Design, synthesis, and biological evaluation of 4-(5-nitrofuran-2-yl)prop- 2-en-1-one derivatives as potent antitubercular agents

Article abstract of DOI:10.1016/j.bmcl.2010.08.127

Based on stereoelectronic feature analysis using density functional theory (DFT) at B3LYP/3-21G level, a series of 4-(5-nitrofuran-2-yl)prop-2-en-1-one derivatives with low LUMO energies (<-0.10 eV); concentrated over the nitro group, furan moiety and α,β-unsaturated carbonyl bridge were envisaged as potential antitubercular agents. The target compounds were prepared by condensation of 5-nitro-2-furaldehyde with various ketones under acidic condition. The compounds were evaluated for antitubercular activity against Mycobacterium tuberculosis H37Rv and their cytotoxicity in VERO cell line. Several synthesized compounds showed good antitubercular activity of <5 μM along with low cytotoxicity. In particular, compound ((E)-3-(5-nitrofuran-2-yl) -1-(4-(piperidin-1-yl)phenyl)prop-2-en-1-one) (3v) was found to be very potent (MIC: 0.19 μM) with good selectivity index (MIC₉₀/CC₅₀: >1800). Thus, this study shows the potential of stereoelectronic property analysis in developing improved nitroaromatics as antitubercular agents.

Full text of DOI:10.1016/j.bmcl.2010.08.127

Bioorganic & Medicinal Chemistry Letters 20 (2010) 6175–6178
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis, and biological evaluation of 4-(5-nitrofuran-2-yl)prop-
2-en-1-one derivatives as potent antitubercular agents
Nilesh R. Tawari a, Ranjeet Bairwa a, M. K. Ray b, M. G. R. Rajan b, Mariam S. Degani a,
⇑
a Department of Pharmaceutical Sciences and Technology, Institute of Chemical Technology, Matunga (E), Mumbai 400019, India
b Radiation Medicine Center, Bhabha Atomic Research Centre, Tata Memorial Hospital Annex, Parel, Mumbai 400012, India
a r t i c l e i n f o
a b s t r a c t
*
Article history:
Received 27 March 2010
Revised 29 July 2010
Accepted 27 August 2010
Available online 16 September 2010
Based on stereoelectronic feature analysis using density functional theory (DFT) at B3LYP/3-21 G level, a
series of 4-(5-nitrofuran-2-yl)prop-2-en-1-one derivatives with low LUMO energies (<À0.10 eV); concen-
trated over the nitro group, furan moiety and a,b-unsaturated carbonyl bridge were envisaged as potential
antitubercular agents. The target compounds were prepared by condensation of 5-nitro-2-furaldehyde
with various ketones under acidic condition. The compounds were evaluated for antitubercular activity
againstMycobacterium tuberculosis H37Rv and their cytotoxicity in VERO cell line. Several synthesized com-
Keywords:
4-(5-Nitrofuran-2-yl)prop-2-en-1-one
Antitubercular
Molecular electrostatic profile (MESP)
Lowest unoccupied molecular orbital
(LUMO)
pounds showed good antitubercular activity of <5
((E)-3-(5-nitrofuran-2-yl)-1-(4-(piperidin-1-yl)phenyl)prop-2-en-1-one) (3v) was found to be very potent
(MIC: 0.19 M) with good selectivity index (MIC90/CC50: >1800). Thus, this study shows the potential of ste-
lM along with low cytotoxicity. In particular, compound
l
reoelectronic property analysis in developing improved nitroaromatics as antitubercular agents.
Ó 2010 Elsevier Ltd. All rights reserved.
Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb), a
obligate pathogen remains one of the most fatal infectious dis-
ease.1,2 World Health Organization (WHO) estimates that1.3 million
people died due to TB in 2008.3 Despite elucidation of the genomic
sequence of Mtb and emergence of multiple targets, TB continues
to be a serious public-health problem at the global level.4–7 Hence,
there is a need to develop novel therapeutics to treat TB.
Currently, two molecules PA-824 and OPC-67683 belonging to
the nitroimidazole class, having good in vitro and in vivo activity
against Mtb in both its active and persistent forms, are being eval-
uated in phase II clinical trials to treat tuberculosis.8 Recently, con-
siderable research is focused on understanding the mechanistic
details of this class of molecules.9 It is proposed that the release
of toxic intermediates, formed by the one electron and two elec-
tron bioreduction of the nitroimidazole moiety is the key compo-
nent responsible for the high activity of these molecules.10 Two
series of nitrofurans have also been reported as antitubercular
agents,11 while other nitrofurans are reported to be antibacterial
agents.13
Becke’s three-parameter exchange potential and the Lee–Yang–Parr
correlation functional (B3LYP) using basis set 3-21G , on the re-
*
ported antitubercular nitrofurans12 suggested that all the very ac-
tive compounds possess characteristic localized negative potential
regions near both the oxygen atoms of nitro group which extend lat-
erally to the isoxazole ring system/amide bond.14 Furthermore,
higher negative values of LUMO energies located over the nitro
group in the most active compounds were indicative of the electron
acceptor capacity of the compounds, suggesting that these com-
pounds are prodrugs and must be activated by TB-nitroreductase.
Thus, this study indicated that compounds with good antitubercular
activity could be designed by variation in LUMO energies that can be
affected by changing the electronic features of the nitro heterocycle.
It is worthy to note that, recently, while current work was under pro-
gress in our laboratory, Valdez et al. reported that molecules having
olefins with a conjugated bridge connecting the nitro heterocycle
and a substituted phenyl or heterocyclic ring showed a great in-
crease in antigiardial activityand lower toxicity as compared to met-
ronidazole in a murine giardiasis model.15
Interactions between molecules are the consequence of their ste-
reoelectronic properties that govern strength of bonds, strength of
nonbonded interactions and molecular reactivity. Molecular elec-
trostatic profile (MESP) is a very useful tool in understanding the
chemical reactivity of molecules. Pharmacophore mapping and
MESP analysis employing hybrid density functional theory with
Considering the apparent importance of nitro reduction in the
activation of nitroaromatic compounds, compounds were designed
by coupling electronegative a,b-unsaturated carbonyl bridge to the
nitrofuran ring and substituted phenyl or heterocyclic ring to
minimize LUMO energies (Fig. 1). The designed compounds were
studied by complete geometry optimization at DFT level of theory
*
*
using the B3LYP functional and 3-21 G basis set (B3LYP/3-21 G).
All calculations were carried in aqueous environment using the
Poisson–Boltzmann solver as implemented in Jaguar.14,16
⇑
Corresponding author. Tel.: +91 22 33612213; fax: +91 22 24145414.
E-mail address: ms.degani@ictmumbai.edu.in (M.S. Degani).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.08.127

6176
N. R. Tawari et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6175–6178
Figure 1. Chemical structures of the compounds synthesized.
Corroborating with our design postulate, DFT results showed that
all compounds posses localized negative potential regions near
the oxygen atom of carbonyl group and both the oxygen atoms
of nitro group that extend laterally to the furan ring oxygen
(Fig. 2a and b). Furthermore, the compounds have smaller values
of LUMO energies (<À0.1 eV) located over the nitro group, furan
matic substitutions and changing the aromatic moiety with hetero-
aromatic moiety; using acid catalyzed reaction (Scheme 1),17 as the
starting material, 5-nitro-2-furaldehyde is alkali sensitive and
standard Claisen–Schmidt condensation under basic condition
could not be used. The purity of the compounds was checked by
HPLC and the structures were confirmed by spectral data (FT-IR,
1H NMR, 13C NMR, and MS). The synthesized compounds (3a–k)
were screened against Mtb H37Rv using twofold dilution, in order
to determine the actual minimum inhibitory concentration (MIC)
with Resazurin microtiter assay (REMA).18,19 MIC is the lowest con-
centration at which complete inhibition was observed and was
ring and
Fig. 2c).
a,b-unsaturated carbonyl bridge attached to it (Table 1,
Thereafter, these compounds were considered for synthesis and
biological activity evaluation against Mtb. Initially, compounds,
3a–k (Fig. 1) were synthesized as exploratory set, with various aro-

N. R. Tawari et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6175–6178
6177
Figure 2. (a) MESP (À75 to 110 kcal/mol) superimposed onto a surface of constant electron density (0.01 e/au3) for compound 3a showing the most positive potential region
(deepest blue color) by the protons of C3 and C4 of the furan ring and protons of a,b-unsaturated system and the most negative potential region (deepest red color) by the
carbonyl oxygen and oxygen atoms of the nitro group extending laterally over the oxygen atom of furan ring. (b) Three-dimensional isopotential contours of MESP at
À30 kcal/mol for compound 3a showing negative potential regions around the carbonyl oxygen and oxygen atoms of the nitro group extending laterally over the oxygen atom
of furan ring. (c) LUMO energy distribution for compound 3a. (d) Superimposition of compound 3a (violet color) and OPC67683 (cyan color).
Table 1
determined by visual inspection (color change from blue to pink)
(Table 1). Isoniazid (INH) was used as the reference drug. The com-
pounds were also evaluated for toxicity in a mammalian VERO cell
Results of antitubercular activity and cytotoxicity values for the synthesized
compounds, along with their calculated electronic properties
a
Sr.
no.
Code MW
MICa
CC50
HOMO
(eV)
LUMO
(eV)
line (C1008) in 96-well microtitre plates and the CC50 values were
determined.19
(lM)
(lM)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
3a
3b
4
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
3o
3p
3q
3r
243.21
261.21
216.21 >93.89
5.51
4.17
444.06
463.23
661.39
439.28
558.24
353.92
570.93
416.53
384.05
742.26
776.22
359.84
515.26
430.13
311.05
339.53
307.47
334.62
392.59
299.43
342.74
334.03
349.32
386.82
403.62
À0.25
À0.25
À0.23
À0.25
À0.25
À0.25
À0.24
À0.25
À0.22
À0.25
À0.25
À0.25
À0.24
À0.24
À0.22
À0.24
À0.24
À0.24
À0.24
À0.20
À0.20
À0.20
À0.20
À0.20
À0.20
ND
À0.12
À0.12
À0.09
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.12
À0.11
À0.12
À0.12
ND
Among the synthesized compounds, 3a–k; five compounds
showed good antitubercular activity of <5 lM. The p-methoxy
substituted compound, 3f, was found to be the most active com-
pound. It was observed that there was little difference in activity
between unsubstituted and p-monosubstituted aromatic ring in
compounds, 3a–f. Di- and tri-substitution led to lower activity as
seen in compounds, 3g,h. Replacing the aromatic ring by heteroar-
omatic moiety, 3i–k, led to decrease in activity. To evaluate the ef-
fect of nitro group on the activity; des-nitro derivative, 4, of
compound, 3b, was synthesized and evaluated. The des-nitro com-
257.24
277.66
322.11
273.24
312.10
333.29
249.24
233.18
311.25
349.34
367.33
379.36
4.35
4.50
3.88
3.84
14.13
6.06
5.86
8.28
29.94
3.29
19.79
1.85
pound was found to be inactive at high concentration (ꢀ90
lM),
thus, indicating the importance of nitro group for the activity of
these compounds. This observation is in accordance with the re-
cently published results on PA-824 and OPC-67683, where the cor-
responding des-nitro derivatives were devoid of antitubercular
activity.20,21
418.23 >46.63
383.78
433.33
313.30
417.46 >31.74
443.49 >32.70
314.34
326.35
328.32
344.38
137.10
6.10
1.66
5.59
3s
3t
To further modify and enhance the potency of these compounds,
the unsubstituted compound, 3a, was aligned on the structure of
OPC-67683 using PHASE ( Fig. 2d).22 Alignment studies revealed
the scope for introduction of a substituted benzyloxy moiety at the
para-position. In this light, compounds, 3l–q, were synthesized
(Scheme 1). Commercially unavailable substituted acetophenone
derivatives required for the synthesis of 3l–q were prepared by
etherification of p-hydroxyacetophenone with corresponding
3u
3v
3w
3x
INH
0.48
0.19
0.38
15.62
1.80
>500.0
ND—not determined.
a
Results of three independent experiments performed in duplicate.
Scheme 1. Synthesis of 4-(5-nitrofuran-2-yl)prop-2-en-1-one derivatives. Reagents and conditions: acetic acid, catalytic H2SO4, 60–100 °C, 3–6 h.

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N. R. Tawari et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6175–6178
benzyl halide in presence of K2CO3 using CH3CN as solvent.23 Within
these molecules, compounds with p-methoxy, 3n (MIC: 1.85 M)
and p-trifluromethoxy, 3q (MIC: 1.66 M) substitution showed
good activity. It is interesting to note that both PA-824 and OPC-
67683 posses p-trifluromethoxy substitution in the distal part of
their structures. The compound, 3r, with a flexible methyallyloxy
substitution at the distal part was found to be less active than the
compound with a distal aromatic ring, 3l.
References and notes
l
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l
Results of Tangallapally et al.24 study and OPC-67683 SAR
revealed the importance of various tertiary amines as important fea-
tures responsible for potent antitubercular activity of nitroaromat-
ics. In this light, compounds, 3s,t, were synthesized. The required
ketones for the synthesis of 3s,t were synthesized by substituting
the fluorine of 4-fluoroacetophenone by conventional aromatic
nucleophilic displacement with secondary amines 1-benzylpipera-
zine and 1-cinnamylpiperazine.25 However, the biological activity
evaluation revealed that both compounds were less active, this
could be due to increase in bulk at the para-position. Thereafter,
compounds, 3u–w, were synthesized by introducing small tertiary
amine substitutions at the para-position. All of these compounds
6. Cole, S. T.; Brosch, R.; Parkhill, J.; Garnier, T.; Churcher, C.; Harris, D.; Gordon, S.
V.; Eiglmeier, K.; Gas, S.; Barry, C. E., III; Tekaia, F.; Badcock, K.; Basham, D.;
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Gentles, S.; Hamlin, N.; Holroyd, S.; Hornsby, T.; Jagels, K.; Krogh, A.; McLean, J.;
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exhibited potent antitubercular activity at <0.5 lM. Of particular
Letter, compound 3v ((E)-3-(5-nitrofuran-2-yl)-1-(4-(piperidin-1-
yl)phenyl)prop-2-en-1-one) was found to be most potent (MIC:
11. Dharmarajan, S.; Perumal, Y.; Prathiba, D.; Palaniappan, S.; Debjani, B.;
Thimmappa, H. M. Bioorg. Med. Chem. Lett. 2009, 19, 1152.
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21.
0.19
l
M). The compound, 3v, was ꢀ10 times potent than the first
line antitubercular agent INH.
Dramatic decrease in activity was found for compound, 3x, which
was synthesized by replacing the furan moiety of 3w, with thiophene
moietyfromcorresponding5-nitrothiophene-2-carbaldehyde, point-
ing tothe importance ofthe electronegative furan moiety in the vicin-
ity of the nitro group. Thus, compounds with small tertiary amine
feature as structural features (3u, 3v, and 3w) were more active than
other compounds. Fittingofligandinthe receptor siteplays an impor-
tant role for activity. This has been achieved by actual experiments
with systematic modificationof substitutions. This coupled with low-
er LUMO energies is responsible for overall activity picture and there-
fore no direct correlation between the LUMO, HOMO energies and
biological activity was observed.
The compounds were also evaluated for toxicity in a mamma-
lian VERO cell line (C1008) in 96-well microtitre plates and the
CC50 values were determined (Table 1). All compounds showed
good safety profile, particularly, the most potent compound, 3v,
had a selectivity index >1800.
In summary, several 4-(5-nitrofuran-2-yl)prop-2-en-1-one
derivatives were designed and synthesized using stereoelectronic
feature analysis. Screening of the antimycobacterial activity of these
compounds identified ((E)-3-(5-nitrofuran-2-yl)-1-(4-(piperidin-1-
yl)phenyl)prop-2-en-1-one) (3v) as a lead endowed with high anti-
tubercular activity. This study also shows the potential of combining
stereoelectronic property analysis with synthetic approaches and
biological evaluation in developing improved nitroaromatics as
antitubercular agents. Further structure–activity relationship and
mechanistic studies are under progress in our laboratory.
14. Tawari, N. R.; Degani, M. S. J. Comput. Chem. 2010, 31, 739.
15. Valdez, C. A.; Tripp, J. C.; Miyamoto, Y.; Kalisiak, J.; Hruz, P.; Andersen, Y. S.;
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16. Jaguar, version 7.5 Schrödinger, LLC, New York, NY.
17. Typical procedure for the condensation of 5-nitro-2-furaldehyde with ketones,
synthesis of ((E)-3-(5-nitrofuran-2-yl)-1-(4-(piperidin-1-yl)phenyl)prop-2-en-1-
one) (3v): 5-nitro-2-furaldehyde (500 mg, 3.5 mmol), 1-(4-(piperidin-1-
yl)phenyl)ethanone (0.71 g, 3.5 mmol) and concd sulfuric acid (0.2 mL) in
acetic acid (5 mL) were stirred at 60–100 °C until completion of the reaction
(3–24 h). The cooled mixture was diluted with methanol (5–10 mL) and the
precipitated solid was filtered off. Column purification using CombiFlashÒ
RETRIEVEÒ system (SiO2, 3 g, 30% EtOAc/hexanes) of the crude product yielded
pure 0.49 g (42%) of 3v; mp: 225 °C; IR (KBr): Vmax, cmÀ1 3097, 2939, 2853,
1641, 1607, 1577, 1512, 1474, 1391, 1353, 1234, 1124, 1025; 1H NMR
(300 MHz, CDCl3): d, ppm 8.02 (s, 2H), 7.79 (d, 1H, J 18 Hz), 7.51 (d, 1H, J 15 Hz),
7.37 (s, 1H), 6.93 (s, 2H), 6.78 (s, 1H), 3.43 (s, 4H), 1.69 (s, 6H). 13C NMR
(300 MHz, CDCl3): d, ppm 185.72, 153.84, 153.82, 153.80, 131.17, 131.13,
126.54, 126.50, 125.62, 115.73, 113.64, 113.55, 113.42, 48.81, 48.69, 25.22,
25.20, 24.21; ESI MS: 327 (M+1), 266.9 (M+–NO2); purity of the final
compounds was confirmed by analytical liquid chromatographic system
Jasco LC-900 coupled with a Jasco MD 2015 plus intelligent Photo Diode
Array detector. The mobile phase comprised of mixed methanol/THF/water in
the ratio of 40:25:35. Detection was done at kmax of 425 nm. purity—99.58%,
tR—5.46.
18. Palomino, J. C.; Martin, A.; Camacho, M.; Guerra, H.; Swings, J.; Portaels, F.
Antimicrob. Agents Chemother. 2002, 46, 2720.
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R.; Degani, M. S. Bioorg. Med. Chem. Lett. 2010, 20, 1623.
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Sasaki, H.; Shimokawa, Y.; Komatsu, M. PLoS Med. 2006, 3, 2131.
21. Kim, P.; Zhang, L.; Manjunatha, U. H.; Singh, R.; Patel, S.; Jiricek, J.; Keller, T. H.;
Boshoff, H. I.; Barry, C. E., III; Dowd, C. S. J. Med. Chem. 2009, 52, 1317.
22. Phase, version 2.0 Schrödinger, LLC, New York, NY.
23. S. Bag, N.R. Tawari, S.F. Queener, M.S. Degani, J. Enzyme Inhib. Med. Chem.
doi:10.3109/14756360903179443.
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Acknowledgment
This work was supported by Department of Biotechnology,
India; Grant number: BT/PR7858/Med/14/1142/2006.