Design, synthesis, and structure-activity relationships of pyrazole derivatives as potential FabH inhibitors

Article abstract of DOI:10.1016/j.bmcl.2010.05.105

Fatty acid biosynthesis is essential for bacterial survival. FabH, β-ketoacyl-acyl carrier protein (ACP) synthase III, is a particularly attractive target, since it is central to the initiation of fatty acid biosynthesis and is highly conserved among Gram-positive and -negative bacteria. Fifty-six 1-acetyl-3,5-diphenyl-4,5-dihydro-(1H)-pyrazole derivatives were synthesized and developed as potent inhibitors of FabH. This inhibitor class demonstrates strong antibacterial activity. Escherichia coli FabH inhibitory assay and docking simulation indicated that the compounds 1-(5-(4-fluorophenyl)- 3-(4-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)ethanone (12) and 1-(5-(4-chlorophenyl)-3-(4-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)ethanone (13) were potent inhibitors of E. coli FabH.

Full text of DOI:10.1016/j.bmcl.2010.05.105

Bioorganic & Medicinal Chemistry Letters 20 (2010) 4657–4660
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis, and structure–activity relationships of pyrazole derivatives
as potential FabH inhibitors
*
Peng-Cheng Lv, Juan Sun, Yin Luo, Ying Yang, Hai-Liang Zhu
State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, Nanjing 210093, People’s Republic of China
a r t i c l e i n f o
a b s t r a c t
Article history:
Fatty acid biosynthesis is essential for bacterial survival. FabH, b-ketoacyl-acyl carrier protein (ACP) syn-
thase III, is a particularly attractive target, since it is central to the initiation of fatty acid biosynthesis and
is highly conserved among Gram-positive and -negative bacteria. Fifty-six 1-acetyl-3,5-diphenyl-
4,5-dihydro-(1H)-pyrazole derivatives were synthesized and developed as potent inhibitors of FabH. This
inhibitor class demonstrates strong antibacterial activity. Escherichia coli FabH inhibitory assay and dock-
ing simulation indicated that the compounds 1-(5-(4-fluorophenyl)-3-(4-methoxyphenyl)-4,5-dihydro-1
H-pyrazol-1-yl)ethanone (12) and 1-(5-(4-chlorophenyl)-3-(4-methoxyphenyl)-4,5-dihydro-1H-pyrazol-
1-yl)ethanone (13) were potent inhibitors of E. coli FabH.
Received 1 March 2010
Revised 24 May 2010
Accepted 29 May 2010
Available online 8 June 2010
Keywords:
Design
Synthesis
FabH inhibitors
Antibacterial
Ó 2010 Elsevier Ltd. All rights reserved.
Pyrazole derivatives
Structure–activity relationship
The emergence of bacterial resistance to most of all antibiotics
poses a threat to health care, and novel therapeutics are needed.
Recently, the research has been focused toward development of
new antibacterial agents with novel target so as to prevent this
serious medical problem. A promising target is the fatty acid syn-
thase (FAS) pathway in bacteria. Fatty acid biosynthesis (FAB) is an
essential metabolic process for prokaryotic organisms and is re-
quired for cell viability and growth.1 b-Ketoacyl-acyl carrier protein
(ACP) synthase III, also known as FabH or KAS III, plays an essential
and regulatory role in bacterial FAB.2,3 The enzyme initiates the fatty
acid elongation cycles,4,5 and is involved in the feedback regulation
of the biosynthetic pathway via product inhibition.6 Therefore, it
represents a promising target for the design of novel antimicrobial
drugs. Because of this, various kinds of compounds were screened
by enzymatic assays to generate leads that were co-crystallized with
various pathogenic FabH proteins and subsequently optimized
using structure guided drug design methods.7–11
Many pyrazole derivatives are acknowledged to possess a wide
range of bioactivities. The pyrazole motif makes up the core struc-
ture of numerous biologically active compounds. Thus, some repre-
sentatives of this heterocycle exhibit anti-viral/anti-tumor,12–14
antibacterial,15–18 antiinflamatory,19 analgesic,20 fungistatic,21
and anti-hyperglycemic activity.22,23 Much attention was paid to
pyrazole as a potential antimicrobial agent after the discovery of
the natural pyrazole C-glycoside, pyrazofurin which demonstrated
a broad spectrum of antimicrobial activity.24
However, to our knowledge, few reports have been dedicated to
the synthesis and FabH inhibitory activity of 1-acetyl-3,5-diphe-
nyl-4,5-dihydro-(1H)-pyrazole derivatives. Herein, in continuation
to extend our research on antibacterial compounds with FabH
inhibitory activity,25,26 we report in the present work the synthesis
and structure–activity relationships of a series of pyrazole deriva-
tives. This combination was suggested in an attempt to investigate
the inhibitory activity against Escherichia coli FabH. Biological eval-
uation indicated that compounds 12 and 13 displayed not only sig-
nificant antibacterial activity against E. coli ATCC 35218, but also
favorable activity against other five bacterial strains, indicating
that they possessing broad-spectrum antibacterial activity. Be-
sides, further E. coli FabH inhibitory assay was undertaken and
the results suggested that compounds 12 and 13 were potent
E. coli FabH inhibitors. In addition, docking simulation were per-
formed to position compound 12 into the E. coli FabH active site
to determine the probable binding conformation and the results
confirmed that compound 12 was potential inhibitor of E. coli
FabH.
Fifty-six 1-acetyl-3,5-diphenyl-4,5-dihydro-(1H)-pyrazole deriv-
atives were synthesized to evaluate their antibacterial activity
and inhibitory activity against E. coli FabH. The synthesis of
compounds 4–59 followed the general pathway outlined in
Scheme 1. They are prepared in two steps. Firstly, the chalcones
were obtained by direct condensation between the aromatic alde-
hydes and the substituted acetophenone, using 20% potassium
* Corresponding author. Tel./fax: +86 25 8359 2672.
E-mail address: zhuhl@nju.edu.cn (H.-L. Zhu).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.05.105

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P.-C. Lv et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4657–4660
i
R1
R2
R2
R1
O
CH3
O
O
H
3ah-3go
1a-1g
2h-2o
R1
A
B
ii
R2
N
N
O
4-59
H3C
Scheme 1. Reagents and conditions. (i) NaOH, CH3CH2OH, 25 °C, 4–10 h; (ii) N2H4ÁH2O, CH3COOH, 120 °C, reflux, 12 h.
hydroxide as catalyst in ethanol. Secondly, a solution of chalcones
(5 mmol) in 30 mL of acetic acid was added dropwise to 0.6 mL of
hydrazine hydrate (12.5 mmol) and kept under stirring at 120 °C
for 24 h. The mixture was then poured into ice-water, obtaining
the 1-acetyl-3,5-diphenyl-4,5-dihydro-(1H)-pyrazole derivatives
pyrazole derivatives 4–59. All of the synthetic compounds gave
satisfactory analytical and spectroscopic data, which were in full
accordance with their depicted structures.
All the synthesized compounds were screened for antibacterial
activity against three Gram positive bacterial strains and three
Gram negative bacterial strains by MTT method. The MICs (mini-
mum inhibitory concentrations) of the compounds against six
strains are presented in Table 1. The antibacterial activity of refer-
ence compounds kanamycin B and penicillin G were also included.
Studies were performed by modification of the A ring and B ring of
the parent compounds to determine how the substituents of the
subunits affected the antibacterial activities. As shown in Table 1,
compounds 12 and 13 displayed potent activity with MIC values
Bacillus subtilis ATCC 6633. For example, Compounds 37 showed
potent activity with MIC values of 0.78 g/mL, which were supe-
rior to the positive control penicillin G. Compounds 36 and 38 sug-
gested favorable activity with MIC values of 1.562 g/mL, which
was comparable to the positive control penicillin G. Compounds
39–43 showed moderate activity with MIC values ranging from
l
l
3.125
Compounds 4, 12, 20, and 36 displayed potent activity with MIC
values of 1.562 g/mL against Streptococcus faecalis ATCC 9790,
lg/mL to 12.5 lg/mL against B. subtilis ATCC 6633. Besides,
l
which were comparable to the positive control penicillin G.
Then, the E. coli FabH inhibitory potency of the selected com-
pounds 12, 13, 14, 15, 16, 17, and 18 was examined and the results
are summarized in Table 2. As shown in Table 2, among the tested
compounds, compounds 12 and 13 showed potent inhibitory activ-
ity with IC50 of 4.2
pounds displayed moderate inhibitory activity with IC50 ranging
from 10.4 M to 46.7 M.
lM and 7.6 lM, respectively. Other tested com-
l
l
of 0.39
were superior to the positive control kanamycin B. Compound 14
exhibited significant activity with MIC values of 1.562 g/mL
lg/mL and 0.78 lg/mL against E. coli ATCC 35218, which
l
against E. coli ATCC 35218, which was comparable to the positive
control kanamycin B. Based on the data obtained, we found that
compounds bearing a methoxy group at 4-position at A ring (com-
pounds 12–19) showed potent antibacterial activities against E. coli
ATCC 35218. Among them, a comparison of the substitution on
B-ring demonstrated that 4-position-substituted derivatives with
halogen atoms have more potent activity against E. coli ATCC
35218. Most significantly, the stronger electron-withdrawing sub-
stituents the compound contained on B-ring at 4-position, the
more potent it showed (compounds 12, 13, and 14), which was
illustrated by the potency order F > Cl > Br. Meanwhile, the deriva-
tives which have electron-donating substituents (such as CH3,
OCH3) on B-ring exhibited less potent activity against E. coli ATCC
Figure 1. Binding model of compound 12 and E. coli FabH. For clarity, only
interacting residues are displayed. The H-bond (purple) is displayed as line. Asn247
forms hydrogen bond with the nitrogen atom of the pyrazole ring of compound 12,
meanwhile, Arg249 forms hydrogen bond with the oxygen atom of the carbonyl
group of compound 12.
35218, and their MICs values range from 3.125 lg/mL to 25 lg/mL
(compounds 15, 16, and 19).
In addition, compounds 36–43 which bear two chlorine atoms
at 3- and 4-position at A ring exhibited significant activity against

P.-C. Lv et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4657–4660
4659
Table 1
Physical properties and MICs (minimum inhibitory concentrations) (lg/mL) of compounds 4–59
Compounds
Microorganisms
Gram positive
Gram negative
Entry
R1
R2
B. subtilis
S. aureus
S. faecalis
P. aeruginosa
E. coli
E. cloacae
ATCC 6633
ATCC 6538
ATCC 9790
ATCC 13525
ATCC 35218
ATCC 13047
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-OCH3
4-OCH3
4-OCH3
4-OCH3
4-OCH3
4-OCH3
4-OCH3
4-OCH3
4-Br
4-Br
4-Br
4-Br
4-Br
4-F
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
4-F
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
4-F
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
4-F
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
4-F
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
4-F
12.5
12.5
6.25
6.25
12.5
6.25
12.5
6.25
6.25
6.25
12.5
25
6.25
25
6.25
6.25
25
25
12.5
25
25
25
50
50
25
12.5
25
12.5
25
25
12.5
>50
25
>50
6.25
6.25
25
12.5
6.25
6.25
>50
>50
12.5
1.562
3.125
3.125
6.25
12.5
25
6.25
6.25
>50
>50
25
>50
25
12.5
25
25
12.5
25
>50
>50
25
25
>50
25
1.562
1.562
12.6
3.125
3.125
25
25
>50
25
12.5
>50
>50
12.5
>50
25
25
6.25
3.125
1.562
6.25
6.25
25
25
50
12.5
6.25
>50
12.5
25
12.5
>50
25
12.5
25
12.5
12.5
>50
25
25
25
12.5
12.5
12.5
25
12.5
25
12.5
0.39
0.78
1.562
6.25
3.125
6.25
25
>50
12.5
>50
12.5
12.5
>50
12.5
25
>50
3.125
12.5
6.25
>50
6.25
12.5
12.5
25
6.25
3.125
3.125
>50
12.5
>50
>50
12.5
12.5
25
12.5
>50
>50
12.5
>50
12.5
25
25
50
1.562
6.25
3.125
25
12.5
12.5
50
>50
12.5
12.5
3.125
3.125
6.25
1.562
1.562
6.25
12.5
3.125
12.5
12.5
12.5
>50
25
12.5
25
12.5
12.5
25
50
12.5
12.5
25
25
25
1.562
6.25
25
50
6.25
50
25
25
50
12.5
25
50
4-Br
4-Br
4-Br
2-F
2-F
2-F
2-F
2-F
12.5
12.5
50
50
25
25
25
25
25
12.5
25
6.25
6.25
12.5
6.25
1.562
0.78
1.562
6.25
3.125
3.125
6.25
12.5
25
12.5
12.5
>50
12.5
12.5
25
12.5
>50
>50
12.5
12.5
>50
25
2-F
2-F
2-F
25
12.5
1.562
6.25
3.125
12.5
6.25
12.5
12.5
50
12.5
50
50
12.5
12.5
50
25
12.5
50
12.5
12.5
6.25
50
25
12.5
25
1.562
3.125
3,4-2Cl
3,4-2Cl
3,4-2Cl
3,4-2Cl
3,4-2Cl
3,4-2Cl
3,4-2Cl
3,4-2Cl
3,4-2CH3
3,4-2CH3
3,4-2CH3
3,4-2CH3
3,4-2CH3
3,4-2CH3
3,4-2CH3
3,4-2CH3
4-H
4-H
4-H
4-H
4-H
4-H
4-H
4-H
—
25
25
12.5
12.5
25
25
25
25
12.5
6.25
6.25
12.5
3.125
3.125
6.25
12.5
3.125
12.5
12.5
25
12.5
12.5
25
12.5
25
12.5
12.5
6.25
3.125
>50
12.5
>50
>50
25
12.5
25
12.5
25
>50
25
12.5
>50
3.125
1.562
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
4-F
4-Cl
4-Br
4-CH3
4-OCH3
2-Cl
4-H
3,5-2OCH3
—
>50
25
12.5
25
>50
25
12.5
12.5
>50
>50
12.5
25
>50
25
>50
>50
25
25
>50
>50
25
25
12.5
25
>50
>50
25
25
3.125
1.562
25
12.5
1.562
0.39
Penicillin G
Kanamycin B
6.25
3.125
—
—
Table 2
E. coli FabH inhibitory activity of the selected compounds
Furthermore, molecular docking of compound 12 and E. coli
FabH were performed on the binding model based on the E. coli
FabH–CoA complex structure (1HNJ.pdb).27 The binding model of
compounds 12 and E. coli FabH are depicted in Figure 1. It can be
seen from Figure 1 that, in the binding model of compound 12
and E. coli FabH, there are two hydrogen bonds. Asn247 forms
hydrogen bond with the nitrogen atom of the pyrazole ring of com-
pound 12, meanwhile, Arg249 forms hydrogen bond with the oxy-
gen atom of the carbonyl group of compound 12. These results,
along with the data of E. coli FabH inhibitory activity assay
a
Compounds
E. coli FabH IC50
(lM)
Hemolysis LC30 (mg/mL)
12
13
14
15
16
17
18
4.2
7.6
>10
>10
>10
>10
>10
>10
>10
10.4
17.3
19.6
31.4
46.7
a
Lytic concentration 30%.

4660
P.-C. Lv et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4657–4660
9. Daines, R. A.; Pendrak, I.; Sham, K.; Aller, G. S. V.; Konstantinidis, A. K.; Lonsdale,
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E. coli FabH with potent antibacterial activity.
In conclusion, 56 1-acetyl-3,5-diphenyl-4,5-dihydro-(1H)-pyra-
zole derivatives were synthesized and evaluated for their inhibi-
tory activity against E. coli FabH. Data obtained indicated that
compounds 12 and 13 displayed potent antibacterial activity
against E. coli ATCC 35218. Further E. coli FabH inhibitory activity
assay suggested that they are potent E. coli FabH inhibitors. Be-
sides, docking simulation were performed to position compound
12 into the E. coli FabH active site to determine the probable bind-
ing conformation and the results confirmed that compound 12 was
potential inhibitor of E. coli FabH.
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Acknowledgment
The work was financed by Grants from National Natural Science
Foundation of China (Project 30772627), the Jiangsu National Sci-
ence Foundation (No. BK2009239) and Anhui National Science
Foundation (No. 070416274X).
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