Fluorescent diagnosis and treatment agent compound with near-infrared II region fluorescence imaging and chemical kinetics treatment performance as well as nanoparticles and preparation method of fluorescent diagnosis and treatment agent compound

Abstract

The invention relates to a fluorescent diagnosis and treatment agent compound with near-infrared II region fluorescence imaging and chemical kinetics treatment properties, and nanoparticles and a preparation method thereof. The compound is obtained by chelating organic fluorescent micromolecules with copper ions. The compound is prepared into nanoparticles under the action of a surfactant. The organic fluorescent micromolecule has a triphenylamine substituted benzothiadiazole structure, and catechol groups are introduced to two ends of the molecule, so that the organic fluorescent micromolecule has NIR-II fluorescence imaging and metal ion chelating properties. Due to intramolecular charge transfer between the copper ions and the organic fluorescent small molecules, fluorescence quenching of the prepared compound and the nanoparticles thereof is realized; under a weak acidic condition, the compound can be subjected to chemical kinetics reaction with glutathione to generate reactive oxygen species and o-benzoquinone products. Under the irradiation of 660 nm or 830 nm laser, the compound shows excellent NIR-II fluorescence characteristics, realizes high spatial resolution and contrast fluorescence imaging, and is used for real-time dynamic imaging and monitoring of a chemical kinetic reaction process mediated by the diagnosis and treatment agent compound. The fluorescent diagnosis and treatment agent compound can also be used for tumor chemical kinetics treatment.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical chemistry and pharmaceutical preparation technology, and relates to a near-infrared II region fluorescent diagnostic and therapeutic agent for real-time monitoring of Fenton-like reactions and tumor chemodynamic therapy. Specifically, it relates to the preparation of organic small molecules used to construct fluorescent diagnostic and therapeutic agents, the preparation of diagnostic and therapeutic agent complexes and diagnostic and therapeutic agent nanoparticles, near-infrared II region (1000-1700 nm) fluorescence imaging and tumor chemodynamic therapy, and the application of the diagnostic and therapeutic agent nanoparticles in real-time imaging of copper ion-mediated Fenton-like reactions and tumor chemodynamic therapy in living tumors. Background Art

[0002] The prior art discloses that near-infrared light has been widely used in the biomedical field due to its advantages such as deeper tissue penetration and lower tissue background interference. Studies have shown that the longer the excitation wavelength used in biological imaging, the weaker the tissue light scattering and autofluorescence. At the same time, tissue absorption reaches a minimum in the range of 650-1450 nm. Therefore, compared with traditional visible light region (400-750 nm) and near-infrared I window (750-950 nm) fluorescence imaging, near-infrared II region fluorescence imaging can achieve higher spatial resolution and contrast due to its longer wavelength.

[0003] Chemodynamic therapy is a new cancer treatment method that mainly uses Fe 2+ , Cu + and Mn 2+ Transition metal ions such as hydroxyl radicals (•OH) induce Fenton or Fenton-like reactions in the tumor microenvironment, catalyzing hydrogen peroxide (H2O2) to generate cytotoxic hydroxyl radicals (•OH), thereby inducing cell apoptosis and achieving anti-tumor effects. Compared with traditional cancer treatments, chemodynamic therapy is non-invasive and highly effective, and exhibits low toxic side effects.

[0004] Electron spin resonance and a variety of fluorescence or Raman imaging probes are often used in current research to detect the various reactive oxygen species produced by the reaction, thereby indirectly monitoring the occurrence of chemical kinetic reactions. In addition, magnetic resonance imaging and other technologies can also be used to monitor Fe3O4 and MnO2 or their released Fe 3+ and Mn 2+ The distribution of Fenton or Fenton-like reaction catalysts in the tumor can be used to predict the therapeutic effect of chemodynamic therapy. However, since chemical reactions are affected by many factors, only detecting reactants or reactive oxygen products cannot accurately reflect the process of chemodynamic reactions and make accurate predictions on the efficacy of chemodynamic therapy.

[0005] In addition, current research lacks real-time dynamic imaging of the chemical kinetic reaction process of chemical kinetic drugs at the living cell level. Therefore, it is urgent to design a diagnostic agent that can directly participate in and monitor the dynamic process of chemical kinetic reactions in real time, so as to provide direct information for predicting the efficacy of chemical kinetic drugs. Summary of the invention

[0006] The object of the present invention is to provide a fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic treatment properties.

[0007] Another object of the present invention is to provide application of the above-mentioned composite.

[0008] The object of the present invention is achieved by the following scheme: a fluorescent diagnostic and therapeutic agent complex with near-infrared II region fluorescence imaging and chemodynamic treatment performance, the complex includes an organic fluorescent small molecule and a copper ion chelate, the organic fluorescent small molecule and the copper ion have a binding molar ratio of 1:0.1-1:1.64, the fluorescence is quenched due to the intramolecular charge transfer between the copper ion and the fluorescent small molecule, and a chemodynamic reaction occurs with glutathione in a weakly acidic environment to generate active oxygen clusters and generate o-benzoquinone products, emitting near-infrared II region fluorescence, wherein the organic fluorescent small molecule can emit near-infrared II region fluorescence (1000-1700 nm), and its structure is as follows: Its structure is shown in general formula I: One of them.

[0009] The organic fluorescent small molecule uses triphenylamine-substituted coupled benzothiadiazole as the parent core, and introduces catechol groups at both ends through Schiff base reaction, and has near-infrared II region fluorescence imaging and metal ion chelation properties. The organic small molecule of general formula I has strong ultraviolet absorption at 650-700 nm, and exhibits a strong fluorescence emission peak at 1050 nm under 660 nm laser irradiation.

[0010] Preferably, the molar ratio of the organic fluorescent small molecule to the copper ion is 1:1.64.

[0011] Based on the above scheme, the complex is a nanoparticle formed by an organic fluorescent small molecule, copper ions and a surfactant, wherein the molar ratio of the organic fluorescent small molecule, copper ions and surfactant content is 1: (0.1-1.64): (0.16-1.6), and the average particle size of the nanoparticles is 140-300 nm. For example, when the amount of the organic fluorescent small molecule is 1 mol, the amount of copper ions can be 0.1-1.64 mol, and the amount of the surfactant can be 0.16-1.6 mol.

[0012] In the present invention, the complex formed by the chelation of the organic fluorescent small molecule and the copper ion has the following structural formula II: , Due to the intramolecular charge transfer between copper ions and fluorescent small molecules, fluorescence quenching occurs. In a weakly acidic environment, a chemical kinetic reaction occurs with glutathione to produce reactive oxygen species and generate o-benzoquinone products, emitting near-infrared II region fluorescence.

[0013] Cu 2+ The complex formed by chelation with the above-mentioned organic fluorescent small molecules almost completely quenches the fluorescence of the organic small molecules due to the charge transfer within the molecule. Taking IR26 (Quantum yield, QY = 0.5%) as a reference, the quantum efficiency of the complex under 660 nm laser excitation is only about 0.10-0.17%.

[0014] Preferably, the surfactant is Pluronic F-127 or Pluronic F-68 or distearoylphosphatidylethanolamine-methoxypolyethylene glycol 2000 (DSPE-PEG 2000 ) or distearoylphosphatidylethanolamine-methoxypolyethylene glycol 5000 (DSPE-PEG 5000 )

[0015] The present invention also provides an organic fluorescent small molecule as described above, which uses aldehyde-substituted triphenylamine as a raw material, and undergoes bromination, Suzuki reaction, and Schiff base reaction to obtain a target product shown in general formula I. The synthesis route is as follows: Among them, R1 is , One of the following; R2 is hydrogen or methoxy.

[0016] The reagents used in the above steps are: (a) N-bromosuccinimide, N,N-dimethylformamide, chloroform, 60°C; (b) p-Toluenesulfonic acid monohydrate, triethylamine, toluene, room temperature; (c) n-butyllithium, -78°C; (d) Tetrakistriphenylphosphine palladium, dioxane, 100 °C; (e) Glacial acetic acid, methanol, tetrahydrofuran, 74°C.

[0017] The prepared diagnostic and therapeutic agent nanoparticles are characterized by.

[0018] The present invention also provides an application of the fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapeutic properties in chemodynamic drugs.

[0019] Furthermore, the complex undergoes a chemical kinetic reaction with glutathione in a weakly acidic environment to produce reactive oxygen species and generate o-benzoquinone products, which can emit near-infrared II region fluorescence under 660 nm and 830 nm laser irradiation, which can be used for real-time dynamic imaging and monitoring of the chemical kinetic reaction process mediated by the complex.

[0020] The present invention further provides an application of the fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapeutic properties in cancer treatment drugs.

[0021] The present invention relates to a near-infrared II region fluorescent diagnostic and therapeutic agent, which is a stable complex formed by chelating an organic fluorescent small molecule with a copper ion, can directly participate in a chemodynamic reaction, generate active oxygen clusters, and produce o-benzoquinone products, and emit near-infrared II region fluorescence, and is used for real-time monitoring of chemodynamic reactions mediated by the complex and chemodynamic treatment.

[0022] The complex of the present invention can also be in the form of nanoparticles, which are prepared from organic fluorescent small molecules, copper ions and surfactants. After intravenous injection, the diagnostic and therapeutic agent nanoparticles can be used for chemical dynamic reaction monitoring at the cellular level of living tumors and tumor chemical dynamic treatment due to their good tumor accumulation ability.

[0023] The complex of the present invention has the effect of integrated tumor diagnosis and treatment, and is a complex formed by chelating organic fluorescent small molecules with copper ions, such as structural formula II.

[0024] Furthermore, the present invention provides a method for preparing the above-mentioned diagnostic and therapeutic agent complex and diagnostic and therapeutic agent nanoparticles containing the fluorescent molecule, which comprises the steps of: (1) dissolving the fluorescent molecule and Cu(OAc)2 in DMF in a certain ratio; (2) The organic solution 60 o C for 2 h to form a chelate; (3) Centrifuging the organic solution in step (2) and collecting the precipitate, and washing it with DMF three times to obtain the desired diagnostic and therapeutic agent complex; (4) dissolving the diagnostic and therapeutic agent complex and the surfactant in step (3) in DMF; (5) Under ultrasonic and ice-water bath conditions, add the organic solution of step (4) into pure water and mix well; (6) Stir the mixture obtained in step (5) at room temperature for 4-12 hours; (7) concentrating the nanoparticle solution obtained in step (6) to obtain the desired nanoparticles; Preferably, the molar ratio of the fluorescent molecule to Cu(OAc)2 in step (1) is 1:1.64.

[0025] Preferably, in step (3), Pluronic F-127 or Pluronic F-68 or DSPE-PEG 2000 or DSPE-PEG 5000 The molar ratio of the fluorescent molecule described in step (1) is 0.8:1.

[0026] Preferably, the ratio of the organic solvent to pure water in step (5) is 1:10.

[0027] Preferably, the stirring time in step (6) is 8-12 h.

[0028] The water solubility of the composite nanoparticles prepared by the present invention is significantly improved, and can be applied to intravenous administration in mice. Animal experimental results show that after intravenous injection in mice, the diagnostic and therapeutic agent nanoparticles can accumulate at the tumor site, and under the stimulation of the low pH value and high concentration of reduced GSH unique to the tumor microenvironment, specifically mediate the chemical kinetic reaction to generate o-benzoquinone products, and produce near-infrared II region fluorescence after 660 nm and 830 nm laser irradiation, which can be used to monitor the dynamic process of the chemical kinetic reaction in real time. Further tumor chemodynamic therapy experimental results show that the diagnostic and therapeutic agent nanoparticles have good chemodynamic therapy ability in vivo and can effectively kill tumor cells.

[0029] The present invention adopts the diagnostic and therapeutic agent complex prepared by the organic small molecule and copper ion, and utilizes Pluronic F-127 or Pluronic F-68 or DSPE-PEG 2000 or DSPE-PEG 5000As a surfactant, the prepared therapeutic agent nanoparticles can be uniformly dispersed in aqueous solution. The prepared therapeutic agent complex and therapeutic agent nanoparticles participate in chemical kinetic reactions under weak acidic pH and glutathione (Glutathione, GSH) catalysis to generate o-benzoquinone products, emit near-infrared II region fluorescence, and can be used for real-time monitoring of the chemical kinetic reaction. The prepared therapeutic agent nanoparticles have good tumor accumulation properties and can be used for real-time imaging of copper ion-mediated chemical kinetic reactions in living tumors, providing dynamic information of chemical kinetic reactions at the tissue / organ and cell levels, and used for tumor chemokine therapy. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 , SEM images of Cu-BADA complex before and after chemical kinetic reaction at pH 5.5 and GSH (10 mM); Figure 2 , transmission electron microscopy images and particle size distribution of Cu-BADA nanoparticles; Figure 3 , (a) Fluorescence spectrum of compound BADA (10 μM) in DMF under 660 nm excitation; (b) Fluorescence excitation spectrum of Cu-BADA nanoparticles (100 μM) solution before and after reaction with GSH (10 mM) under acidic conditions (before and after dissolution) at 1000 nm; (c) Visible light photograph of Cu-BADA nanoparticles (100 μM) after dissolution and the solution irradiated with lasers at 660 nm and 830 nm (laser power density was 0.1 W / cm 2 ) Fluorescence photograph of near-infrared II region; (d) Fluorescence spectra of Cu-BADA nanoparticles (100 μM) before and after dissolution under excitation at 660 nm and 830 nm, respectively; Figure 4 The generation of reactive oxygen species was detected by electron spin resonance technology, which showed that after the aqueous solution of Cu-BADA nanoparticles (100 μM) reacted with GSH (2.5 mM) at pH 5.5, the generation of ·OH and superoxide anions (O2 ·- ) generation; Figure 5 , 0-24h after intravenous administration of Cu-BADA nanoparticles (40 mg / kg) to mice bearing CT26-Luc orthotopic tumor model, the copper ion content in plasma and tumor, the total amount of BADA and the amount of BADA conversion; Figure 6, CT26-Luc subcutaneous tumor (a) and orthotopic tumor (b) model mice, 0-24 h after intravenous administration of Cu-BADA nanoparticles (40 mg / kg), visible light photos of the tumor site and corresponding near-infrared II fluorescence imaging (660 / 830 nm, laser power density 0.1 W / cm 2 ); Arrows indicate the location of subcutaneous tumors or tumors in situ; Figure 7 , 3-4h after intravenous administration of Cu-BADA nanoparticles (40 mg / kg) to mice bearing CT26-Luc subcutaneous tumor model, near-infrared II fluorescence in vivo microscopic imaging of the tumor site under 660 or 830 nm laser irradiation; Figure 8 , The chemodynamic treatment effect of mice bearing CT26-Luc orthotopic tumor model, showing the survival curves of mice in the phosphate-buffered saline group, the single-dose Cu-BADA nanoparticle treatment group, and the multiple-dose Cu-BADA nanoparticle treatment group (5 doses) (n = 5); **P<0.01, ***P<0.001, ****P<0.0001. DETAILED DESCRIPTION

[0031] The present invention is further described below with reference to the embodiments, but is not limited thereto.

[0032] Example 1 An organic fluorescent small molecule 8a (BADA) with the structural formula: Prepared according to the aforementioned organic fluorescent small molecule synthesis method: (1) Synthesis of compound 2 3.88 g of 21.80 mmol N-bromosuccinimide (NBS) was added dropwise to 5.00 g of 18.29 mmol 4-(diphenylamino)benzaldehyde in a mixed solution of 40 mL of chloroform / N,N-dimethylformamide (DMF) (v / v, 1:1) at room temperature. The mixed solution was stirred at 60°C for 12 h under nitrogen protection, quenched with water, and extracted with dichloromethane. The organic phase was washed with water and saturated brine, and dried over anhydrous sodium sulfate overnight. After removing dichloromethane, the residue was purified by silica gel chromatography and eluted with ethyl acetate / petroleum ether (1:30) to obtain 5.92 g of the desired product compound 2 with a yield of 92%.

[0033] 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J= 8.0 Hz, 2H), 7.57 (d, J =8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J = 7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0034] (2) Synthesis of compound 6 2.00 g of compound 2 (5.68 mmol), 1.93 g of triethyl orthoformate (6.65 mmol), and 16 mg of p-toluenesulfonic acid monohydrate (0.084 mmol) were dissolved in 5 mL of toluene, and then 2-3 drops of triethylamine were added. The reaction solution was stirred at room temperature for 30 min. After the reaction was completed, the excess solvent was removed by rotary evaporation, and the obtained solid was vacuum dried at room temperature overnight. The solid product was dissolved in 20 mL of THF. After the temperature was lowered to -78°C using dry ice, 7.1 mL of 11.36 mmol n-butyl lithium (1.6 M in Hexanes) was added dropwise to the THF solution. The reaction mixture was stirred under nitrogen for 1 h, and 4.62 g of 14.20 mol Bu3SnCl was added. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation. The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2. CH2Cl2 was removed under reduced pressure, and the obtained solid was dried in vacuo at room temperature overnight. The product was transferred to a 50 mL two-necked flask, and 10 mL of 1,4-dioxane was added. 0.33 g of 0.284 mmol Pd(PPh3)4 and 0.60 g of 1.70 mmol of 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole); the reaction solution was heated in an oil bath at 100°C for 12 h under nitrogen protection; after the reaction, the solvent was removed by rotary evaporation and purified by silica gel column chromatography using CH2Cl2 as the eluent to obtain the desired product compound 6 (0.35 g) with a yield of 28%.

[0035] 1 H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J =8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0036] (3) Synthesis of BADA The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 through an acid-catalyzed Schiff base reaction.

[0037] Compound 6 (73.6 mg, 0.10 mmol) was dissolved in 10 mL of THF to obtain a compound 6 solution; dopamine hydrochloride (56.9 mg, 0.30 mmol) was dissolved in 10 mL of anhydrous methanol to obtain a dopamine hydrochloride solution, and the two solutions were mixed to obtain a mixed solution; 2-3 drops of glacial acetic acid were added to the mixed solution as a catalyst, and the reaction solution was reacted in an oil bath at 74°C for 8 h; after the reaction was completed, the solvent was removed by rotary evaporation, and the product was purified by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent to obtain the desired product BADA (91.3 mg, 91%).

[0038] 1H NMR (600 MHz, DMSO-d6, δ): 8.89 (dd, J = 28.7, 14.1 Hz, 4H), 8.16 (d, J = 3.1 Hz, 2H), 7.42 (m, 3H), 7.36 (d, J = 8.4 Hz, 2H), 7.30 (d, J = 4.5 Hz, 1H), 7.17-7.13 (m, 14H), 6.82 (d, J = 8.1 Hz, 1H), 6.68 (d, J = 8.0 Hz, 5H), 6.63 (s, 4H), 6.61 (s, 1H), 6.56 (d, J = 8.4 Hz, 1H), 6.46 (dd, J = 8.0, 1.9 Hz, 2H), 2.90 - 2.86 (m, 4H), 2.71 (s, 4H). 13 C NMR (151 MHz, DMSO- d 6 , δ): 152.15, 146.51, 145.46, 144.22, 133.28, 131.41, 130.90, 130.09, 130.04, 128.13, 125.43, 123.73, 122.16, 119.35, 116.23, 115.95, 115.34, 114.57, 58.11, 55.09, 32.52. MALDI-TOF MS (ESI) m / z :[M+H] + Calculate for: C 60 H 47 N8O4S2 + , 1007.31; found: 1006.51.

[0039] Example 2 Preparation of a fluorescent diagnostic and therapeutic agent complex containing the compound BADA and its nanoparticles and evaluation of the complex 1) Cu-BADA complex was prepared according to the following steps: Cu(OAc)2 (39.6 μM, 7.2 mg) and BADA (9.9 μM, 10.0 mg) were dissolved in 5 mL DMF; the mixture was heated at 60°C for 2 h; after the reaction, the Cu-BADA precipitate was collected by centrifugation and washed three times with DMF to obtain a Cu-BADA complex, which could be uniformly dispersed in DMF by ultrasound.

[0040] The Cu-BADA complex was quenched due to the charge transfer within the molecule, and the quantum efficiency under 660 nm laser excitation was about 0.10. The Cu-BADA complex reacted chemically with glutathione (GSH) under acidic conditions and was degraded.

[0041] SEM images of Cu-BADA complex before and after chemical kinetic reaction at pH 5.5 and GSH (10 mM), as shown in Figure 1 shown.

[0042] 2) Cu-BADA nanoparticles: Pluronic F-127 (100 mg) was dissolved in 1 mL DMF, added to the Cu-BADA complex obtained in step 1), and ultrasonicated to make it uniformly dispersed; under magnetic stirring, the mixed solution was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and stirred at room temperature for 8 h to form a stable Cu-BADA nanoparticle solution.

[0043] The particle size of the prepared Cu-BADA nanoparticles was 190.1 ± 4.3 nm and the PDI was 0.11 ± 0.02. Figure 2 The transmission electron micrograph and particle size distribution are shown.

[0044] 3) Fluorescence characteristics of Cu-BADA nanoparticles before and after dissolution, see Figure 3 As shown: 10 μM compound BADA was excited at 660 nm in the benign polar solvent DMF, and its emission wavelength was 900 – 1400 nm, which was near-infrared II region fluorescence, such as Figure 3 As shown in a.

[0045] After the compound BADA was chelated with copper ions to prepare Cu-BADA nanoparticles, the fluorescence was quenched due to the charge transfer within the molecule; 100 μM Cu-BADA nanoparticle solution could react with 10 mM GSH in an acidic environment (pH 5.5) to cause the nanoparticles to dissolve, and the fluorescence was restored as shown in Figure 2. Figure 3 b to Figure 3 As shown in d.

[0046] Figure 3 b is the fluorescence excitation spectrum of Cu-BADA nanoparticles at 1000 nm before and after the chemical kinetic reaction (before and after dissolution) under acidic and GSH conditions. The o-benzoquinone products after the reaction have fluorescence intensity excitation peaks at 403 / 634 / 836 nm, respectively.

[0047] Figure 3c is the visible light photograph of Cu-BADA nanoparticles (100 μM) after dissolution and the solution irradiated with laser at 660 nm and 830 nm (laser power density was 0.1 W / cm 2 ) is a near-infrared II fluorescence photograph, which shows that the fluorescence is quenched before dissolution and recovered after dissolution.

[0048] like Figure 3 As shown in Figure d, the fluorescence of Cu-BADA nanoparticles was quenched before dissolution and recovered after dissolution. The fluorescence was excited by 660 nm and 830 nm, emitting near-infrared II region fluorescence with a wavelength of 1000-1300 nm, with quantum efficiencies of 0.70% and 1.2%, respectively. The fluorescence intensity excited by 830 nm was greater than that excited by 660 nm.

[0049] 4) Detection of active oxygen in the Cu-BADA nanoparticle solution obtained in step 2): In vitro • OH and O2 -· It can be detected by electron spin resonance technology.

[0050] 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was used as the •OH in water and O2 in methanol. -• The experimental group includes: (1) Cu-BADA nanoparticles (100 μM), pH = 7.4; (2) Cu-BADA nanoparticles (100 μM), pH = 5.5; (3) Cu-BADA nanoparticles (100 μM) + GSH, pH = 5.5.

[0051] The results showed that the aqueous solution of Cu-BADA nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· (like Figure 4 as shown).

[0052] Electron spin resonance technology is used to detect the generation of reactive oxygen species. Figure 4 It is shown in the figure that the aqueous solution of Cu-BADA nanoparticles (100 μM) reacted with GSH (2.5 mM) at pH 5.5, and the intensity was stronger, and the detection of OH and superoxide anions (O2 ·- ) is generated.

[0053] The nanoparticles under neutral conditions have the weakest strength.

[0054] 5) Effect of aqueous solution of Cu-BADA nanoparticles on cancer: Mouse subcutaneous tumor model and orthotopic colon cancer model were established using mouse colon cancer CT26-Luc cell line, and Cu-BADA nanoparticles (40 mg / kg) were injected intravenously.

[0055] For the mouse orthotopic colon cancer model, the copper ion content, total BADA and BADA conversion in mouse plasma and orthotopic tumors were measured 0-24 hours after injection. The results of copper ion content, total BADA and BADA conversion in plasma and tumors are as follows Figure 5 As shown in Figure 2, in plasma, the amount of BADA conversion remains at a relatively low level within 0-24 h after injection, which helps to reduce the interference of background fluorescence signals in normal tissues. Figure 5 As shown in the right figure, 4 h after injection, the total amount of BADA and the amount of BADA conversion in the tumor increased by about 6.6 times and 5.8 times compared with 3 h, respectively, indicating that copper ions underwent a chemokinetic reaction in the tumor. The total amount of BADA, the amount of BADA conversion, and the copper ion concentration in the tumor all reached a peak at 6 h after injection and then gradually decreased. After 6 h of intravenous injection, the amount of BADA conversion was close to the total amount, indicating that the Cu-BADA nanoparticles were almost completely dissociated and converted into o-benzoquinone products. 12 h after injection, the copper ion concentration in the tumor was close to that at 2 h after injection, indicating that the chemokinetic reaction in the tumor was terminated.

[0056] For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results are shown in Figure 6 As shown in the figure, in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. The fluorescence intensity reached the highest level 6 h after injection and then gradually weakened until 12 h, when the fluorescence disappeared.

[0057] Cu-BADA nanoparticles (40 mg / kg) were intravenously injected into mice bearing CT26-Luc orthotopic tumors for 3-4 hours, and near-infrared II region fluorescence microscopy imaging was performed using 660nm and 830nm lasers. The results showed that 3 hours after injection, fluorescence signals began to appear in tumor cells and gradually increased over time, and the fluorescence images obtained under 830nm laser irradiation had higher fluorescence intensity and clarity than those obtained under 660nm laser irradiation (e.g. Figure 7 as shown).

[0058] The chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of Cu-BADA nanoparticles (40 mg / kg). The cure rate of the tumor-bearing mice in the multi-dose treatment group (5 doses) was 80%, and the tumor did not recur within 60 days. The single-dose treatment group could significantly prolong the survival time of the tumor-bearing mice. All the mice in the phosphate buffered saline solution group relapsed and died within 60 days, which proved the effectiveness of the Cu-BADA nanoparticles of the present invention in the treatment of tumors (such as Figure 8 as shown).

[0059] Example 3 The binding formula of the organic fluorescent small molecule 8b is as follows: Prepared by the following synthetic method: (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature, and the mixture was stirred at 60 °C for 12 h under nitrogen protection, then quenched with water and extracted with dichloromethane; the organic phase was washed with water and saturated brine, and dried over anhydrous sodium sulfate overnight; after removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%).

[0060] 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J = 8.0 Hz, 2H), 7.57 (d, J =8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J = 7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO +, 351.03; found: 351.07.

[0061] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol), triethyl orthoformate (1.93 g, 6.65 mmol), and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol) were dissolved in 5 mL of toluene, and then 2-3 drops of triethylamine were added. The reaction solution was stirred at room temperature for 30 min. The excess solvent was removed by rotary evaporation, and the obtained solid was dried in vacuum at room temperature overnight; The above product was dissolved in 20 mL of THF; After the temperature was lowered to -78°C using dry ice, n-butyllithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution to obtain a reaction mixture; The reaction mixture was stirred under nitrogen for 1 h, and then Bu3SnCl (4.62 g, 14.20 mol) was added. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation, leaving a viscous solid; The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2; CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuo at room temperature overnight; The dried solid product was transferred to a 50 mL double-necked flask, and 10 mL of 1,4-dioxane was added; Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added to obtain a reaction solution; The reaction solution was heated in an oil bath at 100°C under nitrogen protection for 12 h; the solvent was removed by rotary evaporation, and the product was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product compound 6 (0.35 g, 28%).

[0062] 1 H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J =8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0063] (3) Synthesis of organic fluorescent small molecule 8b The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction according to the following steps: Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), and 4-aminocatechol (37.5 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed to obtain a mixed solution; 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was reacted in a 74°C oil bath for 8 h; the solvent was removed by rotary evaporation; and the product was purified by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent to obtain the desired product 8b (81.3 mg, 85%).

[0064] MALDI-TOF MS (ESI) m / z :[M+H] + Calculate for: C 56 H 38 N8O4S2 + ,950.25;found:950.67.

[0065] Example 4 Preparation and evaluation of the therapeutic agent complex and its therapeutic agent nanoparticles containing organic fluorescent small molecule 8b 1) Preparation of complex Cu(OAc)2 (39.6 μM, 7.2 mg) and organic fluorescent small molecule 8b (9.9 μM, 8.05 mg) were dissolved in 5 ml of DMF to obtain a mixed solution; the mixed solution was heated at 60°C for 2 h, the precipitate was collected by centrifugation, and washed three times with DMF to obtain a diagnostic and therapeutic agent complex.

[0066] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, emitting near-infrared II region fluorescence.

[0067] 2) Preparation of diagnostic and therapeutic nanoparticles Surfactant DSPE-PEG 2000 (23 mg) was dissolved in 1 mL DMF and added to the diagnostic and therapeutic agent complex prepared in step 1), and ultrasonicated to make it uniformly dispersed; under magnetic stirring, the mixed solution was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and stirred at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles.

[0068] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0069] 3) Chemical kinetic reaction characteristics of diagnostic and therapeutic nanoparticles: In vitro • OH and O2 -· It can be detected by electron spin resonance technology, using DMPO as the •OH in water and O2 in methanol -• The results showed that the aqueous solution of the therapeutic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0070] 4) Effects of therapeutic nanoparticles in cancer diagnostic and therapeutic drugs: The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model.

[0071] The therapeutic nanoparticles (40 mg / kg) were injected intravenously, and near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers for subcutaneous tumor models in mice 0-24 h after injection.

[0072] For the mouse orthotopic colon cancer model, the skin and peritoneum at the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging of the orthotopic tumor was performed using 660 nm and 830 nm lasers 0–24 h after injection.

[0073] The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 hours after injection. The fluorescence intensity reached its peak 6 hours after injection and then gradually weakened until 12 hours after injection, when the fluorescence disappeared.

[0074] This proves that the therapeutic nanoparticles can mediate specific chemokinetic reactions in the mouse colon cancer model, and the o-benzoquinone products with near-infrared II region fluorescence generated by the therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemokinetic reaction process of the therapeutic nanoparticles in mouse colon cancer, achieving the effect of integrated diagnosis and treatment.

[0075] The CT26-Luc orthotopic tumor mice were treated with chemodynamic therapy by intravenous injection of therapeutic nanoparticles (40 mg / kg). In the multiple-dose treatment group, the tumor did not recur within 60 days after the treatment after 5 administrations; the single-dose treatment group significantly prolonged the survival time of tumor-bearing mice, while all the mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0076] Example 5 Organic fluorescent small molecule 8c, the chemical structure is as follows: Prepared by the following synthetic method: (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature; The mixed solution was stirred at 60 °C for 12 h under nitrogen protection, quenched with water, and extracted with dichloromethane; the organic phase was washed with water and saturated brine, and dried over anhydrous sodium sulfate overnight; after removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%).

[0077] 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J = 8.0 Hz, 2H), 7.57 (d, J =8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J = 7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J= 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0078] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol), triethyl orthoformate (1.93 g, 6.65 mmol), and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol) were dissolved in 5 mL of toluene, and 2-3 drops of triethylamine were added. The reaction solution was stirred at room temperature for 30 min, and then the excess solvent was removed by rotary evaporation. The obtained solid was dried in vacuum at room temperature overnight. The solid product was dissolved in 20 mL of THF. After the temperature was lowered to -78°C using dry ice, n-butyl lithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution to obtain a reaction mixture. The reaction mixture was stirred under nitrogen for 1 h, and then Bu3SnCl (4.62 g, 14.20 mol) was added. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation, leaving a viscous solid. The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2; CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuo at room temperature overnight; The solid product was transferred to a 50 mL double-necked flask, and 10 mL of 1,4-dioxane was added; Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added to obtain a reaction solution; The reaction solution was heated in an oil bath at 100°C under nitrogen protection for 12 h; the solvent was removed by rotary evaporation, and the mixture was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product compound 6 (0.35 g, 28%); 1 H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J =8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J= 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0079] (3) Synthesis of compound 8c The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction: Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), and drotaverine impurity 20 (41.7 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed to obtain a mixed solution; Add 2-3 drops of glacial acetic acid as a catalyst to the mixture, and react the reaction solution in an oil bath at 74°C for 8 h; then remove the solvent by rotary evaporation; The product was purified by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent to obtain the desired product 8c (82.2 mg, 84%).

[0080] MALDI-TOF MS (ESI) m / z :[M+H] + Calculate for: C 58 H 42 N8O4S2 + , 978.28; found: 977.97.

[0081] Example 6 Preparation and evaluation of theranostic complexes and nanoparticles containing the organic fluorescent small molecule 8c 1) The diagnostic and therapeutic agent complex containing the organic fluorescent small molecule 8c was prepared according to the following method: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8c (9.9 μM, 9.68 mg) were dissolved in 5 ml DMF to obtain a mixed solution; After the mixture was heated at 60°C for 2 h, the precipitate was collected by centrifugation and washed three times with DMF to obtain the diagnostic and therapeutic agent complex.

[0082] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, emitting near-infrared II region fluorescence.

[0083] 2) Preparation of diagnostic and therapeutic nanoparticles 47 mg of DSPE-PEG 5000 Dissolve in 1 mL DMF, add to the diagnostic and therapeutic agent complex prepared in step 1), and ultrasonicate to uniformly disperse the mixture; under magnetic stirring, drop the mixture into 10 mL ultrapure water, ultrasonicate for 10 min, and stir at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles.

[0084] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0085] 3) Chemical kinetic reaction characteristics of diagnostic and therapeutic nanoparticles: In vitro • OH and O2 -· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O2 in methanol. -• The results showed that the aqueous solution of the diagnostic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0086] 4) Effects of diagnostic and therapeutic nanoparticles in cancer treatment The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model.

[0087] The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously, and for the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers at 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut to expose the intestinal segment, and near-infrared II fluorescence imaging of the orthotopic tumor was performed using 660 nm and 830 nm lasers at 0-24 h after injection.

[0088] The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 hours after injection. The fluorescence intensity reached its peak 6 hours after injection and then gradually weakened until 12 hours, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the o-benzoquinone products with near-infrared II fluorescence generated by the diagnostic and therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemical kinetic reaction process of the diagnostic and therapeutic nanoparticles in mouse colon cancer. It can be used as a diagnostic agent to achieve high spatial resolution and contrast fluorescence imaging of cancer.

[0089] The chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of diagnostic and therapeutic nanoparticles (40 mg / kg). In the multiple-dose treatment group, the cure rate of tumor-bearing mice was 80% after 5 doses of the drug, and the tumor did not recur within 60 days. As a drug for chemodynamic therapy, it has a significant effect; the single-dose treatment group can significantly prolong the survival time of tumor-bearing mice; all mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0090] Example 7 Organic fluorescent small molecule 8d, the chemical structure is as follows: Prepared by the following synthetic method: (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature to obtain a mixture; The mixture was stirred at 60 °C for 12 h under nitrogen protection, quenched with water, and extracted with dichloromethane; the organic phase was washed with water and saturated brine, and dried over anhydrous sodium sulfate overnight; after removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%).

[0091] 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J = 8.0 Hz, 2H), 7.57 (d, J =8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J = 7.3 Hz, 1H), 7.19 (d, J= 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0092] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol), triethyl orthoformate (1.93 g, 6.65 mmol), and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol) were dissolved in 5 mL of toluene, and 2-3 drops of triethylamine were added. The reaction solution was stirred at room temperature for 30 min, and then the excess solvent was removed by rotary evaporation. The obtained solid was dried in vacuum at room temperature overnight. The solid product was dissolved in 20 mL of THF. After the temperature was lowered to -78°C using dry ice, n-butyl lithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution. After the reaction mixture was stirred for 1 h under nitrogen protection, Bu3SnCl (4.62 g, 14.20 mol) was added to obtain a mixture. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation; the remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2; CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuo at room temperature overnight; The solid product was transferred to a 50 mL double-necked flask, and 10 mL of 1,4-dioxane was added, followed by Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) to obtain a reaction solution; The reaction solution was heated in an oil bath at 100°C for 12 h under nitrogen protection. After the reaction, the solvent was removed by rotary evaporation and the mixture was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product compound 6 (0.35 g, 28%).

[0093] 1 H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J= 8.3 Hz, 4H), 7.79 (d, J =8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0094] (3) Synthesis of compound 8d The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction: Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), 1,2-Benzenediol,5-(aminomethyl)-3-methoxy-, hydrochloride (1:1) (50.3 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed to obtain a mixed solution; 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was reacted in an oil bath at 74°C for 8 h; the solvent was removed by rotary evaporation; and the product was purified by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent to obtain the desired product 8d (75.9 mg, 73%).

[0095] MALDI-TOF MS (ESI) m / z :[M+H] + Calculate for: C 60 H 46 N8O6S2 + , 1038.30; found: 1038.89.

[0096] Example 8 Preparation and evaluation of theranostic complexes and nanoparticles containing organic fluorescent small molecule 8d 1) The diagnostic and therapeutic agent complex containing the organic fluorescent small molecule 8d was prepared according to the following method: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8d (9.9 μM, 10.3 mg) were dissolved in 5 ml DMF. The mixture was heated at 60 °C for 2 h. The precipitate was collected by centrifugation and washed three times with DMF to obtain a diagnostic and therapeutic agent complex.

[0097] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, emitting near-infrared II region fluorescence.

[0098] 2) The diagnostic and therapeutic nanoparticles containing the organic fluorescent small molecule 8d were prepared according to the following method: DSPE-PEG 5000 (47 mg) was dissolved in 1 mL of DMF and added to the diagnostic and therapeutic agent complex prepared in step 1), and ultrasonically dispersed to obtain a mixed solution; under magnetic stirring, the mixed solution was dropped into 10 mL of ultrapure water, ultrasonically treated for 10 min, and then stirred at room temperature for 8 h to form a stable diagnostic and therapeutic agent nanoparticle solution.

[0099] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0100] 3) Chemical kinetic reaction characteristics of diagnostic and therapeutic nanoparticles In vitro • OH and O2 -· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O2 in methanol. -• The results showed that the aqueous solution of the diagnostic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0101] 4) Application of therapeutic nanoparticles in the treatment and diagnosis of cancer The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model.

[0102] The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously. For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. After 6 h after injection, the fluorescence intensity reached the highest level and then gradually weakened until 12 h, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the o-benzoquinone products with near-infrared II fluorescence generated by the diagnostic and therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemical kinetic reaction process of the diagnostic and therapeutic nanoparticles in mouse colon cancer.

[0103] The chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of diagnostic and therapeutic nanoparticles (40 mg / kg). In the multiple-dose treatment group, the drug was administered five times, and the tumor did not recur in the tumor-bearing mice within 60 days after treatment. The chemodynamic therapy for mice bearing CT26-Luc orthotopic tumors was effective; the single-dose treatment group could significantly prolong the survival time of tumor-bearing mice, and all mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0104] Embodiment 9: Organic fluorescent small molecule 8e, the chemical structure is as follows: Prepared by the following synthetic method: (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature; the mixed solution was stirred at 60 °C for 12 h under nitrogen protection, then quenched with water and extracted with dichloromethane; the organic phase was washed with water and saturated brine, and dried over anhydrous sodium sulfate overnight; after removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%).

[0105] 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J = 8.0 Hz, 2H), 7.57 (d,J =8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J = 7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0106] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol), triethyl orthoformate (1.93 g, 6.65 mmol), and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol) were dissolved in 5 mL of toluene, and 2-3 drops of triethylamine were added. The reaction solution was stirred at room temperature for 30 min, and then the excess solvent was removed by rotary evaporation. The obtained solid was dried in vacuum at room temperature overnight. The solid product was dissolved in 20 mL of THF. After the temperature was lowered to -78°C using dry ice, n-butyl lithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution to obtain a reaction mixture. The reaction mixture was stirred for 1 h under nitrogen protection, and then Bu3SnCl (4.62 g, 14.20 mol) was added; the mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation; the remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2; CH2Cl2 was removed under reduced pressure, and the obtained solid was dried in vacuum at room temperature overnight; The solid product was transferred to a 50 mL double-necked flask, and 10 mL of 1,4-dioxane was added; then Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added to obtain a reaction solution; The reaction solution was heated in an oil bath at 100°C for 12 h under nitrogen protection. After the reaction, the solvent was removed by rotary evaporation and the mixture was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product compound 6 (0.35 g, 28%).

[0107] 1 H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J =8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0108] (3) Synthesis of compound 8e The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction: Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), and DL-norepinephrine (50.7 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed to obtain a mixed solution; 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was reacted in a 74°C oil bath for 8 h. After the reaction, the solvent was removed by rotary evaporation. The product was purified by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent to obtain the desired product 8e (77.9 mg, 75%).

[0109] MALDI-TOF MS (ESI) m / z :[M+H] +Calculate for: C 60 H 46 N8O6S2 + , 1038.30; found: 1037.79.

[0110] Embodiment 10: Preparation and evaluation of diagnostic and therapeutic agent complexes and diagnostic and therapeutic agent nanoparticles containing organic fluorescent small molecule 8e 1) The diagnostic and therapeutic agent complex containing the organic fluorescent small molecule 8e was prepared according to the following steps: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8c (9.9 μM, 10.3 mg) were dissolved in 5 ml of DMF to obtain a mixed solution; The mixed solution was heated at 60°C for 2 h. After the reaction was completed, the precipitate was collected by centrifugation and washed three times with DMF to obtain a diagnostic and therapeutic agent complex.

[0111] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, emitting near-infrared II region fluorescence.

[0112] 2) The diagnostic and therapeutic nanoparticles containing the organic fluorescent small molecule 8e were prepared according to the following steps: DSPE-PEG 5000 (47 mg) was dissolved in 1 mL DMF and added to the above diagnostic and therapeutic agent complex, and ultrasonicated to make it uniformly dispersed. Under magnetic stirring, the mixture was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and then stirred at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles.

[0113] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0114] 3) Chemical kinetic reaction characteristics of diagnostic and therapeutic nanoparticles: In vitro • OH and O2 -· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O2 in methanol. -• The results showed that the aqueous solution of the therapeutic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0115] 4) Application of diagnostic and therapeutic nanoparticles in cancer diagnostic and therapeutic drugs The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model. The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously. For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. After 6 h after injection, the fluorescence intensity reached the highest level and then gradually weakened until 12 h, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the o-benzoquinone product with near-infrared II fluorescence generated by the diagnostic and therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemical kinetic reaction process of the diagnostic and therapeutic nanoparticles in mouse colon cancer.

[0116] The CT26-Luc orthotopic tumor mice were treated with chemodynamic therapy by intravenous injection of therapeutic nanoparticles (40 mg / kg). The cure rate of the tumor-bearing mice in the multi-dose treatment group (5 doses) was 80%, and the tumor did not recur within 60 days, indicating a significant therapeutic effect; the single-dose treatment group significantly prolonged the survival time of the tumor-bearing mice, and all the mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0117] Example 11: Organic fluorescent small molecule 8f, the chemical structure is: Prepared by the following synthetic method (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature. The mixture was heated at 60 °C under nitrogen. o C was stirred for 12 h, then quenched with water and extracted with dichloromethane. The organic phase was washed with water and saturated brine and dried over anhydrous sodium sulfate overnight. After removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%). 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J= 8.0 Hz, 2H), 7.57 (d, J = 8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J =7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0118] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol) was dissolved in 5 mL of toluene with triethyl orthoformate (1.93 g, 6.65 mmol) and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol). Then 2-3 drops of triethylamine were added and the reaction solution was stirred at room temperature for 30 min. After the reaction was completed, the excess solvent was removed by rotary evaporation and the obtained solid was vacuum dried at room temperature overnight. The above product was dissolved in 20 mL of THF. The temperature was lowered to -78 °C using dry ice. o C, n-butyllithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution. After the reaction mixture was stirred for 1 h under nitrogen protection, Bu3SnCl (4.62 g, 14.20 mol) was added. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation. The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2. CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuo at room temperature overnight. The above product was transferred to a 50 mL two-necked flask and 10 mL of 1,4-dioxane was added. Then Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added. The reaction solution was stirred at 100 °C under nitrogen protection. oThe reaction was heated in an oil bath at C for 12 h. After the reaction was completed, the solvent was removed by rotary evaporation and the mixture was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product, compound 6 (0.35 g, 28%). 1 H NMR (400MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J = 8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0119] (3) Synthesis of compound 8f The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction. Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), 1,2-Benzenediol, 4-(2-amino-1-methylethyl)-, (±)-,methanesulfonate (salt) (50.4 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed. 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was heated at 74 o The reaction was carried out in an oil bath of C for 8 h. After the reaction was completed, the solvent was removed by rotary evaporation. The desired product 8f (95.1 mg, 92%) was obtained by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent. MALDI-TOF MS (ESI) m / z :[M+H] + Calculate for: C 62H 50 N8O4S2 + , 1034.34; found: 1035.37.

[0120] Example 12: Preparation and evaluation of diagnostic and therapeutic agent complexes and diagnostic and therapeutic agent nanoparticles containing organic fluorescent small molecule 8f 1) The diagnostic and therapeutic agent complex containing the organic fluorescent small molecule 8f was prepared according to the following method: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8f (9.9 μM, 10.2 mg) were dissolved in 5 ml DMF. o After the reaction was completed, the precipitate was collected by centrifugation and washed three times with DMF to obtain the diagnostic and therapeutic agent complex.

[0121] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, and emits near-infrared II region fluorescence.

[0122] 2) The diagnostic and therapeutic nanoparticles containing the organic fluorescent small molecule 8f were prepared according to the following method: Pluronic F-68 (67 mg) was dissolved in 1 mL DMF and added to the above-mentioned diagnostic and therapeutic agent complex, and ultrasonicated to make it uniformly dispersed. Under magnetic stirring, the mixture was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and then stirred at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles.

[0123] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0124] 3) Chemical kinetic reaction characteristics of diagnostic and therapeutic nanoparticles: In vitro • OH and O2 -· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O2 in methanol. -• The results showed that the aqueous solution of the therapeutic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0125] 4) Application of diagnostic and therapeutic nanoparticles in cancer diagnostic and therapeutic drugs The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model. The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously. For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. After 6 h after injection, the fluorescence intensity reached the highest level and then gradually weakened until 12 h, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the o-benzoquinone products with near-infrared II fluorescence generated by the diagnostic and therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemical kinetic reaction process of the diagnostic and therapeutic nanoparticles in mouse colon cancer.

[0126] The chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of diagnostic and therapeutic nanoparticles (40 mg / kg). The cure rate of mice bearing tumors in the multi-dose treatment group (5 doses) was 80% after treatment, and the tumor did not recur within 60 days. The chemodynamic therapy had a significant effect on mice bearing CT26-Luc orthotopic tumors; the single-dose treatment group could significantly prolong the survival time of tumor-bearing mice; all mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0127] Example 13: The chemical structure of the organic fluorescent small molecule 8g is: Prepared by the following synthetic method (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature. The mixture was heated at 60 °C under nitrogen. o C was stirred for 12 h, then quenched with water and extracted with dichloromethane. The organic phase was washed with water and saturated brine and dried over anhydrous sodium sulfate overnight. After removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%). 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J= 8.0 Hz, 2H), 7.57 (d, J = 8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J =7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0128] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol) was dissolved in 5 mL of toluene with triethyl orthoformate (1.93 g, 6.65 mmol) and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol). Then, 2-3 drops of triethylamine were added and the reaction solution was stirred at room temperature for 30 min. After the reaction was completed, the excess solvent was removed by rotary evaporation and the obtained solid was dried in vacuum at room temperature overnight. The above product was dissolved in 20 mL of THF. After the temperature was lowered to -78 °C using dry ice, n-butyl lithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution. After the reaction mixture was stirred for 1 h under nitrogen protection, Bu3SnCl (4.62 g, 14.20 mol) was added. The mixture was stirred at room temperature for 12 h and all volatile liquids were removed by rotary evaporation. The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2. CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuum at room temperature overnight. The above product was transferred to a 50 mL two-necked flask and 10 mL of 1,4-dioxane was added. Then Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added. The reaction solution was heated at 100 °C under nitrogen protection. o The reaction was heated in an oil bath at C for 12 h. After the reaction was completed, the solvent was removed by rotary evaporation, and the product was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product, compound 6 (0.35 g, 28%).1 H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J = 8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131 .33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0129] (3) Synthesis of compound 8g The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction. Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), and norepinephrine impurity 41 (54.9 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed. 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was heated at 74 o C oil bath for 8 h. After the reaction, the solvent was removed by rotary evaporation. Silica gel column chromatography was used as the eluent to obtain 8 g (83.1 mg, 78%) of the desired product. MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 62 H 50 N8O6S2 + , 1066.33; found: 1066.70.

[0130] Example 14: Preparation and evaluation of diagnostic and therapeutic agent complexes and diagnostic and therapeutic agent nanoparticles containing 8 g of organic fluorescent small molecules 1) A diagnostic and therapeutic agent complex containing 8 g of an organic fluorescent small molecule was prepared according to the following steps: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8g (9.9 μM, 10.6 mg) were dissolved in 5 ml DMF. o C for 2 h. After the reaction, the precipitate was collected by centrifugation and washed three times with DMF to obtain the diagnostic and therapeutic agent complex. The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex undergoes a chemical kinetic reaction with GSH and is degraded, and emits near-infrared II region fluorescence.

[0131] 2) The diagnostic and therapeutic nanoparticles containing 8 g of organic fluorescent small molecules were prepared according to the following steps: Pluronic F-127 (100 mg) was dissolved in 1 mL DMF and added to the above-mentioned diagnostic and therapeutic agent complex, and ultrasonicated to make it uniformly dispersed. Under magnetic stirring, the mixture was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and then stirred at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles.

[0132] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0133] 3) Chemical kinetic reaction characteristics of diagnostic and therapeutic nanoparticles: In vitro • OH and O2 -· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O2 in methanol. -• The results showed that the aqueous solution of the therapeutic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0134] 4) Application of diagnostic and therapeutic nanoparticles in cancer diagnostic and therapeutic drugs The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model. The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously. For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. After 6 h after injection, the fluorescence intensity reached the highest level and then gradually weakened until 12 h, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the o-benzoquinone products with near-infrared II fluorescence generated by the diagnostic and therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemical kinetic reaction process of the diagnostic and therapeutic nanoparticles in mouse colon cancer.

[0135] The chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of diagnostic and therapeutic nanoparticles (40 mg / kg). The cure rate of mice bearing tumors in the multi-dose treatment group (5 doses) was 80% after treatment, and the tumor did not recur within 60 days. The chemodynamic therapy was effective in mice bearing CT26-Luc orthotopic tumors; the single-dose treatment group could significantly prolong the survival time of tumor-bearing mice; all mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0136] Example 15: Organic fluorescent small molecule 8h, the chemical structure is: Prepared by the following synthetic method: (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature. The mixture was heated at 60 °C under nitrogen. o C was stirred for 12 h, then quenched with water and extracted with dichloromethane. The organic phase was washed with water and saturated brine and dried over anhydrous sodium sulfate overnight. After removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%). 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J= 8.0 Hz, 2H), 7.57 (d, J = 8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J =7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0137] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol) was dissolved in 5 mL of toluene with triethyl orthoformate (1.93 g, 6.65 mmol) and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol). Then 2-3 drops of triethylamine were added and the reaction solution was stirred at room temperature for 30 min. After the reaction was completed, the excess solvent was removed by rotary evaporation and the obtained solid was vacuum dried at room temperature overnight. The above product was dissolved in 20 mL of THF. The temperature was lowered to -78 °C using dry ice. o C, n-butyllithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution. After the reaction mixture was stirred for 1 h under nitrogen protection, Bu3SnCl (4.62 g, 14.20 mol) was added. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation. The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2. CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuo at room temperature overnight. The above product was transferred to a 50 mL two-necked flask and 10 mL of 1,4-dioxane was added. Then Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added. The reaction solution was stirred at 100 °C under nitrogen protection. oThe reaction was heated in an oil bath at C for 12 h. After the reaction was completed, the solvent was removed by rotary evaporation and the mixture was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product, compound 6 (0.35 g, 28%). 1 H NMR (400MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J = 8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151 MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131.33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0138] (3) Synthesis of compound 8h The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction. Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), and 4-(3-aminopropyl)-1,2-benzenediol (50.2 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed. 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was heated at 74 o The reaction was continued in a C oil bath for 8 h. After the reaction was completed, the solvent was removed by rotary evaporation. The desired product 8h (84.0 mg, 81%) was obtained by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent. MALDI-TOF MS (ESI) m / z :[M+H] + Calculate for: C 62 H 50 N8O4S2 +, 1034.34; found: 1035.34.

[0139] Example 16: Preparation and evaluation of diagnostic and therapeutic agent complexes and diagnostic and therapeutic agent nanoparticles containing organic fluorescent small molecule 8h 1) The diagnostic and therapeutic agent complex containing the organic fluorescent small molecule 8h was prepared according to the following steps: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8g (9.9 μM, 10.6 mg) were dissolved in 5 ml DMF. o After the reaction was completed, the precipitate was collected by centrifugation and washed three times with DMF to obtain the diagnostic and therapeutic agent complex.

[0140] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, emitting near-infrared II region fluorescence.

[0141] 2) The diagnostic and therapeutic nanoparticles containing organic fluorescent small molecules 8h were prepared according to the following method: Pluronic F-127 (100 mg) was dissolved in 1 mL DMF and added to the above-mentioned diagnostic and therapeutic agent complex, and ultrasonicated to make it uniformly dispersed. Under magnetic stirring, the mixture was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and then stirred at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles. The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to intramolecular charge transfer, which can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. It can be excited at 660 nm and 830 nm, and emits near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0142] 3) Chemical kinetics of diagnostic and therapeutic nanoparticles containing organic fluorescent small molecules for 8h In vitro • OH and O2 -· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O2 in methanol. -• The results showed that the aqueous solution of the therapeutic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O2 -· .

[0143] 4) Application of diagnostic and therapeutic nanoparticles containing organic fluorescent small molecules 8h in cancer diagnosis and treatment The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model. The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously. For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. After 6 h after injection, the fluorescence intensity reached the strongest and then gradually weakened until 12 h, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the diagnostic and therapeutic nanoparticles generate o-benzoquinone products with near-infrared II fluorescence during the reaction process. The diagnostic and therapeutic nanoparticles can be used for real-time monitoring of the chemical kinetic reaction process of mouse colon cancer.

[0144] Chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of therapeutic nanoparticles (40 mg / kg). The cure rate of tumor-bearing mice in the multi-dose treatment group (5 doses) was 80%, and the tumor did not recur within 60 days. The single-dose treatment group significantly prolonged the survival time of tumor-bearing mice, and all mice in the phosphate buffered saline solution group relapsed and died within 60 days.

[0145] Example 17: Organic fluorescent small molecule 8i, the chemical structure is: Prepared by the following synthetic method: (1) Synthesis of compound 2 NBS (3.88 g, 21.80 mmol) was added dropwise to a mixed solution of 4-(diphenylamino)benzaldehyde (5.00 g, 18.29 mmol) in chloroform / DMF (40 mL, v / v, 1:1) at room temperature. The mixture was heated at 60 °C under nitrogen. o C was stirred for 12 h, then quenched with water and extracted with dichloromethane. The organic phase was washed with water and saturated brine and dried over anhydrous sodium sulfate overnight. After removing dichloromethane, the residue was purified by silica gel chromatography, eluted with ethyl acetate / petroleum ether (1:30) to obtain the desired product compound 2 (5.92 g, yield 92%). 1 H NMR (400 MHz, DMSO- d6 , δ): 9.79 (s, 1H), 7.79 (t, J = 8.0 Hz, 2H), 7.57 (d, J= 8.5 Hz, 3H), 7.43 (t, J = 7.3 Hz, 1H), 7.24 (t, J =7.3 Hz, 1H), 7.19 (d, J = 8.1 Hz, 1H), 7.10 (t, J = 8.5 Hz, 3H), 7.00 (d, J = 8.0 Hz, 1H), 6.93 (t, J = 8.7 Hz, 1H). MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 19 H 15 BrNO + , 351.03; found: 351.07.

[0146] (2) Synthesis of compound 6 Compound 2 (2.00 g, 5.68 mmol) was dissolved in 5 mL of toluene with triethyl orthoformate (1.93 g, 6.65 mmol) and p-toluenesulfonic acid monohydrate (16 mg, 0.084 mmol). Then 2-3 drops of triethylamine were added and the reaction solution was stirred at room temperature for 30 min. After the reaction was completed, the excess solvent was removed by rotary evaporation and the obtained solid was vacuum dried at room temperature overnight. The above product was dissolved in 20 mL of THF. The temperature was lowered to -78 °C using dry ice. o C, n-butyl lithium (1.6 M in Hexanes) (7.1 mL, 11.36 mmol) was added dropwise to the THF solution. After the reaction mixture was stirred for 1 h under nitrogen protection, Bu3SnCl (4.62 g, 14.20 mol) was added. The mixture was stirred at room temperature for 12 h, and all volatile liquids were removed by rotary evaporation. The remaining viscous solid was separated by neutral alumina column chromatography and eluted with CH2Cl2. CH2Cl2 was removed under reduced pressure, and the resulting solid was dried in vacuo at room temperature overnight. The above product was transferred to a 50 mL two-necked flask and 10 mL of 1,4-dioxane was added. Then Pd(PPh3)4 (0.33 g, 0.284 mmol) and 4,7-dibromobenzo[1,2-c:4,5-c']bis([1,2,5]thiadiazole) (0.60 g, 1.70 mmol) were added. The reaction solution was stirred at 100 °C under nitrogen protection. o The reaction was heated in an oil bath at C for 12 h. After the reaction was completed, the solvent was removed by rotary evaporation and the mixture was purified by silica gel column chromatography using CH2Cl2 as eluent to obtain the desired product, compound 6 (0.35 g, 28%). 1H NMR (400 MHz, CDCl3, δ): 9.90 (s, 2H), 8.32 (d, J = 8.3 Hz, 4H), 7.79 (d, J = 8.3 Hz, 4H), 7.44 (d, J = 8.6 Hz, 8H), 7.34 (d, J = 8.0 Hz, 4H), 7.29 (s, 2H), 7.28 (s, 4H). 13 C NMR (151MHz, CDCl3, δ): 190.50, 152.81, 152.73, 146.81, 145.93, 133.13, 131 .33, 130.86, 129.96, 127.00, 125.67, 124.57, 124.47, 120.90, 120.31, 119.1. MALDI-TOF MS (ESI) m / z :[MH] - Calculate for: C 44 H 27 N6O2S2 - ,735.16;found:733.99.

[0147] (3) Synthesis of compound 8i The amino group in dopamine hydrochloride was connected to the aldehyde group at the end of compound 6 by an acid-catalyzed Schiff base reaction. Compound 6 (73.6 mg, 0.10 mmol) was dissolved in THF (10 mL), and 4-(4-aminobutyl)benzene-1,2-diol (54.3 mg, 0.30 mmol) was dissolved in anhydrous methanol (10 mL), and the two were mixed. 2-3 drops of glacial acetic acid were added to the mixture as a catalyst, and the reaction solution was heated at 74 o The reaction was carried out in a C oil bath for 8 h. After the reaction was completed, the solvent was removed by rotary evaporation. The desired product 8i (94.4 mg, 89%) was obtained by silica gel column chromatography using CHCl2 / methanol (5:1) as the eluent. MALDI-TOFMS (ESI) m / z :[M+H] + Calculate for: C 64 H 54 N8O4S2 + , 1062.37; found: 1062.74.

[0148] Example 18: Preparation and evaluation of diagnostic and therapeutic agent complexes and diagnostic and therapeutic agent nanoparticles containing organic fluorescent small molecule 8i 1) The diagnostic and therapeutic agent complex containing the organic fluorescent small molecule 8i was prepared according to the following method: Cu(OAc)2 (39.6 μM, 7.2 mg) and compound 8g (9.9 μM, 10.5 mg) were dissolved in 5 ml DMF. o After the reaction was completed, the precipitate was collected by centrifugation and washed three times with DMF to obtain the diagnostic and therapeutic agent complex.

[0149] The diagnostic and therapeutic agent complex can be uniformly dispersed in DMF by ultrasound, and the fluorescence is quenched due to the charge transfer within the molecule. Under acidic conditions, the complex reacts chemically with GSH and is degraded, emitting near-infrared II region fluorescence.

[0150] 2) The diagnostic and therapeutic nanoparticles containing the organic fluorescent small molecule 8i were prepared according to the following method: Pluronic F-127 (100 mg) was dissolved in 1 mL DMF and added to the above-mentioned diagnostic and therapeutic agent complex, and ultrasonicated to make it uniformly dispersed. Under magnetic stirring, the mixture was dropped into 10 mL ultrapure water, ultrasonicated for 10 min, and then stirred at room temperature for 8 h to form stable diagnostic and therapeutic agent nanoparticles.

[0151] The diagnostic and therapeutic agent nanoparticles have fluorescence quenching due to charge transfer within the molecule, but can be dissolved after reacting with GSH in an acidic environment (pH 5.5), and the fluorescence is restored. They can be excited at 660 nm and 830 nm, and emit near-infrared II region fluorescence with a wavelength of 1000-1300 nm.

[0152] 3) Chemical kinetics of diagnostic and therapeutic nanoparticles In vitro •OH and O 2-· It can be detected by electron spin resonance technology, so DMPO is used as •OH in water and O in methanol. 2-• The results showed that the aqueous solution of the therapeutic agent nanoparticles (100 μM) reacted with GSH at pH 5.5 to generate a large amount of •OH and O 2-· .

[0153] 4) Application of diagnostic and therapeutic nanoparticles in cancer diagnostic and therapeutic drugs The CT26-Luc cell line was used to establish a mouse subcutaneous tumor model and an orthotopic colon cancer model. The diagnostic and therapeutic nanoparticles (40 mg / kg) were injected intravenously. For the mouse subcutaneous tumor model, near-infrared II fluorescence imaging was performed directly on the tumor site using 660 nm and 830 nm lasers 0-24 h after injection. For the mouse orthotopic colon cancer model, the skin and peritoneum of the cecum were cut open to expose the intestinal segment, and near-infrared II fluorescence imaging was performed on the orthotopic tumor using 660 nm and 830 nm lasers 0-24 h after injection. The results showed that in both models, near-infrared II fluorescence began to appear in the tumor site 4 h after injection. After 6 h after injection, the fluorescence intensity reached the highest level and then gradually weakened until 12 h, when the fluorescence disappeared. This proves that the diagnostic and therapeutic nanoparticles can mediate specific chemical kinetic reactions in the mouse colon cancer model, and the o-benzoquinone products with near-infrared II fluorescence generated by the diagnostic and therapeutic nanoparticles during the reaction can be used for real-time monitoring of the chemical kinetic reaction process of the diagnostic and therapeutic nanoparticles in mouse colon cancer.

[0154] The chemodynamic therapy was performed on mice bearing CT26-Luc orthotopic tumors by intravenous injection of diagnostic and therapeutic nanoparticles (40 mg / kg). The tumor did not recur in the mice bearing tumors in the multi-dose treatment group (5 doses) within 60 days after treatment, and the chemodynamic therapy was effective in mice bearing CT26-Luc orthotopic tumors; the single-dose treatment group could significantly prolong the survival time of tumor-bearing mice; all mice in the phosphate buffered saline solution group relapsed and died within 60 days.

Claims

1. A fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapeutic properties, characterized in that: The complex includes an organic fluorescent small molecule and a copper ion chelate, and the combination molar ratio of the organic fluorescent small molecule and the copper ion is 1:0.1-1:1.64, wherein the organic fluorescent small molecule can emit near-infrared II region fluorescence (1000-1700 nm), and its structure is shown in general formula I: Among them, R is selected from , or One of them.

2. The fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapy performance according to claim 1, characterized in that: The complex is a nanoparticle formed by an organic fluorescent small molecule, copper ions and a surfactant, wherein the molar ratio of the organic fluorescent small molecule, the copper ion and the surfactant is 1: (0.1-1.64): (0.16-1.6), and the average particle size of the nanoparticle is 140-300 nm.

3. The fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapy performance according to claim 1 or 2, characterized in that: The complex formed by the chelation of the organic fluorescent small molecule and the copper ion has the following structural formula II: Due to the intramolecular charge transfer between copper ions and fluorescent small molecules, fluorescence quenching occurs. In a weakly acidic environment, a chemical kinetic reaction occurs with glutathione to produce reactive oxygen species and generate o-benzoquinone products, which emit near-infrared II region fluorescence, which is used for real-time dynamic imaging and monitoring of the chemical kinetic reaction process mediated by the diagnostic and therapeutic agent complex.

4. The fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapy performance according to claim 2, characterized in that: The molar ratio of the organic fluorescent small molecule to the copper ion is 1:1.

64.

5. The fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapy performance according to claim 2, characterized in that: The surfactant is one of Pluronic F-127, Pluronic F-68, distearoyl phosphatidylethanolamine-methoxy polyethylene glycol 2000, or distearoyl phosphatidylethanolamine-methoxy polyethylene glycol 5000.

6. The fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapy performance according to claim 2, characterized in that: The method for preparing nanoparticles comprises the steps of: (1) dissolving the organic fluorescent small molecule and Cu(OAc)2 in DMF, wherein the molar ratio of the organic fluorescent small molecule to Cu(OAc)2 is 1:1.64, to obtain an organic solution; (2) heating the organic solution in step (1) at 60° C. for 2 h to generate a chelate solution; (3) centrifuging the chelate solution in step (2) and collecting the precipitate, and washing it with DMF three times to obtain the desired complex; (4) The complex and surfactant in step (3) are dissolved in DMF, and the molar ratio of the complex and the organic fluorescent small molecule in step (1) is 0.8:1, and the surfactant is Pluronic F-127 or Pluronic F-68 or DSPE-PEG 2000 or DSPE-PEG 5000 ; (5) Under ultrasonic and ice-water bath conditions, the organic solution of step (4) was added to pure water and mixed evenly, with the ratio of organic solvent to pure water being 1:10; (6) Stir the mixture obtained in step (5) at room temperature for 4-12 h; (7) Concentrating the nanoparticle solution obtained in step (6) to obtain the desired nanoparticles.

7. An organic fluorescent small molecule according to any one of claims 1 to 6, characterized in that: Using aldehyde-substituted triphenylamine as raw material, the target product is obtained through bromination, Suzuki reaction, and Schiff base reaction; the synthetic route is as follows: Among them, R1 is , One of the following; R2 is hydrogen or methoxy. The reagents used in the above steps are: (a) N-bromosuccinimide, N,N-dimethylformamide, chloroform, 60°C; (b) p-Toluenesulfonic acid monohydrate, triethylamine, toluene, room temperature; (c) n-butyllithium, -78°C; (d) Tetrakistriphenylphosphine palladium, dioxane, 100 °C; (e) Glacial acetic acid, methanol, tetrahydrofuran, 74°C.

8. The organic fluorescent small molecule according to claim 7, characterized in that: The chemical structural formula of the organic fluorescent small molecule is: ， or, ， or, ， or, ， or, ， or, ， or, ， or, ， or, 。 9. Use of the fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapy properties according to any one of claims 1 to 6 in chemodynamic therapy.

10. The use according to claim 8, characterized in that: The complex undergoes a chemical kinetic reaction with glutathione in a weakly acidic environment to produce reactive oxygen species and generate o-benzoquinone products, which can emit near-infrared II region fluorescence under 660 nm and 830 nm laser irradiation, and can be used for real-time dynamic imaging and monitoring of the chemical kinetic reaction process mediated by the diagnostic and therapeutic agent complex.

11. Use of the fluorescent diagnostic and therapeutic agent complex having near-infrared II region fluorescence imaging and chemodynamic therapeutic properties according to any one of claims 1 to 6 in cancer treatment drugs.

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