Highly antiproliferative neutral Ru(ii)-arene phosphine complexes

Article abstract of DOI:10.1039/c7nj04442a

Six ruthenium(ii)- and four gold(i)-phosphine based complexes were synthesized and fully characterized. Some of them displayed strong antiproliferative properties for several types of cancer including colon, breast, and lung. Notably, two of the Ru(ii) complexes displayed an IC₅₀ of around 2 μM, which is exceptional for these types of complexes. The dramatic impact of the nature of the arene coordinated on the ruthenium center was clearly evidenced.

Full text of DOI:10.1039/c7nj04442a

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ISSN 1144-0546
PAPER
Jason B. Benedict et al.
The role of atropisomers on the photo-reactivity and fatigue of
diarylethene-based metal–organic frameworks
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Highly antiproliferative neutral Ru(II)-arene phosphine complexes
Florian Chotard,a Lucile Dondaine,a,b Cédric Balan,a Ali Bettaïeb,b Catherine Paul,b Pierre Le
Gendrea and Ewen Bodioa*
Received 00th January 20xx,
Accepted 00th January 20xx
Six ruthenium(II)- and four gold(I)-phosphine based complexes have been synthesized and fully characterized. Some of
them display strong antiproliferative properties on several types of cancer including colon, breast, and lung. Notably, two
of Ru(II) complexes display IC50 around 2 µM, which is exceptional for these kinds of complexes. The dramatic impact of
DOI: 10.1039/x0xx00000x
www.rsc.org/
the
nature
of
the
arene
coordinated
on
ruthenium
center
was
clearly
evidenced.
by synthesizing auranofin analogues – auranofin is a gold(I)
complex used initially as an antirheumatic agent, which is
under current phase 2 clinical tests to fight against specific
types of leukemia and non-small cell lung cancer – (Figure 1).
Introduction
These last decades, numerous researchers have looked for
alternative to cisplatin and its analogues for treating cancer.
They have one aim in mind: design a compound more efficient
and displaying less side effects than platinum derivatives.
Among the different strategies explored, the use of other
metals such as titanium, gold, ruthenium… has given promising
results.1–15 During the last ten years, we have chosen to
conceive heterobimetallic complexes to combine their
biological properties.16–19 In a study dealing with the design of
Figure 2: structures of the Ru(II)-based complexes investigated in this study.
Ti-Ru derivatives, we synthesized
a
diene-phosphine
ruthenium complex as a control (Figure 1).16 Surprisingly, this
complex displayed strong antiproliferative properties on some
cancer cell lines.
Concerning, Ru(II)-based-arene-phosphine complexes, another
point is still under debate: the impact of coordinated arene.
Indeed, some studies indicate a low impact of arene, while
other articles reported that substitution of arene by lipophilic
groupement increases significantly the antiproliferative
properties.22 In the present study, we decided to evaluate the
effect of the substitution of the arene by an electron-
withdrawing group.
Figure 1: structure of the diene-phosphine ruthenium complex synthesized previously
as a control compound and of auranofin.
Results and discussion
Synthesis of complexes.
Most of reported Ru(II)-based-arene-phosphine complexes
derivatives display poor cytotoxicity in vitro. That is why such a
result was unexpected. A possible explanation relies on some
Synthesis of the Ru(II)-based-arene-phosphine complexes is
divided in three part: i- synthesis of phosphine derivatives; ii-
preparation of ruthenium precursors; iii- final complexation.
studies, which claim that
a prenyl group can vectorise
compounds and facilitate their internalization in cells.20,21 R-2a
does not display a true prenyl moiety, but the butadiene group
may lead to the same kind of properties. Thus, we decided to
investigate the impact of this diene pattern by synthesizing its
saturated and phenyl analogues (Figure 2). We take advantage
of this study to investigate the impact of these different
phosphines on the corresponding gold(I) complexes, especially
Phosphine
reported:23 4-methylenehex-5-enol
isoprene by Brandsma’s procedure,24 activated thanks to
mesyl chloride, and reacted with lithium diphenylphosphide.
can be synthesized at gram scale. One additional step is
needed for the synthesis of “saturated” phosphine . Indeed,
3
is obtained following a procedure we recently
1
is obtained from
3
7
before reacting with mesyl chloride, 4-methylenehex-5-enol
is hydrogenated.
1
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Scheme 1: synthetic paths of the two phosphine ligands 3 and 7.
Gold(I) complexes are synthesized by a similar strategy. The
reaction of the two phosphines and with [AuCl(tht)] –
synthesized by reduction of HAuCl4 by THT
(tetrahydrothiophene) –27 leads to the two chlorido-gold
complexes A-1a and A-2a. The two auranofin analogues A-1b
and A-2b can be obtained by ligand exchanges. Indeed,
thioglucose tetraacetate (thiolate form) readily substitutes
chlorido ligand. The ligand exchange can be checked by Far-IR
spectroscopy (ꢀŬAu-Cl ≈ 320 cm-1 and ꢀŬAu-S ≈ 370 cm-1).
Moreover, due to purification issue, the air-sensitive
3
7
phosphine
7 have to be protected by BH3 to be isolated, and
then can be quantitavely deprotected in presence of a slight
excess of DABCO (1,4-diazabicyclo[2.2.2]octane) and used
directly in complexation reactions.
Ruthenium dimers [RuCl2(η6-arene)]2 (arene
BzOEt) are synthesized by reacting RuCl3.nH2O with α-
phellandrene25 – commercially available
or ethyl
1,4 cyclohexadiene-3-carboxylate26
synthesized by Birch
= p-cymene,
–
–
Inexplicably, complex A-2a quickly degrades, its colour turns
reaction starting from ethyl benzoate – in ethanol at reflux.
from white to violet – typical colour of gold nanoparticles –,
while A-1a, A-1b, and A-2b are totally air-stable. The same
[RuCl2(κP-phosphine)(η6-arene)] complexes are obtained by
behaviour is observed when A-2a is stored at -10°C in
glovebox. Consequently, we decided to remove this complex
from the biological study.
reacting 7, 3, or triphenylphosphine with ruthenium dimers.
The reaction can be monitored by 31P-NMR thanks to the
significant shift of the resonance observed going from the free
phosphine (singlet between -16 and -6 ppm) to the complexes
(singlets between 23 and 27 ppm). Noteworthy, no trace of
phosphine oxide was observed and the ruthenium complexes
appeared to be air-stable.
Scheme 3: syntheses of the gold(I) complexes
Scheme 2: syntheses of the ruthenium(II) complexes
2 | J. Name., 2012, 00, 1-3
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Biological evaluation
2.5 times more than R-2b but display 2.5 higher IC50. The same
behavior can be observed on HEK 293T cell line.
Table 1: Determination of the IC50 values (µM) of Ru(II) complexes and of Au(I)
complexes by MTS assay (Promega®) at 48 h.
The antiproliferative properties of the nine selected Ru(II) and
Au(I) complexes have been investigated on colon (CT26),
breast (4T1), and lung (LLC1) murine cancer cell lines. A model
of non-cancerous human cell line (HEK293T) has also been
included in the study in order to evaluate if the complexes
display or not intrinsic selectivity for cancerous cells. The
cytotoxicity of the complexes has been compared to cisplatin,
which still is the benchmark compound for metal complexes
anticancer agents.
IC50 (µM)
Compounds
CT26
4T1
LLC1
HEK 293T
Ruthenium
p-cymene
R-1a
R-2a
17.3±0.2
40.0±0.3
14.8±0.2
34.9±0.4
15.3±0.8
30.2±0.3
99.8±0.2
4.6±0.3
10.0±0.3
49.8±0.4
Contrary to cisplatin, most of the complexes display the same
antiproliferative properties on the different cell lines.
R-3a
100.0±0.4 121.7±0.8
The three gold complexes A-1a, A-1b, and A-2b display very
Benzoate
R-1b
low IC50. They are even 2 to 10 times more cytotoxic than
cisplatin. These values are in the range of auranofin. Thus, for
these compounds, no impact of the ligands (phosphines,
chlorido, thiolato) has been observed.
2.2±0.2
2.0±0.2
5.0±0.7
2.2±0.2
2.7±0.4
9.9±0.1
2.2±0.2
2.6±0.5
10.0±0.1
2.2±0.3
2.7±0.4
9.9±0.2
R-2b
R-3b
On the contrary, for Ru(II) complexes, the results are very
different depending on the ligands borne by the ruthenium
centre.
Gold
A-1a
1.9±0.4
2.7±0.2
2.3±0.2
3.7±0.4
2.3±0.2
2.3±0.3
3.9±0.3
2.6±0.3
2.4±0.2
3.9±0.4
2.3±0.4
2.4±0.4
Indeed, the p-cymene-based-Ru(II)-complexes R-1a, R-2a, and
R-3a display very different antiproliferative properties. R-3a
displays very low cytotoxicity on the different cell lines, R-2a is
twice less potent on CT26, 4T1, and LLC1 than cisplatin, while
R-1a is more efficient on CT26 and 4T1 than cisplatin. In this
case, there is a strong impact of the phosphine ligand on the
cytotoxicity of the complexes.
A-1b
A-2b
Cisplatin
25.3±0.3
24.8±0.2
10.4±0.2
50.1±0.3
Complexes highlighted in blue bear saturated phosphine 7, in
orange, diene phosphine 3, and in white, PPh3. Values written in
red correspond to IC50 superior to cisplatin one.
Concerning ruthenium-complexes bearing an ethyl benzoate
as η6-arene ligand R-1b
,
R-2b, and R-3b, they all display very
low
IC50,
significantly lower than the other
Table 2: Cellular uptake of Ru(II) complexes in CT26 and HEK 293T cells treated with 5
[RuCl2(phosphine)arene] complexes reported in literature. R-
1b and R-2b are even in the range of gold(I) complexes and,
thus, they are 4 to 12 times more potent than cisplatin. These
complexes are as cytototoxic on cancer cell lines as on
HEK 293T. However, it could be interesting to take advantage
of the low impact of the nature of the phosphine to graft a
vector at this position to target selectively the complexes to
cancer cell lines.
CT26
HEK 293T
[Ru]
Compounds
[Ru]
IC50
(µM)
IC50
(µM)
(ppt/µg prot.)
(ppt/µg prot.)
R-3a
R-2a
R-1a
R-1b
R-2b
R-3b
13.1
100.0
40.0
17.3
2.2
13.9
49.8
10.0
4.6
23.4
43.5
84.2
90.1
219.0
35.9
32.1
Many hypotheses can be found to explain such impact of the
ligands of ruthenium complexes. Among them, structural
variation of the complex can modify its cell uptake and/or its
mechanism of action (arene removal, chlorido aquation…). To
get insight into the mechanism of action, we decided to
quantify ruthenium uptake of ruthenium complexes in CT26
and HEK 293T by ICP-MS.
206.6
202.9
317.6
2.2
2.0
2.7
5.0
9.9
μM of metal complexes for 1 h at 37 °C analysed by ICP-MS (in pmole per µg of protein)
compared to their respected IC50
.
The results of this study have been gathered in Table 2 and
compared to the corresponding IC50. The compounds have
been ranked according to their uptake in CT26 in order to
facilitate the interpretation of the results. According to this
study, a clear correlation can be made between cell uptake of
ruthenium-complexes and their antiproliferative properties,
apart from R-3b. Indeed, surprisingly, R-3b accumulate around
This study is too preliminary to determine the mechanism of
action of the different arene-ruthenium derivatives. However,
we obtained answers to our questions:
-
There is no “diene effect”, indeed the nature of the
phosphine ligand influences the internalization of the
complex in cells, but it is not yet predictable: an
aliphatic chain increases the internalization of a
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p-cymene-ruthenium complex, while it decreases the
one of an ethyl benzoate-ruthenium complex.
There is a strong impact of the nature of arene: ethyl
benzoate dramatically increases both internalization
and antiproliferative properties of complexes.
It seems that antiproliferative properties of our
complexes are more linked to their cell uptake than
to the lability of arene.
-
-
Syntheses:
4: 4-methylhexanol:
A solution of alcohol 1 (2.0 g, 17.8 mmol) in degassed methanol (50
mL) was loaded in an autoclave with 10 wt% Pd/C (10 wt%, 0.2 g).
The autoclave atmosphere was flushed with dihydrogen (10 bar)
and the mixture was stirred for 16 h at room temperature. Pd/C
was filtered off with Millipore filter. The filtrate was concentrated
to give 4 as a yellow oil (1.67 g, 80 %).
R-2b, R-1b, and even R-3b display cytotoxicity in the range of
the most cytotoxic arene-ruthenium complexes.22,28 However,
these highly cytotoxic complexes are really different than our
complexes: they are cationic, ours are neutral; their ruthenium
centres bear “L2” diamino ligand, instead of a chlorido and a
phosphino ligands for our complexes. It is worth noting that
several studies conclude that increasing the lipophilicity of a
Ru(II)-complex leads to the improvement of its cytotoxic,22,29
while, in our case, we observe the opposite effect when we
replace benzoate ligand by p-cymene one.
1H NMR (300 MHz, CDCl3): δ (ppm) = 3.62 (t, 2H, JHH = 6.6 Hz,
3
CH2OH), 1.65 – 1.44 (m, 2H, CH2CH2OH), 1.43 – 1.26 (m, 1H + 1H +
1H, CH, CHaHbCH2CH2OH, CHaHbCH3), 1.23 – 1.07 (m, 1H + 1H,
3
CHaHbCH2CH2OH, CHaHbCH3), 0.87 (d, 3H, JHH = 6.3 Hz, CHCH3
3
overlapping with CH2CH3), 0.86 (t, 3H, JHH = 7.1 Hz, CH2CH3
overlapping with CHCH3).
13C{1H} NMR (75 MHz, CDCl3): δ (ppm) = 63.6 (CH2OH), 34.4 (CH),
32.7 (CH2CH2CH2OH), 30.5 (CH2CH2OH), 29.6 (CH3CH2), 19.3
(CHCH3), 11.4 (CH2CH3).
Experimental
Otherwise specified, all reactions were carried out under Argon
using conventional Schlenk techniques. CH2Cl2, diethyl ether,
toluene, THF and pentane were dried using an MBRAUN SPS 800
solvent purification system. The precursors ruthenium dimers
[RuCl2(η6-arene)]2 (arene = p-cymene, BzOEt) and [AuCl(tht)] have
been synthesized according to literature procedure.25–27 All other
reagents were commercially available and used as received.
5: 4-methylhexyl methanesulfonate
In a round-bottom flask, to a solution of alcohol 4 (1 eq., 13.8
mmol, 1.60 g) in CH2Cl2 (60 mL) was added triethylamine (1.2 eq.,
16.5 mmol, 2.30 mL). The yellow solution was cooled down to 0°C
and methanesulfonyl chloride (1.2 eq., 16.5 mmol, 1.28 mL) was
slowly added. The mixture was stirred for 4 h while warming to rt.
The mixture was treated with water (30 mL) and sodium
bicarbonate saturated solution (30 mL), and extracted by CH2Cl2 (3 x
60 mL). The gathered organic layers were washed with water (3 x
50 mL) and brine (50 mL), dried with anhydrous magnesium sulfate,
and concentrated to give 5 as an orange oil (2.40 g, 90 %).
All of the analyses were performed at the “Plateforme d’Analyse
Chimique et de Synthèse Moléculaire de l’Université de Bourgogne”
(PACSMUB). The identity and purity of the compounds were
unambiguously established using elemental analyses, multinuclear
NMR spectroscopy, high-resolution mass spectrometry, and
Infrared. Elemental analyses were obtained on a Flash EA 1112
CHNS-O Thermo Electron Flash instrument. High resolution mass
spectra were recorded on a Thermo LTQ Orbitrap XL ESI-MS
spectrometer. NMR spectra (1H, 31P, 13C) were recorded on Bruker
300 Avance III or 500 Avance III spectrometers. Chemical shifts are
HR-MS (ESI-pos): calcd for [C8H18O3SNa]+ [M + Na]+: 217.08689.
Found: 217.08593 (-4.4 ppm).
quoted in parts per million (δ) relative to TMS (for H and 13C) or
1
85% H3PO4 (for 31P). For 1H and 13C spectra, values were determined
by using solvent residual signals (e.g. CHCl3 in CDCl3) as internal
standards. For 31P, 85% H3PO4 was used as an external standard.
The coupling constants (J) are reported in Hertz (Hz). The list of
abbreviations for signals in NMR are the following: s = singlet, d =
doublet, t = triplet, q = quadruplet, dd = doublet of doublets, ddd =
doublet of doublet of doublets, m = multiplet, and bs = broad
3
1H NMR (300 MHz, CDCl3): δ (ppm) = 4.20 (t, 2H, JHH = 6.8 Hz,
CH2O), 2.98 (s, 3H, SO2CH3), 1.84 – 1.63 (m, 2H, CH2CH2OH), 1.46 –
1.27 (m, 1H + 1H + 1H, CH, CHaHbCH2CH2OH, CHaHb CH3), 1.26 – 1.08
(m, 1H + 1H, CHaHbCH2CH2OH, CHaHbCH3), 0.91 – 0.82 (m, 3H + 3H
overlapping, CH2CH3, CHCH3).
13C{1H} NMR (75 MHz, CDCl3): δ (ppm) = 70.6 (CH2O), 37.5 (SO2CH3),
34.1 (CH), 32.2 (CH2CH2CH2OH), 29.4 (CH2CH2OH), 26.9 (CH3CH2),
19.1 (CHCH3), 11.4 (CH2CH3).
singlet. Assignment of H and 13C signals (when possible) was done
1
thanks to DEPT and 2D experiences (COSY and HSQC) (data not
shown). Infrared spectra were recorded on a Bruker Vertex 70v
spectrophotometer fitted with a Globar MIR source, a Ge/KBr (MIR)
or silicon (FIR) beam splitter, a DLaTGS detector and a diamond ATR
module.
6: (4-methylenehexyl)diphenylphosphine borane
Diphenylphosphine (1eq., 7.76 mmol, 1.35 mL) was diluted in
diethyl ether (30 mL). n-Butyllithium (1 eq., 7.76 mmol, 1.6 M in
hexanes, 4.85 mL) was slowly added at room temperature, a
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yellow/orange color appeared, and the resulting mixture was 1H NMR (500 MHz, CDCl3): δ (ppm) = 7.92 – 7.85 (m, 4H, o-Ph), 7.51
stirred 1 h. In another schlenk, mesylate 5 (1 eq., 7.76 mmol, 1.51 g) – 7.43 (m, 4H + 2H, m-Ph, p-Ph), 5.26 (d, 2H, 3JHH = 5.8 Hz, iPrCqCH p-
was dissolved in diethyl ether (30 mL), and cooled to 0°C. The first cymene), 5.07 (d, 2H, 3JHH = 6.0 Hz, MeCqCH p-cymene), 2.58 – 2.46
i
solution was slowly transferred to the second one, and the resulting (m, 2H + 1H, PCH2, CH Pr), 1.88 (s, 3H, Me p-cymene), 1.18 – 1.02
reacting mixture was stirred 2 h while warming to room (m, 1H
+ 1H + 1H + 1H, CHaHbCH3, CHCH3 PCH2CHaHb,
temperature. A white solid appeared. The resulting mixture was PCH2CH2CHaHb), 0.86 – 1.01 (m, 1H + 1H + 1H, CHaHbCH3,
i
filtered over Celite®, and the filtrate was concentrated to give the PCH2CHaHb, PCH2CH2CHaHb), 0.80 (d, 3JHH = 7.1 Hz, 6H, CH3 Pr), 0.69
crude phosphine 7 as a yellow oil.
(t, 3JHH = 7.2 Hz, 3H, CH2CH3), 0.58 (d, 3JHH = 6.3 Hz, 3H, CHCH3).
As the product was not pure enough, it was further purified. To
carry it out, we used the phosphine-borane route. The crude was 13C{1H} NMR (126 MHz, CDCl3): δ (ppm) = 133.43 (d, 2JCP = 8.5 Hz, o-
2
1
dissolved in THF (30 mL). Borane dimethyl sulfide (1.1 eq., 8.54 Pha), 133.41 (d, JCP = 8.3 Hz o-Phb), 133.1 (d, JCP = 42.1 Hz, i-Pha),
1
4
mmol, 0.809 mL) was diluted in THF (30 mL), and transferred. The 133.0 (d, JCP = 42.0 Hz, i-Phb), 130.6 (d, JCP = 1.9 Hz, p-Ph), 128.4
3
i
resulting mixture was stirred 16 h at room temperature. The (pseudo d, JCP ≈ 9.9 Hz, m-Ph), 108.1 (s, Cq Pr p-cymene), 93.6 (s,
mixture was treated by water (few drops), dried with anhydrous CqMe p-cymene), 90.7 (d, 2JCP = 3.4 Hz, MeCqCH p-cymene), 85.7 (d,
3
magnesium sulfate, and filtered. The filtrate was concentrated and 2JCP = 5.3 Hz, iPrCqCH p-cymene), 37.8 (d, JCP = 11.7 Hz,
purified by silica column chromatography (Pentane/Et2O; 95:5) to PCH2CH2CH2), 33.8 (s, CHCH3), 30.1 (s, CH iPr), 29.4 (s, CH2CH3), 22.9
1
give the pure phosphine-borane 6 product as a colorless oil (1.55 g, (d, JCP = 28.5 Hz PCH2), 21.41 (s, CH3a iPr), 21.40 (s, CH3b iPr), 20.6
2
67 %).
(d, JCP = 8.9 Hz, PCH2CH2), 19.0 (s, CHCH3), 17.4 (s, Me p-cymene),
The phosphine 7 can be obtained from the phosphine-borane 6 by 11.3 (s, CH2CH3).
using DABCO (1.2 eq.) at 50°C in toluene during 16h. After reaction,
the crude is filtered on a small silica plug and directly used as a 31P{1H} NMR (202 MHz, CD2Cl2): δ (ppm) = 24.0 (s).
toluene solution.
Selected IR Bands (ATR): wavenumber (cm-1) = 290 (νRu–Cl).
HR-MS (ESI-pos): calcd for [C19H28BPNa]+ [M + Na]+: 321.19139.
Found: 321.19029 (-3.4 ppm).
R-1b: RuCl2(η6-BzOEt)[(4-methylhexyl)diphenylphosphine-κP]
1H NMR (300 MHz, CDCl3): δ (ppm) = 7.72 - 7.62 (m, 4H, o/m-Ph),
7.51 – 7.39 (m, 4H + 2H, o/m-Ph, p-Ph), 2.24 – 2.11 (m, 2H, PCH2),
1.63 – 1.46 (m, 2H, PCH2CH2), 1.42 – 1.00 (m, 5H, PCH2CH2CH2, CH,
CH2CH3), 0.82 (t, 2H, 3JHH = 7.2 Hz, CH2CH3 overlapping with CHCH3),
0.79 (d, 3H, 3JHH = 6.1 Hz, CHCH3 overlapping with CH2CH3).
[RuCl2(η6-BzOEt)]2 (1 eq., 343 mg, 0.532 mmol) and phosphine 7
(2.2 eq., 333 mg, 1.17 mmol) in toluene (5 mL) were stirred at room
temperature for 16 h in the dark. The solvent was evaporated. The
residue was triturated and washed with pentane, and dried to give
R-1b as an orange solid (465 mg, 72%).
13C{1H} NMR (75 MHz, CDCl3): δ (ppm) = 132.3 (d, JCP = 9.0 Hz, o/m-
Elemental Analysis: calcd for C28H35Cl2O2PRu: C, 55.45; H, 5.82.
4
1
Ph), 131.2 (d, JCP = 2.5 Hz, p-Ph), 130.0 (d, JCP = 54.5 Hz, i-Ph),
Found: C, 55.89; H, 6.53.
3
128.9 (d, JCP = 9.9 Hz, o/m-Ph), 38.3 (d, JCP = 13.3 Hz, PCH2CH2CH2),
1
HR-MS (ESI-pos): calcd for [C28H35Cl2O2PRuNa]+ [M
+
Na]+:
34.1 (s, CH), 29.5 (s, CH3CH2), 26.1 (d, JCP = 36.7 Hz, PCH2), 20.8 (s,
PCH2CH2), 19.1 (s, CH3CH), 11.4 (s, CH3CH2).
629.06893. Found: 629.06977 (-1.6 ppm).
31P{1H} NMR (121 MHz, CDCl3): δ (ppm) = 17.1 – 15.0 (m).
1H NMR (500 MHz, CDCl3): δ (ppm) = 7.89 – 7.82 (m, 4H, o-Ph), 7.54
– 7.46 (m, 4H + 2H, m-Ph, p-Ph), 6.34 (d, 2H, 3JHH = 5.6 Hz, o-BzOEt),
3
R-1a: RuCl2(η6-p-cymene)[(4-methylhexyl)diphenylphosphine-κP]
5.47 – 5.43 (m, 1H, p-BzOEt), 5.03 (t, JHH = 5.1 Hz, 2H, m-BzOEt),
3
4.30 (q, JHH = 7.0 Hz, 2H, OCH2), 2.61 – 2.49 (m, 2H, PCH2), 1.35 (t,
[RuCl2(η6-p-cymene)]2 (1 eq., 245 mg, 0.400 mmol) and phosphine 7 3JHH = 7.0 Hz, 3H, OCH2CH3), 1.19 – 1.00 (m, 1H + 1 H + 1H + 2H,
(2.2 eq., 250 mg, 0.879 mmol) in toluene (10 mL) were stirred at CHaHbCH3, PCH2CH2CHaHb, CH, PCH2CH2), 0.99 – 0.87 (m, 1H + 1H,
3
room temperature for 16 h in the dark. The solvent was CHaHbCH3, PCH2CH2CHaHb), 0.70 (t, JHH = 7.0 Hz, 3H, CH2CH3), 0.60
evaporated. The residue was triturated and washed with pentane, (d, 3JHH = 6.3 Hz, 3H, CHCH3).
and dried to give R-1a as an orange powder (392 mg, 83%).
2
13C{1H} NMR (127 MHz, CDCl3): δ (ppm) = 133.3 (d, JCP = 8.6 Hz, o-
2
1
Elemental Analysis: calcd for C29H39Cl2PRu: C, 58.98; H, 6.66. Found: Pha), 133.2 (d, JCP = 8.6 Hz, o-Phb), 132.3 (d, JCP = 45.4 Hz, i-Pha),
C, 59.08; H, 6.52.
132.6 (d, 1JCP = 45.4 Hz, i-Phb), 130.9 (d, 4JCP = 2.2 Hz, p-Ph), 128.6 (d,
3JCP = 9.7 Hz, m-Ph), 94.8 – 94.7 (m, o-BzOEt), 89.8 (s, p-BzOEt), 85.9
HR-MS (ESI-pos): calcd for [C29H39ClPRu]+ [M - Cl]+: 555.15159. (d, JCP = 6.9 Hz, i-BzOEt), 84.7 – 84.6 (m, m-BzOEt), 62.6 (s, OCH2),
2
3
Found: 555.15037 (-2.2 ppm).
37.8 (d, JCP = 12.2 Hz, PCH2CH2CH2), 33.8 (s, CH), 29.4 (s, CH2CH3),
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24.1 (d, 1JCP = 30.2 Hz, PCH2), 20.9 (d, 2JCP = 8.0 Hz, PCH2CH2), 19.0 (s, washed with pentane (2 x 5 mL) and dried to afford A-1a as a beige
CHCH3), 14.6 (s, OCH2CH3), 11.3 (s, CH2CH3).
solid (180 mg, 70% yield).
31P{1H} NMR (202 MHz, CDCl3): δ (ppm) = 25.3 (s).
Elemental Analysis: calcd for C19H25AuClP: C, 44.16; H, 4.88. Found:
C, 44.97; H, 5.57.
Selected IR Bands (ATR): wavenumber (cm-1) = 283 (νRu–Cl), 1103
(νO–C–C), 1268 (νC–C(=O)–O), 1728 (νC=O).
HR-MS (ESI-pos): calcd for [C19H25AuClPNa]+ [M + Na]+: 539.09401;
found: 539.09204 (–3.7 ppm).
R-2b: RuCl2(η6-BzOEt)[(4-methylenehex-5-enyl)diphenylphosphine
-κP]
1H NMR (500 MHz, CDCl3): δ (ppm) = 7.70 – 7.64 (m, 4H, o-Ph), 7.53
– 7.43 (m, 4H + 2H, m-Ph, p-Ph), 2.47 – 2.35 (m, 2H, PCH2), 1.71 –
1.53 (m, 2H, PCH2CH2 overlapping with H2O), 1.49 – 1.41 (m, 1H,
PCH2CH2CHaHb), 1.21 – 1.34 (m, 1H + 1H + 1H, PCH2CH2CHaHb,
CHCH3, CHaHbCH3), 1.03–1.13 (m, 1H, CHaHbCH3), 0.81 (t,
3JHH = 7.4Hz, 3H, CH2CH3), 0.79 (d, 3JHH = 6.3 Hz, 3H, CHCH3).
[RuCl2(η6-BzOEt)]2 (1 eq., 383 mg, 0.594 mmol) and phosphine 3
(2.1 eq., 350 mg, 1.25 mmol) in toluene (6 mL) were stirred at room
temperature for 16 h in the dark. The solvent was evaporated. The
residue was triturated and washed with pentane, and dried to give
R-2b as an orange solid (522 mg, 73%).
2
13C{1H} NMR (126 MHz, CDCl3): δ (ppm) = 133.32 (d, JCP = 12.9 Hz,
o-Pha), 133.31 (d, 2JCP = 12.9 Hz, o-Phb), 131.9 (d, 4JCP = 2.1 Hz, p-Ph),
Elemental Analysis: calcd for C28H31Cl2O2PRu: C, 55.82; H, 5.19.
1
1
129.91 (d, JCP = 58.2 Hz, i-Pha), 129.90 (d, JCP = 58.2 Hz, i-Phb),
Found: C, 56.07; H, 5.74.
129.3 (d, JCP = 11.4 Hz, m-Ph), 37.9 (d, 3JCP = 15.7 Hz, PCH2CH2CH2),
3
34.07 (s, CH), 29.4 (s, CH2CH3), 28.6 (d, 1JCP = 36.9 Hz, PCH2), 23.3 (d,
2JCP = 2.6 Hz, PCH2CH2), 19.1 (s, CHCH3), 11.4 (s, CH2CH3).
HR-MS (ESI-pos): calcd for [C28H31Cl2O2PRuNa]+ [M
+
Na]+:
625.03744. Found: 625.03905 (-2.6 ppm).
31P{1H} NMR (202 MHz, CDCl3): δ (ppm) = 29.4 (s, PPh2).
Selected IR Bands (ATR, cm-1): 323 (νAu–Cl).
1H NMR (500 MHz, CDCl3): δ (ppm) = 7.88 – 7.80 (m, 4H, o-Ph), 7.54
– 7.44 (m, 4H + 2H, m-Ph, p-Ph), 6.35 (d, 3JHH = 6.0 Hz, 2H, o-BzOEt),
3
3
6.18 (dd, JHHtrans = 17.9 Hz, JHHcis = 10.6 Hz, 1H, HC=CH2), 5.45 (m,
3
3
1H, p-BzOEt), 5.04 (t, JHH = 5.8 Hz, 2H, m-BzOEt), 4.95 (d, JHHtrans
=
3
17.6 Hz, 1H, HC=CHcisHtrans), 4.88 (d, JHHcis = 10.6 Hz, 1H,
A-2a: AuCl[(4-methylenehex-5-enyl)diphenylphosphine]
HC=CHcisHtrans overlapping with C=CHaHb), 4.87 (s, 1H, C=CHaHb,
3
overlapping with HC=CHcisHtrans), 4.73 (s, 1H, C=CHaHb), 4.30 (q, JHH AuCl(tht) (1 eq., 0.246 mmol, 79 mg) and phosphine 3 (1 eq., 0.246
= 7.1 Hz, 2H, OCH2), 2.67 – 2.60 (m, 2H, PCH2CH2CH2), 2.06 (t, 3JHH
=
mmol, 69 mg) in frozen-degassed benzene (4 mL) were stirred at
3
7.5 Hz, 2H, PCH2), 1.35 (t, JHH = 7.1 Hz, 3H, OCH2CH3, overlapping room temperature for 16 h. The reacting mixture was concentrated
with PCH2CH2), 1.33 – 1.26 (m, 2H, PCH2CH2, overlapping with and washed with pentane (2 x 5 mL) and dried to give A-2a as a
OCH2CH3).
white solid (93 mg, 74% yield). The product has to be used quickly
in further reactions because it is readily degraded and goes from
13C{1H} NMR (126 MHz, CDCl3): δ (ppm) = 164.1 (s, C=O), 145.3 (s, white to purple quickly (even when stored in a glovebox).
2
C=CH2), 138.5 (s, HC=CH2), 133.2 (d, JCP = 8.4 Hz, o-Ph), 132.2 (d,
1JCP = 45.3 Hz, i-Ph), 131.0 (d, 4JCP = 2.4 Hz, p-Ph), 128.6 (d, 3JCP = 9.8 1H NMR (500 MHz, CDCl3): δ (ppm) = 7.67 – 7.62 (m, 4H, o-Ph), 7.53
Hz, m-Ph), 116.3 (s, C=CH2), 113.4 (s, HC=CH2), 94.8 (d, 2JCP = 3.4 Hz, – 7.44 (m, 4H + 2H, m-Ph, p-Ph), 6,31 (dd, 3JHHtrans = 17.7 Hz, 3JHHcis
=
o-BzOEt), 89.8 (s, p-BzOEt), 86.1 (d, JCP = 7.0 Hz, i-BzOEt), 84.7 (d, 10.9 Hz, 1H, CH=CH2), 5.13 (d, 3JHHtrans = 17.7 Hz, 1H, CH=CHcisHtrans),
2
2JCP = 3.2 Hz, m-BzOEt), 62.6 (s, OCH2CH3), 32.5 (d, JCP = 13.5 Hz, 5.05 (m, 1H, C=CH2), 5.04 (d, 1H, JHHcis = 10.9 Hz, CH=CHcisHtrans),
3
3
PCH2CH2CH2), 24.1 (d, JCP = 30.7 Hz, PCH2), 21.9 (d, JCP = 7.5 Hz, 4.95 (m, 1H, C=CH2), 2.45 – 2.39 (m, 2H, PCH2), 2.38 (t, 2H, 3JHH = 6,9
1
2
PCH2CH2), 14.6 (s, OCH2CH3).
31P{1H} NMR (202 MHz, CDCl3): δ (ppm) = 24.8 (s).
Hz, PCH2CH2CH2), 1.89 – 1.81 (m, 2H, PCH2CH2).
13C{1H} NMR (126 MHz, CDCl3): δ (ppm) = 144.5 (s, Cq, C=C), 138.3
4
Selected IR Bands (ATR): wavenumber (cm-1) = 289 (νRu–Cl), 1100 (s, Cq, C=C), 133.3 (d, JCP = 13.1 Hz, o/m-Ph), 132.1 (d, JCP = 2.7 Hz,
(νO–C–C), 1267 (νC–C(=O)–O), 1718 (νC=O).
p-Ph), 129.5 (d, 1JCP = 60.0 Hz, i-Ph), 129.4 (d, JCP = 11.1 Hz, o/m-Ph),
3
117.0 (s, CH, C=C), 114.0 (s, CH, C=C), 32.2 (d, JCP = 16.8 Hz,
1
2
PCH2CH2CH2), 27.8 (d, JCP = 38.2 Hz, PCH2), 23.8 (d, JCP = 4.4 Hz,
A-1a : AuCl[(4-methylhexyl)diphenylphosphine]
PCH2CH2).
AuCl(tht) (1 eq., 0.50 mmol, 160 mg) and phosphine 7 (1.1 eq., 0.55
mmol, 156 mg) in toluene (4 mL) were stirred at room temperature
for 3 h. The reacting mixture was concentrated, and the residue was
31P{1H} NMR (202 MHz, CDCl3): δ (ppm) = 29.1 (s, PPh2).
6 | J. Name., 2012, 00, 1-3
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A-1b: Au[(4-methylhexyl)diphenylphosphine][3,4,5-triacetyloxy-6- Gold complex A-2a (1 eq., 0.117 mmol, 60 mg,) was dissolved in
(acetyloxymethyl)oxane-2-thiolate]
degassed acetone (3 mL) and cooled to 0°C. In another schlenk, 1-
thio-β-D-glucose tetraacetate (1 eq., 0.117 mmol, 38 mg) was
dissolved in degassed acetone (3 mL), and in the dark 1 M NaOH (1
eq., 0.117 mmol, 117 µL) was added. The mixture was stirred 10
min at room temperature, cooled to 0°C, and slowly added to the
first solution. The mixture was stirred 3 h in the dark while warming
to room temperature. The reacting mixture was settled and the
supernatant was filtered off. The filtrate was concentrated and
DCM (4 mL) was added. The supernatant was filtered off,
concentrated, and washed with pentane to give A-2b as a white
solid (m = 73 mg, 74% yield).
Gold complex A-1a (1 eq., 0.193 mmol, 100 mg,) was dissolved in
degassed acetone (4 mL) and cooled to 0°C. In another schlenk, 1-
thio-β-D-glucose tetraacetate (1 eq., 70 mg, 0.193 mmol) was
dissolved in degassed acetone (3 mL), and in the dark 1 M NaOH (1
eq., 0.193 mmol, 193 µL) was added. The mixture was stirred 10 mn
at room temperature, cooled to 0°C, and slowly added to the first
solution. The resulting mixture was stirred 3 h in the dark while
warming to room temperature. The reacting mixture was settled
and the supernatant was filtered off. The filtrate was concentrated
and DCM (4 mL) was added. The supernatant was filtered off,
concentrated and washed with pentane to give A-1b as a white
powder (m = 100 mg, 62% yield).
Elemental Analysis: calcd for C33H40AuO9PS: C, 47.15; H, 4.80; S,
3.81. Found: C, 46.28; H, 5.24; S, 3.06.
HR-MS (ESI-pos): calcd for [C33H40AuO9PSNa]+ [M
+
Na]+:
Elemental Analysis: calcd for C33H44AuO9PS: C, 46.92; H, 5.25; S,
3.80. Found: C, 46.60; H, 5.58; S, 3.72.
863.16884; found: 863.16883 (0.01 ppm).
HR-MS (ESI-pos): calcd for [C33H45AuO9PS]+ [M + H]+: 845.21819;
1H NMR (500 MHz, CDCl3): δ (ppm) = 7.72 – 7.63 (m, 4H, o-Ph), 7.52
3
found: 845.21505 (-3.7 ppm).
– 7.42 (m, 4H + 2H, m-Ph + p-Ph), 6,31 (dd, 1H, JHHtrans =17.7 Hz,
3JHHcis = 10.8 Hz, CH=CH2), 5.18 – 5.01 (m, 2H + 1H + 3H + 1 H,
CH=CH2+ C=CH2 + 3 x CHOAc + CHS), 4.96 (bs, 1H, C=CH2), 4.20 (dd,
1H, 3JHH = 12.2 Hz, 3JHH = 4.9 Hz, CHaHbOAc), 4.09 (dd, 1H, 2JHH = 12.2
1H NMR (500 MHz, CDCl3): δ (ppm) = 7.75 – 7.66 (m, 4H, o-Ph), 7.52
3
– 7,45 (m, 4H + 2H, m-Ph + p-Ph), 5.18 (d, 1H, JHH = 9.3 Hz, CHS),
3
3
3
3
5.14 (t, 1H, JHH = 9.2 Hz, CH(OAc)CH(OAc)CH(OAc), 5.09 (t, 1H, JHH
Hz, JHH = 2.3 Hz, CHaHbOAc), 3.74 (ddd, 1H, JHH = 9.8, 4.9, 2.3 Hz,
3
3
= 9.4 Hz, CHCHCH2OAc), 5.04 (t, 1H, JHH = 9.3 Hz, CHSCHOAc), 4.21
(dd, 1H, 2JHH = 12.2 Hz, 3JHH = 4.8 Hz, CHaHbOAc), 4.11 (dd, 1H, 2JHH
CHCH2OAc), 2.48 – 2.41 (m, 2H, PCH2), 2.38 (t, 2H, JHH = 7.0 Hz, P
=
CH2CH2CH2), 2.04 (s, 3H, OAc), 2.00 (s, 3H, OAc), 1.97 (s, 3H,
CH2OAc), 1.92 (s, 3H, OAc), 1.89 – 1.81 (m, 2H, PCH2CH2).
12.2, 3JHH = 2.4 Hz, CHaHbOAc), 3.74 (ddd, 1H, JHH = 2.4, 4.8, 9.8 Hz,
CHCH2OAc), 2.46 – 2.37 (m, 2H, PCH2), 2.06 (s, 3H, OAc), 2.00 (s, 3H,
OAc), 1.98 (s, 3H, OAc), 1.92 (s, 3H, CH2OAc), 1.73 – 1.56 (m, 2H,
PCH2CH2), 1.50 – 1.42 (m, 1H, PCH2CH2CHaHb), 1.36 – 1.22 (m, 1H +
1H + 1H, PCH2CH2CHaHb, CHCH3, CHaHb), 1.14 – 1.05 (m, 1H,
3
13C{1H} NMR (126 MHz, CDCl3): δ (ppm) = 170.9 (s, C=O), 170.4 (s,
C=O), 169.7 (s, C=O), 169.6 (s, C=O), 144.6 (s, C=CH2), 138.4 (s,
2
2
HC=CH2), 133.4 (d, JCP = 12.8 Hz, o-Pha), 133.3 (d, JCP = 12.8 Hz, o-
3
1
CHaHbCH3), 0.82 (t, 3H, JHH = 7.4 Hz, CH2CH3 overlapping with
CHCH3), 0.81 (d, 3H, 3JHH = 6.2 Hz, CHCH3 overlapping with CH2CH3).
Phb), 131.8 (pseudo t, J = 2.8 Hz, p-Pha+b), 130.3 (d, JCP = 55.2 Hz, i-
1
3
Pha), 130.2 (d, JCP = 55.2 Hz, i-Phb), 129.3 (d, JCP = 11.0 Hz, m-Pha),
3
129.3 (d, JCP = 11.0 Hz, m-Phb), 116.9 (s, HC=CH2), 113.9 (s, C=CH2),
13C{1H} NMR (126 MHz, CDCl3): δ (ppm) = 170.9 (s, C=O), 170.5 (s,
C=O), 169.8 (s, C=O), 169.7 (s, C=O), 133.42 (m, o-Pha+b), 131.7 (m,
83.2 (s, CHS), 77.6 (s, CHOAc), 75.9 (s, CHCH2OAc), 74.4 (s, CHOAc),
69.1 (s, CHOAc), 62.9 (s, CH2OAc), 32.3 (d, ³JCP = 16.5 Hz,
3
3
1
2
p-Pha+b), 130.8 (d, JCP = 52.6 Hz, i-Pha), 130.7 (d, JCP = 54.2 Hz, i-
PCH2CH2CH2), 27.7 (d, JCP = 35.0 Hz, PCH2), 23.7 (d, JCP = 4.4 Hz,
PCH2CH2), 21.3 (s, C(=O)CH3), 20.9 (s, C(=O)CH3), 20.8 (s, C(=O)CH3),
20.7 (s, C(=O)CH3).
3
3
Phb), 129.3 (d, JCP = 11.0 Hz, m-Pha), 129.2 (d, JCP = 11.0 Hz, Cmeta
Pha), 83.3 (s, CHS), 77.7 (s, CHSCHOAc), 75.9 (s, CHCH2OAc), 74.4 (s,
CH(OAc)CH(OAc)CHOAc), 69.1 (s, CHCHCH2OAc), 63.0 (s, CH2OAc),
3
31P{1H} NMR (202 MHz, CDCl3): δ (ppm) = 33.3 (s, PPh2)
38.0 (d, JCP = 15.6 Hz, PCH2CH2CH2), 34.0 (s, CH CH3), 29.4 (s,
1
CH2CH3), 28.5 (d, JCP = 33.9 Hz, PCH2), 23.2 (m, PCH2CH2), 21.3 (s,
Selected IR Bands (ATR, cm-1): 372 (νAu–S), 1030 (νO–C–C, OAc), 1216
(C=O)CH3), 20.9 (s, (C=O)CH3), 20.8 (s, (C=O)CH3), 20.7 (s,
CH2(C=O)CH3), 19.1 (s, CHCH3), 11.4 (s, CH2CH3).
(νC–C(=O)–O), OAc), 1742 (νC=O, OAc)
31P{1H} NMR (202 MHz, CDCl3): δ (ppm) = 34.8 (s, PPh2)
Antiproliferative properties evaluation:
Murine colon (CT26), mammary (4T1) and lung (LLC1) cancer cell
lines but also human HEK293T cell line were obtained from the
ATCC. All cells were maintained at 37°C in DMEM with 4.5 g/l
glucose supplemented with 10% FBS and antibiotics (Dutscher,
Brumath, France). Cells were grown at a density of 1x104 cells per
well in 96-well cell culture plates (Dutscher) the day before
treatment with different concentrations of cisplatin, ruthenium, or
Selected IR Bands (ATR, cm-1): 371 (νAu–S), 1030 (νO–C–C, OAc), 1217
(νC–C(=O)–O), OAc), 1743 (νC=O, OAc)
A-2b: AuCl[(4-methylenehex-5-enyl)diphenylphosphine][3,4,5-
triacetyloxy-6-(acetyloxymethyl)oxane-2-thiolate]
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gold complexes during 48 h. Thereafter, 10 µL of MTS (3-(4,5-
dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-
Support was provided by the Conseil Régional de Bourgogne
Franche-Comté, the Ministère de l’Enseignement Supérieur et
de la Recherche, the Centre National de la Recherche
Scientifique (CNRS), and the French Research National Agency
(ANR) via project JCJC “SPID” ANR-16-CE07-0020, the
Université de Bourgogne, the École Pratique des Hautes
Études (EPHE). This work is part of the project
PHARMACOIMAGERIE ET AGENTS THERANOSTIQUES and of
the project CHIMIE DURABLE, ENVIRONNEMENT ET
AGROALIMENTAIRE, supported by the Université de
Bourgogne, Conseil Régional de Bourgogne through the plan
d'actions régional pour l'innovation (PARI) and the European
Union through the PO FEDER-FSE Bourgogne 2014/2020
programs. FrenchBIC is acknowledged for fruitful discussion.
Dr F. Picquet and Ms M.-J. Penouilh are gratefully
acknowledged for HR-MS analyses, and Dr M. Heydel for the
ICP-MS analyses.
sulfophenyl)-2H-tetrazolium, Promega, Charbonnieres, France) was
added in 200 µL of medium and absorbance at 490 nm was
measured after 3h incubation at 37°C. DMSO at comparable
concentrations did not show any effects on cell cytotoxicity. IC50
was calculated using GraphPad7.0 Prism software.
ICP-MS measurement:
Determination of uptake of ruthenium was performed in CT26 and
HEK293T cells by ICP-MS. All of these cells (1.105 cells) were
incubated at 37 °C for 1 hour with 15 µM of different ruthenium
complexes. Then, cells were rinsed twice with phosphate buffer
saline (PBS) and lysed in water. All samples were digested in ICP-MS
grade concentrated hydrochloric acid (OPTIMA Grade, Fisher
Chemical) for 3 h at room temperature and filled to a total volume
of 5 mL with ultrapure water. Yttrium was added as an internal
standard at a concentration of 1 ppb. Determinations of total metal
contents were achieved using a Thermo Element 2 ICP-MS. The
instrument was tuned daily using a solution provided by the
manufacturer containing 1 ppb each of Ba, B, Co, Fe, Ga, In, K, Li, Lu,
Na, Rh, Sc, Tl, U, Y (Tune-Up Solution ELEMENT, Thermo Fisher
Scientific, Bremen, Germany). The external standard was prepared
by dilution of a gravimetrically prepared ruthenium solution single
element standard at 1000 ppm (ASSURANCE, SPEX CertiPrep) in an
identical matrix to the samples (with regard to the internal standard
and hydrochloric acid).
Notes and references
1 O. Florès, A. Trommenschlager, S. Amor, F. Marques, F. Silva, L.
Gano, F. Denat, M. P. C. Campello, C. Goze, E. Bodio and P. L.
Gendre, Dalton Trans., , DOI:10.1039/C7DT01981E.
2 Y. Ellahioui, S. Prashar and S. Gómez-Ruiz, Inorganics, 2017, 5, 4.
3 M. Cini, T. D. Bradshaw and S. Woodward, Chem. Soc. Rev., 2017,
46, 1040–1051.
4 Y. F. Mui, J. Fernández-Gallardo, B. T. Elie, A. Gubran, I. Maluenda,
M. Sanaú, O. Navarro and M. Contel, Organometallics, 2016, 35,
1218–1227.
5 C. Roder and M. J. Thomson, Drugs RD, 2015, 15, 13–20.
6 C. Nardon, G. Boscutti and D. Fregona, Anticancer Res., 2014, 34,
487–492.
7 T. Zou, C. T. Lum, C.-N. Lok, J.-J. Zhang and C.-M. Che, Chem. Soc.
Rev., 2015, 44, 8786–8801.
8 S. A. Patil, S. A. Patil, R. Patil, R. S. Keri, S. Budagumpi, G. R.
Balakrishna and M. Tacke, FUTURE Med. Chem., 2015, 7, 1305–
1333.
9 B. Bertrand and A. Casini, Dalton Trans., 2014, 43, 4209–4219.
10 M. A. Cinellu, I. Ott and A. Casini, in Bioorganometallic
Chemistry, eds. G. Jaouen and M. Salmain, Wiley-VCH Verlag
GmbH & Co. KGaA, 2014, pp. 117–140.
11 A. Bergamo and G. Sava, Chem. Soc. Rev., 2015, 44, 8818–8835.
12 B. S. Murray, M. V. Babak, C. G. Hartinger and P. J. Dyson,
Coord. Chem. Rev., 2016, 306, 86–114.
13 C. G. Hartinger, M. A. Jakupec, S. Zorbas-Seifried, M. Groessl, A.
Egger, W. Berger, H. Zorbas, P. J. Dyson and B. K. Keppler, Chem.
Biodivers., 2008, 5, 2140–2155.
Conclusions
In this study, we reported the synthesis and the
characterization of several new ruthenium(II) and gold(I)
complexes. The six air stable complexes – along with three
already reported ruthenium complexes and cisplatin – were
included in a biological evaluation of their antiproliferative
properties. Contrary to some previous studies, we observed a
clear impact of the ligands, especially of the arene. Our
findings enable us to design two ruthenium complexes, which
are 50 to 60 times more cytotoxic than the initial [Ru(p-
cymene)PPh3] complex, and more than 10 times more
cytotoxic than cisplatin. A preliminary investigation of the
mechanism of action highlights the strong correlation between
IC50 of the complexes and their cell uptake. Moreover, impact
of the nature of the phosphine on the more cytotoxic
complexes was null to very low. Altogether, these results
prompt us to investigate – in future studies – the possibility to
14 C. P. Popolin and M. R. Cominetti, Mini-Rev. Med. Chem., 2017,
17, 1435–1441.
15 T. Lazarević, A. Rilak and Ž. D. Bugarčić, Eur. J. Med. Chem., ,
DOI:10.1016/j.ejmech.2017.04.007.
introduce
a vector on the phosphine ligand to target
selectively cancer cells.
16 F. Pelletier, V. Comte, A. Massard, M. Wenzel, S. Toulot, P.
Richard, M. Picquet, P. Le Gendre, O. Zava, F. Edafe, A. Casini and
P. J. Dyson, J. Med. Chem., 2010, 53, 6923–6933.
17 M. Wenzel, B. Bertrand, M.-J. Eymin, V. Comte, J. A. Harvey, P.
Richard, M. Groessl, O. Zava, H. Amrouche, P. D. Harvey, P. Le
Gendre, M. Picquet and A. Casini, Inorg. Chem., 2011, 50, 9472–
9480.
Conflicts of interest
There are no conflicts to declare.
Acknowledgements
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Journal Name
ARTICLE
18 M. Wenzel, E. Bigaeva, P. Richard, P. Le Gendre, M. Picquet, A.
Casini and E. Bodio, J. Inorg. Biochem., 2014, 141, 10–16.
19 M. Wenzel, A. de Almeida, E. Bigaeva, P. Kavanagh, M. Picquet,
P. Le Gendre, E. Bodio and A. Casini, Inorg. Chem., 2016, 55,
2544–2557.
20 L. Zhang, C.-C. Chen, T.-P. Ko, J.-W. Huang, Y. Zheng, W. Liu, I.
Wang, S. R. Malwal, X. Feng, K. Wang, C.-H. Huang, S.-T. D. Hsu, A.
H.-J. Wang, E. Oldfield and R.-T. Guo, Angew. Chem. Int. Ed., 2016,
55, 4716–4720.
21 J. D. Hildebrandt, in The Enzymes, eds. F. Tamanoi, C. A. Hrycyna
and M. O. Bergo, Academic Press, 2011, vol. 29, pp. 97–124.
22 M. J. Chow, M. V. Babak, D. Y. Q. Wong, G. Pastorin, C. Gaiddon
and W. H. Ang, Mol. Pharm., 2016, 13, 2543–2554.
23 J. Baraut, A. Massard, F. Chotard, E. Bodio, M. Picquet, P.
Richard, Y. Borguet, F. Nicks, A. Demonceau and P. Le Gendre,
Eur. J. Inorg. Chem., 2015, 2015, 2671–2682.
24 P. A. A. Klusener, L. Tip and L. Brandsma, Tetrahedron, 1991, 47,
2041–2064.
25 M. A. Bennett, T.-N. Huang, T. W. Matheson, A. K. Smith, S. Ittel
and W. Nickerson, in Inorganic Syntheses, ed. J. P. F. Jr, John
Wiley & Sons, Inc., 1982, pp. 74–78.
26 B. Therrien, T. R. Ward, M. Pilkington, C. Hoffmann, F. Gilardoni
and J. Weber, Organometallics, 1998, 17, 330–337.
27 R. Uson, A. Laguna, M. Laguna, D. A. Briggs, H. H. Murray and J.
P. Fackler, in Inorganic Syntheses, ed. H. D. Kaesz, John Wiley &
Sons, Inc., 1989, pp. 85–91.
28 S. M. Guichard, R. Else, E. Reid, B. Zeitlin, R. Aird, M. Muir, M.
Dodds, H. Fiebig, P. J. Sadler and D. I. Jodrell, Biochem.
Pharmacol., 2006, 71, 408–415.
29 F. Caruso, R. Pettinari, M. Rossi, E. Monti, M. B. Gariboldi, F.
Marchetti, C. Pettinari, A. Caruso, M. V. Ramani and G. V.
Subbaraju, J. Inorg. Biochem., 2016, 162, 44–51.
This journal is © The Royal Society of Chemistry 20xx
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The introduction of a benzoate ligand
dramatically increases the ruthenium
uptake in cells, leading to a low micromolar
IC50.