Hypoxia-Targeting Organometallic Ru(II)-Arene Complexes with Enhanced Anticancer Activity in Hypoxic Cancer Cells

Article abstract of DOI:10.1021/acs.inorgchem.8b01070

As hypoxia is an important factor to limit chemotherapeutic efficacy in tumors, we herein report three ruthenium(II)-arene complexes containing a hypoxia inducible factor-1α inhibitor (YC-1), which endow the organometallic complexes with potential for hyp

Full text of DOI:10.1021/acs.inorgchem.8b01070

Article
Cite This: Inorg. Chem. XXXX, XXX, XXX−XXX
pubs.acs.org/IC
Hypoxia-Targeting Organometallic Ru(II)−Arene Complexes with
Enhanced Anticancer Activity in Hypoxic Cancer Cells
†
,‡
†
,†,‡
†
†
†
Jian Zhao, Wanchun Li, Shaohua Gou,*
Shuang Li, Shengqiu Lin, Qianhui Wei,
,†,‡
and Gang Xu*
†
Pharmaceutical Research Center and School of Chemistry and Chemical Engineering, Southeast University, Nanjing 211189, China
Jiangsu Province Hi-Tech Key Laboratory for Biomedical Research, Southeast University, Nanjing 211189, China
‡
*
S Supporting Information
ABSTRACT: As hypoxia is an important factor to limit
chemotherapeutic efficacy in tumors, we herein report three
ruthenium(II)−arene complexes containing a hypoxia inducible
factor-1α inhibitor (YC-1), which endow the organometallic
complexes with potential for hypoxia targeting. In vitro tests
showed the resulting complexes had higher anticancer activities
in hypoxia than in normoxia against the tested cancer cell lines.
Western blot analysis revealed that complexes 1−3 blocked HIF-
1
α protein accumulation under hypoxic conditions. Moreover,
these complexes displayed much less cytotoxicity toward the
normal human umbilical vein endothelial cell line (HUVEC),
indicating that complexes 1−3 may be selectively cytotoxic for human cancer cell lines. These findings proved that ligation with
YC-1 endowed these organometallic ruthenium(II) complexes with potential for hypoxia targeting in addition to enhancing
their anticancer activities.
INTRODUCTION
■
The therapeutic values of metal-based agents have gained
increasing interest since the discovery of cisplatin in 1965.
1
Platinum(II) drugs, particularly cisplatin, carboplatin, and
2
−4
oxaliplatin, are now widely used in clinical practice,
which
exhibit their activities by covalently binding to DNA and
forming stable DNA adducts with guanines and adenine
5
,6
bases. However, their irreversible binding to DNA and the7
lack of the selectivity have resulted in severe side-effects.
Therefore, ruthenium complexes with relatively lower toxicity
were considered to be an attractive alternative to platinum
8
−16
drugs.
So far two ruthenium(III) compounds, [IndH]-
[
trans-Ru(Ind) Cl ] (KP1019, Ind = indazole) and [ImH]-
2
4
[
trans-Ru(DMSO)(Im)Cl ] (NAMIA, Im = imidazole), have
4
17,18
Figure 1. Representative anticancer ruthenium(II) complexes.
entered clinical trials,
especially KP1019 and its sodium
salt KP1339 (Figure 1), which have successfully finished a
19,20
phase I clinical trial with promising activity.
Besides,
result in the stabilization and accumulation of hypoxia
organometallic ruthenium(II)−arene complexes also show
great potential for cancer therapy because the coordination
ligands in arene−ruthenium(II) complexes can provide more
opportunity to modulate their pharmacological properties such
37,38
inducible factor-1α (HIF-1α) protein,
and the resultant
HIF-1α can dimerize with its β subunit to form a transcription
factor that binds to hypoxia response elements (HREs) and
activates a number of genes involved in vascular epidermal
21−29
as cellular accumulations and kinetic reactivities.
For
39
growth factor (VEGF) and erythropoietin (EPO). Thus,
HIF-1α plays an essential role in tumorigenesis by regulating
the expression of various genes associated with tumor
metabolism, angiogenesis, metastasis, proliferation, and differ-
examples, [(C H Ph)Ru(en)Cl][PF ] (RM175, en = ethyl-
6
5
6
i
enediamine) and [(p PrC H Me)Ru(pta)Cl ] (RAPTA-C, pta
6
4
2
=
1,3,5-triaza-7-phosphatricyclo[3.3.1.1]-decane) (Figure 1)
1
9,30−32
are currently at an advanced preclinical stage.
Hypoxia, caused by the imbalance between oxygen supply
and consumption, is an important hallmark of cancer
Received: April 18, 2018
3
3−36
progression.
The reduced oxygen levels in tumor tissues
©
XXXX American Chemical Society
A
DOI: 10.1021/acs.inorgchem.8b01070
Inorg. Chem. XXXX, XXX, XXX−XXX

Inorganic Chemistry
Article
a
Scheme 1. Preparation of Ligand L1 and Complexes 1−3
a
Reagents and conditions: (a) KOH, DMF, I , rt, 4 h; (b) benzyl bromide, t-BuOK, THF, rt, 4 h; (c) (5-formylfuran-2-yl)boronic acid, Pd(PPh ) ,
2
3 4
Na CO , DMF, 90 °C, 12 h; (d) NaBH , MeOH, rt, 2 h; (e) succinic anhydride, Et N, DMF, 4 h; (f) 5-amino-1,10-phenanthroline, HATU, Et N,
2
3
4
3
3
DMF, 60 °C, 12 h; (g) [(arene)RuCl ] , DCM, rt, 4 h.
2
2
3
9
54
entiation. Overexpression of HIF-1α was detected in more
than 90% of colon, lung, and prostate cancers, but no
expression was detected in their corresponding normal
et al. Thus, the hydrolysis behavior of complexes 1−3 was
studied using UV−vis spectroscopy. It was observed that the
hydrolysis was accompanied by a gradual increase of the
absorption band around 275 nm, which was chosen for kinetic
calculation (Figure 2). The experimental data fitted well to a
monoexponential function, and the hydrolysis rate constants
(k) and half-times (t1/2) of complexes 1−3 were shown in
Table 1. Obviously, the hydrolysis rate of complex 3 is the
fastest (t1/2 = 13.8 min) as compared with complexes 1 and 2.
The relative order of the hydrolysis rates of these complexes is
4
0
tissues. In addition, HIF-1α overexpression was found to
be associated with resistance to treatment and poor patient
prognosis in the hypoxic region around solid tumors.
Consequently, HIF-1α becomes a compelling drug target for
the treatment of tumor angiogenesis and cancer.
3
8
41−44
1
-Benzyl-3-(5′-hydroxymethyl-2′-furyl)indazole (YC-1) has
been reported as an effective HIF-1α inhibitor by stimulation
of factor inhibiting HIF (FIH)-dependent p300 dissociation
from HIF-1α,
chemotherapy drugs like cisplatin and sorafenib.
fore, we designed and prepared three organometallic
ruthenium(II) compounds bearing a YC-1 moiety, which are
expected to target the hypoxic tumor microenvironment and
exhibit their anticancer activity through multiple mechanisms
so as to improve the therapeutic efficacy of chemotherapeutic
drugs.
3
> 1 > 2, which is due to the fact that increased electron
density at Ru(II) from the aromatic ligand with more alkyl
groups can improve the Cl lability and facilitate the hydrolysis
reaction.
important role in controlling the hydrolysis rates of the arene−
Ru(II) complexes, which may further affect their biological
activities.
4
5−47
which can enhance chemosensitivity of
48,49
There-
−
55,56
The study indicated that the arene groups play an
In Vitro Cytotoxicity. The cytotoxic activities of
complexes 1−3 and ligand L1 have been investigated against
two human cancer cell lines (HCT-116 and A549 cancer cells)
and human umbilical vein endothelial cell line (HUVEC)
under normoxic or hypoxic conditions together with cisplatin
as a positive agent. Each complex was measured at eight
different concentrations against the tested cell lines, and the
corresponding IC50 values (dose required to inhibit 50%
cellular growth) were determined from dose−survival curves
RESULTS AND DISCUSSION
■
Synthesis and Characterization. Complexes 1−3 were
prepared by following the procedure shown in Scheme 1 in
which YC-1 was obtained as described previously. The
resulting arene−ruthenium(II) complexes were characterized
50
1
13
by elemental analysis and H and C NMR spectra along with
ESI−MS spectrometry (Figures S1−S11). All the spectral data
were compatible with the proposed molecular structures of
complexes 1−3.
(
Figure S12).
According to the IC50 values (Table 2), ligand L1 exhibited
little cytotoxicity under normoxia, while complexes 1−3
showed dose-dependent inhibition against the tested cell
lines with IC50 values ranging from 22.8 to 76.3 μM. Notably,
complex 3 (22.8 ± 1.2 μM) showed comparable cytotoxicity to
that of cisplatin (19.9 ± 0.9 μM) in HCT-116 cells, which may
partly attribute to its increased hydrolysis rate. Moreover, all
complexes exhibited markedly improved cytotoxic activity
Hydrolysis Studies. Hydrolysis of the organometallic
6
Ru(II)−arene complexes with the type of [( η-arene)Ru-
+
(
NN)Cl] (NN = chelating diamine ligand) is believed as a
51−53
key activation step before they bind to the biomolecules,
and a relationship between the aquation rate for arene-−Ru(II)
complexes and cytotoxicity was proposed by Wang and Sadler
B
DOI: 10.1021/acs.inorgchem.8b01070
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Figure 2. Time-dependent UV−vis spectra for the aquation of 0.05 mM complexes 1 (a), 2 (c), and 3 (e) (95% H O/5% MeOH). Absorbance−
2
time trace (275 nm) and monoexponential fit obtained for the hydrolysis of complexes 1 (b), 2 (d), and 3 (f) at 310 K.
Table 1. Hydrolysis Rate Constants (k) and Half-Times
t1/2) of Complexes 1−3 (95% H O/5% MeOH) at 310 K
under hypoxia against HCT-116 and A549 cells with HF
(hypoxia factor: the toxicity under normoxia vs hypoxia) values
range from 1.8 to 3.8, indicating that YC-1 can enhance the
antiproliferative activity of the Ru(II)−arene complexes against
hypoxic cancer cells. Additionally, significant morphological
changes to the HCT-116 cells were observed when they were
treated with complex 3 at different concentrations (12.5, 50,
and 100 μM) under hypoxia (Figure S13). In contrast,
(
2
complex
1
2
3
k (10− s−1
4
)
5.03 ± 0.23
3.89 ± 0.19
8.39 ± 0.29
t1/2 (min)
23.0
29.7
13.8
Table 2. Cytotoxicity of the Compounds against HCT-116 and A549 Cancer Cells under Normoxic and Hypoxic Conditions
IC50 values (μM)
a
b
c
HCT-116
A549
HUVEC
d
d
compound
normoxia
hypoxia
HF
normoxia
hypoxia
HF
normoxia
1
2
3
L1
65.7 ± 0.9**
57.5 ± 2.5**
22.8 ± 1.2*
112.9 ± 8.6**
19.9 ± 0.9
17.5 ± 1.4
17.9 ± 0.8
12.7 ± 0.6**
53.7 ± 4.5**
19.0 ± 1.3
3.8
3.2
1.8
2.1
1.0
76.3 ± 3.2**
65.5 ± 4.8**
36.3 ± 1.6**
147.1 ± 8.7**
20.9 ± 1.5
31.7 ± 2.5**
24.7 ± 0.9*
15.7 ± 0.6**
63.8 ± 2.3**
21.7 ± 1.3
2.4
2.7
2.3
2.3
1.0
129.1 ± 10.6**
92.6 ± 5.7**
153.2 ± 9.8**
128.9 ± 11.9**
16.6 ± 1.5
cisplatin
a
b
c
Human colorectal cancer cell line. Human nonsmall-cell lung cancer cell line. Human umbilical vein endothelial cell line. Data are expressed as
d
the mean (±SD) for three independent experiments. HF (hypoxia factor): the toxicity under normoxia vs hypoxia. *p < 0.05 and **p < 0.01
compared with the value of cisplatin.
C
DOI: 10.1021/acs.inorgchem.8b01070
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cisplatin showed similar cytotoxicity under both normoxic and
hypoxic conditions against the tested cell lines. Notably under
hypoxia, the IC50 values of complexes 1−3 were lower than
that of cisplatin against HCT-116 cells, implying that these
complexes were more active than cisplatin in hypoxic HCT-
1
16 cells. Overall, complexes 1−3 showed selective and
markedly improved cytotoxic activity against hypoxic cancer
cells, proving that the introduction of YC-1 to the Ru(II)−
arene complexes is an effective way to enhance anticancer
activity in hypoxic cancer cells.
Cellular Accumulation. The intracellular ruthenium
accumulation of complexes 1−3 in HCT-116 cells was studied
to investigate the possible relationship between cellular
accumulation and cytotoxicity. As shown in Figure 3 (Table
Figure 4. (a) HCT-116 cells treated with complexes 1−3 under
hypoxia for 12 h were examined for the expression of HIF-1α proteins
using Western blot analysis. (b) Densitometric analysis of the
expression of apoptosis-regulated proteins normalized with GAPDH.
The relative expression of each protein was represented by the density
of the protein band/density of GAPDH band. The data are
representative of three independent experiments. *p < 0.05 and **p
<
0.01 compared with the value of control.
staining and flow cytometry assay, with cisplatin as positive
control. As demonstrated in Figures 5 and 6, HCT-116 cells
exhibited numerous apoptotic cells with condensed or
Figure 3. Intracellular accumulation of complexes 1−3 in HCT-116
cells after 24 h of incubation under normoxic and hypoxic conditions.
Data are expressed as the mean (±SD) for three independent
experiments. **p < 0.01.
S1), the relative order of the cellular accumulation of
complexes 1−3 in normoxia is 3 > 1 > 2, which is in
accordance with the hydrophobicity of the arene ligands
(
hexamethylbenzene > cymene > benzene). Thus, the higher
accumulation of complex 3 may be another reason for its
superior cytotoxicity to complexes 1 and 2. While under
hypoxia, the ruthenium contents of complexes 1−3 in HCT-
1
16 cells dramatically increased, and all these complexes had
much higher Ru amount than they did under normoxia,
indicating their ability to target hypoxic cancer cells. The
results showed that the introduction of YC-1 can promote the
cellular accumulation of the Ru(II)−arene complexes in
hypoxic cancer cells and lead to more potent anticancer
activity for these compounds.
Western Blot. In order to determine the effect of these
complexes on the expression of HIF-1α under hypoxic
conditions, Western blot analysis was applied to detect the
HIF-1α protein expression in HCT-116 cells. As shown in
Figure 4, the expression of HIF-1α decreased after treatment
with complexes 1−3 and L1 that all blocked HIF-1α protein
accumulation in a dose-dependent manner. This study implied
that our metal complexes could effectively inhibit HIF-1α
protein accumulation under hypoxic conditions in HCT-116
cells.
Figure 5. Cell morphological observation for cell apoptosis induction
on the HCT-116 cells treated with complexes 1−3, L1, and cisplatin
at 30 μM for 48 h, respectively: (a) normoxia and (b) hypoxia. Cells
were stained by Hoechst 33342 (size bar = 20 μm).
Apoptosis Studies. Apoptotic analysis of complexes 1−3
and L1 against HCT-116 cells was performed under both
normoxic and hypoxic conditions by the Hoechst 33342 DNA
D
DOI: 10.1021/acs.inorgchem.8b01070
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Article
complexes containing a hypoxia inducible factor-1α inhibitor
(
YC-1) were first designed and synthesized. In vitro tests
showed the resulting complexes had higher anticancer activities
in hypoxia than in normoxia against the tested cancer cell lines,
especially complex 3, with more alkyl groups than complexes 1
and 2, exhibiting superior cytotoxicity to cisplatin under a
hypoxic condition. Significantly, these complexes are much less
cytotoxic than cisplatin against human umbilical vein
endothelial cells (HUVEC), suggesting that they have
selectivity for cancer cells over normal cells. Both kinetic
study on the hydrolysis and cellular accumulation tests
indicated that the enhancing cytotoxicity of these complexes
under either normoxia or hypoxia matched their hydrolysis
rates and cellular accumulations in HCT-116 cancer cells
positively. Moreover, complex 3 was the most active to induce
apoptosis among the tested compounds under hypoxia,
particularly in early stage apoptosis. Western blot analysis
revealed that complexes 1−3 as well as the ligand containing a
YC-1 moiety blocked HIF-1α protein accumulation in a dose-
dependent manner. These findings proved that ligation with
YC-1 endowed these organometallic ruthenium(II) complexes
with potential for hypoxia targeting in addition to enhancing
their anticancer activities. Consequently, our study provides a
useful way for further development of ruthenium(II)
anticancer agents with selective identification of cancer cells
and enhanced antitumor activity under hypoxia.
EXPERIMENTAL SECTION
Materials and Measurements. All chemicals and solvents were
■
[
Figure 6. Flow cytometry analysis for apoptosis of HCT-116 cells
induced by complexes 1−3, L1, and cisplatin at the concentration of
of analytical reagent grade and used without further purification.
6
Ru(η -arene)Cl ] (arene = cymene, benzene, and hexamethylben-
2
2
3
0 μM for 48 h: (a) normoxia and (b) hypoxia.
57,34
1
zene) and YC-2 were prepared according to previous reports.
H
1
3
and C NMR spectra were measured on Bruker Avance III-HD 600
MHz spectrometer. Mass spectra were recorded an Agilent Q-TOF
fragmented nuclei cells, while nuclei of the control cells
retained the regular round contours, exhibiting that the tested
compounds induced more apoptotic cells than the control
group. Moreover, under a hypoxic condition, cells treated with
complex 3 were more condensed and brighter than those in
other groups (Figure 5), suggesting the significant cell
apoptosis induction of complex 3 on HCT-116 cells.
The results were further verified by flow cytometry assay.
Under normoxia, the apoptotic rates of HCT-116 cells treated
with complexes 1−3 increased as compared with that of the
untreated cells (Figure 6). The apoptotic rate of complex 3
6
540 spectrometer. Elemental analysis of C, H, and N used a Vario
MICRO CHNOS elemental analyzer (Elementar). Hydrolysis study
was performed on a Shimadzu UV2600 instrument equipped with a
thermostatically controlled cell holder. Cancer cells were obtained
from Jiangsu KeyGEN BioTECH company (China). Cell apoptosis
experiments were measured by flow cytometry (FAC Scan, Becton
Dickenson) and analyzed by Cell Quest software.
Synthesis of YC-2. To a solution of YC-1 (0.84 g, 2.76 mM)
dissolved in DMF (50 mL) were added succinic anhydride (0.55 g,
5.54 mM) and TEA (0.56g, 5.53 mM). The reaction mixture was
refluxed for 4 h, and then DMF was removed under reduced pressure.
The product was purified by column chromatography (silica gel,
methanol/dichloromethane 1:9) to give YC-2. Yield: 1.04 g (93.3%).
White solid. Anal. Calcd (%) for C H N O : C 68.31, H 4.98, N
6.93. Found: C 68.20, H 4.91, N 6.79; H NMR (600 MHz, DMSO)
δ 2.50−2.52 (m, 2H), 2.57−2.59 (m, 2H), 5.19 (s, 2H), 5.73 (s, 2H),
(
44.88%) was compatible to that of cisplatin (46.87%), and the
relative order of inducing apoptosis against HCT-116 cells is
2
3
20
2
5
1
cisplatin (46.87%) ≈ 3 (44.88%) > 1 (35.87%) ≈ 2 (32.54%)
>
L1 (19.69%), which is in line with the result of the
6
7
1
.71−6.72 (d, 1H, J = 3.4 Hz), 7.02−7.03 (d, 1H, J = 3.4 Hz), 7.24−
.28 (m, 4H), 7.30−7.33 (m, 2H), 7.45−7.47 (m, 1H), 7.76−7.77 (d,
H, J = 8.5 Hz), 8.10−8.12 (d, 1H, J = 8.2 Hz), 12.24 (s, 1H) ppm.
Synthesis of L1. YC-2 (0.50 g, 1.24 mM), HATU (0.51 g, 1.34
cytotoxicity assay to some extent. Once hypoxia was
conducted, both the complexes and their ligand L1 induced
an improved incidence of early to late stage apoptosis in HCT-
1
16 cells compared with that in normoxia. Notably, complex 3
mM), TEA (0.14g, 1.40 mM), and DMF (7.5 mL) were added into a
5 mL round-bottom flask. The mixture was stirred at room
had a higher apoptotic rate than cisplatin, proving that it had
advantages over cisplatin to induce cell apoptosis under a
hypoxic condition. This further verified the considerable
antiproliferative effect of complex 3 in hypoxic HCT-116 cells.
2
temperature for 2 h. Then 5-amino-1,10-phenanthroline (0.20 g,
1.02 mM) was added, and the resulting mixture was stirred at 60 °C
for 12 h. The solvent was then removed by evaporation under reduced
pressure. Column chromatography (eluent 30:1 DCM/methanol)
gave L1. Yield: 0.32 g (54.0%). Light yellow solid. Anal. Calcd (%) for
C H N O : C 72.28, H 4.68, N 12.04. Found: C 72.20, H 4.81, N
CONCLUSION
■
3
5
27
5
4
In view of hypoxia as an important factor to limit chemo-
therapeutic efficacy in clinical practice, we purposed to develop
organometallic ruthenium(II)−arene complexes targeting the
hypoxic tumor microenvironment so as to enhance their
anticancer activity. In this study, three ruthenium(II)−arene
1
1
2
1.96; H NMR (600 MHz, DMSO) δ 2.77−2.79 (m, 2H), 2.88−
.90 (m, 2H), 5.24 (s, 2H), 5.70 (s, 2H), 6.74−6.75 (d, 1H, J = 3.4
Hz), 7.01−7.02 (d, 1H, J = 3.3 Hz), 7.18−7.26 (m, 4H), 7.29−7.31
(m, 2H), 7.39−7.42 (m, 1H), 7.72−7.74 (d, 1H, J = 8.2 Hz), 7.75−
7.77 (q, 1H, J = 4.4 Hz), 7.84−7.86 (q, 1H, J = 4.4 Hz), 8.08−8.09
E
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(
d, 1H, J = 8.2 Hz), 8.18 (s, 1H), 8.45−8.47 (1H, d-d, J = 8.1, 1.4
MTT Assay. Cytotoxicity of complexes 1−3, ligand L1, and
cisplatin against HCT-116, A549, and HUVEC cells was determined
by means of the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl
tetrazolium bromide) assay. Cells (5000−10000 per well) with better
vitality were seeded in 96-well plates. The compounds were dissolved
by DMF (water for cisplatin) and diluted with medium to various
concentrations (the final concentration of DMF was less than 0.4%).
After being incubated under a normoxic condition (20% O , 5% CO ,
Hz), 8.69−8.70 (d, 1H, J = 7.4 Hz), 9.01 (m, 1H), 9.03−9.04 (1H, d-
d, J = 4.3, 1.6 Hz), 9.12−9.13 (1H, d-d, J = 4.2, 1.5 Hz), 10.27 (m,
1
H) ppm.
General Procedure for Synthesis of Complexes 1−3. A solution of
L1 (0.29 g, 0.50 mM) in CH Cl (10 mL) was added to a suspension
of [Ru(arene)Cl ] (0.21 mM) in CH Cl (15 mL) dropwise. The
mixture was stirred at room temperature for 8 h. Then the reaction
mixture was concentrated to 3 mL. The crude product was separated,
2
2
2
2
2
2
2
2
and 75% N ) or hypoxic condition (1% O , 5% CO , and 94% N ) at
2
2
2
2
washed with Et O (3 × 10 mL) and cold methanol (3 × 10 mL), and
37 °C for 72 h, cells were stained with MTT (5 mg/mL) for another
5 h, and then the medium was thrown away and replaced by 150 mL
of DMSO. The inhibition of cell growth induced by the tested
complexes was detected by measuring the absorbance of each well at
570/630 nm using enzyme labeling instrument. The IC50 values were
calculated by SPSS software after three parallel experiments.
2
dried in a vacuum-dryer.
Complex 1. Yield: 0.23 g (61.3%). Yellow-green powder. Anal.
Calcd (%) for C H Cl N O Ru: C 60.88, H 4.65, N 7.89. Found: C
4
5
41
2
5
4
+
1
6
(
2
2
0.73, H 4.74, N 7.66. ESI−MS: m/z [M − Cl] = 852.2. H NMR
600 MHz, DMSO) δ 0.89−0.91 (t, 6H, J = 6.6 Hz), 2.17 (s, 3H),
.59−2.64 (m, 1H), 2.76−2.78 (m, 2H), 2.95−2.97 (m, 2H), 5.23 (s,
H), 5.71 (s, 2H), 6.12−6.15 (m, 2H), 6.36−6.38 (t, 2H,, J = 6.4
Cellular Accumulation. HCT-116 cells were seeded in six-well
6
plates at a density of 10 cells/well. After overnight incubation, 50 μM
Hz), 6.73−6.74 (d, 1H, J = 3.4 Hz), 7.02−7.03 (d, 1H, J = 3.4 Hz),
complexes 1−3 were added, respectively. After incubation under a
7
7
8
.19−7.26 (m, 4H), 7.30−7.32 (m, 2H), 7.39−7.42 (m, 1H), 7.73−
.75 (d, 1H, J = 8.5 Hz), 8.06−8.11 (m, 2H), 8.18−8.21 (m, 1H),
.48 (s, 1H), 8.79−8.80 (d, 1H, J = 8.0 Hz), 9.27−9.28 (d, 1H, J = 8.0
normoxic condition (20% O , 5% CO , and 75% N ) or hypoxic
2
2
2
condition (1% O , 5% CO , and 94% N ) at 37 °C for 24 h, cells were
2
2
2
collected and washed three times with ice-cold PBS, then centrifuged
at 1000 rpm for 10 min and resuspended in 1 mL of PBS. A volume of
100 μL was taken out to determine the cell density. Then the
1
3
Hz), 9.89 (m, 1H), 10.02 (m, 1H), 11.12 (m, 1H) ppm; C NMR
150 MHz, DMSO-d ) δ 18.69, 22.12, 22.14, 29.17, 30.86, 31.23,
(
6
5
1
1
1
1
2.44, 58.31, 84.23, 84.39, 86.42, 86.57, 103.23, 104.48, 108.41,
10.75, 113.28, 120.73, 121.50, 122.14, 126.05, 126.45, 126.85,
27.37, 127.73, 128.05, 129.08, 130.12, 133.95, 135.48, 135.65,
37.76, 138.50, 140.77, 143.18, 145.87, 149.06, 149.59, 155.35,
56.60, 172.02, 172.61 ppm.
remaining cells were digested by HNO (200 μL, 65%) at 65 °C for 4
h. The Ru level in cells was measured by ICP−MS after three parallel
experiments.
3
Apoptosis Assessment by Hoechst 33342 Staining. HCT-116
5
cells were seeded in 24-well plates at 1 × 10 cells/well and incubated
Complex 2. Yield: 0.21 g (59.6%). Yellow-green powder. Anal.
overnight. Cells were incubated with 30 μM of the tested compounds
Calcd (%) for C H Cl N O Ru: C 59.21, H 4.00, N 8.42. Found: C
under a normoxic condition (20% O , 5% CO , and 75% N ) or
41
33
2
5
4
2
2
2
+
1
5
9.03, H 4.14, N 8.20. ESI−MS: m/z [M − Cl] = 796.2. H NMR
hypoxic condition (1% O , 5% CO , and 94% N ) at 37 °C for 48 h.
2 2 2
(
2
7
600 MHz, DMSO) δ 2.82−2.84 (m, 2H), 3.04 (m, 2H), 5.28 (s,
Then the cells were rinsed twice in PBS and stained with Hoechst
33342 fluorescent dye for 10 min in the dark at 37 °C. Cell apoptosis
was examined under the fluorescence microscope with excitation
wavelength of 330−380 nm, and data were collected from three
independent experiments.
H), 5.76 (s, 2H), 6.40 (s, 6H), 6.79−6.80 (d, 1H, J = 3.1 Hz), 7.06−
.07 (d, 1H, J = 3.1 Hz), 7.22−7.25 (d, 1H, J = 7.5 Hz), 7.29−7.38
(
m, 5H), 7.44−7.46 (m, 1H), 7.78−7.80 (d, 1H, J = 8.5 Hz), 8.11 (m,
1H), 8.13−8.15 (d, 1H, J = 8.2 Hz), 8.23 (m, 1H), 8.50 (s, 1H),
.82−8.83 (d, 1H, J = 8.0 Hz), 9.30−9.31 (d, 1H, J = 8.2 Hz), 10.03
8
Apoptosis Analysis by Flow Cytometry. HCT-116 cells were
1
3
5
(
m, 1H), 10.17 (m, 1H), 11.18 (m, 1H) ppm; C NMR (150 MHz,
grown in a six-well plate at a density of 2 × 10 cells/well and cultured
DMSO-d ) δ 29.19, 31.32, 52.41, 58.29, 87.11, 108.38. 110.71,
overnight. The tested complexes were added, which were diluted to a
concentration of 30 μM. After incubation under a normoxic condition
(20% O , 5% CO , and 75% N ) or hypoxic condition (1% O , 5%
6
1
1
1
1
13.29, 120.69, 121.49, 122.11, 125.85, 126.56, 126.65, 127.35,
27.73, 128.04, 129.07, 130.12, 133.92, 135.46, 135.77, 137.76,
38.46, 140.73, 143.33, 146.02, 149.06, 149.57, 155.56, 156.82,
72.03, 172.61 ppm.
2
2
2
2
CO , and 94% N ) for 48 h, cells were collected by centrifugation (5
2
2
min, 25 °C, 2000 rpm). Then, the cells were washed twice with cold
water and resuspended in binding buffer (10 mM HEPES, 140 mM
Complex 3. Yield: 0.25 g (65.2%). Yellow-green powder. Anal.
Calcd (%) for C H Cl N O Ru: C 61.64, H 4.95, N 7.65. Found: C
NaCl, 2.5 mM CaCl , pH 7.4). The cells were stained with 5 μL of
47
45
2
5
4
2
+
1
6
1.38, H 5.11, N 7.53. ESI−MS: m/z [M − Cl] = 880.3. H NMR
Annexin V- FITC and then with 5 μL of propidium iodide (20 μg/
mL) for 15 min in the dark at room temperature. The fluorescence of
cells was detected by an annexin V-FITC apoptosis detection kit
(Roche) according to the manufacturer’s protocol, and cells were
quantified by system software (Cell Quest; BD Biosciences).
Western Blot. HCT-116 cells were grown in a six-well plate at a
(
(
7
2
600 MHz, DMSO) δ 2.10 (s, 18H), 2.75−2.77 (m, 2H), 2.98−3.00
m, 2H), 5.21 (s, 2H), 5.69 (s, 2H), 6.72−6.73 (d, 1H, J = 2.9 Hz),
.00−7.01 (d, 1H, J = 2.9 Hz), 7.18−7.25 (m, 4H), 7.28−7.31 (m,
H), 7.38−7.40 (m, 1H), 7.72−7.73 (d, 1H, J = 8.4 Hz), 8.08−8.09
(
9
m, 2H), 8.16 (m, 1H), 8.45 (s, 1H), 8.73−8.75 (d, 1H, J = 7.8 Hz),
13
5
.24−9.27 (m, 2H), 9.37−9.38 (m, 1H), 11.16 (m, 1H) ppm;
C
density of 2 × 10 cells/well and cultured until the cell density
NMR (150 MHz, DMSO-d ) δ 15.75, 29.19, 31.23, 52.42, 58.29,
reached 80%. The solutions of complexes 1−3 and L1 were diluted
down in media to give the required concentration (0.1 , 1 , or 5 μM)
for addition to the cells, and the cells were cultured under a normoxic
condition (20% O , 5% CO , and 75% N ) or hypoxic condition (1%
6
9
1
1
1
5.81, 108.41. 110.75, 113.28, 120.71, 121.50, 122.14, 126.24, 126.27,
27.06, 127.36, 127.73, 128.05, 129.07, 129.87, 134.05, 135.36,
35.46, 137.75, 138.05, 140.74, 143.23, 145.92, 149.04, 149.58,
53.05, 154.24, 172.06, 172.62 ppm.
2
2
2
O , 5% CO , and 94% N ) for 12 h at 37 °C. HCT-116 cells were
2
2
2
Hydrolysis Studies. Hydrolysis of complexes 1−3 was recorded on
lysed in cell lysis buffer and collected by centrifugation at 13 000 rpm
for 20 min at 4 °C. Proteins from cell lysates were separated by 8−
12% sodium dodecyl sulfate−polyacrylamide gel electrophoresis
(SDS-PAGE) and transferred onto a polyvinylidine difluoride
(PVDF) membrane (Amersham Biosciences). The membrane was
blocked with PBST containing 5% nonfat dry milk for 1 h and further
incubated with monoclonal antihuman HIF-1α antibody (Santa Cruz
Biotechnology, USA) overnight at 4 °C under gentle shaking. After
that, the membrane was incubated with the secondary antibody
(1:2000) for 1 h at RT (25 °C). Protein blots were detected with
chemiluminescence reagent (Thermo Fischer Scientifics Ltd.).
GAPDH was used as loading control.
a Shimadzu UV2600 instrument equipped with a thermostatically
controlled cell holder. The UV−vis spectra were recorded by scanning
from 185 to 600 nm every 5 min at 37 °C, and 275 nm was selected
for the kinetic study. The time-dependent absorbance was fitted using
Origin 9.0 to give the first order rate constant k.
Cell Culture. HCT-116 (human colorectal cancer cell line) and
A549 (human nonsmall cell lung cancer cell line) were maintained in
a humidified atmosphere of 5% CO at 37 °C. Cells were cultured in
2
RPMI-1640 medium with 10% fetal bovine serum (FBS). All media
were also supplemented with 100 mg/mL of penicillin and 100 mg/
mL of streptomycin.
F
DOI: 10.1021/acs.inorgchem.8b01070
Inorg. Chem. XXXX, XXX, XXX−XXX

Inorganic Chemistry
Article
Statistical Analysis. Differences among samples were calculated
with the two-tailed Student’s t-test using an independent samples t-
test in SPSS 16.0. Differences among groups were considered
statistically significant at P < 0.05.
single molecule compounds to nanomaterials. Chem. Soc. Rev. 2017,
46, 5771−5804.
(12) Hartinger, C. G.; Zorbas-Seifried, S.; Jakupec, M. A.; Kynast, B.;
Zorbas, H.; Keppler, B. K. From bench to bedside-preclinical and
early clinical development of the anticancer agent indazolium trans-
[tetrachlorobis (1H-indazole) ruthenate (III)](KP1019 or FFC14A).
J. Inorg. Biochem. 2006, 100, 891−904.
ASSOCIATED CONTENT
Supporting Information
■
*
S
(13) Han Ang, W.; Dyson, P. J. Classical and non-classical
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.inorg-
chem.8b01070.
ruthenium-based anticancer drugs: Towards targeted chemotherapy.
Eur. J. Inorg. Chem. 2006, 20, 4003−4018.
(14) Zhao, J.; Zhang, D.; Hua, W.; Li, W.; Xu, G.; Gou, S.
1
13
H and C NMR and ESI−MS spectra of complexes 1−
Anticancer Activity of Bifunctional Organometallic Ru (II) Arene
Complexes Containing a 7-Hydroxycoumarin Group. Organometallics
3
, dose-dependent cell viability curves, and cellular
2
(
018, 37, 441−447.
accumulation data (PDF)
15) Tian, M.; Li, J.; Zhang, S.; Guo, L.; He, X.; Kong, D.; Zhang,
̂
H.; Liu, Z. Half-sandwich ruthenium (ii) complexes containing N N-
chelated imino-pyridyl ligands that are selectively toxic to cancer cells.
Chem. Commun. 2017, 53, 12810−12813.
(16) Li, J.; Tian, M.; Tian, Z.; Zhang, S.; Yan, C.; Shao, C.; Liu, Z.
Half-Sandwich Iridium (III) and Ruthenium (II) Complexes
AUTHOR INFORMATION
Corresponding Authors
E-mail: sgou@seu.edu.cn.
E-mail: xugang@seu.edu.cn.
■
*
*
̂
Containing PP-Chelating Ligands: A New Class of Potent Anticancer
ORCID
Agents with Unusual Redox Features. Inorg. Chem. 2018, 57, 1705−
Shaohua Gou: 0000-0003-0284-5480
Notes
1
(
716.
17) Hartinger, C. G.; Jakupec, M. A.; Zorbas-Seifried, S.; Groessl,
The authors declare no competing financial interest.
M.; Egger, A.; Berger, W.; Zorbas, H.; Dyson, P. J.; Keppler, B. K.
KP1019, a new redox-active anticancer agent-preclinical development
and results of a clinical phase I study in tumor patients. Chem.
Biodiversity 2008, 5, 2140−2155.
ACKNOWLEDGMENTS
■
We are grateful to the National Natural Science Foundation of
China (Grant No. 21601034) and Jiangsu Province Natural
Science Foundation (Grant No. BK20160664) for financial
aids to this work. We also thank the Fundamental Research
Funds for the Central Universities (Project 2242016K30020
and 2242017K41025) and Priority Academic Program
Development of Jiangsu Higher Education Institutions for
the construction of fundamental facilities are also appreciated.
(
18) Bergamo, A.; Sava, G. Ruthenium anticancer compounds:
myths and realities of the emerging metal-based drugs. Dalton Trans.
011, 40, 7817−7823.
19) Riedl, C. A.; Flocke, L. S.; Hejl, M.; Roller, A.; Klose, M. H.;
2
(
Jakupec, M. A.; Kandioller, W.; Keppler, B. K. Introducing the 4-
Phenyl-1, 2, 3-Triazole Moiety as a Versatile Scaffold for the
Development of Cytotoxic Ruthenium (II) and Osmium (II) Arene
Cyclometalates. Inorg. Chem. 2017, 56, 528−541.
(20) Trondl, R.; Heffeter, P.; Kowol, C. R.; Jakupec, M. A.; Berger,
REFERENCES
W.; Keppler, B. K. NKP-1339, the first ruthenium-based anticancer
drug on the edge to clinical application. Chem. Sci. 2014, 5, 2925−
2932.
■
(
1) Rosenberg, B.; Van Camp, L.; Krigas, T. Inhibition of cell
division in Escherichia coli by electrolysis products from a platinum
electrode. Nature 1965, 205, 698−699.
(21) Guerriero, A.; Oberhauser, W.; Riedel, T.; Peruzzini, M.;
Dyson, P. J.; Gonsalvi, L. New Class of Half-Sandwich Ruthenium
(II) Arene Complexes Bearing the Water-Soluble CAP Ligand as an
in Vitro Anticancer Agent. Inorg. Chem. 2017, 56, 5514−5518.
(22) Wang, F.; Chen, H.; Parsons, S.; Oswald, I. D.; Davidson, J. E.;
Sadler, P. J. Kinetics of Aquation and Anation of Ruthenium (II)
Arene Anticancer Complexes, Acidity and X-ray Structures of Aqua
Adducts. Chem. - Eur. J. 2003, 9, 5810−5820.
(
2) Wheate, N. J.; Walker, S.; Craig, G. E.; Oun, R. The status of
platinum anticancer drugs in the clinic and in clinical trials. Dalton
Trans. 2010, 39, 8113−8127.
(
3) Johnstone, T. C.; Suntharalingam, K.; Lippard, S. J. The Next
generation of platinum drugs: targeted Pt (II) agents, nanoparticle
delivery, and Pt (IV) prodrugs. Chem. Rev. 2016, 116, 3436−3486.
(
4) Kelland, L. The resurgence of platinum-based cancer chemo-
therapy. Nat. Rev. Cancer 2007, 7, 573−584.
5) Todd, R. C.; Lippard, S. J. Inhibition of transcription by
platinum antitumor compounds. Metallomics 2009, 1, 280−291.
6) Zhao, J.; Wang, D.; Xu, G.; Gou, S. Improve the anticancer
(23) Batchelor, L. K.; Paunescu, E.; Soudani, M.; Scopelliti, R.;
(
Dyson, P. J. Influence of the Linker Length on the Cytotoxicity of
Homobinuclear Ruthenium (II) and Gold (I) Complexes. Inorg.
Chem. 2017, 56, 9617−9633.
(
potency of the platinum (II) complexes through functionalized
(24) Ma, L.; Ma, R.; Wang, Z.; Yiu, S.-M.; Zhu, G. Heterodinuclear
leaving group. J. Inorg. Biochem. 2017, 175, 20−28.
Pt (IV)-Ru (II) anticancer prodrugs to combat both drug resistance
(
7) Galanski, M.; Keppler, B. K. Searching for the magic bullet:
and tumor metastasis. Chem. Commun. 2016, 52, 10735−10738.
anticancer platinum drugs which can be accumulated or activated in
(25) Prior, T. J.; Savoie, H.; Boyle, R. W.; Murray, B. S. pH-
the tumor tissue. Anti-Cancer Agents Med. Chem. 2007, 7, 55−73.
Dependent Modulation of Reactivity in Ruthenium (II) Organo-
(
8) Yan, Y. K.; Melchart, M.; Habtemariam, A.; Sadler, P. J.
metallics. Organometallics 2018, 37, 294−297.
Organometallic chemistry, biology and medicine: ruthenium arene
(26) Caruso, F.; Pettinari, R.; Rossi, M.; Monti, E.; Gariboldi, M. B.;
anticancer complexes. Chem. Commun. 2005, 0, 4764−4776.
Marchetti, F.; Pettinari, C.; Caruso, A.; Ramani, M. V.; Subbaraju, G.
V. The in vitro antitumor activity of arene-ruthenium (II)
curcuminoid complexes improves when decreasing curcumin polarity.
J. Inorg. Biochem. 2016, 162, 44−51.
(
9) Meier-Menches, S. M.; Gerner, C.; Berger, W.; Hartinger, C. G.;
Keppler, B. K. Structure-activity relationships for ruthenium and
osmium anticancer agents-towards clinical development. Chem. Soc.
Rev. 2018, 47, 909−928.
(27) Ma, L.; Lin, X.; Li, C.; Xu, Z.; Chan, C.-Y.; Tse, M.-K.; Shi, P.;
(
10) Alessio, E.; Mestroni, G.; Bergamo, A.; Sava, G. Ruthenium
antimetastatic agents. Curr. Top. Med. Chem. 2004, 4, 1525−1535.
11) Zeng, L.; Gupta, P.; Chen, Y.; Wang, E.; Ji, L.; Chao, H.; Chen,
Z.-S. The development of anticancer ruthenium (II) complexes: from
Zhu, G. A cancer cell-selective and low-toxic bifunctional hetero-
dinuclear Pt(IV)-Ru(II) anticancer prodrug. Inorg. Chem. 2018, 57,
2917−2924.
(
G
DOI: 10.1021/acs.inorgchem.8b01070
Inorg. Chem. XXXX, XXX, XXX−XXX

Inorganic Chemistry
Article
(
28) Paunescu, E.; Soudani, M.; Martin, P.; Scopelliti, R.; Lo Bello,
transcription 3 (STAT3) in hepatocellular carcinoma. Mol. Cancer
2014, 13, 7.
M.; Dyson, P. J. Organometallic Glutathione S-Transferase Inhibitors.
Organometallics 2017, 36, 3313−3321.
(49) Lau, C. K.; Yang, Z. F.; Lam, S. P.; Lam, C. T.; Ngai, P.; Tam,
K. H.; Poon, R. T.-P.; Fan, S.-T. Inhibition of Stat3 activity by YC-1
enhances chemo-sensitivity in hepatocellular carcinoma. Cancer Biol.
Ther. 2007, 6, 1900−1907.
(
29) Zhang, Y.; Zheng, W.; Luo, Q.; Zhao, Y.; Zhang, E.; Liu, S.;
Wang, F. Dual-targeting organometallic ruthenium(II) anticancer
complexes bearing EGFR-inhibiting 4-anilinoquinazoline ligands.
Dalton Trans 2015, 44, 13100−13111.
(50) Xiao, J.; Jin, C.; Liu, Z.; Guo, S.; Zhang, X.; Zhou, X.; Wu, X.
The design, synthesis, and biological evaluation of novel YC-1
derivatives as potent anti-hepatic fibrosis agents. Org. Biomol. Chem.
(
30) Guichard, S.; Else, R.; Reid, E.; Zeitlin, B.; Aird, R.; Muir, M.;
Dodds, M.; Fiebig, H.; Sadler, P.; Jodrell, D. Anti-tumour activity in
non-small cell lung cancer models and toxicity profiles for novel
ruthenium (II) based organo-metallic compounds. Biochem. Pharma-
col. 2006, 71, 408−415.
2
(
015, 13, 7257−7264.
51) Zheng, W.; Luo, Q.; Lin, Y.; Zhao, Y.; Wang, X.; Du, Z.; Hao,
X.; Yu, Y.; Lv, S.; Ji, L. Complexation with organometallic ruthenium
pharmacophores enhances the ability of 4-anilinoquinazolines
inducing apoptosis. Chem. Commun. 2013, 49, 10224−10226.
(
31) Scolaro, C.; Bergamo, A.; Brescacin, L.; Delfino, R.; Cocchietto,
M.; Laurenczy, G.; Geldbach, T. J.; Sava, G.; Dyson, P. J. In vitro and
in vivo evaluation of ruthenium(II)-arene PTA complexes. J. Med.
Chem. 2005, 48, 4161−4171.
(52) Du, J.; Zhang, E.; Zhao, Y.; Zheng, W.; Zhang, Y.; Lin, Y.;
Wang, Z.; Luo, Q.; Wu, K.; Wang, F. Discovery of a dual-targeting
organometallic ruthenium complex with high activity inducing early
stage apoptosis of cancer cells. Metallomics 2015, 7, 1573−1583.
(
32) Murray, B. S.; Babak, M. V.; Hartinger, C. G.; Dyson, P. J. The
development of RAPTA compounds for the treatment of tumors.
(53) Liu, Z.; Habtemariam, A.; Pizarro, A. M.; Fletcher, S. A.;
Coord. Chem. Rev. 2016, 306, 86−114.
Kisova, A.; Vrana, O.; Salassa, L.; Bruijnincx, P. C. A.; Clarkson, G. J.;
Brabec, V.; Sadler, P. J. Organometallic Half-sandwich Iridium
Anticancer Complexes. J. Med. Chem. 2011, 54, 3011−3026.
(
33) Moyer, M. W. Targeting hypoxia brings breath of fresh air to
cancer therapy. Nat. Med. 2012, 18, 636−637.
34) Xu, Z.; Zhao, J.; Gou, S.; Xu, G. Novel hypoxia-targeting Pt
IV) prodrugs. Chem. Commun. 2017, 53, 3749−3752.
35) Wilson, W. R.; Hay, M. P. Targeting hypoxia in cancer therapy.
Nat. Rev. Cancer 2011, 11, 393−410.
36) Purkait, K.; Karmakar, S.; Bhattacharyya, S.; Chatterjee, S.;
(
(54) Wang, F.; Habtemariam, A.; van der Geer, E. P.; Fernandez, R.;
(
Melchart, M.; Deeth, R. J.; Aird, R.; Guichard, S.; Fabbiani, F. P.;
Lozano-Casal, P. Controlling ligand substitution reactions of organo-
metallic complexes: tuning cancer cell cytotoxicity. Proc. Natl. Acad.
Sci. U. S. A. 2005, 102, 18269−18274.
(
(
Dey, S. K.; Mukherjee, A. A hypoxia efficient imidazole-based Ru(II)
arene anticancer agent resistant to deactivation by glutathione. Dalton
Trans. 2015, 44, 5969−5973.
(55) Dougan, S. J.; Melchart, M.; Habtemariam, A.; Parsons, S.;
Sadler, P. J. Phenylazo-pyridine and Phenylazo-pyrazole Chlorido
Ruthenium(II) Arene Complexes: Arene Loss, Aquation, and Cancer
Cell Cytotoxicity. Inorg. Chem. 2006, 45, 10882−10894.
(
37) Ryan, H. E.; Lo, J.; Johnson, R. S. HIF-1α is required for solid
tumor formation and embryonic vascularization. EMBO J. 1998, 17,
005−3015.
38) Bos, R.; van der Groep, P.; Greijer, A. E.; Shvarts, A.; Meijer, S.;
(56) Bugarcic, T.; Habtemariam, A.; Deeth, R. J.; Fabbiani, F. P. A.;
3
(
Parsons, S.; Sadler, P. J. Ruthenium(II) arene anticancer complexes
with redox-active diamine ligands. Inorg. Chem. 2009, 48, 9444−9453.
Pinedo, H. M.; Semenza, G. L.; van Diest, P. J.; van der Wall, E.
Levels of hypoxia-inducible factor-1α independently predict prognosis
in patients with lymph node negative breast carcinoma. Cancer 2003,
(57) Bennett, M. A.; Smith, A. K. Arene ruthenium(II) complexes
formed by dehydrogenation of cyclohexadienes with ruthenium(III)
trichloride. J. Chem. Soc., Dalton Trans. 1974, 2, 233−241.
9
(
7, 1573−1581.
39) Harris, A. L. Hypoxia - a key regulatory factor in tumour
growth. Nat. Rev. Cancer 2002, 2, 38−47.
40) Semenza, G. L. Expression of hypoxia-inducible factor 1:
mechanisms and consequences. Biochem. Pharmacol. 2000, 59, 47−53.
41) Xia, Y.; Choi, H.-K.; Lee, K. Recent advances in hypoxia-
inducible factor (HIF)-1 inhibitors. Eur. J. Med. Chem. 2012, 49, 24−
0.
42) Semenza, G. L. Targeting HIF-1 for cancer therapy. Nat. Rev.
Cancer 2003, 3, 721−732.
43) Wang, J.; Kou, J.; Hou, X.; Zhao, Z.; Chao, H. A ruthenium(II)
(
(
4
(
(
anthraquinone complex as the theranostic agent combining hypoxia
imaging and HIF-1α inhibition. Inorg. Chim. Acta 2017, 454, 176−
1
(
83.
44) Lv, W.; Zhang, Z.; Zhang, K. Y.; Yang, H.; Liu, S.; Xu, A.; Guo,
S.; Zhao, Q.; Huang, W. A Mitochondria-targeted Photosensitizer
Showing Improved Photodynamic Therapy Effects Under Hypoxia.
Angew. Chem., Int. Ed. 2016, 55, 9947−9951.
(
45) Kim, H.-L.; Yeo, E.-J.; Chun, Y.-S.; Park, J.-W. A domain
responsible for HIF-1α degradation by YC-1, a novel anticancer agent.
Int. J. Oncol. 2006, 29, 255−260.
(
46) Shin, D. H.; Kim, J.-H.; Jung, Y.-J.; Kim, K.-E.; Jeong, J. M.;
Chun, Y.-S.; Park, J.-W. Preclinical evaluation of YC-1, a HIF
inhibitor, for the prevention of tumor spreading. Cancer Lett. 2007,
2
(
55, 107−116.
47) Chen, C.-J.; Hsu, M.-H.; Huang, L.-J.; Yamori, T.; Chung, J.-G.;
Lee, F.-Y.; Teng, C.-M.; Kuo, S.-C. Anticancer mechanisms of YC-1 in
human lung cancer cell line, NCI-H226. Biochem. Pharmacol. 2008,
7
(
5, 360−368.
48) Kong, J.; Kong, F.; Gao, J.; Zhang, Q.; Dong, S.; Gu, F.; Ke, S.;
Pan, B.; Shen, Q.; Sun, H. YC-1 enhances the anti-tumor activity of
sorafenib through inhibition of signal transducer and activator of
H
DOI: 10.1021/acs.inorgchem.8b01070
Inorg. Chem. XXXX, XXX, XXX−XXX