BODIPY-phosphane as a versatile tool for easy access to new metal-based theranostics

Article abstract of DOI:10.1039/c2dt32055j

A new BODIPY-phosphane was synthesized and proved to be a versatile tool for imaging organometallic complexes. It also led to easy access to a new family of theranostics, featuring gold, ruthenium and osmium complexes. The compounds' cytotoxicity was test

Full text of DOI:10.1039/c2dt32055j

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BODIPYꢀphosphane as a versatile tool for an easy access to new metalꢀ
based theranostics†
Semra Tasan,a Olivier Zava,b Benoît Bertrand,a Claire Bernhard,a Christine Goze,a Michel Picquet,a
Pierre Le Gendre,a Pierre Harvey,c Franck Denat,a Angela Casini,*d and Ewen Bodio*a
5
Received (in XXX, XXX) Xth XXXXXXXXX 20XX, Accepted Xth XXXXXXXXX 20XX
DOI: 10.1039/b000000x
A new BODIPYꢀphosphane was synthesized and proved a versatile tool as imaging organometallic
complexes. It also led to an easy access to a new family of theranostics, featuring gold, ruthenium and
osmium complexes. The compounds’ cytotoxicity was tested on cancer cells, and their cell uptake was
10 followed by fluorescence microscopy in vitro.
45 quantum yields (ca. фF = 0.5−0.8), excitation and emission in
visible to near infrared spectral region (ca. 500ꢀ700 nm), narrow
and intense emission bands, tunable solubility, nanosecond
fluorescence lifetimes, and weak tripletꢀstate formation render
these fluorescent dyes appealing.11 Moreover, the optical
50 properties of BODIPY are easily tunable via chemical
modifications. Because of these traits, BODIPYs have already
found many applications such as laser dyes, donorꢀacceptor
systems, artificial light harvesters, fluorescent sensors, electronꢀ
transfer reagents, light emitting devices and biological labeling.12
Introduction
Since the pioneer discovery of cisplatin for biological
applications by Rosenberg in the 1960's,1 metal complexes have
become the most currently investigated and used class of
15 compounds in cancer chemotherapy.2 Being far more than just
cisplatin analogues, the thousands of potential metallodrugs
tested so far have opened the way to a still increasing number of
proposed targets and possible mechanisms of action.3 However in
most cases, these mechanisms are still poorly understood.
20 Imaging drugs aimed at understanding their mechanism of action
and studying their pharmacokinetics is thus one of the key
challenges of medicinal chemists today.4 Moreover, it now
appears necessary to determine their sideꢀeffects and provide
some clues to suppress them.
Organometallicꢀarene complexes are indeed very promising
anticancer agents,3,5 as well as gold complexes.6 In several cases,
these new generation anticancer agents are thought active through
different mechanisms with respect to classic platinumꢀbased
drugs involving targeting proteins’ activities rather than DNA,7
30 therefore offering a different approach to therapy. Nowadays,
very few reports describe the labeling of Ruꢀarene
compounds,4cꢀe,8 and none using a fluorescent phosphane ligand.
Phosphanes have been extensively studied for their coordination
chemistry and are notorious to strongly bind metal ions.
35 Noteworthy, phosphanes were scarcely used in labeling metal
complexes for imaging purposes,9 maybe due to their sensitivity
towards oxygen. To overcome this problem, we chose a less
sensitive triarylphosphane residue. The latter was coupled to a
55
Noteworthy, water soluble metallated BODIPY derivatives as
potential probes for bimodal imaging (optical and nuclear) were
recently described by some of us.13 Within this frame, we now
report the synthesis of a new BODIPYꢀphosphane ligand and its
coupling to different metalꢀbased moieties (including RuII, OsII
60 and AuI ions) exhibiting anticancer properties. The study of these
new complexes are discussed in relation to the possible
applications in both imaging and therapeutic applications, and,
therefore, as new theranostic agents.
25
Results and discussion
65 Synthesis of the theranostics
The key synthesis step consists of an efficient coupling of the
phosphane
with
the
BODIPY
moieties.
The
aryldiphenylphosphane activated ester 1 and ethylene diamineꢀ
functionalized BODIPY 2 were prepared by adapting literature
70 procedures (Scheme 1).13,14 1 was obtained in two steps using a
palladoꢀcatalysed coupling reaction between pꢀiodobenzoic acid
and diphenylphosphine, followed by an activation of acid moiety
with Nꢀhydroxysuccinimide (NHS). The synthesis of 2 proceeds
in four steps: formation of the BODIPY backbone from a reaction
75 between 3ꢀethylꢀ2,4ꢀdimethylpyrrole and pꢀformylbenzoic acid,
boronation with BF3, activation of the acid function using Nꢀ
dipyrromethene fluorophore (BODIPY) to form
40 fluorescent coordinating ligand.
Within the rich library of fluorescent dyes, the BODIPYs have
attracted a spectacular growing interest during the last years, due
to their rich photophysical properties.10 Chemical and
photochemical stability, relative high absorption coefficients and
a highly
hydroxysuccinimide and
a condensation of the BODIPYꢀ
activated ester derivative on ethylenediamine yielding 2.
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After coupling 1 and 2, the BODIPYꢀphosphine 3 was 10 ruthenium, its electronic effect on phosphorus resonance is
metallated with areneꢀruthenium, areneꢀosmium and gold
derivatives in mild conditions. The complexation reactions were
monitored by 31P NMR. A significative shift of the resonance was
observed going from the free ligand (singlet at 5.5 ppm) to the
significantly different to that of ruthenium. The osmium and
ruthenium shift the signal to high and down field, respectively.
Ruthenium and osmium dimeric precursors were obtained from
the reaction of their trichloride precursors with 1ꢀisopropylꢀ4ꢀ
5
complexes (singlets at 25.2 ppm for Ru derivative, at ꢀ12.4 ppm 15 methylcyclohexaꢀ1,3ꢀdiene. All the steps were optimized to
for the Os one and at 32.9 ppm for Au complexes). Noteworthy,
no trace of phosphine oxide was observed (no signal around 27ꢀ
28 ppm). Although osmium is assumed to behave similarly to
obtain an excellent overall yield (28ꢀ30% for 10 steps)
(Scheme 1).
20
Scheme 1 Synthetic routes for the synthesis oft he BODIPYꢀPhosphane derivatives
Photophysical properties of compounds 3, 4, 5 and 6
Nonetheless, 4 and 5’s ΦF remain relatively high for these
fluorescent probes. For all fluorophores, the Stokes’ shifts (ꢀν =
40 458ꢀ464 cmꢀ1) compare to those reported for other mesoꢀaryl
substituted BODIPY dyes.17 The fluorescence of 3ꢀ6 were also
measured in water and the emission maxima were almost the
same as for DMSO.
Ligand 3 in DMSO exhibits a typical fluorescence emission band
25 at 539 nm with фF = 95 % (Fig. 1), this emission wavelength
being insensitive to complexation (λem = 539 nm for 3ꢀ6 in
DMSO). Interestingly, the fluorescence is not affected upon
coordination to AuI (ΦF = 85%) but is moderately quenched in
the RuII and OsII cases (ΦF = 57 and 52% respectively), which are
30 prone to photoinduced electron transfer,15 as well as to promote
phosphorescence of the BODIPY.16
45 Preliminary stability studies of 3, 4, 5 and 6
The stability of 4ꢀ6 in DMSO and aqueous solutions was studied.
After 24 h in DMSO, almost no change was detected in the
absorption and emission spectra. In phosphate buffer (PBS
(Phosphate Buffered Saline), pH 7.4), the three species showed
50 different stabilities as the UVꢀvisible spectra changed with time,
most likely due to aggregation processes. The absorption spectra
are characterized by a strong absorption band in the 500ꢀ600 nm
range. The most stable complex in the series in physiologicalꢀ
type conditions is the AuI derivative 6, and its absorption
55 spectrum remained unaltered for 12 h incubation (data not
shown). Conversely, the OsII and RuII complexes displayed a
decrease of signal after 9 h only. Under the same conditions, the
spectrum of the BODIPY ligand alone remained unaltered over
24 h
60
Fig. 1 Emission spectra of the BODIPYꢀphosphane 3 and its different
31P and 1HꢀNMR studies combined with mass spectrometry
experiments were performed in order to evaluate the stability of
35
complexes in DMSO at 298 K
2
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the organometallic residue of compounds 4ꢀ6 (the details of this
stability study are described in the ESI). Interestingly, these
compounds appeared stable in the presence of cell culture
medium: RPMI 1640 + 10% fetal calf serum and antibiotics at
37°C. Only little degradation was observed after several days
with the loss of one chloride. Consequently, one can assume that
modifications of optical properties after 9 or 12 h are not due to a
degradation of the complexes, but most likely to aggregation
processes.
5
10
IC50 determinations on cancer cells
Complexes 4ꢀ6 were further tested for their antiproliferative
effects on human ovarian cancer cell lines sensitive (A2780S)
and resistant to cisplatin (A2780cisR) (Table 1). Generally, the
15 compounds showed moderate cytotoxic properties with respect to
cisplatin but markedly more pronounced with respect to the free
BODIPY ligand 3, the most active compound being the Au
derivative 6. Interestingly, an improved cytotoxic effect for the
Ru derivative 4 with respect to the organometallic analogue
20 RAPTAꢀC, was noted.
50
Fig. 2 Confocal microscopy images of A2780S cells incubated 2 h at
37 °C with 3ꢀ6. Pictures show cells in transmitted light (Trans light),
green channel (exc 488 nm, emission 520 nm), red channel (exc 515 nm,
emission 550 nm) and coꢀlocalised pixel map, where grey pixels mean
complete coꢀlocalisation.caption
55 Conclusions
BODIPYꢀphosphane derivatives proved promising agents for
Table 1. Comparison of the IC50 values of 4ꢀ6 after 72 h incubation with
human ovarian carcinoma cell lines cisplatin sensitive (A2780S) and
resistant (A2780cisR), with those for cisplatin and RAPTAꢀC.
labeling metals and organometallic entities. Indeed, three metalꢀ
containing BODIPY compounds based on Ru, Os and Au have
been prepared and studied for their spectroscopic and biological
60 properties in vitro. The BODIPY ligand and the metal complexes
exhibit fluorescence and water solubility, thus partially solving
one of the problems preventing the use of BODIPY agents for in
vivo optical imaging. Indeed, the fluorescence characteristics of
the BODIPYꢀphosphane moiety were maintained upon metal
65 complexation in a way that allowed monitoring the compounds
uptake in cancer cells in vitro. The compounds were moderately
cytotoxic in the selected cancer cell lines, and showed a
preference for accumulation in the cell membranes without being
able to reach the nuclei. Moreover, the cell uptake does not seem
70 to be mediated by active transport, and the compounds’
fluorescence staining pattern appears to be determined by the
BODIPYꢀphosphane moiety. The latter feature could be
exploited, for example, to target metal complexes with affinity
towards specific membrane proteins and enzymes. Interestingly,
75 these BODIPYꢀbased compounds show similar membrane stains
to the amphiphilic styryl dyes FM4ꢀ64 and FM1ꢀ43, widely used
in microscopy studies of endomembrane trafficking, and
therefore could also be used in the study of membrane trafficking
or of lipid metabolism disorders in cells.20
Compound
A2780S
> 150
A2780cisR
> 150
3
4
5
6
50 ± 12
141 ± 11
32 ± 16
2.5 ± 0.9
> 300
49 ± 15
67 ± 12
37 ± 10
40 ± 7.0
> 300
cisplatin
RAPTAꢀC18
25
In vitro confocal microscopy experiments
The detection of the BODIPYꢀphosphane ligand 3 and its
complexes 4ꢀ6 in living cells by fluorescence microscopy was
also performed. The complexes were incubated with A2780S
30 cells for 1 and 2 h at 37°C and then visualized by confocal
fluorescence microscopy. The in vitro imaging of 3ꢀ6 showed that
they all rapidly bind to the biological membranes, with no clear
specificity (Fig. 2), this observation being consistent with
previous cell studies on BODIPYꢀbased fluorophores.19
35 Moreover, no difference was noted between the BODIPYꢀ
phosphane ligand and its metal derivatives implying that the
uptake and distribution properties of the compounds are mainly
determined by the BODIPY moiety. Moreover, similar staining
and fluorescence properties are also observed in cells incubated at
40 4°C, temperature at which active transport mechanisms are
commonly assumed to be inhibited.
80
Thus, although further studies are in progress in order to
optimize the BODIPYꢀphosphane tool for its application in vivo
(e.g. improvement of water solubility and modification of optical
properties to be suitable for near infrared fluorescence), this
approach to theranostic agents is valuable and worth exploring.
Noteworthy, in addition to the observed fluorescence (Fig. 1),
we noticed a second redꢀshifted fluorescence signal (550 nm).
45 This unexpected result is not unprecedented as previous reports
on different BODIPYs showed that upon local high
concentrations, two fluorophores can stack and form a dimer with
a marked shift of fluorescence.20
85 Acknowledgments
Support was provided by the Conseil Régional de Bourgogne
(PARI & 3MIM programs), the Ministère de l’Enseignement
Supérieur et de la Recherche, the Centre National de la Recherche
Scientifique (CNRS), and the University of Groningen (Rosalind
90 Franklin fellowship to AC). COST Action CM0902
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"Translocation of metallic ions across the biomembrane and their
fate after uptake" is acknowledged for fruitful discussion. Dr. F.
Picquet and Ms M.ꢀJ. Penouilh are gratefully acknowledged for
(s, 1C, CC6H4ꢀCO), 133.3 (d, 2C, 2JCꢀP = 20.1 Hz, CHortho), 134.2 (d,
4C, 2JCꢀP = 20.1 Hz, CHortho), 136.1 (d, 2C, 1JCꢀP = 10.2 Hz, Cipso),
1
145.5 (d, 1C, JCꢀP = 14.3 Hz, Cipso), 171.9 (s, 1C, CCOOH).
HRꢀMS analysis and Mr M. Pirrotta for his help during the 60 31P{1H} NMR (202.5 MHz, CD2Cl2): δ (ppm) ꢀ 4.7. HRꢀMS
5
synthesis of BODIPY derivatives.
(ESI): m/z calcd for C19H15O2P ꢀ H+: 305.07259 [MꢀH]ꢀ. Found:
305.07199. IR: (cmꢀ1) = 1296 (νCꢀO), 1687 (νC=O), 2551ꢀ3057
(νOꢀH).
Experimental
General information
65 Second
(diphenylphosphino)benzoate (1).
The reaction was carried out under Ar atmosphere
NꢀHydroxysuccinimide (1.34 g, 11.6 mmol) and 1ꢀethylꢀ3ꢀ(3ꢀ
dimethylaminopropyl) carbodiimide hydrochloride (EDC.HCl,
step:
synthesis
of
succinimid-N-yl
4-
All solvents were dried and distilled under argon before use.
BODIPYꢀacid derivative was synthesized as reported by
.
10 Tomasulo et al.21 and pꢀcymeneꢀruthenium and osmium dimers
were obtained following Bennett’s procedures.22 All other
reagents were commercially available and used as received. The 70 2.22 g, 11.6 mmol) were added to
a
solution of
analyses were performed at the " Plateforme d’Analyses
Chimiques et de Synthèse Moléculaire de l’Université de
15 Bourgogne ". The identity and purity (≥ 95%) of the complexes
were unambiguously established using highꢀresolution mass
pꢀ(diphenylphosphino)benzoic acid (1.77 g, 5.8 mmol) in
dichloromethane (15 mL). The reaction was stirred at room
temperature for 1 h. The mixture was washed with degassed
water (3×10 mL) and the organic layers were dried over
spectrometry and multinuclear NMR spectroscopy. The exact 75 magnesium sulfate. The solvent was removed under reduced
mass of the complexes were obtained on a Thermo LTQ Orbitrap
XL ESIꢀMS. 1H (300.13, 500.13, or 600.23 MHz), 13C (75.5,
20 125.8, or 150.9 MHz), 11B (192.5 MHz) and 31P (121.5, 202.5, or
242.9 MHz) NMR spectra were recorded on Bruker 300 Avance
pressure to obtain a light yellow powder. The residue was
purified by silica gel column chromatography (eluent:
dichloromethane) to afford the desired product as a white powder
(2.14 g, 92%).
1
III, Bruker 500 Avance III, or
Bruker 600 Avance II 80 Mp: 174ꢀ175 °C. H NMR (600.23 MHz, CD2Cl2): δ (ppm) 2.87
spectrometers. Chemical shifts are quoted in parts per million (δ)
relative to tetramethylsilane, TMS (1H and 13C), using the
25 residual protonated solvent (1H) or the deuterated solvent (13C) as
an internal standard. Alternatively, 85% H3PO4 (31P) and
(s, 4H, CH2), 7.34ꢀ7.43 (m, 12H, Ph), 8.05 (m, 2H, C6H4ꢀCOOR
H
ortho). 13C{1H} NMR (125.8 MHz, CDCl3): δ (ppm) 25.8 (s, 2C,
3
CH2), 124.9 (s, 1C, CPhCO), 129.0 (d, 4C, JCꢀP = 7.3 Hz, CHmeta),
3
129.6 (s, 2C, CHpara), 130.3 (d, 2C, JCꢀP = 6.2 Hz, CHmeta), 133.5
BF3.OEt2 (11B) were used as external standards. The coupling 85 (d, 2C, JCꢀP = 18.2 Hz, CHortho), 134.2 (d, 4C, JCꢀP = 20.1 Hz,
2
2
1
constants are reported in Hertz. All aromatic positions: ortho,
meta, para are defined using phosphorus as main groupment.
30 Infrared spectra were recorded on a Bruker Vector 22 FTꢀIR
spectrophotometer (transmission mode) with 1% sample mixed
with potassium bromide. The melting points were determined on 90 Found: 426.08484. IR:
a BÜCHI Melting Point Bꢀ545 instrument. 1761 (νC=O).
CHortho), 135.8 (d, 2C, JCꢀP = 10.3 Hz, Cipso), 147.6 (d, 1C,
1JCꢀP = 15.9 Hz, Cipso), 161.9 (s, 1C, CCOOR), 169.3 (s, 2C, CCON).
31P{1H} NMR (242.9 MHz, CD2Cl2): δ (ppm) ꢀ 4.3. HRꢀMS
(ESI): m/z calcd for C23H18NO4P + Na+: 426.08657 [M+Na]+.
(cmꢀ1) = 1202 (νCꢀO), 1738 (νC=O),
35 Synthesis
benzoate (1).
of
succinimidꢀNꢀyl
4ꢀ(diphenylphosphino)
Synthesis of 4,4ꢀdifluoroꢀ8ꢀ(4ꢀ((2ꢀaminoethyl)carbamoyl)
phenyl)ꢀ1,3,5,7ꢀtetramethylꢀ2,6ꢀdiethylꢀ4ꢀboraꢀ3a,4aꢀdiazaꢀsꢀ
95 indacene (2)
First step: synthesis of p-(diphenylphosphino)benzoic acid.
The reaction was carried out under Ar atmosphere. p-Iodobenzoic
acid (2.00 g, 8.1 mmol) and Pd(OAc)2 (3.3 mg, 0.015 mmol)
40 were dissolved in acetonitrile (20 mL). Distilled triethylamine
(2.25 mL, 16.1 mmol) and diphenylphosphine (1.50 mL, 8.6
mmol) were added and the mixture was refluxed for 72 h. The
solvent was removed under reduced pressure to obtain a light
yellow solid. The solid was dissolved in a 1 M KOH aqueous
45 solution (50 mL) and the solution was washed three times with
diethyl ether (3×30 mL). The aqueous layer was acidified with a
2 M HCl aqueous solution (until pH = 2ꢀ3). The phosphine
precipitated as a yellow solid. Aqueous layer was extracted three
times with dichloromethane (3×30 mL). Dichloromethane organic
50 layers were combined and dried over magnesium sulfate. The
solvent was removed under reduced pressure to afford the desired
pure product as a light yellow powder (2.22 g, 90%).
First step: synthesis of 4,4-difluoro-8-(4-((succinimid-N-
yl)oxycarbonyl)phenyl)-1,3,5,7-tetramethyl-2,6-diethyl-4-bora-
3a,4a-diaza-s-indacene.
The reaction was carried out under Ar atmosphere. BODIPYꢀ
100 acid derivative (566 mg, 1.3 mmol), EDC.HCl (513 mg, 2.7
mmol), Nꢀhydroxysuccinimide (308 mg, 2.7 mmol) were
dissolved in dichloromethane (150 mL). The mixture was stirred
at room temperature for 3 h. The reaction was monitored by TLC
(silica gel, eluent: ethyl acetate/heptane (1/1)). The solution was
105 washed twice with distilled water (2×100 mL), dried over
magnesium sulfate and solvent was removed under reduced
pressure. The residue was purified by silica gel column
chromatography (eluent: ethyl acetate/heptane (1/1)) to afford the
BODIPYꢀester activated derivative as a red powder with green
110 glints (693 mg, > 99%).
1
Mp: 150ꢀ151 °C. 1H NMR (500.13 MHz, CD2Cl2): δ (ppm) 7.33ꢀ
Mp: 288ꢀ289 °C. H NMR (300.13 MHz, CDCl3): δ (ppm) 0.96
(t, 6H, 3J = 7.5 Hz, CH2CH3), 1.25 (s, 6H, CH3), 2.27 (q, 4H, 3J =
7.5 Hz, CH2CH3), 2.52 (s, 6H, CH3), 2.93 (s, 4H, CH2CO), 7.44
(d, 2H, 3J = 8.3 Hz, C6H4), 8.21 (d, 2H, 3J = 8.3 Hz, C6H4).
7.41 (m, 12H, Ph), 8.03 (m, 2H, C6H4ꢀCO). 13C{1H} NMR (75.5
3
55 MHz, CDCl3): δ (ppm) 128.9 (d, 4C, JCꢀP = 7.1 Hz, CHmeta),
3
129.4 (s, 2C, CHpara), 130.0 (d, 2C, JCꢀP = 6.6 Hz, CHmeta), 131.7
4
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CH3), 2.31 (q, 4H, 3J = 7.7 Hz, CH2CH3), 2.50 (s, 6H, CH3), 3.72
13C{1H} NMR (75.5 MHz, CDCl3): δ (ppm) 12.1 (s, 2C, CH3),
12.7 (s, 2C, CH3), 14.6 (s, 2C, CH3CH2), 17.3 (s, 2C, CH3CH2), 60 (m, 4H, NHCH2), 7.16 (br s, 1H, NH), 7.29ꢀ7.39 (m, 12H,
25.7 (s, 2C, CH2CO), 125.6 (s, 1C, CArꢀBODIPY), 129.3 (s, 2C,
CHArꢀBODIPY), 131.2 (s, 2C, CPyrrole), 133.3 (s, 2C, CHArꢀBODIPY),
137.7 (s, 2C, CPyrrole), 138.0 (s, 2C, CPyrrole), 138.7 (s, 1C,
CBODIPY), 143.0 (s, 1C, CArꢀBODIPY), 154.6 (s, 2C, CPyrrole), 161.4
C6H4PPh2), 7.41 (d, 2H, 3J = 8.4 Hz, C6H4), 7.76 (dd, 2H, 3J = 8.3
3
Hz, JHꢀP = 1.4 Hz, C6H4CONH), 7.95 (d, 2H, 3J = 8.2 Hz,
5
C6H4CONH). 13C{1H} NMR (125.8 MHz, CDCl3): δ (ppm) =
12.0 (s, 2C, CH3), 12.7 (s, 2C, CH3), 14.7 (s, 2C, CH3CH2), 17.2
(s, 1C, CO2), 169.2 (s, 2C, CON). 11B NMR (192.5 MHz, 65 (s, 2C, CH3CH2), 41.0 (s, 1C, CH2NH), 41.8 (s, 1C, CH2NH),
CDCl3): δ (ppm) 0.8 (t, 1JB-F = 33.1 Hz). HRꢀMS (ESI): m/z calcd
127.0 (d, 2C, JCꢀP = 6.8 Hz, CHArꢀP), 128.0 (s, 2C, CHArꢀBODIPY),
128.8 (d, 4C, 3JCꢀP = 7.3 Hz, CHPhꢀP), 129.0 (s, 2C, CHPhꢀP), 129.3
(s, 2C, CHArꢀBODIPY), 130.5 (s, 2C, CPyrrole), 133.2 (s, 2C, CPyrrole),
3
for C28H30BF2N3O4
+
Na+: 544.21896 [M+Na]+. Found:
10 544.21783. UVꢀVis (CH3CN): λmax (nm) (ε, molꢀ1.cmꢀ1) 378
(7900), 492 (sh, 23200), 525 (73000). IR: (cmꢀ1) 1190 (νCꢀO),
1744 (νC=O), 1776 (νC=O).
2
133.6 (d, 2C, JCꢀP = 18.7 Hz, CHArꢀP), 133.8 (s, 1C, CArꢀP), 134.0
2
70 (d, 4C, JCꢀP = 20.0 Hz, CHPhꢀP), 134.3 (s, 1C, CArꢀBODIPY), 136.3
(d, 2C, 1JCꢀP = 10.7 Hz, CPhꢀP), 138.3 (s, 2C, CPyrrole), 138.9 (s, 1C,
1
Second step: synthesis of 4,4-difluoro-8-(4-((2-aminoethyl)
15 carbamoyl)phenyl)-1,3,5,7-tetramethyl-2,6-diethyl-4-bora-3a,4a-
diaza-s-indacene (2).
CBODIPY), 139.5 (s, 1C, CArꢀBODIPY), 142.8 (d, 1C, JCꢀP = 13.7 Hz,
CArꢀP), 154.3 (s, 2C, CPyrrole),167.9 (s, 1C, CO), 168.9 (s, 1C, CO).
31P{1H} NMR (242.9 MHz, CD2Cl2): δ (ppm) ꢀ 5.5. 11B NMR
1
BODIPYꢀactivated ester (314 mg, 0.6 mmol) was dissolved in 75 (192.5 MHz, CDCl3): δ (ppm) 0.79 (t, JB-F = 33.7 Hz). HRꢀMS
dichloromethane (30 mL). Ethylenediamine (1.60 mL,
23.9 mmol) was added. The resulting solution was stirred at room
20 temperature for 16 h. The reaction was monitored by TLC (silica
gel, eluent: ethyl acetate/heptane (1/1)). The solution was washed
twice with water (2×50 mL), dried over magnesium sulfate and
the solvent was removed under reduced pressure. The residue was
purified by silica gel column chromatography (eluent:
25 ethanol/NH4OH (4/1)) to afford compound 2 as a red powder
with green glints. (262 mg, 93%).
(ESI): m/z calcd for C45H46BF2N4O2P +
Na+: 777.33117
[M+Na]+. Found: 777.32996. UVꢀVis (DMSO): λmax (nm) (ε, Mꢀ1
cmꢀ1) 378 (6500), 492 (sh, 17900), 526 (55300). IR: (cmꢀ1)
1636ꢀ1684 (νC=O), 3415 (νNꢀH).
80 Synthesis
of
dichloro[(η6ꢀpꢀcymene)(4,4ꢀdifluoroꢀ8ꢀ(4ꢀ((2ꢀ(4ꢀ(diphenylphos
phino)benzamido)ethyl)carbamoyl)phenyl)ꢀ1,3,5,7ꢀ
tetramethylꢀ2,6ꢀdiethylꢀ4ꢀboraꢀ3a,4aꢀdiazaꢀsꢀindacene)ꢀ
ruthenium(II)] (4).
Mp (decomposition): 177ꢀ179 °C. 1H NMR (300.13 MHz,
85 The reaction was carried out under under Ar atmosphere
BODIPYꢀphosphine derivative 3 (52 mg, 0.07 mmol) and
[RuCl2(pꢀcymene)]2 (19 mg, 0.03 mmol) were dissolved in
degassed benzene (3 mL). The resulting mixture was stirred at
room temperature for 17 h. The reaction was monitored by 31P
90 NMR (242.9 MHz, 300 K). The solvent was removed under
reduced pressure. The ruthenium complex 4 was isolated as a
bright red powder (65 mg, 89%).
.
3
CDCl3): δ (ppm) 0.99 (t, 6H, J = 7.6 Hz, CH2CH3), 1.30 (s, 6H,
CH3), 2.32 (q, 4H, 3J = 7.6 Hz, CH2CH3), 2.50 (s, 6H, CH3), 2.97
30 (t, 2H, 3J = 5.8 Hz, CH2NH2), 3.51 (q, 2H, 3J = 5.8 Hz,
CONHCH2), 6.91 (br s, 1H, NH), 7.41 (d, 2H, 3J = 8.1 Hz, C6H4),
7.95 (d, 2H, 3J = 8.1 Hz, C6H4). 13C{1H} NMR (125.8 MHz,
CDCl3): δ (ppm) 11.8 (s, 2C, CH3), 12.5 (s, 2C, CH3), 14.6 (s,
2C, CH3CH2), 17.0 (s, 2C, CH3CH2), 42.2 (s, 1C, CH2NH2), 43.8
35 (s, 1C, CONHCH2), 129.1 (s, 2C, CHArꢀBODIPY), 129.9 (s, 1C, CArꢀ
BODIPY), 130.4 (s, 2C, CPyrrole), 131.0 (s, 2C, CHArꢀBODIPY), 133.3
(s, 2C, CPyrrole), 138,2 (s, 2C, CPyrrole), 138.6 (s, 1C, CBODIPY),
141.8 (s, 1C, CArꢀBODIPY), 154.6 (s, 2C, CPyrrole),171.4 (s, 1C, CO).
11B NMR (192.5 MHz, CDCl3): δ (ppm) 0.8 (t, 1JB-F = 32.9 Hz).
40 HRꢀMS (ESI): m/z calcd for C26H33BF2N4O + H+: 467.27762
Mp (decomposition): 203ꢀ205 °C.
3
1H NMR (500.13 MHz, CD2Cl2): δ (ppm) 0.98 (t, 6H, J = 7.7
3
95 Hz, CH2CH3), 1.12 (d, 6H, J = 7.0 Hz, (CH3)2CH), 1.27 (s, 6H,
CH3), 1.86 (s, 3H, CH3pꢀcymeneꢀRu), 2.31 (q, 4H, 3J = 7.7 Hz,
CH2CH3), 2.50 (s, 6H, CH3), 2.77 (hept, 1H, 3J = 7.0 Hz,
3
(CH3)2CH), 3.69 (m, 4H, NHCH2), 5.00 (d, 2H, J = 6.4 Hz, pꢀ
[M+H]+. Found: 467.27883. IR:
(cmꢀ1) 1186 (νCꢀN), 1542
3
cymeneꢀRu), 5.23 (d, 2H, J = 6.4 Hz, pꢀcymeneꢀRu), 7.27 (br s,
(δNH2), 1699ꢀ1636 (νC=O), 3419 (νNꢀH).
100 1H, NH), 7.39ꢀ7.48 (m, 9H, Ph + NH), 7.73ꢀ7.81 (m, 6H,
C6H4CONH + Ph), 7.89 (t, 2H, 3J = 8.7 Hz, PPh2), 7.95 (d, 2H, 3J
= 8.2 Hz, C6H4CONH). 13C{1H} NMR (125.8 MHz, CD2Cl2): δ
(ppm) 12.1 (s, 2C, CH3), 12.7 (s, 2C, CH3), 14.8 (s, 2C,
CH3CH2), 17.4 (s, 2C, CH3CH2), 18.0 (s, 1C, CH3 pꢀcymeneꢀRu), 22.1
105 (s, 2C, CH(CH3)2 pꢀcymeneꢀRu), 30.8 (s, 1C, CH(CH3)2 pꢀcymeneꢀRu),
Synthesis of 4,4ꢀdifluoroꢀ8ꢀ(4ꢀ((2ꢀ(4ꢀ(diphenylphosphino)
45 benzamido)ethyl)carbamoyl)phenyl)ꢀ1,3,5,7ꢀtetramethylꢀ2,6ꢀ
diethylꢀ4ꢀboraꢀ3a,4aꢀdiazaꢀsꢀindacene (3).
The reaction was carried out under Ar atmosphere. BODIPYꢀ
41.2 (s, 1C, CH2NH), 41.8 (s, 1C, CH2NH), 87.8 (s, 2C, CH
pꢀ
amine derivative
2 (151 mg, 0.32 mmol) and distilled
cymeneꢀRu), 89.4 (s, 2C, CH pꢀcymeneꢀRu), 97.0 (s, 1C, C pꢀcymeneꢀRu),
triethylamine (0.23 mL, 1.65 mmol) were dissolved in degassed
50 chloroform (15 mL). Compound 1 (157 mg, 0.39 mmol) was
added. The resulting mixture was refluxed for 19 h. The reaction
was monitored by 31P NMR (242.9 MHz, 300 K) and TLC (silica
gel, eluent: ethyl acetate). The solvent was removed under
reduced pressure. The residue was purified by filtration over a
55 silica plug (eluent: dichloromethane/ethyl acetate (7/3)) to afford
BODIPYꢀderivative 3 as a red powder (179 mg, 73%).
3
111.1 (s, 1C, C pꢀcymeneꢀRu), 126.5 (d, 2C, JCꢀP = 9.9 Hz, CHArꢀP),
128.3 (s, 2C, CHArꢀBODIPY), 128.6 (d, 4C, 3JCꢀP = 10.1 Hz, CHPhꢀP),
110 129.1 (s, 2C, CHArꢀBODIPY), 130.8 (s, 2C, CPyrrole), 131.0 (s, 2C,
1
CHPhꢀP), 133.5 (s, 2C, CPyrrole), 133.9 (d, 2C, JCꢀP = 44.9 Hz,
2
CHPhꢀP), 134.7 (d, 4C, JCꢀP = 9.6 Hz, CHPhꢀP), 134.9 (s, 1C, CArꢀ
BODIPY), 135.0 (d, 2C, 2JCꢀP = 9.6 Hz, CHArꢀP), 135.9 (s, 1C, CArꢀP),
3
137.9 (d, 1C, JCꢀP = 44.5 Hz, CArꢀP), 138.8 (s, 2C, CPyrrole), 139.3
Mp (decomposition): 175ꢀ177 °C. 1H NMR (300.13 MHz,
CD2Cl2): δ (ppm) 0.98 (t, 6H, 3J = 7.7 Hz, CH2CH3), 1.27 (s, 6H,
115 (s, 1C, CBODIPY), 139.6 (s, 1C, CArꢀBODIPY), 154.4 (s, 2C, CPyrrole),
167.7 (s, 1C, CO), 168.3 (s, 1C, CO). 31P{1H} NMR (242.9 MHz,
This journal is © The Royal Society of Chemistry [year]
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CD2Cl2):
δ
(ppm) 25.2. HRꢀMS (ESI): m/z calcd for
Mp (decomposition): 205ꢀ207 °C. 1H NMR (500.13 MHz,
C55H60BCl2F2N4O2PRu + Na+: 1083.28278 [M+Na]+. Found:
CD2Cl2): δ (ppm) 0.98 (t, 6H, 3J = 7.5 Hz, CH2CH3), 1.27 (s, 6H,
1083.28357. UVꢀVis (DMSO): λmax (nm) (ε, Mꢀ1 cmꢀ1) 492 (sh, 60 CH3), 2.31 (q, 4H, 3J = 7.5 Hz, CH2CH3), 2.50 (s, 6H, CH3), 3.74
21500), 378 (7900), 526 (70500). IR: (cmꢀ1) 1653ꢀ1700 (νC=O),
3419 (νNꢀH).
(m, 4H, NHCH2), 7.16 (br s, 1H, NH), 7.42 (d, 2H, J = 8.3 Hz,
C6H4), 7.47ꢀ7.66 (m, 13H, Ph + NH), 7.92ꢀ7.95 (m, 4H,
C6H4CONH). 13C{1H} NMR (125.8 MHz, CD2Cl2): δ (ppm) 12.1
(s, 2C, CH3), 12.7 (s, 2C, CH3), 14.8 (s, 2C, CH3CH2), 17.4 (s,
65 2C, CH3CH2), 41.4 (s, 1C, CH2NH), 42.1 (s, 1C, CH2NH), 128.1
(d, 2C, 3JCꢀP = 11.8 Hz, CHArꢀP), 128.2 (s, 2C, CHArꢀBODIPY), 128.6
(d, 2C, 1JCꢀP = 62.6 Hz, CHPhꢀP), 129.3 (s, 2C, CHArꢀBODIPY), 129.8
3
5
Synthesis of dichloro[(η6ꢀpꢀcymene)(4,4ꢀdifluoroꢀ8ꢀ(4ꢀ((2ꢀ(4ꢀ
(diphenylphosphino)benzamido)ethyl)carbamoyl)phenyl)ꢀ1,3,
5,7ꢀtetramethylꢀ2,6ꢀdiethylꢀ4ꢀboraꢀ3a,4aꢀdiazaꢀsꢀindacene)ꢀ
osmium(II)] (5).
10 The reaction was carried out under Ar atmosphere. BODIPYꢀ
phosphine derivative 3 (51 mg, 0.07 mmol) and [OsCl2(pꢀ
cymene)]2 (24 mg, 0.03 mmol) were dissolved in degassed
benzene (5 mL). The resulting mixture was stirred at room
temperature for 15 h. The reaction was monitored by 31P NMR
15 (242.9 MHz, 300 K). The solvent was removed under reduced
pressure. The osmium complex 5 was isolated as a red powder
(73 mg, 94%).
3
(d, 4C, JCꢀP = 12.2 Hz, CHPhꢀP), 130.8 (s, 2C, CPyrrole), 132.7 (s,
1
2C, CHPhꢀP), 133.0 (d, 1C, JCꢀP = 61.0 Hz, CArꢀP), 133.6 (s, 2C,
70 CPyrrole), 134.6 (d, 4C, 2JCꢀP = 13.6 Hz, CHPhꢀP), 134.7 (d, 2C, 2JCꢀP
= 14.5 Hz, CHArꢀP), 134.8 (s, 1C, CArꢀBODIPY), 137.9 (s, 1C, CArꢀP),
138.8 (s, 2C, CPyrrole), 139.4 (s, 1C, CBODIPY), 139.7 (s, 1C, CArꢀ
BODIPY), 154.5 (s, 2C, CPyrrole), 167.4 (s, 1C, CO), 168.4 (s, 1C,
CO). 31P{1H} NMR (242.9 MHz, CD2Cl2): δ (ppm) 32.9. HRꢀMS
75 (ESI): m/z calcd for C45H46BClF2N4O2PAu + Na+: 1009.26658
[M+Na]+. Found: 1009.26575. UVꢀVis (DMSO): λmax (nm) (ε, Mꢀ
Mp (decomposition): 195ꢀ197 °C. 1H NMR (500.13 MHz,
CD2Cl2): δ (ppm) 0.98 (t, 6H, 3J = 7.6 Hz, CH2CH3), 1.16 (d, 6H,
20 3J = 7.0 Hz, (CH3)2CH), 1.27 (s, 6H, CH3), 1.97 (s, 3H, CH3 pꢀ
cymeneꢀOs), 2.31 (q, 4H, J = 7.6 Hz, CH2CH3), 2.50 (s, 6H, CH3),
2.68 (hept, 1H, J = 7.0 Hz, (CH3)2CH), 3.70 (m, 4H, NHCH2),
1
cmꢀ1) 378 (7000), 492 (sh, 17800), 526 (59200). IR: (cmꢀ1) =
3335 (νNꢀH), 1653 (νC=O).
3
3
80 Fluorescence measurements
3
3
5.18 (d, 2H, J = 5.8 Hz, C6H4ꢀOs), 5.42 (d, 2H, J = 5.8 Hz,
C6H4ꢀOs), 7.23 (br s, 1H, NH), 7.37 (br s, 1H, NH), 7.39ꢀ7.46 (m,
The steadyꢀstate fluorescence emission and excitation spectra
were obtained by using a JASCO FP8560 spectrofluorometer
instrument. All fluorescence spectra were corrected for
85 instrument response.
3
4
25 8H, Ph), 7.71 (td, 4H, J = 8.6 Hz, J = 1.8 Hz, PPh2), 7.76 (dd,
2H, 3J = 8.4 Hz, 3JHꢀP = 1.9 Hz, C6H4CONH), 7.87 (t, 2H, 3J = 8.4
Hz, PPh2), 7.95 (d, 2H, 3J = 8.2 Hz, C6H4CONH). 13C{1H} NMR
(125.8 MHz, CD2Cl2): δ (ppm) 12.1 (s, 2C, CH3), 12.7 (s, 2C,
CH3), 14.8 (s, 2C, CH3CH2), 17.4 (s, 2C, CH3CH2), 18.0 (s, 1C,
30 CH3 pꢀcymeneꢀOs), 22.4 (s, 2C, CH(CH3)2 pꢀcymeneꢀOs), 30.7 (s, 1C,
CH(CH3)2 pꢀcymeneꢀOs), 41.2 (s, 1C, CH2NH), 41.9 (s, 1C, CH2NH),
80.6 (s, 2C, CH pꢀcymeneꢀOs), 81.0 (s, 2C, CH pꢀcymeneꢀOs), 89.6 (s, 1C,
The fluorescence quantum yield ( ΦF
) was calculated from
equation 1.
∞
I (λ ,λ )dλ
∫
F
E
F
F
(
λE )
R
3
ΦF
n2
1−10−A
0
∞
C pꢀcymeneꢀOs), 103.3 (s, 1C, C pꢀcymeneꢀOs), 126.5 (d, 2C, JCꢀP = 10.4
=
×
×
λE
)
ΦFR nR2
1−10−A(
Hz, CHArꢀP), 128.3 (s, 2C, CHArꢀBODIPY), 128.5 (d, 4C, 3JCꢀP = 10.1
I (λ ,λ )dλ
∫
FR
E
F
F
35 Hz, CHPhꢀP), 129.1 (s, 2C, CHArꢀBODIPY), 130.8 (s, 2C, CPyrrole),
0
1
131.0 (s, 2C, CHPhꢀP), 133.5 (d, 2C, JCꢀP = 52.2 Hz, CHPhꢀP),
133.5 (s, 2C, CPyrrole), 134.9 (d, 4C, 2JCꢀP = 9.9 Hz, CHPhꢀP), 134.9
(s, 1C, CArꢀBODIPY), 135.2 (d, 2C, 2JCꢀP = 9.1 Hz, CHArꢀP), 135.8 (s,
90 Equation 1
3
1C, CArꢀP), 137.8 (d, 1C, JCꢀP = 49.9 Hz, CArꢀP), 138.9 (s, 2C,
ΦF
and ΦFR are fluorescence quantum yields of the compound and
40 CPyrrole), 139.3 (s, 1C, CBODIPY), 139.6 (s, 1C, CArꢀBODIPY), 154.4
(s, 2C, CPyrrole), 167.7 (s, 1C, CO), 168.3 (s, 1C, CO). 31P{1H}
NMR (242.9 MHz, CD2Cl2): δ (ppm) ꢀ 12.4. HRꢀMS (ESI): m/z
calcd for C55H60BCl2F2N4O2POs + Na+: 1173.33991 [M+Na]+.
Found: 1173.33826. UVꢀVis (DMSO): λmax (nm) (ε, Mꢀ1 cmꢀ1)
45 378 (7900), 492 (sh, 20100), 526 (66900). IR: (cmꢀ1) 1653
(νC=O), 3420 (νNꢀH).
the reference respectively. A( E) and AR( E) are the absorbance at the
λ
λ
excitation wavelength, and n is the refractive index of the medium. IF and
95 IFR are fluorescent intensities of the compound and the reference
respectively. The reference system used was rhodamine 6G (φ = 0.78 in
H2O, λex = 488 nm).23 The data are shown in Table 2.
Table 2 Spectroscopic data of Bodꢀacid (BODIPYꢀacid derivative), 3 and
100 the different complexes 4, 5, 6 at 298 K
Synthesis
of
chloro[(4,4ꢀdifluoroꢀ8ꢀ(4ꢀ((2ꢀ(4ꢀ(diphenylꢀ
phosphino)benzamido)ethyl)carbamoyl)phenyl)ꢀ1,3,5,7ꢀtetraꢀ
methylꢀ2,6ꢀdiethylꢀ4ꢀboraꢀ3a,4aꢀdiazaꢀsꢀindacene)gold(I)] (6).
Stokes’
ε
/M-1
shift / cm-
λ
abs/nm
λ
em/nm
Φ
F(%)a,b
Compound Solvent
.cm-1
1
50 The reaction was carried out under Ar atmosphere. BODIPYꢀ
phosphine derivative 3 (56 mg, 0.07 mmol) and [Au(tht)Cl] (tht =
tetrahydrothiophene) (23 mg, 0.07 mmol) were dissolved in
degassed benzene (7 mL). The resulting mixture was stirred at
room temperature for 4 h. The reaction was monitored by 31P
55 NMR (242.9 MHz, 300 K). The solvent was removed under
reduced pressure. The gold complex 6 was isolated as a red
powder (63 mg, 89%).
Bodꢀacid
Bodꢀacid
CH3CN
DMSO
DMSO
DMSO
DMSO
DMSO
523
523
526
526
526
526
72900
83400
55300
70500
66900
59200
536
536
539
539
539
539
464
464
458
458
458
458
66
100
95
57
52
3
4
5
6
85
a
λ
exc = 488 nm ; b Using Rhodamine 6G as reference, ΦF = 0.78 in water,
λ
exc = 488 nm. All ΦF are corrected from refractive index.
6
|
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Antiproliferative assays
60
65
1
a) B. Rosenberg, L. VanCamp, T. Krigas, Nature, 1965, 205, 698–
699; b) B. Rosenberg, L. VanCamp, J. E. Trosko, V. H. Mansour,
Nature, 1969, 222, 385–386.
C. X. Zhang, S. J. Lippard, Curr. Opin. Chem. Biol., 2003, 7, 481ꢀ
489.
Human A27807S and A2780cisR cells were obtained from the
European Centre of Cell Cultures (ECACC, Salisbury, UK). All
cell culture reagents were obtained from GibcoꢀBRL (Basel,
Switzerland). Cells were grown in RPMI 1640 medium
containing 10% fetal calf serum (FCS) and antibiotics. Cisplatin
was purchased at SigmaꢀAldrich, and auranofin at Vinci
Biochem. Stock solutions of the complexes (in DMSO) were
diluted in complete medium to the required concentration. DMSO
2
3
5
a) A. Bergamo, C. Gaiddon, J. H. M. Schellens, J. H. Beijnen,
G. Sava, J. Inorg. Biochem., 2012, 106, 90ꢀ99; b) A. Casini, C. G.
Hartinger, A. A. Nazarov, P. J. Dyson, Top. Organomet. Chem.,
2010, 32, 57ꢀ80; c) M. A. Jakupec, M. Galanski, V. B. Arion, C. G.
Hartinger, B. K. Keppler, Dalton Trans., 2008, 183ꢀ194.
a) Numerous studies in “theranostics field” are dealing with
nanoparticles,[4b] but there are only few papers describing the
labeling of single metalꢀatom complexes;[4cꢀ4e] b) Special issue
“theranostic nanomedicine”: Acc. Chem. Res., 2011, 44, 841ꢀ1134;
c) F. Schmitt, P. Govindaswamy, O. Zava, G. SüssꢀFink, L.
JuilleratꢀJeanneret, B. Therrien, J. Biol. Inorg. Chem., 2009, 14,
101ꢀ109; d) Y. Q. Tan, P. J. Dyson, W. H. Ang, Organometallics,
2011, 30, 5965ꢀ5971; e) M. A. Furrer, F. Schmitt, M. Wiederkehr,
L. JuilleratꢀJeanneret, B. Therrien, Dalton Trans., 2012, 41, 7201;
f) S. Wu, C. Zhu, C. Zhang, Z. Yu, W. He, Y. He, Y. Li, J. Wang,
Z. Guo, Inorg. Chem., 2011, 50, 11847ꢀ11849.
10 at comparable concentrations did not show any effects on cell
cytotoxicity.
70
4
For cytotoxicity screening cells were grown in 96ꢀwell cell
culture plates (Corning, NY) at a density of 25x103 cells per well.
The culture medium was replaced with fresh medium containing
15 the complexes at concentrations varying from 20 to 200 ꢁM, with
an exposure time of 72 h. Thereafter, the medium was replaced
by fresh medium and cell survival was measured using the MTT
((3ꢀ(4,5ꢀDimethylthiazolꢀ2ꢀyl)ꢀ2,5ꢀdiphenyltetrazolium bromide)
test as previously described. Briefly, 3ꢀ(4,5ꢀdimethylꢀ2ꢀthiazoyl)ꢀ
20 2,5ꢀdiphenyltetrazolium bromide (MTT, Merck) was added at
250 ꢁg/mL and incubation was continued for 2 h. Then the cell
culture supernatants were removed, the cell layer was dissolved
in DMSO, and absorbance at 540 nm was measured in a 96ꢀwell
multiwellꢀplate reader (iEMS Reader MF, Labsystems,
25 Bioconcept, Switzerland) and compared to the values of control
cells incubated in the absence of complexes. The experiments
were conducted in quadruplicate wells and repeated at least twice.
75
80
5
6
M. J. Clarke, Coord. Chem. Rev., 2002, 232, 69ꢀ93.
S. Nobili, E. Mini, I. Landini, C. Gabbiani, A. Casini, L. Messori,
Med. Res. Rev., 2010, 30, 550ꢀ580.
7
a) A. Casini, L. Messori, Curr. Top Med. Chem., 2011, 11, 2647ꢀ
2660 ; b) A. Casini, A. Guerri, C. Gabbiani, L. Messori, J. Inorg.
Biochem. 2008, 102, 995ꢀ1006; c) W.H. Ang, A. Casini, G. Sava,
P.J. Dyson, J. Organomet. Chem., 2011, 696, 989ꢀ998.
a) A. A. Nazarov, J. Risse, W. H. Ang, F. Schmitt, O. Zava,
A. Ruggi, M. Groessl, R. Scopelitti, L. JuilleratꢀJeanneret,
C. G. Hartinger, P. J. Dyson Inorg. Chem., 2012, 51, 3633–3639.
b) Q. Zhou, W. Lei, Y. Chen, C. Li, Y. Hou, B. Zhang, X. Wang,
Chem. Eur. J., 2012, 18, 8617ꢀ8621. c) F. Zobi, B. B. Mood, P. A.
Wood, F. P. A. Fabbiani, S. Parsons, P. J. Sadler, Eur. J. Inorg.
Chem., 2007, 2783–2796.
85
8
90
Determination of CellꢀAssociated Drug Fluorescence by
30 Fluorescence Microscopy
A2780S Cells were grown for 24 h on chambered coverglass
(LabꢀTek, NUNC) slides in complete medium at a density of
1×104 and later exposed to compounds 3ꢀ6 (10 and 20 ꢁM) at
37 °C in the dark. Excess complex was washed away with PBS
35 after 1 and 2 h incubation, and before observation. The
measurement chamber was placed on the stage of an inverted
confocal fluorescence microscope (LS M700, Zeiss) equipped
with a 40× oil immersion objective (Zeiss). The laser line for
excitation of BODIPY compounds were 488 nm and 515 nm,
40 respectively. Fluorescence signal intensities were evaluated by
using Zeiss LSM software. Colocalization map was obtained with
ImageJ software, using its colocalization plugin.
95
9
L. H. Davies, B. Stewart, R. W. Harrington, W. Clegg, L.J.
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Notes and references
a
Institut de Chimie Moléculaire de l’Université de Bourgogne, UMR
45 6302 CNRS, Université de Bourgogne, 9 avenue A. Savary, BP47870,
21078 Dijon, France, Fax: +33 380396117; Tel:+33 380393773; E-mail:
ewen.bodio@u-bourgogne.fr
13
115
b
Institut des Sciences et Ingénierie Chimiques, École Polytechnique
14
Fédérale de Lausanne (EPFL), CH-1015 Lausanne, Switzerland
c
50 Département de Chimie, Université de Sherbrooke, Sherbrooke, JIK
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2R1, Québec, Canada
d
Research Institute of Pharmacy, University of Groningen, Antonius
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† Electronic Supplementary Information (ESI) available: synthetic
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An unprecedented family of Au, Ru and Os theranostics was synthesized starting from a new
BODIPY-phosphane tool. They were tested in vitro for cytotoxicity and imaging.