Platinum(ii)-triarylpyridines complexes with electropositive pendants as efficient G-quadruplex binders

Article abstract of DOI:10.1039/c0dt01161d

Herein we reported three new platinum(ii)-triarylpyridines complexes with peralkylated ammonium pendants that strongly stabilize G-quadruplex DNA.

Full text of DOI:10.1039/c0dt01161d

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Platinum(II)-triarylpyridines complexes with electropositive pendants as
efficient G-quadruplex binders†
Jin-Tao Wang,a Yi Li,a Jia-Heng Tan,b Liang-Nian Jia and Zong-Wan Mao*a,c
Received 3rd September 2010, Accepted 10th November 2010
DOI: 10.1039/c0dt01161d
Herein we reported three new platinum(II)-triarylpyridines
complexes with peralkylated ammonium pendants that
strongly stabilize G-quadruplex DNA.
surfaces that were more compatible with G-quartet motif.6d The
same case also took place in the Pt(II) complexes reported by us.6i
Recently, a triarylpyridine ligand family has been found to have
a general capacity to interact with G-quadruplex DNA sequences.7
Their elementary unit is composed of four aromatic rings, which
form a large planar p-conjugated surface. To become more elec-
trophilic and extend planarity to the ligands, herein, we add differ-
ent peralkylated ammonium pendants to the triarylpyridine ligand
and synthesize their platinum(II) complexes, [Pt(L1)Cl](PF6)2·H2O
(1), [Pt(L2)Cl](PF6)2·H2O (2), and [Pt(L3)Cl] (PF6)2·2H2O
(3), where L1 = 4¢-(4-(trimethylamino)methylphenyl)-2,2¢:6¢,2¢¢-
terpyridine, L2 = 4¢-(4-(triethylammonio)methylphenyl)-2,2¢:6¢,2¢¢-
terpyridine, and L3 = 4¢-(4-(tributylammonio)methylphenyl)-
2,2¢:6¢,2¢¢-terpyridine (see Scheme 1).
G-quadruplex DNA sequences, formed by biologically significant
four-stranded nucleic acid, have received considerable attention in
several research fields including chemistry, biology, and pharma-
cology in the last decade.1 Such DNA sequences have an unusual
planar structure with four guanines in a circle through Hoogsteen
hydrogen bonds.2 In human telomeric DNA, the G-quadruplex
DNA has just a short sequence 5¢-d(TTAGGG)-3 and is tandemly
repeated in rich guanine residues domains. The formation of
G-quadruplex complexes can prevent telomerase elongation of
telomeres by interrupting the interaction between the enzyme
and unfolded guanine-rich single strand.3 Therefore, the use of
small molecules that promote the formation and/or stabilize G-
quadruplex structures has become attractive for anticancer drug
design.4
Many reports have shown that an effective G-quadruplex-
stabilizing molecule exhibits unique characteristics, including a
large electron-deficient p-aromatic surface that can stack on the
quadruplex, a positively charged area that can reside closely
to the center of the guanine quartet, or positively charged
substituents that can interact with both the grooves and loops of
the quadruplex and negatively charged phosphates backbone.5 In
order to optimize the above-mentioned features, metal complexes
are usually employed. For example, a platinum(II) ion has positive
divalent charges and usually forms a planar square complex with
an appropriate ligand. These features make them desirable for G-
Quadruplex stabilization. However, only a few Pt(II) complexes
have been applied in this field.6 Sleiman and co-workers found
Pt(II) complexes with extended aromatic ligands to provide p-
Scheme 1 Synthesis of the ligands L1–L3 and Pt(II) complexes 1–3.
These complexes were obtained through a synthetic route
summarized in Scheme 1. 4¢-(4-Bromomethylphenyl)-2,2¢:6¢,2¢¢-
terpyridine, as a reactant,8 directly reacts with trimethylamine,
triethylamine, and tributylamine to give L1, L2, and L3, respec-
tively. The reaction of the ligands with Pt(DMSO)2Cl2 gave the
final products 1, 2 and 3 with suitable yields. Detailed experimental
procedures for the preparation of ligands and complexes can be
found in the ESI.†
During the investigation of stabilizing G-quadruplexes, struc-
tural conversion between various kinds of the human telomeric
quadruplexes, including intra- and intermolecularly in parallel,
and mixed arrangements, depending on the strand orientation
were initially observed.9 In our current studies, a human telomeric
DNA sequence (22AG, 5¢-AG3(T2AG3)3-3¢) is applied to study
the stabilization of G-quadruplex by CD titration with ligands
and their Pt(II) complexes that are monitored by CD assays.
Obtained results are compared in Fig. 1 and S1.† Evidenced by
aMOE Key Laboratory of Bioinorganic and Synthetic Chemistry, School of
Chemistry and Chemical Engineering, Sun Yat-Sen University, Guangzhou,
510275, China. E-mail: cesmzw@mail.sysu.edu.cn; Fax: +86 20 84112245;
Tel: +86 20 84113788
bSchool of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou,
510080, China
cState Key Laboratory of Natural and Biomimetic Drug/Department of
Chemical Biology, School of Pharmaceutical Science, Peking University,
Beijing, 100191, China
† Electronic supplementary information (ESI) available: Synthesis of the
ligands L1–L3 and complexes 1–3, ESI-MS spectra, 1H NMR spectra,
CD titration, FRET melting experiments, PCR stop assay and Molecular
Modelling. See DOI: 10.1039/c0dt01161d
564 | Dalton Trans., 2011, 40, 564–566
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Table 1 Stabilization temperatures (DTm) determined by FRET
experiment
DTm/◦C at 1 mM compound concentrationa
Compounds
F21T
F10T
L1
L2
L3
1
21.0
23.2
20.0
26.2
30.5
25.3
2.0
2.0
1.9
5.6
6.2
3.8
2
3
a DTm = Tm (DNA + compound) - Tm (DNA). The concentrations of
F21T and F10T were both 0.2 mM.
(ICD) signals in the region of 310–360 nm (Fig. 1a and 1c) and a
minor positive peak at near 345 nm (Fig. 1b) were observed. These
ICD signals indicated that besides the end-stacking mode, other
binding modes (groove binding, loop binding, or phosphates back-
bone binding) are also possible. Considering the multiple binding
modes, the sizes of the peralkylated ammonium groups may affect
the binding abilities of these compounds. So the potential reason
that the complexes 1 and 2 show stronger capability than 3 is that
steric hindrance from the largest group tributylamine inhibits the
interaction between 3 and G-quadruplexes.
Fig. 1 CD titration spectra of 22AG sequence (3 mM) at increasing
concentrations of 2 and L2 in 10 mM Tris-HCl buffer, pH 7.4, 100 mM
KCl and no metal cations, rt. Arrows indicate the increasing amounts of
complexes (r = compound/DNA strand concentration). (a) 2: r of 0.2, 0.5,
1.0, 1.2, 1.4, 1.8 and 2.0 in 100 mM KCl; (b) L2: r of 0.5, 1.0, 2.0, 4.0, 6.0
and 8.0 in the absence of KCl; (c) 2: L2: r of 0.2, 0.5, 0.8, 1.0 and 1.2 in
100 mM KCl; (d) L2: r of 0.5, 1.0, 2.0, 4.0 and 6.0 in the absence of KCl.
The binding of all the ligands and complexes to G-quadruplex
DNA F21T (sequence: 5¢-FAM-G3[T2AG3]3-TAMRA-3¢, mimic-
ing the human telomeric repeat) and
a hairpin duplex
DNA F10T (5¢-FAM-dTATAGCTATA-HEG-TATAGCTATA-
TAMRA-3¢) were also investigated by a FRET (Fluorescence
Resonance Energy Transfer) melting assay.11 Table 1 provides
the effect of the ligands and complexes on the enhanced melt-
ing temperature (DTm) of two labelled oligonucleotides in K+
solution.
the CD spectra in Fig. 1a, in the presence of potassium cations, the
quadruplex molecules exist as a mixture of parallel and antiparallel
G-quadruplex conformations at the very beginning. When 2 is
added at 0–6.0 mM, a positive band at about 290 nm and a
negative band at about 260 nm are observed and reach saturation.
These changes suggest that 2 strongly stabilizes antiparallel
G-quadruplex conformation.10 In the absence of any salt, the
quadruplex molecules not only exist as a mixture of parallel and
antiparallel G-quadruplex conformations, but also are dissociated
partially to single-stranded molecules.9 Therefore, larger band
changes are observed in the CD spectrum with the addition of
2 (0–3.6 mM) to 22AG sequence. As illustrated in Fig. 1c, the
maximum at 254 nm is gradually suppressed and shifted to
245 nm, while the bands centred at about 293 nm and 263 nm
increasing sharply along with the addictions of 2. Finally, the
induced CD spectra of 2 virtually resemble those of antiparallel
G-quadruplexes. Relative to 2, smaller band changes induced
by L2 and require higher compound concentrations, 24.0 and
18.0 mM, to reach saturation, are observed in both the presence
and absence of K+ ions. The results indicate that the complex
2 is more efficient at inducing the formation of antiparallel G-
quadruplexes than its ligand L2. The same results are also obtained
during the investigation on 1 and 3 and their ligands (Fig. S1).
As expected, complexes containing larger square p-aromatic
surfaces with Pt(II) present more effective interaction with G-
quadruplex DNA sequences. In principle, the positively charged
substituents provide potential interactions with the negatively
charged sugar–phosphate backbone, the grooves and loops of the
quadruplex.5,7a,10c Two hints concerning such interactions also can
be found from the CD spectra. Raised induced circular dichroism
All of the ligands and complexes have high DTm values. At
1.0 mM of them, the DTm values are in the range of 20–30 ◦C.
On the other hand, none of these compounds are observed to
increase the melting temperature of F10T significantly, suggesting
their poor binding to the duplex DNA. Likewise, The FRET-
melting data demonstrate that the complexes are more capable
than the ligands, and all the three complexes are better G-
quadruplexes binders than another Pt(II) complex (11.3 ◦C at
1.0 mM, FRET) coordinated by a similar ligand but lacking of
any electropositive pendants, Pt-ttpy (ttpy = p-tolyl-terpyridine).6b
This reveals that the introduced electropositive pendants are
beneficial to G-quadruplex stabilization by these Pt(II) complexes.
Besides, a consistent result from this experiment is that L2 and
2 show little higher DTm values than other two ligands and
complexes. This may implied that moderate triethylamine group
can better favor the binding to G-quadruplex.
In order to further evaluate how well the ligands and the
complexes stabilize G-quadruplex DNA, a polymerase chain
reaction (PCR)-stop assay was used to ascertain whether these
compounds are bound to a test oligomer (5¢-G3[T2AG3]3-3¢) and
therefore stabilize the G-quadruplex structure.12 Fig. 2 illustrates
that in the presence of the complexes, the template sequence
forms G-quadruplex structures undetectable to the PCR product.
The inhibitory effect of these complexes are enhanced clearly as
the concentration increases from 1.0 to 8.0 mM with even no
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20821001), Guangdong Provincial Natural Science Foundation
(No. 9351027501000003), National Basic Research Program of
China (973 Program No. 2007CB815306), and Fundamental
Research Funds for the Central Universities.
Notes and references
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Fig. 2 Effect of complexes 1, 2 and 3 (0–8.0 mM) on the hybridization of
HTG21 in a PCR-stop assay.
PCR product detected at 8.0 mM. But the three ligands hardly
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with positively charged tetraalkylammonium groups are efficient
G-quadruplex binders.
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Fig. 3 Predicted interaction between 2 and the intermolecular G-quadru-
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In this work, our purpose was to design and evaluate a
series of platinum(II)-triarylpyridines complexes with tetraalky-
lammonium pendants that would show effective binding to G-
quadruplex. All the experimental results clearly show that these
ligands and complexes are capable of inducing the stabilization of
G-quadruplexes, while the complexes are more efficient than their
free ligands. Furthermore, complex 2 exhibits highest stabilization
potential for G-quadruplex due to its appropriate triethylamine
pendant. Our experiments also suggest that Pt(II) complexes could
provide planar p-surfaces that promise them with G-quadruplex
binding potentials, therefore the introduction of proper positively
charged pendants is a practical approach for G-quadruplex binder
design.
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This work was supported by the National Natural Science
Foundation of China (Nos. 30770494, 20725103, 20831006, and
566 | Dalton Trans., 2011, 40, 564–566
This journal is
The Royal Society of Chemistry 2011
©