Discovery of high in vitro and in vivo antitumor activities of organometallic ruthenium(ii)-arene complexes with 5,7-dihalogenated-2-methyl-8-quinolinol

Article abstract of DOI:10.1039/C9DT00866G

This paper reports the synthesis, structure characterization, and anticancer properties of 13 organometallic Ru(ii)-arene complexes: [Ru(η⁶-p-cymene)Cl-(L1)] (1), [Ru(η⁶-p-cymene)Cl-(L2)] (2), [Ru(η⁶-p-cymene)Cl-(L3)] (3), [Ru(η⁶-p-cymene)Cl-(L4)] (4), [Ru(η⁶-p-cymene)Cl-(L5)] (5), [Ru(η⁶-p-cymene)I-(L1)] (6), [Ru(η⁶-p-cymene)I-(L2)] (7), [Ru(η⁶-p-cymene)I-(L3)] (8), [Ru(η⁶-p-cymene)I-(L4)] (9), [Ru(η⁶-p-cymene)I-(L5)] (10), [Ru(η⁶-p-cymene)I-(L6)] (11), [Ru(η⁶-p-cymene)I-(L7)] (12), and [Ru(η⁶-p-cymene)Cl-(L8)] (13) respectively containing deprotonated 5,7-dichloro-2-methyl-8-quinolinol (H-L1), 5,7-dibromo-2-methyl-8-quinolinol (H-L2), 5-chloro-7-iodo-8-hydroxy-quinoline (H-L3), 5,7-dibromo-8-quinolinol (H-L4), 5,7-diiodo-8-hydroxyquinoline (H-L5), 8-hydroxy-2-methylquinoline (H-L6), 2,8-quinolinediol (H-L7), or 6,7-dichloro-5,8-quinolinedione (H-L8). MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay showed that 13 organometallic Ru(ii)-arene complexes 1-13 are more selective for HeLa cells than normal HL-7702 cells. In addition, 1, 2, 5, and 6, which contain the active ligands H-L1 and H-L2, showed remarkable cell cytotoxicity, giving the respective IC₅₀ values of 2.00 ± 0.20 nM, 0.89 ± 0.62 μM, 25.00 ± 0.30 nM, and 2.18 ± 0.35 μM on HeLa cancer cells. These values indicated higher activity than 6,7-dichloro-5,8-quinolinedione and other 8-hydroxyquinoline derivative Ru(ii)-arene complexes. Interestingly, all these Ru(ii)-arene complexes 1-13 were significantly less toxic to human hepatic (HL-7702) cells. Moreover, 1- and 2-induced HeLa cell apoptosis was mediated by the inhibition of telomerase activity and dysfunction of mitochondria, and resulted in DNA damage and increased anti-migration activity on HeLa cells. The organometallic Ru(ii)-arene complex 1 exhibited evident priority to the antitumor activity compared to 2, which should be highly associated with the key roles of the 5,7-dichloro substituted groups in the L1 ligand of organometallic Ru(ii)-arene complexes 1. Remarkably, 1 showed higher inhibitory activity against the xenograft tumor growth of human cervical cells (HeLa) in vivo (tumor growth inhibition rate (TGIR) = 58.5%) than cisplatin. This study was the first to show that the 5,7-dihalogenated-2-methyl-8-quinolinol organometallic Ru(ii)-arene complexes 1 and 2 are novel Ru(ii) anticancer drug candidates.

Full text of DOI:10.1039/C9DT00866G

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ARTICLE
Discovery of a high in vitro and in vivo antitumor activities of organometallic
ruthenium(II)-arene complexes with 5,7-dihalogenated-2-methyl-8-quinolinol
Received 00th January 20xx,
Accepted 00th January 20xx
Ting Menga, Qi-Pin Qina,b,*, Zi-Lu Chena, Hua-Hong Zoua,*, Kai Wanga,c, and Fu-Pei Lianga,c,*
This paper reports the synthesis, structures characterization, and anticancer properties of 13 organometallic Ru(II)-arene
complexes: [Ru(η6-p-cymene)Cl-(L1)] (1), [Ru(η6-p-cymene)Cl-(L2)] (2), [Ru(η6-p-cymene)Cl-(L3)] (3), [Ru(η6-p-
cymene)Cl-(L4)] (4), [Ru(η6-p-cymene)Cl-(L5)] (5), [Ru(η6-p-cymene)I-(L1)] (6), [Ru(η6-p-cymene)I-(L2)] (7), [Ru(η6-p-
cymene)I-(L3)] (8), [Ru(η6-p-cymene)I-(L4)] (9), [Ru(η6-p-cymene)I-(L5)] (10), [Ru(η6-p-cymene)I-(L6)] (11), [Ru(η6-p-
cymene)I-(L7)] (12), and [Ru(η6-p-cymene)Cl-(L8)] (13) respectively containing deprotonated 5,7-dichloro-2-methyl-8-
quinolinol (H-L1), 5,7-dibromo-2-methyl-8-quinolinol (H-L2), 5-chloro-7-iodo-8-hydroxy-quinoline (H-L3), 5,7-dibromo-
8-quinolinol (H-L4), 5,7-diiodo-8-hydroxyquinoline (H-L5), 8-hydroxy-2-methylquinoline (H-L6), 2,8-quinolinediol (H-
L7), or 6,7-dichloro-5,8-quinolinedione (H-L8). MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay
showed that 13 organometallic Ru(II)-arene complexes 1−13 are more selective for HeLa cells than normal HL-7702 cells.
In addition, the 1, 2, 5, and 6, which contain the active ligands H-L1 and H-L2, showed remarkable cell cytotoxicity, giving
the respective IC50 values of 2.00 ± 0.20 nM, 0.89 ± 0.62 μM, 25.00 ± 0.30 nM, and 2.18 ± 0.35 μM on HeLa cancer cells.
These values indicated higher activity than 6,7-dichloro-5,8-quinolinedione and the other 8-hydroxyquinoline derivative
Ru(II)-arene complexes. Interestingly, all these Ru(II)-arene complexes 1−13 were significantly less toxic to human hepatic
(HL-7702) cells. Moreover, 1- and 2-induced HeLa cell apoptosis was mediated by inhibition of telomerase activity and
dysfunction of mitochondria, and resulted in DNA damage and increased anti-migration activity on HeLa cells. The
organometallic Ru(II)-arene complex 1 exhibited evident priority on antitumor activity than 2, which should be highly
associated with the key roles of the 5,7-dichloro substituted groups in L1 ligand of organometallic Ru(II)-arene complexes
1. Remarkably, 1 showed higher inhibitory activity against xenograft tumor growth of human cervical cells (HeLa) in vivo
(tumor growth inhibition rate (TGIR) = 58.5%) than cisplatin. This study was the first to show that the 5,7-dihalogenated-2-
methyl-8-quinolinol organometallic Ru(II)-arene complexes 1 and 2 are novel Ru(II) anticancer drug candidates.
DOI: 10.1039/x0xx00000x
www.rsc.org/
[{Ru(phen)2}2tpphz]4+,9 Δ-/Λ-[Ru(phen)2(p-MOPIP)]2+ and Λ-
/Δ-[Ru(phen)2(p-HPIP)]2+ Ru complexes10, and more recently,
several types of nickel(II), copper(II), iron(II,III),
ruthenium(II,III), cobalt(II,III), tin(IV), vanadium(IV,V),
zinc(II), osmium(VI), rhodium(III), platinum(II,IV), and Ln(III)
Introduction
Cisplatin and its derivatives (e.g., oxaliplatin, nedaplatin,
carboplatin, heptaplatin, and lobaplatin) have remained popular
for their anticancer activities to date1. Practically, however, the
problems of drug resistance and systemic toxicity of Pt-drugs
stimulated the design for an alternative transition metal
antitumor drugs1: NAMI-A2, DW1/23, KP10194, RM1755 and
antitumor
metal
complexes
of
8-hydroxy-quinoline
derivatives11−20. Turel reported Cl–Ru complex ([Ru(η6-p-
cymene)Cl-(Cq)]) induced cell apoptosis via NFκB signaling
pathway, which was different from the clioquinol (H-Cq)16. A
series of Ru(II) coordination complexes with the 2,9-dimethyl-
1,10-phenanthroline (or 2,2’-bipyridine) and various
hydroxyquinoline mixed chelating ligands were designed but
failed to inhibit the proteasome at IC50 value19. On one hand,
KP13394,
RAPTA-T6,7
,
PTS
(RAPTA)
complexes8,
aState Key Laboratory for the Chemistry and Molecular Engineering of Medicinal
Resources, School of Chemistry and Pharmacy, Guangxi Normal University, 15
Yucai Road, Guilin 541004, PR China. E-mail: gxnuchem@foxmail.com (H.-H.
Zou); fliangoffce@yahoo.com (F.-P. Liang).
Liu investigated 8-hydroxyquinoline [Ru(phen)2(8-HQ)]+ (PQ
and [Ru(bpy)2(8-HQ)]+
BQ) ruthenium(II) complexes induce
)
bGuangxi Key Lab of Agricultural Resources Chemistry and Biotechnology, College
of Chemistry and Food Science, Yulin Normal University, 1303 Jiaoyudong Road,
Yulin 537000, PR China. qpqin2018@126.com (Q.‐P. Qin).
(
Hep-G2 cell apoptosis via binding of bFGF and remarkably
inhibited Hep-G2 tumor growth in vivo15. On the other hand,
Heidary and coauthors reported that 5-chloro-7-iodo-8-
hydroxy-quinoline (H-ClIQ) and 5,7-dibromo-8-quinolinol (H-
BrBrQ) Ru(II) complexes exhibited promising cytotoxic
activity against HL60 cancer cells, with IC50 values of 0.12 ±
0.002 μM and 0.08 ± 2.0 nM, respectively12. To date, a highly
tumor-selective organometallic Ru(II)-arene complexes with
cGuangxi Key Laboratory of Electrochemical and Magnetochemical Functional
Materials, College of Chemistry and Bioengineering, Guilin University of
Technology, Guilin 541004, China.
†Electronic Supplementary Information (ESI) available: The IR and X-ray
crystallization data, and
number for the Ru(II)-arene complexes
1895411, 1895280, 1895281, 1895282, 1895283, 1895284, 1895285, 1895412,
1895286, 1895287, 1895288, 1896981 and 1896982. The data can be obtained free
of charge viahttp://www.ccdc.cam.ac.uk, or from the Cambridge Crystallographic
Data Centre, 12 Union Road, Cambridge CB21EZ, UK (Fax: (+44) 1223-336-033;
E-mail: deposit@ccdc.cam.ac.uk). See DOI: 10.1039/x0xx00000x
1
suppressed HeLa tumor growth in vivo. The CCDC
13 and p-cymene-RuCl were 1895279,
1−
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5,7-dichloro-2-methyl-8-quinolinol (H-MClClQ) and 5,7- solvent was rotary-evaporated and replaced by CH3OH, and
dibromo-2-methyl-8-quinolinol (H-MBrBrQ) ligands have yet CH2Cl2 mixture solution was removed by filtration. The red-
to be reported, and the detailed in vitro and in vivo anticancer brown precipitate solution of organometallic Ru(II)-arene
mechanisms of these organometallic Ru(II)-arene complexes complexes collected by filtration and washed with n-hexane
remain unexplored.
Therefore, we synthesized the new organometallic Ru(II)- Ru(II)-arene complexes
arene complexes ([Ru(η6-p-cymene)Cl-(L1)] ( ), [Ru(η6-p- obtained by slow evaporation of a CH3OH and CH2Cl2 (v:v =
(5.0 mL, yield: 85.1%−95.2%). Crystals of 13 organometallic
1
−
13 and p-cymene-RuCl were
1
cymene)Cl-(L2)] (
cymene)Cl-(L4)] (
2
4
), [Ru(η6-p-cymene)Cl-(L3)] (
), [Ru(η6-p-cymene)Cl-(L5)] (
3
5
), [Ru(η6-p- 5:2) solution (Scheme 1) and were suitable for X-ray analysis.
), [Ru(η6-p- All the organometallic Ru(II)-arene complexes were fully
cymene)I-(L1)] (
6
), [Ru(η6-p-cymene)I-(L2)] (
), [Ru(η6-p-cymene)I-(L4)] (
7
9
), [Ru(η6-p- characterized by IR spectroscopy, ESI-MS spectra and NMR
), [Ru(η6-p- spectroscopy, single-crystal X-ray diffraction analyses, and
cymene)I-(L3)] (
8
cymene)I-(L5)] (10), [Ru(η6-p-cymene)I-(L6)] (11), [Ru(η6-p- elemental analysis (Fig.
cymene)I-(L7)] (12), and [Ru(η6-p-cymene)Cl-(L8)] (13)) with Information).
5,7-dichloro-2-methyl-8-quinolinol (H-L1), 5,7-dibromo-2-
1
and S1–S55, Supporting
methyl-8-quinolinol
(H-L2),
5-chloro-7-iodo-8-hydroxy-
quinoline (H-L3), 5,7-dibromo-8-quinolinol (H-L4), 5,7-diiodo-
8-hydroxyquinoline (H-L5), 8-hydroxy-2-methylquinoline (H-
L6),
quinolinedione (H-L8), respectively. In addition, the Ru(II)-
arene complexes 1- and -induced HeLa cell apoptosis was
2,8-quinolinediol
(H-L7),
or
6,7-dichloro-5,8-
2
mediated by inhibition of telomerase activity, dysfunction of
mitochondria, and evidently inhibition of HeLa xenograft tumor
growth (tumor growth inhibition rate (TGIR) = 58.5%) in vivo.
Scheme 1. General synthetic pathway for 13 organometallic
Ru(II)-arene complexes 13 Reagents and conditions:
1
−
.
CH3OH and CH2Cl2 mixture (v:v = 1:1), 65 °C, 6.0 h.
Crystal structure and stability of 13 organometallic Ru(II)-arene
complexes 1−13
The 13 organometallic Ru(II)-arene complexes 1−13 comprises
a Ru(II) center with an η6-pcymene ring, a deprotonated H-L1,
H-L2, H-L3, H-L4, H-L5, H-L6, H-L7, or H-L8 (O^N-QX), and
one Cl ligand (Fig. 2, S14–S16 and Table S1–S42). The X-ray
diffraction analysis of 13 organometallic Ru(II)-arene
complexes revealed that all the 13 organometallic Ru(II)-arene
Fig. 1. The chemical structure of anticancer Ru complexes
Results and discussion
complexes 1−13 and p-cymene-RuCl also featured the pseudo-
tetrahedral piano stool structure (Figs. 2 and S14–S16) with the
Ru(II) atom center coordinated to the pyridine N atom
(N^ligand) and the adjacent O atom (O^ligand) in a bidentate
fashion.
Furthermore, the solution behavior of 13 organometallic
Ru(II)-arene complexes 1−
13 (3.0 × 10–5 M) in 10 mM Tris-
Synthesis and characterization
The synthesis of 6,7-dichloro-5,8-quinolinedione (H-L7) was
carried out according to a procedure reported by Mulchin and
Kubanik et al.21,22 In addition, 0.049 mmol of dichloro(p-
cymene)Ru(II) dimer
(
p-cymene-RuCl
)
or diiodo(p-
HCl buffer (pH = 7.35, TBS, containing 5% DMSO) or DMSO
solution was further studied by ESI-MS spectra and NMR
spectroscopy. The NMR result showed that no other peaks
appeared (Figs. S31–S55), indicating no structural transitions
and/or decompositions on the 13 organometallic Ru(II)-arene
cymene)Ru(II) dimer (p-cymene-RuI) and 0.098 mmol of 5,7-
dichloro-2-methyl-8-quinolinol (H-L1), 5,7-dibromo-2-methyl-
8-quinolinol (H-L2), 5-chloro-7-iodo-8-hydroxy-quinoline (H-
L3),
hydroxyquinoline (H-L5), 8-hydroxy-2-methylquinoline (H-
L6), 2,8-quinolinediol (H-L7) or 6,7-dichloro-5,8-
5,7-dibromo-8-quinolinol
(H-L4),
5,7-diiodo-8-
complexes
1
−
13. The ESI-MS assay suggested that the ESI-MS
of 13 organometallic Ru(II)-arene complexes
1
−
13 (3.0 × 10–5
quinolinedione (H-L8) was dissolved in CH3OH and CH2Cl2
mixture solution, and the solution was refluxed for 6.0 h. The
M) had the base peak for [M – Cl]+ at m/z = 463.4, [M – Cl +
DMSO]+ at m/z = 626.9, [M – Cl]+ at m/z = 539.4, [M – Cl]+ at
2 | J. Name., 2019, 00, 1-9
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m/z = 537.3, [M – Cl]+ at m/z = 631.3, [M – Cl]+ at m/z = 461.4, and HL-7702 normal cells were performed to evaluate the
[M – Cl]+ at m/z = 551.4, [M – Cl]+ at m/z = 539.4, [M – Cl]+ at antitumor activity of 13 organometallic Ru(II)-arene complexes
m/z = 537.3, [M – Cl]+ at m/z = 631.3, [M – Cl]+ at m/z = 393.4,
[M – Cl]+ at m/z = 395.4 and [M – Cl]+ at m/z = 463.5 in the cisplatin. Table 1 revealed that organometallic Ru(II)-arene
TBS for 0 h (Figs. S17–S30), respectively. No change in the complexes and showed stronger antitumor potency than 11
1−13, H-L1, H-L2, H-L3, H-L4, H-L5, H-L6, H-L7, H-L8, and
1
2
m/z values (Figs. S17–S30) was observed after 48-h incubation organometallic Ru(II)-arene complexes and their corresponding
in 10 mM Tris-HCl buffer (pH = 7.35, TBS), suggesting that 13 H-L1, H-L2, H-L3, H-L4, H-L5, H-L6, H-L7, and H-L8
organometallic Ru(II)-arene complexes
this condition. In conclusion, 13 organometallic Ru(II)-arene following order:
complexes 11 12. In addition,
13 (3.0 × 10–5 M) were stable in 10 mM TBS for
48 h at 37 °C.
1
−
13 were stable under ligands, and the in vitro different antitumor activity were in the
13 10, and
was most cytotoxicities in HeLa
1
>
2
>
3
>
4
1
>
5
>
, 6 > 7 > 8 > 9 >
1
−
1
>
2
>
>
cells, with IC50 values 2.00 ± 0.20 nM, which was 40.0−9010.0
times more potent than that of clinical medicine cisplatin (IC50
= 15.02 ± 1.85 μM), 5,7-dichloro-2-methyl-8-quinolinol Fe(III)
complex (IC50 = 5.04 ± 0.62 μM)17, 5-chloro-7-iodo-8-hydroxy-
quinoline (H-ClIQ) and 5,7-dibromo-8-quinolinol Pt(II), and
Dy(III) and Ru(II) complexes (IC50 = 5.02 ± 0.62 μM, 4.09 ±
1.06 μM, 1.53 ± 0.59 μM, 18.02 ± 1.05 μM, 0.12 ± 0.002 μM,
and 0.08 ± 2.0 μM)12,18,20. Interestingly, these Ru(II)-arene
complexes
normal cells (IC50 > 100 μM), indicating the selectivity of
Ru(II)-arene complexes and on HeLa. This study was the
first report to show that 5,7-dihalogenated-2-methyl-8-
quinolinol organometallic Ru(II)-arene complexes and were
1 and 2 were significantly less toxic to HL-7702
Fig. 2. Molecular structure of organometallic Ru(II)-arene
complexes (A) and (B), respectively.
1
2
1
2
1
2
remarkably cytotoxic to HeLa tumor cells but significantly less
toxic to HL-7702 normal cells.
In vitro cytotoxicity
MTT assays with HeLa (human cervical cancer), T-24 (human
bladder cancer), SK-OV-3 (human ovarian carcinoma cancer),
Table 1. Cytotoxicities (IC50, μM) of 13 organometallic Ru(II)-arene complexes (H-L1, H-L2, H-L3, H-L4, H-L5, H-L6, H-L7, H-
L8) and cisplatin toward human HeLa, T-24, SK-OV-3, and HL-7702 cell linesa
Compounds
HeLa
T-24
SK-OV-3
HL-7702
H-L1
> 100
2.00 ± 0.20 nMc
0.89 ± 0.62
> 100
> 100
65.36 ± 1.69
1.03 ± 0.52
2.15 ± 1.13
82.23 ± 1.22
2.03 ± 0.51
2.55 ± 0.76
15.06 ± 0.77
3.19 ± 1.06
6.10 ± 0.97
88.01 ± 1.65
3.65 ± 0.46
6.83 ± 1.11
97.56 ± 1.28
4.44 ± 0.63
> 100
> 100
2.69 ± 0.74
5.36 ± 1.09
75.03 ± 1.02
8.36 ± 0.11
11.25 ± 0.63
25.03 ± 1.01
20.81 ± 1.45
> 100
1
> 100
6
H-L2
88.96 ± 0.45
> 100
25.00 ± 0.30 nMc
2.18 ± 0.35
50.26 ± 0.58
4.43 ± 0.52
8.31 ± 1.17
54.03 ± 1.18
4.96 ± 0.39
8.95 ± 0.76
86.99 ± 1.85
5.73 ± 0.44
2
> 100
7
H-L3
75.25 ± 1.43
85.69 ± 1.02
49.03 ± 0.75
> 100
3
8
H-L4
4
71.29 ± 2.06
20.31 ± 0.33
32.92 ± 1.25
> 100
77.69 ± 0.36
65.97 ± 0.86
75.03 ± 1.09
80.12 ± 0.28
9
H-L5
5
7.19 ± 0.23
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9.36 ± 1.45
13.99 ± 1.02
28.11 ± 0.57
17.83 ± 1.45
85.03 ± 1.01
10.09 ± 1.85
61.85 ± 0.74
5.01 ± 1.07
16.36 ± 0.95
7.22 ± 0.59
40.23 ± 1.33
16.57 ± 0.58
57.32 ± 0.35
4.36 ± 0.69
65.82 ± 1.16
6.17 ± 0.15
12.23 ± 0.41
70.15 ± 1.36
60.36 ± 2.21
61.23 ± 0.74
89.63 ± 1.52
88.23 ± 1.56
77.98 ± 1.08
90.22 ± 1.73
16.09 ± 1.05
10
H-L8
13
35.06 ± 1.44
10.53 ± 0.19
90.22 ± 0.55
6.11 ± 0.56
70.14 ± 1.21
2.03 ± 0.22
15.02 ± 1.85
H-L6
11
H-L7
12
Cisplatinb
aCancer and normal cells were treated with 13 organometallic Ru(II)-arene complexes, the corresponding ligands, and cisplatin at
different concentrations for 48 h. IC50 values were equal to the mean value ± SD value from five independent assays. bA total of 1.0
mM cisplatin was prepared in 0.154 M NaCl23−26. cThe concentration was in nM.
HeLa cells incubated with organometallic Ru(II)-arene
complexes
1 (2.0 nM) and 2 (25.0 nM) for 24 h exhibited
Migration assay
significant G0 peaks (sub-G1 peaks), which indicated HeLa cell
apoptosis and the apoptotic peaks of 26.54% and 18.68% in
sub-G1 phase of the cell cycle distribution, respectively (Fig.
4a−c). Such is the result further verified in the apoptosis
experiment by flow cytometry with FITC-Annexin V and PI
(propidium iodide) double staining. HeLa tumor cells were
Cancer cell migration was an important characteristic in cancer
metastasis27,28. The transwell migration assay in vitro was
performed on HeLa cells to study anti-migration effect of
organometallic Ru(II)-arene complexes
nM). The organometallic Ru(II)-arene complexes
and (25.0 nM) showed excellent anti-migration activities on
HeLa cells at 2.0 nM (low concentrations) (Fig. 3). Results
indicated that organometallic Ru(II)-arene complex (2.0 nM)
more significantly inhibited HeLa cancer cell migration at 2.0
nM than that of (25.0 nM).
1
(2.0 nM) and
2 (25.0
1
(2.0 nM)
2
treated with organometallic Ru(II)-arene complexes
1 (2.0 nM)
and (25.0 nM) for 24 h, and then these cancer cells were
2
1
harvested and analyzed by flow cytometry after staining with PI
and Annexin V-FITC dye (Fig. 4d−f) to confirm this property.
The percentages of apoptosis cells were 28.30% and 19.55%,
respectively, when the HeLa cancer cells were incubated with
2
organometallic Ru(II)-arene complexes
nM) for 24 h.
1 (2.0 nM) and 2 (25.0
Fig. 4. The organometallic Ru(II)-arene complexes
and (25.0 nM) induced the cell cycle distribution (a−c) and
apoptosis (d−f) of HeLa cells for 24 h
1 (2.0 nM)
2
Fig. 3. Anti-migration effect of organometallic Ru(II)-arene
complexes (2.0 nM, c) and (25.0 nM, b) on HeLa cells (a)
1 and 2 caused DNA damage and mitochondrial dysfunction
1
2
for 24 h via transwell migration assay. (d) The statistical result
of anti-migration assays were equal to the mean value ± SD
value from three independent assays (magnification 200 ×)
H2A.X and cleaved PARP proteins were closely related to
DNA damage response factors29−32. Organometallic Ru(II)-
arene complexes
1 (2.0 nM) and 2 (25.0 nM) treatment were
suggested to induce HeLa cell apoptosis, which was highly
associated with DNA damage or the appearance of sub-G1 peak
(the hypodiploid DNA content peak)29−32. Therefore, the
1- and 2-Induced Apoptosis
4 | J. Name., 2019, 00, 1-9
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immunofluorescence assay and Western blot were used to
clarify the expression of the H2A.X and cleaved-PARP proteins
in the HeLa cells. Treatment of organometallic Ru(II)-arene
complexes
1 (2.0 nM) and 2 (25.0 nM) remarkably enhanced
the levels of H2A.X and cleaved-PARP proteins expression
(Figs. 5 and 6). Thus, our data suggested that organometallic
Ru(II)-arene complexes
1 (2.0 nM) and 2 (25.0 nM)
substantially induced DNA damage. Moreover, Bcl-2 family
apoptosis-related proteins (e.g., bad, bcl-2, and bax) and
mitochondria-mediated pathway (e.g., cytochrome c, caspase-3,
and caspase-9) were activated by DNA damage29,33–35. Further
evidence from Western blot suggested that organometallic
Fig. 6. (A and B) Western blot analysis to detect the levels of
the DNA damage, mitochondria-mediated pathway, and Bcl-2
family-related proteins in HeLa cells treated with
Ru(II)-arene complexes
1 (2.0 nM) and 2 (25.0 nM)
organometallic Ru(II)-arene complexes
nM) for 24 h
1 (2.0 nM) and 2 (25.0
upregulated the expression of bad and bax proteins and the
correspondingly downregulated the level of bcl-2 protein (Fig.
6). These stimuli could decrease the ΔΨm (mitochondrial
membrane potential (MMP)) level (Fig. 7), and the
correspondingly green fluorescence intensity (JC-1 monomers)
increases from 8.76% to 45.51% or 36.65%, respectively,
which also could activate the level of reactive oxygen species
(ROS) (Fig. 8 and Table S43) and increase the apoptotic
cytochrome c and active caspase-3 and caspase-9 proteins (Fig.
6) in HeLa cells.
1 and 2 inhibit telomerase activity and decreased the related
proteins
Previous studies demonstrated that the cell cycle distribution in
sub-G1 phase by organometallic Ru(II)-arene complexes
nM) and (25.0 nM) was treated, which was related to the
inhibition of telomerase36−39. First, the effect of organometallic
Ru(II)-arene complexes (2.0 nM) and (25.0 nM) on
telomerase in HeLa cells was determined. Fig. 9A showed that
organometallic Ru(II)-arene complexes (2.0 nM) and (25.0
1 (2.0
2
1
2
1
2
nM) evidently inhibited telomerase activity (i.e., 49.51% and
30.61%, respectively). As predicted, organometallic Ru(II)-
arene complexes
1 (2.0 nM) and 2 (25.0 nM) significantly
decreased the level of the related (e.g., c-myc and hTERT)
proteins in HeLa cells (Fig. 9B and C). Results clearly revealed
that organometallic Ru(II)-arene complexes
1 (2.0 nM) may
have higher affinity toward telomerase activity and the related
proteins than
results.
2 (25.0 nM), which was in accord with the above
Fig. 5. The organometallic Ru(II)-arene complexes
and (25.0 nM) induced DNA damage in HeLa cells for 24 h.
The HeLa cells were treated with organometallic Ru(II)-arene
complexes (2.0 nM) and (25.0 nM) for 24 h, respectively,
and followed staining by H2A.X (green, primary antibodies)
and DAPI (blue) for 40.0 min. Thereafter, these cancer cells
were visualized by LeicaTCS-SP5 confocal microscope
(Germany, magnification 400 ×).
1 (2.0 nM)
Fig. 7. The organometallic Ru(II)-arene complexes
1 (2.0 nM)
2
and (25.0 nM) decreased the ΔΨm of HeLa cells for 24 h and
2
consequently were analyzed by flow cytometry after incubation
with JC-1 staining (the fluorescence probe JC-1) for 30.0 min
1
2
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1
(10.0 mg/kg/q2d) shows no significant toxicity within the 21-
day treatment. Fig. 10 and Tables S44−S46 show that treatment
with organometallic Ru(II)-arene complex (10.0 mg/kg/q2d)
1
resulted in a significant reduction in tumor volume (T/C =
35.6%) in comparison with the vehicle group. In addition,
inhibition of HeLa TGIR by organometallic Ru(II)-arene
complex
1 (10.0 mg/kg/q2d) with treated versus vehicle group
of 58.5% was observed on day 21.0 after treatment, which was
significantly higher than that of cisplatin (35.2%)43. The data
testified more potent inhibitory effect of organometallic Ru(II)-
arene complex
1 (10.0 mg/kg/q2d) on HeLa tumor growth
(TGIR = 58.5%) in vivo and higher safety than cisplatin.
Fig. 8. The organometallic Ru(II)-arene complexes
1 (2.0 nM)
and
2 (25.0 nM) which increased the level of ROS generation in
HeLa cells for 24.0
photometer
h were analyzed by fluorescence
Fig. 10. The organometallic Ru(II)-arene complex
HeLa tumor growth in vivo. Effect (A) and photographs (B) of
organometallic Ru(II)-arene complex (10 mg/kg/q2d) and
1 suppressed
1
vehicle (10% DMSO in saline, v/v) on HeLa tumor growth
(mean tumor volume (mm3)) ± SD (n = 6)
Structure–Activity Relationships (SAR)
Certain SARs trends in the different substituted quinolinedione
and 8-hydroxyquinoline derivative ligands, and the differences
antitumor activities (Fig. 11) and their mechanisms were
observed based on the in vitro and in vivo anticancer activity
results of 13 organometallic Ru(II)-arene complexes
i) The in vitro different cytotoxicity studies were in the
following order: 13 10 and
11 12
ii) The in vitro antitumor activity of organometallic Ru(II)-
arene complexes and follow the order of
iii) The in vivo anticancer activity of organometallic
Ru(II)-arene complex and cisplatin in HeLa tumor xenograft
1−13.
1
>
2
>
3
>
4
>
5
>
, 6 > 7 > 8 > 9 >
1
>
2
>
>
.
1
2
1 > 2.
Fig. 9. The organometallic Ru(II)-arene complexes
and (25.0 nM) inhibit telomerase activity and the related
proteins. (A) The influence of organometallic Ru(II)-arene
complexes (2.0 nM) and (25.0 nM) on the telomerase
1 (2.0 nM)
1
2
(SARs trend
1
> cisplatin) was also observed.
1
2
activity of HeLa. (B and C) Western blot analysis of the related
(e.g., c-myc and hTERT) proteins in organometallic Ru(II)-
arene complexes
24 h
1 (2.0 nM) and 2 (25.0 nM) treated cells for
1 suppressed HeLa tumor growth in vivo
Model nude mice with HeLa tumor received a possible highest
administration value of organometallic Ru(II)-arene complex
1
(10.0 mg/kg every two days (q2d), 1.0 mL/20 g, 5% v/v
DMSO/saline) by intraperitoneal injection40−42, and no signs of
peritonitis (other adverse effects) or damage to organs were
observed, suggesting that organometallic Ru(II)-arene complex
6 | J. Name., 2019, 00, 1-9
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organometallic Ru(II)-arene complexes 1−13 suitable for X-ray
analysis were obtained by slow evaporation of
CH3OH/CH2Cl2 (6.0 mL, v:v = 5:2) solution (Scheme 1).
a
Data for 6: Yield (95.2%). Elemental analysis calcd (%)
for C20H20Cl2INORu: C 40.77, H 3.42, N 2.38. Found: C 40.72,
1
H 3.45, N 2.34. ESI-MS: m/z = 461.4 for [M – Cl]+. H NMR
(500 MHz, DMSO-d6) δ 8.22 (d, J = 8.6 Hz, 1H), 7.67 (d, J =
8.7 Hz, 1H), 7.51 (s, 1H), 5.95 (d, J = 5.9 Hz, 1H), 5.87 – 5.80
(m, 2H), 5.55 (d, J = 5.9 Hz, 1H), 3.14 (s, 3H), 2.65 (dq, J =
13.9, 6.9 Hz, 1H), 2.47 (s, 3H), 1.09 (d, J = 6.9 Hz, 3H), 0.93
(d, J = 6.9 Hz, 3H). 13C NMR (126 MHz, DMSO-d6) δ 163.06,
162.75, 144.85, 134.30, 128.06, 125.48, 124.26, 117.64,
110.37, 104.15, 98.48, 85.38, 81.48, 80.69, 80.38, 31.13, 30.31,
22.19, 21.82, 21.05.
Data for 1: Yield (85.1%). Elemental analysis calcd (%)
for C20H20Cl3NORu: C 48.25, H 4.05, N 2.81. Found: C 48.28,
1
H 4.07, N 2.78. ESI-MS: m/z = 463.4 for [M – Cl]+. H NMR
(500 MHz, DMSO-d6) δ 8.20 (d, J = 8.6 Hz, 1H), 7.74 (s, 1H),
7.69 (d, J = 8.7 Hz, 1H), 5.98 (dd, J = 5.9, 1.1 Hz, 1H), 5.88
(dd, J = 6.1, 1.2 Hz, 1H), 5.77 (dd, J = 6.2, 1.2 Hz, 1H), 5.55
(dd, J = 5.9, 1.2 Hz, 1H), 3.32 (s, 1H), 3.19 (s, 3H), 2.20 (s,
3H), 1.04 (d, J = 7.0 Hz, 3H), 0.91 (d, J = 6.9 Hz, 3H). 13C
NMR (126 MHz, DMSO-d6) δ 162.68, 161.18, 143.94, 133.80,
127.60, 124.96, 123.72, 116.58, 109.83, 100.39, 100.10, 86.31,
79.87, 79.78, 78.47, 30.25, 28.25, 21.64, 21.54, 18.27
Fig. 11. Proposed anticancer mechanisms for organometallic
Ru(II)-arene complexes
1 and 2 (e.g., mechanisms of 1)
Data for 2: Yield (87.5%). Elemental analysis calcd (%)
for C20H20Br2ClNORu: C 40.94, H 3.44, N 2.39. Found: C
40.90, H 3.47, N 2.36. ESI-MS: m/z = 626.9 for [M – Cl +
DMSO]+. 1H NMR (500 MHz, DMSO-d6) δ 8.20 (d, J = 8.6 Hz,
1H), 7.74 (s, 1H), 7.69 (d, J = 8.7 Hz, 1H), 5.98 (dd, J = 5.9, 1.1
Hz, 1H), 5.88 (dd, J = 6.1, 1.1 Hz, 1H), 5.77 (dd, J = 6.1, 1.2
Hz, 1H), 5.55 (dd, J = 5.9, 1.2 Hz, 1H), 3.19 (s, 3H), 2.20 (s,
3H), 1.04 (d, J = 7.0 Hz, 3H), 0.91 (d, J = 6.9 Hz, 3H). 13C
NMR (126 MHz, DMSO-d6) δ 164.87, 161.71, 156.92, 144.33,
136.84, 133.52, 125.88, 107.31, 100.79, 100.75, 99.41, 86.80,
80.51, 80.37, 79.36, 30.82, 28.79, 22.19, 22.15, 18.82.
Experimental materials and methods
Synthesis of 6,7-dichloro-5,8-quinolinedione (L7)
The synthesis of 6,7-dichloro-5,8-quinolinedione (L7) was
carried out according to a procedure reported by Mulchin and
Kubanik et al.21,22 Yield: 17.0%. ESI-MS: m/z = 241.2 for [M +
Na]+. Elemental analysis: calcd (%) for C9H5Cl2NO2: C 46.99,
H 2.19, N 6.09; found: C 46.95, H 2.21, N 6.06. 1H NMR (500
MHz, DMSO-d6) δ 9.05 (dd, J = 4.6, 1.7 Hz, 1H), 8.46 (dd, J =
7.9, 1.7 Hz, 1H), 7.89 (dd, J = 7.9, 4.6 Hz, 1H). 13C NMR (126
MHz, DMSO-d6) δ 176.46, 174.71, 154.93, 147.52, 143.59,
142.04, 135.38, 128.99, 128.75.
Data for 7: Yield (90.1%). Elemental analysis calcd (%)
for C20H20Br2INORu: C 35.42, H 2.97, N 2.07. Found: C 35.45,
1
H 2.94, N 2.02. ESI-MS: m/z = 551.4 for [M – Cl]+. H NMR
(500 MHz, DMSO-d6) δ 8.15 (d, J = 8.6 Hz, 1H), 7.73 (s, 1H),
7.68 (d, J = 8.7 Hz, 1H), 5.93 (dd, J = 5.9, 1.2 Hz, 1H), 5.86
(dd, J = 6.3, 1.2 Hz, 1H), 5.83 – 5.81 (m, 1H), 5.56 (dd, J = 6.0,
1.2 Hz, 1H), 3.15 (s, 3H), 2.65 (p, J = 7.0 Hz, 1H), 2.46 (s, 3H),
1.10 (d, J = 7.0 Hz, 3H), 0.93 (d, J = 6.9 Hz, 3H). 13C NMR
(126 MHz, DMSO-d6) δ 164.66, 162.69, 144.63, 136.75,
133.39, 125.98, 125.80, 107.73, 104.31, 99.37, 98.17, 85.17,
81.68, 81.05, 80.29, 31.09, 30.29, 22.25, 21.76, 20.95.
Synthesis and characterization of 13 organometallic Ru(II)-
arene complexes
A total of 0.049 mmol of dichloro(p-cymene) Ru(II) dimer (p-
cymene-RuCl) or diiodo(p-cymene) Ru(II) dimer (p-cymene-
RuI) and 0.098 mmol of 5,7-dichloro-2-methyl-8-quinolinol
(H-L1), 5,7-dibromo-2-methyl-8-quinolinol (H-L2), 5-chloro-7-
iodo-8-hydroxy-quinoline (H-L3), 5,7-dibromo-8-quinolinol
(H-L4), 5,7-diiodo-8-hydroxyquinoline (H-L5), 8-hydroxy-2-
methylquinoline (H-L6), 2,8-quinolinediol (H-L7), or 6,7-
dichloro-5,8-quinolinedione (H-L8) were dissolved in 10.0 mL
of CH3OH/CH2Cl2 mixture (v:v = 1:1), and the solution was
refluxed for 6.0 h. The solvent was rotary evaporated, replaced
by CH3OH/CH2Cl2 (5.0 mL, v:v = 1:1), and was removed by
filtration. The red-brown precipitate solution of organometallic
Ru(II)-arene complexes were collected by filtration and washed
with n-hexane (5.0 mL, yield: 85.1%−95.2%). Crystals of 13
Data for 13: Yield (93.5%). Elemental analysis calcd (%)
for C19H19Cl2NO3Ru: C 47.41, H 3.98, N 2.91. Found: C 47.36,
1
H 4.03, N 2.88. ESI-MS: m/z = 463.5 for [M – Cl]+. H NMR
(500 MHz, DMSO-d6) δ 9.44 (d, J = 5.6 Hz, 1H), 8.32 (d, J =
7.8 Hz, 1H), 7.83 (dd, J = 7.8, 5.6 Hz, 1H), 6.07 (d, J = 6.1 Hz,
1H), 6.02 (d, J = 6.0 Hz, 1H), 5.85 (d, J = 6.2 Hz, 1H), 5.84 –
5.79 (m, 2H), 2.77 (q, J = 6.9 Hz, 1H), 2.20 (s, 3H), 1.21 (d, J =
7.0 Hz, 3H), 1.17 (s, 3H).
Data for 3: Yield (89.6%). Elemental analysis calcd (%)
for C19H18Cl2INORu: C 39.67, H 3.15, N 2.43. Found: C 39.63,
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H 3.19, N 2.41. ESI-MS: m/z = 539.4 for [M – Cl]+. H NMR
Data for
8
:
Yield (88.9%). Elemental analysis calcd (%)
(500 MHz, DMSO-d6) δ 9.30 (dd, J = 4.9, 1.3 Hz, 1H), 8.36 for C19H18Cl2INORu: C 34.23, H 2.72, N 2.10. Found: C 34.19,
1
(dd, J = 8.6, 1.2 Hz, 1H), 7.79 (s, 1H), 7.71 (dd, J = 8.6, 4.9 Hz, H 2.74, N 2.07. ESI-MS: m/z = 539.4 for [M – Cl]+. H NMR
1H), 5.92 (dd, J = 6.1, 1.1 Hz, 1H), 5.78 (dd, J = 6.0, 1.1 Hz, (500 MHz, DMSO-d6) δ 9.25 (dd, J = 5.0, 1.3 Hz, 1H), 8.32
1H), 5.71 (dd, J = 6.0, 1.1 Hz, 1H), 5.64 (dd, J = 6.1, 1.1 Hz, (dd, J = 8.6, 1.2 Hz, 1H), 7.78 (s, 1H), 7.68 (dd, J = 8.6, 5.0 Hz,
1H), 2.71 (hept, J = 6.9 Hz, 1H), 2.19 (s, 3H), 1.17 (d, J = 6.9 1H), 5.86 (dd, J = 6.0, 1.2 Hz, 1H), 5.80 (ddd, J = 9.1, 6.0, 1.2
Hz, 3H), 1.09 (d, J = 6.9 Hz, 3H). 13C NMR (126 MHz, Hz, 2H), 5.73 (dd, J = 5.9, 1.2 Hz, 1H), 2.82 (p, J = 6.9 Hz,
DMSO-d6) δ 167.73, 152.06, 141.96, 136.23, 134.52, 127.01, 1H), 2.33 (s, 3H), 1.22 (d, J = 6.9 Hz, 3H), 1.13 (d, J = 6.9 Hz,
124.37, 111.02, 101.17, 97.92, 82.60, 82.51, 82.15, 80.70, 3H). 13C NMR (126 MHz, DMSO-d6) δ 167.50, 153.30, 142.43,
80.35, 30.94, 22.31, 22.28, 18.41.
Data for Yield (92.3%). Elemental analysis calcd (%) 82.01, 81.99, 81.31, 80.52, 31.31, 22.31, 22.11, 19.56.
for C19H18Br2ClNORu: C 39.85, H 3.17, N 2.45. Found: C Data for 11: Yield (85.8%). Elemental analysis calcd (%)
136.12, 134.39, 126.93, 124.44, 110.92, 103.92, 96.21, 84.17,
4
:
1
39.81, H 3.20, N 2.43. ESI-MS: m/z = 537.3 for [M – Cl]+. H for C20H22INORu: C 46.16, H 4.26, N 2.69. Found: C 46.12, H
1
NMR (500 MHz, DMSO-d6) δ 9.33 (dd, J = 5.0, 1.2 Hz, 1H), 4.30, N 2.67. ESI-MS: m/z = 393.4 for [M – Cl]+. H NMR
8.31 (dd, J = 8.6, 1.2 Hz, 1H), 7.81 (s, 1H), 7.72 (dd, J = 8.6, (500 MHz, DMSO-d6) δ 8.04 (d, J = 8.5 Hz, 1H), 7.45 (d, J =
5.0 Hz, 1H), 5.96 – 5.90 (m, 1H), 5.83 (dd, J = 6.1, 1.2 Hz, 1H), 8.5 Hz, 1H), 7.13 (t, J = 7.9 Hz, 1H), 6.71 (dd, J = 7.9, 1.2 Hz,
5.71 – 5.63 (m, 2H), 2.69 (p, J = 6.9 Hz, 1H), 2.18 (s, 3H), 1.13 1H), 6.65 (dd, J = 7.8, 1.1 Hz, 1H), 5.87 (dd, J = 5.8, 1.2 Hz,
(d, J = 6.9 Hz, 3H), 1.08 (d, J = 6.9 Hz, 3H). 13C NMR (126 1H), 5.77 (dd, J = 6.2, 1.2 Hz, 1H), 5.69 (dd, J = 6.1, 1.2 Hz,
MHz, DMSO-d6) δ 164.73, 151.63, 143.87, 136.27, 133.97, 1H), 5.47 (dd, J = 5.8, 1.2 Hz, 1H), 2.63 (td, J = 6.9, 2.5 Hz,
127.15, 123.96, 105.98, 100.48, 98.60, 97.87, 82.21, 81.72, 1H), 2.42 (s, 3H), 1.04 (d, J = 6.9 Hz, 3H), 0.92 (d, J = 6.9 Hz,
81.38, 79.95, 30.35, 21.80, 21.55, 17.89.
Data for Yield (88.3%). Elemental analysis calcd (%) 137.62, 128.53, 128.34, 124.17, 114.33, 109.81, 103.04, 98.66,
for C19H18Br2INORu: C 34.36, H 2.73, N 2.11. Found: C 34.40, 85.81, 81.48, 80.37, 79.94, 31.21, 30.01, 22.42, 21.91, 21.19.
3H). 13C NMR (126 MHz, DMSO-d6) δ 168.72, 160.40, 144.58,
9
:
1
H 2.71, N 2.13. ESI-MS: m/z = 537.3 for [M – Cl]+. H NMR
Data for 12
:
Elemental analysis calcd (%) for
(500 MHz, DMSO-d6) δ 9.28 (dd, J = 5.0, 1.2 Hz, 1H), 8.28 C19H20INO2Ru: C 43.69, H 3.86, N 2.68. Found: C 43.67, H
(dd, J = 8.6, 1.2 Hz, 1H), 7.80 (s, 1H), 7.69 (dd, J = 8.6, 5.0 Hz, 3.90, N 2.66. ESI-MS: m/z = 395.4 for [M – Cl]+.
1H), 5.87 (dd, J = 6.1, 1.2 Hz, 1H), 5.84 (dd, J = 6.0, 1.3 Hz,
1H), 5.79 (dd, J = 6.0, 1.2 Hz, 1H), 5.72 (dd, J = 5.9, 1.2 Hz,
Materials and methods
1H), 2.83 – 2.78 (m, 1H), 2.33 (s, 3H), 1.17 (d, J = 7.0 Hz, 3H),
1.12 (d, J = 6.9 Hz, 3H). 13C NMR (126 MHz, DMSO-d6
)
The X-ray crystallography structure analysis method and
antitumor mechanism the detailed procedures of 13
organometallic Ru(II)-arene complexes
ESI (supporting information).
164.50, 152.86, 144.33, 136.15, 133.86, 127.07, 124.03,
106.27, 103.23, 98.48, 96.10, 83.12, 81.52, 81.22, 81.00, 30.74,
21.75, 21.54, 19.10.
1−13 were described in
Data for 5: Yield (87.0%). Elemental analysis calcd (%)
for C19H18ClI2INORu: C 34.23, H 2.72, N 2.10. Found: C
Conclusions
1
34.20, H 2.75, N 2.08. ESI-MS: m/z = 631.3 for [M – Cl]+. H
NMR (500 MHz, DMSO-d6) δ 9.25 (dd, J = 4.9, 1.2 Hz, 1H), A total of 13 organometallic Ru(II)-arene complexes
8.16 (dd, J = 8.6, 1.2 Hz, 1H), 8.06 (s, 1H), 7.68 (dd, J = 8.6, been synthesized and characterized. Cytotoxicity studies
4.9 Hz, 1H), 5.92 (dd, J = 6.1, 1.2 Hz, 1H), 5.77 (dd, J = 5.9, showed that organometallic Ru(II)-arene complexes
and
1−13 have
1
2
1.2 Hz, 1H), 5.70 (dd, J = 5.9, 1.2 Hz, 1H), 5.64 (dd, J = 6.1, have higher antiproliferative activity than other 11 Ru(II)-arene
1.2 Hz, 1H), 2.70 (hept, J = 6.9 Hz, 1H), 2.19 (s, 3H), 1.17 (d, J complexes on HeLa cells, with IC50 values 2.00 ± 0.20 nM and
= 6.9 Hz, 3H), 1.08 (d, J = 7.0 Hz, 3H). 13C NMR (126 MHz, 25.00 ± 0.30 nM, respectively. Interestingly, all these Ru(II)-
DMSO-d6) δ 168.92, 151.98, 145.55, 142.86, 141.31, 131.02, arene complexes were significantly less toxic to HL-7702
124.89, 101.15, 97.91, 83.15, 82.64, 82.54, 82.11, 80.74, 73.45, normal cells. Moreover, organometallic Ru(II)-arene complexes
30.94, 22.33, 22.29, 18.43.
Data for 10 Yield (91.0%). Elemental analysis calcd (%) inhibition of telomerase activity (Fig. 11) and dysfunction of
for C19H18I3INORu: C 30.10, H 2.39, N 2.11. Found: C 30.05, mitochondria. The organometallic Ru(II)-arene complexes
1- and 2-induced HeLa cell apoptosis was mediated by the
:
1
1
H 2.43, N 2.09. ESI-MS: m/z = 631.3 for [M – Cl]+. H NMR exhibited evident priority on antitumor activity than
2, which
(500 MHz, DMSO-d6) δ 9.20 (dd, J = 4.9, 1.2 Hz, 1H), 8.12 should be highly associated with the key roles of the 5,7-
(dd, J = 8.6, 1.2 Hz, 1H), 8.05 (s, 1H), 7.65 (dd, J = 8.6, 5.0 Hz, dichloro substituted groups in L1 ligand of organometallic
1H), 5.86 (dd, J = 6.1, 1.3 Hz, 1H), 5.79 (td, J = 6.5, 1.2 Hz, Ru(II)-arene complexes
1. Remarkably, organometallic Ru(II)-
2H), 5.73 (dd, J = 6.0, 1.2 Hz, 1H), 2.81 (p, J = 6.9 Hz, 1H), arene complex
1
also evidently inhibited human cervical cells
2.33 (s, 3H), 1.22 (d, J = 7.0 Hz, 3H), 1.13 (d, J = 6.9 Hz, 4H). (HeLa) xenograft tumor growth (TGIR = 58.5%) in vivo. In
13C NMR (126 MHz, DMSO-d6) δ 168.70, 153.23, 145.43, conclusion, this study might imply the first 5,7-dihalogenated-
143.32, 141.20, 130.94, 124.94, 103.93, 96.17, 84.19, 83.33, 2-methyl-8-quinolinol organometallic Ru(II)-arene complex
1
82.06, 81.97, 81.33, 73.39, 31.31, 22.31, 22.12, 19.57.
as novel Ru(II) anticancer drug candidates.
8 | J. Name., 2019, 00, 1-9
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15 L. Yang, J. Zhang, C. Wang, X. Qin, Q. Yu, Y. Zhou and J.
Liu, Metallomic, 2014, , 518–531.
16 M. Gobec, J. Kljun, I. Sosic, I. Mlinaric-Rascan, M. Ursis, S.
Gobec and I. Turel, Dalton Trans., 2014, 43, 9045–9051.
17 B.-Q. Zou, Q.-P. Qin, Y.-X. Bai, Q.-Q. Cao, Y. Zhang, Y.-C.
Liu, Z.-F. Chen and H. Liang, Med. Chem. Commun., 2017,
Conflicts of interest
6
No conflicts to declare.
Acknowledgements
8
, 633–639.
We thank the National Natural Science Foundation of China
(Nos. 21771043, 51572050, 21867017, 21601038, and
21261025), Guangxi Natural Science Foundation (Nos.
18 Q.-P. Qin, S.-L. Wang, M.-X. Tan, Y.-C. Liu, T. Meng, B.-Q.
Zou and H. Liang, Eur. J. Med. Chem., 2019, 161, 334–342.
19 D. K. Heidary, B. Howerton and E. C. Glazer, J. Med. Chem.
,
2014, 57, 8936–8946.
2016GXNSFAA380085,
2015GXNSFDA139007,
and
20 H.-H. Zou, T. Meng, Q. Chen, Y.-Q. Zhang , H.-L. Wang, B.
2018GXNSFBA138021), Guangxi Key Laboratory of
Electrochemical and Magnetochemical Functional Materials
(EMFM20162107), the Key Foundation Project of Colleges and
Universities in Guangxi (No. ZD2014108), the PhD Research
Startup Program of Yulin Normal University (No. G2017009),
State Key Laboratory for Chemistry and Molecular Engineering
of Medicinal Resources (Guangxi Normal University)
(CMEMR2018-C8 and CMEMR2017-A11) and the Innovative
Li, K. Wang, Z.-L. Chen and F.-P. Liang, Inorg. Chem.
2019, 58, 2286−2298
,
21 B. J. Mulchin, C. G. Newton, J. W. Baty, C. H. Grasso, W. J.
Martin, M. C. Walton, E. M. Dangerfield, C. H. Plunkett, M.
V. Berridge, J. L. Harper, M. S. M. Timmer and B. L.
Stocker, Bioorg. Med. Chem., 2010, 18, 3238−3251.
22 M. Kubanik, N. Y. S. Lam, H. U. Holtkamp, T. Sohnel, R. F.
Anderson, S. M. F. Jamieson and C. G. Hartinger, Chem.
Commun., 2018, 54, 992−995.
23 R. Cao, J.-L. Jia, X.-C. Ma, M. Zhou and H. Fei, J. Med.
Team
& Outstanding Talent Program of Colleges and
Chem., 2013, 56, 3636−3644.
Universities in Guangxi (2014-49) for the financial support.
24 C. M. Clavel, E. Păunescu, P. Nowak-Sliwinska, A. W.
Griffioen, R. Scopelliti and P. J. Dyson, J. Med. Chem.
,
2014, 57, 3546−3558.
References
25 M. Gobec, J. Kljun, I. Sosič, I. Mlinarič-Raščan, M. Uršič, S.
Gobec and I. Turel, Dalton Trans., 2014, 43, 9045–9051.
26 Y. Gothe, T. Marzo, L. Messori and N. Metzler-Nolte, Chem.
Eur. J., 2016, 22, 12487−12494.
27 S.-W. Mao, H. Chen, L.-F. Yu, F. Lv, Y.-J. Xing, T. Liu, J.
Xie, J. Tang, Z. Yi and F. Yang, Eur. J. Med. Chem., 2016,
122, 574−583.
1
I. Romero-Canelón, L. Salassa and P. J. Sadler, J. Med.
Chem., 2013, 56, 1291−1300.
2
V. Novohradsky, A. Bergamo, M. Cocchietto, J. Zajac, V.
Brabec, G. Mestroni and G. Sava, Dalton Trans., 2015, 44
,
1905−1913.
3
K. S. M. Smalley, R. Contractor, N. K. Haas, A. N. Kup, G.
E. Atilla-Gokcuman, D. S. Williams, H. Bregman, K. T.
Flaherty, M. S. Soengas, E. Meggers and M. Herlyn, Cancer
Res., 2007, 67, 209−217.
28 J. Yan, J. Chen, S. Zhang, J. Hu, L. Huang and X. Li, J. Med.
Chem., 2016, 59, 5264−5283.
29 Z.-F. Chen, Q.-P. Qin, J.-L. Qin, Y.-C. Liu, K.-B. Huang, Y.-
L. Li, T. Meng, G.-H. Zhang, Y. Peng, X.-J. Luo and H.
Liang, J. Med. Chem., 2015, 58, 2159−2179.
4
5
F.-Z. Sadafi, L. Massai, G. Bartolommei, M. R. Moncelli, L.
Messori and F. Tadini-Buoninsegni, ChemMedChem, 2014,
30 H. Takai, A. Smogorzewska and T. de Lange, Curr. Biol.
,
9
, 1660−1664.
2003, 13, 1549−1556.
R. E. Morris, R. E. Aird, P. S. Murdoch, H. Chen, J.
Cummings, N. D. Hughes, S. Parsons, A. Parkin, G. Boyd,
31 G. B. Celli and T. de Lange, Nat. Cell Biol., 2005,
7
,
712−718.
D. I. Jodrell and P. J. Sadler, J. Med. Chem., 2001, 44
3616−3621.
,
32 Y. Zhang, D. Li, Q. Jiang, S. Cao, H. Sun, Y. Chai, X. Li, T.
Ren, R. Yang, F. Feng, B.-A. Li and Q. Zhao, Cell Death
6
P. Nowak-Sliwinska, J. R. van Beijnum, A. Casini, A. A.
Nazarov, G. Wagnières, H. van den Bergh, P. J. Dyson and
A. W. Griffioen, J. Med. Chem., 2011, 54, 3895−3902.
L. Zeng, P. Gupta, Y. Chen, E. Wang, L. Ji, H. Chao and Z.-
S. Chen, Chem. Soc. Rev., 2017, 46, 5771−5804.
K. J. Kilpin, S. M. Cammack, C. M. Clavel and P. J. Dyson,
Dalton Trans., 2013, 42, 2008−2014.
Dis., 2018, 9, 743. Doi: 10.1038/s41419-018-0804-6.
33 M.-C. Wei, W.-X. Zong, E. H. Y. Cheng, T. Lindsten, V.
Panoutsakopoulou, A. J. Ross, K. A. Roth, G. R.
7
8
9
MacGregor, C. B. Thompson and S. J. Korsmeyer, Science
2001, 292, 727−730.
,
34 K. Sinha, J. Das, P. B. Pal and P. C. Sil, Arch. Toxicol., 2013,
87, 1157−1180.
35 Q. Zhang, Y. Teng, Y. Yuan, T. Ruan, Q. Wang, X. Gao, Y.
Zhou, K. Han, P. Yu and K. Lu, Eur. J. Med. Chem., 2018,
156, 800–814.
T. Wilson, P. J. Costa, V. Félix, M. P. Williamson and J. A.
Tomas, J. Med. Chem., 2013, 56, 8674−8683.
10 D. Sun, Y. Liu, D. Liu, R. Zhang, X. Yang and J. Liu, Chem.
Eur. J., 2012, 18, 4285−4295.
36 A. K. P. Taggart, S.-C. Teng and V. A. Zakian, Science
,
11 B. Deka, T. Sarkar, S. Banerjee, A. Kumar, S. Mukherjee, S.
2002, 297, 1023−1026.
Deka, K. K. Saikia and A. Hussain, Dalton Trans., 2017, 46
,
37 Q.-P. Qin, J.-L. Qin, T. Meng, W.-H. Lin, C.-H. Zhang, Z.-Z.
Wei, J.-N. Chen, Y.-C. Liu, H. Liang and Z.-F. Chen, Eur. J.
Med. Chem., 2016, 124, 380−392.
396−409.
12 D. Havrylyuk, B. S. Howerton, L. Nease, S. Parkin, D. K.
Heidary and E. C. Glazer, Eur. J. Med. Chem., 2018, 156
,
38 Q. Yu, Y. Liu, J. Zhang, F. Yang, D. Sun, D. Liu, Y. Zhou
790−799.
and J. Liu, Metallomics, 2013, 5, 222−231.
13 C. M. Santos, S. Cabrera, C. Rios-Luci, J. M. Padron, I. L.
Solera, A. G. Quiroga, M. A. Medrano, C. Navarro-
Ranninger and J. Aleman, Dalton Trans., 2013, 42, 13343–
13348.
14 S. Tardito, A. Barilli, I. Bassanetti, M. Tegoni, O. Bussolati,
R. Franchi-Gazzola, C. Mucchino and L. Marchio, J. Med.
Chem., 2012, 55, 10448–10459.
39 L. Xu, X. Chen, J. Wu, J. Wang, L. Ji and H. Chao, Chem.
Eur. J., 2015, 21, 1−14.
40 K.-H. Diehl, R. Hull, D. Morton, R. Pfister, Y.
Rabemampianina, D. Smith, J.-M. Vidal and C. Van De
Vorstenbosch, J. Appl. Toxicol., 2001, 21, 15−23.
41 Z.-F. Chen, Q.-P. Qin, J.-L. Qin, J. Zhou, Y.-L. Li, N. Li, Y.-
C. Liu and H. Liang, J. Med. Chem., 2015, 58, 4771−4789.
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Journal Name
42 Y. Wang, J. Xiao, H.-P. Zhou, S.-L. Yang, X.-P. Wu and G.
Liang, J. Med. Chem., 2011, 54, 3768−3778.
43 J.-Q. Wang, P.-Y. Zhang, L.-N. Ji and H. Chao, J. Inorg.
Biochem., 2015, 146, 89–96.
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