Accelerating the multifunctionalization of therapeutic nanoparticles by using a multicomponent reaction

Article abstract of DOI:10.1002/chem.201103743

Simpler and faster! Multifunctionalized nanoparticles are crucial for nanobiomedical applications, but their syntheses are tedious and time-consuming. A multicomponent reaction on nanostructures is an excellent way to prepare such nanomaterials. The gold nanosystem illustrated in the scheme was built and shown to enhance cancer cell targeting and killing by combining the effects of a therapeutic drug with X-ray radiation.

Full text of DOI:10.1002/chem.201103743

COMMUNICATION
DOI: 10.1002/chem.201103743
Accelerating the Multifunctionalization of Therapeutic Nanoparticles by
Using a Multicomponent Reaction
Hongyu Zhou,[a] Gaoxing Su,[a, b] Peifu Jiao,[a, b] and Bing Yan*[a, b]
The sluggishness of the drug-discovery pipeline in recent
decades suggests limitations to the building of many desira-
ble drug properties into a small molecule with a molecular
weight of around 500. These desirable properties include at
the least anti-disease activity, low toxicity, target selectivity,
and optimal absorption, distribution, metabolism and excre-
tion properties. Because of the unique properties of nano-
particles (NPs), including large surface area and flexibility
for surface modifications, it is highly feasible to incorporate
many drug-like properties into a single nanoparticle.[1,2]
Such multifunctionalized nanoparticles (MFNPs) can be
equipped with features such as cell targeting, drug delivery,
diagnostics, and radiotherapy enhancement. Cancer target-
ing NPs for drug delivery and radiation enhancement have
been reported by others[3–6] and our own laboratory.[7–9] With
the use of either a small molecule or protein as targeting
moiety, NPs selectively enter cancer cells and enhance kill-
ing by both drug action and radiation. Strategies for assem-
bling MFNPs include: 1) multiple functions that are individ-
ually attached to NP,[10–12] and 2) multiple functions that are
attached to NP with a single attachment point.[13] Modifica-
tion strategies have been reported for carbon nano-
tubes,[10–12] quantum dots,[14] gold nanoparticles (GNPs),[15]
and supermagnetic NPs.[16,17] However, further applications
of both approaches are hindered primarily by the tedious
multistep synthesis and the associated difficulties in analyti-
cal quality control of each synthesis step. The lack of effi-
cient and well-controlled synthesis approaches to make
MFNPs has become a bottleneck limiting the exploration of
MFNPs in biological and biomedical applications. Therefore,
simpler, faster, more efficient and well-controlled ap-
proaches for preparing MFNPs are in urgent need. Here we
report a one-step multifunctionalization of nanoparticles
employing Ugi multicomponent reaction (MCR).
The four starting components for Ugi MCR are thioctic
acid, b-cyclodextrin (b-CD), folic acid, and cyclohexyl isocy-
anide. Thioctic acid serves as a linker to connect the de-
À
signed multifunctional ligand to GNPs with strong Au S
bonds. The folic acid enables MFNPs to target cancer cells
that over-express folate receptor (FR). The isocyanide
group is an extra functional group reserved for adding other
functions. b-CD is used as the drug carrier because it has
been widely used for drug formulation with good biocom-
patibility.[18] Hydrophobic drugs are easily adsorbed into b-
CDꢀs cavity through hydrophobic interactions and are then
released at a lower pH. Folic acid functions as the targeting
molecule because FR is over-expressed on the surface of
several human cancer cells compared to normal cells.
MFNPs with folic acid-targeting moieties enter cells through
receptor-mediated endocytosis.[19,20] The drug can then be re-
leased intracellularly to kill cancer cells. As an additional
lethal attack, X-ray irradiation enhanced by GNPs further
accelerates cancer cell killing at a dose that is safe to
normal cells.[3–9]
Ugi MCRs are powerful tools for building target scaffolds
with maximal diversities through a simple reaction process.
To make reagents suitable for the Ugi reaction, folic acid
was first treated with excess diamines to form an amine, and
then one of the hydroxyl groups in b-CD was converted to
an aldehyde (detailed information on the synthesis of inter-
mediates is included in the Supporting Information). The
Ugi MCR was performed at room temperature in MeOH/
DMF with thioctic acid, folic acid-derived amine, b-CD-de-
rived aldehyde, and cyclohexylisocyanide. A similar Ugi
product was obtained by using Boc-protected diethylamine
to replace folic acid-derived amine and was used as a nega-
tive control. The as-synthesized ligands were prepared in
mild conditions with high yield through a single step that
combined the linker, targeting, and drug-loading functions
into one molecule. The ligands were attached to GNPs in
situ during the synthesis of multifunctionalized GNPs by
treatment with chloroauric acid in the presence of NaBH4.
GNPs coupled with Ugi products having either a folic acid
group (MFGNP-I) or a Boc group (MFGNP-II) were syn-
thesized. MFGNP-II had the same drug-loading property as
MFGNP-I but lacked the targeting capability of MFGNP-I.
The anticancer drug doxorubicin (Dox) was loaded onto
MFGNP-I and MFGNP-II through noncovalent binding
(Scheme 1).
[a] Dr. H. Zhou, G. Su, P. Jiao, Prof. Dr. B. Yan
Department of Chemical Biology and Therapeutics
St. Jude Childrenꢀs Research Hospital
Memphis, Tennessee, 38105 (USA)
Fax : (+1)901-595-5715
E-mail: dr.bingyan@gmail.com
[b] G. Su, P. Jiao, Prof. Dr. B. Yan
School of Chemistry and Chemical Engineering
Shandong University, Jinan, 250100 (P.R. China)
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/chem.201103743.
Chem. Eur. J. 2012, 18, 5501 – 5505
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
5501

Figure 1. Characterization of multifunctionalized GNPs. a) TEM image
of MFGNP-I; scale bar: 20 nm. b) Fourier transform infrared (FTIR)
spectra of free ligand I and MFGNP-I obtained by using the KBr pellet
method. The FTIR spectrum of MFGNP-I shows similar features to that
of ligand I.
ic properties on the surface of these GNPs were similar
(Figure S3 in the Supporting Information). Both MFGNP-I
and -II had characteristic peaks at approximately 520 nm in
UV–visible spectra, which is typical for GNPs. MFGNP-I
also showed a peak for folic acid at approximately 270 nm
(Figure S4 in the Supporting Information). After analyzing
the elemental analysis data to determine the nitrogen con-
tent, we concluded that 85 and 132 molecules per nanoparti-
cle were loaded on MFGNP-I and MFGNP-II, respectively
(Table S1 in the Supporting Information).
We next explored the drug-loading and -release character-
istics of the GNPs. Drug loading was calculated by subtract-
ing the number of Dox molecules remaining in the superna-
tant after incubating GNPs with Dox for 4 h from the
number of Dox molecules initially added to the media. Ap-
proximately 50 Dox molecules were loaded onto each GNP.
About 23% of the Dox payloads were released from GNPs
within 32 h at pH 7.4. However, more than 70% of bound
Dox was released within 6 h at pH 5.5, a pH value found in
endosomes (Figure S5 in the Supporting Information).
To evaluate the cancer cell targeting and killing functions
of the GNPs, we performed a series of investigations in cul-
tured cells. HeLa cells are cervical cancer cells that over-ex-
press FR. A549 cells, which express very low levels of FR,
were used as a negative control. TEM images show that
after an 8 h incubation of cells with MFGNP-I, GNP aggre-
gates were prominent in endosome- and lysosome-like vesi-
cles in HeLa cells (Figure 2a) but rare in A549 cells (Fig-
ure 2b). We then quantitatively determined the cell binding
and uptake rates of GNPs by performing inductively cou-
pled plasma mass spectrometry (ICP-MS) analysis.
Scheme 1. Scheme of the use of a multicomponent reaction to fabricate
multifunctionalized drug-delivery nanosystems.
GNPs were characterized by a variety of techniques, in-
cluding transmission electron microscopy (TEM), Fourier
transform infrared (FTIR) spectroscopy, UV–visible spec-
troscopy, z potentials, and elemental analysis. TEM images
show that the GNPs had an average diameter of 4 nm (Fig-
ure 1a) and dynamic light scattering analyses demonstrate
that the particles had a hydrodynamic diameter of 12 nm
(Figure S1 in the Supporting Information); this indicates
a slight aggregation in aqueous solution. The FTIR spectra
of MFGNP-I and -II had similar features as did those of the
free ligands (Figure 1b and Figure S2 in the Supporting In-
formation). Specifically, all spectra contained characteristic
À1
À
C H stretching vibrations between 2850 and 2950 cm , C=
À1
À
O stretching vibrations at 1650 cm , and C O stretching vi-
brations between 800 and 1200 cmÀ1 (Figure 1b). When sus-
pended in water, MFGNP-I and -II had z potentials of À18
and À24 mV, respectively; this indicates that the electrostat-
5502
www.chemeurj.org
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2012, 18, 5501 – 5505

Multifunctionalized Therapeutic Nanoparticles
COMMUNICATION
MFGNP-II, MFGNP-I also revealed a sevenfold increase in
cell recognition enhancement in HeLa cells (Figure 3a). The
data demonstrate that MFGNP-I had a significantly en-
hanced cell recognition capability in FR positive cells. To
test whether folic acid was responsible for the enhanced cell
recognition, HeLa cells were pretreated with folic acid and
the competing free ligand partially blocked cell binding and
uptake (Figure 3b). Cell uptake of MFGNP-I was also par-
tially blocked when HeLa cells were pretreated with sodium
azide, a metabolic inhibitor, or incubated with MFGNP-I at
48C (Figure 3b); this indicates an energy-dependent endocy-
tosis process. Here we conclude that MFGNP-I achieved
cellular delivery through FR-mediated endocytosis.
GNPs have been shown to enhance both X-ray computed
tomography (CT)[21,22] and radiation-induced cell death.[23,24]
With the confirmation of drug-loading property and cell-rec-
ognition enhancement of the MFNPs, we next tested the cell
killing induced by the combined drug therapeutic efficacy
and X-ray irradiation (Figure 4). At a concentration of
Figure 2. TEM images of: a) HeLa, and b) A549 cells incubated with
MFGNP-I for 8 h. The GNP concentration was 50 mgmLÀ1. GNP aggre-
gates were trapped in organelles, such as the endosome and lysosome, as
seen from the magnified section in (a).
MFGNP-II in general showed low rates of cell binding and
uptake in both cell lines (Figure 3a), whereas MFGNP-I ex-
hibited a fivefold increase in cell binding and uptake in
HeLa cells compared to A549 cells. When compared with
Figure 4. Cell viability of: a) A549, and b) HeLa cells incubated with Dox
and GNP–Dox after X-ray irradiation. The Dox concentration for all ex-
periments was 500 nm. The X-ray experiments were performed by using
the Minishot X-ray cabinet at 160 kV for a single dose of 5 Gy. The data
represent the mean Æstandard deviation of the results from three inde-
pendent experiments; *: p<0.05.
500 nm, Dox induced 11 and 20% cell death in A549 and
HeLa cells, respectively. Both MFGNP-I and -II caused low
cytotoxicity (i.e., only 2–5% cell death) in A549 cells com-
pared with HeLa cells, because the lack of FR reduced the
GNP recognition of A549 cells. MFGNP-I showed a similar
toxic profile (21% cell death) whereas MFGNP-II had
lower cytotoxicity (10% cell death) in HeLa cells. Following
X-ray irradiation, A549 cells treated with either of the
Figure 3. a) Cellular uptake of GNPs was quantitatively determined in
&
&
HeLa ( ; FR positive) and A549 ( ; FR negative) cells by performing
ICP-MS. b) Cellular uptake of MFGNP-I in HeLa cells under different
conditions. The GNP concentration was 50 mgmLÀ1. The data represent
the mean Æstandard deviation of the results from three independent ex-
periments. FA: folic acid, SA: sodium azide.
Chem. Eur. J. 2012, 18, 5501 – 5505
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
5503

B. Yan et al.
Characterization: TEM images of GNPs were taken by using a JEOL
1200 EX transmission electron microscope (JEOL, Tokyo, Japan) at
80 kV. The images were acquired by using an AMT 2k CCD camera. The
dynamic diameter of the GNPs was measured by using the Dynapro
Titan system (Wyatt Technology, Santa Barbara, CA). FTIR spectra were
recorded on a Nicolet 6700 FT-IR spectrometer (Thermo Scientific). The
stock solution (3.0 mL) was dried under vacuum by using GeneVac sol-
vent evaporator. The dry samples were collected for FTIR analysis. Then
z potentials of GNPs were measured in a Malvern Nano Z Zetasizer.
GNPs were suspended in water. Each material was tested three times.
The UV/Vis absorption spectra of the GNPs were obtained with
a Varian 5000 UV/Vis spectrometry (Varian, Santa Clara, CA). All the
spectra had subtracted background with baseline correction by the ab-
sorption of deionized water. The MFGNPs used for characterization and
following cellular uptake experiments did not contain Dox.
GNPs did not show any clear signs of toxicity. However,
with the same dose of X-ray irradiation, HeLa cells treated
with MFGNP-I underwent 85% cell death and those treated
with MFGNP-II only showed 25% cell death (Figure 4).
Our data show that MFGNP-I significantly enhanced cell
death; this was caused by the combination of the therapeu-
tic drug and X-ray irradiation in FR positive cells.
In summary, we have demonstrated an accelerated synthe-
sis of a multifunctionalized drug-delivery nanosystem by
using Ugi MCR. To our knowledge, this is the first report of
the application of MCR in assembling a multifunctionalized
nanosystem. The nanosystem enhanced cancer cell targeting
selectivity and greatly improved cancer cell killing by com-
bining the effects of the therapeutic drug with those of radi-
ation. We expect that this expedited synthesis method will
break the bottleneck for preparation of a wide range of
MFNPs for biomedical applications.
Cell culture: HeLa cells were grown in Eagleꢀs minimum essential
medium (MEM, Invitrogen, IL). A549 cells were grown in RPMI 1640
(Gibco, Invitrogen, IL). Each cell culture medium was supplemented
with fetal bovine serum (FBS, 10%), penicillin (10 UmLÀ1) and strepto-
mycin (10 mgmLÀ1).
TEM experiments with cells: TEM images of cells were taken by using
a
JEOL 1200 EX transmission electron microscope (JEOL, Tokyo,
Japan) at 80 kV. The images were acquired by using an AMT 2k CCD
camera. To obtain the TEM images, both cell lines were cultured in 6-
well plates with a density of 100000 cells per well. Cultures were incubat-
ed at 378C under a humidified atmosphere with CO2 (5%). After 24 h in-
cubation, old medium was removed and fresh medium containing GNPs
(50 mgmLÀ1) was added. The cells were incubated with GNPs for 8 h and
washed three times with cold PBS to remove unbound GNPs. The cells
were fixed with glutaraldehyde (2.5%) in Na-cacodylate buffer (0.1m,
Tousimis Research Corporation) for 30 min at room temperature. The
fixed cells were collected for TEM study.
Experimental Section
Synthesis of ligand I: FA-Et-NH2 (4-1, 20 mg, 0.05 mmol) was dissolved
in MeOH (1 mL). 6-O-(4-Formylphenyl)-b-CD (2, 40 mg, 0.03 mmol) was
dissolved in deionized H2O (1 mL) and was mixed with the solution of
FA-Et-NH2. The mixture was stirred at room temperature for 1 h. Thio-
ctic acid (15 mg, 0.07 mmol) and 1-isocyanocyclohexane (10 mL,
0.07 mmol) were dissolved in MeOH (1 mL) and added to the mixture in
sequence. The resulting solution was stirred at room temperature for
2 days until the completion of the reaction. The mixture was evaporated
under vacuum and the final oil was poured into acetone (20 mL). The
precipitate was collected by centrifugation at 4000 rpm for 5 min and
washed twice with acetone. The crude product was purified by reversed-
phase flash chromatography using 20% acetonitrile/water as eluent to
give ligand I (31 mg, 43.6%). 1H NMR (400 MHz, [D6]DMSO with one
drop of [D2]H2O, 25 8C): d=9.85 (s, 1H), 8.69–8.29 (m, 1H), 7.85 (d, J=
8.5 Hz, 2H), 7.72–7.52 (m, 2H), 7.13 (d, J=8.6 Hz, 2H), 6.96 (d, J=
8.0 Hz, 1H), 6.69–6.52 (m, 1H), 5.79 (m, 1H), 4.95–4.74 (m, 7H), 4.52 (d,
J=31.1 Hz, 1H), 4.27 (d, J=40.2 Hz, 2H), 3.99 (d, J=9.6 Hz, 2H), 3.84–
3.57 (m, 29H), 3.31 (m, 16H), 3.14 (dt, J=24.6, 6.0 Hz, 3H), 2.81 (m,
2H), 2.41 (dd, J=12.7, 6.3 Hz, 2H), 2.20 (t, J=7.2 Hz, 3H), 1.87 (dq, J=
13.4, 6.5 Hz, 3H), 1.74–1.45 (m, 8H), 1.44–1.25 ppm (m, 6H).
Cellular uptake of GNPs: Both cell lines were cultured in 12-well plates
with a density of 50000 cells per well. Cultures were maintained at 378C
under a humidified atmosphere with CO2 (5%). After 24 h incubation,
the cells were washed once with cold PBS, and the solutions of GNPs
(50 mgmLÀ1) were added. The cells were incubated with GNPs for 8 h
and then washed three times with cold PBS to remove unbound GNPs.
The cells were detached with trypsin-EDTA solution (0.25% trypsin,
1 mm EDTA). The detached cells were counted by using Cellometer cell
counter Auto T4 (Nexcelom Bioscience, Lawrence, MA) and then pre-
pared for ICP-MS. Each experiment was repeated three times.
ICP-MS sample preparation and measurements: All ICP-MS measure-
ments were performed on a Varian 820. GNPs were incubated with dif-
ferent cell lines separately, as described above. After detaching and
counting the cells, the resulting cell lysate (100 mL) was digested for 4 h
at 378C by adding Aqua Regia (200 mL). The solution (50 mL) was dilut-
ed to 5.0 mL with a 209Bi internal standard (50 ppb) solution in HNO3
(1.0%) and used for ICP-MS measurements. Cellular uptake experiments
with each GNP were repeated three times, and each replicate was mea-
sured five times by ICP-MS. A series of gold standard solutions (1000,
500, 100, 50, 10, 5, and 1 ppb) with 209Bi internal standard (50 ppb) were
prepared before each measurement. The resulting calibration curve was
used to calculate the gold concentration taken up by the different cell
lines. Two injections of 209Bi internal standard solution in HNO3 were
used to wash the instrument between analyses to remove trace amounts
of gold.
Synthesis of ligand II: Ligand II was synthesized by treatment of 4-2, 6-
O-(4-formylphenyl)-b-CD (2), thioctic acid, and 1-isocyanocyclohexane
according to the procedure described above; yield 56.2%. 1H NMR
(400 MHz, [D4]MeOD, 258C): d=7.27 (d, J=8.1 Hz, 2H), 7.01 (d, J=
8.9 Hz, 2H), 5.86 (s, 1H), 4.96 (m, 7H), 4.57 (m, 7H), 4.26–4.03 (m, 2H),
3.90–3.65 (m, 23H), 3.65–3.37 (m, 15H), 3.25–2.98 (m, 3H), 2.70–2.40 (m,
3H), 1.98–1.58 (m, 8H), 1.47 (s, 9H), 1.46–1.10 ppm (m, 6H).
Synthesis of multifunctionalized GNPs: In a typical experiment, water
(1.0 mL) containing chloroauric acid (15.0 mg, 0.038 mmol) was added to
a solution of either ligand I or II (15.0 mg) in water (8.0 mL). After being
stirred for 15 min at room temperature, NaBH4 (6.0 mg, 0.15 mmol) in
water (6.0 mL) was added to the mixture dropwise. The solution turned
red immediately and was stirred for 4 h at room temperature. HCl (1N)
was added to the reaction mixture dropwise to neutralize the excess
sodium tetrahydroborate until the pH reached 7.0. To remove the free
ligand from the nanoparticles, the reaction mixture was centrifuged at
4000 rpm for 30 min with Millipore centrifugal filters with molecule
weight cut-off 10 k (50 mL tube). The colorless supernatant was decanted
and the solid was dissolved in deionized water (10 mL) and centrifuged
again at 4000 rpm for 30 min. This wash/centrifugation cycle was repeat-
ed five times. After the final washing step, the GNPs were dissolved in
Millipore water (5–10 mL) and used as stock solution. The gold concen-
tration of the stock solution was determined by ICP-MS method.
X-ray irradiation: Cells were cultured in 96-well plates with a density of
5000 cells per well. After 24 h incubation, the cells were washed once
with cold PBS. Dox and GNP–Dox solutions were added with a specified
final Dox concentration (500 nm). The cells were incubated for 8 h and
then washed three times with cold PBS to remove unbound GNPs. Fresh
medium (100 mL) was then added to the plates. The AXR Minishot 160
X-ray cabinet irradiator working with 1 mm aluminum at 160 kV and
3 mA, and yielding a mean dose rate of 27.0 RminÀ1 was used for X-ray
irradiation. The cells were exposed to X-ray irradiation with a total dose
of 5 Gy; this corresponds to an irradiation time of 18.5 min. The XTT
measurements were performed 48 h after X-ray irradiation to test cell vi-
5504
www.chemeurj.org
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2012, 18, 5501 – 5505

Multifunctionalized Therapeutic Nanoparticles
COMMUNICATION
ability. For all experiments, a duplicate 96-well plate treated identically
but without X-ray irradiation was used as control. Each experiment was
performed three times.
[6] Y.-A. Shieh, S.-J. Yang, M.-F. Wei, M.-J. Shieh, ACS Nano 2010, 4,
1433–1442.
[7] H. Zhou, P. Jiao, L. Yang, X. Li, B. Yan, J. Am. Chem. Soc. 2011,
133, 680–682.
[8] P. Jiao, H. Zhou, M. Otto, Q. Mu, L. Li, G. Su, Y. Zhang, E. R.
Butch, S. E. Snyder, G. Jiang, B. Yan, J. Am. Chem. Soc. 2011, 133,
13918–13921.
[9] X. Li, H. Zhou, L. Yang, G. Du, A. S. Pai-Panandiker, X. Huang, B.
Yan, Biomaterials 2011, 32, 2540–2545.
Cytotoxicity assay: Cell proliferation kit II (XTT, Roche Diagnostics Cor-
poration, Indianapolis, IN) was used to evaluate the viability of the cells
after uptake of GNPs and X-ray irradiations. The assay is based on the
cleavage of the water-soluble yellow tetrazolium salt XTT to form an
orange formazan dye by metabolically active cells. The formazan was di-
rectly quantified by using a microplate reader, and correlated with the
number of viable cells. Suspension medium was removed 48 h after the
X-ray irradiation and XTT solution (50 mL, 1 mgmLÀ1, Sigma) and fresh
medium (100 mL) were added to each well and incubated at 378C for 4 h.
The absorbance of all samples was measured at 480 and 650 nm separate-
ly by using a SpectraMax M5 microplate reader (Molecular Devices Sun-
nyvale, CA) and the cell viability was calculated following the standard
procedure. Each experiment was performed three times.
[10] J. Chen, S. Chen, X. Zhao, L. V. Kuznetsova, S. S. Wong, I. Ojima, J.
Am. Chem. Soc. 2008, 130, 16778–16785.
[11] G. Pastorin, W. Wu, S. Wieckowski, J. P. Briand, K. Kostarelos, M.
Prato, A. Bianco, Chem. Commun. 2006, 1182–1184.
[12] W. Wu, S. Wieckowski, G. Pastorin, M. Benincasa, C. Klumpp, J. P.
Briand, R. Gennaro, M. Prato, A. Bianco, Angew. Chem. 2005, 117,
6516–6520; Angew. Chem. Int. Ed. 2005, 44, 6358–6362.
[13] E. Garanger, E. Aikawa, F. Reynolds, R. Weissleder, L. Josephson,
Chem. Commun. 2008, 4792–4794.
[14] X. H. Gao, Y. Y. Cui, R. M. Levenson, L. W. K. Chung, S. M. Nie,
Nat. Biotechnol. 2004, 22, 969–976.
[15] S. Kumar, J. Aaron, K. Sokolov, Nat. Protoc. 2008, 3, 314–320.
[16] M. Liong, J. Lu, M. Kovochich, T. Xia, S. G. Ruehm, A. E. Nel, F.
Tamanoi, J. I. Zink, ACS Nano 2008, 2, 889–896.
[17] R. Weissleder, K. Kelly, E. Y. Sun, T. Shtatland, L. Josephson, Nat.
Biotechnol. 2005, 23, 1418–1423.
[18] M. E. Davis, M. E. Brewster, Nat. Rev. Drug Discovery 2004, 3,
1023–1035.
Acknowledgements
We thank Sharon Frase and Linda Mann for technical assistance, and
Cherise M. Guess for editing assistance. This work was supported by the
National Basic Research Program of China (2010CB933504), National
Natural Science Foundation of China (90913006, 21077068, and
21137002), National Cancer Institute (P30A027165), and the American
Lebanese Syrian Associated Charities (ALSAC).
[19] P. S. Low, A. C. Antony, Adv. Drug Delivery Rev. 2004, 56, 1055–
1058.
[20] R. J. Lee, P. S. Low, J. Biol. Chem. 1994, 269, 3198–3204.
[21] W. Eck, A. I. Nicholson, H. Zentgraf, W. Semmler, S. Bartling, Nano
Lett. 2010, 10, 2318–2322.
Keywords: drug delivery
multifunctionalization · nanoparticles · X-ray irradiation
· multicomponent reactions ·
[22] R. Popovtzer, A. Agrawal, N. A. Kotov, A. Popovtzer, J. Balter,
T. E. Carey, R. Kopelman, Nano Lett. 2008, 8, 4593–4596.
[23] T. Kong, J. Zeng, X. P. Wang, X. Y. Yang, J. Yang, S. McQuarrie, A.
McEwan, W. Roa, J. Chen, J. Z. Xing, Small 2008, 4, 1537–1543.
[24] C. J. Liu, C. H. Wang, S. T. Chen, H. H. Chen, W. H. Leng, C. C.
Chien, C. L. Wang, I. M. Kempson, Y. Hwu, T. C. Lai, M. Hsiao,
C. S. Yang, Y. J. Chen, G. Margaritondo, Phys. Med. Biol. 2010, 55,
931–945.
[1] M. Ferrari, Nat. Rev. Cancer 2005, 5, 161–171.
[2] V. P. Torchilin, Adv. Drug Delivery Rev. 2006, 58, 1532–1555.
[3] T. Kong, J. Zeng, X. Wang, X. Yang, J. Yang, S. McQuarrie, A.
McEwan, W. Roa, J. Chen, J. Z. Xing, Small 2008, 4, 1537–1543.
[4] N. Chattopadhyay, Z. Cai, J.-P. Pignol, B. Keller, E. Lechtman, R.
Bendayan, R. M. Reilly, Mol. Pharm. 2010, 7, 2194–2206.
[5] P. Huang, L. Bao, C. Zhang, J. Lin, T. Luo, D. Yang, M. He, Z. Li,
G. Gao, B. Gao, S. Fu, D. Cui, Biomaterials 2011, 32, 9796–9809.
Received: November 28, 2011
Published online: March 21, 2012
Chem. Eur. J. 2012, 18, 5501 – 5505
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
5505