Phosphinogold(I) dithiocarbamate complexes: Effect of the nature of phosphine ligand on anticancer properties

Article abstract of DOI:10.1021/ic4025926

The reactions of potassium salts of the dithiocarbamates L {where L = pyrazolyldithiocarbamate (L1), 3,5-dimethylpyrazolyldithiocarbamate (L2), or indazolyldithiocarbamate (L3)} with the gold precursors [AuCl(PPh₃)], [Au₂Cl₂(dppe)], [Au₂Cl₂(dppp)], or [Au₂Cl₂(dpph)] lead to the new gold(I) complexes [AuL(PPh₃)] (1-3), [Au₂L₂(dppe)] (4-6), [(Au₂L₂)(dppp)] (7-9), and [Au₂(L) ₂(dpph)] (10-12) {where dppe = 1,2-bis(diphenylphosphino)ethane, dppp = 1,3-bis(diphenylphosphino)propane, and dpph = 1,6-bis(diphenylphosphino) hexane}. These gold compounds were characterized by a combination of NMR and infrared spectroscopy, microanalysis, and mass spectrometry; and in selected cases by single-crystal X-ray crystallography. Compounds 4-6, which have dppe ligands, are unstable in solution for prolonged periods, with 4 readily transforming to the Au₁₈ cluster [Au₁₈S ₈(dppe)₆]Cl₂ (4a) in dichloromethane. Compounds 1-3 and 7-12 are all active against human cervical epithelioid carcinoma (HeLa) cells, but the most active compounds are 10 and 11, with IC₅₀ values of 0.51 μM and 0.14 μM, respectively. Compounds 10 and 11 are more selective toward HeLa cells than they are toward normal cells, with selectivities of 25.0 and 70.5, respectively. Further tests, utilizing the 60-cell-line Developmental Therapeutics Program at the National Cancer Institute (U.S.A.), showed 10 and 11 to be active against nine other types of cancers.

Full text of DOI:10.1021/ic4025926

Article
pubs.acs.org/IC
Phosphinogold(I) Dithiocarbamate Complexes: Effect of the Nature
of Phosphine Ligand on Anticancer Properties
Frankline K. Keter,†,⊥ Ilia A. Guzei,†,‡ Margo Nell,§ Werner E. van Zyl,†,¶ and James Darkwa*,†
†Department of Chemistry, University of Johannesburg, P.O. Box 524, Auckland Park 2006, South Africa
‡Department of Chemistry, University of Wisconsin-Madison, 1101 University Avenue, Madison, Wisconsin 53706, United States
§Department of Pharmacology, University of Pretoria, Pretoria 0002, South Africa
⊥Advanced Materials Division, Mintek, Private Bag X3015, Randburg 2125, South Africa
S
* Supporting Information
ABSTRACT: The reactions of potassium salts of the dithiocarbamates L
{where L = pyrazolyldithiocarbamate (L1), 3,5-dimethylpyrazolyldithio-
carbamate (L2), or indazolyldithiocarbamate (L3)} with the gold
precursors [AuCl(PPh3)], [Au2Cl2(dppe)], [Au2Cl2(dppp)], or [Au2Cl2-
(dpph)] lead to the new gold(I) complexes [AuL(PPh3)] (1−3),
[Au2L2(dppe)] (4−6), [(Au2L2)(dppp)] (7−9), and [Au2(L)2(dpph)]
(10−12) {where dppe = 1,2-bis(diphenylphosphino)ethane, dppp = 1,3-
bis(diphenylphosphino)propane, and dpph
= 1,6-bis-
(diphenylphosphino)hexane}. These gold compounds were characterized
by a combination of NMR and infrared spectroscopy, microanalysis, and
mass spectrometry; and in selected cases by single-crystal X-ray
crystallography. Compounds 4−6, which have dppe ligands, are unstable
in solution for prolonged periods, with 4 readily transforming to the Au18
cluster [Au18S8(dppe)6]Cl2 (4a) in dichloromethane. Compounds 1−3 and 7−12 are all active against human cervical epithelioid
carcinoma (HeLa) cells, but the most active compounds are 10 and 11, with IC50 values of 0.51 μM and 0.14 μM, respectively.
Compounds 10 and 11 are more selective toward HeLa cells than they are toward normal cells, with selectivities of 25.0 and 70.5,
respectively. Further tests, utilizing the 60-cell-line Developmental Therapeutics Program at the National Cancer Institute
(U.S.A.), showed 10 and 11 to be active against nine other types of cancers.
shown to possess anticancer properties. These include the
report by Tiekink and co-workers that triorganophosphinogold-
(I) dithiocarbamates of general formula [(R3P)Au(S2CNR2)]
were active against seven human cancer cell lines.13 Of these,
[(Et3P)Au(S2CNEt2)] (Et = ethyl) was the most active against
the IGROV ovarian cell line (ID50 = 12 ng/mL).13
INTRODUCTION
■
Gold compounds in the oxidation states of +1 and +3 continue
to be of interest in medicinal chemistry because their efficacies
toward certain diseases can be fine-tuned.1,2 For instance, a
number of gold(III) dithiocarbamates have been reported as
anticancer agents by Fregona and co-workers.3 This includes
the classics [Au(DMDT)X2] and [Au(ESDT)X2] (where X =
Cl, Br; DMDT = N,N-dimethyldithiocarbamate; ESDT =
ethylsarcosinedithiocarbamate), which were shown to be 1−4
times more active than cis-platin and particularly potent against
the lymphoproliferative-type HL-60 cell line.3,4 Various modes
of action have been proposed for these compounds, from weak
DNA binding to proteasome inhibition.5−7 A second
generation of gold(III) dithiocarbamate (dtc) anticancer
compounds includes the gold(III)-dipeptidedithiocarbamato
derivatives AuX2(dtc-Sar-AA-O(t-Bu))] (X = Cl, Br; Sar =
Sarcosine; AA = Gly, α-aminoisobutyric acid, and L-Phe)8 and
several [AuX2(pdtc)] (X = Cl, Br; pdtc = oligopeptidedithio-
carbamate) compounds, which are active against four human
tumor cell lines at lower IC50 values than is cis-platin.9
An early report by Mirabelli et al.,14 supported by others,12,13
on various gold(I) compounds of structural formula R3P−Au−
X (R = ethyl; X = thioglucose (SR)) has demonstrated that the
presence of a P−Au−S motif in these compounds enhances
their anticancer activity. Notably, phosphinogold(I) thiolate
complexes of type [Au(PR3)(SR)] are more active than gold(I)
thiolates of type [Au(SR)], suggesting that the presence of
phosphine ligands increases the lipophilicity and membrane
permeability of the phosphinogold(I) complexes that make
them active.12
It has been established that gold compounds, such as
auranofin, act against cancer cells via the mitochondria by
inhibiting thioredoxin reductase.11−18 In doing so, gold(I)
binds the C-terminal of the redox-active selenocysteine, leading
Gold(I) thiolate compounds have also been of interest as
anticancer agents since the first reports on the antiarthritic
agent auranofin,1,10,11 and other gold(I) thiolates12−14 were
Received: October 16, 2013
© XXXX American Chemical Society
A
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to cytotoxic effects.18−20 For these reasons, several
phosphinogold(I) thiolate complexes have been prepared and
investigated for their anticancer activities. Examples include the
hydrophilic tetrakis((tris(hydroxymethyl)phosphino)gold(I)
complex, which was reported to be cytotoxic toward HCT-15
tumor cells.21 Other examples are [Au(PPh3)(sppa)] (sppa =
sulfanylpropenoate),22 [Au(PEt3)(K3TSC)] (K3TSC = vitamin
K3 derivative),23 and the gold(I) 7-azacoumarin complex,24
which exhibit activity against HeLa and A2780cis cells,
[(AuOmS)2(Ph2P(CH2)2PPh2)] (OmS = lupinylsulfide), ex-
hibiting activity against ovarian carcinomas,25 and [Et3PAu-
(S2CNEt2)] and [Ph3PAu(S2CNC2C4H8)], which exhibit
activity against breast, ovarian, and colon cancer.12,13a This
strongly suggests that one strategy to make gold(I) anticancer
drugs is to prepare molecules that have a phosphine and a
sulfur-containing ligand bound to gold(I) to have the P−Au−S
motif.
RESULTS AND DISCUSSION
Triphenylphosphinogold(I) Dithiocarbamato Com-
plexes (1−3). The gold(I) complexes 1−3 were prepared
from the reaction of [AuCl(PPh3)] and the dtc ligands L1−L3
(Scheme 1) and were isolated as orange-yellow solids. The
■
Scheme 1. Preparations of (Monophosphino)gold(I)
Dithiocarbamate Complexes 1−3
For phosphinogold(I) complexes, the nature of the
phosphine ligand appears to be important in regulating their
anticancer behavior. For example a report on the anticancer
properties of dinuclear di(phosphino)alkane gold(I) chloride
by Mirabeli et al.26 suggested two key factors determine the
activity of these gold(I) compounds. The first is to have phenyl
groups on the phosphorus, and the second is the effect of CH2
linkers, with the compound with two CH2 linkers being more
active than compounds with three, four, five, or six CH2 linkers.
However, a recent report by Raubenheimer and co-workers
indicates that dinuclear di(phosphino)alkane gold(I) azole
complexes with longer CH2 linkers are highly active for a
number of cancer cell lines, and at very low dosage.27 Clearly,
from these two reports, the nature of the di(phosphino)-
alkanegold(I) and the ancillary ligand bound to the gold(I) is
important in determining the anticancer activity of these
compounds.
We report here new phosphinogold(I) dithiocarbamate
complexes, using triphenylphosphine, 1,2-bis-
( d i p h e n y l p h o s p h i n o ) e t h a n e ( d p p e ) , 1 , 3 - b i s -
(diphenylphosphino)-propane (dppp), 1,6-bis-
(diphenylphosphino)hexane (dpph), and the pyrazol-1-yl- and
indazol-1-yl-based dithiocarbamate ligands (dtcs) to test the
hypothesis that combining alkane chain length in the
diphosphino ligand and using dithiocarbamate as the thiolate
can lead to new gold anticancer compounds. Our choice of
pyrazoles and indazole as the backbone of the dithiocarbamate
is based on their medicinal properties.28,29 This approach to
prepare phosphinogold(I) dithiocarbamato compounds has
resulted in compounds that have excellent anticancer activities
against several cancer cell lines. Several reports on the
chemistry of gold(I) dithiocarbamate compounds,30−39 gold(I)
thiosemicarbazones, and gold(I) thiosemicarbazone com-
plexes40,41 have appeared in the literature. Some of these
compounds have anticancer activity that has been attributed to
the synergistic effect of the metals and the thiolate ligands, but
none of these compounds has phosphine ligands.
complexes were characterized by a combination of spectrosco-
py, mass spectrometry, and microanalysis; in the case of 3,
single-crystal X-ray crystallography was also used. Complex 1
1
displays a typical H NMR spectrum for these compounds,
which shows dtc protons for 4-pz and 5-pz proton peaks at 6.27
and 7.63 ppm, respectively. The 31P{1H} NMR spectra of
complexes 1−3 displayed broad signals between 29.9 and 36.3
ppm.
Crystal data, together with the data collection and refinement
parameters, are presented in Table 1. Compound 3 exhibits the
expected near-linear geometry about the Au(I) center with the
P1−Au1−S1 angle spanning 175.36(2)°. A molecular drawing
of 3 is shown in Figure 1. The Au−P distance of 2.2533(6) Å is
in excellent agreement with the average value of 2.26(2) Å
obtained by averaging 628 Au−PPh3 distances from 501
complexes reported to the Cambridge Structural Database
(CSD).44 The Au−S distance in 3 (2.3272(6) Å) is slightly
longer than the Au−P distance, but is somewhat shorter than
the average distance of 2.38(16) Å computed based on 89 Au−
S distances for 58 dithiocarbamate complexes reported to the
CSD. However, the difference with the latter is not statistically
significant as Au−S bond lengths fall in a broad range. In
closely related compounds these distances are as follows:
[Au(S2CNEt2)(PPh3)], Au−S (2.338(4) Å) and Au−P
(2.251(3) Å)33; [Au(S2CNC10H20O4)(PPh3)] (where
C10H20O4 = aza-15-crown-5), Au−S (2.339(3) Å) and Au−P
(2.253(3) Å).31 The C−S distances are different, with the C1−
S1 distance to the coordinated S atom being longer (single
bond, 1.732(3) Å) than the C1−S2 bond that clearly shows
multiple character (1.663(3) Å).
(Diphosphino)ethanegold(I) Dithiocarbamato Com-
plexes (4−6). Complexes 4−6 (Scheme 2) were prepared
from the reaction of [Au2Cl2(dppe)] with 2 equiv of L1−L3
and were isolated as flaky yellow solids in moderate yields. In
the 1H NMR spectra of 4−6, the CH2 protons of the
di(phosphino)alkane ligand appeared upfield (1.8−2.9 ppm) as
second-order multiplets, and the 13C{1H} NMR spectra
showed peaks at ca. 215.0 and ca. 129.2−133.5 ppm, which
were assigned to C(CS) and the phenyl carbons,
We have further shown for the first time that dithiocarbamate
can act as a sulfur transfer reagent to form the high-nuclearity
gold cluster [Au18S8(dppe)6]2+ from [Au(L1)(dppe)], a
previously known cluster that was erroneously reported as
neutral [Au18S8(dppe)6]·H2O.42 The heavier congener of this
cluster, [Au18Se8(dppe)6]2+, is known.43
B
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Table 1. Crystallographic Data for Compounds 3, 4a, and 12
3
4a
12
empirical formula fw
temp (K)
C26H20AuN2PS2 652.50
C156H144Au18Cl2P12S8 6263.13
C46H42Au2N4P2S4 1234.95
150(2)
100(2)
105(2)
a
wavelength (Å)
cryst syst
0.710 73
0.710 73
0.774 90
monoclinic
monoclinic
monoclinic
space group
a (Å)
P21/n
P21/n
P21/n
9.1382(12)
17.6854(7)
13.499(11)
b (Å)
22.487(3)
34.4361(14)
11.560(19)
c (Å)
11.3760(14)
29.1894(12)
15.396(15)
β (deg)
volume (Å3)
91.541(2)
92.4570(10)
108.60(6)
2336.8(5)
17760.5(12)
2277(5)
Z
4
4
2
density (calc) (Mg/m3)
1.855
2.342
1.801
abs. coeff. (mm−1
)
6.560
15.077
8.334
F(000)
1264
11376
1196
final R indices (R1)
reflections collected
completeness to θ
0.0587
0.0473
0.0404
36 929
188 093
31 033
99.3%
99.8%
99.2%
goodness-of-fit on F2
final R indices [I > 2σ(I)]
R indices (all data)
1.19
1.02
1.02
R1 = 0.0210, wR2 = 0.0542
R1 = 0.0216, wR2 = 0.0544
1.15 and −0.88
R1 = 0.0473, wR2 = 0.1258
R1 = 0.0677, wR2 = 0.1347
2.14 and −1.44
R1 = 0.0404, wR2 = 0.0984
R1 = 0.0566, wR2 = 0.1071
1.45 and −0.86
larg. diff. peak hole (e.Å−3
)
a
Using synchrotron radiation tuned to λ = 0.7749 Å.
Scheme 2. Preparations of (Diphosphino)alkylgold(I)
Dithiocarbamato Complexes 4−12
Figure 1. Molecular structure of 3 drawn with 30% probability
ellipsoids. H atoms are omitted for clarity. Selected bond lengths [Å]
and angles [deg]: Au1−P1, 2.2533(6); Au1−S1, 2.3272(6); P1−C1,
1.811(2); N1−C1, 1.399(3); S2−C1, 1.663(3); P1−Au1−S1,
175.36(2); C1− S1−Au1, 100.04(9); C9−P1−Au1, 118.20(8); S2−
C1−S1, 124.85(15).
(pzH)2]2ClO4 and [μ-dppmAu2(pzH)2]2ClO4 (pzH = pyr-
azole).45
Attempts to grow crystals of compound 4 were unsuccessful.
On prolonged standing, a solution of 4 in a mixture of
dichloromethane and diethylether at room temperature led to
the isolation of the chloride salt of the cationic gold(I). cluster
[Au18S8(dppe)6]2+ (4a), whose molecular structure was
established by single crystal X-ray crystallography (Table 1,
respectively; the 31P{1H} NMR spectra showed broad singlets
between 29.0 and 33.0 ppm. The spectroscopic data for 4−6
are similar to the data for the bis(diphenylphosphino)digold(I)
cations with azotate neutral heterocycles [μ-(dppe)Au2-
C
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Figure 2. Molecular structure of 4a. H atoms are omitted for clarity. Selected bond lengths [Å] and angles [deg]: Au1−P1, 2.2533(6); Au1−S2,
2.343(3); Au1−S3, 2.368(4); Au1−Au2, 2.9263(7); Au1−Au17, 2.9452(7); Au1−Au7, 3.1395(7); Au7−P10, 2.268(4); S2−Au1−S3, 175.80(12);
Au10−Au2−Au3, 77.409(18); S2−Au1−Au2, 92.05(8); S1−Au2−Au1, 109.71(8); P8−Au9−S4, 109.71(8).
Figure 2). The molecular structure of 4a contains 18 gold
atoms, 8 sulfur atoms, and 6 dppe units, with 2 chloride
counterions and some unidentified solvents of crystallization
(see Supporting Information for more details). It is
isomorphous with the Se analogue [Au18Se8(dppe)6]2+.43 The
geometry of 4a is similar to that of the selenium analogue
[Au18Se8(dppe)6]2+.43 At the core of the cluster is a Au6S2
cubane composed of atoms Au1−Au6, S1, and S2 in such a
fashion that each face contains three Au and one S atoms; the S
atoms are positioned across the body diagonal of the cubane.
The three Au atoms of each face are capped with a Au2S
triangle. Each S atom is μ3-bridging: atoms S1 and S2
coordinate to three Au atoms of the cubane, whereas the
other S atoms ligate one Au atom from the cubane and two Au
atoms from a capping Au2S triangle. Each of the Au atoms in
the capping triangles is ligated by one P atom of the six
bidentate dppe ligands. The Au−Au distances vary between
2.9147(7) and 3.2838(8) Å; the Au−S bond length falls within
a 2.328(4)−2.374(3) Å range, and the 12 Au−P distances
average 2.260(7) Å, a value similar to that in compound 3.
Other high nuclearity gold(I) sulfide complexes with bridging
diphosphines have been reported.46−48 As for the neutral
hydrated cluster [Au18S8(dppe)6]·H2O,32 its formulation
implies a mixed-valent Au0−AuI compound, contrary to our
findings and unlike the related cluster [Au18Se8(dppe)6]2+.43,48
The formation of 4a is unique because dithiocarbamate as a
sulfur source is clearly unusual in forming high-nuclearity
clusters. The formation of 4a from 4 is likely to occur via a C−S
bond cleavage, triggered by the heterocycle attached to the CS2
in the dithiocarbamate. This is supported by [Au-
(dithiocarbamate)(dppe)] complexes that are stable in solution
and have solid-state structures where the dithiocarbamate
ligands remain intact, for example, [Au2(S2CNEt2)2(dppe)],49
[Au2 (S2 CNEt2 )2 {μ-(PPh2 )2 CCH2 }],4 9 and[Au-
(S2CNC10H20O4)(dppe)] (where C10H20O4 = aza-15-crown-
5).31 The dppe backbone and aurophilicity of the two Au atoms
in 4 must have aided the cluster formation since increase in the
alkyl chain length in the diphosphine gave stable
diphosphinogold(I) dithiocarbamate complexes (vide infra).
(Diphosphino) Propyl and Hexyl Gold(I) Dithiocarba-
mato Complexes (7−12). To avoid the cluster formation
observed during crystallization of 4, the −CH2− linker count
within the diphosphines was increased to three in dppp and six
in dpph. Their respective diphosphinogold(I) dithiocarbamates
(7−12) were synthesized from the reaction between
[Au2Cl2(dppp)] or [Au2Cl2(dpph)] and 2 equiv of L1, L2,
or L3 (Scheme 2). The 1H and 31P{1H} NMR spectra of 7−12
showed similar patterns as those of 4−6, with the CH2 protons
in the backbone of the diphosphine ligands appearing upfield in
the region of 1.49−2.93 ppm and singlets for phosphorus
between 29.0 and 33.0 ppm. These complexes were stable in
solution for several weeks, with no signs of decomposition. ESI-
MS of 9 (m/z = 1190.88, Figure S1, Supporting Information)
showed molecular ions that correspond to a binuclear complex
with a molecular ion peak at m/z = 1190.88, and the crystal
structure of 12 (Figure 3) further indicates that the length of
the alkyl linker in the diphosphine ligand is important in
preventing compounds 7−12 from transforming into clusters.
Crystal data, together with the data collection and refinement
parameters, are presented in Table 1. The molecular drawing of
12 is shown in Figure 3. The dinuclear complex resides on a
crystallographic inversion center, and only one-half of it is
symmetry independent; the most important distances are
discussed. The symmetry-independent half of complex 12 can
D
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bonding distances and angles are in the same range and
correspond with the expected values. Crystal data, together
with the data collection and refinement parameters, are
presented in Table 1.
Biological Activity. Fifteen compounds (L1−L3, 1−12)
were initially screened for their ability to inhibit cell growth on
human cervical epitheloid carcinoma (HeLa) cells in vitro. All
data were acquired in triplicate, and the final values were
recorded as averages. The dose values that caused 50%
inhibition of cell growth (IC50) are listed in Table 2. To
establish the activities of phosphine gold(I) complexes it was
important to first establish the activities of the ligands (L1−L3)
to determine whether activities of the metal complexes could be
due to the presence of the dithiocarbamate ligands. All the free
ligands were inactive against HeLa cells.
The gold(I) complexes were grouped into three sets
according to ligand type, PPh3 (1−3), dppp (7−9), and
dpph (10−12), for testing. Complexes 4−6 (the dppe set)
were not tested because of the instability of 4 in solution
discussed earlier. Although complexes 1−3, 7−9, and 12 had
quite good IC50 values (2.2−7.0 μM), their tumor specificity
(TS) values were very low (Table 2), and they were less active
than cis-platin (0.476 μM). The low TS factors suggest that
these compounds were toxic to both tumor and normal cells
and were thus not further investigated. Compounds 10 and 11
had IC50 values of 0.51 μM and 0.14 μM, respectively,
compared to 0.45 μM for cis-platin. These two gold compounds
(10 and 11) displayed very good TS values of 25.0 and 70.5,
respectively (Table 2). Our findings are comparable to and in
some cases much better than activities reported for phosphine
gold(I) thiolate complexes,12,13,23 which further buttresses the
conception that a P−Au−S motif enhances the therapeutic
effect of phosphine gold(I) thiolate compounds as anticancer
agents. Complexes 10 and 11 were further investigated against
a panel of 60 cancer cell lines of the Developmental
Therapeutics Program (DTP) at the National Cancer Institute
(U.S.A.), where the concentration that inhibits growth of cells
by 50% (GI50), the concentration that causes total tumor
growth inhibition (TGI), and the least concentration required
to kill 50% of tumor cells (LC50) were determined. The 60 cell
lines were organized into subpanels, representing various
histologies, for example, nonsmall cell lung-, colon-, breast-,
ovarian-, renal-, prostrate-, and CNS cancers, leukemia, and
melanoma. Because of the extensive data, we have highlighted
only the most important findings in Tables 3 and 4, but more
data are provided as Supporting Information (Figures S2 and
S3; Tables S1 and S2). Complexes 10 and 11 were, in general,
Figure 3. Molecular structure of 12 drawn with 50% probability
ellipsoids. H atoms are omitted for clarity. Selected bond length [Å]
and angles [deg]: Au1−P1, 2.255(2); Au1−S1,2.312(3); P1−C9,
1.818(6); N1−C8, 1.390(8); S1−C1, 1.720(7); S2−C1, 1.656(6);
P1−Au1−S1, 173.41(6); C1−S1−Au1, 100.9(2); C15−P1−Au1,
112.6(8); S2−C1−S1, 124.1(4). Selected symmetry-related atoms
are labeled with a superscript.
be considered a congener of complex 3. In complex 12 the
coordination environment about the Au atoms is nearly linear,
with the P1−Au1−S1 angle spanning 173.41(6)°; the Au1−S1
(2.312(3) Å) and Au1−P1 (2.255(2) Å) distances are typical
and in good agreement with the ones observed in complex 3.
The single C1−S1 distance to the ligating atom S1 (1.720(7)
Å) is longer than the formally double C1S2 bond of 1.656(6)
Å, but both values closely match the corresponding values in
complex 3. In the lattice, pairs of clusters are packed with an
inversion center at the center of the Au−S···Au−S
parallelepiped. Within each parallelepiped, the nonbonding
Au···Au distance measures 3.986(3) Å, and the Au···S distance
is 3.598(4) Å. Both distances exceed the sum of the van der
Waals radii of the involved elements; thus, there are no
aurophilic interactions in the lattice. The structural findings for
complex 12 are similar to those reported by Uson et al.50 and
Cookson et al.51 for the gold(I) complexes [Au(PPh3)(S2C-aza-
15-crown-5)] and [Au(C5H5NS)2], respectively. The other
Table 2. Growth Inhibition Data of Complexes 1−3, 7−9, and 10−12 against HeLa Cells and Corresponding Tumor
Specificities
drug
HeLa IC50 (μM)
lymph (resting) IC50 (μM)
lymph (stimulated) IC50 (μM)
tumor specificity (TS)
PPh3 derivatives
dppp derivatives
dpph derivatives
1
2
3
7
8
9
2.56 0.12
2.63 0.10
2.56 0.17
3.07 0.48
6.96 0.51
3.38 0.30
0.51 0.08
0.14 0.01
4.04 0.49
0.45 0.09
9.05 3.50
9.53 3.18
13.98 10.50
7.05 2.02
6.13 2.92
18.48 3.88
18.89 5.66
18.23 15.90
37.74 12.60
8.55 3.37
8.63 3.77
5.08 2.48
5.02 1.72
2.94 0.91
8.16 4.10
6.68 3.40
1.38 0.39
30.25 11.06
3.4
3.5
3.7
1.9
0.7
3.9
10
25.0
70.5
8.4
11
12
cis-platin
E
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Inorganic Chemistry
Article
Figures S5 and S7, Supporting Information), which in the DTP
program indicates high activity across most of the panel of cell
lines.
Following on the in vitro results, compound 11 was further
tested in vivo in hollow fiber studies, but the activity data
suggested that the compound could not be delivered to the
targets. We are therefore investigating the use of various drug
delivery vehicles, including the use of β− and γ−cyclodextrin,
to deliver this drug in vivo.
Table 3. Selected DTP Antitumor Inhibition Results of
Complex 10
type of cancer
leukemia
panel/cell line GI50 (μM) TGI (μM) LC50 (μM)
RPMI-8226
SR
0.19
0.19
0.38
0.31
0.43
0.38
0.70
0.45
0.53
>100
>100
>100
>100
>100
>100
>100
>100
2.41
nonsmall cell lung
colon
A549/ATCC
COLO205
KM12
>100
>100
1.73
melanoma
ovarian
renal
UACC-62
OVCAR-3
RXF393
>100
14.20
CONCLUSIONS
■
We have prepared the first examples of phosphinogold(I)
dithiocarbamates derived from heterocycles. The stability of
these phosphinogold(I) dithiocarbamate complexes depends
on the nature of the phosphine ligand used. Triphenylphos-
phino and diphenylphosphinoalkyl ligands with alkyl chains
longer than ethyl produce stable gold dithocarbamates in
solution, but the diphenylphosphinoethanegold(I) dithiocarba-
mates are unstable and were found to transform to a Au18
cluster. All the phosphinogold(I) dithiocarbamates that are
stable in solution are active against HeLa cancer cells,
suggesting the importance of the P−Au−S moiety in conferring
activity to the compounds. Compounds with hexyl chain were
found to be the most active and extremely selective; in
particular compounds 10 and 11 were 25.0 and 70.5 times
more selective for HeLa cells than normal cells. Bis-
(diphosphines) alkanes with longer CH2 linkers appear to
hold better promise as anticancer agents than their shorter
CH2-linker counterparts, similar to the observation by Mirabeli
et al.27 We also found compounds 10 and 11 to have excellent
activities for nine other cancer cell lines in vitro. Of the nine
cancer cell lines tested, the best activity against RPMI-8226 was
found for 10 (GI50 = 0.19 μM), while the best activity against
SR cells (GI50 = 0.03 μM) was for 11. Although activities for 10
and 11 in vivo were not so great, we believe finding drug
delivery vehicles to transport these two compounds would
improve their activities in vivo. We are therefore investigating
the use of various delivery vehicles for these compounds.
Table 4. Selected DTP Antitumor Inhibition Results of
Complex 11
type of cancer
leukemia
panel/cell line GI50 (μM) TGI (μM) LC50 (μM)
RPMI-8226
SR
0.14
0.03
0.35
0.11
0.25
0.25
0.60
0.38
0.41
0.52
>100
>100
90
nonsmall cell lung
colon
A549/ATCC
COLO205
HCC-2998
UACC-62
OVCAR-8
A498
>100
>100
2.99
0.80
melanoma
ovarian
renal
>100
>100
>100
>100
active against all of the 60 cell lines in the subpanels mentioned
above, with GI50 values in the range of 0.03−100 μM.
Compound 10 showed good activity against all cell lines
tested but was particularly highly active against the leukemia
RPMI-8226 and SR cell lines, with a GI50 value of 0.19 μM for
each cell line and TGI values of 0.53 and 2.41 μM for these cell
lines, respectively (Table 3). This indicates the inhibitory
property of 10 at very low molar concentrations. However,
what was interesting is that the LC50 for 10 was greater than
100 μM for most cell lines, suggesting minimal cytotoxic
properties of 10. It, therefore, implies that this complex is
cytostatic. Similar results have been reported in literature,
where a compound is cystostatic/antiproliferative and not
necessarily cytotoxic.52
We observed a similar activity pattern for complex 11 against
all the cell lines, but 11 was even more potent than 10. For
instance, the GI50 values of 11 against the leukemia cell lines
RPMI-8226 and SR were 0.14 μM and 0.03 μM, respectively. In
fact, the best activity registered for all the compounds tested
was that of 11 against the SR cell line (0.03 μM) (Table 4).
The TGI concentrations recorded were similarly quite low,
RPMI-8226 (0.41 μM) and SR (0.52 μM), which corroborates
the cytostatic properties of 11. Of interest is that compound 11,
unlike 10, showed very good cytotoxic activity against the
HCC-2998 cell line (LC50 = 2.99 μM). Compound 11 also
showed activity against the colon cancer COLO205 and HCC-
2998 cell lines, with GI50 of 0.11 μM and 0.25 μM, respectively.
In general, complexes 10 and 11 recorded high cytostatic
median values (GI50 = −6.25; TGI = −4.51) and cytotoxic
median value (LC50 = −4.05) against the 60 cell lines (Table 5,
EXPERIMENTAL SECTION
■
Materials and Instrumentation. All manipulations were
performed under a dry, deoxygenated nitrogen atmosphere using
Schlenk techniques. All commercially available chemicals were used as
received. Pyrazol-1-yldithiocarbamate (L1), 3,5-dimethylpyrazol-1-
yldithiocarbamate (L2), and indazol-1-yldithiocarbamate (L3) were
synthesized according to literature methods.53,54 Gold starting
materials [Au2Cl2(dppe)], [Au2Cl2(dppp)], and [Au2Cl2(dpph)]
{dppe = 1,2-bis(diphenyl-phosphino)ethane; dppp = 1,3-bis-
(diphenylphosphino)propane; and dpph = 1,6-bis(diphenyl-
phosphino)hexane} were synthesized according to the literature
procedures.55 Infrared (IR) spectra were recorded as KBr pellets on
a Bruker Tensor27 spectrophotometer. 1H, 13C{1H}, and 31P{1H}
NMR spectra were recorded on a Varian 2000 spectrometer (1H, 300
MHz; 13C, 75.4 MHz; and 31P, 121.5 MHz) in CDCl3 or D2O at room
temperature. Elemental analysis was performed on a Fisons elemental
analyzer at the University of Cape Town, South Africa. ESI-MS spectra
Table 5. Cytostatic and Cytotoxic Median Data of 10 and 11
GI50
TGI
LC50
drug
MG-MID
Δ
range
MG-MID
Δ
range
MG-MID
Δ
range
10
11
−5.93
−6.25
0.8
2.73
1.96
−4.36
−4.51
1.92
1.87
2.28
2.38
−4.02
−4.05
0.69
1.47
0.71
1.52
1.22
F
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Inorganic Chemistry
Article
were recorded on a Waters API Quattro Micro spectrometer at the
University of Stellenbosch, South Africa. The mass spectra were
collected using 3.0 s cyclical scans and applying the sample cone
voltage of 15 V at the source block temperature of 100 °C.
Desolvation temperature was 350 °C at desolvation cone gas flow rate
of 350 L/h.
Anal. Calc. for C26H20AuCl2N2PS2: C 29.71, H 2.32, N 4.10. Found: C
29.80, H 2.00, N 3.80%.
Compounds 5−12 were prepared using the procedure described for
4 above, using the appropriate starting materials.
Bis-(3,5-dimethylpyrazolyl-1-dithiocarbamato)-bis-
(diphenylphosphino)ethane Dinuclear Gold(I) (5). L2 (0.04 g, 0.19
mmol), [Au2Cl2(dppe)] (0.1 g, 0.09 mmol). Yield = 0.09 g (88%). 1H
NMR (CDCl3): δ 7.64 (m, 8H, Ph2P(CH2)2PPh2); 7.35 (m, 12H,
Ph2P(CH2)2PPh2); 6.09 (s, 2H, 4-pz); 2.69 (bs, 4H, Ph2P-
(CH2)2PPh2); 2.26 (d, 12H, CH3, 3,5-pz). 13C{1H} NMR (CDCl3):
δ 216.5 (C(CS)); 152.7 (C(5-pz); 143.1 (C(3-pz)); 135.1−128.6
(phenyl region); 112.6 (C(4-pz)); 14.0 (C(5-pPz); 12.4 (C(3-pz));
Synthesis of Triphenylphosphinegold(I) Complexes. Pyra-
zolyl-1-dithiocarbamato-triphenylphosphinogold(I) (1). Complex 1
was prepared by dissolving L1 (0.05 g, 0.3 mmol) in deionized water
(10 mL), after which a solution of [AuCl(PPh3)] (0.15 g, 0.3 mmol)
in dichloromethane (10 mL) was added. The resultant biphasic
mixture was vigorously stirred at room temperature for 30 min, during
which time the color of the organic layer changed from colorless to
red. The aqueous and organic layers were separated, and the organic
layer was dried over anhydrous MgSO4. The solvent was removed
from the organic extract in vacuo to afford an orange-yellow solid.
Yield = 0.17 g (93%). 1H NMR (CDCl3): δ 7.63 (s, 1H, 5-pz); 7.57 (s,
1H, 3-pz); 7.49 (m, 9H, PPh3); 7.35 (m, 6H, PPh3); 6.27 (s, 1H, 4-
pz). 13C{1H} NMR (CDCl3): δ 213.4 (C(CS)); 144.0 (C(5-pz);
134.2−129.1 (phenyl region); 140.8 (C(3-pz)); 106.1 (C(4-pz)). IR
(KBr, cm−1): υCN = 1620, υC−S = 1121, υCS = 884. 31P{1H} NMR
(CDCl3): δ 35.2 (PPh3). Anal. Calc. for C24H22AuN2PS2: C 43.86, H
3.01, N 4.65. Found: C 43.53, H 3.05, N 4.62%.
25.7 (C(Ph2P(CH2)2PPh2). IR (KBr, cm−1): υCN = 1603, υC−S
=
1263, υCS = 989. 31P{1H} NMR: (CDCl3): δ 35.7 (Ph2P-
(CH2)2PPh2). Anal. Calc. for C38H38Au2N4P2S4: C 40.22, H 3.37, N
4.94, S 11.30. Found: C 39.89, H 3.59, N 4.67, S 11.15%.
Bis-(indazolyl-1-dithiocarbamato)-bis-(diphenylphosphino)-
ethane Dinuclear Gold(I) (6). L3 (0.04 g, 0.19 mmol),
[Au2Cl2(dppe)] (0.08 g, 0.09 mmol). Yield = 0.07 g (63%). 1H
NMR (CDCl3): δ 9.16 (s, 1H, Ha); 8.06 (s, 1H, Hb); 7.78 (m, 8H,
(Ph)2P(CH2)2P(Ph)2); 7.49 (m, 12H, (Ph)2P(CH2)2P(Ph)2); 7.67 (d,
3
4
1H, JHH = 8.4 Hz, He); 7.71 (1H, Hd); 7.43 (t, 1H, JHH = 15 Hz,
Hc); 2.87 (bs, 4H, CH2, (Ph)2P(CH2)2P(Ph)2). 13C{1H} NMR
(CDCl2): δ 213.9 (C(CS)); 142.1 (C(5-Pz); 139.0 (C(3-Pz));
134.1−129.7 (phenyl region); 126.1 (C(7C-Ph)); 123.1 (C(4C-Ph));
122.2 (C(8C-Ph, 9C-Ph)); 117.5 (C(6C-Ph) 25.2 (C(Ph2P-
Compounds 2 and 3 were prepared using the procedure described
for 1 above, using the reagents indicated for each compound.
3 , 5 - d i m e t h y l p y r a z o l y l - 1 - d i t h i o c a r b a m a t o -
triphenylphosphinogold(I) (2). L2 (0.1g, 0.47 mmol), [AuCl(PPh3)]
(0.24 g, 0.47 mmol). Yield = 0.17 g (57%). 1H NMR (CDCl3): δ 7.44,
7.29 (m, 15H, PPh3); 6.12, 5.82 (s, 1H, 4-pz); 2.29 (m, 6H, 3, 5-pz).
13C{1H} NMR (CDCl3): δ 215.5 (C(CS)); 149.7 (C(5-pz); 134.1−
(CH2)2PPh2). IR (KBr, cm−1): υCN = 1612, υC−S = 1137, υCS
=
915. 31P{1H} NMR: (CDCl3): δ 34.9 (Ph2P(CH2)2PPh2). Anal. Calc.
for C42H34Au2N4P2S4: C 42.79, H 2.91, N 4.75, S 10.88. Found: C
43.24, H 2.82, N 4.65, S 10.57%.
Bis(pyrazol-1-yldithiocarbamato)-bis-(diphenylphosphino)-
propane Dinuclear Gold(I) (7). L1 (0.08 g, 0.46 mmol),
[Au2Cl2(dppp)] (0.2 g, 0.23 mmol). Yield = 0.15 g (59%). 1H
128.7 (phenyl region); 142.1 (C(3-pz)); 112.4, 104.1 (C(5-pz)); 13.9
(C(CH3, 5-pz); 12.31 (C(CH3, 3-pz). IR (KBr, cm−1): υCN = 1580,
υC−S = 1154, υCS = 854. 31P{1H} NMR (CDCl3): δ 33.8 (PPh3).
Anal. Calc. for C24H22AuN2PS2: C 45.72, H 3.52, N 4.44. Found: C
45.89, H 3.89, N 3.29%.
3
NMR (CDCl3): δ 8.73 (s, 2H, JHH = 2.7 Hz, 5-pz); 7.73 (m, 8H,
3
Ph2P(CH2)3PPh2); 7.51 (s, 4H, JHH = 1.8 Hz, 3-pz); 7.41 (m, 12H,
3
Ph2P(CH2)3PPh2); 6.41 (s, 2H, JHH = 2.7 Hz, 4-pz); 2.93 (m, 2H,
Indazolyl-1-dithiocarbamato-triphenylphosphinogold(I) (3). L3
(0.1 g, 0.4 mmol), [AuCl(PPh3)] (0.2 g, 0.4 mmol). Yield = 0.15 g
(58%). 1H NMR (CDCl3): δ 9.21 (d, 1H, 4JHH = 9.0 Hz, Ha); 8.17 (s,
1H, Hb); 7.73 (d, 1H, 3JHH = 8.4 Hz, He); 7.61, 7.50 (m, 15H, PPh3);
Ph2PCH2CH2CH2PPh2); 1.99 (m, 4H, Ph2PCH2CH2CH2PPh2).
13C{1H} NMR (CDCl3): δ 218.9 (C(CS)); 147.5 (C(5-pz);
140.8 (C(3-pz)); 133.2−129.2 (phenyl region); 106.2 (C(4-pz)); 28.5
1
(2C, JC−P = 62.7 Hz, P−CH2, Ph2PCH2CH2CH2PPh2); 21.4 (C,
4
(7.58 (1H, Hd); 7.35 (t, 1H, JHH = 15 Hz, Hc). 13C{1H} NMR
Ph2PCH2CH2CH2PPh2). IR (KBr, cm−1): υCN = 1628, υC−S = 1088,
υCS = 891. 31P{1H} NMR (CDCl3): δ 32.8. Anal. Calc. for
C35H32Au2N4P2S4: C 38.47, H 2.95, N 5.13, S 11.74. Found: C 38.99,
H 3.08, N 5.09, S 11.45%.
(CDCl3): δ 214.2 (C(CS)); 139.5 (C(5C-pz); 134.7−129.5
(phenyl region); 129.5 (C(3C-pz)); 126.9 (C(7C-Ph)); 124.6
(C(4C-Ph); 121.4 (C(8C, 9C-Ph)); 118.7 (C(6C-Ph)). IR (KBr,
cm−1): υCN = 1601, υC−S = 1148, υCS = 905. 31P{1H} NMR
(CDCl3): δ 36.8 (PPh3). Anal. Calc. for C26H20AuN2PS2: C 47.86, H
3.09, N 4.29. Found: C 47.41, H 3.04, N 3.85%.
Bis(3,5-dimethylpyrazol-1-yldithiocarbamato)-bis-
(diphenylphosphino)propane Dinuclear Gold(I) (8). L2 (0.1 g, 0.46
mmol), [Au2Cl2(dppp)] (0.2 g, 0.23 mmol). Yield = 0.13 g (50%). 1H
NMR (CDCl3): δ 7.66 (m, 8H, Ph2P(CH2)3PPh2); 7.34 (m, 12H,
Ph2P(CH2)3PPh2); 5.85 (s, 2H, 4-pz); 2.91 (m, 2H, Ph2PCH2-
CH2CH2PPh2); 1.88 (m, 4H, Ph2PCH2CH2CH2PPh2); 2.24, 2.26 (d,
12H, CH3, 3,5-pz). 13C{1H} NMR (CDCl3): δ 215.9 (C(CS));
153.3 (C(5-pz); 144.2 (C(3-pz)); 133.4−129.2 (phenyl region); 112.5
Synthesis of Dinuclear Di(phosphino)alkylgold(I) Com-
p l e x e s . B i s - ( p y r a z o l y l - 1 - d i t h i o c a r b a m a t o ) - b i s -
(diphenylphosphino)ethane Dinuclear Gold(I) (4) and
[Au18S8(dppe)6]Cl2 (4a). To a solution of [Au2Cl2(dppe)] (0.08 g,
0.09 mmol) in CH2Cl2 (10 mL) was added L1 (0.03 g, 0.19 mmol)
previously dissolved in water (10 mL). The resultant mixture was
stirred at room temperature for 20 min during which time the CH2Cl2
layer changed color from a white suspension to a clear orange solution,
while the yellowish water layer became clear. The two layers were
separated and the organic fraction dried over anhydrous MgSO4. The
organic fraction was evaporated in vacuo to obtain 4 as an orange
1
(C(4-pz)); 27.9 (2C, JC−P = 58.4 Hz, P−CH2, Ph2PCH2-
CH2CH2PPh2); 20.2 (C, Ph2PCH2CH2CH2PPh2); 13.8 (C(CH3, 5-
pz); 13.2 (C(CH3, 3-pz). IR (KBr, cm−1): υCN = 1653, υC−S = 1099,
υCS = 957. 31P{1H} NMR (CDCl3): δ 29.2. Anal. Calc. for
C39H40Au2N4P2S4: C 40.77, H 3.51, N 4.88, S 11.16. Found: C 40.99,
H 3.74, N 4.79, S 11.10%.
solid. Yield = 0.07 g (73%). 1H NMR (CDCl3): δ 8.78 (d, 2H, 3JHH
=
Bis(indazol-1-yldithiocarbamato)-bis-(diphenylphosphino)-
propane Dinuclear Gold(I) (9). L3 (0.04 g, 0.22 mmol),
[Au2Cl2(dppp)] (0.1 g, 0.11 mmol). Yield = 0.07 g (54%). 1H
NMR (CDCl3): δ 9.13 (s, 1H, Ha); 8.16 (s, 1H, Hb); 7.73 (1H, Hd);
7.69 (d, 1H, 3JHH = 8.0 Hz, He); 7.66 (m, 8H, Ph2P(CH2)3PPh2); 7.45
2.7 Hz, 5-pz); 7.80 (s, 2H, 3-pz); 7.78 (m, 8H, (Ph)2P(CH2)2P(Ph)2);
7.49 (m, 12H, (Ph)2P(CH2)2P(Ph)2); 6.43 (s, 2H, 4-pz); 2.87 (bs, 4H,
CH2, (Ph)2P(CH2)2P(Ph)2). 13C{1H} NMR (CDCl3): δ 214.2
(C(CS)); 149.7 (C(5-pz); 142.0 (C(3-pz)); 134.1−128.7 (phenyl
region); 111.4 (C(4-pz)); 24.6 (C(Ph2P(CH2)2PPh2). IR (KBr,
cm−1): υCN = 1590, υC−S = 1190, υCS = 889. 31P{1H} NMR:
(CDCl3): δ 36.1 (Ph2P(CH2)2PPh2). Anal. Calc. for C34H30Au2-
N4P2S4: C 37.85, H 2.80, N 5.14, S 11.89. Found: C 38.18, H 3.01, N
4.57, S 11.73%.
4
(t, 1H, JHH = 16.8 Hz, Hc); 7.42 (m, 12H,Ph2P(CH2)3PPh2); 2.89
(m, 2H, Ph2PCH2CH2CH2PPh2); 1.92 (m, 4H, Ph2PCH2-
CH2CH2PPh2). 13C{1H} NMR (CDCl3): δ 213.8 (C(CS)); 140.3
(C(5C-pz); 139.1 (C(3C-pz)); 133.5−129.2 (phenyl region); 127.9
(C(7C-Ph)); 124.2 (C(4C-Ph)); 121.1 (C(8C-Ph, 9C-Ph)); 118.3
1
Attempts to crystallize 4a in a solution of dichloromethane/ether
led to the formation of the gold cluster, [Au18S8(dppe)6]Cl2, (4a).
(C(6C-Ph)); 28.6 (2C, JC−P = 64.2 Hz, P−CH2, Ph2PCH2-
CH2CH2PPh2); 19.8 (Ph2PCH2CH2CH2PPh2). IR (KBr, cm−1):
G
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Inorganic Chemistry
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υCN = 1606, υC−S = 1150, υCS = 993. 31P{1H} NMR (CDCl3): δ
29.2. ESI-MS: m/z 1190.9 [Au2(L3)2(dppp)]+ (5%). Anal. Calc. for
C43H36Au2N4P2S4: C 43.29, H 3.04, N 4.70, S 10.75. Found: C 43.23,
H 3.05, N 4.62, S 10.56%.
calculation at idealized positions and were allowed to ride on the
neighboring atoms with relative isotropic displacement coefficients.57
TESTING OF COMPOUNDS FOR ANTICANCER
ACTIVITY
■
Synthesis of Dinuclear Bis(pyrazol-1-yldithiocarbamato)-bis-
(diphenylphosphino)hexane Gold(I) (10). L1 (0.02 g, 0.11 mmol),
[Au2Cl2(dpph)] (0.05 g, 0.05 mmol). Yield = 0.04 g (70%). 1H NMR
(CDCl3): δ 8.78 (d, 2H, 3JHH = 2.7 Hz, 5-pz); 7.80 (s, 2H, 3-pz); 7.78
(m, 8H, (Ph)2P(CH2)6P(Ph)2); 7.49 (m, 12H, (Ph)2P(CH2)6P(Ph)2);
6.43 (s, 2H, 4-pz); 2.87 (bs, 4H, Ph2P(CH2)2(CH2)2(CH2)2PPh2);
1.49 (bs, 8H, Ph2P(CH2)2(CH2)2(CH2)2PPh2). 13C{1H} NMR
(CDCl3): δ 213.8 (C(CS)); 140.3 (C(5C-pz); 139.1 (C(3C-pz));
133.5−129.2 (phenyl region); 127.9 (C(7C-Ph)); 124.2 (C(4C-Ph));
121.1 (C(8C-Ph, 9C-Ph)); 118.3 (C(6C-Ph)); 30.1 (4C,
PPh2(CH2)2(CH2)2(CH2)2PPh2); 25.9 (2C, Ph2P(CH2)2-
(CH2)2(CH2)2PPh2). IR (KBr, cm−1): υCN = 1620, υC−S = 1108,
υCS = 960. 31P{1H} NMR (CDCl3): δ 32.3. Anal. Calc. for
C38H38Au2N4P2S4: C 40.22, H 3.37, N 4.94, S 11.30. Found: C 40.17,
H 3.45, N 4.36, S 10.28%.
Biological Reagents and Instrumentation. All commer-
cial reagents were used as received. Phosphate-buffered saline
(PBS), Eagle’s RPMI-1640 medium, 3-(4,5-dimethylthiazol-2-
yl)-2,5-diphenyltetrazolium bromide (MTT) kit, phytohemag-
glutinin-protein form (PHA-P) and the 96-well flat-bottomed
culture plates were all purchased from BD Biosciences Ltd. Cis-
platin was purchased from Sigma Aldrich. Eagle’s medium with
0.1 mM nonessential amino acids was prepared by adding 2
mM L-glutamine, 1.0 mM sodium pyruvate, and 5% bovine fetal
calf serum to the pure Eagle’s RPMI-1640 medium. A total of
fifteen compounds (L1−L3, 1−3, and 7−12) were screened for
their anticancer activities. Human cervix epithelial carcinoma
(HeLa) cells and human lymphocytes (PBMCs), from
preservative-free heparinised peripheral blood, were obtained
from the Department of Pharmacology and Pretoria Medical
Hospital, University of Pretoria, South Africa. The absorbance
values were recorded on a Whittaker Microplate Reader 2001
spectrophotometer at 570 nm and the reference wavelength of
630 nm.
Two complexes (10 and 11) were further tested against the
60-cell-line panel at the DTP using their internal proce-
dures.58,59 From this study, three important (cytostatic and
cytotoxic) properties are evident, namely, cytostatic values that
include molar concentrations of drug required for 50% growth
inhibition (GI50), cytostatic values that include molar
concentrations of drug required for total growth inhibition
(TGI), and cytostatic values that include molar concentrations
of drug required to kill 50% of the cell population (LC50).
Cell Culture and Drug Treatment. HeLa cells were
cultured in Eagle’s medium with 0.1 mM nonessential amino
acids, 2 mM L-glutamine, 1.0 mM sodium pyruvate, and 5%
bovine fetal calf serum at 37 °C in an atmosphere of 5% CO2.
Cells were placed in 96-well sterile plates, at a density of 1 ×
104 cells/well in 100 μL of medium, and incubated for 1 h.
Subsequently, the ligands or gold compounds were added, with
concentrations ranging from 0 to 100 μM. Cytotoxicity was
determined by using MTT to stain treated HeLa cells after 7 d,
according to literature methods.60 MTT dye is reduced by
living cells to yield a soluble formazan product that can be
assayed colorimetrically.60 A 20 μL volume of freshly prepared
MTT (5 mg/mL) was added to each well, and the cells were
incubated for another 4 h. Cell survival was evaluated by
measuring absorbance at 570 nm, using a Whittaker Microplate
Reader 2001. All experiments were performed in triplicate.
The inhibition of the growth of normal cells by the
complexes tested was also measured by employing human
lymphocytes (PBMC) cells. The same procedure described
above was used, except that the treated PBMC cells were
incubated for 3 d as opposed to 7 d for HeLa cells. The aim of
testing these compounds on normal cells was to determine
whether the compounds could target the cancerous (HeLa)
cells specifically and not the normal cells. Lymphocytes were
divided into two groups, namely, (i) normal cells that were
stimulated using PHA-P so as to increase their proliferation rate
(stimulated lymphocytes) and (ii) unstimulated normal cells
(resting lymphocytes).
Synthesis of Dinuclear Bis(3,5-dimethylpyrazol-1-yldithiocarba-
mato)-bis-(diphenylphosphino)hexane Gold(I) (11). L2 (0.09 g, 0.42
1
mmol), [Au2Cl2(dpph)] (0.2 g, 0.2 mmol). Yield = 0.12 g (50%). H
NMR (CDCl3): δ 7.67 (m, 8H, Ph2P(CH2)6PPh2); 7.40 (m, 12H,
Ph2P(CH2)6PPh2); 6.00 (s, 2H, 4-pz); 2.76 (bs, 4H, Ph2P(CH2)2-
(CH2)2(CH2)2PPh2); 2.18, 2.27 (d, 12H, CH3, 3,5-pz); 1.60, 1.52 (bs,
8H, Ph2P(CH2)2(CH2)2(CH2)2PPh2). 13C{1H} NMR (CDCl3): δ
215.9 (C(CS)); 153.3 (C(5-pz); 144.2 (C(3-pz)); 133.4−129.2
(phenyl region); 112.5 (C(4-pz)); 31.2 (4C, PPh2(CH2)2-
(CH2)2(CH2)2PPh2); 26.7 (2C, Ph2P(CH2)2(CH2)2(CH2)2PPh2);
13.8 (C(5-pz); 13.2 (C(3-pz). IR (KBr, cm−1): υCN = 1653,
υC−S = 1099, υCS = 957. 31P{1H} NMR (CDCl3): δ 32.7. Anal.
Calc. for C42H46Au2N4P2S4: C 42.36, H 3.89, N 4.70, S 10.77. Found:
C 42.12, H 3.84, N 4.62, S 10.56%.
Synthesis of Dinuclear Bis(indazol-1-yldithiocarbamato)-bis-
(diphenylphosphino)hexane Gold(I) (12). L3 (0.1 g, 0.44 mmol),
1
[Au2Cl2(dpph)] (0.2 g, 0.22 mmol). Yield = 0.14 g (52%). H NMR
(CDCl3): δ 9.18 (d, 1H, 4JHH = 8.4 Hz, (Ha)); 8.10 (s, 1H, Hb); 7.49
(d, 1H, 3JHH = 8.7 Hz, He); 7.24 (t, 1H, 4JHH = 15.0 Hz, Hd); 7.32 (t,
4
1H, JHH = 15.0 Hz, Hc); 7.66 (m, 8H, Ph2P(CH2)6PPh2); 7.42 (m,
12H, Ph2 P(CH2 )6 PPh2 ); 2.39 (m, 4H, Ph2 P(CH2 )2 -
(CH2)2(CH2)2PPh2); 1.57 (m, 8H, Ph2P(CH2)2(CH2)2(CH2)2PPh2).
13C{1H} NMR (CDCl3): δ 213.8 (C(CS)); 140.3 (C(5C-pz); 139.1
(C(3C-pz)); 133.5−129.2 (phenyl region); 127.9 (C(7C-Ph)); 124.2
(C(4C-Ph)); 121.1 (C(8C-Ph, 9C-Ph)); 118.3 (C(6C-Ph)); 28.9 (4C,
PPh2(CH2)2(CH2)2(CH2)2PPh2); 25.6 (2C, Ph2P(CH2)2-
(CH2)2(CH2)2PPh2). IR (KBr, cm−1): υCN = 1657, υC−S =
1099, υCS = 994. 31P{1H} NMR CDCl3): δ 31.8. Anal. Calc. for
C46H42Au2N4P2S4: C 44.74, H 3.43, N 4.54, S 10.39. Found: C 44.37,
H 3.46, N 4.50, S 10.32%.
Crystal Structure Determination. Crystals of 3 and 4a were
mounted in oil on a glass fiber, and data collection was performed on a
Bruker CCD-1000 diffractometer with Mo Kα (λ = 0.71073 Å)
radiation at a diffractometer-to-crystal distance of 4.9 cm. The
reflections were successfully indexed by an automated indexing
routine built in the SMART program. The data were corrected for
Lorentz and polarization effects. Intensity data for 12 were collected at
150 K on a D8 goniostat equipped with a Bruker APEXII CCD
detector at beamline 11.3.1 at the Advanced Light Source (Lawrence
Berkeley National Laboratory) using synchrotron radiation tuned to λ
= 0.7749 Å. The data frames were collected using the program APEX2
and processed using the program SAINT routine within APEX2.
Option SQUEEZE of program PLATON56 was used to correct the
diffraction data for diffuse scattering effects and to identify the solvate
molecule.
The absorption correction for the all three complexes was based on
fitting a function to the empirical transmission surface as sampled by
multiple equivalent measurements. A successful solution by the direct
methods provided all non-hydrogen atoms from the E-map. All non-
hydrogen atoms were refined with anisotropic displacement
coefficients. All hydrogen atoms were included in the structure factor
H
dx.doi.org/10.1021/ic4025926 | Inorg. Chem. XXXX, XXX, XXX−XXX

Inorganic Chemistry
Article
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ASSOCIATED CONTENT
* Supporting Information
■
S
Crystallographic information data (CIF); ESI-MS of 9;
biological data for compounds 11 and 12. This material is
available free of charge via the Internet at http://pubs.acs.org.
The deposition numbers CCDC 680273, 949434, and 920397
contain the supplementary crystallographic data for compounds
3, 4a, and 12, respectively. These data can be obtained free of
charge from The Cambridge Crystallographic Data Centre via
www.ccdc.cam.ac.uk/data_request/cif.
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AUTHOR INFORMATION
Corresponding Author
■
*Email: jdarkwa@uj.ac.za.
Present Address
́ ́
(20) Urig, S.; Fritz-Wolf, K.; Reau, R.; Herold-Monde, C.; Toth, K.;
¶School of Chemistry and Physics, University of KwaZulu-
Natal, Westville Campus, Private Bag X54001, Durban, 4000,
South Africa.
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Gonzalez, A.; Sordo, J.; Varela, J. M.; Lopez, E. M. V. J. Inorg. Biochem.
2008, 102, 184.
Notes
The authors declare no competing financial interest.
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A.; Sordo, J.; Taboada, C. J. Inorg. Biochem. 2006, 100, 1858.
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ACKNOWLEDGMENTS
■
We acknowledge financial support for this work from Harmony
Gold under project AuTEK, run by Mintek (South Africa), and
University of Johannesburg. Crystallographic data were
collected through the SCrALS (Service Crystallography at
Advanced Light Source) program at the Small-Crystal
Crystallography Beamline 11.3.1 at the Advanced Light Source
(ALS), Lawrence Berkeley National Laboratory. The ALS is
supported by the U.S. Department of Energy, Office of Energy
Sciences Materials Sciences Division, under contract DE-AC02-
05CH11231. The authors would also like to thank the National
Cancer Institute (U.S.A.) for the anticancer data generated
under their DTP program and Dr. Frederik H. Kriel of Mintek
who arranged for samples to be tested at the NCI.
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