Nanomized tumor-microenvironment-active NIR fluorescent prodrug for ensuring synchronous occurrences of drug release and fluorescence tracing

Article abstract of DOI:10.1039/c8tb03188f

Improving the bioavailability and tumor-targeting ability of a prodrug, as well as monitoring its active ingredient release in vivo, is still a challenge in cancer diagnosis and therapy. Herein, a specific nanomized tumor-microenvironment-active near-infrared (NIR) fluorescent DCM-S-GEM/PEG prodrug was developed as a potent monitoring platform, wherein we conjugated antitumor drug gemcitabine (GEM) and NIR fluorescent chromophore dicyanomethylene-4H-pyran (DCM) via glutathione (GSH)-activatable disulfide linker and encapsulated DCM-S-GEM into an amphiphilic polymer DSPE-mPEG by self-assembly. The nanomized DCM-S-GEM/PEG prodrug exhibits excellent photostability and high biocompatibility, significantly improving the therapeutic efficacy toward lung tumor cells with fewer side-effects toward normal cells. Furthermore, when compared with the standalone DCM-S-GEM prodrug, the micellization with diblock DSPE-mPEG avoids fast metabolism, facilitates the accumulation of drugs in lung tumor tissues, displays longer tumor retention, and realizes precise drug release in lung tumors. The nanomized DCM-S-GEM/PEG prodrug can be developed as a promising tool to monitor prodrug delivery and activation processes in vivo.

Full text of DOI:10.1039/c8tb03188f

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Regulating the stemness of mesenchymal stem cells by tuning
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DOI: 10.1039/C8TB03188F
Journal Name
ARTICLE
Nanomized tumor-microenvironment active NIR fluorescent
prodrug for ensuring synchronous occurrences of drug releasing
and fluorescence tracing†
Received 00th January 20xx,
Accepted 00th January 20xx
a
b
,a
a
a
a
,a
Qiang Li, Jun Cao, Qi Wang,* Jie Zhang, Shiqin Zhu, Zhiqian Guo and Wei-Hong Zhu*
DOI: 10.1039/x0xx00000x
Improving the prodrug bioavailability and tumor targeting ability, as well as monitoring active ingredient release in vivo, is
still a challenge for cancer diagnosis and therapy. Herein, a specific nanomized tumor-microenvironment active near-
infrared (NIR) fluorescent prodrug DCM-S-GEM/PEG as a potent monitoring platform is developed, in which we conjugate
antitumor drug gemcitabine (GEM) and the NIR fluorescent chromophore dicyanomethylene-4H-pyran (DCM) via a
glutathione (GSH) activatable disulfide linker, and encapsulate DCM-S-GEM into the amphiphilic polymer DSPE-mPEG by
self-assembling. The nanomized prodrug DCM-S-GEM/PEG exhibits excellent photostability and high biocompatibility,
significantly improving the therapeutic efficacy to lung tumor cells with low side effects to normal cells. Furthermore,
compared with the free prodrug DCM-S-GEM, the micelliazation with diblock DSPE-mPEG avoids fast metabolism, facilitates
the accumulation of drugs in lung tumor tissues, displays longer tumor retention, and realizes precise drug release in lung
tumor. The nanomized DCM-S-GEM/PEG can be developed as a promising tool to monitor prodrug delivery and activation
process in vivo.
www.rsc.org/
of individualized treatment as well as low toxicity, non-invasive
Introduction
11,12
properties and forceful compliance.
However, the long-real-time
Chemotherapy is an indispensable choice for most cancer cases
because of its high efficiency among various cancer treatments.1
Conventional chemotherapy suffers from poor bioavailability
because of the rapid blood clearance, and nonspecific selectivity
along with low accumulation in tumors.2,3 To address these
drug tracing in vivo is still a huge challenge, owing to the inaccuracy,
instability, and the systematic toxicity in the intricate internal
13
14
environment. Whereas, polymeric micelle, whose nanoscopic
structure was formed through the self-assembly of amphiphilic
copolymer, is helpful for improving the tumor targeting, regulating
the bioavailability as well as increasing stability of prodrug or
fluorescent sensors. As reported, polymeric micelle exhibits good
4
limitations, prodrugs are widely used in the chemotherapies, such
5
as peptide-based prodrugs for improving tumor targeting ability,
6
amino acid-based prodrugs for enhancing both tumor targeting
15
16
17
biodegradability,
stability,
targeting property,
water
7
ability and solubility, aptamer–based prodrugs for higher specificity
18
19
dispersibility, and selectivity, further facilitates blood circulation
of the prodrug vesicles by suppressing protein absorption and
avoiding recognition with macrophages.
Hence, in this work, we design
8
9
as well as affinity, and enzyme cleavable or activated prodrugs for
high specificity and enhanced therapy. Unfortunately, restricted by
the small size, free anti-cancer prodrug still faces the challenges of
fast clearance by blood or kidney, poor bioavailability, adverse
therapeutic effects, especially huge unpredictability, which greatly
20
a
nanomized tumor-
microenvironment active near-infrared (NIR) prodrug DCM-S-
GEM/PEG to trace the drug fate and improve the bioavailability in
vivo owing to the following considerations (Fig. 1A): i) the DSPE-
10
limit the clinical application. Therefore, a strategy for improving the
bioavailability and tumor targeting ability, as well as monitoring
active ingredient release from prodrug is urgently required in tumor
therapy.
21,22
mPEG micelles
encapsulate the prodrug DCM-S-GEM, ensuring
both high water dispersity, tumor targeting, and long time retention
along with low side toxicity, ii) the fluorophore dicyanomethylene-
Fluorophores-labelled prodrugs are helpful for monitoring the
release and eliminating the unpredictability of chemotherapeutic
agent by precise pharmacodynamics evaluation. It is advantageous
23-26
4H-pyran (DCM) chromophore
ensures the long-time low-toxic
tracing because of the excellent emission ability, high photostability
and NIR region, iii) the disulfide bond (-S-S-) is specifically cleavable
by tumor-overexpressed glutathione (GSH) which ensures the tumor
targeting
and
negative
systematic
toxicity,2
7-29
and
a.Shanghai Key Laboratory of Functional Materials Chemistry, Key Laboratory for
Advanced Materials and Institute of Fine Chemicals, Joint International Research
Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel
Prize Scientist Joint Research Center, School of Chemistry and Molecular
Engineering, East China University of Science and Technology, Shanghai 200237,
China. *E-mail: wangqi@ecust.edu.cn, whzhu@ecust.edu.cn
30-32
chemotherapeutic agent gemcitabine (GEM)
is connected to
fluorophore DCM, which ensures the synchronous occurrences of
the drug releasing and fluorescence lighting. The tumor-environment
active prodrug DCM-S-GEM/PEG exhibited high toxicity to tumor
cells, low side effects to normal cells, strong photostability, and
excellent sensitivity in both live A549 cells and lung cancer mice
model, providing an potential approach to real-time and long-time
monitoring of the drug delivery and activation process in vivo.
b.
Department of Interventional Oncology, Dahua Hospital, Xuhui District, Shanghai
2
00237, China.
†
Electronic supplementary information (ESI) available: Reagents and instruments,
1
13
synthetic procedures,
H
and
C NMR, and HRMS information. See
DOI: 10.1039/x0xx00000x
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Fig. 1 Tumor-microenvironment active NIR fluorescent platform DCM-S-GEM/PEG for tumor therapy and drug monitoring. (A) The
tumor-microenvironment sensitive fluorescent platform DCM-S-GEM/PEG generated by assembling the prodrug DCM-S-GEM into
the DSPE-mPEG micelles. (B) Tumor treatment is conducted that, i) injecting the DCM-S-GEM/PEG into the tumor tissue, ii) the DCM-
S-GEM/PEG is internalized, and followed by iii) selectively cleavage of prodrug and light up of fluorescence.
biocompatibility and further enhance the cellular internalization as
well as extend retention time in tumor tissue (Fig. 1B).
Results and discussion
38,39
Micellizing fluorescent prodrug for solving its bioavailability and
tumor targeting ability
GSH-active response of fluorescent prodrug DCM-S-GEM
The disulfide bond is a well-known activatable linker by thiols, such
GEM, a DNA chain terminator, is one of the widely used agents in the
treatment for several solid tumors, especially for non-small-cell lung
27,28,40
as GSH, Cys, Hcy, and so on.
Since the tumor cells express much
4
1-43
more GSH with respect to normal cells,
the fluorescent prodrug
33,34
cancer.
GEM provides an improvement in the quality of life, but
DCM-S-GEM is proposed to be selectively cleaved in tumor tissue,
followed by releasing NIR fluorescence chromophore DCM for tumor
imaging as well as drug tracing, and chemotherapeutic agent GEM
for cancer treatment.
In order to demonstrate whether the fluorophore of DCM and
chemotherapeutic agent of GEM can be concomitantly released from
the disulfide bond-linked prodrug DCM-S-GEM, HPLC spectra analysis
and mass spectroscopy are carried out by incubating DCM-S-GEM
with biologically available thiols GSH. As illustrated in Fig. 2B, the
the side effects and acquired drug resistance seriously limit its clinical
35,36
efficacy.
Here, a fluorescent prodrug DCM-S-GEM was designed
and synthesized to reduce the side effects, improve the tumor
targeting ability and trace the accurate drug distribution. Briefly,
compounds DCM, DCM-NH
to the previous work,
2
and DCM-S were synthesized according
and the final product DCM-S-GEM was
27,37
obtained by mixing DCM-S and GEM in the presence of triphosgene
at room temperature followed by stirring. All intermediates and final
1
13
product DCM-S-GEM were characterized using H, C NMR, and
high-resolution mass spectrometry (HRMS, Fig. S1-S9 in ESI†). The
chemical structure of DCM-S-GEM was described in Fig. 1A that, i)
DCM chromophore with controllable emission, high fluorescent
brightness, and strong photostability ensuring the satisfying tissue
penetration and long time fluorescence imaging in cells, ii) GEM, a
first line chemotherapeutic agent for non-small-cell lung cancer
treatment, iii) disulfide bond, the tumor-environment active linker
between DCM derivative and GEM, ensuring the selectively cleavage
in tumor tissue for eliminating the side effects and tracing the drug
release. Then, DCM-S-GEM was encapsulated into diblock polymer
DSPE-mPEG (abbreviated as DCM-S-GEM/PEG), a amphiphilic
polymer drug carrier with superior biocompatibility, to improve the
aqueous dispersibility, avoid the fast clearance, regulate
2
prodrug DCM-S-GEM, fluorophore DCM-NH , and the antitumor drug
GEM showed narrow peaks with retention time at 16.4, 4.7 and 7.9
min, respectively. After incubating the prodrug DCM-S-GEM with
GSH for 30 min, a new sharp peak at 4.7 min corresponding to DCM-
2
NH , a narrow peak at 7.9 min for GEM and a decreased peak
corresponding to DCM-S-GEM at 16.4 min appeared, indicating the
DCM-S-GEM was cleaved by GSH. Also, after incubating for 20 min,
the mixture were analyzed by mass spectroscopy, showing three m/z
+
peaks 264.0756, 312.0879, and 781.2097 associated with [GEM + H ]
+
+
(
calcd. 264.0790 for C
9
H
12
F
2
N
3
O
4
), [DCM-NH
2
+ H ] (calcd. 312.1131
+
+
for C20
C
H
14
N
3
O ) and [DCM-S-GEM + H ] (calcd.781.1557 for
) (Fig. S10†). Both experiments demonstrated the
+
35
H
31
F
2
N
6
O
9
S
2
fluorescent prodrug DCM-S-GEM is cleaved by GSH, followed by
releasing pharmaceutical agent GEM and fluorescence agent DCM-
2
NH (Fig. 2A).
2
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Fig. 2 GSH-activatable response of fluorescence prodrug DCM-S-GEM. (A) Response mechanism of DCM-S-GEM with GSH under
physiological conditions for drug release, in which the disulfide bond (-S-S-) can be cleaved via GSH. (B) HPLC spectra of DCM-S-GEM,
GEM, DCM-NH , and reaction product of DCM-S-GEM incubated with 2.5 mM GSH for 50 min. (C) Fluorescence spectra and (D)
2
fluorescence intensity (665 nm) towards various amino acids including GSH, Cys, Hcy, Ala, Arg, Asn, Asp, Gln, Glu, Gly, His, Ile, Leu, Phe,
Pro, Ser, Thr, Tho, Tyr, Val, respectively (50 μM). Note: Fluorescence enhancement was observed in thiol-treatment groups, indicating
excellent selectivity to thiol of DCM-S-GEM.
High selectivity is essential for high efficiency as well as low side containing amino acids. Actually, the potential interference of Cys
4
4-46
toxicity in cancer treatment.
Hence, the specificity of the NIR and Hcy can be neglected due to their relatively low concentration
fluorescent prodrug DCM-S-GEM toward GSH was performed. compared to the high physiological concentration of GSH in the
23
Various potential interfere species were tested in this experiment to cytoplasm. Considering with the much higher GSH concentration in
verify the specificity of DCM-S-GEM, including enzyme species, tumor cells to normal cells, this prodrug is supposed with tumor
amino acids, and so on. As shown in Fig. 2C, the fluorescence targeting ability. As demonstrated, the fluorescent prodrug DCM-S-
intensity of DCM-S-GEM was much higher in thiol-containing amino GEM can be specially cleaved by GSH (tumor highly expressed),
acids (glutathione, cysteine and homocysteine) than non-thiol- ensuring the tumor target ability along with limiting side effects.
bearing amino acids, indicative of excellent specificity to thiol-
“
Turn on” fluorescent property of nanomized prodrug DCM-S-
GEM/PEG for drug release monitoring
As shown in Fig. 3, DCM-S-GEM/PEG was well-organized of 110 nm
(
PDI 0.192) with good morphology and dispersity (Fig. 3), and drug
loading effect of 5.21% (Fig. S11†), suggestive of high enhanced
permeability and retention (EPR) effect and low side effects. Owing
to the NIR fluorescent probe for tissue penetrating, the tumor-
environment active disulfide bond for drug tracing, and the micelle
structure for superior biocompatibility and tumor targeting ability, as
well as extended tumor retention, the nanomized tumor-
microenvironment active NIR DCM-S-GEM/PEG is excepted to realize
real-time drug release monitoring and improved tumor lethality.
As demonstrated, the prodrug DCM-S-GEM can be cleaved by
GSH into fluorophore DCM and drug GEM. In order to further study
the fluorescent property, the UV-Vis absorption and fluorescence
Fig. 3 Characterization of DCM-S-GEM/PEG micelles via (A)
transmission electron microscopy (TEM) and (B) dynamic light
scattering (DLS). The size of DCM-S-GEM/PEG is 110 nm with
PDI of 0.192.
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Fig. 4. Tumor-microenvironment sensitive fluorescence ‘turn
on’ prodrug DCM-S-GEM with satisfying photostability.
Absorption (A) and emission spectra (B, λex = 450 nm) of DCM-
S-GEM (10 μM) in aqueous solution (DMSO/PBS = 1/1, v/v; pH
Fig. 5 Internalization and cytotoxicity of DCM-S-GEM/PEG to
lung tumor cells (A549 cells) and normal cells (QSG-7701cells).
Flow cytometry analysis of cellular uptake of (A) DCM-S-GEM
and (B) DCM-S-GEM/PEG at different time intervals from 1 to 4
h in A549 cells. Cell viability of GEM, DCM-S-GEM, DCM-S-
GEM/PEG to (C) QSG-7701 cells and (D) A549 cells. Note: DCM-
S-GEM/PEG markedly facilitated cellular internalization in tumor
cells and exhibited high cytotoxicity to A549 cells along with low
cytotoxicity to QSG-7701 cells in vitro.
=
7.4; 37 °C) upon treatment with increasing concentrations of
GSH (0-250 equiv). Photo-image (inset A) and fluorescence-
image (inset B) of DCM-S-GEM solution before and after
addition of GSH. (C) Fluorescence enhancement at 665 nm with
increasing GSH concentration. (D) Time-dependent
fluorescence intensity of ICG (10 μM in DMSO/PBS = 1:1,
treated with 2.5 mM GSH, monitored at 812 nm, and excited at
be used to predict the treatment effect. Hence, the internalization of
DCM-S-GEM/PEG by lung cancer cells A549 was quantitatively
detected through flow cytometry. As shown in Fig. 5, the
fluorescence cells percent increased on a time-dependent manner
for both DCM-S-GEM/PEG and DCM-S-GEM. Moreover, the DSPE-
mPEG micelles improved the internalization that, the fluorescence
percent is 39.6% for DCM-S-GEM/PEG at 1 h and 92.3% at 6 h (Fig.
B) , compared to 12.3% at 1 h and 83.2% at 6 h for DCM-S-GEM (Fig.
A), suggesting stronger treatment effect toward lung cancer cells.
Standard MTT assay was conducted to investigate the cytotoxicity
7
2
80 nm), DCM-NH , DCM-S-GEM and DCM-S-GEM/PEG (10 μM
in DMSO/PBS = 1:1, treated with 2.5 mM GSH for 1 h at 37 °C,
monitored at 665 nm and excited at 450 nm).
spectra were detected by incubating the prodrug DCM-S-GEM with
biologically available thiols such as GSH (0-250 equiv). As shown in
Fig. 4A, UV absorption gradually red shifted from 450 nm to 485 nm
with an increased GSH concentration, along with a color turning from
pale yellow to pink (inset of Fig. 4A). Also, the fluorescence intensity
at 665 nm became increased after incubating with GSH (Fig. 4B and
C), exhibiting ‘turn-on’ fluorescent characteristic, suggestive of
precise bioimaging possibility of drug release in tumor tissue and
biodistribution in vivo.
5
5
of DCM-S-GEM/PEG against lung cancer cells A549. As illustrated in
Fig. 5C, all GEM, DCM-S-GEM, and DCM-S-GEM/PEG showed obvious
cytotoxicity to A549 cells at a concentration-dependent manner,
with the half-maximal inhibitory concentration (IC50) values of 0.443,
4
0
.511 and 0.465 μM, respectively. Furthermore, limited by the poor
The photostability of NIR fluorophores is a significant portion for
practical application especially in bioimaging.47,48 Hence, the time-
dependent fluorescence intensity of DCM-S-GEM and nanomized
prodrug DCM-S-GEM/PEG were detected after GSH incubation. As
shown in Fig. 4D and Fig. S12†, after exposure to high density light
solubility and biocompatibility, the small molecular prodrug DCM-S-
GEM cannot guarantee a satisfactory treatment efficiency compared
with free GEM. In contrast, owing to the good water dispersity and
excellent biocompatibility, the cell viability of nanomized DCM-S-
GEM/PEG is comparable with the free drug GEM. Obviously, both
DCM-S-GEM and DCM-S-GEM/PEG are low toxic to QSG-7701
because of the low GSH concentration in normal cells compared with
the strong cytotoxicity of free GEM, indicating negative side effects
(
Hamamatsu, LC8 Lightningcure, 300 W), the fluorescence intensity
of the cyanine dye ICG (FDA-approved NIR contrast agent) decreased
almost to vanishing point after 3 min, suggesting photobleaching
vulnerability. In contrast, nearly 50% florescence of DCM-S-
GEM/PEG and DCM-S-GEM were maintained after 10 min, indicative
of higher photostability. Consequently, the DCM-S-GEM/PEG and
DCM-S-GEM guarantee long-time tracing of chemotherapeutic agent
in vivo. Taken together, the nanomized NIR fluorescent prodrug
DCM-S-GEM/PEG is specially ‘turn on’ in tumor site with strong
photostability, ensuring the precise and long time tumor imaging.
(
Fig. 5D). The tumor-microenvironment sensitive disulfide linker and
improved solubility guaranteed high cytotoxicity of DCM-S-GEM/PEG
to cancer cells and low toxicity to normal cells.
The apoptosis property of DCM-S-GEM/PEG was detected using
typical flow cytometry (FCM) analysis with Annexin V-FITC/PI double
staining. The results showed that the percentage of cells with
phosphatidylserine externalization (the typical characteristics of
apoptosis) significantly increased in all GEM, DCM-S-GEM and DCM-
S-GEM/PEG groups compared to control group. As illustrated in Fig.
S13†, the apoptosis ratio increased in a concentration dependent
Enhanced treatment effect of DCM-S-GEM/PEG on lung tumor cells
(A549 cells) in vitro
The intracellular drug concentration of chemotherapeutic drug can manner that, the number are 8.00%, 26.5%, 50.0% and 64.7% after
4
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Fig. 6 Fluorescence images of cancer cells (A549 cells, A) and
normal cells (QSG-7701 cells, B) with the incubation of DCM-S-
GEM and DCM-S-GEM/PEG (10 μM) for 3 h, respectively. Red
channel (DCM-NH ) were obtained with excitation at 485 nm
2
and a long-path (630-690 nm) emission filter. The relative pixel
intensity of DCM-S-GEM and DCM-S-GEM/PEG in (C) tumor cells
Fig. 7 Bioimaging of tumor-bearing mice at various time (0,
0.5, 2, 4, 6, 24 h) after orthotopic injection of (A) DCM-S-GEM
and (B) DCM-S-GEM/PEG. Ex vivo images of major organs of
mice after injection of (C) DCM-S-GEM and (D) DCM-S-
GEM/PEG for 24 h. The color bars corresponding to the
detected fluorescence intensity. Note: The tumor-
microenvironment activable DCM-S-GEM/PEG is suitable for
real-time tracing drug release in vivo.
(
A549 cells) and (D) normal cells (QSG-7701cells). All images
share the same scale bar (30 μm). Note: The DCM-S-GEM/PEG
can real-time trace the drug release and realize tumor diagnose.
treating with 0, 0.5, 1, 5 μM GEM for 48 h, respectively. Similarly, the
apoptosis number are 21.2% (0.5 μM), 30.1% (1 μM) and 48.0% (5
μM) for DCM-S-GEM group, respectively. Limited by the cleavage
rate between the DCM and GEM, the apoptosis ratio of small prodrug
DCM-S-GEM was smaller than free GEM. Notably, the DCM-S-
tracing as well as tumor diagnosis. As a result, the DCM-S-GEM/PEG
platform can selectively lighting the tumor cells compared to the
normal cells as well as labeling the drug releasing, suggesting a good
tumor diagnosis and biodistribution evaluation probe.
GEM/PEG induced the comparable apoptosis ratio to unbounded Long-time and real-time monitoring of drug release in lung
GEM at the concentration of 1 μM (56.2%) and 5 μM (63.0%), which xenograft tumor model
are much higher apoptosis ratio at the concentration of 0.5 μM
As demonstrated in cell experiments, the nanomized DCM-S-
(
43.2%, Fig. S14†). The DCM-S-GEM/PEG induced more cell apoptosis
GEM/PEG is highly internalized by lung tumor cells and cleaved by
GSH in tumor cells, followed by fluorescence ‘turn on’ so as to trace
the drug release. Herein, the DCM-S-GEM and DCM-S-GEM/PEG
were orthotropic injected into the lung xenograft tumor of mice
than free GEM because of the stronger internalization. Compared to
the free GEM and prodrug DCM-S-GEM, the nanomized prodrug
DCM-S-GEM/PEG has higher internalization and toxicity to tumor
cells, owing to the good aqueous dispersity, excellent
biocompatibility and the micelle structure, suggesting improved model and monitored the fluorescence for 24 h to evaluate the drug
medicine efficiency.
tracing ability in vivo.
As illustrated in Fig. 7, no fluorescence was observed in both DCM-
S-GEM and DCM-S-GEM/PEG at 0 h post injection, suggesting no
release of free drug GEM or fluorescent agent DCM. The fluorescence
signal appeared after 0.5 h and the intensity become stronger with
time increasing, indicative of cleavage of -S-S- bond between the
GEM and DCM. Moreover, the fluorescence signal was still visualized
after 24 h, suggestive of long-time tracing ability. That means, the
nanomized prodrug DCM-S-GEM/PEG can long-time trace the drug
release in biological environment. The fluorescence signal spreaded
around the mice body in DCM-S-GEM treatment group after 4 h post
injection, suggestive of poor tumor retention (Fig. 7A). Whereas, the
fluorescence signal of the mice in DCM-S-GEM/PEG treatment group
still remains in the tumor site, guaranteed of longer tumor retention
time and possible stronger treatment effect (Fig. 7B) because the
bigger size not easily to be push out according to the EPR effect.
The longer tumor retention ability of NIR nanomized prodrug
DCM-S-GEM/PEG was also demonstrated by the ex vivo fluorescence
images of main organs of lung xenograft tumor mice after
Real-time monitoring of drug release in lung cancer cells
Due to the extraordinary internalization and GSH cleavable ability of
DCM-S-GEM/PEG in tumor cells, the selectively lighting up of tumor
tissues and drug tracing are proposed through releasing DCM in
tumor-microenvironment. Hence, the fluorescence images of cancer
cells (A549 cells) and normal cells (QSG-7701 cells) are obtained after
incubation of DCM-S-GEM/PEG to evaluate the drug release
visualization and tumor diagnosis. As shown in Fig. 6, the tumor cells
A549 became lighted on upon incubation with either DCM-S-GEM or
DCM-S-GEM/PEG (Fig. 6A) for 3 h. It is noted that, the nanomized
prodrug DCM-S-GEM/PEG exhibited 8-fold stronger fluorescence
intensity than prodrug DCM-S-GEM owing to the better water
dispersity, higher biocompatibility, and stronger internalization (Fig.
6C). In contrast to tumor cells, the fluorescence is neglected in
normal cells QSG-7701 (Fig. 6B and 6D) due to the low GSH
concentration, suggesting precise drug releasing and real-time
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orthotropic injection of DCM-S-GEM and DCM-S-GEM/PEG for 24 h.
As shown in Fig. 7, the fluorescence signal mainly appeared in liver
and kidney rather than in tumor for DCM-S-GEM treatment group
5
6
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(
Fig. 7C), compared with mainly in tumor for DCM-S-GEM/PEG
treatment group (Fig. 7D). This phenomenon claimed the limited side
effect of DCM-S-GEM/PEG compared with free DCM-S-GEM. These
experiments demonstrated that the nanomized prodrug DCM-S-
GEM/PEG is not only perfect for long-time and real-time tracing drug
in vivo, but also improving the therapeutic efficiency.
9
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8
931.
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bioavailability and tumor targeting ability. We exploited diblock
1
1
2 M. H. Lee, J. Y. Kim, J. H. Han, S. Bhuniya, J. L. Sessler, C. Kang
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polymer DSPE-mPEG to nanomize the tumor-microenvironment
active NIR fluorescent prodrug DCM-S-GEM/PEG for avoiding fast
metabolism, displaying longer tumor retention, and achieving
precise drug release in lung tumor. It is well demonstrated that DCM-
S-GEM/PEG exhibits higher photostability, excellent specificity and 14 M. Y. Marzbali and A. Y. Khosroushahi, Cancer Chemoth.
Pharm., 2017, 79, 637-649.
sensitivity towards GSH. Compared with the free prodrug DCM-S-
GEM, the nanomized prodrug DCM-S-GEM/PEG exhibits better
biocompatibility, faster cell endocytosis rate, and 8-fold higher
fluorescence imaging in tumor cells, showing significant toxicity to
A549 tumor cells owing to the improved better aqueous dispersity,
1
1
5 M. L. Pei, X. Jia, X. B. Zhao, J. G. Li and P. Liu, Carbohyd. Polym.,
2
018, 183, 131-139.
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addition, in vivo investigations demonstrate that DCM-S-GEM/PEG
leads to longer retention time than the free small prodrug DCM-S-
GEM in the tumor mice model due to the EPR effect. The longer
retention time in tumor tissue can increase the accumulation and the
subsequent cellular internalization, so as to realize better NIR
fluorescence imaging and sufficient cleavage of prodrug, generating
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9
,
1
1
2
higher
treatment
efficiency.
The
nanomized
tumor-
microenvironment active NIR prodrug DCM-S-GEM/PEG provides a
novel approach to realize real-time and long-time tracking the drug
delivery and activation process without systemic toxicity in vivo.
2
2
2
2
Conflict of interest
4 K. Z. Gu, Y. S. Xu, H. Li, Z. Q. Guo, S. J. Zhu, S. Q. Zhu, P. Shi, T.
D. James, H. Tian and W. H. Zhu, J. Am. Chem. Soc., 2016, 138,
There are no conflicts of interest to declare.
Acknowledgements
5
334-5340.
2
2
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5 X. H. Wang, Z. Q. Guo, S. Q. Zhu, Y. J. Liu, P. Shi, H. Tian and W.
H. Zhu, J. Mater. Chem. B, 2016, 4, 4683-4689.
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(
21788102, 21421004, 21636002 and 81602718), National key
Research and Development Program (2016YFA0200300),
Fundamental Research Funds for the Central Universities
(
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DOI: 10.1039/C8TB03188F
Nanomized
tumor-microenvironment
active NIR fluorescent prodrug for
ensuring synchronous occurrences of
drug releasing and fluorescence tracing †
a
b
,a
a
Qiang Li, Jun Cao, Qi Wang,* Jie Zhang,
a
a
,a
Shiqin Zhu, Zhiqian Guo and Wei-Hong Zhu*
The nanomized NIR fluorescent prodrug for solving its
bioavailability and tumor targeting ability. This
nanomized tumor-microenvironment active NIR prodrug
DCM-S-GEM/PEG provides a novel approach to realize
real-time and long-time tracking the drug delivery and
activation process without systemic toxicity in vivo.