Glycidol-carbohydrate hybrids: A new family of DNA alkylating agents

Article abstract of DOI:10.1016/S0960-894X(03)00659-0

Novel and chiral glycidol-carbohydrate hybrids possessing an epoxy group as a DNA alkylating moiety were designed and synthesized. These artificial hybrids selectively alkylated DNA at the N-7 sites of the guanines and cleaved DNA without any additives. The binding ability of the glycidol was significantly enhanced by the attachment of the carbohydrate.

Full text of DOI:10.1016/S0960-894X(03)00659-0

Bioorganic & Medicinal Chemistry Letters 13 (2003) 3281–3283
Glycidol-Carbohydrate Hybrids: A New Family of
DNA Alkylating Agents
Kazunobu Toshima,* Yukiko Okuno and Shuichi Matsumura
Department of Applied Chemistry, Faculty of Science and Technology, Keio University,
3-14-1 Hiyoshi, Kohoku-ku, Yokohama 223-8522, Japan
Received 21 May 2003;revised 19 June 2003;accepted 19 June 2003
Abstract—Novel and chiral glycidol–carbohydrate hybrids possessing an epoxy group as a DNA alkylating moiety were designed
and synthesized. These artificial hybrids selectively alkylated DNA at the N-7 sites of the guanines and cleaved DNA without any
additives. The binding ability of the glycidol was significantly enhanced by the attachment of the carbohydrate.
# 2003 Elsevier Ltd. All rights reserved.
Studies of the interaction between small molecules and
DNA, especially the effects of the structural character-
istics of the small molecules on the DNA interaction,
are very important in the design of new DNA targeting
drugs.1 A number of DNA interactive small molecules
including DNA oligomers and polyamides have been
already evaluated. In contrast, although carbohydrates
play a key role in various biological processes, the
interaction between carbohydrates and DNA, however,
are still not yet well understood. Only limited studies on
the interaction between certain carbohydrates in DNA
interactive antibiotics and DNA have been reported.2 In
our previous studies on the design of novel and artificial
DNA interactive small molecules possessing carbohy-
drates, it has been found that a suitably deoxygenated
amino sugar showed a high binding affinity to DNA,
and significantly enhanced the intercalating ability of
certain DNA intercalators.3 Furthermore, we have also
demonstrated that the carbohydrate-containing DNA
interactive small molecules showed cell-permeable
properties.3aꢀc DNA alkylating agents as well as DNA
intercalators have attracted considerable attention as
DNA interactive molecules.4 Although many molecules
causing alkylation of DNA have been evaluated, it is
still required to create new molecules having a more
potent biological activity. Therefore, based on our pre-
vious results,3 we had a simple question whether the
deoxygenated amino sugar, that appeared in our pre-
viously reported DNA interactive molecules, enhances
the DNA binding ability of not only DNA intercalators
but also DNA alkylating agents. In this paper, we report
the molecular design, chemical synthesis, DNA alkyl-
ating and cleaving properties of novel and artificial DNA
alkylating agents, that include the glycidol–carbohydrate
hybrids possessing an epoxy function as a DNA alkyl-
ating site (Fig. 1). To the best of our knowledge, this is
first example of DNA alkylating agents constructed from
only an alkylating group and carbohydrate.
In this preliminary study, we noted chiral glycidols 1
and 2 as the DNA alkylating moiety, because the chiral
epoxy structure was found in several DNA alkylating
antitumor antibiotics such as the pluramycins5 and also
in DNA interactive carcinogens such as the aflatoxin B1
oxide.6 Some artificial DNA alkylating agents consist-
ing of a DNA intercalator and epoxide have also been
reported.7 To examine the effect of the deoxygenated
amino sugar on the DNA binding ability of the DNA
alkylating agents, we newly designed the chiral glycidol–
carbohydrate hybrids 3–6 (Fig. 1). Glycidols 1 and 2 are
enantiomers of each other. Accordingly, the glycidol–
carbohydrate hybrids 3 and 4 are the epimers of 5 and 6,
respectively, with respective to the epoxide structure. On
the other hand, the a-glycosides 3 and 5 are the anomers
of the b-glycosides 4 and 6, respectively. The syntheses
of the glycidol–carbohydrate hybrids 3 and 4 are sum-
marized in Scheme 1. The syntheses of 3 and 4 com-
menced with the t-butyldiphenylsilyl (TPS) protected 88
which was easily prepared from the commercially avail-
able chiral alcohol 7. The primary hydroxy group in 8
was selectively protected with a pivaloyl (Pv) group and
the remaining secondary hydroxy group was then
*Corresponding author. Tel.: +81-45-566-1576;fax: +81-45-566-
1576;e-mail: toshima@applc.keio.ac.jp
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/S0960-894X(03)00659-0

3282
K. Toshima et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3281–3283
Figure 1.
Scheme 1. Syntheses of 3 and 4. Reagents and conditions: (a) ref 8;
(b) PvCl, Et3N, CH2Cl2, 35 ꢁC, 7 h, 96%;(c) TsCl, DMAP, Py, 60 ꢁC,
14 h, 72%;(d) HCl, MeOH, 40 ꢁC, 12 h, 72%;(e) TMSOTf, MS 4A,
CH2Cl2, 0 ꢁC, 0.5 h, 84% (13/14=4/1);(f) NaOMe, MeOH, 25 ꢁC, 5 h,
52% for 3, 59% for 4.
masked with a p-toluenesulfonyl (Ts) group by standard
procedures to afford 10 via 9. After the removal of the
TPS group under acidic conditions, the resultant alco-
hol 11 was subjected to glycosidation with the glycosyl
acetate 129 using TMSOTf as the activator to yield the
a- and b-glycosides 13 and 14. At this stage, these
anomers were easily separated by silica-gel column
chromatography. Finally, the treatment of the isolated
glycosides 13 and 14 with NaOMe caused epoxide for-
mation along with deprotection of the acetyl group to
furnish the desired glycidol-carbohydrate hybrids 3 and
4, respectively. The other hybrids 5 and 6 were also
synthesized from the enantiomer of 7 by very similar
protocols.
Figure 2. Cleavage of supercoiled ÈX174 DNA. ÈX174 DNA (50 mM
per base pair) was incubated with various compounds in 20% aceto-
nitrile in Tris–HCl buffer (pH 7.5, 50 mM) at 37 ꢁC for 24 h, and
analyzed by gel electrophoresis (0.9% agarose gel, ethidium bromide
stain): (a), (b), (c), (d), (e) and (f) for the compounds 1, 2, 3, 4, 5 and 6,
respectively: lane 1, DNA alone;lanes 2–7, compound (1000), com-
pound (300), compound (100), compound (30), compound (10) and
compound (3 mM), respectively. Form I: covalently closed supercoiled
DNA, Form II: open circular DNA.
With all the designed glycidol–carbohydrate hybrids 3–6
in hand, the DNA cleaving activities of these hybrids
along with the reference compounds 1 and 2 were next
assayed using supercoiled ÈX174 DNA.10 Based on the
results shown in Figure 2, the glycidol–carbohydrate
hybrids 3–6 caused significant DNA cleavage without
any additives ((c)–f) in Fig. 2). In contrast, glycidols 1
and 2 showed little DNA cleaving activity under similar
conditions ((a) and b) in Fig. 2). These results indicate
that the suitably deoxygenated amino sugar in the
hybrids significantly enhanced the DNA alkylating
ability of the glycidols. Furthermore, these hybrids
showed different DNA cleaving abilities. Thus, the
DNA cleaving ability of the hybrid 6 was found to be
stronger than those of the others, and the strongest 6
cleaved DNA in concentrations over 30 mM ((f) in Fig.
2). From these results, it was found that the DNA
cleaving activity was dependent on the configurations of
both the glycosidic bond in the sugar moiety and the
epoxide structure in the aglycon part in the hybrids.
The DNA cleaving site specificity of the glycidol deri-
vatives 1–6 was next analyzed according to the Sanger
protocol.11 Since the Sanger sequencing reactions result
in base incorporation, cleavage at nucleotide
(sequencing) represents a cleaving site by the agent or
N

K. Toshima et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3281–3283
3283
Acknowledgements
This research was partially supported by Grant-in-Aid for
the 21st Century COE program ‘KEIO Life Conjugate
Chemistry’ and for General Scientific Research from the
Ministry of Education, Culture, Sports, Science, and
Technology, Japan, and Takeda Science Foundation.
References and Notes
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Figure 3. Autoradiogram of 12% polyacrylamide-8 M urea slab gel
electrophoresis for sequence analysis. The 50-end-labeled M13mp18
DNA and 7-deazaguanosine-substituted M13mp18 DNA were treated
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the Maxam-Gilbert reaction at N+1.12 The results
shown in Figure 3 clearly indicated that all the glycidol
derivatives selectively cleaved DNA at the guanine sites,
and the site-selective DNA cleavage was significantly
enhanced upon treatment with hot piperidine13 (lanes
1–6 in Fig. 3). Furthermore, we confirmed that when
7-deazaguanosin-substituted DNA was used in this
assay, no DNA cleavage was observed even with hot
piperidine treatment (lanes 7–12 in Fig. 3). These
results strongly suggested that these hybrids selectively
alkylated DNA at the N-7 sites of the guanines.14
Moreover, it was observed that the intensities of the
glycidol-carbohydrate hybrids induced DNA cleaving
bands at the N-7 sites of the guanines were not always
equal (lanes 1–6 in Fig. 3). These results showed that
the hybrids had a moderate sequence selectivity in
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ation processes. These results suggest the further pos-
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alkylating agents15 based on carbohydrates including
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In summary, the present work demonstrates not only
the molecular design and chemical synthesis of novel
glycidol–carbohydrate hybrids, but also their DNA
alkylating and cleaving profiles. The effect of the car-
bohydrate on the DNA binding ability of the DNA
alkylating agent was also demonstrated. The described
chemistry and biological evaluation provided sig-
nificant information about the molecular design of
novel and selective DNA alkylating agents possessing
carbohydrate(s).
15. (a) For some leading studies Oyoshi, T.;Kawakami, W.;
Narita, A.;Bando, T.;Sugiyama, H. J. Am. Chem. Soc. 2003,
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