Enhanced kinetic stability of [Pd2L4]4+ cages through ligand substitution

Article abstract of DOI:10.1039/c6dt00133e

There is considerable interest in exploiting metallosupramolecular cages as drug delivery vectors. Recently, we developed a [Pd₂L₄]⁴⁺ cage capable of binding two molecules of cisplatin. Unfortunately, this first generation cage was rapidly decomposed by common biologically relevant nucleophiles. In an effort to improve the kinetic stability of these cage architectures here we report the synthesis of two amino substituted tripyridyl 2,6-bis(pyridin-3-ylethynyl)pyridine (tripy) ligands (with amino groups either in the 2-(2A-tripy) or 3-(3A-tripy) positions of the terminal pyridines) and their respective [Pd₂(L_tripy)₄]⁴⁺ cages. These systems have been characterised by ¹H, ¹³C and DOSY NMR spectroscopies, high resolution electrospray mass spectrometry, elemental analysis and, in one case, by X-ray crystallography. It was established, using model palladium(ii) N-heterocyclic carbene (NHC) probe complexes, that the amino substituted compounds were stronger donor ligands than the parent system (2A-tripy > 3A-tripy > tripy). Competition experiments with a range of nucleophiles showed that these substitutions lead to more kinetically robust cage architectures, with [Pd₂(2A-tripy)₄]⁴⁺ proving the most stable. Biological testing on the three ligands and cages against A549 and MDA-MB-231 cell lines showed that only [Pd₂(2A-tripy)₄]⁴⁺ exhibited any appreciable cytotoxicity, with a modest IC₅₀ of 36.4 ± 1.9 μM against the MDA-MB-231 cell line. Unfortunately, the increase in kinetic stability of the [Pd₂(L_tripy)₄]⁴⁺ cages was accompanied by loss of cisplatin-binding ability. FOR VERIFICATION The Royal Society of Chemistry.

Full text of DOI:10.1039/c6dt00133e

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PAPER
4
+
Enhanced kinetic stability of [Pd L ] cages
2
4
through ligand substitution†
Cite this: DOI: 10.1039/c6dt00133e
a
b
a
b
Dan Preston, Samantha M. McNeill, James E. M. Lewis, Gregory. I. Giles and
a
James D. Crowley*
There is considerable interest in exploiting metallosupramolecular cages as drug delivery vectors.
4
+
Recently, we developed a [Pd
2
L
4
]
cage capable of binding two molecules of cisplatin. Unfortunately,
this first generation cage was rapidly decomposed by common biologically relevant nucleophiles. In an
effort to improve the kinetic stability of these cage architectures here we report the synthesis of two
amino substituted tripyridyl 2,6-bis(pyridin-3-ylethynyl)pyridine (tripy) ligands (with amino groups either
4
+
2 4
in the 2-(2A-tripy) or 3-(3A-tripy) positions of the terminal pyridines) and their respective [Pd (Ltripy) ]
1
13
cages. These systems have been characterised by H, C and DOSY NMR spectroscopies, high resolution
electrospray mass spectrometry, elemental analysis and, in one case, by X-ray crystallography. It was
established, using model palladium(II) N-heterocyclic carbene (NHC) probe complexes, that the amino
substituted compounds were stronger donor ligands than the parent system (2A-tripy > 3A-tripy > tripy).
Competition experiments with a range of nucleophiles showed that these substitutions lead to more
4
+
kinetically robust cage architectures, with [Pd
2
(2A-tripy)
4
]
proving the most stable. Biological testing on
4
+
Received 11th January 2016,
Accepted 5th April 2016
the three ligands and cages against A549 and MDA-MB-231 cell lines showed that only [Pd (2A-tripy) ]
2
4
exhibited any appreciable cytotoxicity, with a modest IC50 of 36.4 ± 1.9 µM against the MDA-MB-231 cell
4+
DOI: 10.1039/c6dt00133e
line. Unfortunately, the increase in kinetic stability of the [Pd
cisplatin-binding ability.
2
(Ltripy
)
4
]
cages was accompanied by loss of
www.rsc.org/dalton
1
1d
capable of binding cisplatin (cis-[Pt(NH
3
)
2
Cl
2
]). We hoped to
Introduction
4
+
exploit these [Pd L ] cages as metallosupramolecular drug
2
4
Interest in self-assembled coordination complexes of well- delivery vectors, unfortunately the cage architecture was
−
11a,d,12
defined two- and three-dimensional geometries, or metallo- rapidly decomposed when exposed to chloride (Cl ),
his-
(common nucleophiles in
potential in a range of applications. The molecular recognition biological systems). In order to use these cisplatin binding
1
11a
11a
supramolecular architectures, continues to grow due to their tidine (his)
and cysteine (cys)
4
+
properties of these systems have been used to develop mole- [Pd
2
L
4
]
cages as drug delivery agents a method for increasing
2
3
4
5
7
cular reactions flasks, catalysts and drug delivery agents.
Systems have also been used to sequester reactive species,
the kinetic stability of these metallosupramolecular architec-
tures against nucleophiles was required. There are two
obvious approaches to enhance the kinetic stability of the cage
13
6
and environmental pollutants. Additionally, the biological,
8
9
10
photophysical, electronic and redox properties of these architectures: (1) assemble the cage using more kinetically
metallosupramolecular architectures have been studied. inert metal ions or (2) sterically and electronically tune the
1
1
As part of our interest in the biological properties of tripy ligand framework. Exploiting kinetically inert metals for
metallosupramolecular architectures we have previously the generation of metallosupramolecular architectures can be
4
+
reported the synthesis of a tripyridyl (tripy) [Pd L ] cage difficult due to the formation of kinetically “trapped” inter-
2
4
mediate structures that cannot “error-correct” into the desired
system. Therefore we chose to generate more electron-donating
tripyridyl ligands in an effort to obtain more kinetically robust
a
Department of Chemistry, University of Otago, PO Box 56, Dunedin, New Zealand.
4
+
[Pd
2
L
4
]
cages. Herein we report the synthesis of two new
E-mail: jcrowley@chemistry.otago.ac.nz
b
Department of Pharmacology and Toxicology, University of Otago, PO Box 56,
amino substituted tripyridyl ligands (2A-tripy and 3A-tripy)
and their respective cages (Scheme 1). The addition of the
amino groups to the 2- and 3-positions of the terminal ligating
pyridyl units of the ligand framework was expected to increase
the donor properties of the ligands and lead to more kineti-
Dunedin, New Zealand
†
Electronic supplementary information (ESI) available: The experimental pro-
1
13
cedures, H, C and DOSY NMR, HR-ESMS, crystallographic and computational
data. CCDC 1439952. For ESI and crystallographic data in CIF or other electronic
format see DOI: 10.1039/c6dt00133e
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1
1d,12,14
dard methods
(ESI†) and characterised using NMR
spectroscopy, high resolution electrospray mass spectrometry
HR-ESMS), and elemental analysis (Experimental section and
ESI†).
Addition of one of the ligands (tripy, 2A-tripy, and 3A-tripy)
to a solution of [Pd(CH CN) ](BF in a 2 : 1 ratio resulted in
(
3
4
4 2
)
the formation of the desired cages (Scheme 1). While the tripy
and 3A-tripy containing cages formed instantaneously at room
temperature, the formation of [Pd (2A-tripy) ](BF ) required
2 4 4 4
heating in DMSO solution at 50 °C for six hours to complete
1
the assembly. The cages were characterised using H and
DOSY NMR spectroscopy, HR-ESMS, and elemental analysis,
1
(
Experimental and ESI†). The H NMR spectra (Fig. 1 and
4
+
ESI†) of the [Pd (L
) ] cages show a single set of peaks.
2
tripy 4
The proton resonances due to the terminal pyridyl units of the
cages (Hc–f) are shifted downfield (Δδ = 0.19–0.59 ppm) relative
to the free tripy ligands, consistent with complexation to palla-
dium(II) ions. Interestingly, the proton resonance of the
2
-amino group in the [Pd
large (Δδ(HNH) = 1.56 ppm) shift relative to the free ligand
Fig. 1). Presumably, this large shift is caused by a combi-
2 4 4 4
(2A-tripy) ](BF ) cage displays a very
(
nation of coordination to the palladium(II) ions and intra-
ligand hydrogen bonding interactions between the amino
groups on the adjacent ligands. Consistent with this postulate,
the shift of the corresponding proton resonance in the
[
Pd (3A-tripy) ](BF ) cage is much smaller (Δδ(HNH) =
2
4
4 4
0
.81 ppm, ESI†). The sharp, uncomplicated signals observed
1
4+
in the H NMR spectra of the [Pd
2
(Ltripy
)
4
]
systems are
similar to what was previously observed for the formation of
4
+
other [Pd
2
(L)
4
]
cages and are consistent with the formation
1
1a,d,12
4
+
of complexes of high symmetry in solution.
Scheme 1 Synthesis of the [Pd
2
(Ltripy
)
4
]
cages from the tripyridyl
CN) ](BF
1
ligands (tripy, 2A-tripy, 3A-tripy). Conditions: (i) [Pd(CH
solvent (CH CN, DMF or DMSO), RT or 50 °C.
3
4
4
)
2
,
Diffusion-ordered H NMR spectroscopy (DOSY) provided
3
additional strong support for the selective formation of the
1
cages in solution. H DOSY spectra (d
6
-DMSO, 298 K) were
obtained for ligands (tripy, 2A-tripy, and 3A-tripy) and cages
4
+
4+
4+
(
[Pd
2
(tripy)
4
] , [Pd
2
(2A-tripy)
4
]
and [Pd
2
(3A-tripy)
4
]
(ESI†)).
cally robust cage architectures. Additionally, the presence of
4
+
Each of the proton signals in the individual spectra show the
same diffusion coefficient (D), indicating that there is only one
species present in solution (ESI†). The Dcomplex/Dligand ratios of
the 2-amino units within the [Pd
2
(2A-tripy)
4
]
cage architec-
ture was expected to sterically reduce access to the palladium(II)
ions potentially further enhancing the cage lifetime in the
∼0.50 : 1 are similar to those observed for related literature
presence of biological nucleophiles. Competition experiments
−
with Cl , his and cys nucleophiles showed that the [Pd
2
(2A-
4
+
4+
tripy)4] cage was more stable than the [Pd (3A-tripy) ] cage
2
4
4
+
which in turn was more stable than the [Pd
2
(tripy)
4
]
cage.
Unfortunately, the structural and electronic changes imposed
through amino substitution in the 2-position circumscribe the
ability of the cage to encapsulate cisplatin. Preliminary biologi-
2
cal testing on these systems indicated that only [Pd (2A-
4
+
tripy)4] exhibited an IC50 < 50 µM against either of the two
investigated cell lines.
Results and discussion
Synthesis and characterisation
The electronically and sterically tuned tripyridyl ligands (tripy,
1
Fig. 1 Partial H NMR spectra (298 K, 500 MHz, d
6
-DMSO) of (a) 2A-
2
2 4 4 4
A-tripy, and 3A-tripy, Scheme 1) were synthesised using stan- tripy and (b) the cage [Pd (2A-tripy) ](BF ) .
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1
1a,d,12
15
compounds
and suggest that the palladium(II) cage gen as expected. Additionally, the 2-amino units of the
species are stable in solution (ESI†).
ligands are engaged in intra-ligand hydrogen bonding inter-
4
+
Mass spectra (HR-ESMS) of the [Pd
2
(Ltripy
)
4
]
cage systems actions (N⋯N 3.78(3) Å, N–H⋯N 2.94 Å, Fig. 2b and ESI†) con-
1
in DMF–CH
3
CN solution obtained under pseudo cold-spray sistent with H NMR data described above. In contrast to the
4
+
4+
16
4+
conditions displayed overlapping peaks due to [Pd L ] and [Pd (tripy) ] cage, the [Pd (2A-tripy) ] cation adopts a
2
4
2
4
2
4
2
+
[
PdL
cage displayed a peak due to the [Pd
After considerable effort (>50 crystallisations and data col- to be caused by hydrogen bonding interactions between the
lections) the solid state structure of the [Pd (2A-tripy) ](BF amino groups on the exo faces of the architecture. This is
complex was obtained using X-ray crystallography (Fig. 2 and quite different to what has been previously observed in the
2
]
ions. Additionally, the spectrum of the 3A-tripy based more twisted structure in which the ligands of the 2A-tripy
2
+
2
(3A-tripy)
4
(BF
4
)
2
]
ion.
cages are significantly bent out of planarity and this appears
2
4
4 4
)
4+
ESI†). Small weakly diffracting X-ray quality crystals were gen- solid state structures of unsubstituted [Pd (tripy) ] cations,
2
4
erated by vapour diffusion of diethyl ether into a CH
tion of the [Pd (2A-tripy) ](BF cage. Although the weak adopt a lantern shape, with essentially planar tripy ligands.
diffraction was, at least in part, due to the presence of multiple
3
CN solu- these cages without the 2-amino groups all were found to
1
1a,
2
4
4 4
)
d,12
The coordinated 2A-tripy ligand distorts in two ways: a swi-
disordered solvent molecules and counter anions within the velling of the coordinating pyridine rings relative to the princi-
crystal lattice (vide infra), the cationic framework of the cage pal rotation axis of the molecule (θ = 34.44°–34.64°, compared
4
+
was readily identified (Fig. 2).
with θ = 3.47°–9.42° for [Pd (tripy) ] ) and a twisting of the
2 4
4
+
The solid state structure of the [Pd
2
(2A-tripy)
4
]
cage con- central pyridine out of the plane through which the ligand
firms that the coordination of the 2-amino pyridyl units to the coordinates to the two Pd(II) centres (φ = 35.10° compared with
4
+
palladium(II) ions was monodentate through the pyridyl nitro- φ = 5.61° for [Pd (tripy) ] ). The cavity dimensions also differ
2
4
(
1
a Pd⋯Pd distance of 11.530(9)–11.610(9) Å compared with
1.201(1) Å for [Pd (tripy) ] , and a core-to-core pyridyl N⋯N
2 4
4
+
distance of 10.711(9)–10.732(9) Å compared with 11.07(1)–
4
+
1
1.26(1) Å for [Pd
2 4
(tripy) ] , Table 1).
4
+
Interestingly, the central cavity of the [Pd
2
(2A-tripy)
4
]
cage
is filled in the solid state. The exo-methylene alcohol (CH OH)
2
substituents from the four neighbouring cages in the crystal
lattice penetrate into the cavity of each cage and form a hydro-
gen bonding interaction (O⋯N 2.74(3) Å, O–H⋯N 1.91 Å,
Fig. 3 and ESI†) with the endo-pyridyl unit. These interactions
generate 2D supramolecular sheets of cages through the solid
state structure (Fig. 3 and ESI†).
Competition experiments, cisplatin binding and cytotoxicity
studies
The relative pK
a
values of 2-aminopyridine (6.82), 3-aminopyri-
dine (6.04), and pyridine (5.23) indicate that 2-aminopyridine
1
7
is the most basic ligand. Evidence that 2-aminopyridine was
also the strongest nucleophile was obtained using the palla-
dium(II)-N-heterocyclic carbene (NHC) probe system developed
1
8
by Huynh and coworkers (Table 2 and ESI†). Consistent with
the pK values, the probe complexes indicated that 2-amino-
a
pyridine (161.2 ppm) is a stronger donor than 3-aminopyri-
dine (159.8 ppm) which is a stronger donor than pyridine
(
159.3 ppm). The chemical shift observed for the 2-aminopyri-
dine ligand is very similar to that previously reported for
1
8b
N-methylimidazole (161.1 ppm)
strength of these ligands are similar. To allow direct compari-
son to the literature pK values (Table 2) and for synthetic con-
we have examined the probe complexes of the
cations determined simpler pyridine rather than the tripyridyl ligands. However,
suggesting that the donor
a
1
1a
venience
4
+
Fig. 2 Molecular structure of [Pd
using X-ray crystallography: (a)
2
(2A-tripy)
4
]
a
ball-and-stick representation of
these pyridine model systems can serve as proxies for their
respective tripyridyl ligands (2A-tripy, 3A-tripy, tripy) and
provide indirect experimental evidence for the donor pro-
perties of the tripy ligands because the steric and electronic
changes on going from the pyridine to tripy ligands are the
4+
4+
[
Pd
2
(2A-tripy)
4
]
2 4
cation; (b) a partial view of the [Pd (2A-tripy) ]
cation from the top showing the hydrogen bonding interactions
between the 2-amino substituents of the ligands. Solvent molecules and
counterions omitted for clarity. Colours: magenta = palladium, grey =
carbon, white = hydrogen, blue = nitrogen, red = oxygen.
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13
Table 2 pK
a
values, C NMR chemical shifts of NHC carbene of the
model probe complexes, and half-lives (t1/2) for the decomposition of
4+
the [Pd
nucleophiles (3 : 2
through time-course H NMR spectroscopy
2
(Ltripy
)
4
]
architectures against selected biologically relevant
d
6
-DMSO/D O, 298 K, 500 MHz) as measured
2
1
1
3
a
Cl−
(8 eq.)
C δ
(ppm)
his
(4 eq.)
cys
(4 eq.)
Compound
pK
a
1
1
1
7
7
7
a,18
[Pd
[Pd
[Pd
2
2
2
(tripy)
(2A-tripy)
(3A-tripy)
4
](BF
4
)
4
5.23
6.82
6.04
159.3
161.2
159.8
>1 min 18 min 6 min
2 h 46 h 3 h
10 min 25 min 10 min
a
a
4
](BF
](BF
4
)
)
4
4
4
4
a
Chemical shift (ppm) of the NHC carbene carbon in the model Pd(II)
probe complexes (ESI).
4
+
The kinetic stability of the [Pd (L
) ] architectures in
2
tripy 4
−
the presence of common biological nucleophiles (Cl , his and
cys) was determined using H NMR competition experiments
Table 2 and ESI†). Time-course H NMR competition experi-
1
1
(
6 2
ments were carried out in 3 : 2 d -DMSO/D O where 3 mM
solutions of each cage were treated with 8 equivalents of tetra-
methylammonium chloride or 4 equivalents of his or cys.
4
+
Under these conditions the unsubstituted [Pd
2
(tripy)
4
]
cage
was rapidly decomposed by all the nucleophiles. The half-life
4
+
for the decomposition of the [Pd (tripy) ] complex with his
2
4
4
+
was 18 minutes. Despite the [Pd
2
(3A-tripy)
4
]
architecture con-
taining the slightly more electron rich 3A-tripy ligand the cage
was still quickly decomposed by each of the nucleophiles.
However, the t1/2 were subtly increased against all the nucleo-
philes (for his t1/2 = 25 min) suggesting that the enhanced
ligand donor properties of the 3A-tripy ligand does lead to
increased cage stability relative to the unsubstituted system.
4
+
4+
Fig. 3 The crystal packing of [Pd
2
(2A-tripy)
4
]
cations showing (a) the
The [Pd
2
(2A-tripy)
4
]
cage displayed markedly higher stabi-
interdigitation of the methylene alcohol substituents on adjacent lity against all the nucleophiles studied. The half-lives for the
4+
4+
[
Pd
2
(2A-tripy)
4
]
2 4
cations into the central cavity of the [Pd (2A-tripy) ]
2A-tripy cage decomposition against each nucleophile were all
cation (cage shown in ball-and-stick, methylene alcohol substituents in
over 2 h, whereas the corresponding t1/2 for the other cages
were all less than 30 min. 2A-tripy is only a modestly stronger
donor ligand than the 3A-tripy, and thus the observed large
difference in stability is presumably not predominantly due to
the increase donor ability of the ligand. A more important
spacefilling view), and (b) the extended lattice.
2
Table 1 Selected crystallographic distances and angles from [Pd (2A-
4+
4+ 16
tripy)
4
]
2 4
and [Pd (tripy) ]
element is likely to be the presence of the 2-amino groups on
4
+
4+ 16
4+
[
Pd
2
(2A-tripy)
4
]
2 4
[Pd (tripy) ]
the exo-faces of the [Pd
2
(2A-tripy)
4
]
cage which sterically
a
protect the palladium(II) ions from the incoming nucleophiles.
Additionally, as observed in the X-ray structure (Fig. 3), the
hydrogen bonding interactions between the eight amino
Θ (°)
Φ (°)
34.44°–34.64°
35.10°
11.53(9)–11.610(9) Å
10.71(9)–10.73(9) Å
3.47°–9.42°
5.61°
11.201(1) Å
b
d(Pd(II)⋯Pd(II)) (Å)
d(Ncore⋯Ncore) (Å)
11.07(1)–11.26(1) Å
groups of the four 2A-tripy ligands may further enhance the
a
4+
θ: the swivelling of the coordinating pyridine rings relative to the stability of the [Pd
2
(2A-tripy)
4
]
cage relative to the other tripy
b
−
principal rotation axis of the molecule. φ: twisting of the central
pyridine out of the plane through which the ligand coordinates to the
two Pd(II) centres.
architectures. However, against the stronger nucleophiles (Cl
and cys) the half-lives for the decomposition of the [Pd (2A-
2
4
+
tripy)
4
]
are less than 3 h suggesting that these systems would
need further tuning in order to be useful in a biological
setting.
same across the series. Thus the model complexes provide a
qualitative ranking of the substituents’ effects present in the
tripy ligands and strongly suggest that the donor properties of We
Cisplatin binding
1
1d,12,19
16
and others have previously shown that other
4
+
the tripyridyl ligands follow the order 2A-tripy > 3A-tripy > similar [Pd (tripy) ] cages can encapsulate cisplatin through
2
4
2
0
tripy.
hydrogen bonding interactions in CH CN and DMF solvents.
3
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1
H NMR spectroscopy and recently reported crystallographic CH
3
CN or DMF solvents displayed no shifts relative to the free
1
6b
evidence confirms that despite the presence of the potential cage indicating that the 2-amino substituted cage is not able
hydrogen bond donor CH OH units of the exo surface of the to bind cisplatin. We have previously shown that cisplatin
2
4
+
12
[Pd
2
(tripy)
4
]
cage the system retains the ability to bind cis- binding is very weak and that subtle changes to the size,
2
1
platin. Addition of an excess of cisplatin to a d -DMF solution steric profile and electronic properties of the cage cavity are
7
4
+
of [Pd
.24 ppm) of the internally directed cage proton H
and b) indicative of cisplatin binding within the cage cavity. two factors. Firstly, the presence of the eight amino groups on
2
(tripy)
4
]
resulted in a large downfield shift (Δδ = enough to completely turn off cisplatin binding. Presumably,
0
c
(Fig. 4a the lack of cisplatin binding in this system can be ascribed to
1
6b
4+
The solid state structure
of the [(cisplatin)
2
⊂Pd
2
(tripy)
4
]
the exo faces of the cage has caused a twisting of the architec-
host–guest adduct has recently been reported by Casini, Kuhn ture (as indicated in the crystal structure, Fig. 3, and discussed
and co-workers and is very similar to what we have previously and listed in Table 1). This twisting subtly alters both the size
1
1d,19b
1
observed in related systems.
evidence, the [Pd (tripy) ]
2 4
Consistent with the H NMR of the cisplatin binding cavity and the orientations of the
cation binds two molecules of cis- hydrogen-bond acceptors and donors groups within the cage
4
+
1
6
platin within the cavity of the cage. The guest molecules are cavity, weakening the interaction between the host and the cis-
rotated 180° with respect to each other; hydrogen bonds platin guest. Secondly, the electron donating 2-amino units
between the guests and cage (N–H⋯NPy and Cl⋯H–CPy) as push electron density back onto the terminal pyridyl rings of
well as a metal–metal interaction between the platinum atoms the tripy ligand. This would reduce the polarisation of the
1
1d,16b,19b
of the guests were observed (Fig. 3a).
acidic H
(3A- bonding interaction with chloride ligands of the cisplatin
tripy)4] indicated that the 3-amino substituted cage is also guest. These effects, in concert, appear to be enough to fully
c
protons of the pyridyl unit, weakening the hydrogen
1
A similar H NMR experiment with cisplatin and [Pd
2
4
+
22
able to bind cisplatin in solution, albeit more weakly (Δδ = circumscribe the already weak cisplatin–cage interaction.
.07 ppm for H , Fig. 4c and d) than the parent [Pd (tripy) ]
c 2 4
cage. Conversely, H NMR spectra of the [Pd (2A-tripy) ] cage
in the presence of cisplatin acquired in either deuterated
4
+
0
1
4+
2
4
Cytotoxicity
While the improvement of kinetic stability of the [Pd (2A-
2
4
+
tripy)4] cage has been achieved at the expense of host–guest
4
+
capacity, recent work has shown that [Pd
can act as biological agents in their own right.
2
(L)
4
]
architectures
1
1a,23
Accord-
ingly, a preliminary investigation of the cytotoxic properties
of the three cages (and the corresponding ligands) was under-
taken against two cell lines: A549 (lung cancer) and cisplatin-
resistant MDA-MB-231 (breast cancer) (Table 3). We have
previously determined the IC50 values for cisplatin against
2
4
these two cell lines, which were 9.4 ± 0.3 µM (A549) and
1
1b
41.2 ± 3.9 µM) (MDA-MB-231)
respectively. None of the
ligands exhibited significant cytotoxicity (IC50 > 200 µM)
against these cell-lines. Similarly, neither of the unstabilised
cages, [Pd (tripy) ](BF ) or [Pd (3A-tripy) ](BF ) (IC50
>
>
2
4
4 4
2
4
4 4
5
1
0 µM) nor the palladium(II) tetrafluoroborate salt (IC50
1
1a
00 µM)) [Pd(CH CN) ](BF ) displayed any appreciable cyto-
3
4
4 4
1
Fig. 4 Partial
Pd (tripy) ](BF
tripy) ](BF and (d) [(cisplatin)
H
NMR spectra (298 K, 500 MHz,
(b) [(cisplatin) ⊂Pd (tripy) ](BF and (c) [Pd
⊂Pd (3A-tripy) ](BF
d
7
2 4
-DMF) of (a) toxic effect. The most kinetically stable cage [Pd (2A-tripy) ]
[
2
4
4
)
4
,
2
2
4
4
)
4
2
(3A- (BF
IC50 > 50 µM), and slightly higher cytotoxicity against the
4 4
) displayed low cytotoxicity against the A549 cell line
4
4
)
4
2
2
4
4 4
) .
(
4
+
4+
Table 3 IC50 values for [Pd
2
(Ltripy
)
4
]
and related literature [Pd
IC50 (µM)
2
(L)
4
]
compounds against selected cell lines
A549 (lung)
MDA-MB231 (breast)
HL-60 (leukemia)
SKW-3 (leukemia)
Compound
Ligand
Cage
Ligand
Cage
Ligand
Cage
Ligand
Cage
[
[
[
[
[
Pd
Pd
Pd
Pd
Pd
2
2
2
2
2
(tripy)
(2A-tripy)
(3A-tripy)
4
](BF
4
)
4
>200
>200
>200
28.5 ± 2.6
—
>50
>50
>50
6.9 ± 0.9
—
>200
>200
>200
89.8 ± 10.7
—
>50
36.4 ± 1.9
>50
6.0 ± 0.6
—
41.2 ± 3.9
—
—
—
—
>100
—
—
—
—
—
1.9 ± 0.2
8.1 ± 1.4
—
—
—
—
>80
—
—
—
—
—
1.8 ± 0.2
9.3 ± 2.1
4
](BF
](BF
4
)
)
4
4
4
4
1
1a
(L
(L
a
)
4
](BF
](BF
4
)
4
(L
(L
a
= hextrz)
= dipy-dianthracenyl)
2
3
b
)
4
)
4 4
b
Cisplatin
—
9.4 ± 0.3
—
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2
6
MDA-MB-231 cell line (IC50 = 36.4 ± 1.9 µM). We have pre- Dibromo-4-(hydroxymethyl)pyridine,
5-iodo-2-aminopyridi-
6
a
viously shown that a more kinetically robust and hydrophobic ne,
the dimeric dibromobis(benzimidazolin-2-ylidene)di-
4
+
27
[Pd
2
(L
a
)
4
]
helicate (L
a
= 1,3-bis(1-hexyl-1H-1,2,3-triazol-4-yl) palladium(II) complex,
and trans-dibromo(1,3-diisopropyl-
benzene) was considerably more cytotoxic (IC50 = 6.9 ± 0.9 and benzimidazolin-2-ylidene)(pyridine)palladium(II) were syn-
.0 ± 0.6 µM) against both the A549 and MDA-MB-231 cell thesised according to literature procedures. Solvents were labo-
1
8b
6
1
1a
lines.
Additionally, Yoshizawa and co-workers have recently ratory reagent grade. Petroleum ether refers to the fraction of
4
+
reported that a hydrophobic [Pd
2
(L
b
)
4
]
b
cage (L = 3,3′-((4,5,6- petrol boiling in the range 40–60 °C, isopropyl alcohol (IPA),
tris(2-methoxyethoxy)-1,3-phenylene)bis(anthracene-10,9-diyl))- methanol (CH OH), dichloromethane (CH Cl ), ethylenediamine-
3
2
2
dipyridine) is highly cytotoxic (IC50 = 1.8 and 1.9 µM) against tetraacetate (EDTA), ethynyltrimethylsilane (TMS-acetylene),
23
HL-60 and SKW-3 cancer cell lines. Thus it is presumed that tetrahydrofuran (THF), dimethyl sulfoxide (DMSO), dimethyl-
1
13
low cytotoxicity of the systems described here is connected to the formamide (DMF). H and C NMR spectra were recorded on
combination of both high kinetic lability and the more hydro- either a 400 MHz Varian 400-MR or Varian 500 MHz AR
4+
philic nature of the [Pd
2
(Ltripy
)
4
] architectures and ligands.
spectrometer. Chemical shifts are reported in parts per
million and referenced to residual solvent peaks (CDCl :
3
1
13
1
13
H δ 7.26 ppm, C δ 77.16 ppm; CD
3
CN: H δ 1.94, C δ 1.32,
-DMSO: H δ 2.50 ppm; C δ 39.52 ppm). Coup-
1
13
Conclusions
1
18.26 ppm, d
6
ling constants (J) are reported in Hertz (Hz). Standard abbrevi-
ations indicating multiplicity were used as follows: m =
multiplet, q = quartet, t = triplet, dt = double triplet, d =
doublet, dd = double doublet, s = singlet, br = broad. Full
Two amino substituted tripyridyl 2,6-bis(pyridin-3-ylethynyl)
pyridine (tripy) ligands (with amino groups either in the 2-(2A-
tripy) or 3-(3A-tripy) positions of the terminal pyridines) and
4
+
their respective [Pd (L
) ] cages were synthesised. These
2
tripy 4
1
13
H and C NMR spectra, together with structural labelling are
1
13
systems have been characterised by H, C and DOSY NMR
spectroscopies, high resolution electrospray mass spectro-
metry, elemental analysis and, in one case, by X-ray crystallo-
graphy. It was established, using palladium(II) NHC carbene
probe model complexes, that the amino substituted com-
pounds were moderately stronger donor ligands than the
parent pyridyl system (2A-tripy > 3A-tripy > tripy). Competition
included in the ESI.† IR spectra were recorded on a Bruker
ALPHA FT-IR spectrometer with an attached ALPHA-P
measurement module. Microanalyses were performed at the
Campbell Microanalytical Laboratory at the University of
Otago. Electrospray mass spectra (ESMS) were collected on a
Bruker microTOF-Q spectrometer.
Synthesis of 5-iodopyridin-2-acetamide (1). A solution of
−
experiments with common biological nucleophiles (Cl , his
and cys) showed that the [Pd (2A-tripy) ] cage proved the
5
(
1
-iodopyridin-2-amine (2.5 g, 11.4 mmol) in triethylamine
10 mL) and CH Cl (50 mL) was degassed with N for
5 minutes. Acetic anhydride (10.7 mL, 110 mmol) was added
4
+
2
4
2
2
2
most kinetically stable, presumably due to a favourable combi-
nation of enhanced ligand donor strength, and probably more
importantly, intramolecular hydrogen bonding and steric
shielding. Preliminary biological investigations against two
cell lines (A549 and MDA-MB-231) found that all ligands and
cages had IC values >50 µM, with the exception of [Pd (2A-
and the mixture was stirred at room temperature for 18 hours.
After removal of solvents under vacuum, the crude mixture
was dissolved in 3 : 1 CHCl
saturated aqueous NaHCO3 solution (2 × 75 mL), and brine
75 mL). The organic layer was dried over MgSO , filtered and
3
/IPA (150 mL), and washed with
5
0
2
(
4
4
+
tripy)
.9 µM.
However, while the ligand tuning resulted in more robust
4
]
which against MDA-MB-231 has an IC50 of 36.4 ±
the solvent removed under vacuum. Column chromatography
on silica (1 : 19 acetone/CH Cl ) gave the product as a brown
1
2
2
1
solid (2.3 g, 8.8 mmol, 77%). H NMR (400 MHz, d
6
-DMSO,
), 8.07
4
+
[Pd
2
(Ltripy
)
4
]
architectures the half-lives of the systems
298 K) δ: 10.58 (1H, s, HNH), 8.50 (1H, d, J = 2.3 Hz, H
a
against the stronger nucleophiles were still modest (t1/2
=
(
H
1H, d, J = 8.8 Hz, H ), 7.94 (1H, d, J = 8.8 Hz, H ), 2.08 (3H, s,
b
c
2
–3 h). Furthermore, the subtle structural changes in the most
1
3
). C NMR (100 MHz, d
6
a
-DMSO, 298 K) δ: 169.4, 153.3 (C ),
4
+
d
stabilised cage, [Pd
2
(2A-tripy)
4
]
were found to completely
1
51.2, 145.8 (C
b
), 115.3 (C ), 85.7, 23.9 (C ). HR ESI-MS
c d
destroy the ability of the system to bind cisplatin. Thus, it
appears that in order to exploit these types of metallosupra-
molecular cage architectures as drug delivery vectors, systems
assembled from more kinetically inert metals ions such as
+
(
CHCl ) m/z = 284.95 [M + Na] (calc. For C H IN NaO, 284.95).
3
7
7
2
-
1
IR: ν (cm ) 3215, 3139, 3079, 3018, 1676, 1660, 1520, 1362,
298, 829, 799. Anal. calcd for: C IN O: C, 32.08; H, 2.69;
1
7
H
7
2
N, 10.69%. Found: C, 32.30; H, 2.55; N, 10.71%.
Synthesis of N-(5-((trimethylsilyl)ethynyl)pyridin-2-yl)aceta-
mide (2). A round bottom flask containing 1 (1.00 g,
4
h
4e–g
25
Pt(II) and Ru(II)
and Co(III) will be required. Efforts to
generate more robust systems, composed of kinetically inert
metal ions, capable of binding drug molecules are underway.
3.82 mmol), CuI (0.07 g, 0.38 mmol) and Pd(PPh ) Cl (0.19 g,
3 2 2
0
.27 mmol) was purged with N
2
. Triethylamine (20 mL) was
for
5 minutes. After adding TMS-acetylene (0.68 g, 0.93 mL,
.87 mmol) via syringe, the reaction was heated at reflux under
for 48 hours. The solvent was removed under vacuum, and
added via syringe and the solution was degassed with N
1
6
2
Experimental
General
Unless otherwise stated, all reagents were purchased from
N
2
commercial sources and used without further purification. 2,5- the resulting solid was taken up in 3 : 1 CHCl /IPA (40 mL).
3
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The solution was stirred with aqueous 0.1 M EDTA/NH
4
OH
Synthesis of 5-ethynylpyridin-3-amine (5). A solution of 4
solution (40 mL) for 1.5 hours. After washing with (0.43 g, 2.3 mmol) in methanol (30 mL) with Na CO (0.48 g,
2
3
aqueous 0.1 M EDTA/NH
organic layer was dried with Na
removed under vacuum. The solid was purified through tant solid was purified by column chromatography on silica
column chromatography on silica (1 : 19 acetone/CH Cl ), (1 : 8 acetone/CH Cl ) to give the product as a brown solid
giving the product as a brown solid (0.86 g, 3.72 mmol, 97%). (0.25 g, 2.1 mmol, 96%). H NMR (500 MHz, CDCl
4
OH (50 mL) and brine (100 mL), the 4.5 mmol) was stirred at room temperature for 1 h. After fil-
2
SO , filtered, and the solvent tration, the solvent was removed under vacuum, and the resul-
4
2
2
2
2
1
3
, 298 K)
1
H NMR (500 MHz, CDCl , 298 K) δ: 8.34 (1H, d, J = 1.6 Hz, δ: 8.13 (1H, d, J = 2.7 Hz, H ), 8.05 (1H, d, J = 2.7 Hz, H ), 7.06
3
c
a
H
d
), 8.17 (1H, d, J = 8.7 Hz, H
b
), 8.07 (1H, br, HNH), 7.77 (1H, (1H, dd, J = 2.7 Hz, 2.7 Hz, H
b
), 3.71 (2H, br, HNH), 3.14 (1H, s,
1
3
dd, J = 8.7 Hz, 2.1 Hz, H
c
), 2.21 (3H, s, H
a
), 0.25 (15H, s, H
e
).
d 3 c
H ). C NMR (125 MHz, CDCl , 298 K) δ: 143.0 (C ), 141.8,
1
3
−1
C NMR (125 MHz, CDCl , 298 K) δ: 168.5, 150.9 (C ), 150.3, 137.1 (C ), 124.0 (C ), 119.1, 80.7, 79.8 (C ). IR: ν (cm ) 3328,
3
d
a
b
d
1
41.4 (C
HR ESI-MS (CHCl
C H N NaOSi, 255.09). IR: ν (cm ) 3243, 2955, 2161, 1697, Found: C, 70.96; H, 5.14; N, 22.80%. Mass spectral analysis
c
), 116.0, 112.9 (C
b
), 101.4, 97.1, 24.8 (C
a
), −0.1 (C
e
). 3295, 3163, 3020, 2148, 1644, 1585, 1424, 1329, 1246. Anal.
+
3
) m/z = 255.09 [M + Na] (calc. for calcd for C H N ·0.1 (acetone): C, 70.74; H, 5.37; N, 22.60%.
7 6 2
−
1
1
2
16 2
1
579, 1526, 1380, 1303, 1246, 1030. Anal. calcd for was not successful for this compound.
C
12
H
16
N
2
OSi·0.5 acetone: C, 62.03; H, 7.14; N, 11.35%. Found:
Synthesis of tripy. A round bottom flask containing (2,6-
C, 61.94; H, 7.29; N, 11.35%.
dibromopyridin-4-yl)methanol (336 mg, 1.26 mmol), 3-ethynyl-
Synthesis of 5-ethynylpyridin-2-amine (3). A solution of 2 pyridine (325 mg, 3.15 mmol), CuI (24 mg, 0.12 mmol) and Pd
0.63 g, 2.70 mmol) and NaOH (1.08 g, 27.0 mmol) in metha- (PPh Cl (35 mg, 0.05 mmol) was purged with N , before
(
3
)
2
2
2
nol (30 mL) was heated at reflux for 1.5 hours before removal addition of triethylamine (25 mL) and dry THF (25 mL). The
of solvent under vacuum, with the resultant residue taken up reaction was stirred in the absence of light for 24 hours.
3 2 2 4
in 3 : 1 CHCl /IPA (100 mL) and washed with water (100 mL) CH Cl (50 mL) and aqueous 0.1 M EDTA/NH OH solution
and brine (100 mL). After drying with Na SO and filtration, (100 mL) were added and the mixture stirred at room tempera-
2
4
2 2
the solvent was removed under vacuum, and purification ture for 2 hours. After extraction into CH Cl (2 × 100 mL), the
through column chromatography on silica (1 : 8 acetone/ combined organic layers were washed with brine (100 mL),
CH Cl ) gave the product as a brown solid (0.22 g, 1.90 mmol, dried over MgSO , and then filtered. The solvent was removed
2
2
4
1
7
2
0%). H NMR (500 MHz, CDCl
.2 Hz, 0.8 Hz, H
3
2 2
, 298 K) δ: 8.22 (1H, dd, J = under vacuum. Column chromatography (2 : 3 CH Cl /acetone)
), 7.53 (1H, dd, J = 8.6 Hz, 2.3 Hz, H ), 6.46 gave the product as a colourless solid (280 mg, 0.90 mmol,
b
c
1
(
1H, dd, J = 8.6 Hz, 0.8 Hz, H ), 4.69 (2H, br, HNH), 3.06 (s, Hd). 71%). H NMR (500 MHz, CD CN, 298 K) δ: 8.81 (2H, dd, J =
C NMR (125 MHz, CDCl
), 108.7, 108.0 (C ), 81.2, 77.8 (C ). IR: ν (cm ) 3326, 3295, (2H, ddd, J = 7.9 Hz, 2.2 Hz, 1.7 Hz, H ), 7.58 (2H, t, J = 0.9 Hz,
a
3
1
3
3
, 298 K) δ: 157.6, 151.6 (C
c
), 141.0 2.1 Hz, 0.8 Hz, H
c
f
), 8.61 (2H, dd, J = 4.9 Hz, 1.7 Hz, H ), 7.98
−1
(C
b
a
d
d
3
C
163, 2148, 1645, 1585, 1424, 1329, 1246, 1156. Anal. calcd for H ), 7.41 (2H, ddd, J = 7.9 Hz, 4.9 Hz, 0.9 Hz, H ), 4.67 (2H, dt,
b
e
1
3
7
H
6
N
2
·0.1 acetone: C, 70.74; H, 5.37; N, 22.60%. Found: J = 5.9 Hz, 0.8 Hz, H
a
), 3.55 (1H, t, J = 5.9 Hz, HOH). C NMR
), 150.6 (C ), 143.8,
Synthesis of N-(5-((trimethylsilyl)ethynyl)pyridin-3-yl)acet- 139.9 (C ), 125.3 (C ), 124.4 (C ), 119.9, 91.8, 86.2, 62.4 (C ).
C, 70.44; H, 5.17; N, 22.56%.
(500 MHz, CD
3
CN, 298 K) δ: 154.1, 153.3 (C
c
f
d
b
e
a
+
amide (4). A glass tube containing diisopropylamine (7 mL) HR ESI-MS (acetone) m/z = 645.21 [2M + Na] (calc. for
degassed with N
+
2
was charged with 5-bromopyridin-3-amine
C
40
H
26
N
6
NaO
2
,
645.20), 334.10 [M
+
Na] (calc. for
−
1
(
(
8
500 mg, 2.89 mmol), CuI (55 mg, 0.29 mmol), Pd(dppf) Cl
C H N NaO, 334.10). IR: ν (cm ) 3204, 3032, 2923, 2213,
2
2
20 13 3
85 mg, 0.17 mmol) and TMS-acetylene (1.173 mL, 852 mg, 1685, 1596, 1547, 1475, 1415, 1189, 1076, 1023. Anal. calcd for
.67 mmol) against a positive N flow. The tube was sealed O·0.25 acetone: C, 76.48; H, 4.49; N, 12.90%. Found:
and stirred at 60 °C for 72 hours. 3 : 1 CHCl /IPA (100 mL) and C, 76.70; H, 4.49; N, 13.06%.
2
20 13 3
C H N
3
aqueous 0.1 M EDTA/NH
and the mixture stirred for one hour. The organic layer was amine (20 mL) and THF (20 mL) were degassed with N
washed with water (50 mL) and brine (50 mL), dried over addition of 3 (400 mg, 3.39 mmol), (2,6-dibromopyridin-4-yl)
Na SO , filtered, and the solvent was removed under vacuum. methanol (362 mg, 1.35 mmol), CuI (25 mg, 0.14 mmol), and
The resultant solid was purified by column chromatography Pd(PPh Cl (38 mg, 0.050 mmol) against a positive N flow.
1 : 8 acetone/CH Cl ) to give the product as a brown solid The solution was heated at 50 °C for 48 hours. After removal of
4
OH solution (100 mL) were added
Synthesis of 2A-tripy. In a round bottom flask, diisopropyl-
2
, before
2
4
3
)
2
2
2
(
(
2
2
1
512 mg, 2.71 mmol, 94%). H NMR (500 MHz, CDCl
3
, 298 K) the solvent under vacuum, the resultant solid was taken up in
), 7.04 3 : 1 CHCl /IPA (150 mL) and aqueous 0.1 M EDTA/NH OH
δ: 8.10 (1H, d, J = 1.5 Hz, H
c
), 8.01 (1H, d, J = 2.7 Hz, H
a
3
4
13
(
(
(
1H, dd, J = 2.7 Hz, 1.7 Hz, H ), 0.25 (9H, s, H ). C NMR solution (50 mL) and stirred for 40 minutes. The organic layer
125 MHz, CDCl
b
d
3
, 298 K) δ: 142.9 (C
), 120.3, 101.9, 97.6, 0.0 (C
91.10 [M + H] (calc. for C H N Si, 191.10). Anal. calcd for resultant solid on a silica column deactivated with 3 : 97 tri-
c
a 2 4
), 141.9, 136.8 (C ), 124.1 was washed with brine, dried with Na SO , filtered and then
). HR ESI-MS (MeOH) m/z = the solvent was removed under vacuum. Purification of the
C
b
d
+
1
1
0
15 2
C
10
H
14
N
2
OSi: C, 63.11; H, 7.41; N, 14.62%; found: C, 63.40; ethylamine/CH
2
Cl
2
(0.5/4.5/95 then 1/9/90 saturated aqueous
Cl ) gave the product as a brown
−
1
H, 7.48; N, 14.62%. IR: ν (cm ) 3327, 3295, 3162, 2147, 1585, NH
4
OH solution/MeOH/CH
2
2
1
1
424, 1329, 1245.
solid (364 mg, 1.07 mmol, 79%). H NMR (500 MHz, d -DMSO,
6
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2
8
98 K) δ: 8.18 (2H, dd J = 2.3 Hz, 0.5 Hz, H
c
), 7.57 (2H, dd, J = added and the solution shaken vigorously. After decanting the
.6 Hz, 2.3 Hz, H ), 7.42 (2H, s, H ), 6.55 (4H, s, HNH), 6.49 liquid portion, the precipitate was suspended in CH Cl
d
b
2
2
2
(2H, dd, J = 8.6 Hz, 0.6 Hz, H
e
), 5.51 (1H, t, J = 5.8 Hz, HOH), (5 mL) and isolated by filtration. After washing with CH
.54 (2H, d, J = 5.7 Hz, H ). C NMR (500 MHz, d -DMSO, (5 mL), the solid was dried under vacuum at 60 °C for 4 days
a 6
2
Cl
1
3
4
2
1
=
98 K) δ: 159.7, 153.0, 152.0 (C ), 142.9, 139.7 (C ), 122.6 (C ), to give the product as a red solid (32 mg, 0.017 mmol, 57%).
c
d
b
1
07.7 (C
e
), 104.9, 88.8, 88.0, 60.9 (C
a
). HR ESI-MS (CH
3
CN) m/z
H NMR (500 MHz, d
6
-DMSO, 298 K) δ: 8.77 (8H, d, J = 1.6 Hz,
),
+
683.26 [2M + H] (calc. for C40
H
31
N
10
O
2
, 683.26), 342.13
H
c
), 8.11 (8H, s, HNH), 7.84 (8H, dd, J = 8.9 Hz, 1.5 Hz, H
d
+
−1
[M + H] (calc. for C H N O, 342.14). IR: ν (cm ) 3307, 3145, 7.56 (8H, s, H ), 6.73 (8H, d, J = 8.9 Hz, H ), 4.53 (8H, s, H ).
2
0
16
5
b
e
a
1
3
2
1
920, 2201, 1670, 1634, 1593, 1543, 1505, 1393, 1171, 1140,
055, 1016. Anal. calcd for C20 O·CH Cl : C, 69.00; (C
60.6 (C ). ESI-MS (DMSO/CH CN) m/z = 394.60 [M − 4(BF )]
6
C NMR (500 MHz, d -DMSO, 298 K) δ: 159.8, 153.5, 149.4
H
15
N
5
2
2
c
), 142.7 (C
d
), 142.1, 125.3 (C
b
), 112.6 (C
e
), 107.7, 90.5, 84.6,
4
+
H, 4.38; N, 20.02%. Found: C, 69.12; H, 4.51; N, 20.33%.
Synthesis of 3A-tripy. In a round bottom flask, diisopropyl- (calc. for C80
amine (20 mL) and THF (20 mL) were degassed with N , before (calc. for C40
(365 mg, 3.09 mmol), (2,6-dibromopyridin-4-yl)methanol 2210, 1637, 1546, 1513, 1411, 1025. Anal. calcd for
275 mg, 1.03 mmol), CuI (20 mg, 0.10 mmol), and Pd Pd ·5H O·4DMSO: C, 45.50; H, 4.07;
PPh Cl (36 mg, 0.05 mmol) were added against a positive N, 12.03%. Found: C, 45.24; H, 3.78; N, 12.29%.
Synthesis of [Pd (3A-tripy) ](BF ) . A solution of 3A-tripy
a
3
4
2
+
H
60
N
20
O
4
Pd
2
, 394.58), 394.10 [Pd(2A-tripy)
2
]
−
1
2
H
30
N
10
O
2
Pd, 394.08). IR: ν (cm ) 337, 3194,
5
(
(
C
80
H
60
B
4
F
16
N
20
O
4
2
2
3
)
2
2
N flow. The solution was heated at 40 °C for 48 hours. After
2
2
4
4 4
removal of the solvent under vacuum, the resultant solid was (60 mg, 0.18 mmol) and [Pd(CH
3
CN)
4
](BF
4
)
4
(39 mg,
taken in 3 : 1 CHCl /IPA (150 mL) and aqueous 01. M EDTA/ 0.090 mmol) in d -DMSO (0.75 mL) was sonicated for five
3
6
NH OH solution (50 mL) and stirred for 40 minutes. The minutes. Addition of ethyl acetate (20 mL)) resulted in precipi-
4
organic layer was washed with brine (100 mL), dried with tation of the product. The precipitate was collected by fil-
Na
vacuum. Purification of the resultant solid in a silica column ether (15 mL) to give a brown solid (64 mg, 0.03 mmol, 75%).
deactivated with 3 : 97 triethylamine/CH Cl (0.5/4.5/95 then OR solution of 3A-tripy (60 mg, 0.18 mmol) and
/9/90 saturated aqueous NH OH solution/MeOH/CH Cl ) gave [Pd(CH CN) ](BF (39 mg, 0.090 mmol) in DMF (2 mL) was
the product as a brown solid (197 mg, 0.58 mmol, 56%). sonicated for five minutes. Addition of ethyl acetate (20 mL)
2 4
SO , filtered, and then the solvent was removed under tration, and washed with ethyl acetate (15 mL) and diethyl
2
2
A
1
4
2
2
3
4
4 4
)
1
H NMR (500 MHz, d
), 7.96 (2H, d, J = 1.6 Hz, H
.2 Hz, H ), 5.60 (5H, m, H & HOH), 4.59 (2H, d, J = 5.8 Hz, and diethyl ether (15 mL) to give a brown solid (60 mg,
6
-DMSO, 298 K) δ: 7.99 (2H, d, J = 2.9 Hz, resulted in precipitation of the product. The precipitate was
H
2
H
1
e
c
), 7.59 (2H, s, H ), 7.09 (2H, t, J = collected by filtration, and washed with ethyl acetate (15 mL)
b
d
NH
13
1
a
). C NMR (125 MHz, d
6
-DMSO, 298 K) δ: 153.6, 144.5, 0.03 mmol, 70%). H NMR (500 MHz, d
), 121.3 (C ), 117.8, 89.8, (8H, s, H ), 8.26 (8H, s, H ), 7.63 (8H, s, H
6.41 (8H, s, HNH), 5.59 (4H, br, HOH), 4.55 (8H, s, Ha).
6
-DMSO, 298 K) δ: 8.36
42.2, 139.2 (C
c
), 136.9 (C ), 124.2 (C
e
b
d
e
c
b
), 7.30 (2H, s, H ),
d
+
8
6.4, 60.8 (C ). HR ESI-MS (CH CN) m/z = 340.1190 [M + H]
a
3
−
1
13
(calc. for C20
H
14
N
5
O, 340.1204). IR: ν (cm ) 3326, 3294, 3163,
C NMR (125 MHz, d
6
-DMSO, 298 K) δ: 154.0, 146.9, 141.4,
), 125.1 (C ), 120.7, 119.1, 91.9,
3
020, 2148, 1644, 1585, 1424, 1329, 1246, 1156. Anal. calcd for 138.3 (C
c
), 135.9 (C ), 125.3 (C
e
d
b
C H N O: C, 70.37; H, 4.43; N, 20.52%. Found: C, 70.21; 83.3, 60.7 (C ). HR ESI-MS (DMSO/CH CN) m/z = 394.5820
2
0
15
5
a
3
4
+
H, 4.54; N, 20.52%.
Synthesis of [Pd
[M − 4(BF
4
)
4
]
(calc. for C80
H
60
N
20
O
4
, 394.5797), 342.1401
O, 342.1401). IR: ν (cm
+
−1
2
(tripy)
4
](BF
4
)
4
. A solution of tripy (62 mg, [3A-tripy + H] (calc. for C20
H
16
N
5
)
0
.20 mmol) and [Pd(CH CN) ](BF ) (44 mg, 0.10 mmol) in 3468, 3384, 3230, 2220, 1637, 1592, 1547, 1443, 1020. Anal.
3 4 4 2
acetonitrile (6 mL) was stirred for 1 hour. Vapor diffusion with calcd for
80 60 4 16 20 4 2 2
C H B F N O Pd ·5H O·4DMSO: C, 45.50;
diethyl ether gave a white solid (76 mg, 0.040 mmol, 85%). H, 4.07; N, 12.03%. Found: C, 45.20; H, 3.77; N, 12.28%.
1
H NMR (500 MHz, CD CN, 298 K) δ: 9.32 (8H, d, J = 1.8 Hz,
3
X-ray crystallography [Pd (2A-tripy) ](BF )
2
4
4 4
H
c
), 9.05 (8H, dd, J = 6.1 Hz, 1.1 Hz, H
f
), 8.17 (8H, dt, J =
8
.0 Hz, 1.4 Hz, H
d
), 7.65 (8H, ddd, J = 8.0 Hz, 5.9 Hz, 0.6 Hz, Yellow block crystals of [Pd
2
4 4 4
(2A-tripy) ](BF ) were grown by
H ), 7.64 (8H, t, J = 0.8 Hz, H ), 4.63 (8H, dd, J = 5.6 Hz, 0.8 Hz, vapour diffusion of diethyl ether in a solution of MeCN. X-ray
e
b
1
3
H
a
), 3.57 (4H, t, J = 5.6 Hz, HOH). C NMR (125 MHz, CD
3
CN, data were collected at 100(1) K on an Agilent Technologies
2
98 K) δ: 154.5, 154.4 (C
c
), 151.4 (C ), 144.5 (C ), 143.2, 128.5 Supernova system using Cu Kα radiation with exposures over
f
d
2
8
(
(
C ), 126.5 (C ), 124.1, 94.8, 83.2, 62.2 (C ). HR ESI-MS 1.0°, and data were treated using CrysAlisPro software. The
e
b
a
2
+
29
CH
3
CN) m/z
Pd
921, 2852, 2220, 1685, 1595, 1545, 1483, 1420, 1043, 1027. within the WinGX package. All non-hydrogen atoms were
Anal. calcd for C80 Pd ·3H O: C, 51.67; H, 3.14; refined anisotropically. Hydrogen atoms were placed in calcu-
N, 9.04%. Found: C, 51.87; H, 3.09; N, 8.71%. lated positions and refined using a riding model. The structure
Synthesis of [Pd (2A-tripy) ](BF ) . A solution of 2A-tripy was solved in the primitive tetragonal space group P4/m and
=
816.1313 [M
−
(BF
4
)
2
]
(calc. for structure was solved using Sir-97 and weighted full-matrix
−
1
2
30
C
2
80
H
52
N
12
O
4
2
, 816.1313). IR: ν (cm ) 3211, 3097, 2976, refinement on F was carried out using SHELXL-97 running
3
1
H
52
B
4
F
16
N
12
O
4
2
2
2
4
4 4
(
0
40 mg, 0.12 mmol) and [Pd(CH
.060 mmol) in 1.5 mL d -DMSO in a tube was purged with N
and was heated at 50 °C for 6 hours. Diethyl ether (30 mL) was ions. The counter-anions and solvent molecules present in the
3
CN)
4
](BF
4
)
4
(26 mg, refined to an R
1
value of 14.4%. The asymmetric unit contains
6
2
two half ligands and two quarter occupancy palladium
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solvent lattice were severely disordered and could not be
appropriately modelled. The SQUEEZE routine within PLATON
was employed to resolve this problem, resulting in ten void
spaces (total of 460 electrons), variously assigned to tetra-
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fluoroborate anions (8 in total), H O (5 in total) and MeCN
2
(3 in total) solvent molecules (total of 444 electrons), as
described below. Despite repeated efforts (>50 crystallisations
and data collections over a two year period) to crystallise the
compound, the most suitable candidate was small and a poor
diffractor. The data quality is poor, with two A alerts
(a large Hirshfield difference and high MainMol Ueq. com-
pared to neighbours) and many B alerts, and we emphasise
that metric data cannot be reliably extracted from the structure
and should be treated with caution. However, the connectivity
of the cationic framework is readily apparent. The methylene
alcohol substituents from the four neighbouring cages in the
lattice interpenetrate the cavity of each cage (Fig. 3 and ESI†),
interpenetrating groups shown in spacefilling mode (yellow,
green, blue and pink) and preclude cisplatin encapsulation in
the solid state. Around the coordinating pyridine rings, the
amino groups form a hydrogen bonding network. SQUEEZE
details and crystallographic parameters can be found in the
ESI.†
1
431; (g) B. Therrien, G. Suess-Fink, P. Govindaswamy,
A. K. Renfrew and P. J. Dyson, Angew. Chem., Int. Ed., 2008,
7, 3773–3776; (h) Y.-R. Zheng, K. Suntharalingam,
4
Acknowledgements
T. C. Johnstone and S. J. Lippard, Chem. Sci., 2015, 6, 1189–
1193.
The authors wish to thank Dr Dave McMorran and Dr Warrick
Lo for fruitful discussions, and Christopher Anderson for early
synthetic efforts. DP and JEML thank the University of Otago
for PhD scholarships. JDC (Laurenson Award, LA307) and DP
5 (a) P. Mal, B. Breiner, K. Rissanen and J. R. Nitschke,
Science, 2009, 324, 1697–1699; (b) M. Yamashina, Y. Sei,
M. Akita and M. Yoshizawa, Nat. Commun., 2014, 5.
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16, 12373–12380; (b) I. A. Riddell, M. M. J. Smulders,
J. K. Clegg and J. R. Nitschke, Chem. Commun., 2011, 47,
(MacQueen Summer Scholarship) thank the Otago Medical
Research Fund for financial support. The authors thank the
Department of Pharmacology and Toxicology and the Depart-
ment of Chemistry, University of Otago for additional funding.
4
57–459.
(a) R. A. Kaner and P. Scott, Future Med. Chem., 2015, 7,
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7
1
Notes and references
1
(a) For some selected recent reviews see: (b) T. R. Cook and
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(
c) A. M. Castilla, W. J. Ramsay and J. R. Nitschke, Acc.
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2
013, 42, 1728–1754; (e) N. J. Young and B. P. Hay, Chem.
Commun., 2013, 49, 1354–1379; (f) T. Nakamura, H. Ube
and M. Shionoya, Chem. Lett., 2013, 42, 328–334;
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g) J. E. Beves, B. A. Blight, C. J. Campbell, D. A. Leigh and
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Chem. Rev., 2011, 111, 6810–6918; (i) Y.-F. Han, H. Li and
G.-X. Jin, Chem. Commun., 2010, 46, 6879–6890;
(
(
j) M. D. Ward, Chem. Commun., 2009, 4487–4499;
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Chem. Rev., 2008, 252, 940–963; (l) M. Han, 10 (a) V. Croue, S. Goeb and M. Salle, Chem. Commun., 2015,
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Dalton Transactions
D. Stalke, T. Suenobu, S. Fukuzumi, H. Frauendorf and
G. H. Clever, Angew. Chem., Int. Ed., 2013, 52, 10102–10106.
1 (a) S. M. McNeill, D. Preston, J. E. M. Lewis, A. Robert,
the dibromopalladium(II) complexes, see: (b) H. V. Huynh,
Y. Han, R. Jothibasu and J. A. Yang, Organometallics, 2009,
28, 5395–5404.
1
K. Knerr-Rupp, D. O. Graham, J. R. Wright, G. I. Giles and 19 (a) J. E. M. Lewis, A. B. S. Elliott, C. J. McAdam,
J. D. Crowley, Dalton Trans., 2015, 44, 11129–11136;
b) S. V. Kumar, W. K. C. Lo, H. J. L. Brooks and
J. D. Crowley, Inorg. Chim. Acta, 2015, 425, 1–6;
K. C. Gordon and J. D. Crowley, Chem. Sci., 2014, 5, 1833–
1843; (b) J. E. M. Lewis, C. J. McAdam, M. G. Gardiner and
J. D. Crowley, Chem. Commun., 2013, 49, 3398–3400.
(
4
+
(c) S. K. Vellas, J. E. M. Lewis, M. Shankar, A. Sagatova, 20 Cisplatin binding within the [Pd (tripy) ] cage cavity is
2
4
J. D. A. Tyndall, B. C. Monk, C. M. Fitchett, L. R. Hanton
and J. D. Crowley, Molecules, 2013, 18, 6383–6407;
only observed in CD
water soluble [Pd
3
7
CN and DMF-d . We have generated a
4
+
−
2
(tripy)
4
]
3
cage using nitrate (NO )
(
d) J. E. M. Lewis, E. L. Gavey, S. A. Cameron and
counter anions and examined the binding of cisplatin in
O (ESI†). As we have seen previously in D O (ref. 19a) no
J. D. Crowley, Chem. Sci., 2012, 3, 778–784.
D
2
2
1
2 D. Preston, A. Fox-Charles, W. K. C. Lo and J. D. Crowley,
Chem. Commun., 2015, 51, 9042–9045.
3 (a) Other workers have also shown that nucleophiles will
react with palladium(II) based metallosupramolecular
cages, see: (b) R. Zhu, J. Luebben, B. Dittrich and
G. H. Clever, Angew. Chem., Int. Ed., 2015, 54, 2796–2800;
host–guest interaction between the cage and cisplatin
was observed (ESI†). Similarly, we have also examined the
4
+
1
host–guest interaction between the[Pd
2
(tripy)
4
]
cage and
cisplatin in DMSO-d (ESI†). As with the D
6
2
O experiments
no interaction between the cage and the cisplatin
was observed. Additionally, the DMSO solvent reacts with
(c) J. J. Henkelis, J. Fisher, S. L. Warriner and M. J. Hardie,
cisplatin releasing either (Cl− or NH
3
) ligands which
then react with the [Pd (tripy) ] cation leading to cage
4
+
Chem. – Eur. J., 2014, 20, 4117–4125.
2
4
1
4 (a) A. M. Johnson, O. Moshe, A. S. Gamboa, B. W. Langloss,
decomposition (ESI†).
J. F. K. Limtiaco, C. K. Larive and R. J. Hooley, Inorg. Chem., 21 T. Y. Kim, N. T. Lucas and J. D. Crowley, Supramol. Chem.,
011, 50, 9430–9442; (b) P. Liao, B. W. Langloss, 2015, 27, 734–745.
A. M. Johnson, E. R. Knudsen, F. S. Tham, R. R. Julian and 22 A referee suggested that the hydrogen bonding between the
R. J. Hooley, Chem. Commun., 2010, 46, 4932–4934; exo-methylene alcohol unit and the central endo-pyridyl
c) K. J. Kilpin, M. L. Gower, S. G. Telfer, G. B. Jameson and
2
4
+
(
group observed in the crystal structure of [Pd (2A-tripy) ]
2 4
J. D. Crowley, Inorg. Chem., 2011, 50, 1123–1134.
could be responsible for the lack of cisplatin binding.
1
5 (a) Other workers have previously shown that 2-aminopyri-
dine ligands will coordinate to palladium(II) and platinum
We rule that postulate out for the following reasons: (1)
1
H NMR and DOSY experiments (DMF-d , ESI†) suggest
7
4
+
(II) ions through the pyridyl nitrogen in a monodentate
that the [Pd
2
(2A-tripy)
4
]
cage is monomeric in solution.
4
+
fashion, see ref. 15b–d. However, the 2-aminopyridine unit
can also act as a bridging ligand, see ref. 15e; (b) K. Sakai,
N. Akiyama and M. Mizota, Acta Crystallogr., Sect. E: Struct.
Rep. Online, 2003, 59, m459–m461; (c) C. Gómez-Vaa-
monde, A. Alvarez-Valdés, M. Navarro-Ranninger and
J. Masaguer, Transition Met. Chem., 1984, 9, 52–54;
The diffusion coefficient for the [Pd
2
(2A-tripy)
4
]
cage was
essentially identical to that of cage that does not feature
the exo-methylene alcohol unit suggesting that those two
cages are of very similar size in solution. Additionally, the
1
H NMR spectra of the cage are not concentration depen-
4
+
2 4
dant ruling out any aggregation of the [Pd (2A-tripy) ]
(
d) S. A. Al-Jibori, Q. K. A. Al-Jibori, H. Schmidt,
cage into a larger oligomeric or polymeric architectures; (2)
All the cages studied have the exo-methylene alcohol unit
and the central endo-pyridyl group. Thus if hydrogen
bonding between those substituents was a factor it would
be expected to interfere with cisplatin binding in all the
K. Merzweiler, C. Wagner and G. Hogarth, Inorg. Chim.
Acta, 2013, 402, 69–74; (e) J. M. Casas, B. E. Diosdado,
J. Forniés, A. Martín, A. J. Rueda and A. G. Orpen, Inorg.
Chem., 2008, 47, 8767–8775.
6 (a) While our manuscript was being refereed Casini, Kuhn
and co-workers reported the X-ray crystal structure of the
4
+
1
cage systems. It does not, the [Pd
[Pd (tripy) ]
2 4
2
(3A-tripy)
both interact with cisplatin in DMF solution.
see: 23 A. Ahmedova, D. Momekova, M. Yamashina, P. Shestakova,
4
]
and
4
+
4
+
cisplatin–[Pd (tripy) ]
host
b) A. Schmidt, V. Molano, M. Hollering, A. Pöthig,
A. Casini and F. E. Kühn, Chem. – Eur. J., 2016, 22, 2253–
256.
guest
adduct,
2
4
(
G. Momekov, M. Akita and M. Yoshizawa, Chem. – Asian J.,
2016, 11, 474–477.
24 W. K. C. Lo, G. S. Huff, D. Preston, D. A. McMorran,
G. I. Giles, K. C. Gordon and J. D. Crowley, Inorg. Chem.,
2015, 54, 6671–6673.
2
1
1
7 W. M. Haynes, CRC Handbook of Chemistry and Physics,
Taylor & Francis, 93rd edn, 2012.
8 (a) Huynh and co-workers have shown that benzimidazol-2- 25 P. R. Symmers, M. J. Burke, D. P. August, P. I. T. Thomson,
ylidene-dibromopalladium(II) complexes can be used to
G. S. Nichol, M. R. Warren, C. J. Campbell and P. J. Lusby,
probe the σ-donor strength of the ligands trans to the
Chem. Sci., 2015, 6, 756–760.
benzimidazol-2-ylidene. They have found that there is a 26 C. M. Amb and S. C. Rasmussen, J. Org. Chem., 2006, 71,
direct relationship between the σ-donor strength of the 4696–4699.
trans monodentate ligand and the chemical shift of the 27 H. V. Huynh, Y. Han, J. H. H. Ho and G. K. Tan, Organo-
1
3
benzimidazole carbene carbon in the C NMR spectra of
metallics, 2006, 25, 3267–3274.
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This journal is © The Royal Society of Chemistry 2016
Dalton Trans.