Sulfonium aggregation-induced emission luminophore, preparation and application thereof

Abstract

The application discloses a sulfonium aggregation-induced emission luminophore, and preparation and application thereof. ST The sulfonium fragment is introduced into the side chain of an AIE luminophan to increase spin-orbit coupling SOC and reduce singlet-triplet energy gap ΔE, so as to promote intersystem crossing ISC and enhance the ability of the luminophore to generate reactive oxygen species ROS. In addition, the sulfonium-containing luminophore can realize rapid uptake of tumor cells, and can be anchored to the tumor cell membrane after 10 minutes of incubation, and can be enriched in intracellular lipid droplets with the extension of the incubation time, so that the anti-tumor activity is improved. Under light irradiation, the sulfonium luminophore exhibits good photodynamic anti-tumor activity in vitro and in vivo, can effectively inhibit tumor cell proliferation and tumor growth in tumor-bearing mice, and has good application prospect. The sulfonium AIE luminophore has the following general formula.

Description

Technical Field

[0001] The present invention belongs to the field of tumor photodynamic therapy, and specifically relates to a sulfonium aggregation-induced emission (AIE) photosensitizer and its preparation and application. It is a small molecule AIE photosensitizer containing a sulfonium fragment and its preparation method and application. Background Art

[0002] Photodynamic therapy (PDT) is a novel tumor treatment. Under light, a photosensitizer undergoes a transition from a singlet excited state to a triplet excited state via intersystem crossing (ISC). The photosensitizer then reacts with oxygen and other molecules to generate reactive oxygen species (ROS). ROS can directly react with biomolecules such as lipids, proteins, and nucleic acids, inhibiting tumor cell growth. PDT offers advantages such as minimal invasiveness, high spatiotemporal selectivity, and ease of use.

[0003] Efficient ISC is the key to the anti-tumor activity of photosensitizers. Increasing the spin-orbit coupling (SOC) of photosensitizers or reducing the singlet excited state-triplet excited state (S n -T m ) between the energy difference (ΔE ST ) can effectively promote intersystem crossing of photosensitizers. Introducing traditional heavy atoms (including transition metals, I, and Br) into photosensitizer molecules or fluorophores can increase SOC and improve photosensitivity, but this modification carries the potential for dark cytotoxicity, and the heavy atom effect may also weaken as the chromophore size increases.

[0004] Because sulfur atoms have stronger spin-orbit coupling than oxygen atoms, studies have shown that modifying sulfur atoms into the structure of photosensitizers may improve the intersystem crossing efficiency of photosensitizers. However, this modification strategy currently requires the presence of an oxycarbonyl structure in the photosensitizer molecule, which limits molecular design. Therefore, developing a new sulfur atom modification strategy that is independent of the oxycarbonyl structure and further improves the photosensitizing activity of photosensitizers is of great research significance. AIE-type photosensitizers can effectively overcome the fluorescence quenching phenomenon caused by π-π stacking of traditional porphyrin-based photosensitizers, and therefore show great application potential in the field of PDT. Summary of the Invention

[0005] The present invention aims to provide a sulfonium aggregation-induced emission (AIE) photosensitizer, which is an AIE photosensitizer containing a sulfonium fragment. The AIE photosensitizer has the following general structural formula:

[0006]

[0007] In the formula, Donor is an electron-donating aromatic ring AIE mother nucleus, specifically any one of the following groups:

[0008]

[0009] In the formula, π is a linker aromatic ring, specifically any one of the following groups:

[0010]

[0011] In the formula, X - may be any one of I - , Br - , Cl - , BF4 - , PF6 - .

[0012] Another object of the present application is to provide a preparation method of the sulfonium AIE photosensitizer, which is achieved by the following preparation steps:

[0013] (1) using brominated triphenylamine or brominated tetraphenyl ethylene structure as raw material, and reacting with aryl boronic acid by Suzuki-Miyaura coupling reaction to obtain aldehyde intermediate product or tetraphenyl ethylene-triphenylamine coupling product;

[0014] (2) introducing a single aldehyde group on the aryl group of the tetraphenyl ethylene-triphenylamine coupling product by Vilsmeier-Haack reaction conditions to obtain the aldehyde intermediate product;

[0015] (3) the aldehyde intermediate product is subjected to Knoevenagel condensation with ethyl cyanoacetate under the condition of acetic acid / ammonium acetate to obtain an ester intermediate;

[0016] (4) the ester intermediate is dissolved in a mixed solvent of acetonitrile and methanol, and an aqueous sodium hydroxide solution is added to esterify it under heating conditions to obtain a carboxyl intermediate;

[0017] (5) using 2-(7-azabenzotriazol)-N,N,N',N'-tetramethyl urea hexafluorophosphate (HATU) as a condensation reagent, the carboxyl intermediate is subjected to amide condensation with 3-methyl mercaptopropylamine under alkaline conditions to obtain a sulfide intermediate;

[0018] (6) the sulfide intermediate is reacted with a methylation reagent, and then subjected to anion replacement to obtain the final product sulfonium salt derivative (sulfonium AIE photosensitizer).

[0019] Further, the molar ratio of the bromide raw material to the boronic acid raw material in step (1) is 1:1-1:2.5; the metal catalyst is 1,1'-bis(diphenylphosphino) ferrocene palladium dichloride or tetrakis(triphenylphosphine) palladium, and the amount of the metal catalyst is 1%-10% of the molar amount of the bromide raw material; the base used in the Suzuki-Miyaura coupling reaction is cesium carbonate; the solvent used in the reaction is a mixed solvent of 1,4-dioxane and water, and the volume ratio of 1,4-dioxane to water is 2:1-8:1; the coupling reaction temperature is 60°C-110°C, and the coupling reaction time is 1h-6h.

[0020] Further, the Vilsmeier-Haack reaction conditions in step (2) are a phosphorus oxychloride / N,N-dimethylformamide system, and the molar ratio of phosphorus oxychloride to the aldehyde intermediate product obtained in step (1) or the tetraphenyl ethene-triphenyl amine coupling product is 1:1-10:1; the solvent used in the reaction is N,N-dimethylformamide; the reaction temperature is 0°C for dropwise addition of phosphorus oxychloride, and then the temperature is raised to 40°C-80°C, and the reaction time is 1h-6h.

[0021] Further, the molar ratio of ethyl cyanoacetate to the aldehyde intermediate product obtained in step (1) in step (3) is 1:1-4:1; the Knoevenagel condensation conditions are an acetic acid / ammonium acetate system, and the molar ratio of ammonium acetate to the aldehyde intermediate product is 1:1-4:1; the solvent used in the reaction is acetic acid; the reaction temperature is 90°C-140°C, and the reaction time is 2h-12h.

[0022] Further, the molar ratio of sodium hydroxide to the ester intermediate in step (4) is 1:1-8:1.

[0023] Further, the molar ratio of 3-methylthiopropylamine to the carboxyl intermediate in step (5) is 1:1-3:1; the molar ratio of HATU to the carboxyl intermediate is 1:1-3:1; the base is triethylamine or N,N-diisopropyl ethyl amine, and the molar ratio to the carboxyl intermediate is 1:1-10:1; the reaction solvent is dichloromethane, the reaction temperature is room temperature, and the reaction time is 1h-3h.

[0024] Further, the methylating agent in step (6) is one of trimethyloxonium tetrafluoroborate, iodomethane, bromomethane or chloromethane, and the molar ratio to the sulfide intermediate is 1:1-3:1; the anion replacement agent is AgBF4, AgPF6, KPF6 or NH4PF6; the solvent used in the reaction is dichloromethane, the reaction temperature is room temperature, and the reaction time is 1h-12h.

[0025] Still another object of the present application is to provide the use of the sulfonium AIE photosensitizer in the preparation of an antitumor drug. The sulfonium fragment provided by the present application can significantly improve the photosensitivity of the AIE small molecule photosensitizer, inhibit the proliferation of various tumor cells, and effectively inhibit the growth of tumors in mice in vivo. The tumor cells are prostate cancer cells, breast cancer cells, and skin cancer cells.

[0026] The present application has the following advantages: (1) The present application provides an aggregation-induced emission AIE photosensitizer, which can overcome the quenching problem of traditional macrocyclic photosensitizers in aqueous media. (2) The present application uses a sulfonium fragment to modify the AIE fluorophore, which increases the spin-orbit coupling SOC and reduces the singlet-triplet energy gap ΔE ST , thereby promoting intersystem crossing ISC and enhancing the photosensitivity of the fluorophore. (3) The sulfonium AIE small molecule photosensitizer provided by the present application has good optical properties, including good light stability, large molar absorption coefficient, and Stokes shift, which can realize in vivo tumor fluorescence imaging. (4) The sulfonium AIE small molecule photosensitizer provided by the present application has high ROS generation efficiency and wide photodynamic antitumor range, which can inhibit the proliferation of various tumor cells and effectively inhibit the growth of tumors in mice in vivo. BRIEF DESCRIPTION OF DRAWINGS

[0027] Figure 1 Fig. 6 is the fluorescence spectrum of the DCFH solution of S4, S12, and DMSO under different irradiation times.

[0028] Figure 2 Fig. 7 is a laser confocal image of S4, S12, and cell membrane probe Dio co-staining for 10 minutes.

[0029] Figure 3 Fig. 8 is a laser confocal image of S4, S12, and lipid droplet probe Bodipy493 / 503 co-staining for 3 hours.

[0030] Figure 4 Fig. 9 is a schematic diagram of the proliferation inhibition experiment of S4, S12 on tumor cells under dark and light conditions.

[0031] Figure 5 Fig. 10 is a schematic diagram of the fluorescence in vivo imaging and in vivo antitumor experiment of S12. DETAILED DESCRIPTION

[0032] The technical solutions of the present application will be described in detail below in conjunction with the drawings and examples, but the scope of protection of the present application is not limited thereto.

[0033] Example 1: (E)-(3-(3-(4-((4'-(2,2-bis(4-methoxyphenyl)-1-phenylvinyl)-[1,1'-biphenyl]-4-yl)(phenyl)amino)phenyl)-2-cyanoacrylamidyl)propyl)dimethylsulfonium tetrafluoroborate (Compound No. S4)

[0034] The synthetic route of photosensitizer S4 is as follows:

[0035]

[0036] Synthesis of Compound I: 1-(4-bromophenyl)-2,2-bis(4-methoxyphenyl)-1-phenylvinyl (1.41 g, 3 mmol), 4-boronic acid triphenylamine (1.04 g, 3.6 mmol), Pd(dppf)Cl2(324 mg, 0.45 mmol,) and Cs2CO3(1.46 g, 4.5 mmol) were dissolved in mixed solvents (1,4-dioxane:H2O = 4:1). The reaction was heated at 90 °C for 3 h under nitrogen protection. After the system was cooled to room temperature, the solvent was rotary evaporated under reduced pressure, EA / H2O was used for extraction, and the organic phase was combined and separated by silica gel column chromatography to obtain Compound I (yellow solid, 1.16 g, yield 61%). 1 H NMR (500 MHz, Chloroform-d) δ 7.47-7.43 (m, 2H), 7.35-7.32 (m, 2H), 7.28-7.24 (m, 4H), 7.14-7.09 (m, 9H), 7.08-7.04 (m, 4H), 7.04-7.00 (m, 2H), 7.00-6.93 (m, 4H), 6.69-6.60 (m, 4H), 3.75 (s, 3H), 3.74 (s, 3H). HRMS (ESI): m / z calcd for (C 46 H 37 NO2+H) + : 636.2897; found: 636.2889.

[0037] Synthesis of Compound II: Compound I (635 mg, 1.0 mmol) was dissolved in 10 mL of DMF, and POCl3(746 uL, 8 mmol) was slowly added dropwise at 0 °C. After the system was stirred at 0 °C for 30 min, it was warmed to 60 °C and stirred for 3 h. After the reaction was cooled to room temperature, ice water was slowly added to the system, and EA (50 mL x 3) was used for extraction. The organic phase was combined, dried over anhydrous Na2SO4, and the solvent was rotary evaporated. Compound II was obtained by silica gel column chromatography (yellow solid, 378 mg, yield 57%). 1H NMR (500 MHz, Chloroform-d) δ 9.82 (s, 1H), 7.71 - 7.67 (m, 2H), 7.55 - 7.52 (m, 2H), 7.37 - 7.33 (m, 4H), 7.21 - 7.16 (m, 5H), 7.15 - 7.04 (m, 9H), 7.00 - 6.93 (m, 4H), 6.68 - 6.62 (m, 4H), 3.75 - 3.74 (m, 6H). HRMS (ESI): m / z calcd for (C 47 H 37 NO3+H) + : 664.2864; found: 664.2867.

[0038] Synthesis of compound III: Compound II (663 mg, 1 mmol), ethyl cyanoacetate (524 mg, 4 mmol) and ammonium acetate (308 mg, 4 mmol) were dissolved in acetic acid, the mixture was reacted at 130 °C for 12 h, and the reaction was quenched with water. EA (50 mL x 3) was used for extraction, and the organic phase was combined. After the solvent was rotary evaporated, silica gel column chromatography was used to obtain compound III (yellow solid, 598 mg, yield 79%). 1 H NMR (500 MHz, Chloroform-d) δ 8.11 (s, 1H), 7.87 (d, J = 9.0 Hz, 2H), 7.55 (d, J = 8.5 Hz, 2H), 7.24 - 7.17 (m, 5H), 7.15 - 7.05 (m, 7H), 7.04 - 6.93 (m, 6H), 6.66 - 6.63 (m, 4H), 4.35 (q, J = 8.0 Hz, 2H), 3.75 - 3.74 (m, 6H), 1.38 (t, J = 8.0 Hz, 3H). HRMS (ESI): m / z calcd for (C 52 H 42 N2O4+H) + : 759.3217; found: 759.3212.

[0039] Synthesis of compound IV: Compound III (776 mg, 1 mmol) and sodium hydroxide (48 mg, 1.2 mmol) were added to a three-necked flask, and mixed solvents (CH3CN:MeOH:H2O = 3 mL:3 mL:3 mL) were added to reflux for 2 h. After cooling to room temperature, 1M HCl was used to adjust the pH to 1. Then DCM (50 mL x 3) was used for extraction, and the organic phase was combined. After the solvent was rotary evaporated, silica gel column chromatography was used to separate to obtain compound IV (orange solid, 555 mg, yield 76%). 1H NMR (500 MHz, Chloroform-d) δ 8.14 (s, 1H), 7.89 (d, J = 9.0 Hz, 2H), 7.55 (d, J = 8.5 Hz, 2H), 7.41 - 7.33 (m, 4H), 7.24 - 7.18 (m, 5H), 7.15 - 7.05 (m, 8H), 7.04 - 6.92 (m, 6H), 6.67 - 6.63 (m, 4H), 3.75 (m, 6H). HRMS (ESI): m / z calcd for (C 50 H 38 N2O4+H) + : 731.2904; found: 731.2906.

[0040] Synthesis of compound V: Compound IV (120 mg, 0.16 mmol), HATU (92 mg, 0.24 mmol) and triethylamine were dissolved in dichloromethane, after stirring at room temperature for 30 min, 3- methylthiopropylamine (35 μL, 0.32 mmol) was added to the system and continue to stir at room temperature for 1 h. The reaction was quenched with saturated ammonium chloride solution. The organic phase was collected by extraction, the solvent was rotary evaporated and then separated by silica gel column chromatography to obtain compound V (orange solid, 91 mg, yield 70%) 1 H NMR (500 MHz, DMSO-d6) δ 8.32 (t, J = 5.5 Hz, 1H), 8.02 (s, 1H), 7.85 (d, J = 9.0 Hz, 2H), 7.65 (d, J = 8.5 Hz, 2H), 7.47 (d, J = 8.0 Hz, 2H), 7.42 (t, J = 8.0 Hz, 2H), 7.24 - 7.14 (m, 7H), 7.11 - 7.08 (m, 1H), 7.02 - 6.99 (m, 4H), 6.97 - 6.86 (m, 6H), 6.72 - 6.67 (m, 4H), 3.67 (s, 6H), 3.28 (q, J = 6.5 Hz, 2H), 2.50 - 2.47 (m, 2H), 2.04 (s, 3H), 1.78 - 1.72 (m, 2H). HRMS (ESI): m / z calcd for (C 54 H 47 N3O3S+Na) + : 840.3230; found: 840.3228.

[0041] Synthesis of compound S4: Compound V (100 mg, 0.13 mmol) and trimethyl oxonium tetrafluoroborate (23 mg, 0.16 mmol) were dissolved in DCM under nitrogen protection, stirred at room temperature for 2 h, the solvent was removed under reduced pressure to obtain an oily crude product, then 10 mL of ether was added to the slurry, suction filtered, and the solid was dried under vacuum to obtain compound S4 (orange solid, 93 mg, yield 78%). 1 H NMR (500 MHz, DMSO-d6) δ 8.41 (t, J = 5.5 Hz, 1H), 8.06 (s, 1H), 7.87 (d, J = 9.0 Hz, 2H), 7.67 (d, J = 8.5 Hz, 2H), 7.50 - 7.41 (m, 4H), 7.26 - 7.23 (m, 1H), 7.21 - 7.14 (m, 6H), 7.12 - 7.09 (m, 1H), 7.04 - 6.98 (m, 4H), 6.96 - 6.94 (m, 2H), 6.92 - 6.90 (m, 2H), 6.87 - 6.86 (m, 2H), 6.72 - 6.68 (m, 4H), 3.68 (s, 6H), 3.38 - 3.33 (m, 2H), 3.27 (t, J = 8.0 Hz, 2H), 2.88 (s, 6H), 1.98 - 1.92 (m, 2H). 13 C NMR (125 MHz, DMSO-d6) δ 161.8, 157.8, 157.7, 151.0, 150.1, 145.3, 144.7, 143.7, 142.9, 140.1, 138.2, 136.7, 136.0, 135.6, 135.6, 132.2, 132.0, 132.0, 131.4, 130.8, 130.1, 127.9, 127.8, 126.3, 126.1, 125.6, 125.6, 123.6, 119.1, 117.2, 113.2, 113.1, 100.6, 54.9, 38.1, 24.2, 23.5. HRMS (ESI): m / z calcd for (C 55 H 50 N3O3S) + : 832.3567; found: 832.3561.

[0042] Example 2: (E)-(3-(3-(5-(4-(bis(4-methoxyphenyl)amino)phenyl)thiophen-2-yl)-2- cyanoacrylamido)propyl)dimethylsulfonium tetrafluoroborate (Compound No. S12)

[0043] The synthetic route of photosensitizer S12 is as follows:

[0044]

[0045] Synthesis of compound VI: 4-bromo-N-N-dimethoxyaniline (1.15 g, 3 mmol), 5-formyl-2-thiopheneboronic acid (562 mg, 3.6 mmol), Pd(dppf)Cl2(324 mg, 0.45 mmol,) and Cs2CO3(1.46 g, 4.5 mmol) were dissolved in mixed solvents (1,4-dioxane:H2O = 4:1). The reaction was heated at 90 °C for 3 h under nitrogen protection. The system was cooled to room temperature, the solvent was rotary evaporated under reduced pressure, and EA / H2O was used to extract. The organic phase was combined and separated by silica gel column chromatography to obtain compound VI (orange solid, 759 mg, yield 61%). 1 H NMR (500 MHz, Chloroform-d) δ 9.84 (s, 1H), 7.70 (d, J = 4.0 Hz, 1H), 7.47 - 4.46 (m, 2H), 7.27 (d, J = 2.0 Hz, 1H), 7.12 - 7.08 (m, 4H), 6.91 - 6.90 (m, 2H), 6.89 - 6.86 (m, 4H), 3.82 (s, 6H). HRMS (ESI): m / z calcd for (C 25 H 21 NO3S+H) + : 416.1315; found: 416.1316.

[0046] Synthesis of compound VII: Compound VI (415 mg, 1 mmol), ethyl cyanoacetate (524 mg, 4 mmol) and ammonium acetate (308 mg, 4 mmol) were dissolved in acetic acid. The mixture was reacted at 130 °C for 12 h, and the reaction was quenched with water. EA (50 mL x 3) was used to extract, and the organic phase was combined. After rotary evaporation of the solvent, silica gel column chromatography was used to obtain compound VII (red solid, 357 mg, yield 70%). 1 H NMR (500 MHz, Chloroform-d) δ 8.25 (s, 1H), 7.69 (d, J = 4.5 Hz, 1H), 7.48 - 7.46 (m, 2H), 7.27 (s, 1H), 7.11 - 7.07 (m, 4H), 6.90 - 6.84 (m, 6H), 4.35 (q, J = 7.0 Hz, 2H), 3.81 (s, 6H), 1.38 (t, J = 7.0 Hz, 3H). HRMS (ESI): m / z calcd for (C 30 H 26 N2O4S+H) + : 511.1686; found: 511.1696.

[0047] Synthesis of compound VIII: Compound VII (510 mg, 1 mmol) and sodium hydroxide (48 mg, 1.2 mmol) were added to a three-necked flask, mixed solvent (CH3CN:MeOH:H2O = 3 mL:3 mL:3 mL) was added and refluxed for 2 h. After cooling to room temperature, pH was adjusted to 1 with 1 M HC1. Then extracted with DCM (50 mL x 3), the organic phase was combined, the solvent was evaporated, and compound VIII (red solid, 376 mg, yield 78%) was obtained by silica gel column chromatography. 1 H NMR (500 MHz, Chloroform-d) δ 8.29 (s, 1H), 7.74 (d, J = 4.0 Hz, 1H), 7.52 - 7.46 (m, 2H), 7.29 (d, J = 4.0 Hz, 1H), 7.12 - 7.08 (m, 4H), 6.91 - 6.86 (m, 6H), 3.82 (s, 6H). HRMS (ESI): m / z calcd for (C 28 H 22 N2O4S+H) + : 483.1373; found: 483.1377.

[0048] Synthesis of compound IX: Compound VIII (154 mg, 0.32 mmol), HATU (184 mg, 0.48 mmol) and triethylamine were dissolved in dichloromethane, stirred at room temperature for 30 min, then 3- methylthiopropylamine (70 μL, 0.64 mmol) was added to the system and continued to stir at room temperature for 1 h. The reaction was quenched with saturated ammonium chloride solution. The organic phase was collected by extraction, and the solvent was evaporated after drying to obtain compound IX (orange solid, 124 mg, yield 68%) by silica gel column chromatography. 1 H NMR (500 MHz, Chloroform-d) δ 8.32 (s, 1H), 7.62 (d, J = 4.5 Hz, 1H), 7.49 - 7.44 (m, 2H), 7.24 (d, J = 4.0 Hz, 1H), 7.11 - 7.06 (m, 4H), 6.89 - 6.84 (m, 6H), 6.49 (t, J = 6.0 Hz, 1H), 3.81 (s, 6H), 3.53 (q, J = 7.0 Hz, 2H), 2.58 (t, J = 7.0 Hz, 2H), 2.12 (s, 3H), 1.91 (p, J = 7.0 Hz, 2H). HRMS (ESI): m / z calcd for (C 32 H 31 N3O3S2+H) + : 570.1880; found: 570.1878.

[0049] Synthesis of compound S12: Compound IX (74 mg, 0.13 mmol) and trimethyl oxonium tetrafluoroborate (23 mg, 0.16 mmol) were dissolved in DCM under nitrogen protection, stirred at room temperature for 2 h, the solvent was removed under reduced pressure to obtain an oily crude product, then 10 mL of ether was added to the slurry, suction filtered, and the solid was dried under vacuum to obtain compound S12 (red solid, 70 mg, yield 80%). 1 H NMR (500 MHz, DMSO-d6) δ 8.41 (t, J = 6.0 Hz, 1H), 8.35 (s, 1H), 7.88 (d, J = 4.0 Hz, 1H), 7.60 - 7.56 (m, 2H), 7.54 (d, J = 4.0 Hz, 1H), 7.12 - 7.08 (m, 4H), 6.97 - 6.94 (m, 4H), 6.79 - 6.75 (m, 2H), 3.76 (s, 6H), 3.36 - 3.32 (m, 2H), 3.30 - 3.26 (m, 2H), 2.87 (s, 6H), 1.97 - 1.91 (m, 2H). 13 C NMR (125 MHz, DMSO-d6) δ 161.9, 156.8, 153.1, 150.1, 144.2, 141.1, 139.5, 133.5, 127.9, 127.6, 123.6, 123.6, 118.6, 117.4, 115.5, 99.3, 56.5, 55.7, 38.5, 24.6, 23.9, 19.0. HRMS (ESI): m / z calcd for (C 33 H 34 N3O3S2) + : 584.2036; found: 584.2037.

[0050] Example 3: ROS generation test of sulfonium photosensitizer

[0051] 2',7'-dichlorofluorescein (DCFH) is a classic ROS indicator, which is often used to evaluate the total ROS generation capacity of photosensitizer. ROS can oxidize DCFH to fluorescent compound 2'-7' dichlorofluorescein (DCF), and the maximum emission wavelength of DCF is about 522 nm, which can be detected by fluorescence spectrophotometer. The test concentration of S4 and S12 is 1 μM (1% DMSO in PBS solution), and the test concentration of DCFH is 10 μM. The power of the laser is 20 mW / cm 2The mixed solution was irradiated with white light (400-800 nm), and the change in fluorescence intensity in the range of 490-600 nm under different total irradiation times was recorded as I / I0, where I is the fluorescence intensity after irradiation, I0 is the fluorescence intensity at 0 s without irradiation, and the larger the ratio of I / I0, the stronger the ability of the tested substance to generate ROS. The test results are shown in Figure 1 As shown in FIG. 6, with the extension of the irradiation time, the fluorescence intensity of the DCFH solution in pure DMSO at 522 nm did not change substantially, while the fluorescence of the DCFH solution of sulfonium compounds S4 and S12 was enhanced by 45 and 55 times, respectively, indicating that the sulfonium compounds have strong ability to generate ROS under irradiation, and preliminary verification shows that the modification of AIE photosensitizer with sulfonium salt not only avoids the quenching problem of porphyrin in water, but also significantly improves the photosensitivity of AIE photosensitizer.

[0052] Example 4: Tumor cell uptake and organelle localization test of sulfonium photosensitizer

[0053] Tumor cells 4T1 were cultured with S4 and S12 containing medium respectively, and different commercial organelle localization probes were added for co-incubation. The fluorescence signals in the tumor cells were collected using a laser confocal microscope to observe the uptake of S4 and S12 in the tumor cells. The fluorescence signals of S4 and S12 in 4T1 cells incubated for 10 minutes and 3 hours were collected respectively, and the confocal results are shown in FIG. 7. Figure 2 、 Figure 3 As shown in FIG. 7, the sulfonium AIE photosensitizer shows extremely fast cell uptake, and the fluorescence signals of the above sulfonium AIE photosensitizer can be detected in the tumor cells within 10 minutes ( Figure 2 ), and by comparing with the fluorescence signals of commercial cell membrane green fluorescence probe Dio, it can be found that the sulfonium AIE photosensitizer is anchored on the cell membrane in about 10 minutes, and with the extension of the incubation time, the photosensitizer anchored on the cell membrane gradually enters the cells and is basically enriched in the lipid droplets in the cells in about 3 hours, which is basically consistent with the fluorescence signals of commercial lipid droplet probe Bodipy493 / 503, and the Pearson correlation coefficient is greater than 0.8 ( Figure 3 ). This time-dependent organelle dual targeting phenomenon comes from the positive charge of S4 and S12. The positive charge of sulfonium enables it to bind to the negatively charged cancer cell membrane through electrostatic interaction, and at the same time, the AIE hydrophobic group is embedded in the hydrophobic region of the phospholipid bilayer, at which time the intramolecular motion (RIM) is restricted, and the compound emits fluorescence. With the extension of the incubation time of S4 and S12 in the cells, the photosensitizer may be transported from the cell membrane to the lipid droplets.

[0054] Example 5: Study on the cell level anti-tumor activity of sulfonium photosensitizer

[0055] A431, human prostate cancer cell line PC3 and mouse breast cancer cell line 4T1 were selected as experimental cell lines. Different concentrations of S4 and S12 drug-containing medium were cultured in the above cell lines in 96-well plates, 3h later, fresh cell culture medium was replaced to wash away the photosensitizer that did not enter the cells, then the 96-well plates were irradiated with white light (400-800nm) with a power of 20mW / cm 2 for 15 minutes, and continued to be cultured in the dark for 24 hours. The control group was kept in the dark throughout the process. After 24 hours, the tumor cell survival rate was tested by MTT method. The results are shown in Figure 4 Compared with the dark treatment, the growth of tumor cells under light condition was more strongly inhibited, and the anti-tumor activity of S12 was stronger. The Light-IC 50 of S12 in the three tumor cells were all around 0.9μM, which was more than 10 times higher than that of the non-light group, showing strong anti-tumor photosensitivity.

[0056] Example 6: Small animal fluorescence live imaging study

[0057] The photosensitizer S12 was used for small animal fluorescence imaging using 4T1 tumor-bearing BALB / c mice. S12 with a concentration of 1mg / mL was injected into the mice by intratumoral injection, and the fluorescence signals at different times were collected by a small animal live imaging instrument. As shown in Figure 5 -A, the PBS group as a blank control group did not collect any fluorescence signal, while the mice injected with S12 could clearly observe the fluorescence emission at the tumor site and distinguish it from the surrounding tissues, showing an ideal signal-to-noise ratio. Within 0-3 hours after administration, the fluorescence signal showed an increasing trend in the tumor site, which may be due to the aggregation-induced fluorescence enhancement caused by the uptake of S12 by tumor cells. As the metabolic time continued to extend, the fluorescence signal of S12 was still clearly visible at the tumor site 24 hours after injection, indicating that the photosensitizer could effectively stay in the tumor site and was less likely to cause toxic side effects to other organs.

[0058] Example 7: In vivo anti-tumor activity study

[0059] The photodynamic therapy effect of S12 in vivo tumor was evaluated. 4T1 tumor-bearing BALB / c mice were randomly divided into three groups: A) Control light+ group, B) S12 light- group, C) S12 light+ group. When the tumor volume grew to about 40-50mm 3 , the administration began, the administration method was intratumoral injection, and the administration dose was 5mg / kg (1mg / mL, 100μL). The Controllight+ group and S12 light+ group mice received light irradiation with an intensity of 150mW / cm 2white light treatment, with a 15-minute irradiation length. S12 light- group mice were not treated with light. The light treatment was repeated again after one day, for a total of two treatments. Tumor volume and body weight changes were recorded every two days. The results are shown in Figure 5 As shown in Figure 6, the tumor volume of the Control light+ group and the S12 light- group increased significantly during the treatment, and there was no significant difference between the two groups, indicating that S12 has good biological safety. During the PDT treatment, the body weight of the S12 light+ group was basically consistent with that of the control group (D), and there was no obvious toxic side effect. Under the light condition, the tumor growth of the S12 light+ group was significantly inhibited, which was significantly different from the other two groups, showing good in vivo photodynamic anti-tumor activity. Figure 5

[0060] From the above examples, it can be seen that the present application provides a preparation method and application of a photosensitizer containing a sulfonium fragment. The preparation method is simple to operate and the raw materials are easy to obtain. The AIE photosensitizer modified by the sulfonium fragment has strong ROS generation capacity and time-dependent organelle dual targeting ability, and shows a broad-spectrum anti-tumor effect on multiple tumor cell lines, can effectively inhibit the growth of tumor-bearing mice, and shows good application potential in the field of PDT.

[0061] The method and application of the present application are not limited to the above exemplary embodiments, and for those skilled in the art, other forms of several improvements and modifications made without departing from the purpose or basic characteristics of the present application should be considered as falling within the protection scope of the present application.​

Claims

1. A sulfonium aggregation-induced emission photosensitizer, characterized in that: The sulfonium aggregation-induced luminescence photosensitizers are: 。 2. The method for preparing the sulfonium aggregation-induced emission photosensitizer S4 according to claim 1, characterized in that: This is achieved by following these steps: 。 3. The method for preparing the sulfonium aggregation-induced emission photosensitizer S12 according to claim 1, characterized in that: This is achieved by following these steps: 。 4. Use of the sulfonium aggregation-induced emission photosensitizer according to claim 1 in the preparation of anti-tumor drugs.

5. The use according to claim 4, characterized in that The application is that the sulfonium fragment can significantly improve the photosensitivity of the aggregation-induced luminescence small molecule photosensitizer and inhibit the proliferation of tumor cells.

6. The use according to claim 4, characterized in that The tumor is prostate cancer, breast cancer, or skin cancer.

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