English

• Phototheranostics, as a novel cancer treatment technology, has the advantages of precise spatiotemporal control, efficient diagnosis and treatment, non-invasive, negligible drug resistance, and weak side effects. Upon laser irradiation, phototheranostic agents generate fluorescence for imaging guided diagnosis of tumors [1,2]. These agents also produce cytotoxic reactive oxygen species (ROS) through photodynamic therapy (PDT) or convert photo-energy into heat by photothermal therapy (PTT) to kill tumors [3,4]. Red-shift in absorption and fluorescence spectra, enhancement of ROS generation capability, and increasing photothermal conversion efficiency (PCE) of phototheranostic agents can improve imaging quality and phototheranostic efficacy. Moreover, the combination of PDT and PTT can further improve the therapeutic efficacy because increasing temperature in PTT can accelerate blood flow rate and alleviate oxygen depletion in tumor tissue to improve PDT efficacy [5,6]. Meanwhile, the ROS generation in PDT is advantageous to PTT as it can destroy the structure and biological function of heat shock proteins [7].

  Porphyrins have been clinically approved for use in PDT due to their high singlet oxygen quantum yield (¹O₂ QY) [8,9]. However, their poor water-solubility and large planar π-conjugate structure triggers the formation of aggregates in water through tight intermolecular π-π stacking [10,11], which results in undesirable aggregation-induced quenching (ACQ) that inhibits ROS generation and fluorescence emission [12]. Recent reports suggested that indocyanine green (ICG) can be used as phototheranostic agent for fluorescent imaging-guided PDT and PTT [13,14]. However, its poor photostability, low PCE and ¹O₂ QY lead to decreasing phototherapeutic efficacy [15,16]. Thus, the development of novel phototheranostic agents with strong near-infrared (NIR) absorption and fluorescence, high ¹O₂ QY and PCE, and excellent photostability is urgent.

  Electron-donating and electron-withdrawing groups can be simultaneously introduced into one molecule to construct a donor-acceptor-donor (D-A-D) system, causing a narrow band gap of molecule, thus this is a simple and feasible method for preparing NIR phototheranostic agents [17–19]. For example, Zhao et al. used 4,4′-(2,2-diphenylethene-1,1-diyl)bis(N,N-diphenylaniline) and benzobisthiadiazole as the electron-donating and electron-withdrawing groups, respectively, for constructing an aggregation-induced emission (AIE) fluorogen DDTB (λ_abs = 650 nm, λ_Em = 973 nm), and then prepared it into water-soluble nanoparticles (NPs). The fluorescence and ¹O₂ QYs of DDTB NPs are increased to 0.96% and 1.05%, respectively, which are higher than that of DDTB in molecularly dispersed state. However, the vibration relaxation of DDTB was restricted in aggregates, leading to a low PCE of 30.7% [20]. Aggregate-induced decrease in energy gap between the excited singlet (S₁) and triplet (T₁) state is a promising strategy that can solve decreasing ¹O₂ QY caused by ACQ effect [21,22]. Guo et al. reported a Ru(Ⅱ)-arene compound (λ_abs = 729 nm, λ_Em = 1050 nm) that has no photosensitive activity in molecularly dispersed state but can generate ¹O₂ with a QY of 16.4% in aggregate state [23]. Theoretical calculation results suggested that the enhancement of intersystem crossing (ISC) in aggregate state is caused by decreasing energy gap from S₁ to T₁ (ΔE_ST). However, its PCE is only 24.2%, making it impossible to achieve ideal PTT efficacy.

  The effective intramolecular motion facilitates vibration relaxation, enhancing the PCE of molecule [24]. Chen et al. maximized the intramolecular motion of Py-PS by introducing multiple stretchable and vibrational groups, achieving a high PCE of 86.8% [25]. Peng et al. synthesized tfm-BDP by introducing trifluoromethyl at the meso site of boron-dipyrromethene (BODIPY). Due to the rotation-free mechanism of trifluoromethyl in the aggregation, the PCE of tfm-BDP reaches to 88.3% [26]. In addition, it has been reported that the spatial configuration of excited fluorogen with D-A structure is changed due to intramolecular rotation, resulting in twisted intramolecular charge transfer (TICT) effect [27,28], in which the fluorescence QY of fluorogen decreased and the absorbed photo-energy is released through vibration relaxation. Therefore, TICT effect can significantly improve the photothermal conversion ability of photothermal agents [29]. Tang et al. proposed an “intramolecular motion in aggregate” strategy and designed four compounds (NIR6, NIRb6, NIRb10, and NIRb14) with NIR absorption by introducing molecular rotors and alkane chains into D-A molecules [30]. The long alkane chains served as steric hindrance groups that prevent tight intermolecular π-π stacking [31]. Among all compounds, NIRb14 has the highest PCE of 31.2% due to obvious fluorescence quenching and stable TICT in aggregates. Recently, we designed and synthesized a hemicyanine dye (M1, λ_abs = 714 nm, λ_Em = 1005 nm) with D-π-A structure [32]. The tert-butyl group on cyclohexene and two methyl groups on furan stabilize the rotation of M1 by effectively preventing tight intermolecular π-π stacking [33], in turn increasing the photothermal conversion ability of M1 NPs (PCE = 77.5%). Thus, the combination of “aggregation-induced decrease of ΔE_ST” and “intramolecular motion in aggregate” is an advanced strategy for constructing phototheranostic agents with excellent ROS generation capability and high PCE.

  Based on the above considerations, herein, both aggregation-induced increase in singlet-triplet ISC and TICT effect were simultaneously used to develop phototheranostic agents with strong NIR absorption, aggregation-induced enhancement of ¹O₂ QY and PCE. We selected N,N-diethylaniline and triphenylamine as electron-donating groups, and the core structure of BODIPY as electron-withdrawing group to design and synthesize a D-A-D structured BODIPY derivative (named B-2TPA) with NIR absorption and fluorescence peaks at 729 and 766 nm, respectively, and TICT effect (Fig. 1a). Under 735 nm laser irradiation, B-2TPA had low ¹O₂ QY of 1.8% and PCE of 39.8%. After being assembled with DSPE-PEG-2000, these B-2TPA NPs exhibited excellent water dispersibility, red-shifted absorption and fluorescence spectra. Moreover, the intermolecular aggregation in B-2TPA NPs effectively decreased the ΔE_ST and significantly improved the ISC efficiency, resulting in an increase of ¹O₂ QY to 6.7%. In addition, obvious fluorescence quenching and stable TICT in B-2TPA NPs also enhanced the PCE to 60.1%. Furthermore, B-2TPA NPs had excellent photostability, and strong NIR fluorescence and photoacoustic (PA) signals. Both in vitro and in vivo experiments demonstrated that B-2TPA NPs were suitable for synergistic PDT and PTT, as well as in dual-mode NIR fluorescence and PA imaging of tumor (Fig. 1b).

  Figure 1

  

  Figure 1.  (a) Preparation of B-2TPA NPs, and molecular structure and main optical properties of B-2TPA and B-2TPA NPs. (b) Application of B-2TPA NPs in NIR fluorescence and PA imaging-guided synergistic PDT and PTT of tumor.

  DownLoad: Full-Size Img PowerPoint

  Phototheranostic agents with an aggregation-induced increase in ¹O₂ QY and good photothermal conversion ability are greatly advantageous to cancer therapy. Herein, B-2TPA for phototheranostic application has the following advantages: (1) The introduction of strong electron-donating groups (N,N-diethylaniline and triphenylamine) into the core structure of BODIPY to construct a D-A-D system allows the absorption and fluorescence wavelengths of B-2TPA to locate in the NIR region [34]. (2) As shown in optimized ground state (S₀) geometry figure of B-2TPA (Fig. 2a), the large dihedral angle between triphenylamine and BODIPY causes large non-planar π-conjugated structure of B-2TPA, which not only avoids the tight face-to-face π-π stacking in water [35], but also triggers the formation of B-2TPA NPs through simple intermolecular aggregation, resulting in decreasing energy gap between S₁ and T_n, thus allowing the enhancement of ISC efficiency, in turn increasing the ¹O₂ QY of B-2TPA NPs; and (3) B-2TPA has high liposolubility and multiple rotational structures; thus, in the hydrophobic microenvironment formed by long-chain alkanes of DSPE-PEG-2000, the PCE of B-2TPA aggregates is enhanced due to obvious fluorescence quenching and stable TICT effect [24]. The synthetic route of B-2TPA is shown in Scheme S1 (Supporting information). All the obtained compounds were characterized by ¹H nuclear magnetic resonance (NMR), ¹³C NMR (Figs. S1–S8 in Supporting information), and high-resolution mass spectrometry (HRMS).

  Figure 2

  

  Figure 2.  (a) Optimized ground state (S₀) geometries of B-2TPA. (b) Normalized visible (vis)-NIR absorption and fluorescence spectra of B-2TPA in 1,4-dioxane. (c) Line-fitting of a plot between Stokes shift and Δf. (d) Irradiation time-dependent fluorescence change of SOSG in B-2TPA solution. (e) Irradiation time-dependent temperature variation of B-2TPA solution.

  DownLoad: Full-Size Img PowerPoint

  The absorption and fluorescence spectra of B-2TPA were measured, and the results are shown in Fig. 2b. Due to the strong ICT effect and large π-conjugated structure, the absorption and fluorescence wavelengths of B-2TPA in 1,4-dioxane were 729 and 766 nm, respectively. Upon photoexcitation, the molecules with TICT effect first are in a locally excited state, and then undergo structural twist to enter the TICT state, inducing the locally excited-state and TICT emission of molecule in moderately polar solvents can be observed [36]. Thus, the trailing of fluorescence spectrum of B-2TPA (800–900 nm) in dioxane is caused by its TICT effect.

  To further demonstrate the TICT effect of B-2TPA, the influence of solvent on the optical properties of B-2TPA was studied. As shown in Fig. S9 (Supporting information), with the solvent polarity increased, the absorption and fluorescence wavelengths of B-2TPA were red-shifted from 717 and 759 nm (in diethyl ether) to 749 and 831 nm (in DMSO), respectively, while its fluorescence intensity decreased from 585.5 K (in diethyl ether) to 77.0 K (in DMSO), which is indicative of obvious solvatochromic effect of B-2TPA [37]. To quantitatively evaluate the effect of solvent polarity on fluorescence behavior, the Lippert-Mataga equation, which directly represents the relationship of Stokes shift of fluorogen in different solvents between solvent polarity parameters (Δf) was used [38], and the results are displayed in Fig. 2c and summarized in Table S1 (Supporting information). The Stokes shift gradually increased with increasing solvent polarity parameter, indicating the significant TICT effect of B-2TPA. Additionally, the molar extinction coefficient of B-2TPA was 1.15 × 10⁵ L mol⁻¹ cm⁻¹. Using zinc phthalocyanine (Zn-Pc) as a reference, the fluorescence QY of B-2TPA in 1,4-dioxane was measured to be 22.6% (Fig. S10 and Table S2 in Supporting information).

  The ¹O₂ generation capability of B-2TPA upon 735 nm laser irradiation was investigated using singlet oxygen sensor green (SOSG) as the ¹O₂ trapper. As shown in Fig. 2d, the green fluorescence originated from SOSG in the mixture solution of B-2TPA-SOSG gradually increased with increasing irradiation time. The ¹O₂ QY of B-2TPA was 1.8% using ICG as a reference (Fig. S11 in Supporting information) [39]. Moreover, B-2TPA exhibited excellent photostability. As revealed in Fig. S12 (Supporting information), after irradiation with 735 nm laser with a power of 1.0 W/cm² for 10 min, the absorption spectra of B-2TPA showed almost no change. Moreover, the temperature of B-2TPA solution rose from 25.8 ℃ to 45.7 ℃ (Fig. 2e), and its PCE in dioxane was calculated to be 39.8% ± 0.9% (Fig. S13 in Supporting information).

  B-2TPA was encapsulated in the amphiphilic DSPE-PEG-2000 to prepare water-soluble B-2TPA NPs, and the encapsulation yield of that in NPs is 88.3%. The spherical morphology of B-2TPA NPs was observed by scanning electron microscope (SEM) (Fig. S14a in Supporting information). The size distribution of B-2TPA NPs is ranged from 32 nm to 397 nm (Fig. S14b in Supporting information). The average size of B-2TPA NPs measured by dynamic light scattering (DLS) was ~120 nm, and no obvious changes were observed within one week of observation in aqueous solution, which is indicative of their excellent long-term stability (Fig. S14c in Supporting information). The absorption and fluorescence peaks of B-2TPA NPs red-shifted to 742 and 815 nm, respectively (Fig. 3a). Moreover, the trailing of fluorescence spectrum of B-2TPA NPs (900–1000 nm) caused by TICT effect was also observed. Interestingly, these B-2TPA NPs can still generate ¹O₂ under 735 nm laser irradiation. Compared to B-2TPA in the molecular state, the ¹O₂ QY of B-2TPA NPs was enhanced by 3.7 times, reaching to 6.7% (Fig. 3b and Fig. S15 in Supporting information).

  Figure 3

  

  Figure 3.  (a) Normalized vis-NIR absorption and fluorescence spectra of B-2TPA NPs in aqueous solution. (b) Fluorescence spectra change of SOSG induced by B-2TPA NPs under 735 nm laser irradiation (0.2 W/cm²) for different times. (c) Diagram showing calculated frontier molecular orbital of monomeric and dimeric B-2TPA. ΔE = ELUMO − E_HOMO. (d) Calculated energy levels of the singlet and triplet excited states of monomeric and dimeric B-2TPA. ΔE_ST = E_S − E_T. Irradiation time-dependent temperature variation of B-2TPA NPs aqueous solution at different (e) concentrations and (f) laser powers. (g) Photothermal cycle of B-2TPA NPs and ICG aqueous solutions during laser irradiation. (h) PA intensities and PA images of B-2TPA NPs aqueous solutions at different concentrations.

  DownLoad: Full-Size Img PowerPoint

  To understand the mechanism of aggregation-induced enhancement of ¹O₂ QY, the electronic characteristics and properties in excited state of monomeric and dimeric B-2TPA were investigated by theoretical calculation. The frontier molecular orbitals were acquired via density functional theory (DFT) calculations at B3LYP-D3BJ/6-311G*, and the results were shown in Fig. 3c. As observed, the highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) of monomeric B-2TPA were delocalized in the same molecule, while those of dimeric B-2TPA were delocalized in different molecules. This suggests that both ICT and intermolecular charge transfer exist in dimeric B-2TPA [40]. Moreover, the energy gaps between HOMO and LUMO (ΔE) of monomeric and dimeric B-2TPA were 3.469 eV and 3.256 eV, respectively. The lower ΔE indicates that B-2TPA NPs are more easily excited by laser irradiation [41].

  Furthermore, the lowest excited singlet state (S₁) and excited triplet state (T_n) energy levels of monomeric and dimeric B-2TPA were analyzed via time-dependent density functional theory (TD-DFT) calculations at TD-B3LYP-D3BJ/6-311G*. As shown in Fig. 3d, monomeric B-2TPA only had one ISC channel from S₁ to T₁, while dimeric B-2TPA had two channels (from S₁ to T₁ or T₂). Their smallest energy gap between S₁ and T₁ (ΔE_ST) were 0.937 eV and 0.790 eV, respectively, demonstrating intermolecular aggregation induces lower ΔE_ST, which can effectively improve the ISC efficiency of B-2TPA NPs, enabling its ¹O₂ QY higher than B-2TPA [42].

  The photothermal conversion ability of B-2TPA NPs was also studied. As shown in Figs. 3e and f, the increase in temperature of B-2TPA NPs aqueous solution depended on its concentration and the laser power. The temperature of 8 µmol/L B-2TPA NPs aqueous solution increased from 24.8 ℃ to 55.1 ℃ after laser irradiation (1.0 W/cm²) for 10 min. Next, to evaluate the photostability of B-2TPA NPs, its time-dependent absorption spectra change was recorded under laser irradiation. As shown in Fig. S16 (Supporting information), the NIR absorption peak of B-2TPA NPs only slightly decreased after laser irradiation for 10 min; conversely, ICG was seriously photodegraded after laser irradiation for 3 min, proving that the photostability of B-2TPA NPs is higher than ICG. In addition, after five photothermal conversion cycles, the temperature of B-2TPA NPs aqueous solution reached 63.5 ℃ (Fig. 3g), which is a slight decrease compared to that of ICG and is indicative of the excellent photothermal stability of B-2TPA. Based on its heating and cooling curve, the PCE of B-2TPA NPs was calculated to be 60.1% ± 0.8% (Fig. S17 in Supporting information) [43], which is also higher than that of B-2TPA in dioxane.

  Compared with that of B-2TPA in dioxane, the fluorescence emission of B-2TPA NPs in water was severely quenched (Fig. S18a in Supporting information), indicating that intermolecular aggregation inhibits the radiation decay of B-2TPA NPs, while promoting vibrational relaxation to enhance its photothermal conversion ability. Ma et al. synthesized B3 by introducing two triphenylamine groups into the α position of BODIPY [44]. When water fraction in THF (f_w) increases from 0 to 50%, the fluorescence peak and intensity of B3 gradually redshifted and weakened, which exhibited TICT effect due to free intramolecular motion. Whereas, when f_w continue to rise from 50% to 90%, its fluorescence intensity obviously increases, exhibiting remarkable AIE performance because strong intermolecular aggregation restricted intramolecular motion, inhibiting the formation of TICT state and decreasing the photothermal conversion ability [45]. However, upon the addition of B-2TPA and DSPE-PEG-2000 in water/dioxane mixture solution, as the water content gradually increased, the fluorescence of B-2TPA gradually decreased to quenching (Fig. S18b in Supporting information). This indicates the lipophilic microenvironment formed by alkanes of DSPE-PEG-2000 in NPs protects the stable TICT and effective intramolecular motion of B-2TPA to further enhance the PCE of B-2TPA NPs. Due to its excellent photothermal conversion performance, B-2TPA NPs have high PA imaging capability. As shown in Fig. 3h, the PA intensity of B-2TPA NPs was positively correlated with NPs concentration.

  2,7-Dichlorodihydrofluorescein diacetate (DCFH-DA) was used as an intracellular ¹O₂ fluorescence probe to confirm the ¹O₂ production in 4T1 cells after being treated with different treatments. As shown in Fig. 4a and Fig. S19 (Supporting information), only 4T1 cells incubated with B-2TPA NPs, followed by irradiation with 735 nm laser, showed significant green fluorescence, which indicated that B-2TPA NPs can effectively generate intracellular ¹O₂ under laser irradiation. Calcein AM/propidium iodide (PI) co-staining was employed to visualize the live and death of 4T1 cells. As shown in Fig. 4b and Fig. S20 (Supporting information), after treatment with PBS, PBS + Laser, and B-2TPA NPs, obvious green fluorescence of Calcein AM was observed in 4T1 cells, which suggests that only B-2TPA NPs or laser irradiation did not damage the cells, an indication of the low cytotoxicity of B-2TPA NPs. On the contrary, after incubating with B-2TPA NPs followed by exposure to 735 nm laser, 4T1 cells showed obvious red fluorescence, which confirmed the excellent phototoxicity of B-2TPA NPs. Standard methylthiazolyldiphenyl tetrazolium bromide (MTT) experiment was conducted to further confirm the low cytotoxicity and high phototoxicity of B-2TPA NPs. The results showed that 16 µmol/L B-2TPA NPs + Laser decreased the viability of 4T1 cells to 17% (Fig. 4c), and the corresponding 50% inhibitory concentration (IC₅₀) value was calculated to be 10.4 µmol/L. Flow cytometry results based on annexin V-FITC/7-AAD co-staining further showed that B-2TPA NPs + Laser irradiation increased the early apoptotic and later apoptotic/necrotic rates of 4T1 cells to 5.64% and 25.7%, respectively (Fig. 4d). This indicates that the mechanisms of cell death are early apoptosis and later apoptosis/necrosis.

  Figure 4

  

  Figure 4.  (a) ¹O₂ generation in DCFH-DA incubated 4T1 cells treated with B-2TPA NPs + Laser. (b) Calcein AM/PI co-stained 4T1 cells treated with B-2TPA NPs + Laser. (c) Viability of 4T1 cells incubated with B-2TPA NPs at different concentrations with (B-2TPA NPs + Laser) or without (B-2TPA NPs) laser irradiation. (d) Cell apoptosis observed after B-2TPA NPs + Laser treatment. (e) In vivo PA and (f) NIR fluorescence images of B-2TPA NPs. (g) Infrared thermal images of mice injected with PBS or B-2TPA NPs followed by exposure to laser irradiation for 10 min. (h) Temperature variation of tumor tissues after different treatments. (i) Relative tumor volume and (j) body weight of mice after various treatments for 14 days. (k) H&E-staining and Ki-67 expression of tumor tissues under different treatments. Data represent means ± standard deviation (SD) (n = 5).

  DownLoad: Full-Size Img PowerPoint

  All the animal experiments were approved by the Ethics Committee for Experimental Animal Welfare of Hunan Normal University. B-2TPA NPs were injected into 4T1 tumor-bearing mice through the tail vein, and the in vivo metabolism and distribution were explored. After intravenous injection of B-2TPA NPs, NIR fluorescence images of main organs (heart, liver, spleen, lung, and kidney) and tumor tissues at different times were captured. As shown in Fig. S21 (Supporting information), faint fluorescence was observed in tumor tissue, demonstrating that due to the lack of tumor targeting probe, only a small amount of B-2TPA NPs are accumulated in tumor tissues by enhanced permeability and retention (EPR) effect. Thus, the mice were intratumorally injected with B-2TPA NPs to investigate in vivo optical imaging and anti-tumor ability of B-2TPA NPs. This method can effectively enhance the concentration of B-2TPA NPs in tumor tissue. The strong NIR fluorescence of the liver and spleen proved that B-2TPA NPs accumulated primarily in the liver and spleen, which is attributed to abundant blood perfusion, slow blood flow rate, and active phagocytosis of the reticuloendothelial system [46–48]. Moreover, it gradually increased from 0 to 60 h, indicating that B-2TPA NPs in the body are mainly metabolized by the liver and spleen, and require a longer time to clear from the body (Fig. S22 in Supporting information).

  As shown in Figs. 4e and f, after being intratumorally injected with B-2TPA NPs, the tumor exhibited obvious PA and NIR fluorescence signals, indicating excellent in vivo imaging ability of B-2TPA NPs. Moreover, the monitoring by infrared thermography showed that the temperature of tumors injected with B-2TPA NPs increased from 33.5 ℃ to 61.6 ℃ during the 10 min of illumination, whereas that of tumors injected with PBS only increased from 31.8 ℃ to 38.0 ℃ (Figs. 4g and h). The tumors in the experimental group (B-2TPA NPs + Laser) were completely removed on the fourth day. Conversely, the relative volume of tumor in the control groups (PBS, PBS + Laser, and B-2TPA NPs) has increased by 22-30 times (Fig. 4i). Moreover, the weight of mice gradually increased during monitoring (Fig. 4j), which demonstrates B-2TPA NPs have excellent biocompatibility. The histological analysis of tumor tissues in both the experimental and control groups was conducted by H&E and Ki-67 staining experiments. As shown in Fig. 4k, B-2TPA NPs + laser destroyed the tumor tissues and inhibited their proliferation. In addition, the H&E staining of main organs confirmed that PBS + Laser and B-2TPA NPs groups had no effects on the balance of in vivo physiological activities (Fig. S23 in Supporting information).

  In conclusion, based on aggregation-induced increase in singlet-triplet ISC and TICT effect, we successfully developed phototheranostic agents (B-2TPA NPs) with strong NIR absorption capacity, aggregation-induced enhancement of ¹O₂ QY, and high PCE. B-2PTA was designed and synthesized by introducing N,N-diethyl aniline and triphenylamine into the skeleton of BODIPY, allowing it to exhibit NIR absorption and fluorescence peaks at 729 and 778 nm, respectively. In addition, B-2TPA exhibited a high molar extinction coefficient, excellent photostability, significant TICT effect, and low ¹O₂ QY (1.8%) and PCE (39.8% ± 0.9%). After being wrapped with DSPE-PEG-2000, the obtained B-2TPA NPs showed red-shifted absorption and fluorescence peaks at 751 and 815 nm, respectively. Compared with that of B-2TPA, the ¹O₂ QY of B-2TPA NPs enhanced to 6.7% because the intermolecular aggregation-induced decrease of ΔE_ST improved ISC efficiency. Moreover, obvious fluorescence quenching and stable TICT effect in the aggregation state facilitate vibrational relaxation, increasing the PCE of B-2TPA NPs to 60.1% ± 0.8%. In vitro cell experiments demonstrated that through combined PDT and PTT, B-2TPA NPs efficiently damaged 4T1 cells. Finally, in vivo animal experiments demonstrated that B-2TPA NPs could be utilized in NIR fluorescence and PA imaging-guided synergistic PDT and PTT to eliminate tumor tissues.

  Declaration of competing interest

  The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

  Acknowledgments

  This work was supported by National Key Research and Development Program of China (No. 2022YFA1207600), National Natural Science Foundation of China (Nos. 62175262, 62375289), The Science and Technology Innovation Program of Hunan Province (No. 2022RC1201), The Climb Plan of Hunan Cancer Hospital (No. ZX2021005), The Hunan Provincial Natural Science Foundation of China (No. 2023JJ60464).

  Supplementary materials

  Supplementary material associated with this article can be found, in the online version, at doi:10.1016/j.cclet.2023.109452.

  
  
• 1. [1]

     S. Wang, W. Ren, J.T. Hou, et al., Chem. Soc. Rev. 50 (2021) 8887–8902. doi: 10.1039/D1CS00083G

  2. [2]

     X. Qin, F. Li, Y. Zhang, et al., Anal. Chem. 90 (2018) 9381–9385. doi: 10.1021/acs.analchem.8b01992

  3. [3]

     X. Yue, B. Wang, M. Lan, et al., Chem. Commun. 59 (2023) 4676–4679. doi: 10.1039/D3CC00214D

  4. [4]

     H. Wang, J. Chang, M. Shi, et al., Angew. Chem. Int. Ed. 58 (2019) 1057–1061. doi: 10.1002/anie.201811273

  5. [5]

     K. Yang, F. Long, W. Liu, et al., ACS Appl. Mater. Interfaces 14 (2022) 18043–18052. doi: 10.1021/acsami.1c22444

  6. [6]

     X. Li, F. Fang, B. Sun, et al., Nanoscale Horiz. 6 (2021) 177–185. doi: 10.1039/D0NH00672F

  7. [7]

     Q. Cheng, Z. Li, Y. Xia, et al., NPG Asia Mater. 11 (2019) 63. doi: 10.1038/s41427-019-0164-4

  8. [8]

     L. Zhang, Y. Geng, L. Li, et al., Chem. Sci. 12 (2021) 5918–5925. doi: 10.1039/D1SC00126D

  9. [9]

     A. Kano, Y. Taniwaki, I. Nakamura, et al., J. Control. Release 167 (2013) 315–321. doi: 10.1016/j.jconrel.2013.02.016

  10. [10]

      W. Park, S.J. Park, K. Na, Biomaterials 32 (2011) 8261–8270. doi: 10.1016/j.biomaterials.2011.07.023

  11. [11]

      M. Hou, W. Chen, J. Zhao, et al., Chin. Chem. Lett. 33 (2022) 4101–4106. doi: 10.1016/j.cclet.2022.01.049

  12. [12]

      J. Jin, Y. Zhu, Z. Zhang, et al., Angew. Chem. Int. Ed. 57 (2018) 16354–16358. doi: 10.1002/anie.201808811

  13. [13]

      E.A. Owens, M. Henary, G. El Fakhri, et al., Acc. Chem. Res. 49 (2016) 1731–1740. doi: 10.1021/acs.accounts.6b00239

  14. [14]

      P. Xue, R. Yang, L. Sun, et al., Nano-Micro Lett. 10 (2018) 74. doi: 10.1007/s40820-018-0227-z

  15. [15]

      L. Cheng, W. He, H. Gong, et al., Adv. Funct. Mater. 23 (2013) 5893–5902. doi: 10.1002/adfm.201301045

  16. [16]

      N. Jing, Z. Zhou, W. Xiong, et al., Chin. Chem. Lett. 32 (2021) 3948–3953. doi: 10.1016/j.cclet.2021.06.053

  17. [17]

      Y. Wan, G. Lu, W.C. Wei, et al., ACS Nano 14 (2020) 9917–9928. doi: 10.1021/acsnano.0c02767

  18. [18]

      B. Lu, Y. Huang, Z. Zhang, et al., Mater. Chem. Front. 6 (2022) 2968–2993. doi: 10.1039/D2QM00752E

  19. [19]

      Z. Cheng, T. Zhang, W. Wang, et al., Chin. Chem. Lett. 32 (2021) 1580–1585. doi: 10.1016/j.cclet.2021.02.017

  20. [20]

      R. Jiang, J. Dai, X. Dong, et al., Adv. Mater. 33 (2021) 2101158. doi: 10.1002/adma.202101158

  21. [21]

      J. Zhang, S. Mukamel, J. Jiang, J. Phys. Chem. B 124 (2020) 2238–2244. doi: 10.1021/acs.jpcb.0c00654

  22. [22]

      J. Zhao, K. Yan, G. Xu, et al., Adv. Funct. Mater. 31 (2021) 2008325. doi: 10.1002/adfm.202008325

  23. [23]

      G. Xu, C. Li, C. Chi, et al., Nat. Commun. 13 (2022) 3064. doi: 10.1038/s41467-022-30721-w

  24. [24]

      S. Liu, Y. Li, R.T. Kwok, Chem. Sci. 12 (2021) 3427. doi: 10.1039/D0SC02911D

  25. [25]

      R. Liu, J. Liu, W. Xu, et al., Adv. Mater. 35 (2023) 2303212. doi: 10.1002/adma.202303212

  26. [26]

      D. Xi, M. Xiao, J. Cao, et al., Adv. Mater. 32 (2020) 1907855. doi: 10.1002/adma.201907855

  27. [27]

      Z.R. Grabowski, K. Rotkiewicz, W. Rettig, Chem. Rev. 103 (2003) 3899–4032. doi: 10.1021/cr940745l

  28. [28]

      X. Lv, C. Gao, T. Han, et al., Chem. Commun. 56 (2020) 715–718. doi: 10.1039/C9CC09138F

  29. [29]

      C. Wang, W. Chi, Q. Qiao, et al., Chem. Soc. Rev. 50 (2021) 12656–12678. doi: 10.1039/D1CS00239B

  30. [30]

      S. Liu, X. Zhou, H. Zhang, et al., J. Am. Chem. Soc. 41 (2019) 5359–5368.

  31. [31]

      Z. Zhao, C. Chen, W. Wu, et al., Nat. Commun. 10 (2019) 768. doi: 10.1038/s41467-019-08722-z

  32. [32]

      B. Li, E Pang, S. Zhao, et al., Chem. Biomed. Imaging 1 (2023) 541–549. doi: 10.1021/cbmi.2c00004

  33. [33]

      J. Ma, Z. Liu, Z. Wang, et al., Mater. Chem. Front. 1 (2017) 2547–2553. doi: 10.1039/C7QM00307B

  34. [34]

      Y. Zhang, M. Shi, Z. Yan, et al., Adv. Healthc. Mater. 9 (2020) 2001042. doi: 10.1002/adhm.202001042

  35. [35]

      K. Li, X. Duan, Z. Jiang, et al., Nat. Commun. 12 (2021) 2376. doi: 10.1038/s41467-021-22686-z

  36. [36]

      Z.R. Grabowski, K. Rotkiewicz, A. Siemiarczuk, J. Lumin. 18-19 (1979) 420-424.

  37. [37]

      X.Y. Shen, W. Yuan, Y. Liu, et al., J. Phys. Chem. C 116 (2012) 10541–10547. doi: 10.1021/jp303100a

  38. [38]

      J. Cao, Q. Liu, S. Bai, et al., ACS Appl. Mater. Interfaces 11 (2019) 29814–29820. doi: 10.1021/acsami.9b07677

  39. [39]

      D. Wu, H.C. Daly, E. Conroy, et al., Eur. J. Med. Chem. 161 (2019) 343–353. doi: 10.1016/j.ejmech.2018.10.046

  40. [40]

      W. Hu, X. Miao, H. Tao, et al., ACS Nano 13 (2019) 12006–12014. doi: 10.1021/acsnano.9b06208

  41. [41]

      X. Li, L. Liu, S. Li, et al., ACS Nano 13 (2019) 12901–12911. doi: 10.1021/acsnano.9b05383

  42. [42]

      H. Ma, Y. Lu, Z. Huang, et al., J. Am. Chem. Soc. 144 (2022) 3477–3486. doi: 10.1021/jacs.1c11886

  43. [43]

      X. Xing, K. Yang, B. Li, et al., J. Phys. Chem. Lett. 13 (2022) 7939–7946. doi: 10.1021/acs.jpclett.2c02122

  44. [44]

      H. Gao, Y. Gao, C. Wang, et al., ACS Appl. Mater. Interfaces 10 (2018) 14956–14965. doi: 10.1021/acsami.7b13444

  45. [45]

      S. Liu, C. Chen, Y. Li, et al., Adv. Funct. Mater. 30 (2020) 1908125. doi: 10.1002/adfm.201908125

  46. [46]

      W. Zhang, B. Hao, C. Gao, et al., Eur. J. Pharm. Biopharm. 179 (2020) 206–220.

  47. [47]

      K.M. Tsoi, S. MacParland, X. Ma, et al., Nat. Mater. 15 (2016) 1212–1221. doi: 10.1038/nmat4718

  48. [48]

      Z. Li, Y. Zhu, H. Zeng, et al., Nat. Commun. 14 (2023) 14337.

• 

  Figure 1  (a) Preparation of B-2TPA NPs, and molecular structure and main optical properties of B-2TPA and B-2TPA NPs. (b) Application of B-2TPA NPs in NIR fluorescence and PA imaging-guided synergistic PDT and PTT of tumor.

  下载: 全尺寸图片 幻灯片
  

  Figure 2  (a) Optimized ground state (S₀) geometries of B-2TPA. (b) Normalized visible (vis)-NIR absorption and fluorescence spectra of B-2TPA in 1,4-dioxane. (c) Line-fitting of a plot between Stokes shift and Δf. (d) Irradiation time-dependent fluorescence change of SOSG in B-2TPA solution. (e) Irradiation time-dependent temperature variation of B-2TPA solution.

  下载: 全尺寸图片 幻灯片
  

  Figure 3  (a) Normalized vis-NIR absorption and fluorescence spectra of B-2TPA NPs in aqueous solution. (b) Fluorescence spectra change of SOSG induced by B-2TPA NPs under 735 nm laser irradiation (0.2 W/cm²) for different times. (c) Diagram showing calculated frontier molecular orbital of monomeric and dimeric B-2TPA. ΔE = ELUMO − E_HOMO. (d) Calculated energy levels of the singlet and triplet excited states of monomeric and dimeric B-2TPA. ΔE_ST = E_S − E_T. Irradiation time-dependent temperature variation of B-2TPA NPs aqueous solution at different (e) concentrations and (f) laser powers. (g) Photothermal cycle of B-2TPA NPs and ICG aqueous solutions during laser irradiation. (h) PA intensities and PA images of B-2TPA NPs aqueous solutions at different concentrations.

  下载: 全尺寸图片 幻灯片
  

  Figure 4  (a) ¹O₂ generation in DCFH-DA incubated 4T1 cells treated with B-2TPA NPs + Laser. (b) Calcein AM/PI co-stained 4T1 cells treated with B-2TPA NPs + Laser. (c) Viability of 4T1 cells incubated with B-2TPA NPs at different concentrations with (B-2TPA NPs + Laser) or without (B-2TPA NPs) laser irradiation. (d) Cell apoptosis observed after B-2TPA NPs + Laser treatment. (e) In vivo PA and (f) NIR fluorescence images of B-2TPA NPs. (g) Infrared thermal images of mice injected with PBS or B-2TPA NPs followed by exposure to laser irradiation for 10 min. (h) Temperature variation of tumor tissues after different treatments. (i) Relative tumor volume and (j) body weight of mice after various treatments for 14 days. (k) H&E-staining and Ki-67 expression of tumor tissues under different treatments. Data represent means ± standard deviation (SD) (n = 5).

  下载: 全尺寸图片 幻灯片
•