An efficient one-pot synthesis and photoinduced DNA cleavage studies of 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-yl)quinolines

Article abstract of DOI:10.1016/j.bmcl.2012.08.034

4,5-Dihydroisoxazoles continue to attract considerable interest due to their wide spread biological activities. Here, we identify an efficient protocol for the preparation of 4,5-dihydroisoxazoles (2-isaxazolines) (4a-g) from quinolinyl chalcones. The nuc

Full text of DOI:10.1016/j.bmcl.2012.08.034

Bioorganic & Medicinal Chemistry Letters 22 (2012) 6095–6098
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Bioorganic & Medicinal Chemistry Letters
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An efficient one-pot synthesis and photoinduced DNA cleavage studies
of 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-yl)quinolines
P. J. Bindu a, , K. M. Mahadevan a, T. R. Ravikumar Naik b
⇑
a Department of Studies and Research in Chemistry, Kuvempu University, Shankaraghatt 577 451, India
b Department of Organic Chemistry, Indian Institute of Science, Bangalore 560 012, India
a r t i c l e i n f o
a b s t r a c t
Article history:
4,5-Dihydroisoxazoles continue to attract considerable interest due to their wide spread biological activ-
ities. Here, we identify an efficient protocol for the preparation of 4,5-dihydroisoxazoles (2-isaxazolines)
(4a–g) from quinolinyl chalcones. The nucleolytic activities of synthesized compounds were investigated
by agarose gel electrophoresis. All these compounds were showed the remarkable DNA cleavage activity
(concentration dependent) with pUC19 DNA at 365 nm UV light. The DNA cleavage activity was signifi-
cantly enhanced by the presence of iminyl and carboxy radicals of DIQ.
Received 12 April 2012
Revised 8 August 2012
Accepted 10 August 2012
Available online 17 August 2012
Keywords:
Ó 2012 Elsevier Ltd. All rights reserved.
Quinolinyl chalcones (QCs)
2-Chloro-3-(5-aryl-4,5-dihydroisoxazol-3-
yl)quinolines (DIQs)
DNA
Photo cleavage
The interaction of small molecules with nucleic acids has re-
ceived considerable attention during the past decade.1 Organic
molecules are attractive reagents for the cleavage of nucleic acids
due to their inherently diverse structure and reactivity. Metal-
based photosensitizers are the foremost and widely used antican-
cer drugs for cancer therapy, but these possess inherent side ef-
fects, solubility and acquired drug resistance. Therefore,
considerable attempts are being made to replace these drugs with
suitable alternatives, and numerous small molecules have been
synthesized and tested for their anticancer activities.2 Quinoline
compounds are regarded as the most promising alternatives to me-
tal complexes as anticancer drugs.2
2-Isoxazolines are unique in their chemical behavior not only
among heterocyclic compounds in general, but also among related
azoles. This is because 2-isoxazoline possesses the typical proper-
ties of the aromatic system, which are in fact rather pronounced in
these derivatives, together with the high liability of the ring under
certain conditions, particularly at the nitrogen–oxygen bond.3a,3b
2-Isoxazolines may show interesting medicinal or crop protection
properties or have some other industrial utility. It has been well
established that isoxazole derivatives possess a wide spectrum of
chemotherapeutic activities including antifungal,4a antibacterial,4b
anticonvulsant,4c anti-inflammatory,4d anti-viral,4e analgesic,4f
antitumar,4g chemotherapy activity.4h Isoxazoline derivatives also
show a good potency in animal models of thrombosis.4i Drugs such
as Isocarboxazid, Valdecoxib, Oxacillin, Leflunomide, and Micafun-
gin are the examples5 to substantiate the pharmaceutical accep-
tance of 2-isoxazoline heterocyclic systems.
On the other hand, the quinoline moieties are reported to act as
anticancer and antiviral agents, protein kinase inhibiters for onco-
lytic drugs, PDE4 inhibitors for antiasthmatic and anti-inflamma-
tory agents,6a anticonvulsant agents,6b non-opioid analgesics6c
and antimalarial agents.6d,6e
A considerable number of methods have been reported for the
synthesis of substituted isoxazoles.7,8 The most convenient synthe-
sis of isoxazoline and isoxazole ring systems has been executed in
the literature via 1,3-dipolar cycloaddition7 (1,3-DC) reactions of
alkenes or alkynes with nitrile oxides generated in situ from aldox-
imes.8 Although this three-step procedure is quite general, we
found that the parallel synthesis of isoxazoline due to the number
of extraction, filtration, and solvent evaporation steps, we sought a
one-pot procedure to overcome these limitations. Thus, the devel-
opment of facile and regioselective methods for the synthesis of
DIQ is desirable. We envisioned that, the condensation between
hydroxylamine and
a
,b-unsaturated aldehydes/ketones9 would
be a suitable strategy to achieve this goal (Scheme 1).
Furthermore, there is a real need for discovery of new com-
pounds endowed with anticancer activity, possibly acting through
mechanisms of actions, which are distinct from those of well
known classes of anticancer agents.10 Thus, a single molecule con-
taining more than one pharmacophore, each with different mech-
anism of action could be beneficial for the treatment of cancer. We
have been interested in developing the DNA cleavage methods,
⇑
Corresponding author. Tel.: +91 990 079 2675.
E-mail address: bindu12_naik@rediffmail.com (P.J. Bindu).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.08.034

6096
P. J. Bindu et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6095–6098
O
O
H
N
Cl
R
N
Cl
3a-g
1
NH2OH.HCl
Et3N, DCM, 8-10 hr
+
R = H, CH3, OCH3, OH, Br, Cl, NO2
N
O
O
H3C
N
Cl
R
R
2a-g
4a-g
Scheme 1. Synthesis of 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-yl)quinolines.
Ph
H
Ph
H
O
N
H
N
O
O
hv
N
Cl
Cl
Ph
N
N
Cl
N
Scheme 2. Photo-irradiation of DIQs.
Table 1
particularly by radical species. Taking inspiration from the above
and as a part of our continuing research on the synthesis of biolog-
ically active substituted quinolines.11 In similar manner, we have
synthesized DIQs to cleave the pUC 19 DNA upon UV irradiation
(Scheme 2). The classical synthesis of the title compounds involves
the Claisen–Schmidt condensation of 2-chloroquinoline-3-carbal-
dehyde with substituted acetophenones to give quinolinyl chal-
cones (QC) (3a–g),12 which on cyclisation with hydroxylamine
hydrochloride in presence of Et3N base furnished 2-chloro-3-(5-
aryl-4,5-dihydroisoxazol-3-yl)quinolines (4a–g) in good yields
(Scheme 1). The results showed that, the presence of Et3N in
dichloromethane increases the yield of isoxazolines.
Synthesis of 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-yl)quinolines
Entrya
R
Time (hrs)
Yieldb (%)
M.P(°C)
4a
4b
4c
4d
4e
4f
H
8–10
8–10
8–10
8–10
8–10
8–10
8–10
80
83
85
90
85
91
90
132–134
138–140
141–143
158–160
162–164
152–154
169–171
p-CH3
p-OCH3
p-NO2
p-OH
p-Cl
4g
p-Br
a
All the products were characterized by elemental analysis, 1H NMR, 13C NMR
and mass spectral data.
b
Yields of isolated products.
We have successfully obtained DIQ under the treatment of
hydroxylamine with QC. The most efficient set of conditions were
to employ 1.8 mmol of quinolinyl chalcones (QC), 64.0 mmol of
hydroxylamine and 64.0 mmol of Et3N in DCM at room tempera-
ture (Table 1 and Scheme 1 and 2). However, by using of triethyl
amine in DCM found to be more efficient catalyst in terms of high
yield and easy isolation of the products. As shown in Table 1, a ser-
ies of DIQs were readily obtained in good to excellent yields from
QC and hydroxylamine under the optimal conditions. In general,
substrates containing an electron-withdrawing group were more
active to offer higher yields (entries 4, 6, and 7). This result implied
that electron density at the aromatic ring played an important role
in this process. The synthesized DIQs (4a–g) were characterized by
IR, 1H NMR, 13C NMR and mass spectral data.
It has been reported that most DIQ derivatives upon irradiation
undergo N–O bond fission as the primary process and afford a vari-
ety of products depending on the structure of the starting materi-
als.13 On photo-irradiation, the weak N–O bond of the DIQ can be
selectively cleaved to generate the iminyl and carboxy radicals
which can then cause the cleavage of the DNA as shown in
Scheme 2.
observed for the intact supercoiled Form I.11,14 When scission oc-
curs on one strand (nicking), the supercoil relaxes to generate a
slower-moving, open-circular Form II. If both strands are cleaved,
a linear Form III is generated, migrating between Form I and Form
II.11,14 The 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-yl)quinolines
(DIQs)16 were photolysed at 365 nm, at different concentrations
in the presence of pUC-19 DNA. Solutions were irradiated for 2 h,
in 1:9 DMSO: Trisbuffer (20 lM, pH-7.2) at 365 nm. Figures 1–3
shows the Photo cleavage results of pUC 19 DNA by DIQ, it can
be seen that no DNA cleavage was observed for control experiment
in which the DIQ was absent (Lane 1).
However, the gel separation of pUC19 DNA was observed after
incubation with different concentration of DIQ in the dark (Lane
2–8), while after irradiation at 365 nm for 2 h, in 1:9 DMSO/Trisbuf-
fer (20 lM, pH 7.2). The Micromolar concentrations of the DIQ
showed DNA cleavage as evidenced by the disappearance of Form
I and appearance of Form II and Form III (Lane 2–8), demonstrating
that the DNA cleavage occurred by photoinduced rather than the
hydrolytic reaction pathway. Figure 1 indicates that, there is no mo-
ment of supercoiled Form I to the nicked Form II and it clears that, at
Recently, some small organic molecules were known to cleave
DNA under UV–vis light.11,14,15 When circular plasmid DNA is
subject to electrophoresis, relatively fast migration is generally
lower concentrations (40
lM) DNA cleavage was not effective.
However, with increasing the micromolar concentration of DIQ

P. J. Bindu et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6095–6098
6097
Figure 1. Photo cleavage of DNA by 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-
yl)quinolines were irradiated with UV light at 365 nm. Supercoiled DNA runs at
position I (SC) and nicked DNA at position II (NC). Lane; 1: control DNA (without
compound), Lane; 2: 40
40 M (2d), Lane; 6: 40
l
l
M (2a), Lane; 3: 40
M (2e), Lane; 7: 40
l
l
M (2b), Lane; 4: 40 M (2c), Lane; 5:
M (2f), Lane; 8: 40 lM (2g).
l
l
Figure 4. Plot representation the percentage of pUC 19 DNA cleavage with different
concentrations of DIQs (40–80 lM) at 37 °C.
concentration of 40
of supercoiled DNA to nicked form DNA. Even at 60
tion, the DIQs can exhibit about 50% conversion of form I to II.
However, at the 80 M concentration, the DIQs promoted almost
l
M, DIQs can almost promote 20% conversion
lM concentra-
Figure 2. Light-induced cleavage of DNA by 2-chloro-3-(5-aryl-4,5-dihydroisox-
azol-3-yl)quinolines with UV light at 365 nm. Supercoiled DNA runs at position I
(SC) and nicked DNA at position II (NC). Lane; 1: control DNA (with out compound),
l
85% conversion supercoiled DNA to nicked and linear DNA.
In conclusion, an efficient approach for the highly regioselective
synthesis of a diverse array of DIQ was developed. The synthesized
DIQs were demonstrated as versatile synthons for the DNA Photo
cleavage studies. Study showed that, the iminyl and carboxy radi-
cals resulting from N–O bond homolysis of DIQs are highly reactive
radicals, which abstracts hydrogen atoms efficiently at C-40 of 2-
deoxyribose of B DNA. The presences of aryl and quinoline core
in DIQs were also responsible for spontaneous DNA photocleavage
activity. The details of the mechanism of its DNA photoinduced
cleavage remain to be studied in the future.
Lane; 2: 60
Lane; 6: 60
l
l
M (2a), Lane; 3: 60
M (2e), Lane; 7: 60
l
l
M (2b), Lane; 4: 60 M (2c), Lane; 5: 60
M (2f), Lane; 8: 60 lM (2g).
l
lM (2d),
Figure 3. DNA Photo cleavage of 2-chloro-3-(5-aryl-4,5-dihydroisoxazol-3-yl)quin-
olines at 365 nm with UV light irradiation. Supercoiled DNA runs at position I (SC)
and nicked DNA at position II (NC). Lane; 1: control DNA (without compound), Lane;
Acknowledgments
We are grateful to Prof. H. S. Bhojya Naik for his suggestions
during our research work and Indian Institute of science, Bangalore
for providing the NMR and mass spectra.
2: 80
6: 80
l
l
M (2a), Lane; 3: 80
M (2e), Lane; 7: 80
l
l
M (2b), Lane; 4:80
M (2f), Lane; 8: 80
l
M (2c), Lane; 5: 80
lM (2d), Lane;
lM (2g).
form 40 lM to 60 lM (lanes 2–8), the amount of Form I diminished
gradually with increasing in Form II of pUC 19 DNA as shown in
Supplementary data
Figure 2.
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2012.
08.034.
Further, the nucleolytic efficiencies of DIQs were investigated at
the 80
verted in to form II and form III DNA (Fig. 3). But at higher concen-
trations (120 M) was not examined because of the precipitation
lM concentration, the pUC19 DNA was completely con-
l
References and notes
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the future. The percentage of DNA cleavage with different
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phenylpropen-2-ones (3a–g) (500 mg, 1.8 mmol) in CH2Cl2 (30 mL) was
added hydroxylamine hydrochloride (220 mg, 64.0 mmol) and Et3N (5.0 mL,
64.0 mmol) at room temperature. After being stirred for 8–10 h, the reaction
mixture was quenched with water and extracted with CH2Cl2, which was
washed repeatedly with water and dried over anhydrous NaSO4. Purification
by flash column chromatography on silica gel n-hexane:EtOAc (1:1) gave
(480 mg, 92%) as yellow needles (4a–g).