Discovery of novel antitubercular 3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2- carboxamide/carbothioamide analogues

Article abstract of DOI:10.1016/j.bmcl.2011.07.035

In the present investigation, a series of 3a,4-dihydro-3H-indeno[1,2-c] pyrazole-2-carboxamide/carbothioamide analogues were synthesized and were evaluated for antitubercular activity by two fold serial dilution technique. All the newly synthesized compounds showed low to high inhibitory activities against Mycobacterium tuberculosis H₃₇Rv and INH resistant M. tuberculosis. The compound 3-(4-fluorophenyl)-6,7-dimethoxy-3a,4-dihydro-3H- indeno[1,2-c]pyrazole-2-carbothioamide (4o) was found to be the most promising compound active against M. tuberculosis H₃₇Rv and isoniazid resistant M. tuberculosis with minimum inhibitory concentration 3.12 μM and 6.25 μM, respectively.

Full text of DOI:10.1016/j.bmcl.2011.07.035

Bioorganic & Medicinal Chemistry Letters 21 (2011) 5259–5261
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of novel antitubercular 3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2-
carboxamide/carbothioamide analogues
Mohamed Jawed Ahsan a,b, , Jeyabalan Govinda Samy a, Savita Soni a, Naresh Jain a, Lalit Kumar a,
⇑
Lalit K. Sharma a, Hemant Yadav a, Lokesh Saini a, Rajput Gopal Kalyansing a, Narendra S. Devenda a,
Ravindra Prasad a, Chandra Bhushan Jain c
a New Drug Discovery Research, Department of Medicinal Chemistry, Alwar Pharmacy College, Alwar, Rajasthan 301 030, India
b Department of Pharmaceutical Sciences, National Institute of Medical Sciences University, Jaipur 303 121, India
c Academic, Rajasthan University of Health Sciences, Jaipur 302 022, India
a r t i c l e i n f o
a b s t r a c t
Article history:
In the present investigation, a series of 3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2-carboxamide/carbo-
thioamide analogues were synthesized and were evaluated for antitubercular activity by two fold serial
dilution technique. All the newly synthesized compounds showed low to high inhibitory activities against
Mycobacterium tuberculosis H37Rv and INH resistant M. tuberculosis. The compound 3-(4-fluorophenyl)-
6,7-dimethoxy-3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2-carbothioamide (4o) was found to be the most
promising compound active against M. tuberculosis H37Rv and isoniazid resistant M. tuberculosis with
Received 9 June 2011
Revised 6 July 2011
Accepted 8 July 2011
Available online 19 July 2011
Keywords:
Pyrazolines
Antitubercular agents
minimum inhibitory concentration 3.12
l
M and 6.25
lM, respectively.
Ó 2011 Elsevier Ltd. All rights reserved.
Mycobacterium tuberculosis infections are responsible for one in
four preventable adult deaths in developing countries. The current
strategies for the treatment of tuberculosis (TB) are very compli-
cated as it takes several months of chemotherapy to eliminate per-
sistent bacteria. Also widespread non-compliance has contributed
to the emergence of multidrug-resistant (MDR) and extensively
drug-resistent (XRD) TB.1,2 There are estimated 1.3 million multi/
extensively drug resistant TB (M/XDR-TB) cases will need to be
treated between 2010 and 2015.3 Hence there is a need for fast act-
ing drugs that are capable of eliminating an infection just in a few
weeks.
In an effort to discover new and effective therapeutic agents, we
recently reported the in vitro antimycobacterial activity of novel
diketones and pyrazoline derivatives.4,5 Pyrazoline is a class of het-
erocyclic compounds possess significant pharmacological activities
including anticancer, antitubercular, anticonvulsant, antiamoebic,
antimicrobial, anti-inflammatory, antiviral, antiarrhythmic,
antidepressant, antidiabetic, etc.6–15 In the current work we have
synthesized eighteen 3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2-
carboxamide/carbothioamide analogues based on the structure of
the known antitubercular agent, thiacetazone (Fig 1). All the synthe-
sized compounds were evaluated for their anti-tubercular activities.
The 3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2-carboxamide/car
bothioamide analogues (4a–r) described in this study are shown
in Table 1 and the reaction sequence for the synthesis is summa-
rised in Scheme 1. In the initial step 5,6-dimethoxy-2,3-dihydro-
1H-inden-1-one (0.1 mol) and appropriate aromatic aldehydes
(0.1 mol) in diluted methanolic sodium hydroxide solution stirred
at room temperature giving the (2E)-2-substituted-5,6-dime-
thoxy-2,3-dihydro-1H-indene-1-one derivatives (3a–f). In the
subsequent step 2-substituted-5,6-dimethoxy-2,3-dihydro-1H-in-
dene-1-one derivatives treated with appropriate semicarbazides
furnished the titled compounds (4a–r). The substituted phenyl
semicarbazides were synthesized as per the reported method.16,17
The yields of the titled compounds were ranging from 66% to 87%
after recrystallization with absolute ethanol. The purity of the com-
pounds was checked by TLC using eluant benzene–acetone (9:1)
and elemental analyses. Melting points were determined by open
tube capillary method and are uncorrected. Purity of the com-
pounds was checked on TLC plates (silica gel G) using eluants ben-
zene–acetone (9:1), the spots were located under iodine vapours or
UV light. The entire chemicals were supplied by E. Merck (Ger-
many) and S.D. Fine Chemicals (India). IR spectra were obtained
on a Schimadzu 8201 PC, FT-IR spectrometer (KBr pellets). 1H
NMR spectra were recorded on a Bruker AC 300 MHz spectrometer
using TMS as internal standard in DMSO. Mass spectra were re-
corded on a Bruker Esquire LCMS using ESI and elemental analyses
were performed on Perkin–Elmer 2400 Elemental Analyzer. Both
the analytical and spectral data (IR, 1H NMR) of all the synthesized
compounds were in full accordance with the proposed structures.
In general, the IR spectra of the compounds afforded pyrazoline
C@N stretching at 1501–1576 cmÀ1, C–H deformation at 1362–
1464 cmÀ1, C2–N1 stretching at 1069–1189 cmÀ1, and carbamoyl
⇑
Corresponding author. Tel.: +91 9694087786; fax: +91 144 5121120.
E-mail address: jawedpharma@gmail.com (M.J. Ahsan).
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.07.035

5260
M. J. Ahsan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 5259–5261
Isosteric replacement of 'S' with 'O'
S
S
O
Ar2
N
NH2
N
N
N
N
NH2
N
N
H
O
O
H
O
O
O
Ar1
Title compounds (4a-l)
Ar1
H3C
N
H
Thiacetazone
Title compounds (4m-r)
Figure 1. Design of the title compounds based on known antitubercular drug.
Table 1
Physical constant and antimycobacterial activity of the synthesized compounds
O
S
Ar2
N
N
N
NH2
Ar1
N
N
O
O
O
1H
Ar
O
4a-4l
Ar2
4m-4r
Compound
Ar1
Yield (%)
Mp (°C)
MIC (lM)
MTBa
MTBb
4a
4b
4c
4d
4e
4f
4g
4h
4i
4j
4k
4l
4m
4n
4o
4p
4q
4r
4-Pyridinyl-
2,4-Dimethylphenyl-
2,4-Dimethylphenyl-
2,4-Dimethylphenyl-
2,4-Dimethylphenyl-
2,4-Dimethylphenyl-
2,4-Dimethylphenyl-
2,6-Dimethylphenyl-
2,6-Dimethylphenyl-
2,6-Dimethylphenyl-
2,6-Dimethylphenyl-
2,6-Dimethylphenyl-
2,6-Dimethylphenyl-
–
–
–
–
–
–
–
–
78
72
68
84
67
79
82
87
85
73
66
69
82
84
68
66
76
75
–
176
186
180
188
150
124
190
160
178
174
148
164
194
202
212
262
232
104
–
10
>12.5
10
2-Chlorophenyl-
4-Fluorophenyl-
3,4-Dimethoxyphenyl-
Phenyl-
4-Methoxyphenyl-
4-Pyridinyl-
2-Chlorophenyl-
4-Fluorophenyl-
3,4-Dimethoxyphenyl-
Phenyl-
4-Methoxyphenyl-
4-Pyridinyl-
2-Chlorophenyl-
4-Fluorophenyl-
3,4-Dimethoxyphenyl-
Phenyl-
4-Methoxyphenyl-
–
6.25
6.25
12.5
>12.5
>12.5
>10
6.25
6.25
12.5
12.5
>12.5
6.25
6.25
3.12
12.5
12.5
12.5
0.78
0.006
12.5
>12.5
>12.5
>12.5
>12.5
12.5
10
>12.5
>12.5
>12.5
10
10
6.25
12.5
12.5
>12.5
12.5
3.12
INH
Rifampin
–
–
–
a
M. tuberculosis H37Rv.
INHR-TB.
b
CH
O
3
CH
O
multiplicities and coupling constants in DMSO-d6. The spectra
showed singlet at d 2.21–2.37 ppm corresponding to CH3; triplet
at d 3.20–3.57 ppm corresponding to CH; a doublet at d 3.43–
3.48 ppm corresponding to CH2 group; a singlet at d 3.79–
3.83 ppm corresponding to OCH3 group; a doublet at 4.9–5.4 ppm
corresponding to CH; a broad singlet at 5.2–5.6 ppm corresponding
to NH2; multiplet at d 6.51–8.38 ppm corresponding to aromatic
protons; broad singlet at d8.88–11.52 ppm corresponding to CONH.
The elemental analysis results were within 0.4% of the theoretical
values.
All the synthesized compounds (4a–r) were tested for their
in vitro antimycobacterial activity against MTB and INHR-MTB by
agar dilution method using double dilution technique similar to
that recommended by the National Committee for Clinical Labora-
tory Standards.18 The MTB and INHR-MTB clinical isolate was
obtained from Tuberculosis Research Center, Alwar, India. The
MIC was defined as the minimum concentration of compound re-
quired to inhibit 90% of bacterial growth and MIC of the com-
pounds were reported in Table 1 with standard drug INH and
rifampin for comparison.
O
3
O
CH OH/NaOH
3
1
+
Ar CHO
H C
1
H C
3
3
O
Ar
O
2a-f
1
3a-f
2
Ar NHCXNHNH
AcOH
2
X
2
Ar
CH
O
3
N
N
N
H
1
Ar
H C
3
O
4a-r
4m-r X = S
4a-l X = O
Scheme 1. Protocol for the synthesis.
group N–H stretching at 3112–3481 cmÀ1 and C@O stretching at
1675–1685 cmÀ1 bands. In the Nuclear Magnetic Resonance spectra
(1H NMR) the signals of the respective protons of the synthesized
titled compounds were verified on the basis of their chemical shift,
Among the 18 synthesized compounds, seven compounds were
found to be active with minimum inhibitory concentration of

M. J. Ahsan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 5259–5261
5261
3.12–6.25
MTB and INH-MTB at a MIC of 3.12
l
M. The compound 4o was found to be active against
M and 6.25 M, respectively.
References and notes
l
l
1. Zignol, M.; Hosseini, M. S.; Wright, A.; Weezenbeek, C. L.; Nunn, P.; Watt, C. J.;
Williams, B. G.; Dye, C. J. Infect. Dis. 2006, 194, 479.
2. Wells, C.; Cegielski, J. P. Emerging Infect. Dis. 2007, 13, 380.
3. WHO report 2010, Global Tuberculosis Control Surveillance, Financing and
Planning.
4. Ali, M. A.; Samy, G. J.; Manogaran, E.; Sellappan, V.; Hassan, M. Z.; Ahsan, M. J.;
Pandian, S.; ShaharYar, M. Bioorg. Med. Chem. Lett. 2009, 19, 7000.
5. Ahsan, M. J.; Samy, G. J.; Dutt, K. R.; Agrawal, U. K.; Shankar, B.; Vyas, S.; Kaur,
R.; Yadav, G. Bioorg. Med. Chem. Lett. 2011, 21, 4451.
6. Budakoti, A.; Bhat, A. R.; Athar, F.; Azam, A. Eur. J. Med. Chem. 2008, 43, 1749.
7. Parmar, S. S.; Pandey, B. R.; Dwivedi, C.; Harbison, R. D. J. Pharm. Sci. 1974, 63,
1152.
8. Dawane, B. S.; Konda, S. G.; Mandawad, G. G.; Shaikh, B. M. Eur. J. Med. Chem.
2010, 45, 387.
9. Sharma, P. K.; Kumar, S.; Kumar, P.; Kaushik, P.; Kaushik, D.; Dhingra, Y.; Aneja,
K. R. Eur. J. Med. Chem. 2010, 45, 2650.
10. Bilgin, A. A.; Palaska, E.; Sunal, R. Drug Res. 1993, 43, 1041.
11. Turan-Zitouni, G.; Chevallet, P.; Kiliç, F. S.; Erol, K. Eur. J. Med. Chem. 2000, 35,
635.
12. Mui, M. S.; Siew, B. N.; Buss, A. D.; Crasta, S. C.; Kah, L. G.; Sue, K. L. Bioorg. Med.
Chem. Lett. 2002, 12, 679.
When compared with INH the compound, 4o was four folds less ac-
tive than INH against MTB, while compound, 4o was two folds more
active than INH against INHR-MTB. In the title compounds (4a–r),
both the C3 aryl group and the N-aryl group influenced the antitu-
bercular activity. The 3-substituted compounds with electron with-
drawing groups such as 4-flourophenyl produced more inhibitory
activity than 2-chlorophenyl and 2-pyridyl substitution, while the
electron releasing groups such as 4-methoxyphenyl, 3,4-dime-
thoxyphenyl showed less inhibitory activity. N-Aryl substitution
with electron releasing group showed decreased activity when
compared with reported earlier screening.5 Also substitution of
‘O’ with ‘S’ produced more pronounced activity in the series of title
compounds (4a–4r), might be due to increased lipophilicity. The
compounds 4b, 4c, 4h, 4i, 4m, 4n and 4a showed good to moderate
inhibitory activity against MTB at MIC 6.25
All the active compounds were tested for cytotoxicity (IC50) in
VERO cells at concentrations of 62.5 g/mL or 10 times the MIC.
lM and 10 lM.
13. Manna, F.; Chimenti, F.; Fioravati, R.; Bolasco, A.; Secci, D.; Chimenti, P.; Ferlini,
C.; Scambia, G. Bioorg. Med. Chem. Lett. 2005, 15, 4632.
14. Ali, M. A.; Shaharyar, M.; Siddique, A. A. Eur. J. Med. Chem. 2007, 42,
268.
l
After 72 h 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 method. Most of the ac-
15. Gokhan, N.; Yesßilada, A.; Uçar, G.; Erol, K.; Bilgin, A. A. Arc. Phar. 2003, 336,
362.
tive compounds were found to be non-toxic up to 62.5 l
g/mL.19
16. Pandeya, S. N.; Raja, A. S.; Stables, J. P. J. Pharm Pharm. Sci. 2002, 5, 266.
17. Amir, M.; Ahsan, M. J.; Ali, I. Ind. J. of Chem. 2010, 49B, 1509.
18. Heifets, L. B.; Flory, M. A.; Lindholm-Levy, P. J. Antimicrob. Agents Chemother.
1989, 33, 1252.
All these derivatives can be further modified to exhibit more po-
tency. Further studies to acquire more information about Quantita-
tive Structure Activity Relationships (QSAR) and MDR are in
progress in our laboratory. The pyrazoline derivatives discovered
in this study may provide valuable therapeutic intervention for
the treatment of tubercular disease.
19. Gundersen, L. L.; Nissen-Meyer, J.; Spilsberg, B. J. Med. Chem. 2002, 45, 1383.
Compound
(4o):
5,6-dimethoxy-2,3-dihydro-1H-inden-1-one
(1.92 g,
0.001 mol) with 4-fluorobenzaldehyde (1.24 g, 0.001 mol) in diluted
methanolic sodium hydroxide solution was stirred under room temperature
for 4 h. The reaction mixture was then poured in crushed ice neutralized with
diluted HCl, giving the (2E)-2-(4-fluorobenzylidene)-5,6-dimethoxy-2,3-
dihydro-1H-indene-1-one which was then refluxed with thiosemicarbazide
in glacial acetic acid for 12 h. Excess of solvent was removed and the reaction
mixture was poured in crushed ice then filtered and washed furnished 3-(4-
fluorophenyl)-6,7-dimethoxy-3a,4-dihydro-3H-indeno[1,2-c]pyrazole-2-
carbothioamide (4o). IR (KBr) cmÀ1: 3364 (NH), 1680 (C@O), 1579 (C@N), 1389
(CH), 1180 (C–N), 789 (C–F); 1H NMR (DMSO-d6) ppm: 3.23–3.35 (1H, t, CH),
3.41–3.45 (2H, d, J = 6.3 Hz, CH2), 3.81 (6H, s, OCH3), 5.2 (1H, d, J = 6.1 Hz, CH),
5.4 (2H, br s, NH2), 6.88–8.16 (6H, m, Ar); Mass (m/z) 371 (M+); Cacld/Anal. [C
(61.44) 61.43, H (4.88) 4.87, N (11.31) 11.32].
Acknowledgments
Authors are thankful to the management people of Alwar Phar-
macy College, Alwar, Rajasthan, India for providing research facili-
ties. The authors are also thankful to Dr. K.P. Singh and Dr. (Mrs.)
Shobha Tomar, National Institute of Medical Sciences University
for their guidance.