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Reactivity and biological activity of N,N,S-Schiff-base rhodium pentamethylcyclopentadienyl complexes
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Research paper
Reactivity and biological activity of N,N,S-Schiff-base rhodium pentamethyl-
cyclopentadienyl complexes
Wassila Aboura, Lucinda K. Batchelor, Amine Garci, Paul J. Dyson, Bruno
Therrien
PII: S0020-1693(19)31590-7
DOI: https://doi.org/10.1016/j.ica.2019.119265
Reference: ICA 119265
To appear in: Inorganica Chimica Acta
Received Date: 18 October 2019
Revised Date: 4 November 2019
Accepted Date: 4 November 2019
Please cite this article as: W. Aboura, L.K. Batchelor, A. Garci, P.J. Dyson, B. Therrien, Reactivity and biological
activity of N,N,S-Schiff-base rhodium pentamethylcyclopentadienyl complexes, Inorganica Chimica Acta (2019),
doi: https://doi.org/10.1016/j.ica.2019.119265
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© 2019 Published by Elsevier B.V.
Reactivity and biological activity of N,N,S-Schiff-base rhodium
pentamethylcyclopentadienyl complexes
Wassila Abouraa, Lucinda K. Batchelorb, Amine Garcic, Paul J. Dysonb, and Bruno Therrienc,*
a Laboratoire de Chimie et d’Electrochimie des Complexes Métalliques (LCECM),
Département de Chimie Organique Industrielle, Faculté de Chimie, Université des Sciences et
de la Technologie d’Oran Mohamed Boudiaf, BP 1505, El M’naouer, 31000 Oran, Algérie
b Institut des Sciences et Ingénierie Chimique, Ecole Polytechnique Fédérale de Lausanne
(EPFL), 1015 Lausanne, Switzerland
c Institute of Chemistry, University of Neuchatel, Avenue de Bellevaux 51, 2000 Neuchatel,
Switzerland
*Corresponding author: bruno.therrien@unine.ch
Abstract
Neutral piano-stool complexes of the general formula [(η5-C Me )Rh(L-OR)] (R = Me, 1; R =
5 5
Et, 2; R = Pri, 3) have been prepared in alcohols (methanol, 1; ethanol, 2; isopropanol, 3) from
the Schiff-base 5-methyl-4-{(pyridin-2-ylmethylene)amino}-4H-1,2,4-triazole-3-thiol (L-H)
and the dinuclear precursor [(η5-C Me )RhCl ] . Concomitant with the coordination of the
5 5 2 2
Schiff-base ligand, an alkoxilation occurs on the imine carbon atom of the ligand, thus
forming the corresponding L-OR compounds. In these complexes, the L-OR ligand is N,N,S-
coordinated, introducing chirality at the metal center. The antiproliferative activity of the
piano-stool complexes 1-3 was evaluated on cancerous (A2780 and A2780cisR) and non-
cancerous (HEK293) cell lines, showing no significant activity in vitro (IC > 200 μM),
50
except for the ethanolate derivative 2, which shows an IC of 21.4 μM on the ovarian cancer
50
cell line A2780.
Keywords: Piano-stool complexes; Nucleophilic addition; Schiff-base ligand; Rhodium
complexes; Bio-organometallic chemistry.
1
Highlights
-Mononuclear chiral-at-metal pentamethylcyclopentadienyl rhodium complexes.
-Alkoxylation of the Schiff-base ligand during the formation of the piano-stool complexes.
-Evaluation of the antiproliferative activity of the complexes on cancerous and non-cancerous
cell lines.
Graphical TOC
2
1. Introduction
The biological potential of piano-stool complexes incorporating multidentate ligands
is well-established. The in vitro and in vivo anticancer activity [1], as well as the antimicrobial
activity [2] of piano-stool complexes with symmetrical and unsymmetrical chelating ligands
have been demonstrated. The use of chelating ligands reduces significantly the probability of
ligand exchange [3], and modulate the electronic effects at the metal [4], two factors that play
a major role in the biological activity of piano-stool complexes.
Among chelating ligands, Schiff-base derivatives have been widely explored as these
multidentate ligands are relatively easy to synthesize, and they can be functionalized to
generate chelating ligands with tailored properties [5]. They are generally obtained under mild
reaction conditions, in high yields, by the direct condensation of aldehydes with aliphatic or
aromatic primary amines [6]. Despite showing high stability as isolated organic molecules,
some aromatic Schiff-bases are prone to hydrolyze, to regenerate the starting materials [7].
However, hydrolysis can be avoided by electronic and or steric constraints. Most studies
dealing with piano-stool complexes incorporating multidentate Schiff-bases show the ligand
in a bidentate mode [8], and only a few examples show Schiff-base ligands in a tridentate
mode [9].
Remarkably, some Schiff-base compounds exhibit interesting behavior when
coordinated to metal centers, with the carbon atom of the azomethine group becoming
susceptible to nucleophilic attack. Alcohols [10], water [11] and 2,4-pentadione [12] can react
with the azomethine carbon atom to afford the corresponding alkoxylate, hydroxylate, and
acetylacetonate derivatives. In these systems, the nitrogen atom of the azomethine group is
always coordinated to the metal ion, and is involved in five- or six-membered metallacycles.
Therefore, this particular behavior can be used to introduce additional functionality onto
3
Schiff-based complexes, and accordingly to modulate the biological activity of piano-stool
complexes.
Therefore, to design new organometallic compounds with multidentate ligands and to
confirm the Schiff-base carbon atom activation upon coordination to metal ions, a series of
rhodium-based piano-stool complexes of the general formula [(η5-C Me )Rh(L-OR)] (R =
5 5
Me, 1; R = Et, 2; R = Pri, 3) has been prepared in alcohols (methanol, ethanol or isopropanol)
from the tridentate Schiff-base ligand, 5-methyl-4-{(pyridin-2-ylmethylene)amino}-4H-1,2,4-
triazole-3-thiol (L-H), and the dinuclear precursor [(η5-C Me )RhCl ] . The antiproliferative
5 5 2 2
activity of all complexes was evaluated on various cell lines (A2780, A2780cisR, HEK293),
and the results were compared to those obtained for cisplatin, [(η6-C H )Ru(1,3,5-triaza-7-
10 14
phosphaadamantane)Cl ] (RAPTA-C) and the arene ruthenium analogues [(η6-C H )Ru(L-
2 10 14
OR)] (R = Me, 4; R = Et, 5; R = Pri, 6) [13].
4
2. Results and discussion
The dinuclear complex [(η5-C Me )RhCl ] reacts at room temperature with 5-methyl-
5 5 2 2
4-{(pyridin-2-ylmethylene)amino}-4H-1,2,4-triazole-3-thiol (L-H) to give a series of neutral
complexes of the general formula [(η5-C Me )Rh(L-OR)] (R = Me, 1; R = Et, 2; R = Pri, 3)
5 5
(see Scheme 1). The reactions are conducted in alcohols (methanol, ethanol or isopropanol),
and in parallel to the coordination of L-H to rhodium, an alkoxylation occurs on the imine
carbon atom of L to generate, in accordance with the alcohol used, the L-OR ligands. The
coordination of L-OR in a tridentated fashion and the insertion of the alkoxy group generate
two chiral centers in these mononuclear complexes, one at the metal and the other on the
imine carbon atom.
Scheme 1. Synthesis of the rhodium-based piano-stool complexes 1‒3, with the chiral centers
indicated with the symbol * on the complexes.
The addition of alkoxy groups on the Schiff-base ligand was confirmed by mass
spectrometry. The electrospray ionization mass spectra (methanol, positive mode) of
complexes 1‒3 show [M + H]+ peaks corresponding to mononuclear rhodium-based
complexes (see Experimental section). The presence of rhodium in these compounds was
5
evidenced from the isotopic pattern of the cationic peaks, which correlates exactly with the
calculated theoretical isotopic distributions of rhodium species.
The exact position of the alkoxy group on the L-OR ligand was established by 1H and
13C NMR spectroscopy. Indeed, the azomethine proton, initially observed at 8.7 ppm in the
pre-ligand (L-H) [13,14], is upfield-shifted and appears in the complexes around 5‒6 ppm
(see Table 1). A similar movement in chemical shift is observed for the corresponding carbon
atom, which shifts from 164 ppm in L-H to ca. 106 ppm in the L-OR rhodium-based
complexes. The insertion of an alkoxy group on the imine carbon atom generates an
asymmetric carbon atom on L-OR (see Scheme 1). Therefore, in complexes 2 and 3, the O-
CH moiety of the alkoxy group becomes diastereotopic. In complex 2, two multiplets
2
integrating for one proton each are observed at 4.2 and 3.9 ppm, respectively. The poor
resolution of these signals, as well as others, confirms the existence of diastereoisomers in
solution, which are due to the presence of two chiral centers, one at the metal, and one on the
ligand. Integration of the signals and 2D NMR experiments confirm the presence of two
species in solution with, in both cases, in a 2:3 molar ratio.
6
Table 1. 1H NMR data of complexes 1−3 (including multiplicities and integrations).
1 2 3
H 8.55 (d, 1H) 8.54 (d, 0.6H), 8.52 (d, 0.4H) 8.53 (m, 1H)
1
H 7.91 (dd, 1H) 7.89 (dd, 1H) 7.89 (dd, 1H)
2
H 7.48 (m, 1H) 7.48 (m, 1H) 7.46 (m, 1H)
3
H 7.53 (d, 1H) 7.52 (d, 1H) 7.52 (d, 1H)
4
H 5.26 (s, 1H) 5.31 (s, 0.6H), 5.26 (s, 0.4H) 5.34 (s, 0.4H), 5.26 (s, 0.6H)
5
H - 4.18 (m, 1H), 3.91 (m, 1H) 4.43 (m, 0.6H), 3.50 (m, 0.4H)
6
CH (alkoxyl) 3.72 (s, 3H) 1.30 (dd, 3H) 1.30 (m, 6H)
3
CH (triazine) 2.44 (s, 3H) 2.44 (s, 1.2H), 2.43 (s, 1.8H) 2.44 (s, 1.8H), 2.42 (s, 1.2H)
3
CH (Cp*) 1.70 (s, 15H) 1.71 (s, 9H), 1.70 (s, 6H) 1.72 (s, 6H), 1.70 (s, 9H)
3
7
The cytotoxicity of complexes 1‒3 was evaluated in human ovarian cancer cells,
A2780 and A2780cisR (the latter having acquired resistance to cisplatin), as well as non-
tumorigenic human embryonic kidney (HEK293) cells as a gauge of cancer cell selectivity.
The cytotoxicity of complexes 1‒3 were also compared to arene ruthenium analogues bearing
the same L-OH ligands, i.e. [(η6-C H )Ru(L-OR)] (R = Me, 4; R = Et, 5; R = Pri, 6) [13],
10 14
and benchmarked against cisplatin [15] and RAPTA-C [16] (see Table 2). Cisplatin is a
widely used, cytotoxic alkylating agent and, as can be seen from Table 2, is cytotoxic to the
A2780 cancer cells (IC = 2.3 ± 0.6 μM), but displays limited cancer cell selectivity when
50
compared to the IC value of 8.4 ± 0.9 μM in the HEK293 cell line. In comparison, complex
50
2 is ca. 10 fold less cytotoxic to the A2780 cancer cells exhibitng an IC value of 21 ± 2 μM,
50
but advantageously shows no discernable cytotoxicity towards the non-tumorigenic HEK293
cells at the maximum dose tested, i.e. 200 μM.
Table 2. IC values of complexes 1−6, RAPTA-C and cisplatin in cancerous (A2780,
50
A2780cisR) and non-cancerous cell lines (HEK293).
compound A2780 (μM) A2780cisR (μM) HEK293 (μM)
cisplatin 2.3 ± 0.6 31 ± 3 8.4 ± 0.9
RAPTA-C > 200 > 200 > 200
1 > 200 > 200 > 200
2 21 ± 2 > 200 > 200
3 > 200 > 200 > 200
4 > 200 > 200 > 200
5 > 200 > 200 > 200
6 > 200 > 200 > 200
8
Remarkably, complex 2 is the only complex displaying any appreciable cytotoxicity
below 200 μM. It should be noted, however, that ruthenium complexes which are weakly
cytotoxic to cancer cells have progressed to clinical trials [17]. The half-sandwich complex
RAPTA-C is also not cytotoxic to the cell lines studied, and unlike cisplatin, operates via an
epigenetic mechanism of action [18]. Nonetheless, RAPTA-C show high efficacy in vivo
against primary and metastatic tumors when administered at high doses [19], and when used
in combination with other agents [20], it is effective at low doses, even against chemo-
resistant cancers [21]. Consequently, although complexes 1−6 are not cytotoxic, with the
exception of 2 in A2780 cells, they should not be excluded from further biological screening
for putative anticancer activity.
3. Conclusions
Reactions between [(η5-C Me )RhCl ] and the Schiff-base 5-methyl-4-{(pyridin-2-
5 5 2 2
ylmethylene)amino}-4H-1,2,4-triazole-3-thiol in different primary and secondary alcohols
lead to alkoxylation of the imine carbon atom of the ligand. Coordination of the Schiff-base in
a S,N,N-tridentate fashion coupled to the nucleophilic attack on the imine carbon atom
generate two chiral centers in the complexes. The complexes are not endowed with
antiproliferative activity, except for the ethanolate derivative 2, who possesses an IC of 21.4
50
μM on the ovarian cancer cell line A2780 and a remarkable degree of cancer cell selectivity.
9
4. Experimental section
4.1. Materials and methods
All chemicals were purchased from commercial sources and used as received unless specified
otherwise. The starting materials [(η5-C Me )RhCl ] [22] and 5-methyl-4-{(pyridin-2-
5 5 2 2
ylmethylene)amino}-4H-1,2,4-triazole-3-thiol (L-H) [14] were prepared according to
published methods. The 1H and 13C{1H} NMR spectra were recorded on a Bruker Avance II
400 spectrometer using the residual protonated solvent as internal standard. Infrared spectra
were recorded on a Thermo Scientific iS5 ATR/FTIR spectrometer. Electrospray ionization
mass spectra were obtained in positive ion mode on a Bruker FTMS 4.7T BioAPEX II mass
spectrometer, University of Fribourg (Switzerland). UV-visible absorption spectra were
recorded in methanol on a Perkin Elmer UV/Vis spectrophotometer at 10-5 M concentrations.
The elemental analyzes were carried out by the Mikroelementaranalytisches Laboratorium,
ETH Zürich (Switzerland).
4.2 General synthesis of the complexes [(η5-C Me )Rh(L-OR)(1−3).
5 5
The metal precursor [(η5-C Me )RhCl ] (50.0 mg; 0.08 mmol) and 2 equivalents of L H
5 5 2 2
(35.08 mg; 0.16 mmol) were dissolved in dry alcohol (methanol for 1, ethanol for 2 and
isopropanol for 3) (15 mL), before being stirred at room temperature for 6 h. After removal of
the solvent on a rotary evaporator, the orange precipitate was purified by column
chromatography on silica gel, using a mixture of methanol/ethyl acetate/hexane as eluent
(1:4/2/3; 2:2/3/4; 3:2/2/3). Then the isolated solid was dried under vacuum.
1: Yield: 25 mg, 0.051 mmol (64.8%). 1H NMR (400 MHz, MeOD) δ (ppm): 8.55 (d, 1H, J
= 5.2 Hz, CH ), 7.91 (dd, 1H, J = 7.7 Hz, CH ), 7.53 (d, 1H, J = 8.2 Hz CH ),
pyr pyr pyr
7.48 (m, 1H, CH ), 5.26 (s, 1H, NCH), 3.72 (s, 3H, OCH ), 2.44 (s, 3H, CH ), 1.70
pyr 3 3
(s, 15H, C (CH ) ). 13C{1H} NMR (101 MHz, MeOD) δ (ppm): 166.9 (SC=N), 165.7
5 3 5
(CC=N), 152.8 (CH ), 149.1 (OCHC), 140.1 (CH ), 127.1 (CH ), 124.9 (CH ),
pyr pyr pyr pyr
107.4 (NCH), 97.52 (d, J = 71 Hz, C (CH ) ), 28.0 (OCH ), 11.1 (CH ), 9.0
Rh-C 5 3 5 3 3
(C (CH ) ). IR: ʋ (cm-1): 2913.1 (w, CH), 1417.4 (s, C=C), 1085.3 (m, C-O). UV-visible:
5 3 5
(1.0 × 10-5 M, MeOH, 298 K): λ 322 nm (ε = 48070 M-1 ∙ cm-1), 454 nm (ε = 11330 M-1 ∙
max
cm-1), 756 nm (ε = 4990 M-1 ∙ cm-1). ESI-MS (MeOH): m/z = 488.1 [M + H]+. Anal. (%):
10
Calcd for C
20
H
26
N
5
ORhS·H
2
O: C, 47.53; H, 5.58; N, 13.86: Found: C, 48.03; H, 5.38; N,
13.76.
2: Yield: 30 mg, 0.059 mmol (74.8%).1H NMR (400 MHz, MeOD) δ (ppm): Diastereoisomer
A (60%): 8.54 (d, 1H, J = 4.2 Hz, CH ), 7.89 (dd, 1H, J = 7.6 Hz, CH ), 7.52 (d, 1H, J
pyr pyr
= 8.3 Hz, CH ), 7.48 (m, 1H, CH ), 5.31 (s, 1H, NCH), 4.18 (m, 1H, OCH CH ), 3.91
pyr pyr 2 3
(m, 1H, OCH CH ), 2.43 (s, 3H, CH ), 1.71 (s, 15H, C (CH ) ), 1.30 (dd, 3H, J = 7.0
2 3 3 5 3 5
Hz, OCH CH ). Diastereoisomer B (40%): 8.52 (d, 1H, J = 4.2 Hz, CH ), 7.89 (dd, 1H,
2 3 pyr
J = 7.6 Hz, CH ), 7.52 (d, 1H, J = 8.3 Hz, CH ), 7.48 (m, 1H, CH ), 5.26 (s, 1H,
pyr pyr pyr
NCH), 4.18 (m, 1H, OCH CH ), 3.91 (m, 1H, OCH CH ), 2.44 (s, 3H, CH ), 1.70 (s,
2 3 2 3 3
15H, C (CH ) ), 1.30 (dd, 3H, J = 7.0 Hz, OCH CH ). 13C{1H} NMR (101 MHz, MeOD)
5 3 5 2 3
δ (ppm). Diastereoisomer A (60%): 167.1 (SC=N), 165.6 (CC=N), 152.5 (CH ), 149.0
pyr
(OCHC), 139.9 (CH ), 126.9 (CH ), 124.7 (CH ), 106.5 (NCH), 97.33 (d, J = 68
pyr pyr pyr Rh-C
Hz, C (CH ) ), 64.6 (OCH CH ), 15.6 (OCH CH ), 10.9 (CH ), 8.9 (C (CH ) ).
5 3 5 2 3 2 3 3 5 3 5
13C{1H} NMR (101 MHz, MeOD) δ (ppm). Diastereoisomer B (40%): 166.7 (SC=N), 165.5
(CC=N), 152.7 (CH ), 149.0 (OCHC), 139.9 (CH ), 127.0 (CH ), 124.8 (CH ),
pyr pyr pyr pyr
107.3 (NCH), 97.40 (d, J = 66 Hz, C (CH ) ), 64.6 (OCH CH ), 15.6 (OCH CH ),
Rh-C 5 3 5 2 3 2 3
10.9 (CH ), 8.9 (C (CH ) ). IR: ʋ (cm-1): 2982.5 (w, CH), 1415.6 (s, C=C), 1088.3 (s, C-O).
3 5 3 5
UV-visible: (1.0 × 10-5 M, MeOH, 298 K): λ 434 nm (ε = 10160 M-1 ∙ cm-1), 644 nm (ε =
max
5200 M-1 ∙ cm-1). ESI-MS (MeOH): m/z = 502.1 [M + H]+. Anal. (%): Calcd for
C
21
H
28
N
5
ORhS·H
2
O: C, 48.56; H, 5.82; N, 13.48: Found: C, 49.17; H, 5.57; N, 13.77.
3: Yield: 25 mg, 0.048 mmol (60.7%). 1H NMR (400 MHz, MeOD) δ (ppm): Diastereoisomer
A (40%): 8.53 (m, 1H, CH ), 7.89 (dd, 1H, J = 8.6 & 7.3 Hz, CH ), 7.52 (m, 1H,
pyr pyr
CH ), 7.46 (m, 1H, CH ), 5.34 (s, 1H, NCH), 3.50 (m, 1H, CH(CH ) ), 2.42 (s, 3H,
pyr pyr 3 2
CH ), 1.72 (s, 15H, C (CH ) ), 1.30 (m, 6H, CH(CH ) ). Diastereoisomer B (60%): 8.53
3 5 3 5 3 2
(m, 1H, CH ), 7.89 (dd, 1H, J = 8.6 & 7.3 Hz, CH ), 7.52 (m, 1H, CH ), 7.46 (m,
pyr pyr pyr
1H, CH ), 5.26 (s, 1H, NCH), 4.43 (m, 1H, CH(CH ) ), 2.44 (s, 3H, CH ), 1.70 (s,
pyr 3 2 3
15H, C (CH ) ), 1.30 (m, 6H, CH(CH ) ). 13C{1H} NMR (101 MHz, MeOD) δ (ppm):
5 3 5 3 2
Diastereoisomer A (40%): 168.1 (SC=N), 165.7 (CC=N), 152.8 (CH ), 149.3 (OCHC),
pyr
140.2 (CH ), 127.0 (CH ), 124.7 (CH ), 104.4 (NCH), 97.65 (d, J = 71 Hz,
pyr pyr pyr Rh-C
C (CH ) ), 65.6 (CH(CH ) ), 24.0 (CH(CH ) ), 24.0 (CH(CH ) ), 11.1 (CH ), 9.4
5 3 5 3 2 3 2 3 2 3
(C (CH ) ). Diastereoisomer B (60%): 167.1 (SC=N), 166.0 (CC=N), 152.9 (CH ), 149.3
5 3 5 pyr
11
(OCHC), 140.3 (CH ), 127.3 (CH ), 125.2 (CH ) 107.5 (NCH), 97.72 (d, J = 70 Hz,
pyr pyr pyr , Rh-C
C (CH ) ), 70.6 (CH(CH ) ), 24.0 (CH(CH ) ), 24.0 (CH(CH ) ), 11.3 (CH ), 9.3
5 3 5 3 2 3 2 3 2 3
(C (CH ) ). IR: ʋ (cm 1): 2919.2 (w, CH), 1416.0 (s, C=C), 1026.3 (s, C-O). UV-visible: (1.0
5 3 5
× 10-5 M, MeOH, 298 K): λ 327 nm (ε = 40830 M-1 ∙ cm-1), 453 nm (ε = 9510 M-1 ∙ cm-1),
max
745 nm (ε = 5900 M-1 ∙ cm-1). ESI-MS (MeOH): m/z = 516.2 [M + H]+. Anal. (%): Calcd for
C
22
H
30
N
5
ORhS·AcOET: C, 51.74; H, 6.35, N, 11.60: Found: C, 52.42; H, 6.30; N, 12.38.
4.3 Cell culture and cytotoxicity studies
Human ovarian carcinoma (A2780 and A2780cisR) cell lines were obtained from the
European Collection of Cell Cultures. The human embryonic kidney (HEK-293) cell line was
obtained from ATCC (Sigma, Buchs, Switzerland). Penicillin streptomycin, RPMI 1640
GlutaMAX (where RPMI = Roswell Park Memorial Institute), and DMEM GlutaMAX media
(where DMEM = Dulbecco’s modified Eagle medium) were obtained from Life
Technologies, and fetal bovine serum (FBS) was obtained from Sigma. The cells were
cultured in RPMI 1640 GlutaMAX (A2780 and A2780cisR) and DMEM GlutaMAX (HEK-
293) media containing 10% heat-inactivated FBS and 1% penicillin streptomycin at 37 °C and
CO (5%). The A2780cisR cell line was routinely treated with cisplatin (2 μM) in the media
2
to maintain cisplatin resistance. The cytotoxicity was determined using the 3-(4,5-dimethyl 2-
thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay [23]. Cells were seeded in flat-
bottomed 96-well plates as a suspension in a prepared medium (100 μL aliquots and
approximately 4300 cells/ well) and pre-incubated for 24 h. Stock solutions of compounds
were prepared in MilliQ water. The solutions were sequentially diluted to give a final
compound concentration range (0−200 μM). Cisplatin and RAPTA-C were tested as a
positive (0−100 μM) and negative (200 μM) controls respectively. The compounds were
added to the pre-incubated 96-well plates in 100 μL aliquots, and the plates were incubated
for a further 72 h. MTT (20 μL, 5 mg/mL in Dulbecco’s phosphate buffered saline) was added
to the cells, and the plates were incubated for a further 4 h. The culture medium was aspirated,
and the purple formazan crystals, formed by the mitochondrial dehydrogenase activity of vital
cells, were dissolved in DMSO (100 μL/well). The absorbance of the resulting solutions,
directly proportional to the number of surviving cells, was quantified at 590 nm using a
SpectroMax M5e multimode microplate reader (using SoftMax Pro software, version 6.2.2).
The percentage of surviving cells was calculated from the absorbance of wells corresponding
12
to the untreated control cells. The reported IC values are based on the means from two
50
independent experiments, each comprising four tests per concentration level.
Acknowledgements
W.A. and B.T. thank the University of Neuchatel for financial support.
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16
Highlights
-Synthesis of piano-stool complexes with tridentate Schiff-base ligands
-Insertion of alkoxy group occurs on the Schiff-base ligand
-All metal complexes are chiral, at the metal and on the Schiff-base
17
Graphical Abstract
A series of neutral piano-stool complexes [(η5-C Me )Rh(L-OR)] (R = Me, 1; R = Et, 2; R =
5 5
Pri, 3) has been prepared in alcohols (methanol, 1; ethanol, 2; isopropanol, 3) from 5-methyl-
4-{(pyridin-2-ylmethylene)amino}-4H-1,2,4-triazole-3-thiol (L-H) and the dinuclear
precursor [(η5-C Me )RhCl ] . The antiproliferative activity of the complexes was evaluated
5 5 2 2
on cancerous (A2780 and A2780cisR) and non-cancerous (HEK293) cell lines, showing no
significant activity in vitro, except for the ethanolate derivative, which shows an IC of 21.4
50
μM on the ovarian cancer cell line A2780.
Graphical TOC
18
Conflicts of Interest: The authors declare no conflict of interest.
19