ArticlePDF Available

Tumor‐exocytosed exosome/AIEgen hybrid nano‐vesicles facilitate efficient tumor penetration and photodynamic therapy

Authors:

Abstract and Figures

The development of novel photosensitizing agents with aggregation‐induced emission (AIE) properties have fueled significant advances in the field of photodynamic therapy (PDT). Herein, electroporation method was used to prepare tumor‐exocytosed exosome/AIEgen hybrid nano‐vesicles (termed DES) that could facilitate efficient tumor penetration. Dexamethasone was then used to normalize vascular function within the TME to reduce local hypoxia, thereby significantly enhancing the PDT efficacy of DES nano‐vesicles, allowing them to effectively inhibit tumor growth. We achieved the hybridization of AIEgen and biological tumor‐exocytosed exosomes for the first time, and combine PDT approaches with normalizing the intratumoral vasculature as a means of reducing local tissue hypoxia. Together, this work highlights a new valuable approach to design AIEgen based PDT systems and underscores the potential clinical value of AIEgens.
Content may be subject to copyright.
Anticancer Drugs Hot Paper
Tumor-Exocytosed Exosome/Aggregation-Induced Emission
Luminogen Hybrid Nanovesicles Facilitate Efficient Tumor
Penetration and Photodynamic Therapy
Daoming Zhu+, Yanhong Duo+, Meng Suo, Yonghua Zhao, Ligang Xia, Zheng Zheng, Yang Li,*
and Ben Zhong Tang*
Abstract: The development of novel photosensitizing agents
with aggregation-induced emission (AIE) properties has fueled
significant advances in the field of photodynamic therapy
(PDT). An electroporation method was used to prepare tumor-
exocytosed exosome/AIE luminogen (AIEgen) hybrid nano-
vesicles (DES) that could facilitate efficient tumor penetration.
Dexamethasone was then used to normalize vascular function
within the tumor microenvironment (TME) to reduce local
hypoxia, thereby significantly enhancing the PDT efficacy of
DES nanovesicles, and allowing them to effectively inhibit
tumor growth. The hybridization of AIEgen and biological
tumor-exocytosed exosomes was achieved for the first time,
and combined with PDT approaches by normalizing the
intratumoral vasculature as a means of reducing local tissue
hypoxia. This work highlights a new approach to the design of
AIEgen-based PDT systems and underscores the potential
clinical value of AIEgens.
Introduction
Photodynamic therapy (PDT) is a clinical strategy where-
in the exposure of photosensitizer (PS) compounds to
particular wavelengths of light in the presence of oxygen
(O2) results in the production of cytotoxic free radicals and
reactive oxygen species (ROS), such as singlet oxygen. PDT
can be used as an effective means of treating superficial or
localized tumors and other diseases.[1] The development of PS
agents with aggregation-induced emission (AIE) properties
has enabled the implementation of novel PDT strategies.
AIE-based fluorophores exhibit minimal emissivity when
they exist as isolated molecules in solution, whereas when
they exist as aggregates they produce significantly more
intense emissions.[2] AIE luminogens (AIEgens) are ideal
tools for bioimaging applications, as they are highly photo-
stable, biocompatible, and allow for high-contrast imaging.[3]
Importantly, many AIEgens can readily generate ROS, and
they can be utilized for PDT-based therapeutic interven-
tions.[4] Unfortunately, the majority of AIE PS compounds are
very hydrophobic and therefore cannot readily applied within
in vivo biological contexts. Novel AIEgen-based hybrid
systems, such as lipid nanoparticles, have the potential to
facilitate the direct intratumoral delivery of AIE PS com-
pounds following their encapsulation within the hydrophobic
core of these particles.[5] For example, in one study research-
ers were able to produce AIEgen–lipid conjugates that
formed particles termed “AIEsomes” which could more
efficiently generate ROS and facilitate PDT in vivo.[6] These
previously described AIEgen-based systems, however, are
limited by their poor drug loading content (DLC), low
encapsulation efficiency (EE), limited ability to specifically
target tumors, and poor target tissue permeability.[7] There-
fore, there is a clear need for the development of novel
AIEgen-based platforms as a means of more effectively
penetrating tumors. In addition to the aforementioned
limitations, the hypoxic TME that is common in solid tumors
can compromise the efficacy of PDT owing to a lack of O2,
which is essential for the efficacy of such therapeutic
strategies. Intratumoral hypoxia typically results from unre-
strained tumor cell growth coupled with disorganized and
dysregulated angiogenic activity.[8] This hypoxia can be
further aggravated by PDT-induced microvascular collapse,
further compromising effective tumor treatment. This results
in a negative feedback cycle wherein PDT approaches, which
are already hampered by a poor O2supply, rapidly exhaust
and disrupt the remaining local O2levels so as to significantly
[*] Dr. D. Zhu,[+] Prof. L. Xia, Dr. Y. Li
Department of Gastrointestinal Surgery, Second Clinical Medical
College of Jinan University, Shenzhen People’s Hospital
Shenzhen 518020 (China)
E-mail: 1028168734@qq.com
Dr. Z. Zheng, Prof. B. Z. Tang
Department of Chemistry, Hong Kong Branch of Chinese National
Engineering Research Center for Tissue Restoration and Recon-
struction, Institute for Advanced Study, Department of Chemical and
Biological Engineering and Division of Life Science
The Hong Kong University of Science and Technology (HKUST)
Clear Water Bay, Kowloon, Hong Kong (China)
E-mail: tangbenz@ust.hk
Dr. D. Zhu,[+] Dr. M. Suo
Department of Electronic Science and Technology
School of Physics and Technology, Wuhan University
Wuhan 430072 (China)
Dr. Y. Duo[+]
Department of Microbiology, Tumor and Cell Biology
Karolinska Institute
17177 Stockholm (Sweden)
Prof. Y. Zhao
State Key Laboratory of Quality Research in Chinese Medicine
University of Macau (China)
[
+
] These authors contributed equally to this work.
Supporting information and the ORCID identification number(s) for
the author(s) of this article can be found under:
https://doi.org/10.1002/anie.202003672.
A
ngewandte
Chemi
e
Research Articles
How to cite:
International Edition: doi.org/10.1002/anie.202003672
German Edition: doi.org/10.1002/ange.202003672
&&&&
 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2020,59,210
Ü
Ü
These are not the final page numbers!
impair therapeutic efficacy.[9] This vicious feedback cycle is
highly likely to negatively impact AIE-based PDT
approaches.
Tumor-derived exosomes (EXO) represent a potentially
ideal means of enhancing AIE PS tumor penetration and
efficacy. EXO are small (50–200 nm) particles that are
secreted from cells and that are derived from multivesicular
bodies.[10] Such EXO can be readily collected from biological
samples and the supernatants of cultured cells using com-
mercially available kits.[11] EXO have many properties that
make them promising candidates for drug delivery
approaches, including a lack of immunogenicity, excellent
biocompatibility, and the ability to remain in circulation for
extended periods of time owing to their endogenous origin.[12]
These EXO are also able to cross the blood–brain barrier[13]
and to penetrate deep within structural or otherwise dense
tissue types.[14] Importantly, EXO can be internalized by cells,
and can home to specific tissues or cell types in a specific
manner based on EXO membrane- and cell surface–specific
protein profiles.[11,15] As EXO increases the solubility of
compounds that are otherwise not soluble in aqueous
solutions, they are ideal for clinical drug-delivery applica-
tions.[16] The development of an efficient means to produce
EXO/AIEgen hybrid nanovesicles therefore has the potential
to render PDT treatment of cancer more efficient.
A number of approaches for overcoming intratumoral
hypoxia and enhancing PDT efficacy have been proposed to
date, such as the utilization of catalases, MnO2, or nanoscale
metal–organic frameworks (MOFs).[7e] Preclinical research
suggests that reducing vascular leakage without pruning can
increase intratumoral oxygen delivery.[17] Several different
drugs are effective in blood normalization applications.[18] For
example, the glucocorticosteroid dexamethasone (DEX),
which is commonly administered to patients undergoing
radiotherapy and/or chemotherapy so as to reduce associated
toxicity, can mediate vascular and extracellular matrix (ECM)
normalization. DEX is therefore also able to reduce levels of
intratumoral hypoxia through similar mechanisms.[19] The
combination of EXO/AIEgen hybrid nanovesicles with
vascular normalizing agents may therefore offer a better
PDT than the nanovesicles alone.
Herein, a previously studied AIE PS called (E)-4-(2-(7-
(diphenylamino)-9-ethyl-9H-carbazol-2-yl)vinyl)-1-methyl-
pyridin-1-ium hexafluorophosphate (DCPy) was used.[1a] An
electroporation method was employed to prepare tumor-
exocytosed EXO/AIEgen hybrid nanovesicles (DES) that
could facilitate efficient tumor penetration. DEX was then
used to normalize vascular function within the TME to reduce
local hypoxia, thereby significantly enhancing the PDT
efficacy of DES, and allowing them to effectively inhibit
tumor growth (Scheme 1). The combination of DEX with the
EXO/AIEgen hybrid nanovesicles described herein has the
potential to overcome many of the traditional limitations of
AIE-based PDT strategies, enhancing ROS generation and
improving overall therapeutic efficacy. The hybridization of
EXO/AIEgen nanovesicles, designed by us, have realized
efficient PDT with intratumoral vasculature normalization
therapy, and also highlight a novel AIEgen-based biomimetic
PDT system that has the potential to expedite the clinical
implementation of AIEgens for the treatment of cancer
patients.
Results and Discussion
Preparation and characterization of DES
DCPy was prepared as per our previous study and its AIE
properties were thoroughly discussed in the prior analysis.[1a]
DCPy were purified in excellent yields of 75–85 %. All
Scheme 1. An illustration of DES facilitating efficient tumor penetration and PDT.
A
ngewandte
Chemi
e
Research Articles
&&&&
Angew. Chem. Int. Ed. 2020,59, 2 10  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
Ü
Ü
intermediates and products were fully characterized by
nuclear magnetic resonance (NMR) spectroscopy and with
high-resolution mass spectrometry (Supporting Information,
Figures S1–S3). A commercial isolation kit was then used to
isolate EXO from 4T1 mammary tumor cell supernatants.
Isolated EXO were small (ca. 150 nm) and had a saucer-like
shape (Figure 1A). All of these particles were <500 nm in
size, and electroporation did not significantly alter the
morphology of these particles (Figure 1B). We have also
prepared DES from other types of tumor cells in a similar way
(Figure S4). To confirm successful DCPy coupling in these
DES, which was stained using the membrane fluorescent
probe 3,3-dioctadecyloxacarbocyanine perchlorate (DiO).
When these nanovesicles were analyzed by confocal micro-
scopy, both bright green DiO fluorescence and red DCPy
fluorescence was observed (Figure 1C), confirming that
DCPy had been coupled into these particles. The UV/Vis
absorption spectra of DES nanovesicles exhibited an absorp-
tion peak characteristic of DCPy at 452 nm (Figure 1 D),
which was consistent with the color change of phosphate
buffered saline (PBS) solutions after DCPy coupling. To-
gether, these results all supported the fact that DES nano-
vesicles were successfully coupled with DCPy. Based on
standard curves (Figures S6 and S7), DCPy electroporation
efficiency values were calculated under different initial DCPy
concentrations (Figure S9A), revealing that packaging effi-
ciency slowly rose as starting DCPy concentrations increased,
until reaching a maximum value of 88.2 %—much higher than
for commonly used polymer nanoparticles.[3a,7d,g] The zeta-
potential values of EXO, DCPy, and DES nanovesicles were
25.6 3.1, 28.6 2.8, and 12.8 3.6 mV, respectively (Fig-
ure 1 E). No significant changes in the size or zeta potential of
DES nanovesicles was seen following 3 days of storage in PBS
at 4
8
C, suggesting that these particles are highly stable
(Figures 1F,G). Western blotting further confirmed that these
DES nanovesicles were phenotypically similar to EXO, as
they expressed exosomal biomarkers, including CD9 and
CD63 (Figure 1I). Analyses of photoluminescence (PL)
spectra revealed that the fluorescence of DES nanovesicles
(in PBS) and DCPy (in aggregate in a 90% water–dimethyl
sulfoxide (DMSO) solution) peaked at 614 nm and 698 nm,
respectively (Figure 1H), and poor fluorescence intensity was
observed in a 100% DMSO DCPy solution. This indicated
the AIE properties of DCPy and the PL properties of DCPy
were blue-shifted after DES formation, indicating successful
preparation of hybrid nanovesicles.
DES targets cancer cells in vitro
Owing to the fact that they were prepared using tumor
cells, DES nanovesicles were adept at being internalized by
target tumor cells. Such internalization can be readily traced
owing to the fact that DCPy was able to specifically stain
mitochondria in live cells following its internalization.[1a] To
confirm the ability of DES nanovesicles to be internalized by
tumor cells in vitro, a co-localization experiment wherein 4T1
cells were both incubated with DES nanovesicles and stained
using the commercial mitochondrial MitoTracker Green FM
probe (Figures 2 A,E). After a 30 minute incubation period,
DES nanovesicles were attached to 4T1 cells, and after
2 hours the DCPy within these DES nanovesicles had stained
Figure 1. TEM images of A) EXO and B) DES nanovesicles. C) DiO (green) and DCPy (red) co-localization within DES nanovesicles was assessed
by confocal microscopy. Scale bar=5mm. D) UV/Vis spectra for DCPy, EXO, and DES nanovesicles in PBS, with the inset image showing the
color of DCPy (in DMSO), EXO (in PBS), and DES nanovesicles (in PBS). E) Zeta potential values for EXO, DCPy, and DES nanovesicles. F) DLS
was used to measure the hydrodynamic diameter of EXO and DES nanovesicles. G) The zeta potential of DES nanovesicles suspended in PBS
was assessed after 1, 2, and 3 days. H) PL spectra of DCPy aggregate suspension with a 10% DMSO fraction, 100% DMSO fraction, and a DES
PBS solution. Inset: fluorescence photograph of DCPy taken under 365 nm UV irradiation. I) Exosomal markers, including CD9 (i) and CD63 (ii),
were detected by Western blotting.
A
ngewandte
Chemi
e
Research Articles
&&&&
www.angewandte.org  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2020,59,210
Ü
Ü
These are not the final page numbers!
tumor cell mitochondria. These results confirm that DES are
an ideal platform for targeted tumor cells and mitochondria.
According to the “Trojan exosome hypothesis”, EXO uptake
appears to involve clathrin-mediated endocytosis followed by
back fusion of EXO with the limiting membrane of the
endosome.[20] Subsequently, the mitochondria-specific target-
ing capability of the cationic lipophilic DCPy relies primarily
on the driven force of a very large membrane potential of
around 180 mV across the mitochondrial membrane.[1a] These
provide a possible theoretical basis for the tumor-targeting
ability of DES.
Assessment of in vitro PDT efficacy and biocompatibility of DES
particle
The efficient 1O2generation of these AIEgens and DES is
desirable for realizing satisfactory PDT. We therefore
assessed 1O2generation capability using 9,10-anthracene-
diyl-bis(methylene)dimalonic acid (ABDA), which can un-
dergo oxidation by 1O2to yield endoperoxide, resulting in
a decrease of ABDA absorption. Under white-light irradi-
ation (Figure S8), the absorbance of ABDA solution in the
presence of DCPy and DES decreases gradually with
prolonged irradiation time, whereas a prodigious drop of
ABDA absorption was observed for DCPy and DES under
0.5 Wcm2irradiation dose. Under laser irradiation, DCPy
can be rapidly released from DES, indicating that ROS
produced by DCPy can destroy the DES structure (Fig-
ure S9B). The impact of DES nanovesicles on cell viability
was further evaluated, revealing that they were satisfactorily
biocompatible even at high DCPy concentrations (Fig-
ure S10). Subsequently, the PDTefficacy of DES nanovesicles
and DCPy was evaluated under both normal and hypoxic
conditions (Figure 2 B). Under normoxic conditions, DES or
DCPy treatment led to substantial phototoxicity in response
to appropriate laser irradiation, with viability being reduced
to just 7% in response to a 20 mgmL1DCPy concentration.
Under hypoxic conditions, however, this efficacy was mark-
edly reduced, with cells remaining 50% viable following
treatment with 20 mgmL1DCPy and subsequent laser
irradiation. Since DCPy fluorescence can interfere with
propidium iodide (PI) staining, we used Fluorescein diacetate
(FDA) single staining to observe the survival of tumor cells
(Figure S11A). The results are consistent with those of MTT
assays. When a 2,7-dichlorodihydrofluorescein diacetate
(DCFH-DA) probe was used to quantify ROS production in
these cells, strong green fluorescence consistent with robust
ROS generation under normoxic conditions was observed
following 532 nm laser irradiation (Figures 2C,D). Under
hypoxia-mimicking conditions, however, this fluorescence
intensity was markedly reduced, confirming that oxygen
levels can profoundly shape the efficacy of DES- or DCPy-
mediated antitumor PDT treatment regimens. Figure S11B
shows the opening of mitochondrial permeability transition
pores (MPTP) of 4T1 cancer cells after applying various
formulations monitored by calcein fluorescence in mitochon-
dria. The images showed a direct evidence of the opening of
MPTP through observation of the fluorescence of calcein in
mitochondria. The fluorescence in mitochondria showed the
strongest intensity after applying mitochondrial targeting
DCPy and DES under normoxic condition, as compared to
that after application under hypoxic conditions. This result
indicates that DES can cause mitochondrial damage, and the
degree of damage is related to oxygen concentration.
Dexamethasone alleviates tumor hypoxia
Given that oxygen levels can profoundly alter DES-
mediated PDT efficacy, intratumoral hypoxia is likely to limit
the therapeutic utility of this therapeutic strategy. In an effort
Figure 2. A) Co-localization of MitoTracker Green FM (green) and DCPy (red) or DES over time in 4T1 tumor cells. Scale bar =10 mm.
B) Relative tumor cell viability following PBS, DCPy, or DES treatment and subsequent 532 nm laser irradiation (0.5 Wcm2, 5 min) under normal
or hypoxic conditions, as assessed by an MTT assay. *P<0.05, **P<0.01, ***P<0.005; Student’s t-test. C) Tumor cell fluorescence images and
D) DCFH-DA fluorescence intensity following the indicated treatments. Scale bar =20 mm. *P<0.05, **P<0.01, ***P<0.005; Student’s t-test.
E) DCPy fluorescence intensity in A was quantified using the ImageJ software.
A
ngewandte
Chemi
e
Research Articles
&&&&
Angew. Chem. Int. Ed. 2020,59, 2 10  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
Ü
Ü
to overcome this limitation, DEX was therefore utilized as
a TME-modulating agent as a means of improving the
available oxygen supply within these tumors. To confirm the
efficacy of this approach, BALB/c mice bearing subcutaneous
4T1 tumors were treated with DEX (3 mgkg1) or PBS
(n=3/group) by intraperitoneal injection once per day from
days 11 to 14 post-tumor implantation. DEX dosage was fixed
at 3 mg kg1since this dosage was the lowest dose capable of
reducing interstitial fluid pressure (IFP).[19a] The results
indicated that DEX treatment significantly increased micro-
vessel density and the density of vessels positive for the
pericyte marker a-smooth muscle actin (aSMA+; Figure 3A;
Figures S12 and S13). Most importantly, hypoxia-inducible
factor (HIF)-1 was used as a marker of tumor hypoxia to
assess the antihypoxic efficacy of DEX treatment (Figure 3 B;
Figure S14). The results showed that DEX treatment signifi-
cantly reduced the HIF-1 staining intensity relative to PBS
treatment, confirming that this therapeutic intervention was
sufficient to markedly alleviate intratumoral hypoxia.
Assessment of in vivo pharmacokinetics, biodistribution, and
tumor tissue permeability of DES
In addition to being highly cytotoxic and readily internal-
ized by tumor cells, ideal antitumor platforms need to have
favorable systemic biodistribution profiles.[21] To highlight the
in vivo properties of DES for biodistribution and pharmaco-
kinetics, poly(lactic-co-glycolic) acid (PLGA) was used
to package DCPy and we have successfully prepared
PLGA/DCPy hybrid nanoparticles (DPS) for comparison
with DES. PLGA has been approved by the US Food and
Drug Administration and has been widely studied. The
obtained DPS nanoparticles present a well-defined spherical
shape and homogenous sizes, as revealed by transmission
electron microscopy (TEM; Figure S5A). The average hydro-
dynamic size of DPS was approximately 100 nm, as measured
by dynamic light scattering (DLS; Figure S5B). The UV/Vis
absorption spectrum of DPS showed the characteristic
absorption peak of DCPy, indicating the successful encapsu-
lation of DCPy in the PLGA core (Figure S5C). DCPy
loading efficiency was 13.8 2.6% in our experiments,
measured by UV/Vis spectrophotometry. As expected, DES
shows longer circulation time than DPS (Figure S15). The
DES and DPS biodistribution in 4T1 tumor-bearing mice
(n=3/group) were assessed. At 12, 24, and 48 hours post-
injection, an IVIS small animal imaging system with a Cy5
channel was used to visualize biodistribution profiles in the
animals. Furthermore, another group of mice were sacrificed
after 12 hours and their tumor and major organs were
collected and sampled for tissue-specific imaging (Figure 3 C).
Relatively strong DES and DPS accumulation was observed
within the liver, and DES exhibited more effective tumor
tropism in animals than DPS (Figure 3D). The proteins
within the membranes of tumor-derived EXO generally
govern their efficient homing to and internalization in target
tissues, with recent work suggesting that such protein profiles
can also modulate cargo permeability.[22] This may explain
Figure 3. A) Representative cluster of differentiation 31 (CD31; red) and a-smooth muscle actin (aSMA; green) immunofluorescent images
following treatment with PBS and DEX. B) Representative HIF-1aimmunofluorescent images following treatment with PBS and DEX. C) In vivo
fluorescence imaging and imaging of major organs at 48 h post-injection in 4T1 tumor-bearing mice. The white dotted circle indicates a tumor.
D) Quantification of DCPy tissue biodistribution at 12 h post-injection of DCPy+DEX, DES nanovesicles, DPS +DEX, or DES+DEX at a DCPy
equivalent dose of 5 mgkg1. E) DCPy (red) co-localization with CD31-labeled endothelial cells (green) in tumor tissue sections prepared from
4T1 tumor-bearing mice at 12 h post-treatment. F) DCPy distribution profiles in blood vessels and tumor tissues along the while line highlighted
in (E).
A
ngewandte
Chemi
e
Research Articles
&&&&
www.angewandte.org  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2020,59,210
Ü
Ü
These are not the final page numbers!
why DEX did not significantly alter biodistribution profiles in
these mice. This is a new discovery. According to previous
articles, the accumulation of nanoparticles in tumor sites will
significantly increase after tumor blood vessel normalization,
but this is limited to those nanoparticles without active
targeting ability. There have been no reports on the effect of
tumor vessel normalization on nanoparticles with active
targeting ability. Notably, the free DCPy is not suitable for
direct intravenous injection as it is highly hydrophobic, and
for that reason intraperitoneal injection was employed in the
in vivo experiments. Notably, DCPy fluorescence was detect-
able approximately 200 mm from blood vessels following
DES-mediated treatment (Figures 3E,F), indicating that
DES nanovesicles can readily extravasate from the vascula-
ture so as to penetrate deep into the tumor tissue with or
without DEX. After intraperitoneal injection of pure DCPy,
there was no obvious imaging signal in the tumor tissue within
48 hours, and the content of DCPy in the tumor tissue was far
lower than that of DES, DES +DEX, or the DPS +DEX
group. Additionally, there was no obvious DCPy fluorescence
near and far from tumor blood vessels. This result indicates
the poor tumor-targeting ability of pure DCPy. According to
previous reports,[23] nanoparticles or small molecules with
positive charge or hydrophobicity are not suitable for direct
injection since they tend to bind to plasma proteins, so DES
hybrid nanovesicles are a good improvement on DCPy in
PDT.
Evaluation of in vivo PDT efficacy of DES
To extend these results into an in vivo model system and
more fully explore the PDT efficacy of DES nanovesicles with
or without DEX co-treatment, mice bearing 4T1 breast
cancer tumors were used. These animals were randomly
assigned to six treatment groups: 1) control (PBS), 2) laser (L,
532 nm laser irradiation, 0.5 Wcm2, 20 min and we chose
four points of exposure, each of which was 5 min), 3) DEX,
4) DES, 5) DES+L, and 6) DES +DEX +L groups (Fig-
ure 4A). The combination of DES administration with laser
irradiation mediated a partial inhibition of tumor growth,
whereas this inhibition was sufficiently more potent when
animals were also treated with DEX (Figures 4 B,C; Fig-
ure S16). This suggests that DEX-mediated modulation of the
TME was sufficient to enhance PDT efficacy and overall
therapeutic benefit in these animals. Hematoxylin and eosin
(H&E) stained tissue sections further confirmed that DES +
DEX +L treatment was associated with significant tumor
tissue loss that coincided with extensive necrotic or apoptotic
tumor cell death (Figure 4 D). DCFH-DA was used to
measure intratumoral ROS production in these treated mice,
revealing significantly enhanced staining in the tumors of
mice administered with the DES +DEX +L combination
treatment. This confirms that the enhanced efficacy observed
in these animals coincided with improved free radical
generation consistent with enhanced PDT activity. There is
no significant change in murine body weight in response to
treatment over the 14 day study period, suggesting that these
treatments were associated with minimal systemic toxicity
(Figure S17). This is important, as many nanoplatforms and
medical materials have been limited by associated systemic
toxicity when administered in vivo.[24] To confirm that DES
nanovesicles did not induce any systemic toxicity, the gross
histology of H&E-stained heart, liver, spleen, lung, and
kidney sections from these animals was evaluated. The results
revealed no significant abnormalities in mice in any treatment
groups at any point during the study period (Figure S18).
There was no impairment of liver or kidney function (Fig-
ure S19). In addition, we monitored the temperature changes
of tumor sites in mice during treatment. As shown in the
Figure S19, there was no significant temperature increase,
indicating that the above treatment effect was not generated
by heat. Moreover, this dose of laser irradiation did not
damage the skin (Figures S16 and S20). DES treatment was
thus not associated with any significant adverse events in this
murine model system.
Conclusion
In summary, we developed tumor-exocytosed EXO/AIE-
gen hybrid nanovesicles, called DES. DES were readily able
to penetrate tumors in vivo, and were ideal for PDT
applications when used in combination with DEX as a means
of normalizing the vasculature in the hypoxic TME. The
results of the present study are highly novel for several
reasons. For one, this nanosystem overcomes many limita-
tions associated with the use of AIEgens in PDT. In addition,
a novel combination of these biomimetic nanovesicles with
Figure 4. A) Overview of the DEX+DES+L experimental design.
B) Changes in tumor volume over time in response to the indicated
treatments. *P<0.05, **P<0.01, ***P<0.005; Student’s t-test.
C) Average tumor weight values following the indicated treatments.
*P<0.05, **P<0.01, ***P<0.005; Student’s t-test. D) H&E stained
tumor sections from the indicated treatment groups and ROS mea-
surement by DCFH-DA staining in tumor sections. Scale bar =100 mm.
A
ngewandte
Chemi
e
Research Articles
&&&&
Angew. Chem. Int. Ed. 2020,59, 2 10  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
Ü
Ü
DEX-mediated tumor vascular normalization therapy as
a means of enhancing intratumoral ROS generation upon
laser irradiation, thereby enhancing PDT performance. This
work highlights a novel approach to tumor PDT and
emphasizes the potential clinical use of AIEgens. In the
future, we will make use of these hybrid vesicles in the
application of the patient-derived xenograft model, which will
bring us much closer to clinical trials.
Female BALB/c mice aged 4–5 weeks were purchased
from Vital River Company (Beijing, China). 4T1 cell
suspensions (100 mL; 1106 cells per mL) were subcutane-
ously injected into each mouse to establish the tumor models.
The animal experiments were carried out according to the
protocol approved by the Ministry of Health in Peoples
Republic of P. R. China (document no. 55, 2001) and were
approved by the Administrative Committee on Animal
Research of the second clinical Medicine College of Jinan
University.
Acknowledgements
This work was supported by the National Natural Science
Foundation of China (81901771) and the Science, Technology
& Innovation Commission of Shenzhen Municipality
(JCYJ20190807144209381). The author would like to thank
Fei Huang from Shiyanjia Lab (www.shiyanjia.com) and
Shiyanjia Lab for drawing illustrations.
Conflict of interest
The authors declare no conflict of interest.
Keywords: dexamethasone · exosome/
AIEgen hybrid nanovesicles · photodynamic therapy ·
tumor penetration · tumor vascular normalization
[1] a) Z. Zheng, T. Zhang, H. Liu, Y. Chen, R. T. K. Kwok, C. Ma, P.
Zhang, H. H. Y. Sung, I. D. Williams, J. W. Y. Lam, K. S. Wong,
B. Z. Tang, ACS Nano 2018,12, 8145 –8159; b) W. Li, J. Yang, L.
Luo, M. Jiang, B. Qin, H. Yin, C. Zhu, X. Yuan, J. Zhang, Z. Luo,
Y. Du, Q. Li, Y. Lou, Y. Qiu, J. You, Nat. Commun. 2019,10,
3349; c) D. Xia, P. Xu, X. Luo, J. Zhu, H. Gu, D. Huo, Y. Hu,
Adv. Funct. Mater. 2019,29, 1807294; d) L. Wang, X. Zhang, X.
Yu, F. Gao, Z. Shen, X.Zhang, S. Ge, J. Liu, Z. Gu, C. Chen, Adv.
Mater. 2019,31, 1901965; e) Z. Yang, J. Wang, S. Liu, X. Li, L.
Miao, B. Yang, C. Zhang, J. He, S. Ai, W. Guan, Biomaterials
2020,229, 119580; f) M. Qiu, D. Wang, W. Liang, L. Liu, Y.
Zhang, X. Chen, D. K. Sang, C. Xing, Z. Li, B. Dong, F. Xing, D.
Fan, S. Bao, H. Zhang, Y. Cao, Proc. Natl. Acad. Sci. USA 2018,
115, 501 506; g) J. Liu, T. Liu, P. Du, L. Zhang, J. Lei, Angew.
Chem. Int. Ed. 2019,58, 7808 – 7812 ; Angew. Chem. 2019,131,
7890 –7894 ; h) T. Guo, Y. Wu, Y. Lin, X. Xu, H. Lian, G. Huang,
J.-Z. Liu, X. Wu, H.-H. Yang, Small 2018,14, 1702815; i) W. L.
Liu, T. Liu, M. Z. Zou, W. Y. Yu, C. X. Li, Z. Y. He, M. K.
Zhang, M. D. Liu, Z. H. Li, J. Feng, X. Z. Zhang, Adv. Mater.
2018,30, 1802006.
[2] a) J. Mei, N. L. C. Leung, R. T. K. Kwok, J. W. Y. Lam, B. Z.
Tang, Chem. Rev. 2015,115, 11718–11940 ; b) J. Luo, Z. Xie,
J. W. Y. Lam, L. Cheng, H. Chen, C. Qiu, H. S. Kwok, X. Zhan,
Y. Liu, D. Zhu, B. Z. Tang, Chem. Commun. 2001, 1740 – 1741;
c) M. Yang, D. Xu, W. Xi, L. Wang, J. Zheng, J. Huang, J. Zhang,
H. Zhou, J. Wu, Y. Tian, J. Org. Chem. 2013,78, 10344; d) Y. Cao,
M. Yang, Y. Wang, H. Zhou, J. Zheng, X. Zhang, J. Wu, Y. Tian,
Z. Wu, J. Mater. Chem. C 2014,2, 3686; e) J. F. Lovell, T. W. B.
Liu, J. Chen,G. Zheng, Chem. Rev. 2010,110, 2839– 2857; f) T. J.
Dougherty, C. J. Gomer, B. W. Henderson, G. Jori, D. Kessel, M.
Korbelik, J. Moan, Q. Peng, J. Natl. Cancer Inst. 1998,90, 889 –
905.
[3] a) D. Wang, B. Z. Tang, Acc. Chem. Res. 2019,52, 2559 – 2570 ;
b) Y. Hong, P. Zhang, H. Wang, M. Yu, Y. Gao, J. Chen, Sens.
Actuators B 2018,272, 340 – 347.
[4] a) F. Hu, S. Xu, B. Liu, Adv. Mater. 2018,30, 1801350; b) Y. Gao,
X. Wang, X. He, Z. He, X. Yang, S. Tian, F. Meng, D. Ding, L.
Luo, B. Z. Tang, Adv. Funct. Mater. 2019,29, 1902673; c) D.
Wang, M. M. S. Lee, G. Shan, R. T. K. Kwok, J. W. Y. Lam, H.
Su, Y. Cai, B. Z. Tang, Adv. Mater. 2018,30, 1802105.
[5] a) Z. Sheng, B. Guo, D. Hu, S. Xu, W. Wu, W. H. Liew, K. Yao, J.
Jiang, C. Liu, H. Zheng, B. Liu, Adv. Mater. 2018,30, 1800766;
b) B. Gu, W. Wu, G. Xu, G. Feng, F. Yin, P. H. J. Chong, J. Qu, K.-
T. Yong, B. Liu, Adv. Mater. 2017,29, 1701076; c) D. Wang,
M. M. S. Lee, W. Xu, R. T. K. Kwok, J. W. Y. Lam, B. Z. Tang,
Theranostics 2018,8, 4925 – 4956.
[6] X. Cai, D. Mao, C. Wang, D. Kong, X. Cheng, B. Liu, Angew.
Chem. Int. Ed. 2018,57, 16396 – 16400 ; Angew. Chem. 2018,130,
16634 – 16638.
[7] a) J. Qi, C. Sun, A. Zebibula, H. Zhang, R. T. K. Kwok, X. Zhao,
W. Xi, J. W. Y. Lam, J. Qian, B. Z. Tang, Adv. Mater. 2018,30,
1706856; b) Y. Yang, L. Wang, H. Cao, Q. Li, Y. Li, M. Han, H.
Wang, J. Li, Nano Lett. 2019,19, 18211826; c) W. Wu, D. Mao,
F. Hu, S. Xu, C. Chen, C.-J. Zhang, X. Cheng, Y. Yuan, D. Ding,
D. Kong, B. Liu, Adv. Mater. 2017,29, 1700548; d) Y. Li, Q. Wu,
M. Kang, N. Song, D. Wang, B. Z. Tang, Biomaterials 2020,232,
119749; e) L. Shi, F. Hu, Y. Duan, W. Wu, J. Dong, X. Meng, X.
Zhu, B. Liu, ACS Nano 2020,14, 2183 2190; f) C. Y. Y. Yu, H.
Xu, S. Ji, R. T. K. Kwok, J. W. Y. Lam, X. Li, S. Krishnan, D.
Ding, B. Z. Tang, Adv. Mater. 2017,29, 1606167; g) J. Qi, Y. Fang,
R. T. K. Kwok, X. Zhang, X. Hu, J. W. Y. Lam, D. Ding, B. Z.
Tang, ACS Nano 2017,11, 7177 – 7188.
[8] a) R. Xu, Y. Wang, X. Duan, K. Lu, D. Micheroni, A. Hu, W. Lin,
J. Am. Chem. Soc. 2016,138, 2158–2161; b) C. Zhang, K. Zhao,
W. Bu, D. Ni, Y. Liu, J. Feng, J. Shi, Angew. Chem. Int. Ed. 2015,
54, 1770 – 1774 ; Angew. Chem. 2015,127, 1790 – 1794.
[9] D. Wang, H. Wu, W. Q. Lim, S. Z. F. Phua, P. Xu, Q. Chen, Z.
Guo, Y. Zhao, Adv. Mater. 2019,31, 1901893.
[10] C. Thry, M. Ostrowski, E. Segura, Nat. Rev. Immunol. 2009,9,
581 – 593.
[11] C. Thry, L. Zitvogel, S. Amigorena, Nat. Rev. Immunol. 2002,2,
569 – 579.
[12] J. G. van den Boorn, M. Schlee, C. Coch, G. Hartmann, Nat.
Biotechnol. 2011,29, 325 – 326.
[13] a) L. Alvarez-Erviti, Y. Seow, H. Yin, C. Betts, S. Lakhal,
M. J. A. Wood, Nat. Biotechnol. 2011,29, 341 –345; b) J. Skog, T.
Wrdinger, S. van Rijn, D. H. Meijer, L. Gainche, W. T. Curry,
B. S. Carter, A. M. Krichevsky, X. O. Breakefield, Nat. Cell Biol.
2008,10, 1470 – 1476.
[14] Y. Yang, Y. Hong, G.-H. Nam, J. H. Chung, E. Koh, I.-S. Kim,
Adv. Mater. 2017,29, 1605604.
[15] a) E. V. Batrakova, M. S. Kim, J. Controlled Release 2015,219,
396 405; b) J. G. van den Boorn, J. Daßler, C. Coch, M. Schlee,
G. Hartmann, Adv. Drug Delivery Rev. 2013,65, 331 – 335 ;
c) S. C. Jang, O. Y. Kim, C. M. Yoon, D.-S. Choi, T.-Y. Roh, J.
Park, J. Nilsson, J. Lçtvall, Y.-K. Kim, Y. S. Gho, ACS Nano
2013,7, 7698 – 7710.
[16] A. Fahr, X. Liu, Expert Opin. Drug Delivery 2007,4, 403 – 416.
[17] a) V. P. Chauhan, T. Stylianopoulos, J. D. Martin, Z. Popovic
´,O.
Chen, W. S. Kamoun, M. G. Bawendi, D. Fukumura, R. K. Jain,
A
ngewandte
Chemi
e
Research Articles
&&&&
www.angewandte.org  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2020,59,210
Ü
Ü
These are not the final page numbers!
Nat. Nanotechnol. 2012,7, 383388; b) W. Jiang, Y. Huang, Y.
An, B. Y. S. Kim, ACS Nano 2015,9, 8689 – 8696.
[18] a) R. P. Dings, M. Loren, H. Heun, E. McNiel, A. W. Griffioen,
K. H. Mayo, R. J. Griffin, Clin. Cancer Res. 2007,13, 3395 – 3402 ;
b) C. Rolny, M. Mazzone, S. Tugues, D. Laoui, I. Johansson, C.
Coulon, M. L. Squadrito, I. Segura, X. Li, E. Knevels, S. Costa, S.
Vinckier, T. Dresselaer, P. Akerud, M. De Mol, H. Salomaki, M.
Phillipson, S. Wyns, E. Larsson, I. Buysschaert, J. Botling, U.
Himmelreich, J. A. Van Ginderachter, M. De Palma, M. Dew-
erchin, L. Claesson-Welsh, P. Carmeliet, Cancer Cell 2011,19,
31 – 44.
[19] a) J. D. Martin, M. Panagi,C. Wang, T. T. Khan, M. R. Martin, C.
Voutouri, K. Toh, P. Papageorgis, F. Mpekris, C. Polydorou, G.
Ishii, S. Takahashi, N. Gotohda, T. Suzuki, M. E. Wilhelm, V. A.
Melo, S. Quader, J. Norimatsu, R. M. Lanning, M. Kojima, M. D.
Stuber, T. Stylianopoulos, K. Kataoka, H. Cabral, ACS Nano
2019,13, 63966408; b) S. B. Wang, Z. X. Chen, F. Gao, C.
Zhang, M. Z. Zou, J. J. Ye, X. Zeng, X. Z. Zhang, Biomaterials
2020,234, 119772.
[20] S. J. Gould, A. M. Booth, J. E. Hildreth, Proc. Natl. Acad. Sci.
USA 2003,100, 10592 – 10597.
[21] a) F. Gao, Y. Tang, W. L. Liu, M. Z. Zou, C. Huang, C. J. Liu,
X. Z. Zhang, Adv. Mater. 2019,31, 1904639; b) W. Xie, W. W.
Deng, M. Zan, L. Rao, G. T. Yu, D. M. Zhu, W. T. Wu, B. Chen,
L. W. Ji, L. Chen, K. Liu, S. S. Guo, H. M. Huang, W. F. Zhang,
X. Zhao, Y. Yuan, W. Dong, Z. J. Sun, W. Liu, ACS Nano 2019,
13, 2849 – 2857.
[22] T. Yong, X. Zhang, N. Bie, H. Zhang, X. Zhang, F. Li, A.
Hakeem, J. Hu, L. Gan, H. A. Santos, X. Yang, Nat. Commun.
2019,10, 3838.
[23] a) H. Yang, Q. Wang, Z. Li, F. Li, D. Wu, M. Fan, A. Zheng, B.
Huang, L. Gan, Y. Zhao, X. Yang, Nano Lett. 2018,18, 7909 –
7918; b) R. Wang, C. Zhang, J. Li, J. Huang, Y. Opoku-Damoah,
B. Sun, J. Zhou, L. Di, Y. Ding, Biomaterials 2019,221, 119413;
c) Y.-Y. Yuan, C.-Q. Mao, X.-J. Du, J.-Z. Du, F. Wang, J. Wang,
Adv. Mater. 2012,24, 5476 5480; d) H.-X. Wang, Z.-Q. Zuo, J.-
Z. Du, Y.-C. Wang, R. Sun, Z.-T. Cao, X.-D. Ye, J.-L. Wang,
K. W. Leong, J. Wang, Nano Today 2016,11, 133 – 144.
[24] A. Nel, T. Xia, L. Mdler, N. Li, Science 2006,311, 622 – 627.
Manuscript received: March 11, 2020
Revised manuscript received: April 20, 2020
Accepted manuscript online: May 4, 2020
Version of record online: &&
&&
,
&&&&
A
ngewandte
Chemi
e
Research Articles
&&&&
Angew. Chem. Int. Ed. 2020,59, 2 10  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
Ü
Ü
Research Articles
Anticancer Drugs
D. Zhu, Y. Duo, M. Suo, Y. Zhao, L. Xia,
Z. Zheng, Y. Li,*
B. Z. Tang*
&&&& &&&&
Tumor-Exocytosed Exosome/
Aggregation-Induced Emission
Luminogen Hybrid Nanovesicles
Facilitate Efficient Tumor Penetration and
Photodynamic Therapy
Hybrid AIEgen and biological tumor-
exocytosed exosomes nanovesicles
(DES) were combined with photodynamic
therapy (PDT) in tumor vessel normal-
ization therapy. DES enhance tumor
tissue penetration of AIEgens signifi-
cantly. Dexamethasone (DEX) was used
to normalize vascular function within the
tumor microenvironment, thereby reduc-
ing local hypoxia and enhancing the PDT
effect of DES. Key: aggregation-induced
emission luminogens (AIEgens), photo-
sensitizer (DCPy).
A
ngewandte
Chemi
e
Research Articles
&&&&
www.angewandte.org  2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2020,59,210
Ü
Ü
These are not the final page numbers!
... TEVS-loaded chemotherapy drugs can promote drug uptake and reverse drug resistance in tumor-regenerative or stem-like cancer cells [221][222][223]. Remarkable achievements have been made in photodynamic and photothermal tumor therapy by exploiting the homologous targeting properties of TEVs [77,108,111,224]. Wang used TCMVs to encapsulate ovalbumin-assembled nanoparticles encapsulating the photosensitizer Ce6 [75]. ...
... TEVS-loaded chemotherapy drugs can promote drug uptake and reverse drug resistance in tumor-regenerative or stemlike cancer cells [221][222][223]. Remarkable achievements have been made in photodynamic and photothermal tumor therapy by exploiting the homologous targeting properties of TEVs [77,108,111,224]. Wang used TCMVs to encapsulate ovalbumin-assembled nanoparticles encapsulating the photosensitizer Ce6 [75]. ...
Article
Full-text available
Tumor-derived membrane vesicles (TDMVs) are non-invasive, chemotactic, easily obtained characteristics and contain various tumor-borne substances, such as nucleic acid and proteins. The unique properties of tumor cells and membranes make them widely used in drug loading, membrane fusion and vaccines. In particular, personalized vectors prepared using the editable properties of cells can help in the design of personalized vaccines. This review focuses on recent research on TDMV technology and its application in personalized immunotherapy. We elucidate the strengths and challenges of TDMVs to promote their application from theory to clinical practice.
... The photothermal conversion efficiency (h) of FeS 2 was 33.2% (Figure 2A) in our experiments. Only nanomaterials with good biocompatibility can be applied for subsequent biological experiments (35). As shown in Figure 2B, FeS 2 is stable in blood, and even 200 mg/mL FeS 2 would not cause hemolysis. ...
Article
Full-text available
Single photothermal therapy (PTT) has many limitations in tumor treatments. Multifunctional nanomaterials can cooperate with PTT to achieve profound tumor killing performance. Herein, we encapsulated chemotherapeutic drug camptothecin (CPT) and pyrite (FeS2) with dual enzyme activity (glutathione oxidase (GSH-OXD) and peroxidase (POD) activities) into an injectable hydrogel to form a CFH system, which can improve the level of intratumoral oxidative stress, and simultaneously realize FeS2-mediated PTT and nanozymes catalytic treatment. After laser irradiation, the hydrogel gradually heats up and softens under the photothermal agent FeS2. The CPT then released from CFH to tumor microenvironment (TME), thereby enhancing the H2O2 level. As a result, FeS2 can catalyze H2O2 to produce ·OH, and cooperate with high temperature to achieve high-efficiency tumor therapy. It is worth noting that FeS2 can also deplete excess glutathione (GSH) in the cellular level, further amplifying oxidative stress. Both in vivo and in vitro experiments show that our CFH exhibits good tumor-specific cytotoxicity. The CFH we developed provides new insights for tumor treatment.
Article
Dendritic cell (DC)-derived small extracellular vesicles (DEVs) are recognized as a highly promising alternative to DC vaccines; however, the clinical testing of DEV-based immunotherapy has shown limited therapeutic efficacy. Herein, we develop a straightforward strategy in which DCs serve as a cell reactor to exocytose high-efficient DEV-mimicking aggregation-induced emission (AIE) nanoparticles (DEV-AIE NPs) at a scaled-up yield for synergistic photodynamic immunotherapy. Exocytosed DEV-AIE NPs inherit not only the immune-modulation proteins from parental DCs, enabling T cell activation, but also the loaded AIE-photosensitizer MBPN-TCyP, inducing superior immunogenic cell death (ICD) by selectively accumulating in the mitochondria of tumor cells. Eventually, DEV-AIE synergistic photodynamic immunotherapy elicits dramatic immune responses and efficient eradication of primary tumors, distant tumors, and tumor metastases. In addition, cancer stem cells (CSCs) in 4T1 and CT26 solid tumors were significantly inhibited by the immune functional DEV-AIE NPs. Our work presents a facile method for the cellular generation of EV-biomimetic NPs and demonstrates that the integration of DEVs and AIE photosensitizers is a powerful direction for the production of clinical anticancer nanovaccines.
Article
Fluorescent imaging based on near-infrared (NIR) fluorophores has revolutionized the techniques employed for detecting biological events in depth owing to their advantages referring to diminished photon scattering, high signal-to-noise ratio and better light transparence through tissue. As for conventional luminogens, the nanofabrication of those innately hydrophobic π-conjugated architectures into water-dispersible nanoparticles (NPs) may result in attenuated fluorescent intensity deriving from the detrimental distribution of π-π interactions in the confined space. Oppositely, chromophores possessing aggregation-induced emission (AIE) characteristics emit boosted brightness at aggregate level according to the mechanism of restriction of intramolecular motion (RIM). In this review, we summarize the recent progresses of NIR emissive AIE NPs for multifarious biomedical applications from the viewpoint of different fabricated manners, mainly covering self-assembly and matrices assisted approaches. Furthermore, the current challenges and future research directions of NIR AIE NPs are briefly discussed.
Article
As a non-invasive visualization technique, photoluminescence imaging (PLI) has found its huge value in many biological applications associated with intracellular process monitoring and early and accurate diagnosis of diseases. PLI can also be combined with therapeutics to build imaging-guided theragnostic platforms for achieving early and precise treatment of diseases. Photodynamic therapy (PDT) as a quintessential phototheranostics technology has gained great benefits from the combination with PLI. Recently, aggregation-induced emission (AIE)-active materials have emerged as one of the most promising bioimaging and phototheranostic agents. Most of AIEgens, however, need to be chemically engineered to form versatile nanocomposites with improved their photophysical property, photochemical activity, biocompatibility, etc. In this review, we focus on three categories of AIE-active nanocomposites and highlight their application progresses in the intracellular biological process monitoring and PLI-guided PDT. We hope this review can guide further development of AIE-active nanocomposites and promote their practical applications for monitoring intracellular biological processes and imaging-guided PDT.
Article
Extracellular vesicles (EVs), as natural carriers of bioactive cargo, have a unique micro/nanostructure, bioactive composition, and characteristic morphology, as well as fascinating physical-, chemical- and biochemical- features, which have shown promising application in the treatment of a wide range of diseases. However, native EVs have limitations such as efficient cell targeting, on-demand delivery, and therapeutic feedback. Recently, EVs have been engineered to contain an intelligent core, enabling them to (i) actively target sites of disease, (ii) respond to endogenous and/or exogenous signals, and (iii) provide treatment feedback for optimal function in the host. These advances pave the way for next-generation nanomedicine and offer promise for a revolution in drug delivery. Here, we summarise recent research on intelligent EVs and discuss the use of “intelligent core” based EV systems for the treatment of disease. We provide a critique about the construction and properties of intelligent EVs, and challenges in their commercialization. We compare the therapeutic potential of intelligent EVs to traditional nanomedicine and highlight key advantages for their clinical application. Collectively, this review aims to provide a new insight in the design of next-generation EV-based theranostic platforms for disease treatment.
Article
It is highly desirable to turn “cold” tumors into “hot” ones to improve the efficacy of antitumor immunotherapy. Herein, we develop the peptide-based small interfering RNA (siRNA) micelleplexes (PA7R@siPD-L1) for normalizing vascular-immune crosstalk to establish a positive feedback loop in potentiating antitumor immunotherapy. These micelleplexes are equipped with tumor-microenvironment-responsive property for precise drug delivery and release. The prepared PA7R@siPD-L1-mediated photodynamic therapy could eliminate solid tumors and trigger immunogenic cell death to generate systematic immune responses. Meanwhile, the antiangiogenic peptide A7R normalizes the tumor vasculature by converting chaotic vascular systems into matured and organized ones, thus alleviating tumor hypoxia and promoting intratumoral infiltration by immune cells. Antitumor immunogenicity would be further strengthened with the aid of siPD-L1 by muting the resistance of tumor cells against immune effector cells. This study provides a unique therapeutic strategy in turning “cold” tumors into “hot” tumors enabled by siRNA nanomaterial.
Article
Photodynamic nanomedicines have significantly enhanced the therapeutic efficacy of photosensitizers (PSs) by overcoming critical limitations of PSs such as poor water solubility and low tumor accumulation. Furthermore, functional photodynamic nanomedicines have enabled overcoming oxygen depletion during photodynamic therapy (PDT) and tissue light penetration limitation by supplying oxygen or upconverting light in targeted tumor tissues, resulting in providing the potential to overcome biological therapeutic barriers of PDT. Nevertheless, their localized therapeutic effects still remain a huddle for the effective treatment of metastatic- or recurrent tumors. Recently, newly designed photodynamic nanomedicines and their combination chemo- or immune checkpoint inhibitor therapy enable the systemic treatment of various metastatic tumors by eliciting antitumor immune responses via immunogenic cell death (ICD). This review introduces recent advances in photodynamic nanomedicines and their applications, focusing on overcoming current limitations. Finally, the challenges and future perspectives of the clinical translation of photodynamic nanomedicines in cancer PDT are discussed.
Article
Full-text available
Relative to traditional photosensitizer (PS) agents, those that exhibit aggregation-induced emission (AIE) properties offer key advantages in the context of photodynamic therapy (PDT). At present, PDT efficacy is markedly constrained by the hypoxic microenvironment within tumors and the limited depth to which lasers can penetrate in a therapeutic context. Herein, we developed platelet-mimicking MnO2 nanozyme/AIEgen composites (PMD) for use in the interventional PDT treatment of hypoxic tumors. The resultant biomimetic nanoparticles (NPs) exhibited excellent stability and were able to efficiently target tumors. Moreover, they were able to generate O2 within the tumor microenvironment owing to their catalase-like activity. Notably, through an interventional approach in which an optical fiber was introduced into the abdominal cavity of mice harboring orthotopic colon tumors, good PDT efficacy was achieved. We thus propose that a novel strategy consisting of a combination of an AIEgen-based bionic nanozyme and a biomimetic cell membrane coating represents an ideal therapeutic platform for targeted antitumor PDT. This study is the first to have combined interventional therapy and AIEgen-based PDT, thereby overcoming the limited light penetration that typically constrains the therapeutic efficacy of this technique, highlighting a promising new AIEgen-based PDT treatment strategy.
Article
Full-text available
Regulating the tumor microenvironment (TME) has been a promising strategy to improve antitumor therapy. Here, a red blood cell membrane (mRBC)‐camouflaged hollow MnO2 (HMnO2) catalytic nanosystem embedded with lactate oxidase (LOX) and a glycolysis inhibitor (denoted as PMLR) is constructed for intra/extracellular lactic acid exhaustion as well as synergistic metabolic therapy and immunotherapy of tumor. Benefiting from the long‐circulation property of the mRBC, the nanosystem can gradually accumulate in a tumor site through the enhanced permeability and retention (EPR) effect. The extracellular nanosystem consumes lactic acid in the TME by catalyzing its oxidation reaction via LOX. Meanwhile, the intracellular nanosystem releases the glycolysis inhibitor to cut off the source of lactic acid, as well as achieve antitumor metabolic therapy through the blockade of the adenosine triphosphate (ATP) supply. Both the extracellular and intracellular processes can be sensitized by O2, which can be produced during the decomposition of endogenous H2O2 catalyzed by the PMLR nanosystem. The results show that the PMLR nanosystem can ceaselessly remove lactic acid, and then lead to an immunocompetent TME. Moreover, this TME regulation strategy can effectively improve the antitumor effect of anti‐PDL1 therapy without the employment of any immune agonists to avoid the autoimmunity. A strategy based on intra/extracellular lactic acid exhaustion is reported to achieve synergistic metabolic therapy and immunotherapy of tumors. This strategy is performed by a cascade catalytic nanosystem (PMLR) that integrates a hollow MnO2 nanocarrier with lactate oxidase and a glycolysis inhibitor.
Article
Full-text available
Developing biomimetic nanoparticles without loss of the integrity of proteins remains a major challenge in cancer chemotherapy. Here, we develop a biocompatible tumor-cell-exocytosed exosome-biomimetic porous silicon nanoparticles (PSiNPs) as drug carrier for targeted cancer chemotherapy. Exosome-sheathed doxorubicin-loaded PSiNPs (DOX@E-PSiNPs), generated by exocytosis of the endocytosed DOX-loaded PSiNPs from tumor cells, exhibit enhanced tumor accumulation, extravasation from blood vessels and penetration into deep tumor parenchyma following intravenous administration. In addition, DOX@E-PSiNPs, regardless of their origin, possess significant cellular uptake and cytotoxicity in both bulk cancer cells and cancer stem cells (CSCs). These properties endow DOX@E-PSiNPs with great in vivo enrichment in total tumor cells and side population cells with features of CSCs, resulting in anticancer activity and CSCs reduction in subcutaneous, orthotopic and metastatic tumor models. These results provide a proof-of-concept for the use of exosome-biomimetic nanoparticles exocytosed from tumor cells as a promising drug carrier for efficient cancer chemotherapy.
Article
Full-text available
Immunogenic cell death (ICD)-associated immunogenicity can be evoked through reactive oxygen species (ROS) produced via endoplasmic reticulum (ER) stress. In this study, we generate a double ER-targeting strategy to realize photodynamic therapy (PDT) photothermal therapy (PTT) immunotherapy. This nanosystem consists of ER-targeting pardaxin (FAL) peptides modified-, indocyanine green (ICG) conjugated- hollow gold nanospheres (FAL-ICG-HAuNS), together with an oxygen-delivering hemoglobin (Hb) liposome (FAL-Hb lipo), designed to reverse hypoxia. Compared with non-targeting nanosystems, the ER-targeting naosystem induces robust ER stress and calreticulin (CRT) exposure on the cell surface under near-infrared (NIR) light irradiation. CRT, a marker for ICD, acts as an 'eat me' signal to stimulate the antigen presenting function of dendritic cells. As a result, a series of immunological responses are activated, including CD8+ T cell proliferation and cytotoxic cytokine secretion. In conclusion, ER-targeting PDT-PTT promoted ICD-associated immunotherapy through direct ROS-based ER stress and exhibited enhanced anti-tumour efficacy.
Article
Photodynamic therapy (PDT) has been a well-accepted clinical treatment for malignant tumors owing to its non-invasiveness and high spatiotemporal selectivity. However, the efficiency of PDT is still severely hindered by inherent aggregation caused quenching (ACQ) effect of traditional photosensitizer (PSs), the presence of B-cell lymphoma 2 (Bcl-2), an anti-apoptosis protein in cells and hypoxia in the tumor microenvironment. To address these issues, hybrid nanospheres containing Fe3+, aggregation induced emission (AIE) PS and Bcl-2 inhibitor of sabutoclax were constructed via coordination driven self-assembly in aqueous media. Once the hybrid nanospheres are uptaken by tumor cells, intracellular O2 concentration is observed to increase remarkably via Fenton reaction driven by Fe3+ while intracellular PDT resistance of the AIE PS was mitigated by sabutoclax. The design of the multifunctional hybrid nanospheres demonstrates a prospective nanoplatform for image-guided enhanced PDT of tumors.
Article
Photodynamic therapy (PDT) is a promising treatment modality for tumor suppression. However, the hypoxic state of most solid tumors might largely hinder the efficacy of PDT. Here, a functional covalent organic framework (COF) is fabricated to enhance PDT efficacy by remodeling the tumor extracellular matrix (ECM). Anti-fibrotic drug pirfenidone (PFD) is loaded in an imine-based COF (COFTTA-DHTA) and followed by the decoration of poly(lactic-co-glycolic-acid)-poly(ethylene glycol) (PLGA-PEG) to fabricate PFD@COFTTA-DHTA@PLGA-PEG, or PCPP. After injected intravenously, PCPP can accumulate and release PFD in tumor sites, leading to down-regulation of ECM compenents such as hyaluronic acid (HA) and collagen I. Such depletion of tumor ECM reduces the intratumoral solid stress, a compressive force exerted by the ECM and cells, decompresses tumor blood vessels, and increases the density of effective vascular areas, resulting in significantly improved oxygen supply in tumor. Furthermore, PCPP-mediated tumor ECM depletion also enhances the tumor uptake of subsequently injected Protoporphyrinl IX (PPIX)-conjugated peptide formed nanomicelles (NM-PPIX) due to the improved enhanced permeability and retention (EPR) effect. Both the alleviated tumor hypoxia and improved tumor homing of photosensitizer (PS) molecules after PCPP treatment significantly increase the reactive oxygen species (ROS) generation in tumor and therefore realize greatly enhanced PDT effect of tumor in vivo.
Article
Photosensitizers with aggregation-induced emission (AIE) characteristics are of great interest for cancer theranostics involving both fluorescence imaging and photodynamic therapy (PDT). However, in the purpose of clinical trials of PDT, the development of prominent drug delivery systems for boosting the PDT efficiency of AIE photosensitizers is highly desirable but still remain a challenging task. Herein, a novel strategy is designed and performed for boosting PDT effect based on stimuli-responsive nano-micelles as extraordinary carriers for an AIE photosensitizer, namely MeTTMN. Those presented stimuli-responsive nano-micelles loading MeTTMN exhibit good biocompatibility, excellent stability, appropriate nanoparticle size, high loading efficiency, outstanding imaging quality and significantly promoted PDT performance, eventually making them remarkably impressive and significantly superior to commercially available nano-micelles carried MeTTMN. This study thus offers an ideal template for fluorescence imaging-guided PDT, as well as a promising candidate for clinical trials.
Article
Hypoxia, which frequently reduces the sensitivity to many therapeutic interventions, including chemotherapy, radiotherapy and phototherapy, has been acknowledged as an important reason for poor prognosis. Burgeoning evidences have proved that the tumor hypoxia microenvironment can reduce the therapeutic effect on tumor through inhibiting the drug efficacy, limiting immune cell infiltration of tumors and accelerating tumor recurrence and metastasis. However, the relationship between oxygen supply and the proliferation of cancer cells is still ambiguous and argued. Different from the current commonly used oxygen supply strategies, this study concentrated on the reduction of endogenous oxygen consumption. Specifically, a novel photosensitizers (IR780) and metformin are packaged in PEG-PCL liposomes. Once such nanoparticles accumulated in tumor tissues, the tumor foci were irradiated through 808 nm laser, generated ROS to further release metformin and IR780. Metformin can directly inhibit the activity of complex Ⅰ in the mitochondrial electron transport chain, thus performed a potent inhibitor of cell respiration. After overcoming tumor hypoxia, the combination of mitochondria-targeted photodynamic therapy (PDT) and photothermic therapy (PTT) via IR780 may achieve superior synergistically therapeutic efficacy. Benefit from excellent characteristics of IR780, such synergistic PDT PTT with the inhibition of mitochondrial respiration can be monitored through near-infrared/photoacoustic dual-modal imaging. Such a conception of reducing endogenous oxygen consumption may offer a novel way to solve the important puzzles of hypoxia-induced tumor resistance to therapeutic interventions, not limited to phototherapy.
Article
Fluorescent sensing has emerged as a powerful tool for detecting various analytes and visualizing numerous biological processes by virtue of its superb sensitivity, rapidness, excellent temporal resolution, easy operation, and low cost. Of particular interest is activity-based sensing (ABS), a burgeoning sensing approach that is actualized on the basis of dynamic molecular reactivity rather than conventional lock-and-key molecular recognition. ABS has been recognized to possess some distinct advantages, such as high specificity, extraordinary sensitivity, and accurate signal outputs. A majority of ABS sensors are constructed by modifying conventional fluorogens, which are strongly emissive when molecularly dissolved in solvents but experience emission quenching upon aggregate formation or concentration increase. The aggregation-caused quenching (ACQ) phenomenon leads to a limited amount of labeling of the analyte with the sensor and low photobleaching resistance, which could impede practical applications of the ABS protocol. As an anti-ACQ phenomenon, aggregation-induced emission (AIE) provides a straightforward solution to the ACQ problem. Thanks to their intrinsic advantages, including high photobleaching threshold, high signal-to-noise ratio, fluorescence turn-on nature, and large Stokes shift, AIE-active luminogens (AIEgens) represent a class of extraordinary fluorogen alternatives for the ABS protocol. The use of AIEgen-involved ABS can integrate the advantages of AIEgens and ABS, and additionally, the AIE process offers some unique properties to the ABS approach. For instance, in some cases of water-soluble AIEgen-involved ABS, chemical reaction not only leads to a chang in the emission color of the AIEgens but also causes solubility variations, which could result in specific "light-up" signaling. In this Account, the basic concepts and mechanistic insights of the ABS approach involving the AIE principle are briefly summarized, and then we highlight the new breakthroughs, seminal studies, and trends in the area that have been most recently reported by our group. This emerging sensing protocol has been successfully utilized for detecting an array of targets including ions, small molecules, biomacromolecules, and microenvironments, all of which closely relate to human health, medical, and public concerns. These detections are smoothly achieved on the basis of various reactions (e.g., hydrolysis, boronate cleavage, dephosphorylation, addition, cyclization, and rearrangement reactions) through different sensing principles. In these studies, the AIEgen-involved ABS strategy generally shows good biocompatibility, high selectivity, excellent reliability and high signal contrast, strongly indicating its great potential for high-tech innovations in the sensing field, among which bioprobing is of particular interest. With this Account, we hope to spark new ideas and inspire new endeavors in this emerging research area, further promoting state-of-the-art developments in the field of sensing.
Article
Natural particles ranging from various cell membranes to nascent proteins are highly optimized for their specific functions in vivo and possess features that are desired in drug delivery carriers. However, the current endeavor in research on bioparticles is still seeking the appropriate strategy to shield multiple agents and circumvent biological hurdles. These issues have propelled the advancement of lipid-polymer hybrid nanocarriers, which could be employed as drug reservoirs and strive to meet these expectations. We thereby proposed functionalized biopeptide-lipid hybrid particles, which were applied to encapsulating a PLGA polymeric core together with indocyanine green (ICG) and packaged by a lipoprotein-inspired structural shell. To initiate precision tumor-penetrating performance, tLyP-1-fused apolipoprotein A-I-mimicking peptides (D4F) were exploited to impart tumor-homing and tumor-penetrating biological functions. The sub-100 nm drug vehicle possessed a long circulation time with uniform mono-dispersity but was stable enough to navigate freely, penetrate deeply into tumors and deliver its cargoes to the targeted sites. Moreover, ICG-encapsulated penetrable polymeric lipoprotein particles (PPL/ICG) could realize real-time fluorescence/photoacoustic imaging for monitoring in vivo dynamic distribution. Upon near-infrared (NIR) laser irradiation, PPL/ICG demonstrated a highly efficient phototherapeutic effect to eradicate orthotopic xenografted tumors, resulting in an 88.77% decrease from the initial tumor volume and inhibited tumor metastasis with good biosafety. Therefore, the described bio-strategy opens new avenues for creating polymeric lipoproteins with varied hybrid functionalities, which may be applied to provide a basis and inspiration for improved nanoparticle-based precision theranostic nanoplatforms.