2-Phenylquinoline-carbohydrate hybrids: Molecular design, chemical synthesis, and evaluation of a new family of light-activatable DNA-cleaving agents

Article abstract of DOI:10.1002/(SICI)1521-3773(19991216)38:24<3733::AID-ANIE3733>3.0.CO;2-W

Artificial intercalator-carbohydrate hybrids such as that shown in the scheme cleave DNA at the site of guanine on irradiation with UV light with a long wavelength. The hybrids also exhibit strong cytotoxicity when irradiated. The hybrid system is very im

Full text of DOI:10.1002/(SICI)1521-3773(19991216)38:24<3733::AID-ANIE3733>3.0.CO;2-W

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a
Spex Fluorolog II spectrofluorimeter. Details of the correction of
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timescale were determined with an IBH single photon counting apparatus
(lexc ˆ 337 nm) or a single-shot Nd:YAG laser apparatus (lexc ˆ 532 nm).
Transient absorption spectra and lifetimes with picosecond and nanosecond
resolution were obtained with two pump and probe systems based on
Nd:YAG lasers; excitation with the second (532 nm) or third harmonic
(355 nm) was used. Details of this time-resolved spectroscopy equipment
were reported earlier.[14] Experimental uncertainties were estimated to be
Æ8% for lifetime determination, Æ20% for quantum yields, and Æ3 nm
for emission and absorption peaks.
[9] The accessible surface of the central core to the solvent molecules in
the CuG2 dendrimer was calculated with the MSEED program.[10]
First we calculated the area of exposure to the solvent for CuG0 and
G1CO2tBu, which amounted to 620 Š2 and 840 Š2, respectively. Since
for CuG2 the surface available to the solvent is 6710 Š2, the exposure
of the central core amounts, at most, to about 10%. For CuG3 this can
assumed to be less than 5%. If we consider that in the apolar CH2Cl2
medium the counterion is likely to be in tight vicinity to the
Received: May 31, 1999
Revised version: July 23, 1999 [Z13491IE]
‡
[Cu(phen)2] central core, we can conclude that the interior of the
dendrimer is virtually inaccessible to external contact.
[10] G. Perrot, B. Cheng, K. D. Gibson, J. Vila, K. A. Palmer, A. Nayeem,
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German version: Angew. Chem. 1999, 111, 3895 ± 3899
Keywords: copper ´ dendrimers ´ electrochemistry ´ full-
erenes ´ UV/Vis spectroscopy
[13] N. Armaroli, F. Diederich, C. O. Dietrich-Buchecker, L. Flamigni, G.
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and only in matrices below 5 K or at 77 K in presence of solvents that
contain heavy atoms.
[17] For the calculation of the thermodynamic driving force for electronic
excited states from spectroscopic and/or electrochemical parameters,
see V. Balzani, F. Scandola, Supramolecular Photochemistry, Ellis
Horwood, Chichester, 1991, p. 44.
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Acc. Chem. Res. 1998, 31, 26.
2-Phenylquinoline ± Carbohydrate Hybrids:
Molecular Design, Chemical Synthesis, and
Evaluation of a New Family of Light-
Activatable DNA-Cleaving Agents**
Â
[5] Recent examples: C. J. Hawker, K. L. Wooley, J. M. J. Frechet, J. Am.
Chem. Soc. 1993, 115, 7638; P. J. Dandliker, F. Diederich, M. Gross,
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1995, 107, 2906; Angew. Chem. Int. Ed. Engl. 1995, 34, 2725; H.-F.
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Commun. 1996, 1821; J. P. Collman, L. Fu, A. Zingg, F. Diederich,
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Kazunobu Toshima,* Ryusuke Takano, Yutaka Maeda,
Masataka Suzuki, Akira Asai, and Shuichi Matsumura
The development of photochemical DNA-cleaving agents,
which selectively cleave DNA by irradiation with light with a
specific wavelength under mild conditions and without any
additives such as metals and reducing agents, is very interest-
[*] Prof. Dr. K. Toshima, R. Takano, Y. Maeda, M. Suzuki,
Prof. Dr. S. Matsumura
Â
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Soc. 1998, 120, 10990; F. Cardullo, F. Diederich, E. Echegoyen, T.
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Langmuir 1998, 14, 1955; P. Bhyrappa, G. Vaijayanthimala, K. S.
Suslick, J. Am. Chem. Soc. 1999, 121, 262; M. Plevoets, F. Vögtle, L.
De Cola, V. Balzani, New J. Chem. 1999, 63; M. Enomoto, T. Aida, J.
Am. Chem. Soc. 1999, 121, 874.
Department of Applied Chemistry, Faculty of Science
and Technology
Keio University, 3-14-1 Hiyoshi
Kohoku-ku, Yokohama 223-8522 (Japan)
Fax : (‡ 81)45-563-0446
E-mail: toshima@applc.keio.ac.jp
[6] J.-F. Nierengarten, D. Felder, J.-F. Nicoud, Tetrahedron Lett. 1999, 40,
273.
Dr. A. Asai
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd.
3-6-6 Asahi-machi, Machida-shi, Tokyo 194-8533 (Japan)
[7] a) J.-F. Nierengarten, T. Habicher, R. Kessinger, F. Cardullo, F.
Diederich, V. Gramlich, J.-P. Gisselbrecht, C. Boudon, M. Gross, Helv.
Chim. Acta 1997, 80, 2238; b) J.-F. Nierengarten, C. Schall, J.-F.
Nicoud, Angew. Chem. 1998, 110, 2037; Angew. Chem. Int. Ed. 1998,
37, 1934; c) R. Kessinger, M. Gomez-Lopez, C. Boudon, J.-P.
[**] This research was partially supported by a Grant-in-Aid for Encour-
agement of Young Scientists from the Ministry of Education, Science,
Sports and Culture (Japan).
Angew. Chem. Int. Ed. 1999, 38, No. 24
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ing from both chemical and biological standpoints and offers
considerable potential in medicine.[1] Here, we discuss the
molecular design, chemical synthesis, DNA-photocleaving
properties, and cytotoxicity of two novel and artificial light-
activatable DNA-cleaving agents, namely, the 2-phenylquino-
line ± carbohydrate hybrids 1 and 2.
bromosuccinimide (NBS)[12] in MeCN gave a mixture of the a-
glycoside 5 and the b-glycoside 6 in 79% yield in a ratio of
1.4:1. The azide groups in 5 and 6 were next converted into
N,N-dimethylamino groups by treatment with 35% HCHO
(aq) and a catalytic amount of Pd/C in MeOH under a
hydrogen atmosphere to afford 7 and 8 in 81% yield. After
their separation by column chromatography, 7 (1.0 equiv) and
8 (1.0 equiv) were esterified with 2-phenylquinoline-4-car-
boxylic acid (9, 1.5 equiv) by using N,N-dicyclohexylcarbodi-
imide (DCC) and 4-dimethylaminopyridine (4-DMAP) in
CH2Cl2 to give the hybrids 10 and 11 in 93 and 90% yields,
respectively. Finally, removal of the silyl groups in 10 and 11
with HF/pyridine (Py) furnished the desired 2-phenylquino-
line ± carbohydrate hybrids 1 and 2, respectively, in high
yields.
In our approach to create novel DNA-cleaving molecules,
we designed artificial intercalator± carbohydrate hybrid sys-
tems,[2±4] because many clinically useful antitumor antibiotics
such as anthracyclines[5] and aureolic acids,[6] which interact
with DNA, were commonly found to contain both aromatic
and carbohydrate domains. Since Denny et al.[7] demonstrated
the efficacy of 2‡1 unfused tricyclic aromatic systems such as
phenylquinolines as ªminimal intercalatorsº, 2-phenylquino-
line[8, 9] was selected as the DNA intercalator. The conjugated
The photoinduced DNA-cleaving activities of the hybrids 1
and 2 along with the components of these hybrids, 12 ± 15,[11]
were assayed with supercoiled FX174 DNA. As apparent
ˆ
C N bond in the 2-phenylquinoline unit was also expected to
generate the photoexcited 3(n !p*) state upon photoirradia-
tion, which may have a radical character and could be capable
of cleaving DNA. On the other hand, certain 2,6-dideoxy
amino sugars seemed appropriate as the carbohydrate source,
since they are DNA groove binders in some DNA-binding
antitumor antibiotics[10] and our previously reported artificial
DNA-interactive intercalator± carbohydate molecules.[4]
Therefore, we designed novel, artificial intercalator± carbo-
hydrate hybrids that consist of 2-phenylquinoline and a 2,6-
dideoxy amino sugar, which are connected by an ethylene
glycol linker.
from Figure 1, the 2-phenylquinoline ± carbohydrate hybrids 1
(500mm) and 2 (500mm) caused significant cleavage of DNA,
leading to small fragments upon photoirradiation with long-
wavelength UV light (365 nm), while 12 ± 15 did not show
The designed 2-phenylquinoline ± carbohydrate hybrids 1
and 2 were synthesized by a short reaction sequence
(Scheme 1). Thus, the glycosidation of the phenylthio sugar
3[11] (1.0 equiv) with ethylene glycol (4, 5.0 equiv) using N-
Figure 1. Photocleavage of supercoiled FX174DNA.
FX174DNA (50mm per base pair) was incubated with
various compounds in 20% acetonitrile in Tris ± HCl buffer
(Tris ˆ tris(hydroxymethyl)aminomethane, pH 7.5, 50mm) at
258C for 1 h under UV irradiation (365 nm, 15 W) from a
lamp placed 10 cm from the mixture, and analyzed by gel
electrophoresis (0.9% agarose gel, ethidium bromide stain).
Lane 1: DNA alone; lane 2: DNA with irradiation; lanes 3 ±
8: compounds 1, 2, 12, 13, 14, and 15 (500mm), respectively.
Form I ˆ covalently closed supercoiled DNA, Form IIˆ
open circular DNA, and Form III ˆ linear DNA.
DNA-cleaving activity under the same conditions.
These results clearly indicate the importance of
the hybrid structure for DNA cleaving. These
results also strongly suggest that the 2,6-dideoxy
amino sugar works as the DNA groove binder and
significantly enhances the intercalating ability of
the 2-phenylquinoline. In the absence of light no
DNA cleavage by 1 and 2 was observed. Further-
Scheme 1. Synthesis of 1 and 2. a) NBS (1.5 equiv with respect to 3), 4-Š molecular
sieves, MeCN, 08C, 1 h, 79%, 5:6 ˆ 1.4:1; b) H2, cat. Pd/C, 35% HCHO (aq). MeOH,
258C, 18 h, 81%; c) 9, DCC (1.0 equiv), cat. 4-DMAP, CH2Cl2, 08C, 4.5 h, 93% for 10,
90% for 11; d) HF/Py, Py, 258C, 15 h, 99% for 1, 94% for 2. TBS ˆ tert-butyldimethylsilyl.
more, the DNA-cleaving ability of the b-anomer
hybrid 2 was found to be stronger than that of the
a-anomer hybrid 1 (Figure 2). This result demon-
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activities of 1 and 2 with photoirradiation were much higher
than those without, and correlated with their capacity to
cleave DNA. Furthermore, we found that when the HeLaS3
cells were exposed to 1mm of hybrid 1 or 2 without photo-
irradiation, practically all of the cells survived, while similar
treatment combined with photoirradiation wiped out the cells.
These results clearly show that the DNA-cleaving activity
induced by photoirradiation significantly affects the cytotox-
icity of the hybrids, and the life of the cancer cells can be
controlled by treatment with an appropriate amount of the
2-phenylquinoline ± carbohydrate hybrid with or without the
photoirradiation.
Figure 2. Photocleavage of supercoiled FX174DNA. FX174DNA (50mm
per base pair) was incubated with the hybrid in 20% acetonitrile in Tris ±
HCl buffer (pH 7.5, 50mm) at 258C for 1 h under UV irradiation (365 nm,
15 W) from a lamp placed 10 cm from the mixture, and analyzed by gel
electrophoresis (0.9% agarose gel, ethidium bromide stain). a) Lane 1:
DNA alone; lane 2: DNA with irradiation; lane 3: DNA ‡ 1 without
irradiation; lanes 4 ± 9: 1 (1000), 1 (500), 1 (300), 1 (100), 1 (30), and 1
(10mm), respectively. b) Lane 1: DNA alone; lane 2: DNA with irradiation;
lane 3: DNA ‡ 2 without irradiation; lanes 4 ± 9: 2 (1000), 2 (500), 2 (300), 2
(100), 2 (30), and 2 (10mm), respectively.
Received: May 10, 1999
Revised version: June 24, 1999 [Z13401IE]
German version: Angew. Chem. 1999, 111, 3953 ± 3955
Keywords: antitumor agents ´ carbohydrates ´ cytotoxicity ´
DNA cleavage
[1] B. Armitage, Chem. Rev. 1998, 98, 1171 ± 1200.
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on the configuration of the sugar moiety in the hybrid. Since
the DNA-cleaving activity of 1 and 2 significantly decreased in
the presence of a radical scavenger, dimethyl sulfoxide, the
DNA cleavage must arise from the photoexcited 2-phenyl-
quinoline radical. The DNA-cleaving site specificity of the
hybrids 1 and 2 was also analyzed according to the Sanger
protocol.[13] The results in Figure 3 clearly show the identical
high guanine selectivity.
The cytotoxicity of the DNA-cleaving hybrids 1 and 2 was
next examined using HeLaS3 cells exposed to each agent for
72 h with or without 1 h of photoirradiation.[14] The IC50 values
of 1 and 2 without photoirradiation were 12 and 18mm,
respectively, and those with photoirradiation were 0.44 and
0.25mm, respectively. These results indicate that the cytotoxic
[2] a) D. J. Mincher, G. Shaw, E. D. Clercq, J. Chem. Soc. Perkin Trans. 1
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[11] The synthesis of the compound will be reported in detail elsewhere.
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[13] F. Sanger, S. Nicklen, A. R. Coulsen, Proc. Natl. Acad. Sci. U.S.A.
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48, 4827 ± 4833.
Figure 3. Autoradiogram of slab gel electropho-
resis with 12% polyacrylamide/8m urea for
sequence analysis. The 5'-end-labeled M13mp18
DNA was cleaved by the hybrids at pH 7.5 and
258C for 1 h under UV irradiation (365 nm,
15 W) from
a lamp placed 10 cm from the
mixture (bases 49 ± 105 are shown). Lanes A, G,
C, and T: Sanger A, G, C, and T reactions,
respectively; lanes 1 and 2: 1 and 2 (1000mm),
respectively.
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