Synthesis and in vitro antibacterial activity of 7-(3-alkoxyimino-5-amino/ methylaminopiperidin-1-yl)fluoroquinolone derivatives

Article abstract of DOI:10.1016/j.bmcl.2010.12.073

We report herein the design and synthesis of novel 7-(3-alkoxyimino-5- amino/methylaminopiperidin-1-yl)fluoroquinolone derivatives based on the structures of new fluoroquinolones IMB and DZH. The antibacterial activity of these newly synthesized compounds was also evaluated and compared with gemifloxacin, ciprofloxacin, and levofloxacin. Results revealed that all of the target compounds 10-27 have good potency in inhibiting the growth of Staphylococcus aureus including MSSA (MIC: 0.125-8 μg/mL), Staphylococcus epidermidis including MRSE (MIC: 0.25-16 μg/mL), Streptococcus pneumoniae (MIC: 0.125-4 μg/mL), and Escherichia coli (MIC: 0.25-0.5 μg/mL). In particular, some compounds showed useful activity against several fluoroquinolone-resistant strains, and the most active compound 15 was found to be 16-128, 2-32, and 4-8-fold more potent than the three reference drugs against fluoroquinolone-resistant MSSA, MRSA, and MRSE.

Full text of DOI:10.1016/j.bmcl.2010.12.073

Bioorganic & Medicinal Chemistry Letters 21 (2011) 928–931
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and in vitro antibacterial activity
of 7-(3-alkoxyimino-5-amino/methylaminopiperidin-1-yl)fluoroquinolone
derivatives
⇑
Yibin Zhang, Guoqing Li, Mingliang Liu , Xuefu You, Lianshun Feng, Kai Lv, Jue Cao, Huiyuan Guo
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
We report herein the design and synthesis of novel 7-(3-alkoxyimino-5-amino/methylaminopiperidin-1-
yl)fluoroquinolone derivatives based on the structures of new fluoroquinolones IMB and DZH. The anti-
bacterial activity of these newly synthesized compounds was also evaluated and compared with gemi-
floxacin, ciprofloxacin, and levofloxacin. Results revealed that all of the target compounds 10–27 have
Received 19 October 2010
Revised 14 December 2010
Accepted 15 December 2010
Available online 19 December 2010
good potency in inhibiting the growth of Staphylococcus aureus including MSSA (MIC: 0.125–8
Staphylococcus epidermidis including MRSE (MIC: 0.25–16 g/mL), Streptococcus pneumoniae (MIC:
0.125–4 g/mL), and Escherichia coli (MIC: 0.25–0.5 g/mL). In particular, some compounds showed use-
lg/mL),
l
Keywords:
l
l
Antibacterial activity
Fluoroquinolone
Synthesis
ful activity against several fluoroquinolone-resistant strains, and the most active compound 15 was found
to be 16–128, 2–32, and 4–8-fold more potent than the three reference drugs against fluoroquinolone-
resistant MSSA, MRSA, and MRSE.
Ó 2010 Elsevier Ltd. All rights reserved.
Quinolones represent an extremely successful family of antibi-
otics that have potent, a broad spectrum of antibacterial activity,
and relatively few side effects.1 They inhibit two type-II topoi-
somerases, DNA gyrase and topoisomerase IV, by binding to the
intermediate catalytic enzyme–DNA complex. The stabilization of
the resulting quinolone-enzyme–DNA complex leads to the gener-
ation of double-strand DNA breaks that trigger a cascade of events
leading to cell death.2,3
Most of the quinolones currently on the market or under
development are generally characterized by a broad antibacterial
spectrum, but their activity against clinically important Gram-po-
sitive cocci, including Staphylococci, Streptococci, and Enterococci,
is relatively moderate. This insufficient activity has not only lim-
ited their use in infections caused by these organisms, but has also
contributed to the rapidly developing quinolone resistance. Thus,
recent efforts have been directed toward the synthesis of new
quinolones that can provide improved Gram-positive antibacterial
activity, while retaining the good Gram-negative activity of early
fluoroquinolones, such as ciprofloxacin (CPFX) and ofloxacin.4
The general chemical structure of the quinolone antibacterial
agents consists of a 4-quinolone/naphthyridone-3-carboxylic acid
nucleus and a secondary amino group attached to the C-7 position
of the heterocyclic core, which has a great impact of modulating
potency, spectrum and pharmacokinetics.5 In general, the optimal
substituents have proven to be 5- and 6-membered nitrogen
heterocycles that contain peripheral nitrogens, such as piperazinyl,
pyrrolidinyl, and piperidinyl groups. However, so far the piperidi-
nyl-based quinolone analogs of the three are the least reported
in the literature.6
As part of an ongoing program to find potent new quinolones
displaying strong Gram-positive antibacterial activity, we have fo-
cused our attention on introducing new functional groups to the
piperidine ring and found IMB (Fig. 1), a 8-methoxyl fluoroquino-
lone containing an 3-amino-4-methoxyiminopiperidin-1-yl group
at the C-7 position, have good in vitro and in vivo antibacterial
activity which was comparable to moxifloxacin and gemifloxacin
(GMFX).7–13 It was also reported that DZH, a fluoronaphthyridone
with a 3-methoxyimino-4-methylaminopiperidin-1-yl group at
the C-7 position, showed excellent in vitro antibacterial activity
against Gram-positive and Gram-negative organisms, including
methicillin-sensitive Staphylococcus aureus (MSSA), Streptococcus
pneumoniae, E. faecalis, and P. aeruginosa, which was better than
vancomycin and GMFX.6
⇑
Corresponding author. Tel.: +86 10 63036965; fax: +86 10 63047865.
E-mail address: lmllyx@yahoo.com.cn (M. Liu).
Figure 1. Structures of IMB and DZH.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.12.073

Y. Zhang et al. / Bioorg. Med. Chem. Lett. 21 (2011) 928–931
929
converted into aminopiperidine 3 by Rh/C-catalyzed hydrogena-
tion in water at 80 °C, during which two equivalents of phosphonic
acid was added to avoid the catalyst poisoning caused by the ami-
no group. The amine 3 was subsequently treated with di-tert-butyl
dicarbonate (Boc2O) in NaOH–H2O–EtOH solution to form bis-Boc-
protected amino alcohol 4, which was oxidated to the correspond-
ing piperidone 5 by Jones reagent, with an overall yield of 26% for
the three steps. The alkoxyimino group was introduced into the
ring by coupling 5 with methoxylamine or ethoxylamine at 70 °C
to afford amines 6a,b. Methylation of the amines 6a,b by reaction
with methyl iodide in the presence of sodium hydride produced
methylamines 7a,b.
Removal of the bis-Boc-protecting groups on the amines 6a,b
and methylamines 7a,b turned out to be a challenging task, mainly
due to the instability of the alkoxyimino groups to hydrochloric
acid and trifluoroacetic acid, which are commonly used in the
Boc deprotection reaction. After a number of attempts, we success-
fully removed the Boc groups in neutral condition by reaction with
trimethyl silane iodine (TMSI) to afford the new piperidine deriva-
tives 8a–d.
Figure 2. Design of the new piperidinyl side chain.
Based on these research results, a series of new fluoroquinolone
compounds containing substituted piperidines were designed and
synthesized. These derivatives are structurally novel, having an
alkoxyimino group at the 3-position and an amino/methylamino
group at the 5-position of the piperidine ring (Fig. 2), which are
the position isomers of the side chains at the C-7 position of IMB
and DZH. Our primary objective was to optimize the potency of
these compounds against Gram-positive and Gram-negative
organisms.
The new piperidine derivatives and novel fluoroquinolones de-
scribed herein were synthesized as shown in Schemes 1 and 2,
respectively. Using copper sulfate as the catalyst, amination of
5-bromopyridin-3-ol 1 performed smoothly in ammonia water at
100 °C gave 5-aminopyridin-3-ol 2. The aminopyridine 2 was
Finally, the target compounds 10–27 were obtained by coupling
the new piperidine derivatives 8a–d with various compounds con-
taining quinolone and naphthyridone cores and then hydrolysis
Scheme 1. Reagents and conditions: (i) NH3ꢀH2O, CuSO4ꢀ5H2O, 100 °C, 12 h, 75%; (ii) H3PO4, H2, 5% Rh/C, H2O, 80 °C, 40 h; (iii) NaOH, Boc2O, EtOH–H2O, rt, 2 h; (iv) Jones
reagent, Me2CO, 0 °C, 0.5 h, 26% (three-step yield); (v) R1ONH2ꢀHCl, Et3N, EtOH–H2O, 70 °C, 2 h, 72%; (vi) NaH, MeI, MeCN, 0 °C, 3 h, 81%; (vii) TMSI, CH2Cl2, rt, 0.5 h.
Scheme 2. Reagents and conditions: (viii) (a) Et3N, MeCN, rt to 50 °C, 10–70 h; (b) 2% NaOH/H2O, rt, 2 h; (c) 6 N HCl, H2O, rt, 9–42% (from 9a to 9g).

930
Y. Zhang et al. / Bioorg. Med. Chem. Lett. 21 (2011) 928–931
according to the well-established literature procedures
(Scheme 2).14 In the case of quinolones 10–13, condensation of
ester 9a with 8a–d was performed in the presence of triethyl-
amine. However for 14–27, boric chelates 9b–g were required to
increase reactivity. All of the synthetic compounds were well
characterized through the spectral characteristics.15
Fluoroquinolones 10–27 were evaluated for their in vitro anti-
bacterial activity against representative Gram-positive and Gram-
negative strains using standard techniques and the minimum
inhibitory concentration values (MICs) along with those of the ref-
erence drugs GMFX, CPFX, and levofloxacin (LVFX) for comparison
are shown in Table 1.16
compound 15 was found to be 16–128, 2–32, and 4–8-fold more
potent than the references against fluoroquinolone-resistant
MSSA, MRSA, and MRSE. In addition, compound 14 was 2–32-fold
more potent than the references against fluoroquinolone-resistant
S. hemolyticus, and the same for compound 16 against MRSA.
Generally, the activity of the quinolone nuclei against Gram-po-
sitive strains was in the order: 1-cyclopropyl-8-methoxylquino-
lone > 1-cyclopropyl-8-fluoroquinolone > 1-ethyl-8-fluoroquinolone >
levofloxacin nuclei > 1-cyclopropylquinolone ꢁ 1-cyclopropyl-8-
difluoromethoxylquinolone > 1-cyclopropyl-1,8-naphthyridine. In
addition, fluoroquinolones featuring ethyloxime-incorporated pip-
eridino-substitution at the C-7 position were at least as potent as
the analogs containing methyloxime, and introduction of a methyl
group to the amino group at the 5-position of the piperidine ring
did not change the Gram-positive antibacterial activity remarkably.
As for Gram-negative strains, the novel fluoroquinolones 10–27
did not provided satisfactory potency. All of them were generally
less active than the reference drugs GMFX, CPFX, and LVFX,
although they demonstrated good activity against E. coli (MIC:
All of the target compounds 10–27 have potent antibacterial
activity against the twelve tested strains. They generally exhibited
good potency in inhibiting the growth of most tested Gram-posi-
tive strains, such as S. aureus including MSSA (MIC: 0.125–8
mL), Staphylococcus epidermidis including methicillin-resistant
S. epidermidis (MRSE) (MIC: 0.25–16 g/mL) and S. pneumoniae
(MIC: 0.125–4 g/mL), as well as some of the tested Gram-negative
strains such as Escherichia coli (MIC: 0.25–0.5 g/mL). It is worth
lg/
l
l
l
0.25–0.5
Fluoroquinolones 10–27 were further examined for cytotoxicity
(CC50) in a mammalian Vero cell line from 1024 to 4 g/mL concen-
lg/mL).
noting that some compounds showed useful activity against sev-
eral fluoroquinolone-resistant strains. For example, compounds
l
14–19, 21, 23, 25, and 27 (MICs: 0.125–0.5
fold more potent than the three reference drugs (MICs: 2–16
mL) against MSSA. Compounds 14–17, 19–21, 23, and 25 (MICs:
1–4 g/mL) showed 2–16-fold more potent activity than the refer-
l
g/mL) were 4–128-
trations. After 24 h of exposure, viability was assessed and the re-
sults are reported in Table 1. The eighteen compounds when tested
showed CC50 values ranging from 593 to 1524 lM. A comparison of
the substitution pattern at the C-7 position demonstrated that eth-
lg/
l
ences (MICs: 8–16 lg/mL) against MRSE. The most active
yloxime-incorporated piperidino-substitutions were generally less
Table 1
In vitro antibacterial activity and cytotoxicity of novel fluoroquinolones 10–27
O
F
COOH
R2HN
N
X
N
R3
NOR1
Strains
R1
R2
R3
X
MIC (
S.p
l
g/mL)
CC50 (lM)
Compound
S.a.1
S.a.2
S.a.3
S.e.1
S.e.2
S.h
E.c.1
E.c.2
K.p
P.a.1
P.a.2
10
11
12
13
14
15
16
17
18
19
20
21
22
23
Me
Et
Me
Et
Me
Et
Me
Et
Me
Et
Me
Et
Me
Et
H
H
Me
Me
H
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
c-Pr
N
N
N
N
COMe
COMe
COMe
COMe
CF
CF
CF
CF
COCHF2
COCHF2
4
4
0.5
2
0.25
0.25
0.5
0.5
0.25
0.25
0.25
0.5
2
8
2
4
4
128
16
128
8
4
2
2
8
8
4
2
4
2
8
2
1
1
0.25
4
0.5
2
0.5
2
16
8
16
16
4
2
4
4
8
4
2
2
8
4
4
4
4
128
128
>128
>128
4
8
8
128
128
64
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.5
128
128
>128
>128
64
128
128
128
128
64
128
128
128
128
4
8
64
128
128
8
128
128
822
967
645
731
885
879
1088
1524
739
1323
1023
1013
667
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.5
>128
>128
4
>128
128
0.5
0.125
0.5
0.5
0.5
0.5
2
0.25
8
0.25
0.25
0.125
0.5
1
0.25
0.25
1
1
4
1
H
Me
Me
H
128
128
H
>128
128
>128
>128
>128
Me
Me
H
8
4
128
4
8
128
>128
>128
>128
8
128
>128
128
128
128
128
0.25
0.25
H
0.5
0.25
1
1523
24
Me
H
0.5
2
128
4
8
0.5
64
0.25
0.5
64
8
64
1002
O
O
25
Et
H
0.25
0.5
4
0.5
2
0.25
8
0.25
0.25
64
8
32
1244
26
27
GMFX
LVFX
CPFX
Me
Me
H
H
c-Pr
Et
CH
CF
0.5
4
0.25
2
8
16
64
32
4
32
64
8
16
16
8
8
16
4
0.5
0.008
0.015
0.03
128
128
8
32
64
0.25
0.25
0.015
0.015
0.008
0.25
0.25
0.06
0.5
64
>128
32
32
32
2
8
2
4
>128
>128
1
2
1
593
913
0.25
0.008
0.125
0.25
16
0.25
2
1
0.25
32
Abbreviations: S.a.1: Staphylococcus aureus ATCC25923; S.a.2: methicillin-sensitive Staphylococcus aureus 10-04; S.a.3: methicillin-resistant Staphylococcus aureus 10-05;
S.e.1: methicillin-sensitive Staphylococcus epidermidis 10-4; S.e.2: methicillin-resistant Staphylococcus epidermidis 10-5; S.p: Streptococcus pneumoniae 1001; S.h: Streptococcus
hemolyticus 1002; E.c.1: Escherichia coli ATCC25922; E.c.2: Escherichia coli 10-02; K.p: Klebsiella pneumoniae 10-1; P.a.1: Pseudomonas aeruginosa ATCC27853; P.a.2: Pseu-
domonas aeruginosa 10-1.

Y. Zhang et al. / Bioorg. Med. Chem. Lett. 21 (2011) 928–931
931
7. Wang, X. Y.; Guo, Q.; Wang, Y. C.; Liu, B. Q.; Liu, M. L.; Sun, L. Y.; Guo, H. Y. Acta
Pharm. Sin. 2008, 43, 819.
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cytotoxic than the analogs containing methyloxime. In addition, a
methylamino group in place of the amino group at the 5-position
of the piperidine ring increased cytotoxicity of naphthyridones
(10–13), but contrary for quinolones (14–21).
In summary, we report herein the design and synthesis of some
novel 7-(3-alkoxyimino-5-amino/methylaminopiperidin-1-yl)fluo-
roquinolone derivatives based on the structures of new fluoroquin-
olones IMB and DZH. The antibacterial activity of these newly
synthesized compounds was also evaluated and compared with
GMFX, CPFX, and LVFX. Results reveal that all of the synthesized
fluoroquinolones 10–27 have good potency in inhibiting the
growth of S. aureus including MSSA, S. epidermidis including MRSE,
S. pneumoniae and E. coli. In particular, some compounds showed
useful activity against several fluoroquinolone-resistant strains,
and the most active compound 15 was found to be 16–128,
2–32, and 4–8-fold more potent than the three reference drugs
against fluoroquinolone-resistant MSSA, MRSA, and MRSE.
10. Wang, J. X.; Guo, Q.; Chai, Y.; Feng, L. S.; Guo, H. Y.; Liu, M. L. Chin. Chem. Lett.
2010, 21, 55.
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Y. Bioorg. Med. Chem. Lett. 2010, 20, 5195.
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Eur. J. Med. Chem. 2010, 45, 6101.
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116, 152003e.
15. To a stirring solution of 7b (1.0 g, 2.7 mmol) in dry dichloromethane (20 mL)
was added TMSI (0.8 mL, 5.7 mmol) at room temperature under an atmosphere
of nitrogen and stirred for 0.5 h at the same temperature. Enough methanol
was added to quench the reaction and then the mixture was concentrated
under reduced pressure. The residue containing 8d was dissolved in dry
acetonitrile (20 mL), and to this solution dry triethylamine (1.1 mL, 8.1 mmol)
and 9c (0.9 g, 2.2 mmol) were added. The mixture was stirred at room
temperature under an atmosphere of nitrogen for 20 h and then was
concentrated under reduced pressure. The residue was dissolved in 2%
sodium hydroxide solution (15 mL) and stirred for 2 h at the same
Acknowledgment
temperature. The reaction mixture was adjusted to pH
hydrochloric acid. The resulting solid was collected by suction to afford 21
[0.4 g, 42% (from 9c), purity by HPLC: 54% (X) + 45% (Y) = 99%] as a yellow solid.
7 with 6 N
The work was supported by the National S&T Major Special Pro-
ject on Major New Drug Innovation. Item No. 2009ZX09301-003.
1H NMR (400 MHz, CDCl3):
NOCH2CH3), 2.50–2.60 (4H, m, NCH3, Y-C4 -H), 2.80–2.83 (1H, m, X-C4 -H),
d
1.17–1.32 (7H, m, 2 ꢂ cyclopropyl CH2,
0
0
Supplementary data
0
0
0
3.10–3.21 (2H, m, C5 -H, C4 -H), 3.37 (1H, br, cyclopropyl CH), 3.71 (1H, br, C6
-
0
0
0
H), 3.94–4.14 (6H, m, NOCH2CH3, X-C2 -2H, Y-C2 -H, C6 -H), 4.37 (1H, d,
0
J = 15.6 Hz, Y-C2 -H), 7.89 (1H, d, J = 11.6 Hz, C5-H), 8.77 (1H, s, C2-H). MS (ESI,
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.12.073.
m/z): 435 (M+H)+. HRMS (ESI, m/z): C21H25F2N4O4 (M+H)+. Calcd: 435.18439,
found: 435.18042.
16. Compounds 10–27 were evaluated for their in vitro antibacterial activity using
conventional agar-dilution method in comparison to the reference drugs.
Drugs (10.0 mg) were dissolved in 0.1 N sodium hydroxide solution and water
(10 mL). Further progressive twofold serial dilution with melted Mueller-
Hinton agar was performed to obtain the required concentrations of 128, 64,
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were incubated with 104 colony forming units (cfu) and incubated at 35 °C for
18–24 h. The MIC was the lowest concentration of the test compound, which
resulted in no visible growth on the plate.