.91%.
4.1.11. 6-Acetyl-2-(4-phenoxybenzamido)-4,5,6,7-tetrahydrothieno
4.1.4. Preparation of 6aej
[2,3-c]pyridine-3-carboxamide (7f)
To the stirred solution of Compound 5aej in CH2Cl2 at 0 ꢀC
under N2 atm, was added TFA (2 volumes) and allowed to stir the
reaction mixture at room temperature for 2 h. The reaction mixture
was concentrated to dryness and the obtained solids were washed
with hexanes to afford respective compounds 6aej.
MS (ESI) 436 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 12.27 (s,
1H), 7.92 (s, 2H), 7.81 (d, J ¼ 8.0 Hz, 2H), 7.72 (d, J ¼ 8.4 Hz, 2H),
7.69e7.54 (m, 5H), 4.70 (s, 2H), 3.80 (t, J ¼ 8.4 Hz, 2H), 3.13 (t,
J ¼ 8.0 Hz, 2H), 2.23 (s, 3H); 13C NMR (100 MHz, CDCl3)
d 181.7,
172.7, 169.6, 146.3, 143.7, 132.3(2C), 129.0(2C), 128.4, 127.4(2C),
125.9,125.1(2C),122.6,123.5,121.8,115.9, 46.8, 41.8, 23.6, 20.4. Anal.
calcd for C23H21N3O4S: C, 63.43; H, 4.86; N, 9.65% Found C, 63.52; H,
4.99; N, 9.72%.
4.1.5. General procedure for the preparation of 7aej
To the stirred solution of compounds 6aej in CH2Cl2 at 0 ꢀC
under N2 atm, was added Et3N (2.0 equiv), followed by acetyl
chloride (1.2 equiv) and allowed to stir the reaction mixture at
room temperature for 4 h. The reaction mixture was concentrated
to dryness and the obtained solids were washed with excess
water, cold ethanol and hexanes to afford respective final com-
pounds 7aej.
4.1.12. 2-(1-Naphthamido)-6-acetyl-4,5,6,7-tetrahydrothieno[2,3-
c]pyridine-3-carboxamide (7g)
MS (ESI) 394 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 10.29 (s,
1H), 8.19e8.10 (m, 4H), 7.92e7.74 (m, 5H), 4.57 (s, 2H), 3.90 (t,
J ¼ 8.0 Hz, 2H), 3.14 (t, J ¼ 7.2 Hz, 2H), 2.14 (s, 3H); 13C NMR
(75 MHz, CDCl3) d 179.2, 176.2, 168.2, 141.3, 140.7, 138.4, 137.9, 136.2,
4.1.6. 2-Acetamido-6-acetyl-4,5,6,7-tetrahydrothieno[2,3-c]
135.4, 132.9, 128.7, 126.0, 125.3, 123.3, 122.6, 121.8, 118.1, 44.4, 39.8,
26.2, 21.7. Anal. calcd for C21H19N3O3S: C, 64.10; H, 4.87; N, 10.68%
Found C, 64.12; H, 5.02; N, 10.72%.
pyridine-3-carboxamide (7a)
MS (ESI) 282 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 10.42
(s, 2H), 7.90 (s, 1H), 4.61(s, 2H), 3.91(t, J ¼ 7.6 Hz, 2H), 3.18
(t, J ¼ 8.0 Hz, 2H), 2.19 (s, 3H), 2.10 (s, 3H); 13C NMR (100 MHz,
4.1.13. 6-Acetyl-2-(cyclopropanecarboxamido)-4,5,6,7-
DMSO-d6)
d
172.3, 169.8, 161.6, 133.1, 129.8, 128.5, 112.2, 51.9, 42.8,
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (7h)
26.3, 23.4, 22.5. Anal. calcd for C12H15N3O3S: C, 51.23; H, 5.37; N,
14.94% Found C, 51.32; H, 5.39; N, 15.01%.
MS (ESI) 308 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 9.97
(s, 2H), 6.93 (s, 1H), 4.66 (s, 2H), 3.97(t, J ¼ 8.0 Hz, 2H), 3.21

R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
115
(t, J ¼ 8.0 Hz, 2H), 2.25 (s, 3H), 1.92e1.86 (m, 1H), 1.28e1.03 (m, 4H);
13C NMR (100 MHz, DMSO-d6)
178.4, 172.9, 166.4, 133.3, 131.3,
129.4, 119.1, 43.4, 41.6, 27.4, 24.5, 22.3, 20.7(2C). Anal. calcd for
14H17N3O3S: C, 54.71; H, 5.57; N, 13.67% Found C, 54.82; H, 5.64; N,
4.1.20. 6-Methyl-2-(3-nitrobenzamido)-4,5,6,7-tetrahydrothieno
d
[2,3-c]pyridine-3-carboxamide (8d)
MS (ESI) 361 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 10.21
C
(s, 2H), 9.45 (s, 1H), 8.40 (s, 1H), 8.01 (d, J ¼ 8.0 Hz, 1H), 7.81e7.72
13.71%.
(m, 2H), 4.61 (s, 2H), 3.81 (t, J ¼ 8.0 Hz, 2H), 3.12 (t, J ¼ 8.0 Hz, 2H),
2.34 (s, 3H); 13C NMR (75 MHz, DMSO-d6)
d 181.8, 173.3, 149.8,
144.6, 139.4, 137.4, 136.4, 133.4, 129.9, 126.8, 124.4, 118.3, 51.2, 46.4,
38.9, 26.5. Anal. calcd for C16H16N4O4S: C, 53.32; H, 4.47; N, 15.55%
Found C, 53.42; H, 4.52; N, 15.64%.
4.1.14. 6-Acetyl-2-(cyclopentanecarboxamido)-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (7i)
MS (ESI) 336 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d
9.81 (s,
2H), 7.69 (s, 1H), 4.71 (s, 2H), 3.94 (t, J ¼ 7.6 Hz, 2H), 3.16 (t,
4.1.21. 6-Methyl-2-(4-methylbenzamido)-4,5,6,7-tetrahydrothieno
J ¼ 7.2 Hz, 2H), 2.18 (s, 3H), 2.16e2.10 (m,1H),1.89e1.60 (m, 8H); 13
C
[2,3-c]pyridine-3-carboxamide (8e)
NMR (100 MHz, DMSO-d6)
d 169.9, 167.8, 161.6, 133.2, 132.7, 124.5,
MS (ESI) 330 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 9.47
114.9, 47.9, 46.4, 41.2(2C), 36.9, 31.5(2C), 26.5, 23.4. Anal. calcd for
16H21N3O3S: C, 57.29; H, 6.31; N, 12.53% Found C, 57.32; H, 6.42; N,
(s, 1H), 7.84e7.72 (m, 4H), 7.54 (d, J ¼ 8.0 Hz, 2H), 4.31 (s, 2H), 3.61
C
(t, J ¼ 7.6 Hz, 2H), 3.04 (t, J ¼ 7.6 Hz, 2H), 2.43 (s, 3H), 2.39 (s, 3H);
12.61%.
13C NMR (100 MHz, CDCl3)
d 179.4, 168.3, 148.2, 146.5, 139.0, 136.5,
130.6(2C), 128.4(2C), 126.3, 118.8, 49.8, 43.6, 38.2, 24.5, 21.9. Anal.
calcd for C17H19N3O2S: C, 61.98; H, 5.81; N, 12.76% Found C, 62.02;
H, 5.89; N, 12.81%.
4.1.15. 6-Acetyl-2-(cyclohexanecarboxamido)-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (7j)
ESI-MS showed 350 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d
9.97 (s, 2H), 7.72 (s, 1H), 4.66 (s, 2H), 3.97(t, J ¼ 8.0 Hz, 2H), 3.21 (t,
4.1.22. 6-Methyl-2-(4-phenoxybenzamido)-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (8f)
J ¼ 8.0 Hz, 2H), 2.14e2.09 (m, 4H), 1.98e1.53 (m, 10H); 13C NMR
(100 MHz, DMSO-d6)
d
176.7, 171.8, 166.6, 136.1, 130.8, 122.5, 116.6,
MS (ESI) 408 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 11.97
49.9, 44.8, 38.6(2C), 30.4(2C), 26.3, 24.4, 23.5, 21.4. Anal. calcd for
17H23N3O3S: C, 58.43; H, 6.63; N,12.02% Found C, 58.52; H, 6.73; N,
(s, 2H), 7.83 (d, J ¼ 8.4 Hz, 2H), 7.72e7.63 (m, 3H), 7.54e7.45 (m, 5H),
4.84 (s, 2H), 3.72 (t, J ¼ 8.4 Hz, 2H), 3.21 (t, J ¼ 8.4 Hz, 2H), 2.43 (s,
C
12.13%.
3H); 13C NMR (100 MHz, CDCl3)
d 183.1, 178.7, 148.2, 146.9,
136.4(2C), 136.0, 134.3, 132.0(2C), 129.5, 128.2(2C), 126.9(2C), 125.3,
124.6, 119.9, 52.9, 46.8, 36.9, 24.1. Anal. calcd for C22H21N3O3S: C,
64.85; H, 5.19; N, 10.31% Found C, 64.92; H, 5.21; N, 10.52%.
4.1.16. General procedure for the preparation of 8aej
40% formaldehyde solution (6 vol) was added to the stirred
solution of Compound 6aej in MeOH/H2O (9:1) and stirred for 1 h,
then Na(OAc)3BH (2.0 equiv) was added. The reaction mixture was
stirred at room temperature for 6 h. The product was extracted with
EtOAc and purified by triturating with CH2Cl2, Diethyl ether and
Hexanes to get title compounds 8aej.
4.1.23. 2-(1-Naphthamido)-6-methyl-4,5,6,7-tetrahydrothieno[2,3-
c]pyridine-3-carboxamide (8g)
MS (ESI) 366 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 9.99 (s,
2H), 8.22e8.10 (m, 2H), 7.89e7.72 (m, 6H), 4.54 (s, 2H), 3.79 (t,
J ¼ 8.0 Hz, 2H), 3.13 (t, J ¼ 7.6 Hz, 2H), 2.34 (s, 3H); 13C NMR
(100 MHz, CDCl3)
d 177.2, 168.4, 148.2, 139.3, 138.6, 138.0, 137.4,
4.1.17. 2-acetamido-6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]
135.8, 135.2, 134.5, 132.1, 129.8, 127.2, 126.3, 124.6, 121.1, 51.3, 44.8,
37.8, 22.7. Anal. calcd for C20H19N3O2S: C, 65.73; H, 5.24; N, 11.50%
Found C, 65.82; H, 5.32; N, 11.62%.
pyridine-3-carboxamide (8a)
MS (ESI) 254 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 10.53 (s,
1H), 8.01 (s, 2H), 4.12 (s, 2H), 3.44 (t, J ¼ 7.6 Hz, 2H), 3.15 (t,
J ¼ 8.0 Hz, 2H), 2.41 (s, 3H), 2.16 (s, 3H); 13C NMR (100 MHz, DMSO-
4.1.24. 2-(Cyclopropanecarboxamido)-6-methyl-4,5,6,7-
d6)
d
178.7,167.4,158.7,133.4,129.8,127.6, 51.3, 49.8, 42.7, 25.2, 22.5.
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (8h)
Anal. calcd for C11H15N3O2S: C, 52.15; H, 5.97; N, 16.59% Found C,
52.22; H, 6.02; N, 16.72%.
MS (ESI) 280 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 10.11
(s, 1H), 8.22 (s, 2H), 4.45 (s, 2H), 3.81 (t, J ¼ 8.4 Hz, 2H), 3.18
(t, J ¼ 8.0 Hz, 2H), 2.45 (s, 3H), 2.14e2.10 (m, 1H), 1.54e1.12 (m, 4H);
13C NMR (75 MHz, DMSO-d6)
d 176.4, 171.9, 144.4, 134.3, 132.3,
4.1.18. 2-Benzamido-6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]
120.9, 48.4, 43.6, 38.4, 22.5, 19.3, 18.7(2C). Anal calcd for
13H17N3O2S: C, 55.89; H, 6.13; N, 15.04% Found C, 55.92; H, 6.24; N,
pyridine-3-carboxamide (8b)
C
MS (ESI) 316 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 12.63
15.11%.
(s, 1H), 9.97 (s, 2H), 7.92e7.63 (m, 5H), 4.54 (s, 2H), 3.71 (t,
J ¼ 8.4 Hz, 2H), 3.13 (t, J ¼ 7.6 Hz, 2H), 2.52 (s, 3H); 13C NMR
4.1.25. 2-(Cyclopentanecarboxamido)-6-methyl-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (8i)
MS (ESI) 308 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
(75 MHz, DMSO-d6)
d 176.4, 170.5, 136.2, 129.5, 127.2(2C),
125.8(2C), 125.1, 124.4, 123.6, 115.4, 48.3, 42.6, 39.0, 21.8. Anal.
calcd for C16H17N3O2S: C, 60.93; H, 5.43; N, 13.32% Found C, 61.02;
H, 5.52; N, 13.42%.
d
10.21
(s, 2H), 8.67 (s, 1H), 4.64 (s, 2H), 3.86 (t, J ¼ 8.0 Hz, 2H), 3.12
(t, J ¼ 7.6 Hz, 2H), 2.38 (s, 3H), 2.11e2.0 (m, 1H), 1.69e1.34 (m, 8H);
13C NMR (100 MHz, DMSO-d6)
d 176.9, 169.8, 149.6, 132.2, 130.7,
4.1.19. 2-(2-Methoxybenzamido)-6-methyl-4,5,6,7-
122.5, 52.9, 49.4, 46.2, 39.9, 31.6(2C), 26.3(2C), 21.4. Anal. calcd for
C15H21N3O2S: C, 58.61; H, 6.89; N, 13.67% Found C, 58.72; H, 7.02; N,
13.71%.
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (8c)
MS (ESI) 346 [M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
9.45 (s, 2H),
7.81 (d, J ¼ 7.6 Hz, 1H), 7.72e7.54 (m, 4H), 4.61 (s, 2H), 3.90 (s, 3H),
3.69 (t, J ¼ 7.6 Hz, 2H), 3.21 (t, J ¼ 8.0 Hz, 2H), 2.46 (s, 3H); 13C NMR
4.1.26. 2-(Cyclohexanecarboxamido)-6-methyl-4,5,6,7-
tetrahydrothieno[2,3-c]pyridine-3-carboxamide (8h)
MS (ESI) 322 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
(100 MHz, DMSO-d6) d 178.4, 172.3, 146.9, 142.8, 134.4, 133.9, 132.3,
131.4, 128.4, 126.9, 125.1, 120.0, 60.1, 56.6, 52.9, 39.9, 23.6. Anal.
calcd for C17H19N3O3S: C, 59.11; H, 5.54; N,12.17% Found C, 59.22; H,
5.61; N, 12.16%.
d 10.45
(s, 1H), 7.74 (s, 2H), 4.58 (s, 2H), 3.87 (t, J ¼ 8.0 Hz, 2H), 3.19
(t, J ¼ 8.0 Hz, 2H), 2.61 (s, 3H), 2.16e2.09 (m, 1H), 1.89e1.44

116
R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
(m, 10H); 13C NMR (100 MHz, DMSO-d6)
d
171.2, 169.9, 132.1, 129.8,
CDCl3) d 170.2, 164.9, 152.2, 144.7, 138.3, 136.5, 134.0, 133.2, 130.5,
128.4,119.7, 61.0, 56.5, 47.8, 43.5, 31.6(2C), 28.4, 23.3(2C), 22.4. Anal.
calcd for C16H23N3O2S: C, 59.78; H, 7.21; N, 13.07% Found C, 59.82;
H, 7.29; N, 13.11%.
128.7, 126.9, 123.6, 54.9, 26.8, 25.4, 24.2(2C), 22.9, 22.3. Anal. calcd
for C19H22N2O3S: C, 63.66; H, 6.19; N, 7.82% Found C, 63.72; H, 6.21;
N, 8.01%.
4.1.27. Preparation of 2-amino-4,5,6,7,8,9-hexahydrocycloocta[b]
thiophene-3-carbonitrile (10)
4.1.33. 2-(3-Nitrobenzamido)-4,5,6,7,8,9-hexahydrocycloocta[b]
thiophene-3-carboxamide (12d)
MS (ESI) 374 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 9.21 (s,
To the stirred solution of cyclooctanone (9) (2.5 mL, 19.0 mmol),
malanonitrile (1.38 g, 20.9 mmol) and elemental sulphur (670 mg,
20.9 mmol) in DMF (10 mL) was added Et3N (4.0 mL, 28.5 mmol)
and stirred at room temperature for 9 h. The reaction mixture was
diluted with EtOAc (60 mL), the organic layer was washed with 1 M
HCl (2 ꢂ 20 mL) and brine (2 ꢂ 30 mL). The separated organic layer
was dried over anhyd. Na2SO4 and concentrated under vacuo to get
crude compound. The crude compound was triturated with CH2Cl2,
diethyl ether and hexanes to get compound 10 (2.70 g, 66%) as an
off-white solid. ESI-MS showed 207 [M þ H]þ. Anal. calcd for
2H), 9.00 (s, 1H), 8.39 (s, 1H), 7.99 (d, J ¼ 8.4 Hz, 1H), 7.83e7.69 (m,
2H), 2.81e2.39 (m, 4H),1.59e1.19 (m, 8H); 13C NMR (75 MHz, DMSO-
d6)
125.3, 121.4, 27.9, 27.2, 25.6(2C), 24.3, 22.5. Anal. calcd for
18H19N3O4S: C, 57.89; H, 5.13; N, 11.25% Found C, 57.92; H, 5.22; N,
d 183.8, 175.6, 148.6, 143.5, 138.8, 133.6, 132.6, 130.5, 128.6, 126.7,
C
11.34%.
4.1.34. 2-(4-Methylbenzamido)-4,5,6,7,8,9-hexahydrocycloocta[b]
thiophene-3-carboxamide (12e)
C
11H14N2S: C, 64.04; H, 6.84; N, 13.58% Found C, 64.12; H, 6.89; N,
MS (ESI) 343 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 11.56 (s,
13.64%.
2H), 8.01 (d, J ¼ 8.0 Hz, 2H), 7.84 (s, 1H), 7.72 (d, J ¼ 8.0 Hz, 2H), 2.52
(s, 3H), 2.70e2.29 (m, 4H), 1.87e1.36 (m, 8H); 13C NMR (100 MHz,
4.1.28. Preparation of 2-amino-4,5,6,7,8,9-hexahydrocycloocta[b]
thiophene-3-carboxamide (11)
DMSO-d6)
d 181.2, 173.9, 147.2, 138.4, 137.7(2C), 136.4, 135.3(2C),
127.3, 125.4, 121.3, 26.8, 25.2, 24.9, 24.3(2C), 22.5, 21.4. Anal. calcd
for C19H22N2O2S: C, 66.64; H, 6.48; N, 8.18% Found C, 66.72; H, 6.59;
N, 8.31%.
Compound 10 (2.70 g) was suspended in conc. H2SO4 (10 mL)
and stirred for 12 h. The reaction mixture was quenched with ice,
neutralised with solid Na2CO3, and the product was extracted with
EtOAc, and the combined organic layer was concentrated under
reduced pressure to get compound 11 (1.6 g, 54%) as an off-white
4.1.35. 2-(4-Phenoxybenzamido)-4,5,6,7,8,9-hexahydrocycloocta[b]
solid. MS (ESI) 225 [M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
6.55
thiophene-3-carboxamide (12f)
(bs, 2H), 6.10 (bs, 2H), 2.77e2.62 (m, 4H), 1.70e1.35 (m, 8H); Anal.
calcd for C11H16N2OS: C, 58.90; H, 7.19; N, 12.49% Found C, 58.94; H,
7.29; N, 12.60%.
MS (ESI) 421 [M þ H]þ. 1H NMR (300 MHz, DMSO-d6):
d 12.40 (s,
1H), 7.89 (d, J ¼ 8.7 Hz, 2H), 7.48 (s, 2H), 7.44 (t, J ¼ 7.8 Hz, 2H), 7.24
(t, J ¼ 7.5 Hz, 1H), 7.13 (d, J ¼ 8.4 Hz, 4H), 2.84e2.75 (m, 4H), 1.56e
1.42 (m, 6H), 1.19e1.17 (m, 2H); 13C NMR (75 MHz, DMSO-d6)
4.1.29. General procedure for the preparation of 12aej
To the stirred solution of ReCOOH (1.20 equiv) in CH2Cl2 was
added EDCI (1.30 equiv), HOBt (1.30 equiv) and Et3N (2.50 equiv)
while stirred the mixture for few minutes, then added compound
11 (1.0 equiv). The reaction mixture was stirred at room tem-
perature for 3 h. The reaction mixture was concentrated and
triturated with H2O to get solid compound. The solids were given
cold Ethanol, diethyl ether and hexane washings to get pure
products.
d 167.8, 162.0, 160.4, 155.2, 141.4, 131.9, 130.3(2C), 130.2(2C), 129.4,
127.1, 124.6, 119.8(2C), 117.9(2C), 117.7, 32.2, 29.8, 26.2, 25.6, 25.0,
24.3. Anal. calcd for C24H24N2O3S: C, 68.55; H, 5.75; N, 6.66% Found
C, 68.62; H, 5.81; N, 6.72%.
4.1.36. 2-(1-Naphthamido)-4,5,6,7,8,9-hexahydrocycloocta[b]
thiophene-3-carboxamide (12g)
MS (ESI) 379 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 8.34e
8.19 (m, 4H), 7.92e7.81 (m, 4H), 7.72e7.63 (m, 2H), 2.79e2.38 (m,
4H), 1.53e1.20 (m, 8H); 13C NMR (100 MHz, DMSO-d6)
d 179.2,
4.1.30. 2-Acetamido-4,5,6,7,8,9-hexahydrocycloocta[b]thiophene-
3-carboxamide (12a)
166.3, 146.3, 139.6, 138.2, 137.6, 137.2, 136.0, 135.3, 133.9, 133.2,
130.5, 129.4, 126.3, 122.6, 118.7, 28.9, 27.6, 26.2(2C), 24.2, 19.6. Anal
calcd for C22H22N2O2S: C, 69.81; H, 5.86; N, 7.40% Found C, 69.92; H,
5.94; N, 7.62%.
MS (ESI) 267 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 8.22 (s,
2H), 7.47 (s, 1H), 2.73e2.50 (m, 4H), 2.23 (s, 3H), 1.60e1.35 (m, 8H);
13C NMR (100 MHz, DMSO-d6)
123.4, 29.7, 27.3, 25.4, 24.8(2C), 23.6, 22.5. Anal. calcd for
13H18N2O2S: C, 58.62; H, 6.81; N,10.52% Found C, 58.69; H, 6.84; N,
d 169.4, 166.9, 149.4, 135.3, 133.3,
4.1.37. 2-(Cyclopropanecarboxamido)-4,5,6,7,8,9-
C
hexahydrocycloocta[b]thiophene-3-carboxamide (12h)
10.61%.
MS (ESI) 293 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
d 11.45 (s,
2H), 6.77 (s, 1H), 2.79e2.46 (m, 4H), 2.20e2.07 (m, 1H), 1.69e1.12
4.1.31. 2-Benzamido-4,5,6,7,8,9-hexahydrocycloocta[b]thiophene-
(m, 12H); 13C NMR (100 MHz, DMSO-d6)
d 176.4, 171.9, 156.4, 136.3,
3-carboxamide (12b)
133.3, 121.1, 28.7, 27.2, 26.4(2C), 24.8, 22.3, 21.4, 18.7(2C). Anal. calcd
for C15H20N2O2S: C, 61.62; H, 6.89; N, 9.58% Found C, 61.69; H, 6.94;
N, 9.71%.
MS (ESI) 329 [M þ H]þ. 1H NMR (300 MHz, CDCl3):
d 7.92 (s, 2H),
7.81e7.62 (m, 6H), 2.64e2.36 (m, 4H), 1.71e1.41 (m, 8H); 13C NMR
(75 MHz, CDCl3)
d 178.2, 169.9, 146.2, 139.7, 136.3(2C), 134.4(2C),
129.3, 127.6, 126.7, 120.8, 27.8, 26.9, 26.3, 24.5(2C), 22.9. Anal. calcd
for C18H20N2O2S: C, 65.83; H, 6.14; N, 8.53% Found C, 65.99; H, 6.24;
N, 8.71%.
4.1.38. 2-(Cyclopentanecarboxamido)-4,5,6,7,8,9-
hexahydrocycloocta[b]thiophene-3-carboxamide (12i)
MS (ESI) 321 [M þ H]þ. 1H NMR (300 MHz, CDCl3):
d 8.21 (s, 2H),
4.1.32. 2-(2-Methoxybenzamido)-4,5,6,7,8,9-hexahydrocycloocta
7.27 (s, 1H), 2.73e2.49 (m, 4H), 2.01e1.93 (m, 1H), 1.80e1.33 (m,
[b]thiophene-3-carboxamide (12c)
16H); 13C NMR (75 MHz, CDCl3)
d 178.2, 166.5, 152.1, 136.6, 128.7,
MS (ESI) 359 [M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
11.25 (s,
120.4, 44.8, 33.3(2C), 29.5, 27.2(2C), 25.2, 24.6(2C), 23.4, 22.5. Anal.
calcd for C17H24N2O2S: C, 63.72; H, 7.55; N, 8.74% Found C, 63.82; H,
7.72; N, 8.82%.
1H), 7.90 (d, J ¼ 8.0 Hz, 1H), 7.81 (s, 1H), 7.72e7.63 (m, 4H), 3.81 (s,
3H), 2.81e2.49 (m, 4H), 1.62e1.39 (m, 8H); 13C NMR (100 MHz,

R. Nallangi et al. / European Journal of Medicinal Chemistry 76 (2014) 110e117
117
4.1.39. 2-(Cyclohexanecarboxamido)-4,5,6,7,8,9-
hexahydrocycloocta[b]thiophene-3-carboxamide (12j)
MS (ESI) 335 [M þ H]þ. 1H NMR (400 MHz, DMSO-d6):
into a formazan product using the Promega Cell Titer 96 non-
radioactive cell proliferation assay.
d
7.27 (s,
2H), 6.73 (s, 1H), 2.74e2.41 (m, 4H), 2.18e2.10 (m, 1H), 1.92e1.24
Acknowledgements
(m, 18H); 13C NMR (100 MHz, CDCl3)
d 180.3, 171.9, 149.2, 133.3,
129.6, 119.6, 48.9, 29.8, 27.4, 25.2, 24.9(2C), 22.9(2C), 22.0(2C), 21.6,
20.5. Anal. calcd for C18H26N2O2S: C, 64.64; H, 7.84; N, 8.38% Found
C, 64.72; H, 7.99; N, 8.52%.
The authors are thankful to Department of Science & Technology
(SR/S1/OC-70/2010), New Delhi, India for their financial assistances.
One of the authors RN acknowledges UGC for providing JRF.
4.2. Biological evaluation
Appendix A. Supplementary data
4.2.1. In vitro MTB screening
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.02.028.
Two-fold serial dilutions of each test compound/drug were
prepared and incorporated into Middle-brook 7H11 agar medium
with oleic acid, albumin, dextrose, and catalase (OADC) growth
supplement to get final concentrations of 50, 25, 12.5, 6.25, 3.13,
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