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DOX-loaded pH-sensitive mesoporous silica nanoparticles coated with PDA and PEG induce pro-death autophagy in breast cancer

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The development of multifunctional nano drug delivery carriers has been one of the most effective and prevailing approaches to overcome drug non-selectivity, low cell uptake efficiency and various side effects of traditional chemotherapy drugs. Herein, we report a novel doxorubicin (DOX)-loaded mesoporous silica nanoparticle (MSN) coated with polydopamine (PDA) and polyethylene glycol (PEG) (MSNs-DOX@PDA-PEG) for the treatment of breast cancer. In this system, PDA functions as a pH-sensitive gatekeeper to control the release of DOX from MSNs in response to pH-stimulus and PEG was further grafted on the surface of PDA to increase the stability and biocompatibility under physiological conditions. The in vitro release results suggested that MSNs-DOX@PDA-PEG exhibits a high sensitivity to low pH. A cellular uptake assay showed a high cellular uptake efficiency of MSNs-DOX@PDA-PEG compared to free DOX. Furthermore, MSNs-DOX@PDA-PEG also demonstrated an improved anti-cancer efficacy compared to free DOX both in vivo and vitro breast cancer experiments. Mechanistic studies revealed that MSNs-DOX@PDA-PEG causes a stronger pro-death autophagy compared to free DOX via inhibition of the AKT-mTOR-p70S6K signaling pathway. Taken in concert, our results suggest that the novel material MSNs-DOX@PDA-PEG may represent a promising nanoformulation for breast cancer treatment.
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DOX-loaded pH-sensitive mesoporous silica
nanoparticles coated with PDA and PEG induce
pro-death autophagy in breast cancer
Yanhong Duo,
a
Yang Li,
d
Changke Chen,
c
Baiyun Liu,
a
Xinyu Wang,*
a
Xiaowei Zeng *
c
and Hongbo Chen*
b
The development of multifunctional nano drug delivery carriers has been one of the most eective and
prevailing approaches to overcome drug non-selectivity, low cell uptake eciency and various side
eects of traditional chemotherapy drugs. Herein, we report a novel doxorubicin (DOX)-loaded
mesoporous silica nanoparticle (MSN) coated with polydopamine (PDA) and polyethylene glycol (PEG)
(MSNs-DOX@PDA-PEG) for the treatment of breast cancer. In this system, PDA functions as a pH-
sensitive gatekeeper to control the release of DOX from MSNs in response to pH-stimulus and PEG was
further grafted on the surface of PDA to increase the stability and biocompatibility under physiological
conditions. The in vitro release results suggested that MSNs-DOX@PDA-PEG exhibits a high sensitivity to
low pH. A cellular uptake assay showed a high cellular uptake eciency of MSNs-DOX@PDA-PEG
compared to free DOX. Furthermore, MSNs-DOX@PDA-PEG also demonstrated an improved anti-
cancer ecacy compared to free DOX both in vivo and vitro breast cancer experiments. Mechanistic
studies revealed that MSNs-DOX@PDA-PEG causes a stronger pro-death autophagy compared to free
DOX via inhibition of the AKT-mTOR-p70S6K signaling pathway. Taken in concert, our results suggest
that the novel material MSNs-DOX@PDA-PEG may represent a promising nanoformulation for breast
cancer treatment.
Introduction
Breast cancer represents a leading cause of death among
women.
1
Presently, chemotherapy is one of the main available
treatment options for breast cancer. However, traditional
chemotherapeutic treatments oen result in drug resistance
and frequently cause harmful side eects to non-cancerous
tissues due to the lack of selective tumor targeting.
2
Doxorubicin (DOX) is one of the most ecient anticancer
drugs that is widely used for the treatment of multiple cancer
types including breast, bladder, lung, hematological malig-
nancies, and others.
3
As an anthracycline antibiotic, DOX can
interfere with DNA synthesis, induce DNA damages, produce
reactive oxygen species, and destroy membrane structures.
4,5
Recently, the development of a safe and ecient drug delivery
system to deliver DOX to the tumor site attracted a signicant
amount of attention in the scientic community. With the
development of nanotechnology, nanoformulations have been
widely employed for the delivery of anticancer drugs. However,
drug release from polymer-based nanoparticles takes place
mainly through diusion and/or polymer degradation.
6
The
latter represents a slow process and the therapeutic drug levels
may therefore not promptly reach an eective drug concentra-
tion. Recently, mesoporous silica nanoparticles (MSNs) have
attracted enormous attention in the eld of drug delivery due to
their unique physiochemical properties, including large surface
area and pore volume, tunable particle/pore size, high drug
loading eciency, easy surface modication and remarkable
stability and biocompatibility.
7,8
By modifying the outer surface
of MSNs with various functional groups such as polymers,
9
supramolecules,
10
quantum dots,
11
ligands,
12
or/and by using
a combination with other nanomaterials,
13,14
stimuli-responsive
and active targeting nanosystems can be designed for the tar-
geted delivery of anticancer drugs.
In this study, we developed DOX-loaded mesoporous silica
nanoparticles (MSN) and the nanoparticle surface was further
coated with polydopamine (PDA) and polyethylene glycol (PEG).
a
Key Laboratory of Plant Cell Activities and Stress Adaptation, Ministry of Education,
School of Life Sciences, Lanzhou University, Lanzhou 730000, P. R. China. E-mail:
wangxy@lzu.edu.cn
b
School of Pharmaceutical Sciences (Shenzhen), Sun Yat-sen University, Guangzhou
510275, P. R. China. E-mail: chenhb7@mail.sysu.edu.cn
c
The Shenzhen Key Lab of Gene and Antibody Therapy, Division of Life and Health
Sciences, Graduate School at Shenzhen, Tsinghua University, Shenzhen 518055, P.
R. China. E-mail: zeng.xiaowei@sz.tsinghua.edu.cn
d
Department of Hepatobiliary and Pancreas Surgery, Second Clinical Medical College
of Jinan University, Shenzhen People's Hospital, Shenzhen 518000, P. R. China
Electronic supplementary information (ESI) available. See DOI:
10.1039/c7ra05135b
These authors contribute equally to this work.
Cite this: RSC Adv.,2017,7, 39641
Received 7th May 2017
Accepted 1st August 2017
DOI: 10.1039/c7ra05135b
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In this system, PDA coating not only protects DOX leakage
under physiological conditions (pH 7.4) but also allows for the
sustained-release of the drugs in an acidic environment
(pH 5.0).
15,16
PEG on the surface of MSNs ensures water solu-
bility and further prevents nonspecic interactions with bio-
macromolecules.
17
Previous reports published in the literature
show that the polymer-based nanoparticles may be taken up by
cells through the endocytosis-lysosome pathway.
18,19
Thus, we
anticipated that DOX may be rapidly released when MSNs-
DOX@PDA-PEG enters to lysosomes where the pH value is
about 5.020. Furthermore, our results suggested that MSNs-
DOX@PDA-PEG exhibits improved anti-cancer abilities
compared to free DOX both in vitro and in vivo. Furthermore,
mechanistic studies revealed that MSNs-DOX@PDA-PEG may
cause a stronger pro-death autophagy compared to free DOX via
the AKT-mTOR-p70s6K signaling pathway.
Results and discussion
Synthesis and physicochemical characterization of MSNs-
DOX@PDA-PEG
MSNs-DOX@PDA-PEG was synthesized as described below in
the section Materials and methods.Theparticlesizeand
surface properties of the nanoparticlesplayimportantrolesin
drug release, cellular uptake and pharmacokinetics.
21
Fig. 1A
shows a schematic representation of the MSNs-DOX@PDA-
PEG synthesis. In order to access the morphology of MSNs,
MSNs-DOX@PDA and MSNs-DOX@PDA-PEG, TEM analysis
was performed. Fig. 1B, C and D show representative images
of MSNs, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG,
respectively. As shown in Fig. 1B, the MSNs exhibit a nearly
spherical shape and porous surfaces. Compared with MSNs
(cf. Fig. 1B), MSNs-DOX@PDA (cf. Fig. 1C) and MSNs-
DOX@PDA-PEG (cf. Fig. 1D) show the PDA and PEG coating
on the MSNs surface, with a clear layer found on the periphery
of the particles.
The sizes of MSNs, MSNs-DOX, MSNs-DOX@PDA and MSNs-
DOX@PDA-PEG were determined using dynamic light scattering
(DLS). As shown in Table 1, the diameters of the MSNs, MSNs-
DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG were 125.34
4.42, 130.21 3.37, 170.32 2.49 and 198.75 2.56 nm,
respectively. Although the sizes of MSNs-DOX@PDA and MSNs-
DOX@PDA-PEG were slightly larger than MSNs, they were also
found to be smaller than the cut-osize of tumor neovasculature
pores,
22,23
falling within the range of easy accumulation of the
enhanced permeation and retention (EPR) eect.
The zeta potentials were also determined as shown in Table
1. The zeta potentials of MSNs, MSNs-DOX, MSNs-DOX@PDA
and MSNs-DOX@PDA-PEG were 19.43 5.21, 8.56 2.19,
12.44 3.29 and 2.11 0.97 mV, respectively. Since most
cellular membranes are negatively charged, the zeta potential
can aect the tendency of the nanoparticles to permeate
membranes, with cationic particles exhibiting a higher toxicity
with cell membrane disruption. In general, nanoparticles with
a zeta potential between 10 and +10 mV are considered
neutral. The specic surface area, pore volume and the most
probable pore size of MSNs, MSNs-DOX, MSNs-DOX@PDA and
MSNs-DOX@PDA-PEG are also shown in Table 1. Compared to
MSNs and MSNs-DOX, all pore parameters of MSNs-DOX@PDA
and MSNs-DOX@PDA-PEG were found to be signicantly
decreased. The BET surface area was 264.72 m
2
g
1
, the pore
volume was 0.62 cm
3
g
1
and, as evaluated by the BJH method,
the most probable pore size was about 2.89 nm. Moreover, the
pore size distribution of MSNs was rather narrow. With loading
of DOX, the BET surface area and the most probable pore size
decreased to 121.32 m
2
g
1
and 2.31 nm, respectively. The BET
surface area of MSNs-DOX@PDA and MSNs-DOX@PDA-PEG
was 46.73 and 49.78 m
2
g
1
, respectively. However, the pore
Fig. 1 (A) Schematic representation of MSNs-DOX@PDA-PEG synthesis. (B) TEM image of MSNs. (C) TEM image of MSNs-DOX@PDA. (D) TEM
image of MSNs-DOX@PDA-PEG.
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Table 1 Characterization parameters of MSNs, MSNs-DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG
Polymer Size
a
(nm) ZP (mV)
BET surface
area (m
2
g
1
)
Pore volume
b
(cm
3
g
1
)
Pore size
c
(nm)
MSNs 125.34 4.42 19.43 5.21 264.72 0.62 2.89
MSNs-DOX 130.21 3.37 8.56 2.19 121.32 0.43 2.31
MSNs-DOX@PDA 170.32 4.49 12.44 3.29 46.73 0.15
MSNs-DOX@PDA-PEG 198.75 2.56 2.11 0.97 49.78 0.11
a
NPs size was measured by dynamic light scattering.
b
BJH cumulative pore volume for pores between 1.7 and 300 nm in width.
c
Most probable
pore size.
Fig. 2 (A) In vitro release prole of MSNs-DOX (B) in vitro release prole of MSNs-DOX@PDA-PEG.
Fig. 3 (A) Uptake of free DOX and MSNs-DOX@PDA-PEG detected by confocal microscopy in MCF10A cells (red: DOX; blue: DAPI, scale bar ¼
10 mm). (B) Same treatment as (A) but using MCF7 cells. (C) Flow cytometry (a) free DOX in MCF10A, (b) free DOX in MCF7, (c) MSNs-DOX@PDA-
PEG in MCF10A and (d) MSNs-DOX@PDA-PEG in MCF7 (a black, b blue, c rose red, d red).
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volumes were 0.15 and 0.11 cm
3
g
1
due to the coating of PDA
and PEG onto the surface of DOX-loaded MSNs. Overall, the
structural parameters of MSNs, MSNs-DOX, MSNs-DOX@PDA
and MSNs-DOX@PDA-PEG suggested that DOX occupied the
pore space of MSNs and DOX loaded MSNs were coated with
PDA and PEG. The encapsulation eciency of DOX in MSNs was
about 95.63 2.32.
In addition, as a result of the slightly negative charge of MSNs-
DOX@PDA-PEG, the overall clearance by the reticuloendothelial
system (RES) such as liver was found to be reduced.
24,25
These
data suggested that MSNs-DOX could be successfully synthesized
and modied by introduction of PDA and PEG lms.
A pH-sensitive drug release prole in vitro
Previously published research data shows that the polymer-
based nanoparticles are taken up by cells through the
endocytosis-lysosome pathway.
18,19
The pH value of lysosomes is
about 5.0, which is maintained by proton pumps on the lyso-
some membrane.
20
In addition, low pH conditions are also
considered a hallmark of malignant solid tumor tissues. In
order to verify the drug blocking potency and pH sensitivity of
PDA and PEG coating, we performed a drug release experiment
at pH 5.0 and pH 7.4 at dierent time intervals. As shown in
Fig. 2A, without PDA and PEG coating, DOX was quickly
released at all tested pH values, freely diusing from the pores
Fig. 4 (A, B) Viability of MCF7 and MDA-MB-231 cells cultured with MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG,
respectively, with dierent NPs concentrations after 24 h. Data were expressed as mean SD (*p< 0.05, **p< 0.01). (C, D) Cell proliferation
evaluation of MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG by MTT assay after treatments for 12, 24 and 48 h in
MCF7 and MDA-MB-231 cells, respectively. Data were expressed as mean SD (*p< 0.05, **p< 0.01). (E) MTT assay of normal breast cell line
MCF10A, similar treatment as C and D. (F) BrdU incorporation assay by confocal microscopy (green: BrdU; blue: DAPI, scale bar ¼10 mm). (G)
Clone formation evaluation of MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG.
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of MSNs into solution. However, compared to the rapid release
of MSNs-DOX at pH 7.4, MSNs-DOX@PDA-PEG exhibited
a sustained DOX release rate at pH 7.4, suggesting that PDA-
PEG coating eectively protects DOX leakage from the MSNs
(cf. Fig. 2B). In particular, MSNs-DOX@PDA-PEG exhibited
a signicantly more rapid release rate at pH 5.0 compared to the
slower release rate at pH 7.4 (cf. Fig. 2B), a nding that could be
attributed to the pH sensitivity of PDA under acidic
conditions.
26
Eective internalization of MSNs-DOX@PDA-PEG by cells
To determine the specic recognition and uptake capacity of
MSNs-DOX@PDA-PEG towards target cells, we carried out
Fig. 5 (A) MCF7 cells were treated with PBS, MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG for 12 h and autophagic
vehicles were observed. The cells were imaged under a confocal microscope (blue: DAPI, red: DOX, green: LC3, scale bar ¼10 mm). (B) MCF7
cells were treated similarly to (A) and Western blotting was performed with LC3, beclin 1, p62 and b-actin antibodies. (C) MCF7 cells were treated
similarly to (A) and Western blotting was performed with the indicated antibodies.
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confocal microscopy analysis to compare the cellular uptake
eciencies of free DOX and MSNs-DOX@PDA-PEG (Fig. 3A and
B). The results suggested that cellular uptake of MSNs-
DOX@PDA-PEG in MCF7 cells was signicantly higher than
that of free DOX. Furthermore, cellular uptake of MSNs-
DOX@PDA-PEG in MCF7 cells was remarkably higher than in
MCF10A cells. Flow cytometry analysis also conrmed the
enhanced cellular uptake of MSNs-DOX@PDA-PEG in MCF7
cells (Fig. 3C). Most likely, the increase of cellular uptake was
caused by PDA and PEG. Previous studies have reported similar
results in terms of PDA and PEG being able to increase the
cellular uptake of nanoparticles.
21,27
Inhibition of cell growth and proliferation by MSNs-
DOX@PDA-PEG
MCF7 and MDA-MB-231 cells (2.6 10
4
) were used to study the
in vitro cytotoxicity of MSNs-DOX@PDA-PEG. Fig. 4A and B
show the in vitro cell viability of the drug formulated in MSNs-
DOX@PDA-PEG and DOX at equivalent concentrations of 1, 5,
10, 25 and 35 mgmL
1
, respectively. The percentage of viable
cells was quantitatively assessed by an MTT method. MSNs or
MSNs@PDA-PEG did not exhibit a signicant cytotoxicity
against MCF7 and MDA-MB-231 cells in vitro. As reported
previously, extremely high concentrations of MSNs (about
25 mg mL
1
) at some sizes exhibit cytotoxicity.
28
However,
the required maximum concentration of MSNs herein was
500 mgmL
1
, a concentration at which nearly no cytotoxicity
could be observed. Similarly to our result, PDA and PEG coating
was found to be nontoxic in various cell models and in various
in vivo studies.
29
MCF7 and MDA-MB-231 cells (2.6 10
4
) treated with
10 mgmL
1
MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and
MSNs-DOX@PDA-PEG for 12, 24 and 48 h, respectively. As
shown in Fig. 4C and D, MSNs@PDA-PEG exhibited no signi-
cant eect on the cell viability compared with the PBS control,
providing further evidence that MSNs@PDA-PEG represents
a safe and biocompatible nanocarrier. Both free DOX and
MSNs-DOX@PDA exhibited a signicant cell growth inhibition
while the MSNs-DOX@PDA-PEG demonstrated a stronger
inhibition ability to MCF7 and MDA-MB-231 compared to free
DOX and MSNs-DOX@PDA. However, for the normal breast cell
line MCF10A, both free DOX and MSNs-DOX@PDA exhibited
almost no cell growth inhibition as shown in Fig. 4E.
5-Fluorouridine (BrdU), a thymidine analogue, has been
crucial in the identication of DNA synthesis and the assess-
ment of cell proliferation. In the present research, a BrdU assay
Fig. 6 (A) Tumor growth curves of MCF7 xenografts treated with saline, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG, respectively. (B)
Xenograft tumors were removed and (C) weighted. Values are provided as mean standard error (*p< 0.05, **p< 0.01). (D) Representative H&E
staining of xenograft tumors.
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was performed. The MCF7 cells (6 10
6
) were treated with
10 mgmL
1
MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and
MSNs-DOX@PDA-PEG for 6 h and the BrdU incorporation was
detected by confocal microscopy. As shown in Fig. 4F, MSNs-
DOX@PDA-PEG exhibited the best inhibition eect on BrdU
incorporation.
A colony formation assay was also performed as described
previously.
30
MCF7 cells (1 10
3
) were treated with 10 mgmL
1
MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and MSNs-
DOX@PDA-PEG for 7 days and the obtained colonies were
stained with 0.1% crystal violet. As shown in Fig. 4G, similar to
the MTT assay, MSNs-DOX@PDA-PEG exhibited a stronger
inhibition eect on the colony formation of MCF7 cells
compared to other treatments.
Induction of pro-death autophagy through suppression of
AKT-mTOR-p70S6K pathway by MSNs-DOX@PDA-PEG
Previously conducted research studies have shown that DOX can
induce pro-death autophagy.
31
Autophagy primarily represents
a degradation pathway that clears malfunctioning cellular
components, including intracellular pathogens, in response to
various types of stress. LC3 proteins play a critical role in auto-
phagy. Beclin 1 is considered to be related to the initiation and
progression of autophagy, albeit the underlying mechanism
remains unknown. The autophagy receptor and substrate
SQSTM1/p62 inhibits the E3 ligase ubiquitination of histone in
response to DNA double-strand breaks. Dysregulation of this
process leads to a reduced ability to repair DNA. Some studies
have shown evidence for the accumulation of beclin 1 and LC3B-
II, but degradation of p62. Meanwhile, the percentage of cells
with characteristic LC3B-GFP puncta structure, a characteristic
marker of autophagy, was increased following some autophagy
induced drug treatments.
32
In the present study, we investigated
the eects of MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and
MSNs-DOX@PDA-PEG on autophagy in MCF7 cells. As shown in
Fig. 5A, the number of autophagic vesicles (referred to LC3 dotted
green uorescence) was signicantly higher in MSNs-DOX@PDA-
PEG-treated cells compared to the other groups. Consistently,
MSNs-DOX@PDA-PEG treatment induced an increase in LC3-II
and p62 levels, however, a decrease in beclin 1 level in MCF7
cells was also observed (cf. Fig. 5B).
The AKT/mammalian target of rapamycin (mTOR)/p70 ribo-
somal protein S6 kinase (p70S6K) signaling pathway is known to
regulate autophagy.
33
To investigate whether MSNs-DOX@PDA-
PEG-induced autophagy is involved in the AKT-mTOR-p70S6K
signaling pathway, the phosphorylation levels of the associated
proteins were studied by Western blotting. As shown in Fig. 5C,
all of the treatments with free DOX, MSNs-DOX@PDA and MSNs-
DOX@PDA-PEG signicantly inhibited the phosphorylation
levels of AKT, mTOR, and p70S6K, however, MSNs-DOX@PDA-
PEG exhibited the best inhibition eect.
Signicant inhibition of the MCF7 subcutaneous xenogra
tumor growth by MSNs-DOX@PDA-PEG
The anti-tumor ecacy of MSNs-DOX@PDA-PEG was further
investigated in nude mice bearing subcutaneous MCF7 tumors.
The administration of free DOX, MSNs-DOX@PDA and MSNs-
DOX@PDA-PEG resulted in signicant growth suppression of
MCF7 xenogras compared to the PBS control group. As shown
in Fig. 6A, tumor growth in the treatment groups was
Fig. 7 (A) Body weight changes of mice treated with saline, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG. Data is expressed as mean
value standard error. (B) Representative H&E staining of heart, liver, spleen, lung, kidney.
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signicantly slower than that in the PBS-control group. MSNs-
DOX@PDA-PEG demonstrated the best inhibition ability on
the xenogragrowth. At the end of the experiment, the average
tumor weight in the treatment groups was signicantly lower
than that in the PBS-control group and the MSNs-DOX@PDA-
PEG group featured the lowest tumor weight (cf. Fig. 6B and
C). Furthermore, an immunohistochemical study showed that
the tumor tissues from the treatment of MSNs-DOX@PDA-PEG
exhibited the fewest tumor cells and the highest level of tumor
necrosis compared to the other treatments (cf. Fig. 6D), indi-
cating a higher anti-tumor activity of MSNs-DOX@PDA-PEG.
The latter nding is presumably due to the EPR eect and the
controlled release at tumor sites.
The systematic toxicity of MSNs-DOX@PDA-PEG in vivo was
evaluated by body weight monitoring and H&E tissue section
staining of major organs including heart, liver, spleen, lung,
and kidney. No statistically signicant dierences in body
weight could be observed between the MSNs-DOX@PDA-PEG-
treated group and other groups (p> 0.05, data not shown) (cf.
Fig. 7A) and no noticeable histopathological abnormalities (cf.
Fig. 7B) could be observed in MSNs-DOX@PDA-PEG groups
suggesting that MSNs-DOX@PDA-PEG featured a good
biocompatibility and a low general toxicity in vivo.
Conclusions
In summary, a pH-sensitive drug delivery system involving
mesoporous silica nanoparticles coated with PDA and PEG was
successfully designed for the controlled release of cationic
amphiphilic drug DOX. Furthermore, MSNs-DOX@PDA-PEG
exhibited a good anti-cancer ecacy through the induction of
pro-death autophagy. The obtained results demonstrate that
the DOX-loaded MSNs-DOX@PDA-PEG featured competitive
advantages, including an excellent pH-sensitivity, suitable
cellular uptake and therapeutic ecacy with low side eects (cf.
Fig. 8).
Materials and methods
Materials
3-Mercaptopropyltrimethoxysilane (MPTMS, 95%), cetyl-
trimethyl ammonium bromide (CTAB), tetraethylorthosilicate
(TEOS), amino-terminated poly(ethylene glycol) (H
2
N-PEG,
M
w
¼2000), hydrochloride dopamine and doxorubicin (DOX)
were purchased from J&K Scientic Ltd (Beijing, China).
Ammonium uoride (NH
4
F) was obtained from Aladdin
Industrial Co., Ltd. (Shanghai, China). Acetonitrile was
purchased from EM Science (HPLC grade, Mallinckrodt Baker,
USA). Dulbecco's modied eagle medium (DMEM), trypsin
EDTA solution (0.25%), fetal bovine serum (FBS) and penicillin-
streptomycin were obtained from GIBCO, Invitrogen Co.
(Carlsbad, NM, USA). 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl
tetrazolium bromide (MTT) and bovine serum albumin (BSA)
were purchased from Amresco (Solon, OH, USA). 5-Fluorour-
idine (BrdU) was obtained from Sigma-Aldrich (St. Louis, MO,
USA). 6-Diamidino-2-phenylindole (DAPI) was obtained from
Biyuntian Co., Ltd (Nanjing, China). BrdU mouse monoclonal
antibodies were purchased from Abcam (Cambridge, MA, USA).
LC3 antibody (rabbit source) was purchased from Cell Signaling
Technology (Beverly, MA, USA). P62 antibody was purchased
from Abmart Inc. (Shanghai, China). Beclin 1 was purchased
from Cell Signaling Technology (Beverly, MA, USA) AKT, p-AKT,
mTOR, p-mTOR, p70s6k, p-p70s6k antibodies were purchased
from Santa Cruz (Santa Cruz, CA, USA). Human breast carci-
noma cell line MCF7, MDA-MB-231 and the normal breast cell
line MCF10A were purchased from American Type Culture
Collection (ATCC). Water used throughout the studies was ob-
tained from an ultrapure MilliQ water purication system
(resistance > 18 MUcm). All other chemicals of the highest
available quality were commercially obtained and used as
received.
Synthesis of MSNs
MSNs were synthesized according to our previously reported
procedure with slight modication.
34
Briey, 1.82 g CTAB
(5 mM) and 3 g NH
4
F (81 mM) were dissolved in 500 mL of water
and heated to 80 C in a 1000 mL ask. Under vigorous stirring,
9 mL (8.41 g) TEOS was added dropwise to the mixture and the
temperature was then kept at 80 C for 6 h. The solid product
was centrifuged (12 000 rpm, 10 min), washed with water and
ethanol three times and dried at 40 Cin vacuo. To remove the
surfactant template (CTAB), the product was dispersed in 400
mL of ethanol containing 8 mL of hydrochloric acid (37%) and
reuxed at 80 C for 24 h. This procedure was repeated twice to
ensure that the surfactant was completely removed. The ob-
tained MSNs were centrifuged and washed with deionized
water.
Preparation of MSNs-DOX@PDA-PEG
For DOX loading, 50 mg MSNs were added to a water solution
containing doxorubicin hydrochloride (2.5 mL, 10 mg mL
1
)
and the mixture was stirred for 24 h. The solution was centri-
fuged and washed with water to move the remaining DOX from
Fig. 8 Schematic illustration of the pH-sensitive release of DOX from
MSNs-DOX@PDA-PEG triggering pro-death cell autophagy.
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the surface of MSNs. DOX-loaded MSNs (MSNs-DOX) were dried
at 40 Cin vacuo. Polydopamine-coated NPs (MSNs-DOX@PDA)
were synthesized by incubating 50 mg of MSNs-DOX in
0.5 mg mL
1
dopamine hydrochloride in Tris buer (10 mM,
pH 8.5) for 6 h at room temperature with shaking. Then, MSNs-
DOX@PDA was centrifuged (12 000 rpm, 10 min) and washed
with water to remove any unpolymerized dopamine. The
mixture was then added to 2.5 mg NH
2
-PEG and the resulting
material was stirred for 3 h in the dark at room temperature.
PEG was employed to coat MSNs in order to ensure high
physiological stability.
35
Finally, PEG and PDA-coated MSNs-
DOX (MSNs-DOX@PDA-PEG) were centrifuged, washed three
times with water and dried at 40 Cin vacuo.
The sample was dropped onto a copper grid coated with
a carbon membrane and allowed to dry. Then, the surface
morphology of MSNs-DOX@PDA-PEG was observed by trans-
mission electron microscopy (TEM, Tecnai G2 20, FEI
Company, USA). Drug release from MSNs-DOX@PDA-PEG was
determined as described previously.
36,37
Cell uptake of MSNs-DOX@PDA-PEG
MCF7 cells were cultured in DMEM supplemented with 10% (v/v)
FBS and antibiotics (100 U mL
1
penicillin and 100 mgmL
1
streptomycin) in a humidied 5% CO
2
atmosphere at 37 C.
MCF7 cells were incubated with free DOX and MSNs-DOX@PDA-
PEG (equal quantity of DOX) at 37 C for 0.5 h, washed with cold
PBS three times, and then xed by 4% paraformaldehyde for
20 min. Then, the cells were washed with PBS, stained with 4,6-
diamidino-2-phenylindole (DAPI) for 15 min and observed by
confocal laser scanning microscopy (CLSM, Olympus Fluoview
FV-1000, Japan) with imaging soware. The images of the cells
were determined with dierential interference contrast channel,
blue channel (DAPI) excitation at 358 nm and red channel (DOX)
excitation at 543 nm.
38,39
Immunouorescence observations
MCF7 cells seeded on coverslips were treated with MSNs@PDA-
PEG, free DOX, MSNs-DOX@PDA and MSNs-DOX@PDA-PEG,
respectively. Cells were xed with 4% formaldehyde in PBS for
15 min and then permeabilized in PBS containing 0.1% Triton
X100 for another 7 min. The cells were blocked with 3% BSA for
2 h at RT and probed with an appropriate primary antibody
overnight at 4 C. The coverslips were then incubated with
rhodamine- and uorescein-conjugated secondary antibodies
for 2 h at RT. Aer staining with 0.5 mgmL
1
of DAPI for 10 min,
the cells were observed and imaged using an Olympus FV1000
confocal microscope.
MTT cell proliferation assay
Briey, 1 10
5
MCF7 cells per well seeded in triplicate into a 96-
well plate were treated with certain concentrations of free DOX,
MSNs-DOX@PDA and MSNs-DOX@PDA-PEG. At the appro-
priate time-points, 20 mL MTT (5 mg mL
1
) was added for 4 h at
37 C and all liquid was carefully removed. Optical density (OD)
values were measured with a spectrophotometer at 490 nm
following continuous agitation for 20 min with 130 mL DMSO.
Colony formation assay
MCF7 cells were seeded into a six-well plate (1 10
3
cells per
well) and cultured in DMEM with 10% bovine serum albumin.
MCF7 cells were treated with MSNs@PDA-PEG, free DOX,
MSNs-DOX@PDA and MSNs-DOX@PDA-PEG, respectively.
Aer 7 days, the resulting colonies were stained with 0.01% of
crystal violet and images were captured.
BrdU cell proliferation assay
MCF7 cells seeded in 12-well plates were treated with
MSNs@PDA-PEG, free DOX, MSNs-DOX@PDA and MSNs-
DOX@PDA-PEG for 6 h, respectively. Cells were incubated
with 200 mM 5-uorouridine for 30 min. The medium was
removed and the cells were xed with 4% formaldehyde in PBS
for 15 min and permeabilized in PBS containing 0.1% Triton
X100 for 7 min. The cells were probed with a BrdU antibody
aer blocking with 3% BSA and assessed for BrdU incorpora-
tion by confocal microscopy.
Western blotting
The cells were lysed with lysis buer and equal amounts of
proteins were separated by 10% SDS-PAGE and transferred onto
a PVDF membrane. The membrane was blocked with 5% non-
fat milk in Tris-buered saline containing Tween 20 for 2 h at
room temperature and incubated overnight at 4 C with specic
primary antibodies. Aer washing for a total of three times with
TBST, the membrane was incubated with appropriate HRP-
linked secondary antibodies for 2 h at room temperature and
then detected with ECL reagent.
Xenogratumor growth assay
This study was performed in strict accordance with the NIH
guidelines for the care and use of laboratory animals (NIH
Publication no. 85-23 Rev. 1985) and was approved by the
Institutional Animal Care and Use Committee of Tsinghua
University. 45 week old female nude mice were purchased from
the Medical Experimental Animal Centre of Guangdong Prov-
ince. All animal experiments were approved by the Institutional
Animal Care and Use Committee of Tsinghua University. The
mice were housed in a specic-pathogen-free environment
maintained at 25 1C with 55% relative humidity and food as
well as water were provided. The mice were randomly allocated
into 4 groups with 4 animals per group. MCF7 cells (5 10
5
in
150 mL PBS) in sub-conuent condition were subcutaneously
injected into the mice. Six days aer tumor inoculation, the
mice were administered a daily intraperitoneal injection of DOX
(0.5 mg kg
1
). Tumor volumes were measured every three
days with a caliper and calculated according to the formula: V¼
1/2(LW
2
), where Land Wrepresent the length and width,
respectively. All mice were sacriced 2030 days aer tumor
inoculation and the tumors were excised and weighed.
Statistical analysis
Values are expressed as the mean standard deviation of three
independent experiments. Comparisons were performed using
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a two-tailed paired Student's t-test. Dierences with p< 0.05
were considered signicant.
Conicts of interest
The authors declare no competing nancial interest.
Acknowledgements
This research was supported by the National Natural Science
Foundation of China (No. 81670141), Guangdong Natural
Science Foundation (No. 2014A030313758) and Science, Tech-
nology & Innovation Commission of Shenzhen Municipality
(No. JCYJ20160422170206664).
References
1 R. L. Siegel, K. D. Miller, S. A. Fedewa, D. J. Ahnen,
R. G. Meester, A. Barzi and A. Jemal, Ca-Cancer J. Clin.,
2017, 67, 177193.
2 M. A. Firer and G. Gellerman, J. Hematol. Oncol., 2012, 5, 70.
3 Y. Oh, J.-O. Jin and J. Oh, Nanotechnology, 2017, 28, 125101.
4 O. Tacar, P. Sriamornsak and C. R. Dass, J. Pharm.
Pharmacol., 2013, 65, 157170.
5 A. K. Jain, N. K. Swarnakar, M. Das, C. Godugu, R. P. Singh,
P. R. Rao and S. Jain, Mol. Pharm., 2011, 8, 11401151.
6 H. K. Makadia and S. J. Siegel, Polymers, 2011, 3, 13771397.
7 F. Tang, L. Li and D. Chen, Adv. Mater., 2012, 24, 15041534.
8 Y. Chen, X. Deng, C. Li, F. He, B. Liu, Z. Hou, Z. Cheng,
B. Xing and J. Lin, Nanomedicine: Nanotechnology, Biology
and Medicine, 2017, 13, 875883.
9 Y. Guo, J. Sun, S. Bai and X. Jin, J. Biomater. Appl., 2016,
0885328216653287.
10 N. Song and Y.-W. Yang, Chem. Soc. Rev., 2015, 44, 3474
3504.
11 X. Yao, X. Niu, K. Ma, P. Huang, J. Grothe, S. Kaskel and
Y. Zhu, Small, 2017, 13.
12 N. Li, N. Li, Q. Yi, K. Luo, C. Guo, D. Pan and Z. Gu,
Biomaterials, 2014, 35, 95299545.
13 W. Fang, J. Yang, J. Gong and N. Zheng, Adv. Funct. Mater.,
2012, 22, 842848.
14 W. Gao, W. Cao, Y. Sun, X. Wei, K. Xu, H. Zhang and B. Tang,
Biomaterials, 2015, 69, 212221.
15 W. Tao, X. Zeng, J. Wu, X. Zhu, X. Yu, X. Zhang, J. Zhang,
G. Liu and L. Mei, Theranostics, 2016, 6, 470.
16 J. Hu, X. Zhang, Z. Wen, Y. Tan, N. Huang, S. Cheng,
H. Zheng and Y. Cheng, Oncotarget, 2016, 7, 73681.
17 L. Palanikumar, E. S. Choi, J. Y. Cheon, S. H. Joo and
J. H. Ryu, Adv. Funct. Mater., 2015, 25, 957965.
18 M. Costanzo, F. Carton, A. Marengo, G. Berlier, B. Stella,
S. Arpicco and M. Malatesta, Eur. J. Histochem., 2016, 60.
19 X. Zhang, Y. Dong, X. Zeng, X. Liang, X. Li, W. Tao, H. Chen,
Y. Jiang, L. Mei and S.-S. Feng, Biomaterials, 2014, 35, 1932
1943.
20 T. S. Hnasko and R. H. Edwards, Annu. Rev. Physiol., 2012, 74,
225243.
21 X. Zhu, Y. Xu, L. M. Solis, W. Tao, L. Wang, C. Behrens, X. Xu,
L. Zhao, D. Liu and J. Wu, Proc. Natl. Acad. Sci. U. S. A., 2015,
112, 77797784.
22 C. Zhang, W. Wang, T. Liu, Y. Wu, H. Guo, P. Wang, Q. Tian,
Y. Wang and Z. Yuan, Biomaterials, 2012, 33, 21872196.
23 S. K. Hobbs, W. L. Monsky, F. Yuan, W. G. Roberts,
L. Grith, V. P. Torchilin and R. K. Jain, Proc. Natl. Acad.
Sci. U. S. A., 1998, 95, 46074612.
24 W. Ngamcherdtrakul, J. Morry, S. Gu, D. J. Castro,
S. M. Goodyear, T. Sangvanich, M. M. Reda, R. Lee,
S. A. Mihelic and B. L. Beckman, Adv. Funct. Mater., 2015,
25, 26462659.
25 T. Li, X. Shen, Y. Geng, Z. Chen, L. Li, S. Li, H. Yang, C. Wu,
H. Zeng and Y. Liu, ACS Appl. Mater. Interfaces, 2016, 8,
1374813758.
26 M. Zhang, J. Liu, Y. Kuang, Q. Li, D.-W. Zheng, Q. Song,
H. Chen, X. Chen, Y. Xu and C. Li, Int. J. Biol. Macromol.,
2017, 98, 691700.
27 Q. Zheng, T. Lin, H. Wu, L. Guo, P. Ye, Y. Hao, Q. Guo,
J. Jiang, F. Fu and G. Chen, Int. J. Pharm., 2014, 463,2226.
28 Q. He, Z. Zhang, Y. Gao, J. Shi and Y. Li, Small, 2009, 5, 2722
2729.
29 J. Park, T. F. Brust, H. J. Lee, S. C. Lee, V. J. Watts and Y. Yeo,
ACS Nano, 2014, 8, 33473356.
30 H. Chen, Y. Duo, B. Hu, Z. Wang, F. Zhang, H. Tsai, J. Zhang,
L. Zhou, L. Wang and X. Wang, Oncotarget, 2016, 7, 78747.
31 B. Sivaraman, G. Swaminathan, L. Moore, J. Fox, D. Seshadri,
S. Dahal, I. Stoilov, M. Zborowski, R. Mecham and
A. Ramamurthi, Acta Biomater., 2017, 52, 171186.
32 Q. Zhang, M. Yang, Z. Qu, J. Zhou and Q. Jiang, Biochem.
Biophys. Res. Commun., 2016, 480, 334340.
33 B. Cao, J. Li, X. Zhou, J. Juan, K. Han, Z. Zhang, Y. Kong,
J. Wang and X. Mao, Sci. Rep., 2014, 4, 5749.
34 D. Chang, Y. Gao, L. Wang, G. Liu, Y. Chen, T. Wang, W. Tao,
L. Mei, L. Huang and X. Zeng, J. Colloid Interface Sci., 2016,
463, 279287.
35 Y. Cui, H. Dong, X. Cai, D. Wang and Y. Li, ACS Appl. Mater.
Interfaces, 2012, 4, 31773183.
36 Z. Zhang and S.-S. Feng, Biomaterials, 2006, 27, 40254033.
37 X. Liang, Y. Yang, L. Wang, X. Zhu, X. Zeng, X. Wu, H. Chen,
X. Zhang and L. Mei, J. Mater. Chem. B, 2015, 3, 93839396.
38 M. Zheng, C. Yue, Y. Ma, P. Gong, P. Zhao, C. Zheng,
Z. Sheng, P. Zhang, Z. Wang and L. Cai, ACS Nano, 2013, 7,
20562067.
39 C. Zheng, J. Xu, X. Yao, J. Xu and L. Qiu, J. Colloid Interface
Sci., 2011, 355, 374382.
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