The combined treatment method integrated photothermal therapy (PTT) with chemotherapy is extremely promising owing to the synergistic therapeutic effect as compared to single PTT or chemotherapy. To facilitate more novel and facile photothermal-chemotherapy drugs as well as promote controllable combination therapy, we have developed a mild and facile method to fabricate polymer polypyrrole (PPy)-doxorubicin (DOX) nanoparticles (NPs) as pH-responsive drug nanocarriers for synergistic photothermal-chemotherapy. In the nanoplatform, poly-L-lysine (PLL)-modified PPy serves as the photothermal material, and (DOX) molecules are adopted as the chemotherapy agent. Based on the cross-linking reaction of glutaraldehyde, DOX molecules are flexibly and efficiently assembled on the surface of PLL-modified PPy NPs. The obtained PPy-DOX NPs possess high photothermal effect, superior loading capacity of DOX, and controlled drug release behavior. The combination photothermal-chemotherapy based on PPy-DOX NPs has significantly enhanced the antitumor therapy effect. In general, the designed PPy-DOX NPs may be a potential drug delivery nanoplatform for cancer combination therapy.

Cancer has become a serious threaten to human health at present. Novel and efficient cancer treatment modalities are urgently needed for the development of anticancer research. Single-mode cancer therapy usually suffers from some issues, including drug tolerance, recurrence, metastasis, and limited treatment efficiency.1,2 To resolve these restrictions, the combination therapy has been exploited to obtain synergistic therapeutic effect of various antitumor strategies.3,4 On the other hand, many traditional therapeutic strategies for clinical cancer treatment (e.g., chemotherapy and radiotherapy) often cause serious side effects on normal tissues.5 An emerging therapeutic method, photothermal therapy (PTT), can convert light into heat for inducing tumor cells death. PTT has been widely developed to obtain maximum therapeutic effects by combining the traditional strategies, as well as reduce side effects caused by traditional strategies.6,7 The heat produced during the PTT process can enhance the intracellular delivery and release of chemotherapeutic agents to promote efficient chemotherapy.8,9 Therefore, the platform integration of PTT with chemotherapy holds great potential in optimizing synergistic effect of cancer treatment.

Currently, nanomaterial-based drug delivery platforms have displayed significant prospects in combination cancer treatment owing to its minimal invasiveness, specific selectivity, and low toxicity.10 In addition, nanomaterials usually accumulate in tumors by the passive targeting therapeutic strategy based on enhanced permeability and retention (EPR) effect.11,12 Accordingly, substantial efforts and crucial progress have been carried on with regard to nanomaterial-based systems for the combination tumor therapy, which enhance the anticancer efficiency owing to the integration of multiple tumoricidal mechanisms.8,13

PTT based on near infrared (NIR) light can penetrate into deep tumor tissues owing to its minimal absorption by biological tissues and, thus, show great potential in synergistic photothermal therapy and chemotherapy.14 Numerous NIR-absorbing nanomaterials were developed for photothermal agents, including organic nanomaterials,15,16 inorganic ones,17 and polymer nanoparticles.18 Thereinto, attributing to its excellent optical adsorption in NIR region and high capability of converting light into thermal energy,19 the conjugated polymer polypyrrole (PPy) was regarded as a promising photothermal agent. Moreover, PPy possesses the convenient synthesis route, favorable biocompatibility, and high photostability, which favor biomedical applications.20,21 At present, doxorubicin (DOX) is an attractive chemotherapy drug and is extensively used in clinical anticancer treatment. Benefiting from the acidic conditions in tumor microenvironment, the nanomedicines associated with pH-responsive activation and release of DOX have received considerable attention. Various drug nanocarriers integrating PPy with DOX have been gradually developed for the combination of thermal ablation and chemotherapy.22–25 Dox can be loaded into PPy nanoparticles via π–π stacking interaction26–28 and the hydrophobicity.23 Furthermore, in the presence of various intermediate modification, DOX can be loaded on the surface of PPy-based NPs by forming either a charge complex between the negatively charge and positively charged DOX29 or the acid-sensitive chemical bonds (e.g., Schiff bases).30,31 Overall, PPy-based therapeutic nanoplatforms, with good biocompatibility, excellent photothermal effect, and high drug loading capacity of DOX, are expected as promising nanomedicines for pH-responsive combined photothermal-chemotherapy.

Herein, we have developed a mild and facile route to synthesize PPy-DOX nanoparticles (NPs) as pH-responsive drug nanocarriers for synergistic photothermal-chemotherapy. The nanoplatform is composed of photothermal material poly-L-lysine (PLL)-modified PPy as the core, as well as the chemotherapy agent DOX on the surface. PLL-modified PPy NPs are prepared by the polymerization method, and PLL molecules can be randomly distributed on the PPy NP surface. Then, based on the cross-linking reaction of glutaraldehyde, doxorubicin (DOX) molecules were loaded on the surface of PLL-PPy NPs. The obtained PPy-DOX NPs possess high photothermal effect, superior loading capacity of DOX, and controlled drug release behavior. The antitumor efficacy has been greatly improved by the combined photothermal-chemotherapy of PPy-DOX NPs.

Doxorubicin hydrochloride (DOX HCl), pyrrole (MW = 67.09, 98%), polyvinylpyrrolidone (PVP, MW = 58 000), ferric chloride hexahydrate (FeCl3 · 6H2O, 99%), glutaraldehyde (GA, 50% aqueous solution), and ethyl alcohol were obtained from Aladdin Industrial Co., Ltd. (China). PLL hydrobromide (MW = 30–70 kDa) was from Sigma-Aldrich (China). Doubly distilled water was used in all experiments. All reagents were used as received without further purification.

The morphologies of NPs were detected by transmission electron microscopy (TEM, Hitachi JEM 1400EX). The hydrodynamic size was investigated through dynamic light scattering (DLS, Malvern Instruments). Optical properties of NPs were investigated through the UV–visible absorption spectra (UV-3600plus spectrophotometer, Shimadzu) and fluorescence emission spectra (F-4600 fluorescence spectrophotometer, Hitachi). Intracellular fluorescence imaging was performed using Nikon A1Rsi confocal laser scanning microscopy (Nikon).

The PPy-DOX NPs were prepared by a facile two-step strategy. First, the PLL-modified PPy NPs were synthetized by the modified polymerization method.32 Specifically, the stabilizer PVP (0.4 g) was dissolved in deionized water (10 ml). Then, 0.4 ml of FeCl3 · 6H2O (0.75 g/ml) was pumped into the PVP solution and stirred for ten minutes to provide a stable reaction environment for the subsequent oxidative polymerization. Afterward, 0.07 ml of pyrrole monomer was mixed with 1 ml of PLL (2.5 mg/ml) solution. The mixture of pyrrole and PLL was stirred for 10 min and quickly injected into the above reaction solution composed of PVP and FeCl3. The reactant gradually turned from slightly light yellow to black and was further incubated for 3 h. The synthetic product PLL-modified PPy NPs were purified with ethanol by centrifugation and then re-dispersed in 2 ml deionized water for following experiments.

DOX molecules were loaded on PLL-modified PPy NPs through the Schiff base reaction based on the aldehyde group of glutaraldehyde and the amino group of doxorubicin. In brief, 0.7 ml glutaraldehyde was added into the above PLL-modified PPy NPs and stood still for 2 h. Then, 2 mg of doxorubicin hydrochloride was added into the above reaction solution and stirred for 2 h. The final PPy-DOX NPs were centrifuged to remove unreacted reagents and then stored at −4 °C for subsequent experiments.

To confirm the loading efficiency of DOX within the PPy-DOX NPs, the calibration curves were obtained by detecting the absorption intensity of DOX at various concentrations. The linear calibration plot of DOX fits to the line Y = 0.009 59X + 0.008 71 (Y represents the absorbance intensity at 480 nm and X represents the DOX concentration). In this experiment, the absorption of DOX in PPy-DOX NPs were evaluated by measuring the absorption increase at 480 nm of the PPy-DOX NPs (0.4 mg/ml) and PPy NPs without DOX. Herein, Y is calculated as 0.493 and X is calculated as 50 ppm by the above standard curve. The loading efficiency of DOX within PPy-DOX NPs is obtained as about 12.6% as follows:

To evaluate the photothermal effect of PPy-DOX NPs, the NP aqueous dispersion at various concentrations (0–0.4 mg/ml) in a disposable cuvette was irradiated by an 808 nm laser at a power density of 0.26 W/cm2. Temperature changes were measured by a digital thermometer every 2 min. Phosphate buffered saline (PBS) was employed as the control group.

The DOX release from PPy-DOX NPs was evaluated by dialyzing the sample in PBS (0.1 mg/ml) at pH 5.0, 6.0, and 7.4 for various periods of time under 37 °C. The released DOX in the dialysate was collected and measured by the absorption spectra. The release of DOX was calculated according to standard curve Y = 0.009 59X + 0.008 71.

In terms of NIR-triggered DOX release, PPy-DOX NPs were exposed to 808 nm laser irradiation for 20 min (0.26 W/cm2) at the time points of 12 and 24 h in pH 5.0 and 6.0 solutions. Then, the samples were dialyzed and the released DOX was determined by absorption spectra immediately.

HeLa cells were cultured in DMEM cell medium supplemented containing 10% fetal bovine serum (FBS) at 37 °C under 5% CO2 environment. For cytotoxicity study, MTT assay was used to determine the viability of HeLa cells treated with PPy-DOX NPs. Specifically, HeLa cells were seeded into 96-well plates (1 × 104/well) until adherent and then incubated with PPy-DOX NPs at different concentration (0–20 mg/L) for 24 h. Afterward, 20 μl MTT solutions (5 mg/ml) were added into each well, and the cells were further incubated for 4 h. Finally, the media were taken away and 150 μl DMSO was added to each well. The absorbance intensity at 490 nm was determined by multifunction microplate reader (BioTek, ELx800 TM). The cell viability (%) relative to the control untreated cells was calculated by OD (test)/OD (control) × 100%.

The cellular uptake of PPy-DOX NPs was assessed by laser confocal microscope images of the cells treated with samples. HeLa cells were incubated in 35 mm confocal dishes and then treated with PPy-DOX NPs (10 mg/L) for 24 h. After rinsing three times with PBS, the cells were imaged to visualize the endocytosis of PPy-DOX NPs in HeLa cells.

For in vitro photothermal therapy and combined tumor therapy, HeLa cells were cultured at 1 × 104 cells/well in 96-well plates for 24 h. Then, the cells were treated with PPy NPs and PPy-DOX NPs at 10 mg/l and further incubated for 6 h. Then, the cells were continuously irradiated with 808 nm laser (0.26 W/cm2) for 5 min and further cultured for 24 h. Afterward, the cell viability was determined by MTT assay.

In addition, the cell live/dead staining method was also carried out to evaluate the viability of HeLa cells. Calcein acetoxymethyl ester (CAM) and propidium iodide (PI) were used to stain live cells (green color) and dead cells (red color), respectively. HeLa cells were incubated in 35 mm confocal plates for 24 h, and then, PPy NPs, DOX, or PPy-DOX NPs were added to the plates. After incubation for 6 h, the medium was replaced by fresh medium and was irradiated under the 808 nm laser for 5 min. The cells were incubated for further 24 h and stained with CAM (2 µM) and PI (4 µM) for 5 min. The laser scanning confocal microscope was used to observe the cell viability.

A facile two-step synthesis strategy was used to fabricate PPy-DOX NPs, which were composed of PPy as the core and DOX on the surface (Scheme 1). First, PLL-modified PPy nanoparticles were synthesized via a one-step aqueous dispersion polymerization of pyrrole. In the process of polymerization, PVP acts as a stabilizer and FeCl3 · 6H2O acts as an oxidant. The loading of PLL and the polymerization of pyrrole take place simultaneously, and PLL molecules randomly dispersed over the PPy NP surface. Subsequently, DOX molecules were loaded onto the surfaces of PLL-modified PPy NPs based on the formation of Schiff base. Herein, GA was adopted as a crosslinking agent to consume amino groups from both PLL and doxorubicin through Schiff base. The Schiff base can break in an acidic environment and has been designed as pH-response cancer drugs.33 Therefore, PPy NPs loaded with DOX were synthesized by an extremely facile and efficient synthetic procedure to achieve pH-response combinational photothermal-chemotherapy agents.

SCHEME 1.

Schematic illustration of PPy-DOX NPs for synergistic photothermal-chemotherapy.

SCHEME 1.

Schematic illustration of PPy-DOX NPs for synergistic photothermal-chemotherapy.

Close modal

The TEM images of PPy-DOX and PPy NPs are illustrated in Figs. 1(a) and S1. The PPy-DOX NPs show uniform spherical morphology with a particle size of about 70 nm [Fig. 1(b)], which is larger than that of PPy NPs. The hydrodynamic diameter of PPy-DOX and PPy NPs were determined to be about 180 nm [Fig. 1(c)] and 120 nm (Fig. S2) by dynamic light scattering. These hydrodynamic size data are higher than the data from TEM, probably resulting from the collapse of NPs during the drying process. To verify the effective loading of DOX, the absorption spectra of PPy-DOX NPs were determined [Fig. 1(d)]. Compared to PPy NPs, the absorption intensity of DOX-PPy NPs reveals an obvious improvement at 480 nm that corresponded to the characteristic absorption of DOX, proving the successful loading of DOX onto PPy-DOX NPs. The loading efficiency of DOX within PPy-DOX NPs is obtained as about 12.6% (Fig. S3). Furthermore, both PPy-DOX and single PPy NPs exhibit strong absorption in NIR region from 700 to 1100 nm, suggesting a great potential for high photothermal effects with 808 nm laser irradiation. The stability of nanomaterial-based drugs is critical for tumor treatment. As illustrated in Fig. S4, the PPy-DOX NPs exhibit a well-dispersed state, and the size has no significant change after being dispersed in PBS for three days, indicating the favorable stability.

FIG. 1.

(a) TEM image of PPy-DOX NPs. (b) Size distribution of PPy-DOX NPs from TEM image. (c) Hydrodynamic size of PPy-DOX NPs by dynamic light scattering. (d) The absorption spectra of PPy, DOX, and PPy-DOX NPs.

FIG. 1.

(a) TEM image of PPy-DOX NPs. (b) Size distribution of PPy-DOX NPs from TEM image. (c) Hydrodynamic size of PPy-DOX NPs by dynamic light scattering. (d) The absorption spectra of PPy, DOX, and PPy-DOX NPs.

Close modal

It is well known that PPy possesses excellent photothermal conversion properties and strong NIR absorbance. Thus, PPy-DOX NPs are expected to an attractive photothermal nanomaterial. Figure 2(a) demonstrates the NIR absorption spectra of PPy-DOX NPs solutions at various concentrations. The absorption peak intensity in the NIR region increases linearly as the concentration of PPy-DOX NPs (Fig. S5). Subsequently, the photothermal performance of PPy-DOX NPs at various concentrations is evaluated in Fig. 2(b). Obviously, the PPy-DOX NPs exhibit the concentration-dependent photothermal performance. The temperature of PPy-DOX NPs solutions increased significantly with the prolongation of laser irradiation time, while pure water revealed a little temperature change. The temperature of PPy-DOX NPs at the highest concentration (0.8 mg/ml) has increased by about 21 °C after laser irradiation for 10 min. The above results confirm that PPy-DOX NPs possess better photothermal performance.

FIG. 2.

(a) The absorption spectra of PPy-DOX NPs solution at various concentrations. (b) Temperature variation of PPy-DOX NP solution with different concentrations under 808 nm laser irradiation for 10 min at a power density of 0.26 W cm−2.

FIG. 2.

(a) The absorption spectra of PPy-DOX NPs solution at various concentrations. (b) Temperature variation of PPy-DOX NP solution with different concentrations under 808 nm laser irradiation for 10 min at a power density of 0.26 W cm−2.

Close modal

The release properties of DOX under distinct stimuli were investigated. Figure 3(a) shows the release of DOX at pH 5.0, 6.0, and 7.4. Under pH 7.4, the NPs can release 15% DOX at time point of 6 h. With the decrease in pH to 5.0, 61% DOX was released, which is beneficial to the release of the chemotherapeutic drugs from the acid tumor microenvironment. At the time point of 24 h, the cumulative DOX release runs up to about 73% under pH 5.0. The excellent release performance of DOX at the slightly acidic environment mainly is attributed to the enhanced water solubility of DOX after protonation.34 

FIG. 3.

DOX release from the PPy-DOX NPs at various pHs without (a) or with an 808 nm laser irradiation (b).

FIG. 3.

DOX release from the PPy-DOX NPs at various pHs without (a) or with an 808 nm laser irradiation (b).

Close modal

It is noticed that considerable part of the loaded DOX was still not released at pH 5.0 until 24 h. To improve the drug release, laser irradiation was adopted because the heat energy from laser irradiation usually can trigger and accelerate the release of drug loaded in NPs.28,35 As shown in Fig. 3(b), 90% DOX was released at pH 5.0 when the PPy-DOX NPs were irradiated by 808 nm laser for 40 min. The data suggest that the DOX release can be further enhanced at an acidic tumor microenvironment through the laser irradiation, which can reduce the side effect and enhance tumor therapeutic effect.

Cellular uptake of PPy-DOX NPs into the HeLa cells was evaluated by CLSM. Figure 4(a) displayed that the red DOX fluorescence from PPy-DOX NPs was mainly observed in the cytoplasm. The internalization mechanism of nanomaterial differs from that of the free DOX molecule.26 The passive diffusion mechanism is considered as the main rout of entering the cells for the free DOX, while mostly DOX-loaded nanomaterials enter into the cells via endocytosis. These results demonstrated the efficient endocytosis of PPy-DOX NPs, suggesting further application in cancer therapy.

FIG. 4.

(a) Cellular internalization of PPy-DOX NPs in HeLa cells by confocal laser scanning microscope (Scale bar, 50 μm). (b) Cell viability of HeLa cells treated with PPy, DOX, and PPy-DOX NPs at various concentrations. (c) Cell viability of HeLa cells after incubation with PPy and PPy-DOX NPs under laser irradiation. (d) Confocal fluorescence images of live/dead cell staining assay (scale bar, 50 μm).

FIG. 4.

(a) Cellular internalization of PPy-DOX NPs in HeLa cells by confocal laser scanning microscope (Scale bar, 50 μm). (b) Cell viability of HeLa cells treated with PPy, DOX, and PPy-DOX NPs at various concentrations. (c) Cell viability of HeLa cells after incubation with PPy and PPy-DOX NPs under laser irradiation. (d) Confocal fluorescence images of live/dead cell staining assay (scale bar, 50 μm).

Close modal

The biocompatibility of nanomaterials is crucial in biomedical applications. The cytotoxicity of PPy, DOX, and PPy-DOX NPs were investigated by using the MTT method. HeLa cells were cultured with the NPs at various concentrations for 24 h. The cell viabilities treated with PPy NPs were affected obviously when the concentration is up to 20 mg/l compared to the untreated group [Fig. 4(b)], showing the lower cytotoxicity of PPy NPs. In contrast, the cell viabilities cultured with DOX and PPy-DOX NPs significantly decreased with the increase in concentration. The PPy-DOX NPs have high cytotoxicity, mainly due to the release of DOX from NPs. Under 808 nm laser irradiation, the therapeutic effect of PPy-DOX NPs (10 mg/l) was further estimated. As shown in Fig. 4(c), the cell viabilities treated with PPy NPs under laser irradiation drop down to about 66% compared to the untreated group, which results from the photothermal effect of PPy. Obvious further increase in cell death was determined after treatment with PPy-DOX NPs, benefiting from the combined effect of photothermal therapy and chemotherapy. Chemotherapeutic drugs usually work by disrupting the tumor cell wall, blocking DNA replication, and blocking signaling. For photothermal therapy, high temperature destroys the cancer cell membrane and inhibits the synthesis of DNA and protein, inducing tumor cell death. These results confirmed the enhanced anticancer efficacy of chemotherapy assistant photothermal therapy based on PPy-DOX NPs.

To further investigate the anticancer performance of PPy-DOX NPs, the live/dead staining is adopted to detect the cells death visually. As illustrated in Fig. 4(d), the control cells without DOX, PPy NPs, and PPy-DOX NPs all show strong green fluorescence, revealing excellent cell viability. In the groups dealt with free DOX, some cells show red fluorescence, displaying that parts of cells were killed due to effective chemotherapy. As for the PPy-DOX NPs group, mostly cells display intense red fluorescence, demonstrating that HeLa cells are nearly completely killed. The cell death ratio of combined therapy is obviously higher than the DOX group and PPy group. These results further verified that the combined photothermal-chemotherapy based on PPy-DOX NPs has a remarkable efficacy in inducing cell death, which is consistent with the results derived by the MTT method.

In summary, a type of polypyrrole nanoparticles loaded with DOX were fabricated for chemotherapy-assistant photothermal therapy against cancer cells. The PLL shell provided linkage site for chemotherapy drug DOX. The obtained PPy-DOX NPs possess uniform particle size and spherical shape. The loading efficiency of DOX was optimized to be about 12.6%. This nanocomplex displays high photothermal effect under NIR 808 nm laser irradiation. The release of loaded DOX can be stimulated by acid and further improved by laser irradiation. Under the integration of chemotherapeutics DOX and photothermal drug PPy, the PPy-DOX NPs realized efficient synergistic therapy against tumor cells. In view of the facile synthesis, excellent photothermal effect, high DOX release, and improved anticancer efficiency, the PPy-DOX NPs show great potential application in synergetic photothermal-chemotherapy.

TEM images and size distribution of PPy nanoparticles without DOX, hydrodynamic size of PPy NPs measured by dynamic light scattering, absorption spectra of DOX at different concentrations, absorption peak intensity of DOX plotted against the concentrations, hydrodynamic size of PPy-DOX NPs after being dispersed in PBS for 3 days measured by dynamic light scattering, and linear fitting curve of the absorption intensity of PPy-DOX NP solution at 808 nm vs NP concentrations.

This work was financially supported by the National Natural Science Foundation of China (Grant Nos. 62071178 and 61605014) and the Open Fund of IPOC (BUPT) (Grant No. IPOC2022A06).

The authors have no conflicts to disclose.

Jinhua Liu: Formal analysis (equal); Funding acquisition (equal); Software (equal); Writing – original draft (equal). Guoren Zhu: Formal analysis (equal); Methodology (equal); Resources (equal); Validation (equal). Yuanan Liu: Formal analysis (equal); Validation (equal); Visualization (equal). Xiaohui Wang: Conceptualization (lead); Funding acquisition (equal); Investigation (lead); Project administration (lead); Supervision (lead); Writing – review & editing (lead).

The data that support the findings of this study are available within the article and its supplementary material.

1.
Z.
Xie
,
T.
Fan
,
J.
An
,
W.
Choi
,
Y.
Duo
,
Y.
Ge
,
B.
Zhang
,
G.
Nie
,
N.
Xie
,
T.
Zheng
,
Y.
Chen
,
H.
Zhang
, and
J. S.
Kim
,
Chem. Soc. Rev.
49
,
8065
(
2020
).
2.
3.
X. H.
Huang
,
I. H.
El-Sayed
,
W.
Qian
, and
M. A.
El-Sayed
,
J. Am. Chem. Soc.
128
,
2115
(
2006
).
4.
P.
Wust
,
B.
Hildebrandt
,
G.
Sreenivasa
,
B.
Rau
,
J.
Gellermann
,
H.
Riess
,
R.
Felix
, and
P. M.
Schlag
,
Lancet Oncol.
3
,
487
(
2002
).
5.
F.
Bray
,
J.
Ferlay
,
I.
Soerjomataram
,
R. L.
Siegel
,
L. A.
Torre
, and
A.
Jemal
, “
Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries
,”
Ca-Cancer J. Clin.
68
,
394
(
2018
).
6.
L.
Cheng
,
C.
Wang
,
L.
Feng
,
K.
Yang
, and
Z.
Liu
,
Chem. Rev.
114
,
10869
(
2014
).
7.
D.
Yu
,
Y.
Wang
,
J.
Chen
,
S.
Liu
,
S.
Deng
,
C.
Liu
,
I.
McCulloch
,
W.
Yue
, and
D.
Cheng
,
Acta Biomater.
137
,
238
(
2022
).
8.
J.
Beik
,
Z.
Abed
,
F. S.
Ghoreishi
,
S.
Hosseini-Nami
,
S.
Mehrzadi
,
A.
Shakeri-Zadeh
, and
S. K.
Kamrava
,
J. Controlled Release
235
,
205
(
2016
).
9.
F.
Pierini
,
P.
Nakielski
,
O.
Urbanek
,
S.
Pawlowska
,
M.
Lanzi
,
L.
De Sio
, and
T. A.
Kowalewski
,
Biomacromolecules
19
,
4147
(
2018
).
10.
C. Y.
Zhao
,
R.
Cheng
,
Z.
Yang
, and
Z. M.
Tian
,
Molecules
23
,
826
(
2018
).
11.
K.
Yang
,
H.
Xu
,
L.
Cheng
,
C.
Sun
,
J.
Wang
, and
Z.
Liu
,
Adv. Mater.
24
,
5586
(
2012
).
12.
X.
Wang
,
H.
Li
,
X.
Liu
,
Y.
Tian
,
H.
Guo
,
T.
Jiang
,
Z.
Luo
,
K.
Jin
,
X.
Kuai
,
Y.
Liu
,
Z.
Pang
,
W.
Yang
, and
S.
Shen
,
Biomaterials
143
,
130
(
2017
).
13.
N. R.
Datta
,
S. G.
Ordonez
,
U. S.
Gaipl
,
M. M.
Paulides
,
H.
Crezee
,
J.
Gellermann
,
D.
Marder
,
E.
Puric
, and
S.
Bodis
,
Cancer Treat. Rev.
41
,
742
(
2015
).
14.
Z. J.
Zhang
,
J.
Wang
, and
C. H.
Chen
,
Adv. Mater.
25
,
3869
(
2013
).
15.
R.
Ahmad
,
J.
Fu
,
N. Y.
He
, and
S.
Li
,
J. Nanosci. Nanotechnol.
16
,
67
(
2016
).
16.
B. P.
Jiang
,
B.
Zhou
,
Z. X.
Lin
,
H.
Liang
, and
X. C.
Shen
,
Chem. - Eur. J.
25
,
3993
(
2019
).
17.
X. H.
Wang
,
H. S.
Peng
,
W.
Yang
,
Z. D.
Ren
,
X. M.
Liu
, and
Y. A.
Liu
,
J. Mater. Chem. B
5
,
1856
(
2017
).
18.
M.
Chen
,
X.
Fang
,
S.
Tang
, and
N.
Zheng
,
Chem. Commun.
48
,
8934
(
2012
).
20.
B.
Guo
,
J.
Zhao
,
C.
Wu
,
Y.
Zheng
,
C.
Ye
,
M.
Huang
, and
S.
Wang
,
Colloids Surf., B
177
,
346
(
2019
).
21.
D.
Park
,
Y.
Cho
,
S. H.
Goh
, and
Y.
Choi
,
Chem. Commun.
50
,
15014
(
2014
).
22.
J.
Wang
,
J.
Han
,
C.
Zhu
,
N.
Han
,
J.
Xi
,
L.
Fan
, and
R.
Guo
,
Langmuir
34
,
14661
(
2018
).
23.
W.
Chen
,
J.
Wang
,
L.
Cheng
,
W.
Du
,
J.
Wang
,
W.
Pan
,
S.
Qiu
,
L.
Song
,
X.
Ma
, and
Y.
Hu
,
ACS Appl. Bio Mater.
4
,
1483
(
2021
).
24.
M.
Sun
,
J.
Guo
,
H.
Hao
,
T.
Tong
,
K.
Wang
, and
W.
Gao
,
Theranostics
8
,
2634
(
2018
).
25.
C.
Wang
,
H.
Xu
,
C.
Liang
,
Y.
Liu
,
Z.
Li
,
G.
Yang
,
L.
Cheng
,
Y.
Li
, and
Z.
Liu
,
ACS Nano
7
,
6782
(
2013
).
26.
L.
Wang
,
G.
Liu
,
Y.
Hu
,
S.
Gou
,
T.
He
,
Q.
Feng
, and
K.
Cai
,
Nanoscale
14
,
3097
(
2022
).
27.
S.
Geng
,
H.
Zhao
,
G.
Zhan
,
Y.
Zhao
, and
X.
Yang
,
ACS Appl. Mater. Interfaces
12
,
7995
(
2020
).
28.
T.
Xiao
,
W.
Hu
,
Y.
Fan
,
M.
Shen
, and
X.
Shi
,
Theranostics
11
,
7057
(
2021
).
29.
S.
Lan
,
W.
Xie
,
J.
Wang
,
J.
Hu
,
W.
Tang
,
W.
Yang
,
X.
Yu
, and
H.
Liu
,
J. Nanopart. Res.
20
,
300
(
2018
).
30.
J.
Mao
,
Y.
Li
,
T.
Wu
,
C.
Yuan
,
B.
Zeng
,
Y.
Xu
, and
L.
Dai
,
ACS Appl. Mater. Interfaces
8
,
17109
(
2016
).
31.
X.
Zeng
,
G.
Liu
,
W.
Tao
,
Y.
Ma
,
X.
Zhang
,
F.
He
,
J.
Pan
,
L.
Mei
, and
G.
Pan
,
Adv. Funct. Mater.
27
,
1605985
(
2017
).
32.
X. H.
Wang
,
X. Q.
Chen
,
H. S.
Peng
,
X. F.
Wei
,
X. J.
Wang
,
K.
Cheng
,
Y. A.
Liu
, and
W.
Yang
,
J. Mater. Chem. B
8
,
1033
(
2020
).
33.
S. S.
Han
,
Z. Y.
Li
,
J. Y.
Zhu
,
K.
Han
,
Z. Y.
Zeng
,
W.
Hong
,
W. X.
Li
,
H. Z.
Jia
,
Y.
Liu
,
R. X.
Zhuo
, and
X. Z.
Zhang
,
Small
11
,
2543
(
2015
).
34.
L.
An
,
Y. Y.
Wang
,
J. M.
Lin
,
Q. W.
Tian
,
Y. X.
Xie
,
J. Q.
Hu
, and
S.
Yang
,
ACS Appl. Mater. Interfaces
11
,
15251
(
2019
).
35.
J.
Zhou
,
Y. N.
Han
,
Y.
Yang
,
L.
Zhang
,
H.
Wang
,
Y. T.
Shen
,
J. H.
Lai
, and
J. H.
Chen
,
ACS Appl. Mater. Interfaces
12
,
23311
(
2020
).

Supplementary Material