Synthesis, antimycobacterial activity and docking study of 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives and related hydrazide-hydrazones

Article abstract of DOI:10.1016/j.bmcl.2017.05.011

A new convenient method for preparation of 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives 5b–g and coumarin containing hydrazide-hydrazone analogues 4a–e was presented. The antimycobacterial activity against reference strain Mycobacterium tuberculosis H37Rv and cytotoxicity against the human embryonic kidney cell line HEK-293 were tested in vitro. All compounds demonstrated significant minimum inhibitory concentrations (MIC) ranging 0.28–1.69?μM, which were comparable to those of isoniazid. The cytotoxicity (IC₅₀?>?200?μM) to the “normal cell” model HEK-293T exhibited by 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives 5b–e, was noticeably milder compared to that of their hydrazone analogues 4a–e (IC₅₀ 33–403?μM). Molecular docking studies on compounds 4a–e and 5b–g were also carried out to investigate their binding to the 2-trans-enoyl-ACP reductase (InhA) enzyme involved in M. tuberculosis cell wall biogenesis. The binding model suggested one or more hydrogen bonding and/or arene-H or arene-arene interactions between hydrazones or pyrazole-fused coumarin derivatives and InhA enzyme for all synthesized compounds.

Full text of DOI:10.1016/j.bmcl.2017.05.011

Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis, antimycobacterial activity and docking study of 2-aroyl-[1]
benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives and related
hydrazide-hydrazones
b
c
a
d
Violina T. Angelova a, , Violeta Valcheva , Tania Pencheva , Yulian Voynikov , Nikolay Vassilev ,
⇑
Rositsa Mihaylova a, Georgi Momekov a, Boris Shivachev e
a Faculty of Pharmacy, Medical University of Sofia, 2 Dunav Str., 1000 Sofia, Bulgaria
b ‘‘Stefan Angelov” Institute of Microbiology, Bulgarian Academy of Sciences, 26 Acad. G. Bonchev Str., 1113 Sofia, Bulgaria
c Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, 105 Acad. G. Bonchev Str, 1113 Sofia, Bulgaria
d Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences, 1113 Sofia, Bulgaria
e Institute of Mineralogy and Crystallography, Bulgarian Academy of Sciences, 107 Acad. G. Bonchev Str., 1113 Sofia, Bulgaria
a r t i c l e i n f o
a b s t r a c t
Article history:
A new convenient method for preparation of 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one deriva-
tives 5b–g and coumarin containing hydrazide-hydrazone analogues 4a–e was presented. The antimy-
cobacterial activity against reference strain Mycobacterium tuberculosis H37Rv and cytotoxicity against
the human embryonic kidney cell line HEK-293 were tested in vitro. All compounds demonstrated signif-
Received 28 March 2017
Revised 3 May 2017
Accepted 4 May 2017
Available online 5 May 2017
icant minimum inhibitory concentrations (MIC) ranging 0.28–1.69 lM, which were comparable to those
of isoniazid. The cytotoxicity (IC50 > 200 mM) to the ‘‘normal cell” model HEK-293T exhibited by 2-aroyl-
[1]benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives 5b–e, was noticeably milder compared to that of
their hydrazone analogues 4a–e (IC50 33–403 mM). Molecular docking studies on compounds 4a–e and
5b–g were also carried out to investigate their binding to the 2-trans-enoyl-ACP reductase (InhA) enzyme
involved in M. tuberculosis cell wall biogenesis. The binding model suggested one or more hydrogen
bonding and/or arene-H or arene-arene interactions between hydrazones or pyrazole-fused coumarin
derivatives and InhA enzyme for all synthesized compounds.
Keywords:
Antimycobacterial activity
Chromeno[4,3-c]pyrazol-4-one
Cytotoxicity
Hydrazide-hydrazones
Molecular docking
Ó 2017 Published by Elsevier Ltd.
A reemergence of tuberculosis accompanied by an increasing
number of drug resistant Mycobacterium tuberculosis strains high-
lights the urgent need of searching and developing of new antitu-
bercular drugs, capable of bypassing the resistance mechanisms. In
the last decades, major advances in molecular biology have
increased the knowledge of the mechanisms of resistance to the
main anti-TB drugs, with the identification of specific gene muta-
tions that are associated with drug resistance1,2
Isoniazid (INH), an essential antitubercular agent recommended
by the WHO, is a prodrug that penetrates the tubercle bacilli by
passive diffusion and is bio-activated by the bacterial anti-oxidant
enzyme (KatG).3,4 It exerts its anti-tubercular activity via interfer-
ence with the synthesis of mycolic acids, which comprise crucial
elements of the mycobacterial cell wall. Even with the clinical suc-
cess of isoniazid, severe adverse effects, especially peripheral neu-
ropathy and hepatotoxicity, are associated with INH-based
treatment protocols; moreover its usefulness is further limited by
the occurrence of resistance.5 To overcome the resistance,6 the
drug design strategies frequently employ a combination of the
INH molecule with other pharmacophores, rendering antitubercu-
lar activity. The novel INH hydrazide derivatives appeared to be
promising anti-tubercular agents - more effective and less hepato-
toxic than isoniazid.7–16 In the meantime Ellis at al.,17 have
described the mechanism of action of the pyridoxal isonicotinoyl
hydrazones (PIC) and suggested that hydrazones act as a lipophilic
vehicle for the transport of its intact INH moiety into the mam-
malian cell and the mycobacterium (Fig. 1). The mechanism of
antimycobacterial activity of INH18 and isonicotinoyl hydrazone
derivatives passes through formation of electrophilic intermediate
species (i.e. a hydrazyl radical or ion) (Fig. 1). The acyl radical being
coupled to NADH or NAD + seems to be crucial in yielding adduct
responsible for the inhibition of 2-trans-enoyl-ACP reductase
(InhA), and in restraining the mycobacterial cell wall synthesis.
InhA catalyze the final step in the elongation cycle of the bacterial
⇑
Corresponding author.
E-mail
addresses:
violina@pharmfac.net,
violina_stoyanova@abv.bg
(V.T. Angelova).
http://dx.doi.org/10.1016/j.bmcl.2017.05.011
0960-894X/Ó 2017 Published by Elsevier Ltd.

V.T. Angelova et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
2997
Fig. 1. Schematic representation of the proposed mechanism of action of isoniazid (INH)18 and hydrazone derivative (2-pyridylcarboxaldehydeisonicotinoylhydrazone –
PCIH).17 X-ray structure of the INH-NAD adduct (cyan) bound to InhA (1zid.pdb).27 The substrate binding loop is colored red. This portion of the protein (res 196–211) is
disordered in the structure of triclosan bound to InhA (2b35.pdb).24
.
fatty acid biosynthesis pathway (type-II fatty acid elongation sys-
tem- FAS-II) has been recognized as a primary molecular target
for developing new anti-TB drugs19,20 (Fig. 1). There is also evi-
dence that INH can inhibit non-NAD(P) binding proteins such as
KasA, the b-ketoacyl-ACP synthase in the FASII pathway.21 Impor-
tantly, since mutations of KatG are the most common mechanisms
of resistance to INH,22,23 inhibitors that bind to the final drug target
(s) should be active against the majority of INH-resistant clinicaly
isolated strains.24 Molecular investigation on the genetic basis of
the INH-resistance in M. tuberculosis strains in Bulgaria targeted
two the most frequently reported mutations related to INH resis-
tance, katG 315AGC > ACC and inhA ꢀ15C > T25,26 The global preva-
lence of the katG S315T substitution in INH-resistant strains
highlights the selective advantage conferred by this mutation,
which appears to provide the optimal balance between decreased
catalase activity and a sufficiently high level of peroxidase activity
in KatG. Mutations in the inhA promoter region are thought to
increase the InhA protein expression, thereby elevating the drug
target levels and producing INH resistance by a drug titration
mechanism.
Recently we described a series of hydrazide-hydrazones con-
taining coumarin or 2H-chromene moieties with potent anti-TB
activity and moderate cytotoxicity.32 Having surveyed comprehen-
sively the literature on pyrazole33–36 and coumarin34,37–46 scaffolds
as important structural components in the design of anti-tubercu-
losis agents against Mycobacterium tuberculosis we focused on the
synthesis of pyrazole-fused coumarin derivatives as less toxic
antimycobacterial compounds.
Herein, continuing the research on the biological activity of 2H-
chromene derivatives, we present the one-pot synthesis, antimy-
cobacterial activity and cytotoxicity of novel 2-aroyl-[1]benzopy-
rano[4,3-c]pyrazol-4(1H)-one
derivatives
5b–g.
Molecular
docking studies were carried out with the InhA enzyme from M.
tuberculosis in order to explore the structural requirements con-
trolling the observed antimycobacterial activity of 4a–e and 5b–g.
The synthesis of hydrazide-hydrazones 4a–e and N-2-substi-
tuted coumarin[4,3-c]pyrazoles 5b–g is shown in Scheme 1.
The synthesis of coumarin based hydrazones 4a–e was achieved
by a classical method reported previously.33 It involves a reaction
of 4-chlorocoumarin-3-carbaldehyde 2 with corresponding hydra-
zides 3a–e in abs. EtOH. The detailed investigation of the reaction
showed that fused 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-
one was formed easily when an electron donating group was pre-
sent at p-position in the benzene ring. The reaction of 4-chloro-
coumarin-3-carbaldehyde 2 with 3b–g in EtOH:CH2Cl2 (at a 1:3
ratio) resulted in N-2-substituted chromeno[4,3-c]pyrazol-4-one
analogues 5b–g (Scheme 1) in nearly quantitative yields
(85–92%). Compounds 5b–g were further purified by column
This highlights the crucial need of developing drugs with
shorter, simpler regimens as well as with novel mechanisms of
action that can be used for treatment of drug resistant forms of
the disease. Numerous efforts have been undertaken to develop
new INH derivatives and hydrazones28,29 as anti-TB agents (Salu-
zide, Ftivazide, Salizide).30,31 Indeed, the INH derivative LL-3858
is in initial stages of phase II clinical trial for the treatment of TB
and may be approved to treat TB in the near future.

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V.T. Angelova et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
Scheme 1. Synthetic route of coumarin containing hydrazones 4a–e and 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives 5b–g. Reaction conditions: a) POCl3,
DMF, 60 °C;34 b) abs. EtOH, r.t., 15 min; c) CH2Cl2/EtOH (1:3), r.t., 15 min–1 h.
chromatography (ethyl acetate/hexane at a 1:10 ratio) before being
tested for biological activity. The structure of the newly synthe-
sized coumarin derivatives was confirmed by a combination of
1H NMR, 13C NMR, HR-MS and FTIR (ATR) spectral techniques
(see Supplementary data, Fig. S1). In addition an X-ray crystallo-
graphic study of compound 5d was reported (Fig. 2), (Supplemen-
tary data, Fig. S2, Table S2, Table S3).
The tested series exhibited prominent antimycobacterial activ-
ity against the chosen strain, whereby the MIC values were compa-
rable to those of INH and lower than those of EMB. The cytotoxicity
of the selected compounds was evaluated against the human
embryonal kidney cell line 293 T, after 72 h of continuous expo-
sure, using the MTT-dye reduction assay.48The tested agents
caused a 50% cell growth inhibition within the experimental con-
The newly synthesized compounds 4a–d and 5c–e displayed
chemical stability under the conditions of antimycobacterial activ-
ity determination in DMSO-d6. No changes were detected in their
1H NMR spectra taken in dimethyl sulfoxide-d6 after 1, 2, 3 and
12 days at 20 °C of the NMR tubes with the initially prepared solu-
tions (Supplementary data Fig. S2 and S3). No changes also were
detected in their 1H NMR spectra in 0.6 ml DMSO-d6 and 0.05 ml
assay media Middlebrook 7H9 (pH 6.6) for 16 h and the compound
remained unchanged upon heating to 37 °C the NMR tubes with
the initially prepared solutions (Supplementary data Fig. S4 and
S5).
The in vitro antimycobacterial activity of all compounds against
reference strain Mycobacterium tuberculosis H37Rv was evaluated
using the resazurin microtiter assay (REMA) described by Martin
et al.47 Ethambutol (EMB) and isoniazid (INH) were used as con-
trols. The data for all compounds are shown in Table 1.
centration range (12.5–100 lM). In addition, the selectivity index
values (SI) were determined as the IC50/MIC ratio. All the com-
pounds were found highly potent against the M. tuberculosis
H37Rv demonstrating a nanomolar-to-low-micromolar activity
(MICs ranging 0.32 to 1.69 mM). The most active compounds 4c
(MIC = 0.28 mM) and 5f (MIC = 0.32 mM) with p-methoxyphenyl
and p-hydroxyphenyl substituents, respectively, demonstrated a
remarkable minimal associated cytotoxicity (IC50 = 112.9 mM and
IC50 > 200.00 mM, respectively). The activity of compounds 4a–e
was comparable to that of the hydrazone derivatives presented
in our previous study.22 However, the cytotoxicity of 2-aroyl-[1]
benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives 5b–g, to the
‘‘normal cell” model HEK-293 T (IC50 > 200 mM) was noticeably
milder compared to the one of their open chain hydrazone ana-
logues 4a–e (33–403 mM). The difference in the cytotoxic potential
resulted in much better SI values exhibited by derivatives 5b–g.
Hence,
the
2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one
derivatives 5b–g possess potent antimycobacterial activities com-
bined with low cytotoxicity resulting in SI values higher than those
of their open chain analogues 4a–e (Table 1). It was observed that
the presence of -OH, OMe and -N(Me)2 functional groups at 4th
position in the phenyl ring enhanced the antimycobacterial activity
while any alteration in this substitution pattern caused a substan-
tial decrease of the anti-tubercular potency. The addition of a sec-
ond hydroxyl group in compound 5e gave a derivative slightly less
potent (MIC 0.62 lM) than monohydroxy derivative 5f. All tested
hydrazones, except 4b, were more lipophilic than INH (Table 1)
with log P values higher than those of the controls. It is well known
that increasing the lipophilicity is important for the transport
through the mycobacterial cell.49 We observed no correlation
between the antimycobacterial potency and log P values within
the tested series.
Fig. 2. ORTEP view of the molecular structure of 5d; atomic displacement
parameters are at 50% probability, H atoms are shown as spheres with arbitrary
radii.
To explain the activity order of 4a–e and 5b–g against M. tuber-
culosis the compounds were docked into the binding site of

V.T. Angelova et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
2999
Table 1
Antimycobacterial activity and in vitro cytotoxicity.
Compd.
Structure
MICa(mM)
IC50b(mM)
SIc
33
Log Pd
4a
0.55
21.00
2.18
4b
4c
0.63
0.28
63.62
112.9
236
403
0.61
1.41
4d
0.54
18.70
20
2.75
4e
5b
0.56
0.72
–
–
2.69
1.29
>200.00
>282
5c
0.62
0.59
>200.00
>200.00
>645
>408
2.09
2.61
5d
5e
5f
0.62
0.32
>200.00
>200.00
>308
>625
1.78
2.06
5g
1.69
>200.00
>119
1.54
EMB.2HCle
INHe
1.46
0.79
–
–
–
–
0.77
ꢀ0.63
a
b
c
Antimycobacterial activity against reference strain of Mycobacterium tuberculosis H37Rv.
In vitro cytotoxicity to human embryonal kidney cell line 293 T, IC50 (lM).
Selectivity index, SI ratio = IC50/MIC.
d
e
LogP was calculated by Marvin 16.2.8.0 (http://www.chemaxon.com).
EMB.2HCl (ethambutol dihydrochloride-reference compound) and eINH (Isoniazid - reference compound).
mycobacterial enoyl reductase (InhA). Two docking solutions can
be considered as ‘‘standard” in a typical docking analysis, namely:
1) the top score – the pose with the highest rank; 2) the best pose –
the pose with lowest root mean square deviation (RMSD) to the
reference ligand from the experimentally solved structure. (). The
docking scores (S_value) of ligands 4a–e and 5b–g were found sat-
isfactory in the range of -7.72 to -6.74 (See Supplementary data –
Table S1 lists the essence of docking results for the synthesized
compounds). The results presented in Table S1 showed that the
lowest RMSD was obtained for 2-aroyl-[1]benzopyrano[4,3-c]pyra-
zol-4(1H)-one 5g, followed by one of the most active compounds
5f, while the highest rank was recorded for p-(dimethylamino)phe-

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V.T. Angelova et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
nyl hydrazone derivative 4d. The docking scores of the most active
compounds of the series, 4c and 5f, were found to be ꢀ7.27 and
ꢀ6.89, respectively.
mentioned as important residues were still very close. Compound
5f (Fig. 4a, right) repeated the interaction with Phe149 and demon-
strated one newly appeared interaction with Leu218 (arene-H
interaction with coumarin ring). As in the case of compound 4c,
all the rest mentioned as important residues were still very close.
It should be noted that both most active TB compounds 4c and
5f demonstrated one of the interactions of compound 4d, ranked
by the docking as the top score – the pose with the highest rank
(See Suppl. Table S1).
All synthesized compounds 4a–e and 5b–g as docked in the
ligand-binding domain of M. tuberculosis InhA inhibited by PT70
with Connolly surface are presented on Fig. 5. As seen from the fig-
ure, all synthesized compounds occupied the same binding site as
that of PT70 and NAD+, formed a cluster, fitted well in the ligand-
binding domain of M. tuberculosis InhA and always showed one or
more specific protein-ligand interactions as listed in Supplemen-
tary data Table S1. This result indicated that the coumarin struc-
ture or pyrazole-fused coumarin scaffold was the key fragment
for binding of the compound in the InhA pocket and is crucial to
attaining favorable MIC values, whereas the substituents in the
benzene ring were a key group to enhance MIC values.
The docking study analysis of the tested compounds with
respect to the reward for hydrogen bonding revealed compounds
5b–g to have similar docking scores but protein-ligand interactions
different from those of compounds 4a–e. The results of enzyme
inhibition activity of the substituents at p-position in hydrazone
derivatives showed the following trend: (p-N(Me)2 > (p-MeO phe-
nyl); p-chloro phenyl, o,p-diOH phenyl) > (2-furyl moiety). In the
case of substitution at aroyl moiety in the coumarin fused pyra-
zoles, the substitution with electrondonating groups at the ben-
zene or furyl ring was favored. In addition, any discrepancy
The protein-ligand interactions (PLI) diagram of the co-crystal-
ized ligand of M. tuberculosis InhA inhibited by PT70 in the ligand-
binding domain is presented in Fig. 2. It was obtained using
‘‘Ligand Interactions” tool of Molecular Operating Environment of
Chemical Computing Group (MOE, version 2016.08, https://www.
chemcomp.com/MOE-Molecular_Operating_Environment.htm).
Docking simulations were performed using the only published
X-ray crystallographic structure of InhA from species of
Mycobacterium tuberculosis (ENR) complexed with 5-hexyl-2-(2-
methylphenoxy)phenol (PT70) and co-factor nicotinamide adenine
dinucleotide (NAD+). The crystal structure of M. tuberculosis InhA
inhibited by PT70 was extracted from Protein Data Bank (http://
www.rcsb.org/, PDB ID 2X22). During the docking process, water
molecules and all the ligands in the crystal structures were
removed, except the co-factor NAD+. Protein-ligand interactions
illustrated in Figures 3 and 4 were in the maximum distance of
4.5 Å; between heavy atoms of the ligand and receptor. As seen
from Fig. 3, three residues were involved in interactions, namely
Phe149, Tyr158 and Met161. Residues Ala198, Met199, Ile202
and Val203 should be explicitly mentioned as very important,
since they are involved in the substrate-binding loop.50 Fig. 3
demonstrated all of them at ligand exposure. Fig. 4a presents the
PLI diagrams of the most active compounds 4c and 5f and Fig. 4b
– the docking conformations and corresponding Connolly surface.
As seen from Figure 4a (left), compound 4c demonstrated two
newly appeared interactions with Met199 (HB) and Leu218
(arene-H interaction with coumarin ring), that was at ligand expo-
sure, but not exhibiting any strong interaction in Fig. 3. All the rest
Fig. 3. Interaction diagram of the ligand-binding domain of M. tuberculosis InhA inhibited by PT70 with TCU (5-hexyl-2-(2-methylphenoxy)phenol) (PDB ID 2X22).

V.T. Angelova et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
3001
Fig. 4. Docking results for the most active TB compounds 4c and 5f.
of the antiproliferative effects was thus assessed. All compounds
displayed good SI values ranging from 33 to more than 645. In gen-
eral, 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-one derivatives
5b–g possessed potent antimycobacterial activity combined with
low cytotoxicity what resulted into SI values higher than that of
their open chain analogues 4a–e. The activity of the tested com-
pounds may be attributed to the significant interactions with the
inhibitor binding cavity of M. tuberculosis Enoyl-ACP reductase
and/or related to ability of the tested compounds to penetrate
mycobacterial cells. The above observations show that the novel
hydrazide–hydrazones 4a–e and 2-aroyl-[1]benzopyrano[4,3-c]
pyrazol-4(1H)-one derivatives 5b–g have potential as antimy-
cobacterial agents. The results of the study can be utilized to fur-
ther optimize and improve the potency and selectivity toward
ENR enzyme by varying the basic skeleton and the substituents.
Fig. 5. Docking conformation of all synthesized compounds (in green), PT70 (in
magenta), NAD + (in cyan) and corresponding Connolly surface.
Acknowledgements
The authors are indebted to the Medical Science Council, MU-
Sofia for the partial support of this work under contract No.
7/2016. The authors are indebted to Dr. P. T. Nedialkov for taking
mass spectra and to Dr. Javor Mitkov for IR spectra.
between the docking data and the actual activity of the tested com-
pounds could be ascribed to pharmacokinetic and biopharmaceuti-
cal properties, that make a given compound better accessible by
the receptor site and condition superior activity even when the
binding interaction is a less prominent one. To the best of our
knowledge, this is the first literature report on the antimycobacte-
rial activity of pyrazole-fused chromene derivatives.
In summary, a series of coumarin-linked hydrazide–hydrazones
4a–e and 2-aroyl-[1]benzopyrano[4,3-c]pyrazol-4(1H)-ones 5b–g
were synthesized via an efficient one-pot protocol. All compounds
demonstrated significant minimum inhibitory concentrations
A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2017.05.
011.
References
(MIC) ranging from 0.28 to 1.69 lM against reference M. tuberculo-
sis H37Rv strain. The cytotoxicity against the human embryonic
kidney cell line HEK-293 was also evaluated and the selectivity
1. McBryde ES, Meehan MT, Doan TN, et al. The risk of global epidemic
replacement with drug-resistant Mycobacterium tuberculosis strains. Int
Infect Dis. 2017;56:14–20.
J

3002
V.T. Angelova et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 2996–3002
2. Ramaswamy SV, Amin AG, Göksel S, et al. Molecular genetic analysis of
nucleotide polymorphisms associated with ethambutol resistance in human
isolates of Mycobacterium tuberculosis. Antimicrob Agents Chemother.
2000;44:326–336.
3. Argyrou A, Vetting MW, Aladegbami B, Blanchard JS. Mycobacterium
tuberculosis dihydrofolate reductase is a target for isoniazid. Nat Struct Mol
Biol. 2006;13:408–413.
4. Zhang Y, Heym B, Allen B, Young D, Cole S. The catalase-peroxidase gene and
isoniazid resistance of Mycobacterium tuberculosis. Nature. 1992;358:591.
5. Joshi SD, Kumar D, Dixit SR, et al. Synthesis, characterization and antitubercular
activities of novel pyrrolyl hydrazones and their Cu-complexes. Eur J Med Chem.
2016;121:21–39.
27. Rozwarski DA, Grant GA, Barton DH, Jacobs WR, Sacchettini JC. Modification of
the NADH of the isoniazid target (InhA) from Mycobacterium tuberculosis.
Science. 1998;279:98–102.
28. Maccari R, Ottana R, Vigorita MG. In vitro advanced antimycobacterial
screening of isoniazid-related hydrazones, hydrazides and cyanoboranes: Part
14. Bioorg Med Chem Lett. 2005;15:2509–2513.
29. Ferreira de LM, Candea AL, de OH, et al. Synthesis and cytotoxic evaluation of
disubstituted n-acylhydrazones pyrazinecarbohydrazide derivatives. Lett Drug
Des Discovery. 2010;7(4):275–280.
30. Elhakeem MA, Taher AT, Abuel-Maaty SM. Synthesis and anti-mycobacterial
evaluation of some new isonicotinylhydrazide analogues. Bull Facul Pharm,
Cairo Univ. 2015;53:45–52.
6. Argyrou A, Vetting MW, Blanchard JS. New insight into the mechanism of action
of and resistance to isoniazid: interaction of Mycobacterium tuberculosis
enoyl-ACP reductase with INH-NADP. J Am Chem Soc. 2007;129:9582–9583.
7. Velezheva V, Brennan P, Ivanov P, et al. Synthesis and antituberculosis activity
of indole–pyridine derived hydrazides, hydrazide–hydrazones, and
thiosemicarbazones. Bioorg Med Chem Lett. 2016;26:978–985.
8. Pavan FR, Maia PIdS, Leite SR, et al. Thiosemicarbazones, semicarbazones,
dithiocarbazates and hydrazide/hydrazones: anti-Mycobacterium tuberculosis
activity and cytotoxicity. Eur J Med Chem. 2010;45:1898–1905.
31. Hu Y-Q, Zhang S, Zhao F, et al. Isoniazid derivatives and their anti-tubercular
activity. Eur J Med Chem. 2017.
32. Angelova VT, Valcheva V, Vassilev NG, et al. Antimycobacterial activity of novel
hydrazide-hydrazone derivatives with 2H-chromene and coumarin scaffold.
Bioorg Med Chem Lett. 2017;27:223–227.
33. Velaparthi S, Brunsteiner M, Uddin R, Wan B, Franzblau SG, Petukhov PA. 5-
tert-Butyl-N-pyrazol-4-yl-4, 5, 6, 7-tetrahydrobenzo [d] isoxazole-3-
carboxamide derivatives as novel potent inhibitors of Mycobacterium
tuberculosis pantothenate synthetase: initiating
a
quest for new
9. Kumar P, Narasimhan B, Yogeeswari P, Sriram D. Synthesis and antitubercular
antitubercular drugs. J Med Chem. 2008;51:1999–2002.
activities of substituted benzoic acid N0-(substituted benzylidene/furan-2-
34. Khan MF, Alam MM, Verma G, Akhtar W, Akhter M, Shaquiquzzaman M. The
therapeutic voyage of pyrazole and its analogs: a review. Eur J Med Chem.
2016;120:170–201.
ylmethylene)-N-(pyridine-3-carbonyl)-hydrazides.
2010;45:6085–6089.
Eur
J
Med
Chem.
10. Sriram D, Yogeeswari P, Madhu K. Synthesis and in vitro antitubercular activity
of some 1-[(4-sub) phenyl]-3-(4-{1-[(pyridine-4-carbonyl) hydrazono] ethyl}
phenyl) thiourea. Bioorg Med Chem Lett. 2006;16:876–878.
11. Martins F, Santos S, Ventura C, et al. Design, synthesis and biological evaluation
of novel isoniazid derivatives with potent antitubercular activity. Eur J Med
Chem. 2014;81:119–138.
12. Kumar HN, Parumasivam T, Jumaat F, Ibrahim P, Asmawi MZ, Sadikun A.
Synthesis and evaluation of isonicotinoyl hydrazone derivatives as
antimycobacterial and anticancer agents. Med Chem Res. 2014;23:269–279.
13. Mathew B, Suresh JJ, Ahsan M, et al. Hydrazones as a privileged structural linker
in antitubercular agents: a review. Infect Disord-Drug Targets (Formerly Curr
Drug Targets-Infect Disord). 2015;15(2):76–88.
35. Keri RS, Chand K, Ramakrishnappa T, Nagaraja BM. Recent progress on pyrazole
scaffold-based antimycobacterial agents. Arch Pharm. 2015;348:299–314.
36. Aragade P, Palkar M, Ronad P, Satyanarayana D. Coumarinyl pyrazole
derivatives of INH: promising antimycobacterial agents. Med Chem Res.
2013;22:2279–2283.
37. Keri RS, Sasidhar B, Nagaraja BM, Santos MA. Recent progress in the drug
development of coumarin derivatives as potent antituberculosis agents. Eur J
Med Chem. 2015;100:257–269.
38. Manvar A, Bavishi A, Radadiya A, et al. Diversity oriented design of various
hydrazides and their in vitro evaluation against Mycobacterium tuberculosis
H37 Rv strains. Bioorg Med Chem Lett. 2011;21:4728–4731.
39. Arshad A, Osman H, Bagley MC, Lam CK, Mohamad S, Zahariluddin ASM.
Synthesis and antimicrobial properties of some new thiazolyl coumarin
derivatives. Eur J Med Chem. 2011;46:3788–3794.
14. Negi VJ, Sharma A, Negi J, Ram V. Biological activities of hydrazone derivatives
in the new millennium. Int J Pharm Chem. 2012;2:100–109.
15. Eldehna WM, Fares M, Abdel-Aziz MM, Abdel-Aziz HA. Design, synthesis and
antitubercular activity of certain nicotinic acid hydrazides. Molecules.
2015;20:8800–8815.
40. Ventura LBT, Silva RCD, Lassounskaia E, Maria JE, Muzitano FM, de Oliveira RR.
Coumarine analogues with antimycobacterial and immunomodulatory activity.
Curr Bioact Compd. 2015;11:109–115.
16. Bijev A, Georgieva M. The development of new tuberculostatics addressing the
return of tuberculosis: current status and trends. J Univ Chem Technol Metall.
2010;45:111–126.
41. Karalı N, Kocabalkanlı A, Gürsoy A, Atesß Ö. Synthesis and antitubercular activity
of 4-(3-coumarinyl)-3-cyclohexyl-4-thiazolin-2-one benzylidenehydrazones. Il
Farmaco. 2002;57:589–593.
17. Ellis S, Kalinowski DS, Leotta L, et al. Potent antimycobacterial activity of the
42. Lad HB, Giri RR, Brahmbhatt D. An efficient synthesis of some new 3-
bipyridinyl substituted coumarins as potent antimicrobial agents. Chin Chem
Lett. 2013;24:227–229.
43. Kenchappa R, Bodke YD, Chandrashekar A, Telkar S, Manjunatha K, Sindhe MA.
Synthesis of some 2, 6-bis (1-coumarin-2-yl)-4-(4-substituted phenyl) pyridine
derivatives as potent biological agents. Arab J Chem. 2013.
pyridoxal
isonicotinoyl
hydrazone
analog
2-pyridylcarboxaldehyde
isonicotinoyl hydrazone: a lipophilic transport vehicle for isonicotinic acid
hydrazide. Mol Pharmacol. 2014;85:269–278.
18. Timmins GS, Deretic V. Mechanisms of action of isoniazid. Mol Microbiol.
2006;62:1220–1227.
19. Banerjee A, Dubnau E, Quemard A, et al. InhA, a gene encoding a target for
isoniazid and ethionamide in Mycobacterium tuberculosis. Science-AAAS-
44. Bailly F, Queffelec C, Mbemba G, Mouscadet J-F, Cotelle P. Synthesis and HIV-1
integrase inhibitory activities of caffeic acid dimers derived from Salvia
officinalis. Bioorg Med Chem Lett. 2005;15:5053–5056.
Weekly
Paper
Edition-including
Guide
to
Scientific
Information..
1994;263:227–229.
45. Trivedi JC, Bariwal JB, Upadhyay KD, et al. Improved and rapid synthesis of new
coumarinyl chalcone derivatives and their antiviral activity. Tetrahedron Lett.
2007;48:8472–8474.
46. Basanagouda M, Jambagi VB, Barigidad NN, Laxmeshwar SS, Devaru V.
Synthesis, structure–activity relationship of iodinated-4-aryloxymethyl-
coumarins as potential anti-cancer and anti-mycobacterial agents. Eur J Med
Chem. 2014;74:225–233.
47. Martin A, Camacho M, Portaels F, Palomino JC. Resazurin microtiter assay plate
testing of Mycobacterium tuberculosis susceptibilities to second-line drugs:
rapid, simple, and inexpensive method. Antimicrob Agents Chemother.
2003;47:3616–3619.
20. Joshi SD, Dixit SR, More UA, Aminabhavi TM, Kulkarni VH, Gadad AK. Enoyl ACP
reductase as effective target for the synthesized novel antitubercular drugs: a-
State-of-the-Art. Mini Rev Med Chem. 2014;14(8):678–693.
21. Mdluli K, Slayden RA, Zhu Y, et al. Inhibition of a Mycobacterium tuberculosis
b-ketoacyl ACP synthase by isoniazid. Science. 1998;280:1607–1610.
22. Musser JM, Kapur V, Williams DL, Kreiswirth BN, Van Soolingen D, Van Embden
JD. Characterization of the catalase-peroxidase gene (katG) and inhA locus in
isoniazid-resistant and-susceptible strains of Mycobacterium tuberculosis by
automated DNA sequencing: restricted array of mutations associated with drug
resistance. J Infect Dis. 1996;173:196–202.
23. Ramaswamy SV, Reich R, Dou S-J, et al. Single nucleotide polymorphisms in
genes associated with isoniazid resistance in Mycobacterium tuberculosis.
Antimicrob Agents Chemother. 2003;47:1241–1250.
48. Canetti G, Fox W, Aa Khomenko, et al. Advances in techniques of testing
mycobacterial drug sensitivity, and the use of sensitivity tests in tuberculosis
control programmes. Bull World Health Organ. 1969;41:21.
24. Sullivan TJ, Truglio JJ, Boyne ME, et al. High affinity InhA inhibitors with activity
against drug-resistant strains of Mycobacterium tuberculosis. ACS Chem Biol.
2006;1:43–53.
49. Vinsova J, Imramovsky A, Jampilek J, Monreal JF, Dolezal M. Recent advances on
isoniazide derivatives. Anti-Infect Agents Med Chem (Formerly Curr Med Chem-
Anti-Infect Agents). 2008;7(1):12–31.
25. Valcheva V, Mokrousov I, Narvskaya O, Rastogi N, Markova N. Molecular
snapshot of drug-resistant and drug-susceptible Mycobacterium tuberculosis
strains circulating in Bulgaria. Infect, Genet Evol. 2008;8:657–663.
26. Valcheva V, Markova N, Rastogi N, Narvskaya O, Mokrousov I. Drug Resistant
Tuberculosis in Bulgaria: Molecular Insights. Hauppauge, New York, USA: Nova
Science Publishers Inc; 2009.
50. Luckner SR, Liu N, Am Ende CW, Tonge PJ, Kisker C. A slow, tight binding
inhibitor of InhA, the enoyl-acyl carrier protein reductase from Mycobacterium
tuberculosis. J Biol Chem. 2010;285:14330–14337.