Synthesis and in vitro cytotoxic activity of N-2-(2-furyl)-2- (chlorobenzyloxyimino) ethyl ciprofloxacin derivatives

Article abstract of DOI:10.1155/2011/842982

Quinolone are a class of potent broad-spectrum antibacterial drugs that target the bacterial type II DNA topoisomerases (DNA gyrase) and topoisomerase IV. In the present study, the synthesis and evaluation of cytotoxicity activity of a new series of N-pipearzinyl quinolones containing N-2-(furyl-2-yl)-2- (chlorobenzyloxyimino) ethyl moiety (6a-c) have been studied. Preliminary screening showed that one of the new compounds, namely 7-(4-(2-(3- chlorobenzyloxyimino)-2-(furan-2-yl) ethyl) piperazin-1-yl)-1-cyclopropyl-6- fluoro-4-oxo-1, 4-dihydroquinoline-3-carboxylic acid (6b) showed significant cytotoxic activity against human breast tumor cell lines.

Full text of DOI:10.1155/2011/842982

ISSN: 0973-4945; CODEN ECJHAO
E-Journal of Chemistry
2011, 8(3), 1226-1231
http://www.e-journals.net
Synthesis and In Vitro Cytotoxic Activity of
N-2-(2-Furyl)-2-(chlorobenzyloxyimino)
Ethyl Ciprofloxacin Derivatives
ESKANDAR ALIPOUR, NEGAR MOHAMMADHOSSEINI ,
FATEMEH PANAH#, SUSSAN K. ARDESTANI#, MALIHEH SAFAVI§,
ABBAS SHAFIEE§ and ALIREZA FOROUMADI¶∗
Department of Chemistry, North Tehran Branch
Islamic Azad University, Tehran, Iran
#Institute of Biochemistry and Biophysics,
Department of Biochemistry, University of Tehran, Tehran, Iran
§Pharmaceutical Sciences Research Center
Tehran University of Medical Sciences, Tehran, Iran
¶Department of Medicinal Chemistry
Faculty of Pharmacy and Drug Design & Development Research Center
Tehran University of Medical Sciences, Tehran, Iran
aforoumadi@yahoo.com
Received 21 September 2010; Accepted 22 November 2010
Abstract: Quinolone are a class of potent broad-spectrum antibacterial drugs
that target the bacterial type II DNA topoisomerases (DNA gyrase) and
topoisomerase IV. In the present study, the synthesis and evaluation of
cytotoxicity activity of a new series of N-pipearzinyl quinolones containing N-
2-(furyl-2-yl)-2-(chlorobenzyloxyimino) ethyl moiety (6a-c) have been studied.
Preliminary screening showed that one of the new compounds, namely 7-(4-(2-
(3-chlorobenzyloxyimino)-2-(furan-2-yl) ethyl) piperazin-1-yl)-1-cyclopropyl-
6-fluoro-4-oxo-1, 4-dihydroquinoline-3-carboxylic acid (6b) showed significant
cytotoxic activity against human breast tumor cell lines.
Keywords: Quinolones, Ciprofloxacin, Furyl, Oximes, Cytotoxicity.
Introduction
Quinolone antibacterial agents have been known to inhibit DNA gyrase and topoisomerase IV1.
Type II topoisomerases are the enzymes critical for maintaining and controlling the conformations

1227
ALIREZA FOROUMADI et al.
required for DNA replication and transcription2. Indeed, DNA topoisomerase II enzyme catalyzes
the double-strand breakage of DNA to allow strand passage and thereby control the topology and
conformation of DNA3. DNA topoisomerases are found in both eukaryotic and prokaryotic cells
and are targets for chemotherapeutic intervention in antibacterial and anticancer therapies4,5.
Quinolone antibacterials have been developed for clinical use in human medicine6. Moreover, it
was shown that they could also inhibit mammalian topoisomerase and tubulin polymerization and
thus act as potent antitumor drugs as well7. In view of the mechanistic similarities and sequence
homologies exhibited by the prokaryotic type II topoisomerases (DNA gyrase and
topoisomerase IV) and the eukaryotic type II topoisomerases, tentative efforts to selectively Shift
from an antibacterial to an antitumoral activity was made by synthesizing novel classes of
quinolones8-10. The inhibition of DNA gyrase and cell permeability of quinolones is greatly
influenced by the nature of C-7 substitute on the standard structure of 4-quinolone-3-carboxylic
acid11. In addition, substitution of bulky functional groups is permitted at the C-7 position12.
During recent years, a number of quinolones with substitution on piperazine ring at C-7 position
of the basic structure of quinolones were synthesized and evaluated for antibacterial
activities13-14. The inhibition of DNA topoisomerases and the cell permeability of quinolones
are greatly influenced by the nature of the C-7 substitution. In addition, a position on the
quinolone molecule where substitution of bulky functional groups is permitted is at the C-7
position15. Most of the quinolones that showed anticancer activity contain aryl moiety at the
C-7 position. Introduction of aryl ring at C-7 position of quinolone transforms its selectivity
from bacterial to human topoisomerase II16. A number of 7-pyridinyl or 7-(4-hydroxyphenyl)
quinolone derivatives such as CP-115, 953, WIN572946, A-621767 and A-852268, have been
reported to display potent activity towards the eukaryotic type II enzyme and cytotoxicity
against mammalian cells7-10. Recently, several compounds from N-substitueted piperazinyl
quinolone series (Figure 1) that exhibited significant anticancer activity have been reported by
our group17. In continuation of our research program to find a new cytotoxic agent18-21, herein
we would like to report the preparation and in vitro cytototoxicity of N-2-(2-furyl)-2-
(chlorobenzyloxyimino) ethyl ciprofloxacin (6a-c) derivatives as possible anticancer drug.
Y
F
O
S
N
N
Cl
X
COOH
N
Y= O, NOH, NOCH3, NObenzyl
X=CH, N
R= Cyclopropyl, Et
R
Figure 1. N-substitueted piperazinyl quinolones with cytotoxic activities
Experimental
All starting materials, reagents and solvents were purchased from Merck and Aldrich
companies. The purity of the synthesized compounds was confirmed by thin layer
chromatography (TLC) using various solvents of different polarities. Merck silica gel
60 F254 plates were used for analytical TLC. Column chromatography was performed on
Merck silica gel (70-230 mesh) for purification of intermediate and final compounds.
1H-NMR spectra were recorded using a Broker 500 spectrometer and chemical Shifts are
expressed as δ (ppm) with tetramethylsilane (TMS) as internal standard. The IR spectra were
obtained on a Shimadzu 470 spectrophotometer (potassium bromide disks). Melting points
were determined on a Kofler hot stage apparatus and are uncorrected.

Synthesis and In Vitro Cytotoxic Activity
1228
Chemistry
7-(4-(2-(Chlorobenzyloxyimino)-2-(furan-2-yl)ethyl)piperazin-1-yl)-1-cyclopropyl-6-fluoro-
4-oxo-1,4-dihydroquinoline-3-carboxylic acids (6a-c) were synthesized by reaction of
ciprofloxacin, corresponding 2-bromo-1-(furan-2-yl)ethanone O-chlorobenzyl oxime (5a-c)
in DMF in the presence of NaHCO3 (Scheme 2)15. The intermediate O-benzyloximes 5a-c
were prepared by the reaction of substituted O-benzylhydroxylamine hydrochlorides (3)
with 2-bromo-1-(2-furyl)ehanone (2) (Scheme 2). The structures of synthesized compounds
were characterized by IR, 1H NMR and 13C NMR spectral data.
CH2Cl
Cl
A
HONHCOOEt
HONH2.HCl + ClCOOEt
B
1
O
Cl
NH2.HCl
HN
OEt
O
C
Cl
O
3
2
O
CH2Br
O
N
4
O
Cl
CH2Br
3
D
5a-c
O
Cl
O
F
COOH
O
N
F
COOH
5a-c
O
N
N
N
E
HN
N
N
O
6a-c
Scheme 2. Synthesis of N-2-(2-furyl)-2-(chlorobenzyloxyimino)ethyl ciprofloxacin
derivatives (6a-c). Reagent and condition: A) Na2CO3, H2O; B) Na, EtOH, rt; C) 1. NaOH,
H2O, heat, 2. EtOH, HCl; D) MeOH, rt; E) DMF, NaHCO3, rt.
General Procedure for the synthesis of 7-(4-(2-(chlorobenzyloxyimino)-2-(furan-2-
yl) ethyl) piperazin-1-yl)-1-cyclopropyl-6-fluoro-4-oxo-1, 4-dihydroquinoline-3-
carboxylic acid (6a-c)
A mixture of 2-bromo-1-(furan-2-yl)ethanone O-2-chlorobenzyl oxime derivatives (5a-c)
(0.55 mmol), ciprofloxacin (0.5 mmol) and NaHCO3 (0.5 mmol) in DMF (5 mL) was stirred
at room temperature for 6-9 days. After consumption of ciprofloxacin, water (20 mL) was
added and the precipitate was filtered, washed with water and crystallized from EtOH-
CHCl3 to give compounds 6a-c.
7-(4-(2-(2-Chlorobenzyloxyimino)-2-(furan-2-yl)ethyl)piperazin-1-yl)-1-cyclopropyl
-6-fluoro-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (6a)
°
Yield 47%; m.p. 200-201 C; IR (KBr, cm-1): 1628, 1733 (C=O), 3433 (COOH).
1H NMR(DMSO-d6)δ ppm: 1.13-1.18 (m, 2H, cyclopropyl), 1.26-1.32 (m, 2H, cyclopropyl),

1229
ALIREZA FOROUMADI et al.
2.62-2.68 (m, 4H, piperazine), 3.22-3.25 (m, 4H, piperazine), 3.63(s, 2H, CH2), 3.77-
3.82(m, 1H, cyclopropyl), 5.18 (s, 2H, OCH2), 6.56-6.58 (m, 1H, Hb(furyl)), 6.98 (d, 1H,
Ha(furyl) , J=3.1 Hz), 7.41-7.45 (m, 4H, Hbenzyl), 7.54 (d, 1H, H8, J=7.4 Hz), 7.62-7.64 (m, 1H,
Hc(furyl)), 7.89 (d, 1H, H5, J=13.2 Hz), 8.65 (s, 1H, H2), 15.30 (s, 1H, COOH). 13C NMR (125
MHz, DMSO-d6) δ ppm: 7.50, 35.78, 49.39, 50.59, 52.53, 73.07, 106.43, 106.70, 110.75,
110.93, 111.61, 112.33, 127.20, 129.24, 129.69, 130.36, 132.48, 134.88, 139.08, 144.19,
147.67, 147.91, 148.55, 151.98, 153.95, 165.87, 176.28.
7-(4-(2-(3-Chlorobenzyloxyimino)-2-(furan-2-yl)ethyl)piperazin-1-yl)-1-cyclopropyl-
6-fluoro-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (6b)
Yield 30%; m.p. 279-280 °C; IR (KBr, cm-1): 1627, 1730 (C=O), 3443 (COOH).
1H NMR(DMSO-d6)δ ppm: 1.14-1.20 (m, 2H, cyclopropyl), 1.29-1.36 (m, 2H, cyclopropyl),
2.62-2.73(m, 4H, piperazine), 3.25-3.30 (m, 4H, piperazine), 3.66 (s, 2H, CH2), 3.75-3.84
(m, 1H, cyclopropyl), 5.22 (s, 2H, OCH2), 6.56-6.60(m, 1H, Hb(furyl)), 7.00-7.03 (m, 1H,
H
a(furyl)), 7.37-7.44(m, 3H, Hbenzyl), 7.49(s, 1H, Hbenzyl), 7.56(d, 1H, H8, J=6.8 Hz), 7.72-
7.76(m, 1H, Hc(furyl)), 7.90(d, 1H, H5, J= 13.1 Hz), 8.66(s, 1H, H2), 15.12 (s, 1H, COOH)..
13C NMR (125 MHz, DMSO-d6) δ: 7.50, 35.77, 39.99, 50.69, 52.59, 74.69, 106.00, 106.68,
110.75, 110.93, 111.61, 112.33, 126.30, 127.44, 127.59, 130.22, 139.19, 144.12, 140.45,
147.20, 145.64, 147.86, 150.42, 153.98, 164.23, 165.92, 176.27.
7-(4-(2-(4-Chlorobenzyloxyimino)-2-(furan-2-yl) ethyl) piperazin-1-yl)-1-cyclopropyl-
6-fluoro-4-oxo-1, 4-dihydroquinoline-3-carboxylic acid (6c)
Yield 53%; m.p. 187-189 °C; IR (KBr, cm-1): 1626, 1731 (C=O), 3441 (COOH).
1H NMR(DMSO-d6)δ ppm: 1.10-1.20 (m, 2H, cyclopropyl), 1.30-1.38 (m, 2H, cyclopropyl),
2.60-2.70 (m, 4H, piperazine), 3.18-3.28 (m, 4H, piperazine), 3.66 (s, 2H, CH2), 3.75-3.84
(m, 1H, cyclopropyl), 5.27 (s, 2H, OCH2), 6.52-6.54 (m, 1H, Hb(furyl)), 6.99 (d, 1H, Ha(furyl)
,
J=2.8 Hz), 7.40 (m, 2H, Hbenzyl), 7.42-7.44 (m, 1H, Hc(furyl)), 7.50-7.58 (m, 2H, Hbenzyl-H8),
7.70-7.72 (m, 1H, Hbenzyl), 7.85 (d, 1H, H5, J=13.2 Hz), 8.63 (s, 1H, H2), 15.35 (s, 1H,
COOH). 13C NMR (125 MHz, DMSO-d6) δ: 7.50, 35.75, 49.36, 50.58, 52.52, 74.78, 106.70,
106.74, 110.75, 110.93, 111.58, 112.21, 128.28, 129.73, 132.34, 136.74, 139.10, 144.12,
145.20, 147.26, 147.47, 147.93, 148.62, 165.86, 176.31.
Biological activity
The synthesized compounds were tested against a human breast tumor cell lines. The cell
lines were purchased from National Cell Bank of Iran (NCBI). Cells were seeded in
96-well plates at the density of 8000-10,000 viable cells per well and incubated for 48 h to
allow cell attachment. The cells were then incubated for another 48-96 h (depends to cell
cycle of each cell line) with various concentrations of compounds 6a-c. Cells were then
washed in PBS (Phosphate Buffer Saline) and 20 µL of MTT (3-(4,5-dimethylthiazol-2-yl)-
2,5-diphenyl tetrazolium bromide) solution (5 mg/mL) were added to each well. An
additional 4 h of incubation at 37 °C were done and then the medium was discarded.
Dimethyl sulfoxide (60 µL) was added to each well and the solution was vigorously mixed
to dissolve the purple tetrazolium crystals. The absorbance of each well was measured by
plate reader (Anthous 2020; Austria) at a test wavelength of 550 nm against a standard
reference solution at 690 nm. The amount of produced purple formazan is proportional to
the number of viable cells. Two independent experiments in triplicate were performed for
determination of sensitivity to each compound, the IC50 were calculated by linear
regression analysis, expressed in mean SD22.

Synthesis and In Vitro Cytotoxic Activity
1230
Results and Discussion
The compounds 6a-c were tested in vitro against a panel of a human cancer cell lines
including Human breast tumor cell lines. The percentage of growth was evaluated using
MTT colorimetric assay versus controls not treated with test agents. For each compound,
50% inhibitory concentration (IC50) was determined and reported in Table 1. The data for
tamoxifen was included for comparison. The obtained results revealed that compounds 6a,
6c possessed weaker activity against human cancer cell lines in comparison with 6b, which
possess comparable activity with tamoxifen. The results highlight the relationship between
substitutions at 2, 3 or 4 positions of the phenyl ring and cytotoxic activity. Among these
positions, substitution at 3-chlorophenyl position greatly influences their potency and
spectrum of cytotoxic activity.
Table 1. Cytotoxic activity of compounds 6a-c against human breast tumor cell lines in
comparison with tamoxifen
Cl
O
F
COOH
O
N
N
N
N
O
Compound
6a (2-Cl)
6b (3-Cl)
6c (4-Cl)
Tamoxifen
Human breast tumor cell line a
IC50= 25
IC50=5
IC50=10
IC50=12
a IC50 were presented as µg/mL
Conclusion
In conclusion, we have synthesized a novel series of 7-(4-(2-(chlorobenzyloxyimino)-2-
(furan-2-yl)ethyl)piperazin-1-yl)-1-cyclopropyl-6-fluoro-4-oxo-1,4-dihydroquinoline-3-
carboxylic acids (6a-c), which one of them (6b) showed significant cytotoxic activity
against human breast tumor cell lines.
Acknowledgment
This work was supported by grants from Tehran University of Medical Sciences and Iran
National Science Foundation (INSF).
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