Development of 18F-fluorinatable dendrons and their application to cancer cell targeting

Article abstract of DOI:10.1039/c1nj20417c

¹⁸F-fluorine is an ideal imaging PET (positron emission tomography) nuclide due to the low energy of its β+ (positron) and pure β+ decay. PAMAM dendrimers are branched organic molecules with multiple NH₂ termini each of which can acc

Full text of DOI:10.1039/c1nj20417c

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PAPER
18
Development of F-fluorinatable dendrons and their application to
cancer cell targeting
Laurent Trembleau,b Michael Simpson,ab Richard W. Cheyne,ab Imma Escofet,b
M. Virginia C. A. L. Appleyard,c Karen Murray,c Sheila Sharp,c
Alastair M. Thompsonc and Tim A. D. Smithw*a
Received (in Montpellier, France) 13th May 2011, Accepted 18th July 2011
DOI: 10.1039/c1nj20417c
18F-fluorine is an ideal imaging PET (positron emission tomography) nuclide due to the low
energy of its b+ (positron) and pure b+ decay. PAMAM dendrimers are branched organic
molecules with multiple NH2 termini each of which can accommodate the attachment of
functionalities. The presence of a thiol group also facilitates the conjugation of a targeting
molecule. Here we describe the attachment of boroaryl moieties to the terminal NH2 groups of
dendrimers and after cleavage of the bridging thiol group conjugation of the functionalized
dendrons (half-dendrimers) to biotin. Incubation of the targeted, boroaryl-functionalized
dendrons with 18F-fluoride in acetic acid facilitated their radiolabelling with a labelling efficiency
of about 60%. We then demonstrated that the 18F-dendron–biotin bound to HER-2 expressing
cells in vitro pre-targeted with the avidin–trastuzumab conjugate. The cell-associated 18F
decreased with increasing dendron size suggesting that the larger dendrons sterically hindered
binding of avidin with biotin. This is the first study to demonstrate that dendrimers can be
functionalized with 18F-fluorinatable groups and used to target cell surface receptors.
functionalised by a variety of agents—most notably DTPA
Introduction
(diethylene triamine pentaacetic acid), a chelating group that
Nanoscale platforms utilized in medical imaging and therapy
binds to several metals including the MRI contrast enhancer,
gadolinium (Gd3+ 7
99mTc).8 Due to their potential to carry high drug payloads
delivery systems encompass a broad range of molecules both
inorganic, including Super Paramagnetic Iron oxide (SPIO)1
)
and the SPECT nuclide Technetium
(
and TiO2 crystals, and organic e.g. dendrimers3,4 and
2
dendrimers are also being employed as drug carriers e.g.
Kukowska-Latallo et al.9 have shown that attachment of
methotrexate to dendrimers and folic acid to target tumour
cells increased the antitumor activity of methotrexate and
markedly decreased its toxicity, allowing therapeutic responses
not possible with free drug. Using dendrimers functionalised
for imaging capability facilitates the in vivo detection of the
distribution of the conjugated drug using PET.
18F is an ideal PET isotope, in terms of nuclear character-
istics, for imaging due to the simple decay and emission
properties with high (97%) positron abundance and relatively
low positron emission energy (0.635 MeV max). The latter
means that 18F has a short positron linear range in tissue
producing high resolution PET imaging. In addition, all PET
units use 18F, which is consequently the most readily available
PET isotope.
fullerenes.5 Functionalities appended by chemical modifica-
tion enable the attachment of targeting moieties such as
antibodies and, for PET (positron emission tomography)
and SPECT (single photon emission tomography) imaging
and radiation therapy, the attachment of multiple binding sites
for metal radionuclides. PAMAM (poly(amido amine))
dendrimers are branching polymers built from dendrons
consisting of b-alanine subunits and are water soluble, non-
toxic at moderate concentrations6 and biocompatible. Surface
groups on the outer shell of dendrimers have been
a John Mallard PET Unit, Aberdeen Biomedical Imaging Centre,
School of Medical Sciences, University of Aberdeen, Biomedical
Physics Building, Foresterhill, Aberdeen AB25 2ZD, UK.
E-mail: t.smith@abdn.ac.uk
b School of Natural Sciences and Computing, University of Aberdeen,
Aberdeen AB24 3UE, UK
In this study we describe the functionalizing of the amine
branch termini of PAMAM dendrons (half dendrimers) with
fluorinatable boroaryl groups,10 facilitating [18F]-fluorination
at room temperature compatible with the temperature-
sensitivity of dendrons, conjugation with biotin and
investigate their ability to target cells pre-treated with an
c Centre for Oncology and Molecular Medicine, Division of Medical
Sciences, University of Dundee, Ninewells Hospital & Medical
School, Dundee DD1 9SY, UK
w Biomedical Physics Building, School of Medical Sciences, University
of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, UK. Tel: +44 1224
553481.
c
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avidin-conjugated antibody. The commercially available
dendrimers used in this study possess a disulfide linkage, which
when cleaved produces two dendrons each with thiol groups
available for conjugation.
General procedure for the attachment of boronic esters to
dendrimers
The dendrimer (as a 10% or 20% solution in MeOH, depending
on the generation) was added to a stirred solution of ester 2
(1 eq. per terminal amine in the dendrimer) in CHCl3
(equal volume to the dendrimer solution). The resultant
insoluble material was dissolved by the addition of MeCN
(equal volume to the dendrimer solution) and the mixture was
stirred at room temperature under Ar for 24 hours, or until
TLC analysis showed complete consumption of compound 2.
The solvents were then removed by evaporation.
Methods
Synthesis of 2,6-difluoro-4-[(carboxymethoxyimino)methyl]phenyl-
boronic acid pinacol ester (1)
A mixture of (O-carboxymethyl)hydroxylamine hemihydro-
chloride (171 mg, 1.57 mmol), 2,6-difluoro-4-formylphenyl-
boronic acid pinacol ester (350 mg, 1.31 mmol) and Hunig’s
¨
General procedure for the attachment of potassium
trifluoroborate salts to dendrimers
base (102 mg, 137 mL, 0.787 mmol) was stirred in anhydrous
MeCN (10 mL) containing 3 A molecular sieves at room
temperature under argon (Ar) for 2 hours. The solvent was
evaporated and the residue dissolved in EtOAc. The ethyl
acetate was washed once with H2O then dried (MgSO4),
filtered and evaporated to leave a white solid. This was
recrystallised from a mixture of hot petroleum ether 40/60
and a few drops of EtOAc to leave the product as a white
solid; 339 mg (76%); mp 151–152 1C; dH(250 MHz; CDCl3)
1.37 (12 H, s), 4.77 (2 H, s), 7.06 (2 H, d, J 7.6), 8.09 (1 H, s),
10.40 (1 H, bs); dC(62.5 MHz; CDCl3) 24.7, 70.5, 84.4, 109.8
(d, J 28), 136.5 (t, J 10), 148.5, 166.6 (dd, J 251, 13), 175.4;
dB(192 MHz; CDCl3) 29.1; m/z (ESI) 342.1323 ([M + H]+.
C15H19BF2NO5 requires 342.1324).
The dendrimer solution was concentrated, then dissolved in
CHCl3 and concentrated again to remove traces of MeOH.
The dendrimer was then dissolved in H2O (100 mLper mg
dendrimer) and a solution of trifluoroborate 3 (1 eq. per
terminal amine in the dendrimer) in MeCN (200 mL per mg
dendrimer) was added. The mixture was stirred at room
temperature under Ar for 24 hours, or until TLC analysis
showed complete consumption of compound 3.
General procedure for dendrimer cleavage (producing dendrons)
and attachment of biotin
For dendrimers produced using boronic esters, the crude
material was initially dissolved in a mixture of H2O (equivalent
to 100 mL per mg initial dendrimer) and MeCN (equivalent to
100 mL per mg initial dendrimer). For dendrimers produced
using potassium trifluoroborate salts, no additional solvents
were added. TCEPÁHCl (tris(2-carboxyethyl)phosphine) (1 eq.)
was added as an aqueous solution (10 mL per mg initial
dendrimer) and the reaction mixture was stirred at room
temperature under Ar for 1 hour (Scheme 2). Biotin-maleimide
4 (2 eq., 1 eq. per thiol) was then added and the resultant
suspension stirred at room temperature under Ar for 24 hours,
or until the insoluble material (compound 4) had dissolved.
Product (4 branch dendron): dH(250 MHz; CD3OD) 1.40–1.80
(22 H, m), 2.08–2.33 (8 H, m), 2.40–3.50 (58 H, m), 4.28–4.37
(1 H, m), 4.46–4.56 (1 H, m), 4.64 (7 H, s), 4.76 (1 H, s), 7.03 (2 H,
t, J 8.2), 7.18–7.30 (8 H, m), 8.25 (4 H, s); m/z (TOF MS ES)
567.1989 ([M À 4K]4À. C88H112B4F20N22O19S2 requires 567.1998).
Synthesis of 2,6-difluoro-4-{[2-oxo-2-(pentafluorophenoxy)ethoxy-
imino]methyl}-phenylboronic acid pinacol ester (2)
EDC hydrochloride (26 mg, 0.136 mmol) and pentafluoro-
phenol (22 mg, 0.120 mmol) were added to a solution of 1
(40 mg, 0.117 mmol) in CHCl3 (1.5 mL) and the mixture was
stirred at room temperature under Ar for 15 hours. The
chloroform was evaporated and the residue dissolved in a
mixture of EtOAc and H2O. The layers were separated and
the organic portion dried (MgSO4), filtered and evaporated
to leave the product as a colourless oil; 59 mg (100%);
dH(250 MHz; CDCl3) 1.38 (12 H, s), 5.06 (2 H, s), 7.08 (2 H,
d, J 6.7), 8.12 (1 H, s); dC(62.5 MHz; CDCl3) 24.7, 70.2, 84.5,
109.8 (d, J 28), 136.2 (t, J 11), 149.0, 165.6, 166.6 (dd, J 251, 14);
dB(192 MHz; CDCl3) 29.4; m/z (ESI) 508.1167 ([M + H]+.
C21H18BF7NO5 requires 508.1161).
Synthesis of potassium 2,6-difluoro-4-{[2-oxo-2-(pentafluoro-
phenoxy)ethoxyimino]-methyl}phenyltrifluoroborate (3)
Labelling of trifluoroboroaryl functionalised dendron–biotin
conjugates with 18F-fluoride
A 4 M aqueous solution of KHF2 (792 mL, 3.17 mmol) was
added to a stirred solution of ester 2 (804 mg, 1.59 mmol) in
MeOH (30 mL) and stirring continued at room temperature
for 2 hours (Scheme 1). The solvents were evaporated and the
residue was dissolved in hot acetone and hot MeCN then
filtered. The filtrate was evaporated and the residue suspended
in chloroform. After filtration, the product was obtained as a
white solid; 601 mg (78%); dH(400 MHz; DMSO-d6) 5.25
(2 H, s), 6.94 (2 H, d, J 7.8), 8.33 (1 H, s); dC(62.5 MHz;
acetone-d6) 168.6, 167.1, 150.9, 142.9, 141.4, 139.8, 138.2,
132.2, 125.3, 109.8, 70.9; dF(376 MHz; acetone-d6) À103.4,
À135.8, À154.6, À160.2, À164.6; m/z (TOF MS ES) 448.0224
([M À K]À. C15H5BF10NO3 requires 448.0203).
Dendron preparations (0.1 mmol) were dissolved in 10 mL of
DMSO to which was added 5 mL of glacial acetic acid followed
by 20 MBq (10 mL) of 18F-fluoride in saline. These compo-
nents were mixed on a whirlimix and incubated at room
temperature. The labelling progress was followed by applying
a sample (1 mL) to a silica gel thin layer chromatography sheet
and placing in a covered 100 mL glass beaker containing 5 mL
of a mixture of acetonitrile and water (70 : 30). After the solvent
had reached the top of the sheet it was dried and the location of
radioactivity determined on a Minigita Star radiochromato-
graphy scanner (Raytest UK). After 1 h free 18F-fluoride was
removed by applying to a silica cartridge (Waters UK) and
eluting the dendron–biotin with 25 mL of a mixture of acetonitrile
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Scheme 1 Synthesis of potassium 2,6-difluoro-4-{[2-oxo-2-(pentafluorophenoxy)-ethoxyimino]-methyl}-phenyltrifluoroborate.
and water (70 : 30). TLC confirmed the absence of free
18F-fluoride from the sample. The purified 18F-dendron–biotin
was diluted in the medium for cell incubation studies.
confluent flask of cells was trypsinised and the cells placed
into 25 cm2 growth surface area tissue culture flasks and
allowed to grow for 3 days.
Cells
Preparation of trastuzumab–neutravidin
MDA-MB-435, MDA-MB-468 and SKBr3 breast tumour
cells were obtained from the European Collection of Cell
Cultures (ECACC) and the American Type Culture Collection
(ATCC). Each cell line was cultured in Dulbecco’s Modified
Eagle’s Medium supplemented with foetal bovine serum
(10%) and penicillin (100 units mLÀ1) and streptomycin
(100 mg mLÀ1). Cells were grown to confluence in 75 cm2
growth surface area tissue culture flasks and routinely
passages at a 1 : 10 dilution. For tracer binding studies a
Trastuzumab (Herceptin, Roche UK) (1 mg) was dissolved in
1 mL of water and washed on a 4 mL 100 KDa molecular
weight cut off centrifugal filter (Millipore) by centrifugation at
2000 g for 15 min. The retained trastuzumab was then washed
twice with phosphate buffered saline (PBS) and finally dissolved
in 100 mL of Imject Maleimide conjugation buffer (phosphate
(100 mM), EDTA (10 mM) pH 7.2) (Thermo UK). The
suspension was diluted to 500 mL with water to which was
added 10 mL of dithiothreitol (DTT) and incubated for 1 h at
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Scheme 2 Attachment of boroaryl functionalities to dendrons.
room temperature to allow disulfide reduction to occur. DTT was
removed by centrifugation on a 0.5 mL 50 KDa molecular weight
cut off centrifugal filter (Millipore) by centrifugation at 12 000 g
for 15 min followed by 3 washes with 0.5 mL of Imject Maleimide
conjugation buffer. The retained, reduced, trastuzumab was
recovered in 100 mL of Imject Maleimide conjugation buffer to
which was added 1.5 mg of maleimide activated neutravidin
(Thermo UK) and left overnight at room temperature.
was dissolved in 100 mL of PBS to which was added 0.5 mg of
biotin. The mixture was incubated at 37 1C for 1 h to allow the
biotin to bind to avidin after which excess biotin was removed
by washing on a 0.5 mL 30 KDa molecular weight cut off
centrifugal filter (Millipore) by centrifugation at 12 000 g for
15 min followed by 3 washes with 0.5 mL of PBS. The retained
avidin was dissolved in 100 mL of PBS.
Incubation with trastuzumab–neutravidin and with
18F-dendron–biotin
Preparation of biotin-saturated avidin
To saturate avidin-binding sites on tumour cells, a potential
source of non-HER-2 binding of the avidin–trastuzumab
conjugate, biotin-saturated avidin was prepared. Avidin (3 mg)
Media were removed from flasks of cells then trastuzumab–
neutravidin and biotin-saturated avidin, prepared as described,
were added to 5 mL of medium and 0.3 mL added to each flask
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of cells. Additional control flasks of cells were not incubated
with trastuzumab–neutravidin but were used to determine
non-targeted binding of 18F-dendron–biotin. After 1 h incuba-
tion at 37 1C, the media were removed and cells were washed
three times with PBS. 18F-dendron–biotin with 4, 8 or 16
branched-dendrons was then added to the cells and incubated
at 37 1C for 1 h.
Statistical methods
Results were expressed as mean Æ standard deviation.
Significant differences between means were determined using
the Student t test. Values for ‘t’ and their significance are
stated in each case.
Fig. 2 Association of 18F-dendron–biotin with HER-2 expressing
tumour cells during a 1 h incubation with 18F-dendron–biotin using
dendrons with 4, 8 and 16 branches. The HER-2 receptors were
pre-targeted by incubation of cells with (white) and without (black)
the trastuzumab–neutravidin conjugate for 1 h then washed with PBS
prior to addition of the 18F-dendron–biotin. (Units: cpm per mg protein
per KBq activity.)
Results
Fig. 1 shows the rate of labelling of trifluoro-boroaryl
functionalised 4, 8 and 16 branch dendrons with 18F. The rate
is similar for each dendron, achieving about 55% labelling
after 100 min. The specific activity was 9.6, 10 and 11 MBq mmolÀ1
for the 4, 8 and 16 branch dendrons, respectively, and radio-
chemical purity for each dendron after purification on a silica
cartridge was 499%.
HER-2 positive breast cancer cells pre-incubated with
avidin–trastuzumab cells were incubated with 18F-dendron–
biotin (1 KBq mLÀ1). With increasing dendron size the
amount of 18F-dendron–biotin that binds decreased signifi-
cantly (8 branch vs. 4 branch (t = 4.74, p o 0.01); 16 branch
vs. 8 branch (t = 8.68, p o 0.001)) suggesting that the larger
dendron size interferes with avidin binding (Fig. 2; white
columns). Dye-exclusion measurements using trypan blue
carried out 24 hours after incubating tumour cells with the
16 branch dendron (1 mg mLÀ1) for 90 minutes demonstrated
that 495% of the cells were viable which was comparable
with untreated cells. Further the number of cells in the treated
flasks (2.5 Â 106 Æ 1.9 Â 105) was also comparable with the
number of cells in the control flasks (2.1 Â 106 Æ 0.9 Â 105)
(t = 0.3, not significant) indicating that the lower binding of
18F-dendron(16)-biotin compared with the smaller ones was
not due to toxicity of the larger dendrons.
Fig. 3 Association of 18F-dendron–biotin (4 branch), corrected for
non-specific uptake, with cells expressing HER-2 at high (SKBr3) and
medium (MDA-MB-453) levels and HER-2 negative (MDA-MB468)
breast cancer cells.
uptake represented only 15% of the activity associated with
pre-targeted cells, but about 50% for the 16 branch dendron.
We therefore carried out further experiments using the 4
branch dendron.
The targeting specificity of 18F-dendron–biotin was examined
using cells with high (SKBr3), medium (MDA-MB-453) and
low/zero (MDA-MB-468) HER-2 expression pre-targeted
with avidin–trastuzumab. After correction for non-specific
dendron uptake, the cell associated activity corresponds
to HER-2 expression (Fig. 3). (SKBr3 vs. MDA-MB-453
(t = 10.5, p o 0.001); SKBr3 vs. MDA-MB-468 (t = 14.3,
p o 0.001); MDA-MB-468 (t = 6.95, p o 0.001).
Activity associated with cells after incubation with 18F-dendron–
biotin that had not been pre-incubated (pre-targeted) with
avidin–trastuzumab was similar for each dendron (Fig. 2;
black columns). For the 4 branch dendrons, non-specific
Discussion
Dendrimers have unique structures with a hydrophobic inter-
ior which allows lipophilic-drug encapsulation and a hydro-
philic exterior made up of multiple branch termini facilitating
functionalization with targeting molecules and labels for
detection using imaging techniques. Dendrimers have been used
to enhance the solubilisation of many drugs including the anti-
cancer drugs 5FU, cisplatin, camptothecin and doxorubicin.11,12
To facilitate PET imaging of dendrimer biodistribution we have
Fig. 1 Labelling of trifluoroboroaryl-dendron–biotin with 18F-fluorine
using 4 (triangles), 8 (squares) or 16 (circles) branch dendrons.
c
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synthesised dendrimers that can be readily [18F]-fluorinated in
aqueous solvent. We used low generation dendrimers because
smaller dendrimers exhibit rapid renal excretion but larger
ones accumulate in the liver6,13 which is important to avoid in
imaging and therapeutic agent delivery.
Counterintuitively, we found that with increasing dendron
size the amount of 18F-labelled biotin associated with avidin–
trastuzumab pretargeted cells decreased. This could be due to
the dendron causing steric hindrance in the binding of biotin
to avidin and it may be possible to overcome this problem
using longer length spacer arms between the biotin moiety and
the dendron. With increasing generation and hence size dendri-
mers become less flexible22 which may influence the ability of
conjugated molecules to interact with other macromolecules.
We noted a modest degree of non-receptor associated
binding as some binding was evident by cells that had not
been pre-incubated with avidinised-trastuzumab. Non-specific
tumour uptake of dendrimers has been reported previously.23-25
Saovapakhiran et al.24 demonstrated that PAMAM dendri-
mers were internalised by HT-29 cells by both caveolae-
dependent and macropinocytosis pathways. Although the
smallest dendron used in our study has only 4 branches with
each branch having 1 negatively charged ion, the dendron will
be anionic. A probable route into cells of anionic particles is via
interaction with positively charged end-groups on some cell surface
proteins which can result in internalisation by endocytosis.23
The radiolabelling procedure involves isotopic exchange
with [18F]fluoride so the specific activity of the product is
dependent on the activity of the [18F]fluoride. The radio-
labelling of the dendrons in this study was carried out by
hand limiting the amount of starting activity to 30 MBq.
Future developments of this methodology would include the
translation of the labelling procedure into an automated
system facilitating tracer synthesis using higher levels of radio-
activity for in vivo characterisation. This would facilitate the
use of GBq levels of [18F]fluoride for radiolabelling and
consequently synthesis of specific activities of [18F]dendron–
biotin in the GBq per mmol range. The in vivo testing of
[18F]dendron–biotin using animal–PET systems is an essential
step to establishing the utility of these tracers prior to further
clinical development.
Cells pre-treated with an avidinised antibody were used to
demonstrate the targeting ability of 18F-dendron–biotin
mimicking the pre-targeted approach to radiotracer delivery.
The target antigen is determined by the choice of antibody, in
this case trastuzumab, to target HER2. Trastuzumab can be
substituted for other antibodies allowing this system to be used
to target a range of cell surface receptors. The first report of
the use of a pre-targeted method using radiolabelled biotin to
detect an avidin-conjugated antibody was by Hnatowich and
Rusckowski14 in 1987 in which 111In-DTPA biotin was admi-
nistered to mice which had been inoculated into the peritoneal
cavity with avidin conjugated agarose beads. Subsequently,15
using mice bearing subcutaneous HÁEp-2 tumours and a two
step imaging procedure in which avidin conjugated to the
HMFG-1 antibody was administered prior to 111In-DTPA-
biotin produced superior target/blood 111In ratios compared
with using a directly labelled HMFG-1 antibody.
Using dendrons facilitates the attachment of multiple
nuclides for therapy as well as for imaging. One future strategy
could be to conjugate o4 NH2 branch termini with boroaryl
followed by conjugation of the remaining NH2 groups with
either DTPA or DOTA (1,4,7,10-tetraazacyclododecane-
1,4,7,10-tetraacetic acid) which requires only mild reaction
conditions.16 This would allow the tracer to be utilised for
targeting with cytotoxic nuclides such as 177Lu and 90Y after
allowing the tracer to be used for dosimetry with 18F. The use
of the DOTA chelator also enables pre-targeted radiotherapy
using nuclides such as 90Y. In a study of 43 patients with
adenocarcinomas from several different sites, Breitz et al.17
demonstrated that radioimmunotherapy with 90Y using a
pre-targeted method produced a mean tumour to marrow
absorbed dose ratio of 63 : 1 compared with 6 : 1 for conven-
tional radioimmunotherapy. More recently, a pre-targeted
system has been evaluated in vitro using biotinylated poly-
L-Lysine18 labelled with 211At.
It may be necessary for in vivo work to use biotin that has
been modified to be biotinidase-resistant. Although biotin
hydrolysis is important with the longer-lived tracer to imaging
times or for therapy with long-lived radionuclides this may not
be an issue for imaging with 18F where administration to scan
times needs to be short due to the short t1/2 of this nuclide.
In summary we have demonstrated that dendrimers/
dendrons can be functionalised with fluorinatable moieties,
subsequently labelled with 18F-fluorine and targeted to cell
surface molecules. The decreased cell-associated activity with
increased dendron size was unexpected but may be addressed
by further modification of these conjugates e.g. by altering the
nature of the linker. There remain challenges to optimise this
approach in the preclinical and clinical settings.
Direct cell targeting methods can also be applied to
dendrimers, a number of which have recently been described
and include the exploitation of lactoferrin receptor expres-
sion.19 Folic acid receptors are over-expressed in tumour cells
and the conjugation of folic acid to doxorubicin-loaded
dendrimers to target MCF7 breast cancer cells was demon-
strated by Gupta et al.20 to exhibit greater uptake of doxo-
rubicin than with doxorubicin-dendrimers or doxorubicin
alone. Thomas et al.21 showed that uptake of dendrimers
conjugated with the fibroblast growth factor was greater by
PC3 cells, which overexpress the FGF receptor, than by the
MCF7 cells which are FGF receptor negative. A popular target
in cancer is the RGD binding motif expressed on endothelial
cells and tumour cells derived from endothelial cells. Li et al.4
produced RGD–dendrimer–quantum dot conjugates and
demonstrated their ability to target melanoma cells.
Authors Contributions
TADS, LT and AMT designed the study and drafted the
manuscript. MS, LT, IE and RWC carried out the organic
synthesis. MS drafted the organic synthesis methods section.
TADS carried out radiolabelling and contributed to cell
uptake. SS, VA and KM contributed to tracer uptake studies.
All reviewed and approved the paper.
With increasing generation number and consequently an
increasing number of branches on the dendron, the potential
specific activity delivered would be expected to increase.
c
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10 R. Ting, C. Harwig, U. auf dem Keller, S. McCormick, P. Austin,
C. M. Overall, M. J. Adam, T. J. Ruth and D. M. Perrin, J. Am.
Chem. Soc., 2008, 130, 12045–12055.
11 S. Svenson and A. S. Chauhan, Nanomedicine, 2008, 3,
679–702.
Declaration of Interest
The authors report no declarations of interest.
12 S. Svenson, Eur. J. Pharm. Biopharm., 2009, 71, 445–462.
13 S.-E. Stiriba, H. Frey and R. Haag, Angew. Chem., Int. Ed., 2002,
41, 1329–1334.
14 D. J. Hnatowich and M. Rusckowski, J. Nucl. Med., 1987, 28,
1294–1302.
15 G. Rowlinsonbusza, D. J. Hnatowich, M. Rusckowski, D. Snook
and A. A. Epenetos, Antibody, Immunoconjugates, Radiopharm.,
1993, 6, 97–109.
Acknowledgements
This work was funded by the BBSRC. We are grateful to
Mr. Stuart Craib of the Aberdeen PET facility who prepared
[18F]-Fluoride.
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