Dual-receptor NIR-II ionic fluorescent probe as well as preparation method and application thereof

Abstract

The invention relates to the technical field of fluorescent probes, in particular to a double-receptor NIR-II ionic fluorescent probe as well as a preparation method and application of the double-receptor NIR-II ionic fluorescent probe. According to the present invention, through the double-receptor engineering strategy, the benzothiadiazole receptor unit is ingeniously introduced between the 2-(N, N-diphenylamino) thiophene (DT) unit and the benzo [c, d] indole onium salt (BIN) unit, and the double-receptor NIR-II ionic fluorescent probe DT-BT-BIN is designed and synthesized; the fluorescent probe disclosed by the invention not only has the advantages of aggregation-induced NIR-II fluorescence emission, efficient photodynamic and photothermal activity, excellent light stability, good biocompatibility and the like; and the nano probe DT-BT-BIN NPs formed by self-assembly can be rapidly endocytosed by in-vitro cancer cells, and cell apoptosis is induced by destroying mitochondrial functions under the irradiation of 808nm laser, so that the cancer cells are effectively killed.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of fluorescent probe, in particular to a dual-receptor NIR-II ionic fluorescent probe and its preparation method and application, more particularly to a near-infrared AIE fluorescent probe for destroying mitochondria and having phototherapy activity and its preparation method and application. BACKGROUND

[0002] Cancer has become the second leading cause of death threatening public health, and it is urgent to develop precise and efficient cancer diagnosis and treatment strategies. Light-mediated diagnosis and treatment technology has been widely used in preclinical and clinical applications due to its high spatiotemporal controllability, non-invasiveness, real-time imaging, in-situ treatment, and low toxicity to normal tissues. Notably, ionic probes have attracted much attention in clinical practice, such as indocyanine green (ICG) and methylene blue approved by the US Food and Drug Administration (FDA). Traditional ionic probes have some shortcomings, such as short excitation and emission wavelengths, poor photostability, aggregation-induced fluorescence self-quenching, and reduced aggregation phototherapy (photodynamic and photothermal therapy) activity, making it difficult to meet the complex clinical diagnosis and effective phototherapy needs. Fortunately, ionic aggregation-induced emission (AIE) probes have been repeatedly proven to effectively overcome fluorescence self-quenching and enhance the phototherapy activity in the aggregated state. In addition, near-infrared region II (NIR-II, 1000−1700nm) probes have deep tissue penetration depth, low tissue scattering and absorption rate, and high signal-to-noise ratio, and have attracted much attention in disease diagnosis and treatment. Therefore, it is very promising to develop NIR-II ionic AIE probes for real-time imaging to guide the effective elimination of cancer.

[0003] Quinolinium, pyridinium and benzothiazolium salts are very suitable for constructing NIR-II fluorescent probes with donor-π-acceptor structure due to their strong electron-withdrawing ability. Electron donor groups with twisted conformation (such as tetraphenylethyl and triphenylamine groups) have been widely used to develop NIR-II ionic AIE probes. So far, a variety of NIR-II ionic AIE probes have been developed, such as ionic AIE photosensitizers based on S-π interactions, ionic AIE probes based on quinoline or benzothiazolium salt groups for organelle-targeted phototherapy, ionic AIE probes based on donor engineering strategies, which can induce apoptosis and pyroptosis through photodynamic effects, and systems based on coordinated donor and acceptor engineering strategies. All these methods have been successfully applied to effective tumor phototherapy guided by NIR-II fluorescence imaging. However, most of these ionic probes are only excited by visible light (400−700nm) with small penetration depth. In addition, their emission peaks are mainly located in the near-infrared region I (NIR-I, 700−900nm), with only one tail extending to the NIR-II region, which significantly limits the clinical application conversion. SUMMARY

[0004] In order to overcome the shortcomings and deficiencies existing in the prior art, the purpose of the present application is to provide a dual-receptor NIR-II ionic fluorescent probe and a preparation method and application thereof.

[0005] The purpose of the present application is achieved by the following technical scheme: a dual-receptor NIR-II ionic fluorescent probe, the fluorescent probe has a structure shown in general formula (I): ; Among them, R 1 is any one of hydrogen, halogen, alkyl, alkoxy, perfluoroalkyl, aryloxy, alkenyl, alkynyl, cycloalkyl, alkylthio, amido, amino, monoalkylamino, dialkylamino, diarylamino, triarylethylene, carboxamide, hydroxyl, thiol, aryl or heteroaryl; P is any one of furanyl, thienyl, selenophenyl, epoxy furanyl, epoxy thienyl, epoxy selenophenyl, isobenzofuranyl, benzo[C]thiophenyl, benzo[C]selenophenyl, aryl or heteroaryl; Q is any one of furanyl, thienyl, selenophenyl, epoxy furanyl, epoxy thienyl, epoxy selenophenyl, isobenzofuranyl, benzo[C]thiophenyl, benzo[C]selenophenyl, benzo oxadiazolyl, benzo thiazolyl, benzo selenadiazolyl, aryl or heteroaryl; A is any one of pyridinium salt group, quinolinium salt group, 2,3,3-trimethyl-3H-indolium salt group, 1,1,2-trimethyl-1H-benzo[e]indolium salt group, 2,3,3-trimethyl-3H-benzo[g]indolium salt group or benzo[c,d]indolium salt group.

[0006] The inventors developed a series of ionic AIE probes (DT-In, DT-BIn and DT-BT-BIn) through receptor engineering strategy. By ingeniously introducing a benzothiazole acceptor unit between the donor and the acceptor, a dual-receptor NIR-II ionic probe was designed and synthesized, which showed near-infrared absorption and NIR-II emission, excellent AIE characteristics, high I / II type ROS generation capacity and photo-thermal conversion efficiency. In addition, through the self-assembly of amphiphilic polymer mPEG-PDLLA and DT-BT-BIn, a NIR-II AIE nanoprobe DT-BT-BIn NPs was successfully prepared. The prepared nanoprobe can effectively accumulate in the tumor site, significantly destroy the mitochondrial function under laser irradiation, and then induce cancer cell death and eliminate tumors through the apoptosis mechanism.

[0007] Preferably, the R 1 is hydrogen; the P is thienyl; the Q is benzothiazole group, and the fluorescent probe has a structure shown in general formula (II): ; Wherein, A is any one of pyridinium salt, quinolineium salt, 2,3,3-trimethyl-3H-indoleium salt, 1,1,2-trimethyl-1H-benzo[e]indoleium salt, 2,3,3-trimethyl-3H-benzo[g]indoleium salt, or benzo[c,d]indoleium salt. Their structural formulas are as follows:

[0008] Among them, R 2 It is any one of a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, a sulfonate group attached to a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, a carboxylate group attached to a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, an aryl group, or a heteroaryl group.

[0009] More preferably, A is a benzo[c,d]indolium salt, with the following structural formula: .

[0010] Most preferably, the R 2 The fluorescent probe is methyl and has the structure shown in general formula (Ⅳ): .

[0011] Preferably, the fluorescent probe is self-assembled from methoxy polyethylene glycol-polylactic acid copolymer (mPEG-PDLLA) and a compound represented by general formula (IV) (DT-BT-BIn). The self-assembled NIR-II AIE nanoprobe DT-BT-BInNPs can be rapidly endocytosed by cancer cells in vitro, and through phototherapy activity generated by laser irradiation, it significantly disrupts mitochondrial function and effectively kills cancer cells through apoptosis.

[0012] Preferably, the fluorescent probe is a dual-receptor NIR-II ion probe, which is also a molecular probe with tumor phototherapy activity. Specifically, the dual-receptor NIR-II ion probe refers to an AIE probe that generates NIR-II fluorescence signal under 808nm excitation, locates tumor lesions, and disrupts normal mitochondrial function using highly efficient phototherapy activity. More specifically, the highly efficient phototherapy-active AIE probe that disrupts normal mitochondrial function refers to an AIE probe that generates type I / II ROS and high heat under 808nm light irradiation to achieve a tumor therapeutic effect.

[0013] Another objective of the present invention is achieved by the following technical solution: a method for preparing a dual-receptor NIR-II ionic fluorescent probe, wherein the compound represented by general formula (II) is reacted with 1-ethyl-2-methylbenzo[c,d]indole-1-iodide in an organic solvent in an anhydrous and oxygen-free environment to obtain the fluorescent probe represented by general formula (III).

[0014] The molar ratio of the compound represented by general formula (II) to 1-ethyl-2-methylbenzo[c,d]indole-1-iodide ammonium is 1:1 to 3:1; the organic solvent is at least one of acetonitrile, N,N-dimethylformamide, tetrahydrofuran, methanol, ethanol or dimethyl sulfoxide; the reaction temperature is 60 to 85°C, and the reaction time is 5 to 90 minutes.

[0015] Another objective of this invention is achieved through the following technical solution: the application of a dual-receptor NIR-II ionic fluorescent probe in the preparation of antitumor drugs and / or cellular fluorescence imaging. The NIR-II ionic AIE probe DT-BT-BIn was successfully synthesized using a dual-receptor engineering strategy, exhibiting advantages such as aggregation-induced NIR-II fluorescence emission, excellent photodynamic and photothermal capabilities, high photostability, and superior biocompatibility. Furthermore, the NIR-II AIE nanoprobe DT-BT-BIn NPs, prepared by self-assembly of the amphiphilic polymer mPEG-PDLLA and DT-BT-BIn, can be rapidly endocytosed by cancer cells in vitro. Through phototherapy activity generated by laser irradiation, it significantly disrupts mitochondrial function and effectively kills cancer cells via apoptosis. In addition, DT-BT-BIn NPs can be used to locate tumor sites through NIR-II fluorescence imaging, enabling selective irradiation of tumors with high spatiotemporal controllability. Simultaneously, the phototherapy effect can be monitored in situ in real time through photothermal imaging, ultimately achieving effective in vivo tumor ablation.

[0016] Preferably, the tumor includes tumors formed from cervical cancer cells, breast cancer cells, ovarian cancer cells, lung cancer cells, or skin cancer cells; the drug is a photodynamic and photothermal therapy drug. Specifically, photodynamic and photothermal therapy refers to the selective irradiation of superficial tumors using near-infrared light with high spatiotemporal controllability or the irradiation of deep tumor tissues via optical fiber guidance.

[0017] The beneficial effects of the present invention are as follows: 1. The dual-receptor NIR-II ionic AIE probe of the present invention is based on the dual-receptor engineering strategy. By cleverly introducing a benzothiadiazole receptor unit between strong donor and receptor units, the dual-receptor NIR-II ionic AIE probe DT-BT-BIn was successfully designed and synthesized.

[0018] 2. The dual-receptor NIR-II ionic AIE probe of this invention can be rapidly endocytosed by cancer cells, significantly disrupting mitochondrial function through phototherapy and effectively killing cancer cells via apoptosis. This achieves effective killing of tumor cells both in vivo and in vitro, which is of great significance for tumor phototherapy.

[0019] 3. The mitochondrial-damaging dual-receptor NIR-II ionic AIE probe of the present invention is a tumor therapeutic drug for photodynamic and photothermal therapy. While killing tumor cells, it also has good biocompatibility. Experiments have shown that it has low toxicity to normal cells and will not cause adverse reactions such as hemolysis and inflammation in organisms. Attached Figure Description

[0020] Figure 1 The images show the 1H and 1C NMR spectra of compound DT-BT-BIn in deuterated dimethyl sulfoxide.

[0021] Figure 2 This is the high-resolution mass spectrum of the compound DT-BT-BIn.

[0022] Figure 3 The images show the 1H and 1C NMR spectra of compound DT-In in deuterated dimethyl sulfoxide.

[0023] Figure 4 This is the high-resolution mass spectrum of the compound DT-In.

[0024] Figure 5 Photophysical properties of DT-BT-BIn; (A) UV-Vis absorption spectra of DT-In, DT-BIn and DT-BT-BIn in DMSO solution and (B) normalized fluorescence spectra; (C) Relative maximum fluorescence intensity (I / I0) of DT-In, DT-BIn and DT-BT-BIn in DMSO / toluene mixtures with different toluene contents; (D) Generation of reactive oxygen species in DT-BT-BIn; (E) Generation of superoxide anions; (F) Generation of hydroxyl radicals; (G) Generation of singlet oxygen; Temperature variation curves of DT-BT-BIn at different concentrations (H) and different powers (I).

[0025] Figure 6 The photophysical properties of DT-BT-BIn NPs are as follows: (A) UV-Vis absorption and fluorescence spectra of DT-BT-BIn in aqueous solution; (B) histogram and SEM image of particle size distribution of DT-BT-BIn NPs; (C) generation of reactive oxygen species in DT-BT-BIn NPs; (D) photophysical properties of DT-BT-BIn NPs at different concentrations under an 808 nm laser (power 0.8 W / cm²). 2(E) Temperature change curve under irradiation; (F) Thermal imaging at different powers; (G) Photothermal cycle diagram; (H) Photothermal performance of DT-BT-BIn NPs after cooling to room temperature by linear analysis; (I) Photothermal stability of DT-BT-BIn NPs and ICG.

[0026] Figure 7 Evaluation of the phototherapy and antitumor effects of the fluorescent probe DT-BT-BIn NPs; (A) Cell accumulation of FITC-labeled DT-BT-BIn NPs; (B) Cell viability of breast cancer cells (MDA-MB-231) and (C) normal breast cells (MCF-10A) after treatment with different concentrations of probe; (D) Live and dead staining of cells in different treatment groups.

[0027] Figure 8 The mechanism by which the fluorescent probe DT-BT-BIn NPs induces cancer cell death under laser irradiation; (A) Intracellular reactive oxygen species generation in different treatment groups (A) fluorescence imaging and (B) flow cytometry analysis; Intracellular mitochondrial membrane potential in different treatment groups (C) flow cytometry analysis and (D) fluorescence imaging; (E) CLSM image of DT-BT-BIn NPs and MitoTracker co-treatment; (F) Flow cytometry analysis of apoptosis in different treatment groups.

[0028] Figure 9 For in vivo imaging and evaluation of antitumor activity of fluorescent probe DT-BT-BIn NPs; (A) In vivo NIR-II imaging of tumor-bearing mice; (B) Tumor temperature change of tumor-bearing mice under light irradiation over time; (C) Tumor volume change of nude mice over time; (D) Tumor photographs of mice in each group after 21 days of treatment; (E) Hematoxylin-eosin staining, TUNEL plot and Ki67 immunohistochemical staining of tumor sections.

[0029] Figure 10 The images show hematoxylin-eosin staining of major organs in different treatment groups after 14 days of treatment.

[0030] Figure 11 This is a mechanistic diagram of the dual-receptor NIR-II ionotype AIE probe in tumor and imaging applications. Detailed Implementation

[0031] To facilitate understanding by those skilled in the art, the following description is provided in conjunction with embodiments and appendices. Figures 1-11 The present invention will be further described below, and the content mentioned in the embodiments is not intended to limit the present invention.

[0032] Compound I-1 (84 mg, 0.3 mmol) and compound II-1 (64.6 mg, 0.2 mmol) were added to 10 mL of anhydrous ethanol and refluxed (80 °C) under nitrogen protection, with stirring for 1.5 h. After cooling to room temperature, the reaction mixture was evaporated under reduced pressure. Finally, the residue was further purified by silica gel column chromatography (DCM:MeOH = 15:1) to obtain 69 mg of compound III-1 (DT-BT-BIn) (yield 59%). The 1H and 1C NMR spectra of compound III-1 in deuterated dimethyl sulfoxide are shown below. Figure 1 As shown. 1 HnmR (500 MHz, Chloroform- d ) δ 9.58 (s, 1H), 9.18 – 9.03 (m, 2H), 8.64 (d, J = 14.8 Hz, 1H), 8.09 (dd, J = 21.7, 5.7 Hz, 3H), 7.94 (s,1H), 7.76 (d, J = 7.9 Hz, 1H), 7.64 (d, J = 10.4 Hz, 2H), 7.40 (t, J = 7.8Hz, 4H), 7.31 (d, J = 7.5 Hz, 4H), 7.24 (d, J = 7.4 Hz, 2H), 6.56 (d, J = 4.3Hz, 1H), 4.89 (s, 2H), 1.41 – 1.17 (m, 3H). 13 CnmR (126 MHz, Chloroform- d The high-resolution mass spectra of compound III-1 are as follows: δ 160.68, 160.54, 151.91, 149.40, 146.48, 135.84, 135.11, 133.06, 132.63, 131.63, 129.83, 129.53, 128.96, 128.56, 128.00, 125.70, 125.02, 124.24, 123.69, 122.92, 116.99, 116.26, 115.32, 42.37, 15.52. Figure 2 As shown. m / z [M] + calc,d for C 37 H 27N4S2: 591.1672, found: 591.1676.

[0033] The reaction formula is as follows: .

[0034] Compared with compound III-1 (DT-BT-BIn), the proton and carbon NMR spectra of compound DT-In are as follows: Figure 3 As shown, the high-resolution mass spectrum is as follows: Figure 4 As shown.

[0035] like Figure 5 As shown, the DT-BT-BIn synthesized in Example 1 was subjected to UV-Vis absorption spectroscopy, fluorescence spectroscopy, fluorescence intensity changes of DT-BT-BIn in different proportions of undesirable solvents, and photodynamic and photothermal activities of DT-BT-BIn. Figure 5 A shows the UV-Vis absorption spectra of the molecular probes DT-In, DT-BIn, and DT-BT-Bn. The maximum absorption values ​​for DT-In, DT-BIn, and DT-BT-Bn were observed at 574 nm, 695 nm, and 785 nm, respectively, while their maximum emission values ​​in DMSO solution were located at 658 nm, 787 nm, and 1035 nm (see [reference needed]). Figure 5 B). Subsequently, the fluorescence properties of these three molecules were further investigated. Increasing the fraction of the poor solvent (toluene) from 0 to 90% resulted in increases in fluorescence intensity of 1.28-fold, 1.67-fold, and 3.64-fold, respectively. Figure 5 C), exhibiting superior AIE properties. Subsequently, the phototherapy activity of DT-BT-BIn under 808nm laser irradiation was investigated. DCFH was used as an indicator to detect the generation of reactive oxygen species (ROS) by DT-BT-BIn. Figure 5 As shown in Figure D, after irradiation with an 808 nm laser for 75 seconds, the fluorescence intensity of the DT-BT-BIn+DCFH group increased by 1025.3 times, indicating that DT-BT-BIn possesses excellent photodynamic activity. Meanwhile, Figure 5 E-5G confirmed that DT-BT-BIn can effectively generate type I ROS (O2). •− and •OH) and type II ROS ( 1 The presence of O2 indicates that DT-BT-BIn possesses excellent type I and type II photodynamic activity. Furthermore, the photothermal activity of DT-BT-BIn was evaluated. Figure 5As shown in H-5I, the temperature increase of DT-BT-BIn is correlated with both laser power and its concentration. After irradiation with an 808 nm laser for 300 s, the temperature of DT-BT-BIn (200 μM) in DMSO solution rose to 78.3 °C. This indicates that DT-BT-BIn possesses excellent photothermal activity.

[0036] The DT-BT-BIn synthesized in Example 1 was used to prepare DT-BT-BIn NPs by nanoprecipitation. Figure 6 A shows the UV-Vis absorption and fluorescence spectra of DT-BT-BIn NPs. In aqueous solution, DT-BT-BIn NPs exhibit an absorbance peak at 734 nm and an emission peak at 1174 nm. To detect the formation of DT-BT-BIn NPs, dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used for evaluation. Figure 6 As shown in Figure B, the DT-BT-BIn NPs have a particle size of approximately 139 nm and exhibit a uniform spherical shape. Subsequently, DCFH was used as an indicator to characterize and evaluate the generation of reactive oxygen species (ROS) in DT-BT-BIn NPs under laser irradiation. Figure 6 As shown in Figure C, after irradiation with an 808 nm laser for 80 s, the fluorescence intensity of the DT-BT-BIn NPs+DCFH group significantly increased, indicating that DT-BT-BIn NPs possess excellent photodynamic activity. Furthermore, the photothermal activity of DT-BT-BIn NPs was evaluated by monitoring temperature changes under laser irradiation. Figure 6 As shown in D-6E, the temperature increase of DT-BT-BIn NPs also exhibits both laser power dependence and concentration dependence, and the thermal imaging images also show the same results (see D-6E). Figure 6 F). For example Figure 6 As shown in G-6H, the photothermal conversion efficiency of DT-BT-BIn NPs reaches as high as 55.71%. Meanwhile, compared with ICG, DT-BT-BIn NPs exhibit superior photothermal stability, with negligible temperature decay after ten heating-cooling cycles (see G-6H). Figure 6 I).

[0037] In Example 1, the in vitro uptake efficiency of the synthesized DT-BT-BIn was studied using FITC labeling to evaluate its endocytic behavior. Figure 7 As shown in Figure A, the fluorescence signal gradually increased as the incubation time was extended from 0 hours to 8 hours; subsequently, the phototherapy effect of DT-BT-BInNPs under 808nm laser irradiation was evaluated. Figure 7As shown in Figure B, the cell viability of MDA-MB-231 cells treated with DT-BT-BIn NPs (10 μM) was as low as 10.3% under laser irradiation, while the cell viability was significantly higher (96.4%) under dark conditions, indicating its significant phototherapy activity against cancer cells in vitro. Interestingly, even at high concentrations (100 μM), DT-BT-BIn NPs showed low cytotoxicity against normal MCF-10A cells (see Figure B). Figure 7 C), indicating its excellent biocompatibility. Next, the in vitro anticancer efficacy of DT-BT-BIn NPs was evaluated by co-staining with calcein AM (green, live cells) and propiconazole iodide (red, dead cells). For example... Figure 7 As shown in Figure D, after incubation with DT-BT-BIn NPs, intracellular red fluorescence specifically appeared at the near-infrared laser irradiation site, while other treatment groups were unaffected.

[0038] The intracellular ROS generation capacity was assessed using DCFH-DA as an indicator. Figure 8 As shown in Figure A, DT-BT-BIn NPs exhibited a strong green fluorescence signal in MDA-MB-231 cells under laser irradiation, and flow cytometry analysis also quantitatively confirmed the effectively increased ROS production (see Figure A). Figure 8 B). Due to excessive ROS production, mitochondrial membrane potential decreased significantly, thereby inducing mitochondrial damage and apoptosis. Subsequently, the changes in mitochondrial membrane potential of DT-BT-BIn NPs under different treatment groups were assessed using the JC-1 probe. Flow cytometry analysis showed (see...) Figure 8 C), the content of monomeric JC-1 in DT-BT-BIn NPs was 52.0% after 12 hours of cultivation under 808 nm laser irradiation, and significantly increased to 98.0% after 24 hours of treatment; this was also verified by changes in JC-1 red-green fluorescence imaging (see C). Figure 8 D). Furthermore, mitochondria were labeled Mito Tracker Green, appearing as filamentous green structures in healthy MDA-MB-231 cells. Combined treatment (DT-BT-BIn NPs + laser) induced mitochondrial fragmentation, while no significant changes were observed in MDA-MB-231 cells treated with laser or DT-BT-BIn NPs alone (see [link to treatment]). Figure 8 E). These results indicate that synergistic treatment with DT-BT-BIn NPs and near-infrared irradiation triggers mitochondrial damage, a key initiation event for apoptosis. Subsequently, apoptosis was quantified using Annexin V FITC / PI co-staining assays. Figure 8As shown in Figure F, the apoptosis rate of MDA-MB-231 cells treated with DT-BT-BIn NPs + laser increased to 99.48%, while the apoptosis rates of the laser group and the DT-BT-BIn NPs group were 2.30% and 12.92%, respectively. This indicates that cancer cells have excellent phototherapy activity.

[0039] To verify the in vivo real-time tumor localization ability and anti-tumor efficacy of DT-BT-BIn NPs combined with 808nm laser irradiation, we established an MDA-MB-231 tumor nude mouse model. Figure 9 As shown in Figure A, DT-BT-BIn NPs primarily accumulated in the liver shortly after administration, with tumor-targeted accumulation becoming clearly visible 3 hours post-injection. Maximum fluorescence intensity was reached in the tumor region between 24 and 48 hours. Although the fluorescence signal at the tumor site decreased after 54 hours, significant nanoparticle retention was still observed at the tumor site. These results indicate that DT-BT-BIn NPs can achieve sustained tumor accumulation and allow for gradual metabolic elimination via the liver. Notably, under 808 nm laser irradiation for 10 min, the tumor temperature in mice treated with DT-BT-BIn NPs exceeded 55°C (see Figure A). Figure 9 (B) This high-thermal effect can effectively induce irreversible thermal ablation of tumor tissue. Subsequently, we systematically studied the in vivo antitumor effects of DT-BT-BIn NPs. Figure 9 As shown in Figure C, compared with other groups, the synergistic phototherapy effect of DT-BT-BInNPs under near-infrared light irradiation effectively eliminated tumors in nude mice. In contrast, the tumor volume in all control groups gradually increased over time. Furthermore, tumor tissues from different treatment groups were dissected 21 days after treatment. Photographs of the ex vivo tumor tissues, hematoxylin-eosin staining, TUNEL staining, and Ki67 immunohistochemical staining further validated the significant phototherapy effect of DT-BT-BIn NPs on tumors under laser irradiation. Figure 9 These results collectively demonstrate that DT-BT-BIn NPs possess precise and effective in vivo tumor elimination capabilities under NIR-II fluorescence / photothermal imaging guidance.

[0040] Biosafety verification of DT-BT-BIn NPs. For example... Figure 10 As shown in the hematoxylin-eosin staining image of the fluorescent probe DT-BT-BIn NPs, no obvious inflammation or damage was observed. This confirms that the fluorescent probe DT-BT-BIn NPs possesses excellent biocompatibility.

[0041] like Figure 11The diagram illustrates the mechanism of dual-receptor NIR-II ion-type AIE probes in tumor and imaging applications. These probes can induce aggregation-induced NIR-II emission, leading to the generation of type I and type II reactive oxygen species and improving photothermal conversion efficiency.

[0042] The above embodiments are preferred implementations of the present invention. In addition, the present invention can be implemented in other ways. Any obvious substitutions without departing from the concept of the present invention are within the protection scope of the present invention.

Claims

1. A dual receptor NIR-II ionic fluorescent probe, characterized in that: The fluorescent probe has a structure shown in general formula (I): ； wherein R is any one of hydrogen, halogen, alkyl, alkoxy, perfluoroalkyl, aryloxy, alkenyl, alkynyl, cycloalkyl, alkylthio, amido, amino, monoalkylamino, dialkylamino, diarylamino, triarylethylene, carboxamide, hydroxyl, thiol, aryl, or heteroaryl; 1 wherein R is any one of hydrogen, halogen, alkyl, alkoxy, perfluoroalkyl, aryloxy, alkenyl, alkynyl, cycloalkyl, alkylthio, amido, amino, monoalkylamino, dialkylamino, diarylamino, triarylethylene, carboxamide, hydroxyl, thiol, aryl, or heteroaryl; P is any one of nothing, furan group, thiophene group, selenophene group, epoxy furan group, epoxy thiophene group, epoxy selenophene group, isobenzofuran group, benzo[C]thiophene group, benzo[C]selenophene group, aryl group or heteroaryl group; Q is any one of nothing, furan group, thiophene group, selenophene group, epoxy furan group, epoxy thiophene group, epoxy selenophene group, isobenzofuran group, benzo[C]thiophene group, benzo[C]selenophene group, benzo oxadiazole group, benzo thiadiazole group, benzo selenadiazole group, aryl group or heteroaryl group; A is any one of pyridinium group, quinolinium group, 2,3,3-trimethyl-3H-indolium group, 1,1,2-trimethyl-1H-benzo[e]indolium group, 2,3,3-trimethyl-3H-benzo[g]indolium group or benzo[c,d]indolium group. 2.The dual receptor NIR-II ionic fluorescent probe according to claim 1, characterized in that: The R 1 The P group is hydrogen; the P group is thiophene; the Q group is benzothiadiazole; and the fluorescent probe has the structure shown in general formula (II): ； A is any one of 2,3,3-trimethyl-3H-indolium group, 1,1,2-trimethyl-1H-benzo[e]indolium group, 2,3,3-trimethyl-3H-benzo[g]indolium group or benzo[c,d]indolium group.

3. The dual-receptor NIR-II ionotropic fluorescent probe according to claim 2, characterized in that: The A is benzo[c,d]indolium group, and the fluorescent probe has a structure shown in general formula (III): ； wherein R 2 is any one of a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, an alcohol group linked to a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, a carboxylate group linked to a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, an ester group linked to a saturated or partially saturated alkyl group containing 1 to 16 carbon atoms, an aryl group, or a heteroaryl group.

4. The dual-receptor NIR-II ionotropic fluorescent probe according to claim 3, characterized in that: The R 2 is methyl, the fluorescent probe has the structure shown in general formula (IV): 。 5.The dual receptor NIR-II ionic fluorescent probe according to claim 4, characterized in that: The fluorescent probe is self-assembled by methoxy polyethylene glycol-polylactic acid copolymer and a compound shown in general formula (IV).

6. The dual-receptor NIR-II ionotropic fluorescent probe according to claim 1, characterized in that: The fluorescent probe is a near-infrared two-zone aggregation-induced emission probe for destroying mitochondria, and is also a molecular probe for synergistically killing tumor cells by photodynamic and photothermal therapy.

7. The method for preparing a dual-receptor NIR-II ionic fluorescent probe as described in claim 3, characterized in that: The fluorescent probe shown in general formula (III) is prepared by reacting a compound shown in general formula (II) with 1-ethyl-2-methylbenzo[c,d]indolium-1-iodide in an organic solvent under anhydrous and anaerobic conditions.

8. The method according to claim 7, wherein the method is characterized by: The molar ratio of the compound shown in general formula (II) to 1-ethyl-2-methylbenzo[c,d]indolium-1-iodide is 1:1-3:1; the organic solvent is at least one of acetonitrile, N,N-dimethylformamide, tetrahydrofuran, methanol, ethanol or dimethyl sulfoxide; the reaction temperature is 60-85°C, and the reaction time is 5-90 minutes.

9. Use of the dual-receptor NIR-II ionic fluorescent probe according to any one of claims 1-6 in the preparation of an anti-tumor drug or in vivo tumor fluorescence imaging.

10. Use according to claim 9, characterized in that: The tumor includes a tumor formed by cervical cancer cells, breast cancer cells, ovarian cancer cells, lung cancer cells or skin cancer cells; and the drug is a drug for photodynamic and photothermal therapy.

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