J-Aggregate squaraine nanoparticles with bright NIR-II fluorescence for imaging guided photothermal therapy

Article abstract of DOI:10.1039/c8cc08096h

We introduce a novel strategy to enhance the fluorescence brightness of organic-molecule-based nanoparticles in the second near-infrared window (NIR-II, 1000-1700 nm) by fabricating J-aggregate nanoparticles SQP-NPs_(J). Our prepared J-aggregate

Full text of DOI:10.1039/c8cc08096h

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J-Aggregate squaraine nanoparticles with bright
NIR-II fluorescence for imaging guided
photothermal therapy†
Cite this: Chem. Commun., 2018,
4, 13395
5
Received 10th October 2018,
Accepted 7th November 2018
a
a
a
b
a
a
Pengfei Sun, ‡ Qi Wu,‡ Xiaoli Sun, Han Miao, Weixing Deng, Wansu Zhang,
a
c
Quli Fan * and Wei Huang
DOI: 10.1039/c8cc08096h
rsc.li/chemcomm
1
0
We introduce a novel strategy to enhance the fluorescence bright- lanthanide-based downconversion nanoparticles (DCNPs).
ness of organic-molecule-based nanoparticles in the second near- However, the long-term toxicological profile of inorganic nano-
1
1
infrared window (NIR-II, 1000–1700 nm) by fabricating J-aggregate materials still requires further assessment. In contrast, organic
nanoparticles SQP-NPs(J). Our prepared J-aggregate nanoparticles probes, including conjugated polymers and small molecules,
SQP-NPs(J) show an emission maximum near 1100 nm, and the emission enjoy considerable advantages due to their relatively good bio-
intensity is 4.8-fold higher than that of H-aggregate SQP-NPs(H). In compatibility and easy chemical design, which provide high
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addition, SQP-NPs(J) can be used for NIR-II imaging guided photo- fluorescence quantum yields. However, most of the organic
thermal therapy on MCF-7 tumor-bearing mice due to the fact that molecules are inherently hydrophobic, leading to unavoidable
SQP-NPs(J) have highly effective photothermal properties, which are molecular aggregation and severe fluorescence quenching when
significant for precise tumor diagnostics and treatments.
loaded into hydrophilic polymer matrices or modified with ionic
groups to realize water-solubility. To overcome this limitation,
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Optical imaging can afford high sensitivity and spatial resolu- recently, Liu’s group introduced the aggregation-induced emis-
tion for early diagnosis of tumors and imaging-guided sion (AIE) strategy to solve the fluorescence quenching problem
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therapy.
Nevertheless, optical imaging is limited because in the NIR-II region. However, in the AIE strategy, the AIE
photon scattering by biological tissues at the conventional electron donor group must be strictly selected, which limited its
NIR-I (700–900 nm) ranges leads to small imaging penetration wide range of applications. Therefore, there are still needs for
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depths. In the second near-infrared (NIR-II) window ranges the development of new strategies to solve the molecular aggre-
from 1000 to 1700 nm, light scattering and photon absorption gation and fluorescence quenching problems.
by biological moieties are minimal, which allow for deeper
Among many kinds of molecule aggregations, J-aggregation
penetration, micro-scale resolution and an improved signal to is a type of ordered aggregation that results in a ‘‘slipped stack’’
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,6
15
background ratio in fluorescence imaging.
Thus, NIR-II arrangement (head–tail).
Such J-aggregation molecular
fluorescence imaging shows great promise in meeting the present assembly of dyes results in strong red-shifted absorption and
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unsatisfied optical imaging requirements.
enhanced fluorescence intensities. To date, a few studies have
Currently, many inorganic NIR-II imaging contrast agents reported that J-aggregates can be applied in visible and NIR-I region
have been developed including single-walled carbon nanotubes fluorescence imaging. Research by W u¨ rthner and co-workers
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9
(
SWNTs), transition-metal sulfide/oxide semiconductors and revealed that the J-aggregates of perylene bisimide derivatives
17
exhibit far-red fluorescence and high quantum yields. Xu and
co-workers also have demonstrated that pyrrolopyrrole cyanine
derivative (PCBF) J-aggregates assembled with an amphi-
philic block co-polymer (PCL-b-POEGMA) can enhance the
a
Key Laboratory for Organic Electronics and Information Displays & Jiangsu Key
Laboratory for Biosensors, Institute of Advanced Materials (IAM), Jiangsu National
Synergetic Innovation Center for Advanced Materials (SICAM), Nanjing University
of Posts & Telecommunications, 9 Wenyuan Road, Nanjing 210023, China.
E-mail: iamqlfan@njupt.edu.cn
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light-emission intensity of nanoparticles in the NIR-I region.
Therefore, controlling the J-aggregates of NIR-II molecules may
be a more suitable method to increase the NIR-II fluorescence.
However, to the best of our knowledge, no one has ever tried to
control the J-aggregates of NIR-II molecules to realize brighter
NIR-II fluorescence imaging.
b
c
School of Materials Science and Engineering, Georgia Institute of Technology,
Atlanta, GA 30332, USA
Shaanxi Institute of Flexible Electronics (SIFE), Northwestern Polytechnical
University (NPU), 127 West Youyi Road, Xi’an 710072, Shaanxi, China
Electronic supplementary information (ESI) available. See DOI: 10.1039/
†
In our work, we exploited one NIR-absorptive dye, SQP
(bispyrrole-sq-bispyrrole), as the model molecule to prepare
c8cc08096h
These authors contributed equally to this work.
‡
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Scheme 1 Synthesis route to SQP.
J-aggregate nanoparticles. Squaraine dyes are widely used
fluorescence dyes because they can easily narrow the bandgap to
provide strong light-emission in a long wavelength region and have
19
great capability to form J-aggregates in aqueous solution. After
encapsulating SQP into an amphiphilic copolymer F-127 (PEG-b-
PPG-b-PEG), J-aggregate SQP-NPs(J) were obtained. Meanwhile, we
also prepared H-aggregate (molecules form in a ‘‘vertical stack’’
arrangement, which exhibit a blue-shifted absorption band and
20
weakened fluorescence ) SQP-NPs by changing the assembly
(H)
method. As expected, the NIR-II emission intensity of the SQP-
NPs(J) was 4.8-fold higher than that of the SQP-NPs(H) at the same
concentration. After applying the two NPs in in vitro and in vivo
NIR-II fluorescence imaging, the brighter signal on MCF-7 cells and Fig. 1 (a) The absorption spectra and fluorescence emission spectra of
SQP in THF. The fluorescent emission spectra were obtained under
the sharper image quality of healthy mice proved the superiority
7
30 nm excitation; (b) DLS data of the SQP-NPs(J) and SQP-NPs(H) in
water. (c) UV-vis-NIR absorption spectra of the SQP-NPs(J) and SQP-NPs(H)
d) NIR-II fluorescence emission spectra of the SQP-NPs(J), SQP-NPs(H) and
of the SQP-NPs(J). In addition, the NIR-absorbing SQP-NPs(J)
can convert laser energy into thermal energy, which endows the
SQP-NPs(J) with efficient photothermal therapy (PTT) performance.
;
(
À1
IR1061 NPs at the same concentration of 10 mg mL under 808 nm
Therefore, the SQP-NPs(J) were applied in NIR-II imaging guided excitation; (e) NIR-II fluorescence images of the SQP-NPs(J) and
SQP-NPs(H) at different depths; (f) NIR-II fluorescence intensity of the
photothermal therapy (Scheme 1).
SQP-NPs(J) and SQP-NPs(H) at different depths.
SQP was prepared based on previous literature reports, with
(E,E)-1,4-bis[2-(1-methylpyrrol-2-yl)vinyl]-2,5-didodecyloxybenzene
(
BP) and squaric acid used as the electron donor and acceptor, The TEM images (Fig. S10, ESI†) show that both SQP-NPs(J) and
21
respectively. Meanwhile, each benzene ring was modified with SQP-NPs(H) have stable spherical shapes in water with sizes of
two alkyl (dodecyl) side chains for enhancing the solubility of the approximately 82 and 86 nm. These results revealed that the
SQP in organic solvents. Condensation of squaric acid with two solvent selection of SQP does not have an apparent effect on the
equivalents of BP at 112 1C afforded the target SQP in 13% yield. size of the nanoparticles.
The chemical structure of the SQP was characterized using
H NMR, matrix-assisted laser desorption/ionization time-of- spectra can be used to characterize the J-aggregates or H-aggregates
flight mass spectrometry and FT-IR spectroscopy (Fig. S4, S6 of dyes. As shown in Fig. 1c, the UV-vis-NIR absorption spectra of
and S7, ESI†). The photophysical properties of SQP are presented the SQP-NPs(J) show one blue-shifted absorption band at 695 nm
in Fig. 1a, where the normalized UV-vis-NIR absorption spectra and another sharp, red-shifted absorption band at 901 nm. Similar
It is well known that a red-shift or blue-shift in the absorption
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23,24
of SQP show a sharp absorption band at 766 nm in THF to many other research studies,
this sharp, red-shifted absorp-
tion band at 901 nm suggests that SQP has formed J-aggregates in
Fig. S9, ESI†). The fluorescence emission spectra of SQP the SQP-NP(J) aqueous solution. In contrast, the SQP-NPs(H) only
display main emission peaks at 794 nm and two shoulder exhibit a blue-shifted absorption band at 670 nm, which hints the
À1
À1
with a high mass extinction coefficient of 126.3 L g cm
(
peaks at 855 and 1000 nm.
major H-aggregates of SQP in the SQP-NPs(H) aqueous solution.
To verify the ability of J-aggregate nanoparticles in enhancing
With the SQP dye in hand, an amphiphilic copolymer (PEG-
b-PPG-b-PEG, F-127) was introduced to prepare water-soluble the fluorescence intensity, the fluorescence emission spectrum
SQP nanoparticles by a co-precipitation method. In the self- of the SQP-NPs(J) was investigated and compared with that
assembly process, the SQP dye was dissolved in DCM or THF to of the SQP-NPs(H). As presented in Fig. 1d, the SQP-NPs(J) and
prepare J-aggregate nanoparticles (SQP-NPs ) and H-aggregate SQP-NPs(H) have the same broad emission band extending to
(J)
nanoparticles (SQP-NPs(H)). As we can see from dynamic light 1400 nm, with two peaks at 947 nm and 1036 nm. However, the
scatting (DLS) analysis (Fig. 1b), the hydrodynamic radii (R ) were emission intensity of the SQP-NPs(J) was 4.8-fold higher than
3 Æ 2 and 52 Æ 2 nm, and the polydispersity indices (PDI) were that of the SQP-NPs(H) at the same concentration, proving the
.27 and 0.26 for the SQP-NPs(J) and SQP-NPs(H), respectively. superiority of the J-aggregate SQP-NPs(J). Apparently, it was the
h
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J-aggregates abundantly formed in the SQP-NPs(J) that contributed indicated by the circled regions (labelled 1–2) on the mice
to the enhanced NIR-II emission. In addition, the emission brain, and the NIR-II fluorescence intensity (Fig. 2c) of region
intensity of the SQP-NPs(J) was stronger about 35-fold compared 1 was 4.7-fold higher than that of region 2, illustrating the
to that of the reported NIR-II probe IR1061 NPs, suggesting the superior brightness of the SQP-NPs(J) in vivo. Furthermore, the
exceptionally brighter NIR-II fluorescence of the SQP-NPs .
magnified images of the partial vessel regions (labelled 3–4) on
For further detecting the maximum imaging depth of the the mice body also demonstrated that the vasculature of the
SQP-NPs(J) and comparing the brightness with the SQP-NPs(H) mice injected with the SQP-NPs(J) was much clearer than that of
(J)
,
two tubes filled with the two nanoparticles were placed under the SQP-NPs(H), suggesting that the SQP-NPs(J) can indeed reach
pork with different thicknesses, and the NIR-II images were significantly sharper image quality in vivo.
acquired, respectively (Fig. 1e and f). The images of the SQP-
NPs(J) were clearly much brighter than those of the SQP-NPs(H)
Normally, NIR-absorbing materials can convert laser energy
2
5,26
,
into heat.
Therefore, the photothermal behaviour of the
reflecting the benefit of the J-aggregates. Fig. 1f shows the SQP-NPs(J) was studied. The temperature of the SQP-NPs at
(J)
À1
emission intensity of the two SQP-NPs at different depths. It is a concentration of 100 mg mL was increased from 24 1C
obvious that the SQP-NPs(J) show a much larger fluorescence to 67 1C, while the temperature of deionized water was only
intensity compared to that of the SQP-NPs(H) at 2–6 mm of depth. increased by 3 1C under the same conditions (Fig. S13a, ESI†).
Overall, these results suggest that the NIR-II imaging depth of Furthermore, the photothermal conversion efficiency of the
the two SQP-NPs all reached up to approximately 8 mm.
To study the fluorescence enhancement effect of the SQP- for effective photothermal therapy.
NPs(J) in vitro, the NIR-II fluorescence images of MCF-7 cells To analyse the in vitro PTT efficacy of the SQP-NPs , an MTT
SQP-NPs(J) was 36% (Fig. S13d, ESI†), which is high enough
(J)
incubated with the two NPs were measured. As shown in assay, confocal laser scanning microscopy (CLSM) and flow
Fig. 2a, the NIR-II image of the MCF-7 cells incubated with the cytometry analysis were used to monitor the cytotoxicity to
SQP-NPs(J) was obviously brighter than that of the SQP-NPs(H)
Accordingly, the fluorescence intensity of the MCF-7 cells incu- incubated with 100 mg mL
.
MCF-7 cells. As shown in Fig. S14a (ESI†), the MCF-7 cells only
À1
SQP-NPs(J) exhibited 92% cell
bated with the SQP-NPs(J) was 4.1-fold higher than that of the viability, but showed only 27% cell viability after LED lamp
SQP-NPs(H), reflecting that the SQP-NPs(J) can realize brighter irradiation. In the CLSM images (Fig. S14b, ESI†), the MCF-7
NIR-II imaging at the cellular level.
cells only treated with the SQP-NPs(J) or LED lamp irradiation
Next, in vivo NIR-II fluorescence imaging measurements showed green fluorescence. In contrast, the MCF-7 cells treated
were conducted on healthy BALB/c mice for further comparing with the SQP-NPs(J) and LED lamp irradiation showed red fluores-
the imaging capacity of the two nanoparticles. From Fig. 2b, we cence, which reflected obvious cell death. As shown in the flow
can distinctly see that the brightness of the cerebral vessels of cytometry image (Fig. S14c, ESI†), the amount of late apoptotic cells
the mice injected with the SQP-NPs(J) was much higher than increased from 0.56% to 96.8% after LED lamp irradiation when
that of the SQP-NPs(H). Then, we captured a part of the images incubated with the SQP-NPs(J). These results testify that SQP-NPs(J)
hold great promise to act as PTT agents for tumor treatment.
To further explore the potential application of SQP-NPs(J) in
NIR-II imaging-guided therapy, a MCF-7 tumor bearing mouse
was treated with SQP-NPs(J) and observed under a NIR-II fluores-
cence imaging system. The magnified images of the tumor
regions are presented in Fig. 3a. Shortly after the intravenous
administration of the SQP-NPs(J), the blood vessels of the tumor
could be clearly identified with respect to the surrounding
background tissue. However, as the time increased, the NIR-II
fluorescence signal of the vasculature gradually weakened, but a
bright signal inside the tumor enhanced. 24 h after injection, the
quantified tumor-to-normal tissue ratio (T/NT) (Fig. 3b) reached
the maximum value (3.54 Æ 0.2) and the vessel signal completely
vanished. These subtle signal changes were not possible to be
observed in NIR-I window imaging (Fig. S22, ESI†), which
illustrates that NIR-II imaging can provide more precise infor-
mation about biological diagnoses.
After precise NIR-II fluorescence imaging and diagnosis,
Fig. 2 (a) NIR-II fluorescence intensity of the MCF-7 cells incubated SQP-NPs(J) were further used to conduct in vivo photothermal
with SQP-NPs(J) and SQP-NPs(H). Inset: NIR-II fluorescence images of the
therapy on MCF-7 tumor-bearing nude mice. These mice were
randomly divided into four groups (six mice per group):
(i) saline, (ii) saline with LED lamp treatment, (iii) SQP-NPs(J),
MCF-7 cells incubated with SQP-NPs(J) and SQP-NPs(H); (b) NIR-II fluores-
cence images of mice brain and mice body after intravenous administration
À1
with 2 mg mL SQP-NPs(J) and SQP-NPs(H); (c) magnified images of the
and (iv) SQP-NPs(J) with LED lamp treatment. 24 h after
intravenous injection with SQP-NPs(J), the tumor region was
circled regions (labelled 1–2) and vessel regions (labelled 3–4), as well as the
average fluorescence intensity of regions 1–2.
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This work was financially supported by the National Natural
Science Foundation of China (No. 21604042, 21674048 and
21574064), the Synergetic Innovation Center for Organic
Electronics and Information Displays, the 333 project of
Jiangsu province (No. BRA2016379), and the Primary Research
&
Development Plan of Jiangsu Province (BE2016770).
Conflicts of interest
There are no conflicts to declare.
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