Near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as well as preparation method and application of near-infrared fluorescent photosensitizer

Abstract

The invention relates to the technical field of biomedical materials, and discloses a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as well as a preparation method and application of the near-infrared fluorescent photosensitizer. The near-infrared photosensitizer provided by the invention is synthesized through simple coupling reaction and nucleophilic reaction, has near-infrared fluorescence in a low-polarity solvent, and is high in fluorescence quantum yield. In addition, the compound can specifically target lipid droplets, and still has high light stability under irradiation of strong laser. Importantly, active oxygen species can be efficiently generated under white light irradiation, and inhibition and killing of tumor cells and living mouse tumors are achieved. The invention provides a new way for regulating and controlling a molecular excited state to construct a near-infrared photosensitizer generated by efficient active oxygen.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedical materials, and in particular to a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile, a preparation method and an application thereof, and specifically to a donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile, a preparation method and an application thereof in lipid droplet targeted imaging-mediated photodynamic therapy. Background Art

[0002] Lipid droplets are a type of cell organelle that mainly contains triglycerides and cholesterol esters and is wrapped by a phospholipid monolayer. Many functional proteins, such as structural proteins, membrane transport proteins, and enzymes, are embedded on their surface. As a highly dynamic neutral lipid reservoir, lipid droplets can store excess lipids to prevent lipotoxicity and gradually release lipids according to cellular needs to maintain the energy and redox balance of cells and tissues. These surface proteins are closely related to the generation and degradation of lipid droplets. They can regulate the interaction between lipid droplets and other organelles (such as the endoplasmic reticulum, mitochondria, lysosomes, peroxisomes, and nuclei) to maintain membrane balance and organelle balance. Metabolic disorders and dysfunctions of lipid droplets may cause many metabolic diseases, such as fatty liver disease, diabetes, atherosclerosis, and even cancer. Therefore, lipid droplets are expected to be used as an important target for the treatment of diseases and tumors.

[0003] Photodynamic therapy is a treatment method that uses the interaction of photosensitizers with light of specific wavelengths and oxygen. Photodynamic therapy exhibits high spatial and temporal selectivity, non-invasiveness, low side effects, and negligible drug resistance. These characteristics have achieved breakthroughs in the treatment of superficial tumors and many non-tumor diseases, such as bright erythema, restenosis after angioplasty, and rheumatoid arthritis. However, the lipid droplet fluorescent probes reported so far rarely have the characteristics of photodynamic therapy, mainly because the lipid droplet probes are not targeted enough and the ability of photodynamic therapy to produce reactive oxygen species is not high. Summary of the invention

[0004] Purpose of the invention: In view of the problems existing in the prior art, the present invention provides a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile, a preparation method and application. The near-infrared fluorescent photosensitizer has a donor-acceptor-acceptor structure, high near-infrared fluorescence, fluorescence quantum yield, good photostability, strong lipid droplet targeting, strong ability to generate reactive oxygen species by photodynamic force, and a simple synthesis method.

[0005] Technical solution: In a first aspect, the present invention provides a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile, wherein the near-infrared photosensitizer has a donor-acceptor-acceptor type structure based on benzoselenadiazole and acrylonitrile, and the chemical structural formula is shown in Formula I, ; Wherein, R is independently selected from one of hydrogen, fluorine, chlorine, bromine, iodine, cyano, alkyl, optionally substituted alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, aryl, heterocyclyl, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino; X, Y are independently selected from any one of hydrogen, methyl, methoxy, fluorine, chlorine, bromine, iodine, cyano; Ar1 ​​and Ar2 are each independently a substituted or unsubstituted aromatic ring selected from phenyl, benzene, pyridine, thiophene, furan, pyran, imidazole, quinoline and thiadiazole, and the aromatic ring substituent is selected from hydrogen, alkyl, halogen, hydroxyl, amino, carboxyl, carboxylate, trifluoromethyl and cyano.

[0006] In a second aspect, the present invention provides a method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as described above, and the synthesis route thereof is as follows: .

[0007] Furthermore, the specific steps are as follows: S1, dissolving compound 1 and compound 2 in a mixed solution of tetrahydrofuran and water, adding tetrakis(triphenylphosphine)palladium catalyst to heat for reaction, cooling to room temperature, separating the liquids, removing the organic solvent by vacuum distillation, and separating and purifying by silica gel column to obtain compound 3; S2, compound 3 and compound 4 were dissolved in anhydrous ethanol, potassium tert-butoxide was added and heated for reaction, cooled to room temperature, and separated and purified by silica gel column to obtain a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile.

[0008] Furthermore, in S1, the molar ratio of the compound 1 to the compound 2 is 0.8-1.5:1.

[0009] Furthermore, in S1, the molar ratio of the compound 1 to the tetrakis(triphenylphosphine)palladium is 1:0.05-0.1.

[0010] Furthermore, in S1, the volume ratio of tetrahydrofuran to water is 4-10:1.

[0011] Preferably, in S1, the specific conditions of the heating reaction are: reaction temperature is 60-90°C, and reaction time is 4-12 h.

[0012] Furthermore, in S2, the molar ratio of the compound 3 to the compound 4 is 1:1.2-2.

[0013] Furthermore, in S2, the specific conditions of the heating reaction are: reaction temperature is 40-80°C, and reaction time is 2-10 h.

[0014] In a third aspect, the present invention provides a use of the above-described near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile in lipid droplet targeted imaging-mediated photodynamic therapy.

[0015] In a fourth aspect, the present invention provides an application of a product prepared by the method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as described above in lipid droplet targeted imaging-mediated photodynamic therapy.

[0016] Beneficial effects: Compared with the prior art, the present invention has the following specific beneficial effects: (1) The donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile provided by the present invention is synthesized through a simple coupling reaction and a nucleophilic reaction. It has near-infrared fluorescence and a high fluorescence quantum yield in a low-polarity solvent. In addition, it can specifically target subcellular organelle lipid droplets and still have high photostability under the irradiation of strong lasers.

[0017] (2) Under the irradiation of white light from a xenon lamp, the donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile provided by the present invention can efficiently generate a large amount of reactive oxygen species, thereby inhibiting and killing tumor cells and living tumors.

[0018] (3) The present invention provides a new approach to construct near-infrared photosensitizers for efficient reactive oxygen production by regulating the excited state of molecules. BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 This is the H NMR spectrum of compound 3 in DMSO-d6 in embodiment 2 of the present invention; Figure 2 is a high-resolution mass spectrum of compound 3 in embodiment 2 of the present invention; Figure 3 This is the H NMR spectrum of TSeBT-ANPy in DMSO-d6 in Embodiment 2 of the present invention; Figure 4 is a high-resolution mass spectrum of TSeBT-ANPy in Embodiment 2 of the present invention; Figure 5 is the NMR carbon spectrum of TSeBT-ANQu in DMSO-d6 in embodiment 3 of the present invention; Figure 6 is the high-resolution mass spectrum of TSeBT-ANQu in Embodiment 3 of the present invention; Figure 7 The synthesis route and single crystal structure of TSeBT-ANPy in Embodiment 2 and TSeBT-ANQu in Embodiment 3 of the present invention are as follows: (A) Synthesis route of TSeBT-ANPy and TSeBT-ANQu; (B) Single crystal structure of TSeBT-ANPy and TSeBT-ANQu; Figure 8 Photophysical property diagrams of TSeBT-ANPy prepared in Embodiment 2 and TSeBT-ANQu prepared in Embodiment 3 of the present invention: (A) Absorption spectra of TSeBT-ANPy and TSeBT-ANQu in toluene solution; (B) Fluorescence spectra of TSeBT-ANPy and (C) TSeBT-ANQu in different organic solvents; (D) Fluorescence quantum yields of TSeBT-ANPy and TSeBT-ANQu in different organic solvents; Concentration: 10 μM.

[0020] Fig. 9 Figures showing the photodynamic performance of TSeBT-ANPy prepared in Embodiment 2 and TSeBT-ANQu prepared in Embodiment 3 of the present invention in generating active oxygen: (A) Fluorescence spectrum of active oxygen indicator DCFH in PBS solution containing TSeBT-ANPy changing with time under white light irradiation; (B) Fluorescence spectrum of active oxygen indicator DCFH in PBS solution containing TSeBT-ANQu changing with time under white light irradiation; (C) Spectrum of relative fluorescence intensity of active oxygen indicator DCFH at 525 nm changing with time in PBS solution containing different photosensitizers under white light irradiation; (D) Absorption spectrum of singlet oxygen indicator ABDA in PBS solution containing TSeBT-ANPy changing with time under white light irradiation; (E) Absorption spectrum of singlet oxygen indicator ABDA in PBS solution containing TSeBT-ANQu changing with time under white light irradiation; (F) Absorption spectrum of singlet oxygen indicator ABDA in PBS solution containing different photosensitizers at 378 nm under white light irradiation. Spectrum of relative absorbance at nm changing with time; photosensitizer concentration: 10 μM; indicator concentration: 50 μM.

[0021] Fig.10Figures showing lipid droplet targeting and photodynamic therapy of TSeBT-ANPy prepared in Embodiment 2 and TSeBT-ANQu prepared in Embodiment 3 of the present invention: (A) Confocal imaging of TSeBT-ANPy and TSeBT-ANQu co-stained with commercial lipid droplet green light probe Bodipy 493 / 503 in HeLa cells pretreated with 50 μM oleic acid; (B) Cytotoxicity test results of TSeBT-ANPy at different concentrations in HeLa cells with / without white light irradiation for 3 min; (C) Cytotoxicity test results of TSeBT-ANQu at different concentrations in HeLa cells with / without white light irradiation for 3 min. min; (D) Confocal imaging of TSeBT-ANPy in HeLa cells under white light irradiation for different time periods, the reactive oxygen species indicator DCFH; (E) Confocal imaging of TSeBT-ANPy in HeLa cells over time under continuous laser irradiation; (F) Normalized fluorescence intensity of TSeBT-ANPy in HeLa cells over time under continuous laser irradiation; Photosensitizer concentration: 2 μM (co-staining and photostability test) or 10 μM (photodynamically generated reactive oxygen species imaging); Indicator concentration: 50 μM; Scale bar: 10 μm.

[0022] Fig.11 Figures showing the research of photodynamic therapy of TSeBT-ANPy prepared in Embodiment 2 of the present invention in live mouse tumors: (A) In vivo imaging of the fluorescence intensity of TSeBT-ANPy injected intratumorally in mouse tumors over time; (B) A graph showing the changes in the size of mouse tumors (indicated by white circles) over time after photodynamic therapy after intratumoral injection of PBS and photosensitizer dye TSeBT-ANPy, respectively; (C) A graph showing the changes in the size of mouse tumors over time after photodynamic therapy after intratumoral injection of PBS and TSeBT-ANPy, respectively; (D) A graph showing the changes in the weight of mice over time after photodynamic therapy after intratumoral injection of PBS and TSeBT-ANPy, respectively; (E) Hematoxylin-eosin (HE) staining of different heart, liver, spleen, lung, and kidney tissue sections of mice after photodynamic therapy after intratumoral injection of PBS and TSeBT-ANPy, respectively; photosensitizer concentration: 10 μM. DETAILED DESCRIPTION

[0023] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below in combination with specific implementation methods and comparative examples.

[0024] Implementation 1: This embodiment provides a donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile, and the chemical structure is shown in Formula I. ; Wherein, R is independently selected from one of hydrogen, fluorine, chlorine, bromine, iodine, cyano, alkyl, optionally substituted alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, aryl, heterocyclyl, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino; X, Y are independently selected from any one of hydrogen, methyl, methoxy, fluorine, chlorine, bromine, iodine, cyano; Ar1 ​​and Ar2 are each independently a substituted or unsubstituted aromatic ring selected from phenyl, benzene, pyridine, thiophene, furan, pyran, imidazole, quinoline and thiadiazole, and the aromatic ring substituent is selected from hydrogen, alkyl, halogen, hydroxyl, amino, carboxyl, carboxylate, trifluoromethyl and cyano.

[0025] This embodiment also provides a method for preparing the donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as described above, and the synthesis route thereof is as follows: .

[0026] Implementation 2: According to Embodiment 1, this embodiment provides a donor-acceptor-acceptor type near-infrared fluorescent photosensitizer TSeBT-ANPy based on benzoselenadiazole and acrylonitrile and a preparation method thereof, as follows: In this embodiment, the substituents R, X, and Y are all hydrogen, Ar1 is a benzene ring, Ar1 is pyridine, and the chemical structure of the corresponding donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile (named TSeBT-ANPy) is as follows: .

[0027] The synthetic route of TSeBT-ANPy is as follows Figure 7 As shown in A: First, compound 1 and compound 2 undergo Suzuki coupling under palladium catalysis to obtain intermediate compound 3; then compound 3 and compound 4 (pyridine acetonitrile) undergo nucleophilic reaction to obtain the final photosensitizer compound.

[0028] The specific preparation method of TSeBT-ANPy is as follows: (1) Synthesis of compound 3: Compound 1 (371 mg, 1 mmol), compound 2 (290 mg, 1 mmol) and Pb(PPh3)4 (70 mg) were dissolved in a mixed solution of tetrahydrofuran (10 mL) and water (1 mL), heated to 80°C under nitrogen protection, reacted for 10 h, cooled to room temperature, separated, and the organic phase solvent was distilled off under reduced pressure. Compound 3 was obtained by column separation and purification using an eluent of dichloromethane and n-hexane in a ratio of 1:1. 1 H NMR spectroscopy and high-resolution mass spectrometry such as Figure 1-2 The detailed characterization data are as follows:1 H NMR (400 MHz, DMSO- d 6), δ (ppm): 10.71 (s, 1H), 8.18 (d, J =7.3 Hz, 1H), 7.97‒7.91 (m, 2H), 7.84 (d, J = 7.2 Hz, 1H), 7.37 (t, J = 7.9Hz, 4H), 7.16‒7.10 (m, 6H), 7.09‒7.05 (m, 2H). HRMS (ESI) m / z Calculate for [C 25 H 18 N3O + ] 456.0610 ([M+H] + ), found 456.0654.

[0029] (2) Synthesis of photosensitizer TSeBT-ANPy: Pyridineacetonitrile (142 mg, 1.2 mmol) and potassium tert-butoxide (134 mg, 1.2 mmol) were added to 10 mL of anhydrous ethanol and stirred for 20 min. Compound 3 (455 mg, 1 mmol) was slowly added and reacted at 60°C for 10 h. After the reaction was confirmed to be complete by thin layer chromatography, heating was stopped. After the reaction was cooled to room temperature, the organic solvent was removed by vacuum distillation. The near-infrared photosensitizer TSeBT-ANPy was obtained by column separation and purification using an eluent of dichloromethane: methanol in a ratio of 15:1. The structure was verified by X-ray single crystal diffraction ( Figure 7 B). TSeBT-ANPy 1 H NMR spectroscopy and high-resolution mass spectrometry such as Figure 3-4 The detailed characterization data are as follows: 1 H NMR (400 MHz, DMSO- d 6), δ (ppm): 8.86 (s, 1H), 8.75 (d, J = 6.0 Hz, 2H), 8.46 (d, J = 7.5 Hz, 1H), 7.97(d, J = 8.6 Hz, 2H), 7.89 (d, J = 7.6 Hz, 1H), 7.79 (d, J = 5.8 Hz, 2H), 7.38(t, J= 7.8 Hz, 4H), 7.18–7.05 (m, 8H). HRMS (ESI) m / z calcd for [C 32 H 22 N5 + ]556.1035 ([M+H] + ), found 556.1070.

[0030] Implementation 3: According to Embodiment 1, this embodiment provides a donor-acceptor-acceptor type near-infrared fluorescent photosensitizer TSeBT-ANQu based on benzoselenadiazole and acrylonitrile and a preparation method thereof, as follows: In this embodiment, the substituents R, X, and Y are all hydrogen, Ar1 is a benzene ring, Ar2 is quinoline, and the chemical structure of the corresponding donor-acceptor-acceptor type near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile (named TSeBT-ANQu) is as follows: .

[0031] The synthetic route of TSeBT-ANQu is as follows Figure 7 As shown in A: First, compound 1 and compound 2 undergo Suzuki coupling under palladium catalysis to obtain intermediate compound 3; then compound 3 and compound 4 (quinoline acetonitrile) undergo nucleophilic reaction to obtain the final photosensitizer compound.

[0032] The specific preparation method of TSeBT-ANQu is as follows: (1) The synthesis of compound 3 is shown in Implementation Example 2.

[0033] (2) Synthesis of photosensitizer TSeBT-ANQu: Quinoline acetonitrile (202 mg, 1.2 mmol) and potassium tert-butoxide (134 mg, 1.2 mmol) were added to 10 mL of anhydrous ethanol and stirred for 20 min. Compound 3 (455 mg, 1 mmol) was slowly added and reacted at 60°C for 10 h. After the reaction was confirmed to be complete by thin layer chromatography, heating was stopped. After the reaction was cooled to room temperature, the organic solvent was removed by vacuum distillation. The near-infrared photosensitizer TSeBT-ANQu was obtained by column separation and purification using an eluent of dichloromethane: methanol (15:1). The structure was verified by X-ray single crystal diffraction ( Figure 7 B). TSeBT-ANQu 1 H NMR spectroscopy and high-resolution mass spectrometry such as Figure 3-4 The detailed characterization data are as follows: 1 H NMR (400 MHz, DMSO-d 6), δ (ppm): 9.06 (d, J = 4.5 Hz, 1H), 8.63 (d, J = 7.4 Hz, 1H), 8.47 (s, 1H), 8.36(d, J = 8.2 Hz, 1H), 8.19 (d, J = 8.4 Hz, 1H), 8.04–7.87 (m, 4H), 7.82–7.75(m, 2H), 7.38 (t, J = 7.8 Hz, 4H), 7.25–7.02 (m, 8H). HRMS (ESI) m / z Calculate for [C 36 H 24 N5 + ] 606.1191 ([M+H] + ), found 606.1216.

[0034] Implementation 4: Study on the photophysical properties of near-infrared fluorescent photosensitizers TSeBT-ANPy and TSeBT-ANQu: The TSeBT-ANPy prepared in Embodiment 2 and the TSeBT-ANQu prepared in Embodiment 3 were added to organic solvents of different polarities, respectively, and fluorescence spectra were tested. Figure 8 As shown in Figure A, compared with TSeBT-ANQu, the emission wavelength of TSeBT-ANPy is red-shifted by about 50 nm. TSeBT-ANQu and TSeBT-ANPy have near-infrared emission in low-polarity toluene solution, which are 650 nm and 668 nm, respectively. Figure 8 B and Figure 8 C, and they have a twisted intramolecular charge transfer (TICT) effect, that is, they emit weak fluorescence in polar solvents, while in a low-polarity environment, the TICT effect is suppressed and the fluorescence effect is significantly enhanced.

[0035] The absolute quantum yields of TSeBT-ANPy prepared in Implementation Example 2 and TSeBT-ANQu prepared in Implementation Example 3 in different polar solvents were tested using the integrating sphere method. Figure 8 As shown in D, with the increase of solution polarity, the quantum yield of TSeBT-ANPy prepared in Embodiment 2 and TSeBT-ANQu prepared in Embodiment 3 gradually decreases, and the quantum yield data is consistent with Figure 8 B and Figure 8The fluorescence spectrum of C is consistent with that of daptomycin C, and the fluorescence quantum yield in non-polar solution is as high as 57.2%, which is beneficial for high-contrast imaging and treatment in non-polar environments of cells such as lipid droplets.

[0036] Implementation 5: Study on the photodynamic generation of reactive oxygen species by near-infrared photosensitizers TSeBT-ANPy and TSeBT-ANQu The ROS generation efficiency of photosensitizers TSeBT-ANPy and TSeBT-ANQu was tested using a commercial ROS indicator (2',7'-dichlorodihydrofluorescein, DCFH) under white light irradiation. Fig. 9 As shown in A, as the white light irradiation time increases, the fluorescence intensity of DCFH at 525 nm in the PBS solution containing TSeBT-ANPy gradually increases. This indicates that TSeBT-ANPy has the ability to generate reactive oxygen species under white light irradiation. Similarly, TSeBT-ANQu also has the ability to generate reactive oxygen species under white light irradiation ( Fig. 9 B). The relative fluorescence intensity of DCFH at 525 nm shows that under white light irradiation, the photosensitizer TSeBT-ANPy can produce more reactive oxygen species than TSeBT-ANQu ( Fig. 9 C).

[0037] The commercial singlet oxygen indicator ABDA was used to test the production of singlet oxygen by TSeBT-ANPy and TSeBT-ANQu under white light irradiation. Fig. 9 As shown in D, with the increase of white light irradiation time, the absorbance of the singlet oxygen indicator ABDA in the PBS solution containing TSeBT-ANPy gradually weakened, indicating that TSeBT-ANPy has the ability to generate singlet oxygen under white light irradiation. Similarly, TSeBT-ANQu also has the ability to generate singlet oxygen under white light irradiation ( Fig. 9 E). The relative absorbance of ABDA at 378 nm shows that under white light irradiation, the photosensitizer TSeBT-ANPy can produce more singlet oxygen than TSeBT-ANQu ( Fig. 9 F). In summary, TSeBT-ANPy has better photodynamic ROS generation ability than TSeBT-ANQu, which is beneficial for potential applications in cell and in vivo therapy.

[0038] Implementation 6: Study on lipid droplet targeting and photodynamic therapy of near-infrared photosensitizers TSeBT-ANPy and TSeBT-ANQu In HeLa cells, TSeBT-ANPy and TSeBT-ANQu were counterstained with the commercial lipid droplet probe Bodipy493 / 503. HeLa cells were inoculated into a confocal dish and incubated at 37 degrees and 5% CO2 for 12 hours. HeLa cells were pretreated with 50 μM oleic acid, and then the old culture medium in the confocal dish was aspirated, and 2 μM TSeBT-ANPy and TSeBT-ANQu and commercial probe Bodipy 493 / 503 culture medium solutions were added respectively. After continuing to culture for 30 min, the old culture medium of the confocal dish was aspirated and rinsed twice with PBS. Colocalization experiments confirmed that TSeBT-ANPy and TSeBT-ANQu had high colocalization coefficients with Bodipy 493 / 503, which were 0.87 and 0.82, respectively, thereby confirming that the near-infrared photosensitizer provided by the present invention specifically stains lipid droplets in a low-polarity environment.

[0039] In HeLa cells, cytotoxicity experiments were conducted with different concentrations of TSeBT-ANPy and TSeBT-ANQu with / without white light irradiation. HeLa cells were seeded into 96-well plates and incubated at 37 degrees and 5% CO2 for 12 hours. Then, the old culture medium in the 96-well plates was aspirated, and culture medium with different concentrations of TSeBT-ANPy and TSeBT-ANQu was added. Irradiate with white light for 3 minutes and keep away from light for 3 minutes. Then, the old culture medium in the 96-well plates was aspirated, and culture medium containing 20 μL of MTT was added. The cells were placed at 37 degrees and 5% CO2 for 4 hours. Then, the old culture medium was aspirated, and 100 μL of DMSO was added. The cell viability was tested using an enzyme plate reader. Fig.10 As shown in Figures B and 10C, in the absence of white light irradiation, TSeBT-ANPy and TSeBT-ANQu at different concentrations had almost no effect on the survival rate of HeLa cells. After white light irradiation, as the concentrations of TSeBT-ANPy and TSeBT-ANQu increased, the survival rate of HeLa cells became lower and lower. When the concentrations of TSeBT-ANPy and TSeBT-ANQu were 10 μM, the survival rates of HeLa cells were only 12.2% and 38.2%, indicating that TSeBT-ANPy has a more efficient reactive oxygen species generation ability and photodynamic therapy effect than TSeBT-ANQu.

[0040] The commercial ROS indicator DCFH was used to test the generation of reactive oxygen species by TSeBT-ANPy in HeLa cells under white light irradiation. Fig.10As shown in Figure D, under different durations of white light irradiation, the fluorescence of DCFH, an active oxygen indicator of TSeBT-ANPy in HeLa cells gradually increased. This indicates that TSeBT-ANPy can efficiently produce active oxygen under white light irradiation, thereby achieving the effect of photodynamic therapy. Furthermore, through continuous laser irradiation, it was found that TSeBT-ANPy has excellent photostability. After 6 minutes of irradiation, the fluorescence intensity basically maintained 80% of the initial intensity ( Fig.10 E and Fig.10 F), which is also beneficial for photodynamic therapy under white light irradiation.

[0041] Implementation 7: Study on photodynamic therapy of tumors in living mice using near-infrared photosensitizer TSeBT-ANPy The solution and imaging data confirmed that TSeBT-ANPy can be used as an excellent photosensitizer for photodynamic therapy. This embodiment further verifies its in vivo photodynamic therapy effect on mouse tumors. First, in vivo mouse imaging studies were conducted. Fig.11 As shown in A, TSeBT-ANPy was injected intratumorally into the tumor site of tumor-bearing mice. After 5 minutes, the strongest fluorescence signal appeared in the tumor site, indicating that TSeBT-ANPy successfully differentiated and imaged the tumor tissue. After 2 hours of injection, a strong fluorescence signal was still visible in the tumor site, indicating that TSeBT-ANPy successfully accumulated and retained in the tumor site. After 24 hours of injection, the fluorescence signal basically disappeared, indicating that TSeBT-ANPy can be metabolized and excreted by mice.

[0042] Then, TSeBT-ANPy was verified in the photodynamic therapy of tumors in living mice. Fig.11 B and Fig.11 As shown in C, after intratumoral injection of PBS, the size of mouse tumors gradually increased over time. However, after intratumoral injection of TSeBT-ANPy and photodynamic therapy, the size of mouse tumors only changed slightly over time, indicating that under white light irradiation, TSeBT-ANPy can effectively inhibit tumor growth and has a good photodynamic therapy effect. Fig.11 As shown in D, after intratumoral injection of PBS and TSeBT-ANPy, there was no significant weight loss in mice after 18 days of photodynamic therapy. Fig.11 As shown in E, after intratumoral injection of PBS and TSeBT-ANPy, respectively, no pathological changes or abnormalities were observed in different heart, liver, spleen, lung, and kidney tissues of mice after photodynamic therapy, indicating that TSeBT-ANPy has good biocompatibility and biosafety.

[0043] The above embodiments are only for illustrating the technical concept and features of the present invention, and their purpose is to enable people familiar with the technology to understand the content of the present invention and implement it accordingly, and they cannot be used to limit the protection scope of the present invention. Any equivalent transformation or modification made according to the spirit of the present invention should be included in the protection scope of the present invention.

Claims

1. A near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile, characterized in that: The near-infrared photosensitizer has a donor-acceptor-acceptor structure based on benzoselenadiazole and acrylonitrile, and the chemical structural formula is shown in Formula I. ； Wherein, R is independently selected from one of hydrogen, fluorine, chlorine, bromine, iodine, cyano, alkyl, optionally substituted alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, aryl, heterocyclyl, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino; X, Y are independently selected from any one of hydrogen, methyl, methoxy, fluorine, chlorine, bromine, iodine, cyano; Ar1 ​​and Ar2 are each independently a substituted or unsubstituted aromatic ring selected from phenyl, benzene, pyridine, thiophene, furan, pyran, imidazole, quinoline and thiadiazole, and the aromatic ring substituent is selected from hydrogen, alkyl, halogen, hydroxyl, amino, carboxyl, carboxylate, trifluoromethyl and cyano.

2. A method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as claimed in claim 1, characterized in that: The synthetic route is as follows: 。 3. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 2, characterized in that: The specific steps are as follows: S1, dissolving compound 1 and compound 2 in a mixed solution of tetrahydrofuran and water, adding tetrakis(triphenylphosphine)palladium catalyst to heat for reaction, cooling to room temperature, separating the liquids, removing the organic solvent by vacuum distillation, and separating and purifying by silica gel column to obtain compound 3; S2, compound 3 and compound 4 were dissolved in anhydrous ethanol, potassium tert-butoxide was added and heated for reaction, cooled to room temperature, and separated and purified by silica gel column to obtain a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile.

4. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 3, characterized in that: In S1, the molar ratio of the compound 1 to the compound 2 is 0.8-1.5:

1.

5. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 3, characterized in that: In S1, the molar ratio of the compound 1 to the tetrakis(triphenylphosphine)palladium is 1:0.05-0.

1.

6. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 3, characterized in that: In S1, the volume ratio of tetrahydrofuran to water is 4-10:

1.

7. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 3, characterized in that: In S1, the specific conditions of the heating reaction are: reaction temperature is 60-90°C, and reaction time is 4-12 h.

8. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 3, characterized in that: In S2, the molar ratio of the compound 3 to the compound 4 is 1:1.2-2.

9. The method for preparing a near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile according to claim 3, characterized in that: In S2, the specific conditions of the heating reaction are: reaction temperature is 40-80°C, and reaction time is 2-10 h.

10. Use of a product prepared by the near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as claimed in claim 1 and / or the preparation method of the near-infrared fluorescent photosensitizer based on benzoselenadiazole and acrylonitrile as claimed in claims 2-9 in lipid droplet targeted imaging-mediated photodynamic therapy.

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