Importance of platinum(II)-assisted platinum(IV) substitution for the oxidation of guanosine derivatives by platinum(IV) complexes

Article abstract of DOI:10.1021/ic701868b

Guanosine derivatives with a nucleophilic group at the 5′ position (G-5′) are oxidized by the Pt^IV complex Pt(d,l)(1,2-(NH ₂)₂C₆H₁₀)Cl₄ ([Pt ^IV(dach)Cl₄]). The overall redox reaction is autocatalytic, consisting of the Pt^II-catalyzed Pt^IV substitution and two-electron transfer between Pt^IV and the bound G-5′. In this paper, we extend the study to improve understanding of the redox reaction, particularly the substitution step. The [Pt^II(NH ₃)₂(CBDCA-O,O′)] (CBDCA = cyclobutane-1,1- dicarboxylate) complex effectively accelerates the reactions of [Pt ^IV(dach)Cl₄] with 5′-dGMP and with cGMP, indicating that the Pt^II complex does not need to be a Pt^IV analogue to accelerate the substitution. Liquid chromatography/mass spectroscopy (LC/MS) analysis showed that the [Pt^IV(dach)Cl₄]/[Pt ^II(NH₃)₂(CBDCA-O,O′)]/cGMP reaction mixture contained two Pt^IVcGMP adducts, [Pt^IV(NH ₃)₂(cGMP)(CI)(CBDCA-O,O′)] and [Pt ^IV(dach)(cGMP)Cl₃]. The LC/MS studies also indicated that the trans,cis-[Pt^IV(dach)(³⁷Cl)₂( ³⁵Cl)₂]/[Pt^II(en)(³⁵Cl) ₂]/9-EtG mixture contained two Pt^IV-9-EtG adducts, [Pt^IV(en)(9-EtG)(³⁷Cl)(³⁵Cl)₂] and [Pt^IV(dach)(g-EtG)(³⁷Cl)₂]. These Pt ^IVG products are predicted by the Basolo-Pearson (BP) Pt ^II-catalyzed Pt^IV-substitution scheme. The substitution can be envisioned as an oxidative addition reaction of the planar Pt ^II complex where the entering ligand G and the chloro ligand from the axial position of the Pt^IV complex are added to Pt^II in the axial positions. From the point of view of reactant Pt^IV, an axial chloro ligand is thought to be substituted by the entering ligand G. The Pt^IV complexes without halo axial ligands such as trans,cis-[Pt(en)(OH)₂Cl₂], trans,cis-[Pt(en)(OCOCF ₃)₂Cl₂], and cis,trans,cis-[Pt(NH ₃)(C₆H₁₁NH₂)(OCOCH₃) ₂Cl₂] ([Pt^IV(a,cha)(OCOCH₃) ₂Cl₂], satraplatin) did not react with 5′-dGMP. The bromo complex, [Pt^IV(en)Br₄], showed a significantly faster substitution rate than the chloro complexes, [Pt^IV(en)Cl ₄] and [Pt^IV(dach)Cl₄]. The results indicate that the axial halo ligands are essential for substitution and the Pt ^IV complexes with larger axial halo ligands have faster rates. When the Pt^IV complexes with different carrier ligands were compared, the substitution rates increased in the order [Pt^IV(dach)Cl₄] < [Pt^IV(en)Cl₄] < [Pt^IV(NH ₃)₂Cl₄], which is in reverse order to the carrier ligand size. These axial and carrier ligand effects on the substitution rates are consistent with the BP mechanism. Larger axial halo ligands can form a better bridging ligand, which facilitates the electron-transfer process from the Pt^II to Pt^IV center. Smaller carrier ligands exert less steric hindrance for the bridge formation.

Full text of DOI:10.1021/ic701868b

Inorg. Chem. 2008, 47, 1352-1360
Importance of Platinum(II)-Assisted Platinum(IV) Substitution for the
Oxidation of Guanosine Derivatives by Platinum(IV) Complexes
Sunhee Choi,*,† Livia Vastag,† Yuri C. Larrabee,† Michelle L. Personick,† Kurt B. Schaberg,†
Benjamin J. Fowler,† Roger K. Sandwick,† and Gulnar Rawji‡
Department of Chemistry and Biochemistry, Middlebury College, Middlebury, Vermont 05753, and
Department of Chemistry, Southwestern UniVersity, Georgetown, Texas 78626
Received September 20, 2007
Guanosine derivatives with a nucleophilic group at the 5′ position (G-5′) are oxidized by the PtIV complex Pt(d,l)(1,2-
(NH2)2C6H10)Cl4 ([PtIV(dach)Cl4]). The overall redox reaction is autocatalytic, consisting of the PtII-catalyzed PtIV
substitution and two-electron transfer between PtIV and the bound G-5′. In this paper, we extend the study to
improve understanding of the redox reaction, particularly the substitution step. The [PtII(NH3)2(CBDCA-O,O′)] (CBDCA
) cyclobutane-1,1-dicarboxylate) complex effectively accelerates the reactions of [PtIV(dach)Cl4] with 5′-dGMP and
with cGMP, indicating that the PtII complex does not need to be a PtIV analogue to accelerate the substitution.
Liquid chromatography/mass spectroscopy (LC/MS) analysis showed that the [PtIV(dach)Cl4]/[PtII(NH3)2(CBDCA-
O,O′)]/cGMP reaction mixture contained two PtIVcGMP adducts, [PtIV(NH3)2(cGMP)(Cl)(CBDCA-O,O′)] and
[PtIV(dach)(cGMP)Cl3]. The LC/MS studies also indicated that the trans,cis-[PtIV(dach)(37Cl)2(35Cl)2]/[PtII(en)(35Cl)2]/
9-EtG mixture contained two PtIV-9-EtG adducts, [PtIV(en)(9-EtG)(37Cl)(35Cl)2] and [PtIV(dach)(9-EtG)(37Cl)(35Cl)2].
These PtIVG products are predicted by the Basolo-Pearson (BP) PtII-catalyzed PtIV-substitution scheme. The
substitution can be envisioned as an oxidative addition reaction of the planar PtII complex where the entering
ligand G and the chloro ligand from the axial position of the PtIV complex are added to PtII in the axial positions.
From the point of view of reactant PtIV, an axial chloro ligand is thought to be substituted by the entering ligand
G. The PtIV complexes without halo axial ligands such as trans,cis-[Pt(en)(OH)2Cl2], trans,cis-[Pt(en)(OCOCF3)2Cl2],
and cis,trans,cis-[Pt(NH3)(C6H11NH2)(OCOCH3)2Cl2] ([PtIV(a,cha)(OCOCH3)2Cl2], satraplatin) did not react with 5′-
dGMP. The bromo complex, [PtIV(en)Br4], showed a significantly faster substitution rate than the chloro complexes,
[PtIV(en)Cl4] and [PtIV(dach)Cl4]. The results indicate that the axial halo ligands are essential for substitution and the
PtIV complexes with larger axial halo ligands have faster rates. When the PtIV complexes with different carrier
ligands were compared, the substitution rates increased in the order [PtIV(dach)Cl4] < [PtIV(en)Cl4] < [PtIV(NH3)2Cl4],
which is in reverse order to the carrier ligand size. These axial and carrier ligand effects on the substitution rates
are consistent with the BP mechanism. Larger axial halo ligands can form a better bridging ligand, which facilitates
the electron-transfer process from the PtII to PtIV center. Smaller carrier ligands exert less steric hindrance for the
bridge formation.
Introduction
market, cisplatin, carboplatin, and oxaliplatin, all have an
oxidation state of 2+. Because of severe side effects and
other deficiencies, however, alternative platinum anticancer
drugs are under development. A wide variety PtIV complexes
have potential as powerful anticancer drugs, and they can
be administered orally.2 A prototypical member of this class,
cis,trans,cis-[Pt(NH3)(C6H11NH2)(OC(O)CH3)2Cl2] (satrapl-
Platinum coordination complexes are biologically impor-
tant for their anticancer activity, being effective on tumor
types including testicular, ovarian, and colon cancers.1 The
three platinum anticancer drugs currently available on the
* To whom correspondence should be addressed. E-mail: choi@
middlebury.edu.
(1) (a) Fuertes, M. A.; Alonso, C.; Perez, J. M. Chem. ReV. 2003, 103,
645–662. (b) Lippert, B. Cisplatin: Chemistry and Biochemistry of a
Leading Anticancer Drug; Wiley-VCH: Weinheim, Germany, 1999.
†
Middlebury College.
Southwestern University.
‡
1352 Inorganic Chemistry, Vol. 47, No. 4, 2008
10.1021/ic701868b CCC: $40.75
2008 American Chemical Society
Published on Web 01/26/2008

Platinum(II)-Assisted Platinum(IV) Substitution
atin), is currently undergoing phase III clinical trials. Because
PtIV complexes are kinetically inert,3 it is generally believed
they must be reduced to their PtII analogues in vivo to bind
to DNA and achieve anticancer activity.2 However, while
direct PtIV binding to DNA has been observed in vitro in
several laboratories,4,5 a detailed mechanism was not proposed.
Our laboratory reported that PtIV complexes have a wide
range of reactivity depending on their structures.6 For
example, [PtIV(dach)Cl4] was reduced by ascorbic acid
approximately 700 times faster than cis,trans,cis-[Pt((CH3)2-
CHNH2)2(OH)2Cl2], while trans,cis-[PtIV(en)(OH)2Cl2] resisted
reduction completely. The [PtIV(dach)Cl4] complex reacted
with 5′-GMP approximately 5 times faster than [PtIV(en)Cl4],
which, in turn, was 10-fold faster than trans,cis-[PtIV(en)-
(OCOCH3)2Cl2]. The trans,cis-[PtIV(en)(OH)2Cl2] complex did
not significantly react with 5′-GMP.7
The most reactive complex, [PtIV(dach)Cl4], oxidized 5′-
dGMP,8 3′-dGMP,9 and 5′-d[GTTTT]-3′.9 The redox mech-
anism consists of two steps: a substitution and an electron
transfer.8,9 In the substitution reaction, PtIV binds to the N7
of the guanosine (G) moiety. In the electron-transfer reaction,
the 5′-phosphate or 5′-hydroxyl group attacks C8 of the G
moiety and an inner-sphere, two-electron transfer produces
cyclic (5′-O-C8)-G and a PtII complex. The identity of the
final oxidized G depends on the hydrolysis rate of the cyclic
intermediate. The cyclic phosphodiester intermediate formed
from [PtIV(dach)Cl4]/5′-dGMP is hydrolyzed to 8-oxo-5′-
dGMP.8 However, the cyclic ether intermediate formed from
[PtIV(dach)Cl4]/3′-dGMP (or 5′-d[GTTTT]) does not hydro-
lyze, and this cyclic form is the final oxidation product.9
The lack of a 5′-nucleophilic group in the G moiety, such
as in 3′,5′-cyclic guanosine monophosphate (cGMP), 9-me-
thylxanthine (9-Mxan), 5′-d[TTGTT]-3′, and 5′-d[TTTTG]-
3′, prevents the electron-transfer step, and only the substi-
tution step is observed.9
The kinetics of the redox reactions between [PtIV(dach)Cl4]
and 5′- (and 3′-)dGMP is autocatalytic.10 Catalyzed by PtII
complexes, the reaction seems to follow the classic
Basolo-Pearson (BP) PtII-catalyzed PtIV substitution.10,11
However, several important questions remain unanswered
to confirm this mechanism. First, our previous study showed
that the PtII complexes do not have to be their PtIV analogues
to accelerate the reaction. Furthermore, all of the PtII
complexes studied previously contained at least one leaving
chloro ligand.10 Can PtII without a leaving halo group
accelerate the reaction? Second, the previous studies showed
that a chloro ligand in [PtIV(dach)Cl4] is substituted by G.8,10
Which chloro ligand, axial or equatorial, is substituted? Third,
in the previous studies we used only two PtIV complexes,
namely, [PtIV(dach)Cl4] and [PtIV(en)Cl4].10 Does the BP
mechanism apply to other PtIV complexes? What are the
effects of the PtIV axial and carrier ligands? These questions
are addressed by using several PtIV complexes with different
axial and carrier ligands (Chart 1).
Experimental Section
Sample Preparation. [PtIV(dach)Cl4] and [PtII(NH3)2(CBDCA-
O,O′)] were obtained from the National Cancer Institute, Drug
Synthesis and Chemistry Branch, Developmental Therapeutics
Program, Division of Cancer Treatment. Satraplatin was obtained
through a material transfer agreement with GPC Biotech AG.
Isotopically pure Na35Cl and Na37Cl were obtained from Oak Ridge
National Laboratory, Oak Ridge, TN. The complexes [PtII(en)Cl2],
[PtIV(en)Cl4], [PtIV(en)Br4], trans,cis-[PtIV(en)(OCOCF3)2Cl2],
trans,cis-[PtIV(en)(OH)2Cl2], cis-[PtIV(NH3)2Cl4], and trans-(R,R)-
[PtII(dach)I2] were synthesized following previously published
procedures.12
(2) (a) Hall, M. D.; Hambley, T. W. Coord. Chem. ReV. 2002, 232, 49–
67. (b) Eastman, A. Biochem. Pharmacol. 1987, 36, 4177–4178. (c)
Chaney, S. G. Int. J. Oncol. 1995, 6, 1291–1305. (d) Chaney, S. G.;
Gibbons, G. R.; Wyrick, S. D.; Podhasky, P. Cancer Res. 1991, 51,
969–973. (e) Kelland, L. R.; Murrer, B. A.; Abel, G.; Giandomenico,
C. M.; Mistry, P.; Harrap, K. R. Cancer Res. 1992, 52, 822–828. (f)
Oshita, F.; Eastman, A. Oncol. Res. 1993, 5, 111–118. (g) Yoshida,
M.; Khokhar, A. R.; Zhang, Y.-P.; Siddik, Z. H. Cancer Res. 1994,
54, 4691–4697.
trans-(R,R)-[PtII(dach)35(Cl)2] was prepared from the corre-
sponding diiodo complex according to a literature procedure with
minor modifications.12 trans-(R,R)-Diaminocyclohexanediiodoplat-
inum(II) (0.024 mmol) was suspended in 0.5 mL each of water
and acetone. Solid AgNO3 was added, and the resulting milky
suspension stirred at room temperature in the dark until no more
trans-(R,R)-[PtII(dach)I2] remained (∼2 h). Silver iodide was
removed by filtration through Celite packed in a pasteur pipet. A
10-fold excess of Na35Cl (0.24 mmol) was added to the filtrate and
the solution stirred at 50 °C for about 2 h. A pale-yellow solid that
precipitated was collected, washed with water and acetone, and dried
under vacuum. Yield: ∼78%. LC/MS: m/z 324.9.
(3) (a) Hartley, F. R. Chemistry of the Platinum Group Metals: Recent
DeVelopments; Elsevier: Amsterdam, 1991. (b) Basolo, F.; Pearson,
R. G. Mechanism of Inorganic Reactions; Wiley: New York, 1967.
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J. Biochem. 1995, 228, 616–624. (b) Talman, E. G.; Kidani, Y.;
Mohrmann, L.; Reedijk, J. Inorg. Chim. Acta 1998, 283, 251–255.
(c) Galanski, M.; Keppler, B. K. Inorg. Chim. Acta 2000, 300–302,
783–789.
trans-(R,R)-cis,trans-[PtIV(dach)(35Cl)2(37Cl)2] was prepared by
adapting literature procedures.12 Hydrogen peroxide (0.20 mmol)
was added to a suspension of trans-(R,R)-[PtII(dach)35Cl2] (0.016
mmol) in 0.5 mL of water. The mixture was stirred at 70 °C for
2 h, during which trans-(R,R)-[PtIV(dach)35Cl2(OH)2], a lighter-
(5) (a) Roat, R. M.; Reedijk, J. J. Inorg. Biochem. 1993, 52, 263–274.
(b) Talman, E. G.; Brüning, W.; Reedijk, J.; Spek, A. L.; Veldman,
N. Inorg. Chem. 1997, 36, 854–861. (c) van der Veer, J. L.; Peters,
A. R.; Reedijk, J. J. Inorg. Biochem. 1986, 26, 137–142. (d) Roat,
R. M.; Jerardi, M. J.; Kopay, C. B.; Heath, D. C.; Clark, J. A.; DeMars,
J. A.; Weaver, J. M.; Bezemer, E.; Reedijk, J. J. Chem. Soc., Dalton
Trans. 1997, 3615–3621.
(6) Choi, S.; Filotto, C.; Bisanzo, M.; Delaney, S.; Lagasee, D.; Whitworth,
J. L.; Jusko, A.; Li, C.; Wood, N. A.; Willingham, J.; Schwenker, A.;
Spaulding, K. Inorg. Chem. 1998, 37, 2500–2504.
(10) Choi, S.; Vastag, L.; Leung, C. H.; Beard, A. M.; Knowles, D. E.;
Larrabee, J. A. Inorg. Chem. 2006, 45, 10108–10114.
(11) (a) Basolo, F.; Wilks, P. H.; Pearson, R. G.; Wilkins, R. G. J. Inorg.
Nucl. Chem. 1958, 6, 161. (b) Mason, W. R. 1972, 7, 241–255. (c)
Summa, G. M.; Scott, B. A. Inorg. Chem. 1980, 19, 1079. (d) Cox,
L. T.; Collins, S. B.; Martin, D. S. J. Inorg. Nucl. Chem. 1961, 17,
383.
(7) Choi, S.; Mahalingaiah, S.; Delaney, S.; Neale, N. R.; Masood, S.
Inorg. Chem. 1999, 38, 1800–1805.
(8) Choi, S.; Cooley, R. B.; Hakemian, A. S.; Larrabee, Y. C.; Bunt, R. C.;
Maupaus, S. D.; Muller, J. G.; Burrows, C. J. J. Am. Chem. Soc. 2004,
126, 591–598.
(12) (a) Ellis, L. T.; Er, H. M.; Hambley, T. W. Aust. J. Chem. 1995, 48,
793–806. (b) Al-Baker, S.; Siddik, Z. H.; Khokhar, A. R. J. Coord.
Chem. 1994, 31, 109–116.
(9) Choi, S.; Cooley, R. B.; Voutchkova, A.; Leung, C. H.; Vastag, L.;
Knowles, D. E. J. Am. Chem. Soc. 2005, 127, 1773–1781.
Inorganic Chemistry, Vol. 47, No. 4, 2008 1353

Choi et al.
Chart 1. Structures of the Platinum Complexes and Guanine Derivatives Studied
yellow solid, was formed. The dihydroxo complex generated in
situ was converted to the tetrachloro complex by the addition of
excess Na37Cl (0.23 mmol) and HNO3 (0.04 mmol), followed by
stirring at 70 °C for an additional 2 h. The suspended solid changed
from a yellow to an orange, which redissolved upon the addition
of acetone (0.5 mL) and further stirring for another 30 min. The
resulting pale-yellow solution was filtered and the filtrate evaporated
to dryness, leaving a residue of excess Na37Cl and the desired
product as a yellow solid. The product complex was obtained by
the addition of acetone to the residue and removal of the undissolved
solid (Na37Cl) by filtration. Evaporation of acetone yielded the final
product as yellow crystals. Yield: ∼93%. LC/MS: m/z 454.9.
In every step of the synthesis, amber glassware or glassware
wrapped in aluminum foil was used to avoid photoreduction of the
platinum complexes. The products were characterized by IR (Bruker
Equinox 55 spectrometer), 1H NMR (Bruker 400 Ultrashield
spectrometer), high-performance liquid chromatography (HPLC;
Waters Alliance 2695 equipped with a Waters 2996 photodiode
array detector and a Waters Atlantis dC18 column), and liquid
chromatography/mass spectrometry (LC/MS). Stock platinum com-
plex solutions were prepared by dissolution in saline water (0.1 M
NaCl, pH 8.3–8.6). The pH was adjusted to 8.3–8.6 to fully
deprotonate the phosphate group in 5′-dGMP.10 To prevent hy-
drolysis of the platinum complexes, 0.1 M NaCl and 0.1 M KBr
solutions were used for the chloro and bromo complexes, respec-
tively.
mm), run under isocratic (0.5 mL min-1) conditions with a solvent
of 0.5% formic acid in water. An ion-trap mass spectrometer to
select for specific masses was used with fragmentation both on and
off. The MS parameters were set to scan from m/z 50 to 2200 using
positive or negative detection with electrospray ionization mass
spectrometry. For tuning, the nebulizer was set to 50 psi, the dry
gas flow to 9 L min-1, and the dry temperature to 365 °C.
Kinetic Studies. An appropriate amount of 5′-dGMP was added
to vials containing equal volumes of platinum stock and saline
solutions. The pH values of the solutions were adjusted to pH 8.3
with NaOH using a pH meter (Orion Research 960) equipped with
a Mettler-Toledo Inlab combination pH microelectrode. Attainment
of the desired pH constituted the beginning of the reaction. UV/
vis spectra were obtained in 10-mm cells on a Varian Cary 4000i
spectrophotometer with Cary WinUV kinetic assay software. The
absorbances at 360 and 390 nm were monitored. The sample
temperature was maintained by a Varian-Cary temperature control-
ler. Kinetic experiments were repeated at least three times.
Results
Synthesis of PtII and PtIV Complexes with Isotopically
Pure 35Cl and 37Cl. The key complexes [PtII(en)(35Cl)2],
trans-(R,R)-[PtII(dach)(35Cl)2], and trans-(R,R)-cis,trans-
[PtIV(dach)(35Cl)2(37Cl)2] were synthesized by modification
of published procedures12 and characterized by LC/MS.
In the reactions of the diiodo complexes with AgNO3, the
addition of acetone facilitated better mixing of the suspension
and reduced reaction times for complete precipitation of AgI,
particularly for the complex with dach as the diamine. The
Mass Spectrometry. LC/MS analyses were conducted on an
1100 series LC and LC/MSD Trap XCT Plus from Agilent
Technologies. The LC component was equipped with a photodiode
array detector and an Eclipse X-D8-C8 column (5 µm, 4.6 × 150
1354 Inorganic Chemistry, Vol. 47, No. 4, 2008

Platinum(II)-Assisted Platinum(IV) Substitution
Figure 1. Absorbance at 360 nm vs time of the reactions of 5 mM
[PtIV(dach)Cl4]and20mM5′-dGMPwithandwithout1mM[PtII(NH3)2(CBDCA-
O,O′)] in 100 mM NaCl, pH 8.2 at 40 °C. The absorbance at 360 nm is
due to [PtIV(dach)Cl4], [PtIV(dach)(5′-dGMP)Cl3], and [PtIV(NH3)2(5′-
dGMP)Cl(CBDCA-O,O′)].
Figure 2. A360 and A390 vs time of the reactions of 5 mM [PtIV(dach)Cl4]
and 20 mM cGMP with and without 2 mM [PtII(NH3)2(CBDCA-O,O′)] in
100 mM NaCl at 50 °C and pH 8.2. A360 and A390 are shown in pink and
purple, respectively. The open circle is the calculated A390 only due to
[PtIV(dach)(cGMP)Cl(CBDCA-O,O′)].
target complex [PtIV(dach)(35Cl)2(37Cl)2] obtained from the
reaction of [PtIV(dach)(OH)2Cl2] was generated in situ with
Cl- ions under acidic conditions. This step is usually
accomplished by using HCl as both the source of the acid
as well as the chloride. For this synthesis, however, NaCl and
HNO3 were used instead because Na37Cl was the only
available source for 37Cl-. A larger excess of Na37Cl relative
to the acid was used to ensure a reasonable yield.
ratiosof[PtIV(dach)(cGMP)Cl(CBDCA-O,O′)]and[PtIV(NH3)2-
(cGMP)Cl(CBDCA-O,O′)] and plotted in Figure 2 marked
as open circles. The maximum 3 mM of [PtIV(dach)(cGMP)-
Cl(CBDCA-O,O′)] from 5 mM [PtIV(dach)Cl4] and 2 mM
[PtII(NH3)2(CBDCA-O,O′)] was generated after 1 h. The
same concentration of [PtIV(dach)(cGMP)Cl(CBDCA-O,O′)]
was generated after 2 h from the 5 mM [PtIV(dach)Cl4]
solution without [PtII(NH3)2(CBDCA-O,O′)]. The results
indicate that cGMP simply binds to [PtIV(dach)Cl4] without
a redox reaction. The addition of [PtII(NH3)2(CBDCA-O,O′)]
expedites the binding, indicating that [PtII(NH3)2(CBDCA-
O,O′] accelerates the chloride substitution of [PtIV(dach)Cl4]
by cGMP.
Reactions of [PtIV(dach)Cl4] and 5′-dGMP with and
without [PtII(NH3)2(CBDCA-O,O′)]. Figure 1 compares the
time course of A360 of the [PtIV(dach)Cl] and 5′-dGMP
reaction with and without [PtII(NH3)2(CBDCA-O,O′)]. With-
out [PtII(NH3)2(CBDCA-O,O′)], there is a long induction time
for the reaction to start, which is significantly shortened by
the addition of [PtII(NH3)2(CBDCA-O,O′)]. This clearly
indicates that [PtII(NH3)2(CBDCA-O,O′] accelerates the
redox reaction between [PtIV(dach)Cl4] and 5′-dGMP.
Reactions of [PtIV(dach)Cl4] and cGMP with and
without [PtII(NH3)2(CBDCA-O,O′)]. Figure 2 compares the
time course of A360 and A390 of the [PtIV(dach)Cl4] and cGMP
reaction with and without [PtII(NH3)2(CBDCA-O,O′)]. For
both reactions, A360 does not change while A390 increases to
a maximum without decreasing. A360 is mainly due to the
PtIV complexes and is an excellent indication of the redox
reaction because PtII complexes do not absorb at 360 nm.
A390 is primarily due to the PtIVcGMP adducts. Absorbance
at 390 nm without [PtII(NH3)2(CBDCA-O,O′)] is due to only
[PtIV(dach)(cGMP)Cl(CBDCA-O,O′)]. Absorbance at 390
nm with [PtII(NH3)2(CBDCA-O,O′)] is due to both [PtIV(NH3)2-
(cGMP)Cl(CBDCA-O,O′)] and [PtIV(dach)(cGMP)Cl(CB-
DCA-O,O′)]. The molar absorptivity of [PtIV(NH3)2(cGMP)-
Cl(CBDCA-O,O′)] is approximately twice as big as that of
[PtIV(dach)(cGMP)Cl(CBDCA-O,O′)], which is the reason
why the final A390 with [PtII(NH3)2(CBDCA-O,O′)] is higher
than that without it. A390 only due to [PtIV(dach)(cGMP)
Cl(CBDCA-O,O′)] in the presence of the [PtII(NH3)2(CBDCA-
O,O′)] reaction was calculated from the molar absorptivity
The products of the [PtIV(dach)Cl]/[PtII(NH3)2(CBDCA-
O,O′)]/cGMP mixture were identified by LC/MS (Figure 3).
The peaks at m/z 749.9 and 757.9 correspond to [PtIV(NH3)2-
(cGMP)(Cl)(CBDCA-O,O′)] and [PtIV(dach)(cGMP)Cl3],
respectively.
Reaction of trans,cis-[PtIV(dach)(37Cl)2(35Cl)2], [PtII(en)-
(35Cl)2], and 9-EtG. Figure 4 displays the LC/MS spectra of
the trans,cis-[PtIV(dach)(37Cl)2(35Cl)2]/[PtII(en)(35Cl)2]/9-EtG
reaction mixture. The reactants [PtII(en)(35Cl)2], trans,cis-
[PtIV(dach)(37Cl)2(35Cl)2], and 9-EtG are detected at m/z
324.9, 454.9, and 177.9, respectively. Two main products
are found at m/z 539.9 and 596.0. The 539.9 amu is
equivalent to the mass of [PtIV(en)(9-EtG)(37Cl)(35Cl)2],
which is the sum of 324.9 ([PtII(en)(35Cl)2]), 36.97 (37Cl),
and 177.9 (9-EtG) amu. This means 37Cl and 9-EtG are added
to the axial position of [PtII(en)(35Cl)2], producing trans,cis-
[PtIV(en)(9-EtG) (37Cl)(35Cl)2]. The 596.0 amu is equivalent
to the mass of [PtIV(dach)(9-EtG)(37Cl)(35Cl)2], which is
the subtraction of 36.97 amu (37Cl) from 454.9 amu
([PtIV(dach)(37Cl)2(35Cl)2]) and the addition of 177.9 amu
(9-EtG). This is a result of axial 37Cl breaking from
Inorganic Chemistry, Vol. 47, No. 4, 2008 1355

Choi et al.
Figure 3. MS spectra of (a) [PtIV(NH3)2(cGMP)(Cl)(CBDCA-O,O′)] (m/z 749.9) and (b) [PtIV(dach)(cGMP)Cl3] (m/z 757.9) from the reaction of PtIV(dach)Cl4]/
[PtII(NH3)2(CBDCA-O,O′)]/cGMP (5/2/20 mM).
[PtIV(dach)(37Cl)2(35Cl)2] followed by the addition of 9-EtG,
generating trans,cis-[PtIV(dach)(9-EtG)(37Cl)(35Cl)2]. The
[PtII(dach)(37Cl)2] product could not be detected by LC/MS.
However, [PtII(dach)(37Cl)(9-EtG)] was detected at m/z 524.0
(Supporting Information), which is an indirect evidence of
the generation of [PtII(dach)(37Cl)2].
Determination of the Kinetic Rate Law for the
Substitution. Figure 5a compares the time course of A390 of
[PtIV(dach)Cl4]/cGMP/[PtII(dach)Cl2] at varied concentrations
of each component. The kinetic curves for different concen-
trations were obtained four times, and the average A390 versus
time is displayed in Figure 5b. As seen in Figure 5b and
Table 1, within experimental error, the initial rate was
reduced by half when the concentration of one component
was reduced by half. Therefore, the substitution reaction
follows the third-order rate law with first-order for each
[PtIV], G, and [PtII]. Our result is consistent with the known
rate law for the BP PtII-catalyzed PtIV-substitution reaction.11
The rate law and identification of the PtIVG species is the
sound basis of eq 1 in Scheme 1.
Reactions of 5′-dGMP with [PtIV(en)(OCOCF3)2Cl2],
[PtIV(a,cha)(OCOCH3)2Cl2], [PtIV(en)(OH)2Cl2], [PtIV(en)-
Cl4], and [PtIV(en)Br4]: Effect of Axial Ligands. Figure 6
compares A360 versus time of the reactions of 5′-dGMP
with [PtIV(en)(OCOCF3)2Cl2], [PtIV(a,cha)(OCOCH3)2Cl2],
[PtIV(en)(OH)2Cl2], [PtIV(en)Cl4], and [PtIV(en)Br4]. The A360
of [PtIV(en)(OCOCF3)2Cl2], [PtIV(a,cha)(OCOCH3)2Cl2], and
[PtIV(en)(OH)2Cl2] does not change over time, indicating that
these complexes did not react. The decrease of A360 of
[PtIV(en)Cl4] and [PtIV(en)Br4] reactions indicates that both
of these PtIV complexes oxidized 5′-dGMP. This was
confirmed by detection of 8-oxo-5′-dGMP in the reaction
mixture by HPLC and LC/MS.
The rate constants for the reactions of [PtIV(en)Cl4] and
[PtIV(en)Br4] with 5′-dGMP at pH 6.5 and 30 °C were
obtained by fitting A360 versus time data to eqs 1 and 2 in
Scheme 1 using DynaFit.13 As shown above, the rate law
for the first reaction in Scheme 1 (eq 1) follows the third-
order rate law with first-order in [PtII], [PtIV], and G (entering
group). The close agreement between experimental and
modeled absorbance versus time is shown in Figure 6b,c.
The ks values are 28.5 and 1360 M-2 s-1 for [PtIV(en)Cl4]
and [PtIV(en)Br4], respectively. The ke values are 1.4 × 10-4
and 0.4 × 10-4 s-1 for [PtIV(en)Cl4] and [PtIV(en)Br4],
respectively. The values are shown in Table 2.
Reactions of 5′-dGMP with [PtIV(NH3)2Cl4], [PtIV(en)-
Cl4], and [PtIV(dach)Cl4]: Effect of Carrier Ligands. The
reactions of 5′-dGMP with [PtIV(NH3)2Cl4], [PtIV(en)Cl4], and
[PtIV(dach)Cl4] at 50 °C are compared in Figure 7. The
kinetic rate constants were obtained by fitting A360 versus
time data to eqs 1 and 2 in Scheme 1 using DynaFit.13 The
close agreement between the experimental and modeled A360
versus time is shown in Figure 6. The ks values are 85.8,
25.2, and 11.1 M-2 s-1 for [PtIV(NH3)2Cl4], [PtIV(en)Cl4], and
(13) Kuzmic, P. Anal. Biochem. 1996, 237, 260–273.
1356 Inorganic Chemistry, Vol. 47, No. 4, 2008

Platinum(II)-Assisted Platinum(IV) Substitution
Figure 4. MS spectra of (a) [PtIV(dach)37Cl235Cl2], (b) [PtIV(en)(9-EtG)(37Cl)(35Cl)2], and (c) [PtIV(dach)(9-EtG)(37Cl)(35Cl)2] from the reaction of [PtII(en)35Cl2]/
[PtIV(dach)(37Cl)2(35Cl)2]/9-EtG (0.5/1/1.5 mM) at 50 °C.
[PtIV(dach)Cl4], respectively. The ke values are 1.5 × 10-4,
1.5 × 10-4, and 14.2 × 10-4 s-1 for [PtIV(NH3)2Cl4],
[PtIV(en)Cl4], and [PtIV(dach)Cl4], respectively. The values
are summarized in Table 3.
short-lived intermediate for the PtII-substitution14 and the PtII-
catalyzed PtIV-substitution mechanism.11 In the square-planar
PtII-substitution reaction, the geometry of the five-coordinate
species is thought to be trigonal-bipyramidal because it is
able to account for all of the experimental evidence avail-
able.14 In the PtII-catalyzed PtIV-substitution reaction, the
geometry of the five-coordinate species was not explicitly
discussed in the literature. However, because of the retention
of the geometric configuration for the trans PtIV substrate in
all of the reactions studied, the five-coordinate species is
drawn as square-pyramidal in our scheme.
The five-coordinate [PtIIA2B2G] subsequently binds to
[PtIVC2X2D2] through a halo bridge to form a dimer. This
dimer could not be detected, perhaps because it is a short-
lived intermediate. In the dimer, two electrons transfer from
the PtII center to the PtIV center, generating the new species
[PtIVA2XGB2] and [PtIIC2D2]. The new [PtIIC2D2] complex
was detected as [PtII(dach)Cl2] from the [PtIV(dach)Cl4]/
Discussion
As shown in Figures 1 and 2, [PtII(NH3)2(CBDCA-O,O′)]
accelerates the reaction of [PtIV(dach)Cl4] with 5′-dGMP and
with cGMP. The former is a redox reaction that involves a
substitution followed by an electron transfer between PtIV
and 5′-dGMP, and the latter is only a substitution reaction.
Therefore, it can be deduced that [PtII(NH3)2(CBDCA-O,O′)]
accelerates the substitution step in the redox reaction. This
type of substitution mechanism is consistent with the BP
mechanism (Scheme 2).11 The scheme is fundamentally the
BP scheme11 adapted to our system. The first step is a
binding of the G derivative to [PtIIA2B2] at the fifth position
to produce a five-coordinate [PtIIA2B2G]. Like other re-
searchers in the literature, we could not detect this five-
coordinate species. However, it is generally accepted as a
(14) Langford, C. H.; Gray, H. B. Ligand Substitution Processes; W. A.
Benjamin Inc.: New York, 1965; pp 18–48.
Inorganic Chemistry, Vol. 47, No. 4, 2008 1357

Choi et al.
Figure 5. (a) A390 vs time of the reactions of [PtIV(dach)Cl4], cGMP, and [PtII(dach)Cl2] in 100 mM NaCl at 20 °C and pH 8.2. Because of the spectral noise
during the first 30 min initial period, 30 min was subtracted from the time scale. The absorbance was made zero at 30 min in order to show the first 10 min
(actually 30–40 min after starting the reaction) absorbance difference clearly. (b) Average A390 from the four sets of reactions with the same concentration
of each component.
Table 1. Initial Rates for the Reaction of [PtIV(dach)Cl4]/cGMP/[PtII(dach)Cl2] at Different Concentrations at 25 °C
[PtIV(dach)Cl4]/cGMP/
[PtII(dach)Cl2] mM
rate 1
rate 2
rate 3
rate 4
average rate
× 104 M-1 s-1
× 104 M-1 s-1
× 104 M-1 s-1
× 104 M-1 s-1
× 104 M-1 s-1
4/20/0.4
2/20/0.4
4/10/0.4
4/20/0.2
3.7
2.6
2.6
2.2
3.4
2.0
2.1
1.4
3.6
2.1
1.9
1.8
3.9
1.6
2.5
1.5
3.7 ( 0.2
2.1 ( 0.4
2.3 ( 0.3
1.7 ( 0.3
Scheme 1. Kinetic Model for the Oxidation of 5′-dGMP by
6). The required substitution for the redox reaction could
not occur because OH, OCOCF3, and OCOCH3 are not
suitable bridging ligands for the inner-sphere electron
transfer. Their lowest empty orbitals for the electron transfer
are at a relatively high energy.11 On the other hand, chloro
and bromo complexes readily react, particularly the bromo
complex. As shown in Figures 6b,c, the reaction completion
time of 1 mM [PtIV(en)Cl4] is about 70 h while that of 0.3
mM [PtIV(en)Br4] is about 7 h under the same reaction
conditions. This is due to the 50-fold larger substitution rate
(ks) of the bromo complex in spite of the slower electron-
transfer rate (ke) compared to the chloro complex. The
heavier and more polarizable bromo ligand has lower-energy
orbitals available that can participate in bridge formation and
thus provide an effective path for the electron transfer from
the PtII to PtIV centers.11
It is also interesting to note that the reactivities of these
complexes with 5′-dGMP and with ascorbate are significantly
different. The redox rates for reaction with ascorbate are
dependent on the magnitude of the reduction potentials of
the PtIV complexes, which are in the order trans,cis-
[PtIV(en)(OH)2Cl2] < cis,trans,cis-[PtIV(NH3)(C6H11NH2)(OCO-
CH3)2Cl2] < [PtIV(en)Cl4] < trans,cis-[PtIV(en)(OCOCF3)2Cl2].6
The [PtIV(en)(OCOCF3)2Cl2] complex, which is quickly re-
duced by ascorbate, does not react with 5′-dGMP. Appar-
ently, PtIV reacts with ascorbate via a mechanism different
[PtIVC2Cl2D2]
8
7–9
1
5′-dGMP reaction previously by IR and H NMR.
It
again reacts with G followed by [PtIVC2X2D2], generating
[PtIVC2GXD2] and [PtIIC2D2]. The LC/MS detection of
[PtIV(en)(5′-dGMP)Cl3] and [PtIV(dach)(5′-dGMP)Cl3] from
the [PtIV(dach)Cl4]/[PtII(en)Cl4]/5′-dGMP reaction10 and
[PtIV(NH3)2(cGMP)(Cl)(CBDCA-O,O′)] and [PtIV(dach)(cGMP)-
Cl3] (Figure 3b,c) from the [PtIV(dach)Cl4]/[PtII(NH3)2(CBDCA-
O,O)]/cGMP reaction mixture is consistent with the BP
mechanism.
The detection of trans,cis-[PtIV(en)(9-EtG)(37Cl)(35Cl)2]
and trans,cis-[PtIV(dach)(9-EtG) (37Cl)(35Cl)2] (Figure 4) from
the [PtII(en)(35Cl)2]/trans,cis-[PtIV(dach)(37Cl)2(35Cl)2]/9-EtG
reaction mixture is also consistent with Scheme 2. The
substitution can be thought of as an oxidative addition
reaction of the planar PtII complex in which the entering
ligand G and the halo ligand from the PtIV complex are added
to the PtII complex in the axial positions. From the original
PtIV point of view, the axial halo ligand is substituted by the
G derivative.
The PtIV complexes such as trans,cis-[PtIV(en)(OH)2Cl2],
trans,cis-[PtIV(en)(OCOCF3)2Cl2], and cis,trans,cis-[PtIV(NH3)-
(C6H11NH2)(OCOCH3)2Cl2] do not oxidize 5′-dGMP (Figure
1358 Inorganic Chemistry, Vol. 47, No. 4, 2008

Platinum(II)-Assisted Platinum(IV) Substitution
Table 2. Substitution and Electron-Transfer Rates for the Reaction of
PtIV Complexes with 5′-dGMPa
[PtIV(en)Cl4]
[PtIV(en)Br4]
[PtIV(dach)Cl4]
ks (M-2 s-1
)
28.5 ( 1.6
1.4 ( 0.5
1360 ( 50
0.4 ( 0.2
8.1 ( 2.7
3.2 ( 0.8
ke × 104 (s-1
)
a
The rate constants were obtained from the kinetic curves in Figure
5b,c for reactions [PtIV(en)Cl4/5′-dGMP/NaCl (1/5/100 mM) and [PtIV(en)Br4/
5′-dGMP/KBr (0.3/5/100 mM), pH 6.5 at 30 °C. Values are reported as
(1 standard deviation of the mean for three measurements.
Figure 7. Absorbance at 360 nm vs time of [PtIV(en)Cl4/[PtII(en)Cl2]/5′-
dGMP (1/0.2/20 mM), [PtIV(dach)Cl4/[PtII(dach)Cl2]/5′-dGMP (1/0.2/20
mM), [PtIV(NH3)2Cl4/[PtII(NH3)2Cl2]/5′-dGMP (1/0.2/20 mM) reactions in
100 mM NaCl, pH 8.3 at 50 °C.
Table 3. Substitution and Electron-Transfer Rates for the Reaction of
PtIV Complexes with 5′-dGMPa
[PtIV(NH3)2Cl4]
[PtIV(en)Cl4]
[PtIV(dach)Cl4]
ks (M-2 s-1
)
85.8 ( 6.2
1.5 ( 0.1
25.2 ( 4.1
1.5 ( 0.1
11.1 ( 2.2
14.2 ( 0.8
ke × 104 (s-1
)
a
The rate constants were obtained from the kinetic curves in Figure 6
for the reactions of [PtIV(en)Cl4/[PtII(en)Cl2]/5′-dGMP (1/0.2/20 mM),
[PtIV(dach)Cl4/[PtII(dach)Cl2]/5′-dGMP (1/0.2/20 mM), and [PtIV(NH3)2Cl4/
[PtII(NH3)2Cl2]/5′-dGMP (1/0.2/20 mM) reactions in 100 mM NaCl, pH
8.3 at 50 °C. Values are reported as (1 standard deviation of the mean for
three measurements.
from that with 5′-dGMP. Depending on the structure of PtIV,
reductive elimination,15 and outer- and inner-sphere16 mech-
anisms have been proposed for reduction by ascorbate.
When PtIV complexes with different carrier ligands but
the same chloro axial ligands are compared, the ks values
increase in the order [PtIV(dach)Cl4] < [PtIV(en)Cl4] < cis-
[PtIV(NH3)2Cl4], which is the inverse order of their carrier
ligand size. The smaller the carrier ligand, the less steric
hindrance for the bridge formation. The values for the rate
constant, ke for [PtIV(NH3)Cl4] and cis-[PtIV(en)Cl4] are
(15) (a) Lemma, K.; Shi, T.; Elding, L. I. Inorg. Chem. 2000, 39, 1728–
1734. (b) Lemma, K.; Sargeson, A. M.; Elding, L. I. J. Chem. Soc.,
Dalton Trans. 2000, 1167–1172. (c) Lemma, K.; Berglund, J.; Farrell,
N.; Elding, L. I. J. Biol. Inorg. Chem. 2000, 5, 300–306.
Figure 6. Absorbance at 360 nm vs time of PtIV complexes with 5′-dGMP.
Reaction conditions: (a) [PtIV(en)(OH)2Cl2]/[PtII(en)Cl2]/5′-dGMP/NaCl (2/0.2/50/
100 mM), [PtIV(en)(OCOCF3)2]/[PtII(en)Cl2]/5′-dGMP/NaCl (2/0.2/50/100 mM),
[PtIV(a,cha)(OCOCH3)2Cl2]/5′-dGMP/NaCl (2/50/100 mM), [PtIV(en)Cl4/[PtII(en)Cl2]/
5′-dGMP/NaCl (2/0.2/50/100 mM), [PtIV(en)Br4/5′-dGMP/KBr (0.3/50/100 mM),
pH 6.5 at 30 °C. (b) [PtIV(en)Cl4/5′-dGMP/NaCl (1/5/100 mM), pH 6.5 at 30 °C.
(c) [PtIV(en)Br4/5′-dGMP/KBr (0.3/5/100 mM). pH 6.5 at 30 °C.
(16) (a) Evans, D. J.; Green, M. Inorg. Chim. Acta 1987, 130, 183–184.
(b) Bose, R. N.; Weaver, E. L. J. Chem. Soc., Dalton Trans. 1997,
1797–1799. (c) Hindmarsh, K.; House, D. A.; van Eldik, R. Inorg.
Chim. Acta 1998, 278, 32–42. (d) Weaver, E. L.; Bose, R. N. J. Inorg.
Biochem. 2003, 95, 231–239.
Inorganic Chemistry, Vol. 47, No. 4, 2008 1359

Choi et al.
Scheme 2. Proposed Mechanism for the PtII-Accelerated PtIV-Substitution Reaction for Guanosine Derivatives
The scheme is fundamentally the BP scheme11 adapted to our system. The intermediates were not detected experimentally and are marked in brackets.
Because both [PtIVA2GXB2] and [PtIVC2GXD2] can generate 8-oxo-5′-dGMP, when G is 5′-dGMP, the PtII analogues should be PtIV complexes (A ) C and
B ) D) when ks and ke of [PtIVC2X2D2] are obtained.
similar, but ks of the former is 3-fold larger, resulting in
a faster rate for the overall redox reaction. The cis-
[PtIV(NH3)2Cl4] complex has a 7-fold higher ks but a 10-
fold lower ke than [PtIV(dach)Cl4], resulting in the faster
overall redox rate during the first half of the reaction time
(Figure 7). Taken together, these results strongly suggest that
substitution plays a critical role in the overall redox reaction
between 5′-dGMP and PtIV and is consistent with the BP
mechanism.
the original PtIV point of view, the axial halo ligand is
substituted by the G derivative. The PtIV complex must
possess axial halo ligands for the substitution, and those with
a larger axial halo and a smaller carrier ligand exhibit higher
substitution rates.
Acknowledgment. This work was supported by the
National Science Foundation (Grants CHE-0450060 and
CHE-0707357). Financial support is acknowledged from the
NSF (Grant CHE-0520708) for their support in the purchase
of the LC/MS system used in the course of this work. We
are grateful to Drs. K. Wosikowski and C. Probst at GPC
Biotech AG for satraplatin.
Conclusion
We have shown that the substitution step is essential for
the oxidation of 5′-dGMP by PtIV. The substitution is
accelerated by PtII and follows the BP mechanism. The PtII
complex does not have to possess a halo leaving group and,
therefore, the substitution can be thought of as an oxidative
addition reaction of the planar PtII complex in which the
entering ligand G and the halo ligand from the PtIV complex
are added to the PtII complex in the axial positions. From
Supporting Information Available: MS spectrum of [PtII(dach)-
(37Cl)(9-EtG)]. This material is available free of charge via the
Internet at http://pubs.acs.org.
IC701868B
1360 Inorganic Chemistry, Vol. 47, No. 4, 2008