Aggregation-induced emission materials, methods of making the same, drug delivery systems, methods of making the same, and applications thereof

Abstract

The present application relates to the technical field of fluorescent materials, and particularly relates to an aggregation-induced emission (AIE) material, a preparation method of the AIE material, a drug delivery system, a preparation method of the drug delivery system and application, the AIE material has unique characteristics as a photosensitizer, compared with a traditional photosensitizer with a planar conformation, the AIE photosensitizer can utilize a larger proportion of absorbed energy to emit fluorescence, generate reactive oxygen species (ROS) or photothermal. Therefore, the AIE material provided by the present application can efficiently generate ROS and photothermal in a cell environment; and the AIE material has the characteristics of photodynamic, photothermal therapy and fluorescence, photoacoustic, photothermal imaging; in addition, the drug delivery system comprises the AIE material and bacteria, the bacteria are used as a delivery carrier, so that the accumulation and penetration of the drug in tumor tissue can be enhanced, the curative effect is optimized, the specific killing of tumor tissue is realized, and the multi-modal tumor imaging and anti-tumor efficacy of the AIE material are significantly enhanced.

Description

Technical Field

[0001] This invention relates to the field of fluorescent materials technology, and in particular to an aggregation-induced emission material and its preparation method, a drug delivery system and its preparation method and application. Background Technology

[0002] Malignant tumors have always been a major threat to human health. Traditional treatments, such as surgery, chemotherapy, and radiotherapy, often face challenges of limited efficacy and poor patient prognosis. Therefore, the development of effective cancer treatments is particularly urgent. In recent years, photodynamic therapy (PDT) and photothermal therapy (PTT) have received widespread attention due to their potential to overcome the limitations of traditional therapies. In particular, the thermal effect of PTT can increase oxygen supply, thereby enhancing the therapeutic effect of PDT. However, the strong intermolecular interactions of traditional photosensitizers limit the production of reactive oxygen species (ROS), and the limited diffusion range and short lifespan of ROS in solid tumors further restrict the therapeutic effect.

[0003] Against this backdrop, aggregation-induced emission (AIE) materials offer a new perspective for cancer treatment due to their unique optical properties and broad application potential. AIE materials exhibit enhanced luminescence upon molecular aggregation, a stark contrast to the aggregation-induced quenching (ACQ) phenomenon in traditional organic light-emitting materials. This characteristic makes AIE materials demonstrate enormous application potential in areas such as bioimaging and integrated disease diagnosis and treatment. Particularly in cancer treatment, AIE materials can enhance therapeutic effects by improving the generation efficiency of reactive oxygen species (ROS) and mitigating their diffusion within tumor tissue. However, achieving efficient targeting of tumor sites, inducing sustained anti-tumor effects in vivo, and reducing accumulation in organs such as the liver and spleen remain critical bottlenecks that need to be addressed. The efficiency of current tumor targeting technologies still requires improvement and development.

[0004] Bacteria possess advantages such as targeted colonization at tumor sites, making bacterial therapy a unique treatment approach and a new direction in cancer treatment. However, the mechanism of action of bacterial therapy remains unclear, and issues such as poor controllability, poor safety, and low efficacy when used alone limit its application and development. Therefore, developing NIR fluorescent molecules and new strategies for bacterial-mediated tumor diagnosis and treatment, and integrating the advantages of AIE optical diagnosis and bacterial targeting to improve the precision, safety, and efficiency of cancer diagnosis and treatment, is of great significance. Summary of the Invention

[0005] In view of the shortcomings of the prior art, the purpose of this invention is to provide an aggregation-induced emission material and its preparation method, a drug delivery system and its preparation method and application, aiming to solve the problem that traditional photosensitizers hinder the generation of ROS and weaken the efficacy of photodynamic therapy.

[0006] The technical solution of the present invention is as follows:

[0007] An aggregation-induced emission (AIE) material, the general molecular structure of which is shown in formula (1):

[0008]

[0009] R1 is selected from hydrogen, methyl, and methoxy; R2 is selected from methyl and ethyl.

[0010] In the aggregate-induced emission material, R1 is methoxy; R2 is ethyl.

[0011] A method for preparing an aggregation-induced emission material includes the following steps:

[0012] 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, Sodium dithiosulfate, palladium diacetate, and an organic solvent were heated and stirred to obtain the first product.

[0013] Will The catalyst is mixed with the first product and then refluxed to obtain the second product;

[0014] The second product was mixed with a saturated potassium hexafluorophosphate solution to obtain an aggregation-induced emission material.

[0015] The method for preparing the aggregation-induced emission material, wherein the 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, The molar ratio of sodium dithiosulfate and palladium diacetate is (0.5-1.5):(0.8-1.5):(0.8-1.5):(0.4-1).

[0016] The method for preparing the aggregation-induced emission material, wherein the first product and The molar ratio is (1-2):(1-2).

[0017] The method for preparing the aggregation-induced luminescence material, wherein the heating and stirring treatment is performed at a temperature of 110℃-140℃ for a duration of 36h-54h; and / or the reflux treatment is performed for a duration of 20h-30h.

[0018] A drug delivery system includes bacteria and an aggregation-induced luminescent material; the aggregation-induced luminescent material is loaded on the surface of the bacteria.

[0019] The drug delivery system wherein the bacteria include one or more of Escherichia coli Nissle1917, attenuated transgenic Salmonella Typhimurium strain VNP20009, Bifidobacterium, and lactic acid bacteria.

[0020] A method for preparing a drug delivery system includes the following steps:

[0021] The aggregation-induced emission material was mixed with an organic solvent to obtain a mother liquor;

[0022] The mother liquor is mixed with a bacterial solution and incubated to obtain a drug delivery system.

[0023] Application of a drug delivery system in the preparation of antitumor drugs.

[0024] Beneficial Effects: This invention provides an aggregation-induced emission (AIE) material and its preparation method, a drug delivery system and its preparation method and applications. AIE, as a photosensitizer, possesses unique characteristics. Compared to traditional photosensitizers with a planar conformation, AIE photosensitizers can utilize a larger proportion of absorbed energy to emit fluorescence and generate reactive oxygen species (ROS). Therefore, the AIE provided by this invention can efficiently generate ROS in a cellular environment; and it possesses photodynamic, photothermal therapy, and fluorescence, photoacoustic, and photothermal imaging properties. Furthermore, the drug delivery system includes the AIE material and bacteria. The tumor-targeting ability of bacteria is unaffected by the tumor genome composition and can stimulate innate and adaptive anti-tumor immune responses in the hypoxic, nutrient-deficient, and immunosuppressive environment of tumor tissue. Utilizing bacteria as a delivery carrier can expand the accumulation and penetration of drugs within tumor tissue, thereby optimizing therapeutic efficacy, achieving specific killing of tumor tissue, and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of the AIE material. Attached Figure Description

[0025] Figure 1 Synthetic route diagram for preparing aggregation-induced emission material INX-PF6 in Example 1;

[0026] Figure 2 The aggregation-induced emission material INX-PF6 in Example 1 1 H nuclear magnetic resonance spectrum;

[0027] Figure 3 The aggregation-induced emission material INX-PF6 in Example 1 13 C NMR spectrum;

[0028] Figure 4 The aggregation-induced emission material INX-PF6 in Example 1 19 F nuclear magnetic resonance spectrum;

[0029] Figure 5This is a high-resolution mass spectrum of the aggregation-induced emission material INX-PF6 in Example 1;

[0030] Figure 6 This is a confocal image after co-incubation of EcN and INX-PF6;

[0031] Figure 7 A graph showing the changes in fluorescence intensity after EcN loaded with INX-PF6 detected by flow cytometry.

[0032] Figure 8 Confocal imaging of EcN@INX-PF6 penetrating the tumor sphere on CT26;

[0033] Figure 9 This is a graph showing the changes in tumor volume in mice after in vivo treatment.

[0034] Figure 10 Images of isolated tumor tissues from mice in each group after in vivo treatment. Detailed Implementation

[0035] This invention provides an aggregation-induced emission material and its preparation method, a drug delivery system and its preparation method, and its applications. To make the objectives, technical solutions, and effects of this invention clearer and more explicit, the invention is further described in detail below. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

[0036] It will be understood by those skilled in the art that, unless otherwise defined, all terms used herein (including technical and scientific terms) have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. It should also be understood that terms such as those defined in general dictionaries should be understood to have the same meaning as in the context of the prior art, and should not be interpreted in an idealized or overly formal sense unless specifically defined as herein.

[0037] The strong intermolecular interactions in traditional photosensitizers hinder the generation of reactive oxygen species (ROS), thus weakening the efficacy of photodynamic therapy. Furthermore, the limited diffusion range and short lifespan of ROS within solid tumors also affect treatment outcomes. Therefore, it is imperative to innovate novel photosensitizers with enhanced ROS generation capabilities to improve photodynamic therapy for deep solid tumors.

[0038] Based on this, the present invention provides an aggregation-induced emission material, the general molecular structure of which is shown in formula (1):

[0039]

[0040] R1 is selected from hydrogen, methyl, and methoxy; R2 is selected from methyl and ethyl.

[0041] In this embodiment, the aggregation-induced emission (AIE) material, as a photosensitizer, possesses unique characteristics. Compared to traditional photosensitizers with a planar conformation, AIE photosensitizers can utilize a larger proportion of absorbed energy to emit fluorescence and generate ROS. Therefore, the AIE material provided by this invention can efficiently generate ROS in a cellular environment; and it possesses photodynamic, photothermal therapy, and fluorescence, photoacoustic, and photothermal imaging properties. Furthermore, the drug delivery system includes the AIE material and bacteria, and the tumor-targeting ability of the bacteria is unaffected by the tumor genome composition, enabling the stimulation of innate and adaptive anti-tumor immune responses in the hypoxic, nutrient-deficient, and immunosuppressive environment of tumor tissue. Utilizing bacteria as a delivery carrier can expand the accumulation and penetration of drugs within tumor tissue, thereby optimizing therapeutic efficacy, achieving specific killing of tumor tissue, and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of the AIE material.

[0042] In some embodiments, R1 is methoxy; R2 is ethyl.

[0043] Specifically, when R1 is methoxy and R2 is ethyl, the chemical structural formula of the aggregation-induced emission material is: This aggregation-induced emission material possesses photodynamic, photothermal therapy, and fluorescence, photoacoustic, and photothermal imaging properties. Then, the aggregation-induced emission material is combined through physical mixing or covalent bonding to obtain a bacterial drug delivery system of bacteria-aggregation-induced emission material. The bacterial-mediated bacterial drug delivery system can actively target hypoxic tumor tissue, achieve specific killing of tumor tissue, and significantly enhance the multimodal tumor imaging and anti-tumor efficacy of the aggregation-induced emission material.

[0044] In addition, the present invention also provides a method for preparing an aggregation-induced emission material, comprising the following steps:

[0045] Step S10: Add 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, Sodium dithiosulfate, palladium diacetate, and an organic solvent are heated and stirred to obtain a first product; the chemical structural formula of the first product is as follows.

[0046] Step S20: The catalyst is mixed with the first product and refluxed to obtain the second product; the chemical structural formula of the second product is as follows.

[0047] Step S30: The second product is mixed with a saturated potassium hexafluorophosphate solution to obtain an aggregation-induced emission material.

[0048] In this embodiment, the aggregation-induced emission material prepared by the above method has unique characteristics. Compared with traditional photosensitizers with a planar conformation, the AIE photosensitizer can utilize a larger proportion of absorbed energy to emit fluorescence and generate ROS. Therefore, the aggregation-induced emission material prepared by the above method can efficiently generate ROS in the cellular environment; and it possesses photodynamic, photothermal therapy, and fluorescence, photoacoustic, and photothermal imaging properties. Furthermore, the preparation process is simple, has high synthetic purity, and is suitable for large-scale preparation.

[0049] In some embodiments, the 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, The molar ratio of sodium dithiosulfate carbonate to palladium diacetate is (0.5-1.5):(0.8-1.5):(0.8-1.5):(0.4-1). Controlling the molar ratio within the above range ensures that a first product with high purity and complete reaction is obtained after heating and stirring, which is beneficial for the next synthesis step.

[0050] In a preferred embodiment, the 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, The molar ratio of sodium dithiosulfate and palladium diacetate is 0.68:1:1:0.6.

[0051] Specifically, step S10 includes: mixing 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, Sodium dithiosulfate, palladium diacetate, and an organic solvent were mixed under an inert atmosphere and then heated and stirred. After cooling to room temperature, the solvent was removed by vacuum rotary evaporation. The residue was dissolved again in dichloromethane and washed three times with saturated brine. The organic layer was dried on anhydrous Na2SO4 and then dried under reduced pressure. The crude product was purified by silica gel chromatography using hexane / dichloromethane as the eluent to obtain an orange powdery solid compound, namely 6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonene-4-carboxaldehyde.

[0052] In some embodiments, the volume ratio of n-hexane to dichloromethane in the eluent is (1-2):(1-2).

[0053] In a preferred embodiment, the volume ratio of n-hexane to dichloromethane in the eluent is 1:1.

[0054] In some embodiments, the first product and The molar ratio is (1-2):(1-2). After mixing according to the above molar ratio and reflux treatment, a second product with high purity and relatively complete reaction can be obtained.

[0055] In a preferred embodiment, the first product and The molar ratio is 1:1.

[0056] Specifically, step S20 includes: mixing 6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonoid-4-carboxaldehyde and... The solution was catalyzed by adding 50-100 μL of piperidine dropwise to anhydrous ethanol, refluxed under an inert atmosphere for 24 hours, cooled to room temperature, and the solvent was evaporated under reduced pressure. The residue was purified by silica gel chromatography and eluted with dichloromethane / methanol to give the second product, namely (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonen-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indole-1-onium iodide (INX-I).

[0057] In some embodiments, the heating and stirring treatment is performed at a temperature of 110°C-140°C for a duration of 36-54 hours; and / or, the reflux treatment is performed for a duration of 20-30 hours. Performing the heating and stirring treatment under these conditions allows the raw materials to react rapidly, shortening the reaction time.

[0058] In a preferred embodiment, the heating and stirring treatment is performed at a temperature of 120°C for 48 hours; and / or the reflux treatment is performed for 24 hours.

[0059] In some embodiments, the inert atmosphere includes one or more of argon, nitrogen, and helium.

[0060] In some embodiments, step S30 includes: dissolving INX-I in an organic solvent and mixing it with a saturated potassium hexafluorophosphate solution; stirring at room temperature for 1 hour; evaporating the organic solvent with compressed air; filtering again to obtain a dark red precipitate; washing with water; drying under reduced pressure; purifying the residue by silica gel chromatography; and eluting with dichloromethane / methanol to obtain the aggregation-induced emission material, namely (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonen-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indole-1-onium hexafluorophosphate (V) salt (INX-PF6).

[0061] In some embodiments, the volume ratio of dichloromethane to methanol is (15-25):1; preferably, the volume ratio of dichloromethane to methanol is 20:1.

[0062] In addition, the present invention also provides a drug delivery system comprising bacteria and an aggregation-induced luminescent material; the aggregation-induced luminescent material is loaded on the surface of the bacteria.

[0063] In this embodiment, by combining aggregation-induced emission materials through physical mixing or covalent bonding, a bacterial drug delivery system of bacteria-aggregation-induced emission materials is obtained. The bacterial-mediated bacterial drug delivery system can actively target hypoxic tumor tissue, achieve specific killing of tumor tissue, and significantly enhance the multimodal tumor imaging and anti-tumor efficacy of aggregation-induced emission materials.

[0064] Specifically, a fluorescent material with aggregation-induced emission properties is first synthesized and then combined with engineered bacteria under physical or chemical action to prepare a bacterial drug delivery system of bacteria-aggregation-induced emission material. After being injected via the tail vein, it can be targeted to tumor tissue and, under laser irradiation, generate photodynamic and photothermal effects to exert an anti-tumor effect.

[0065] In some embodiments, the bacteria include, but are not limited to, one or more of Escherichia coli Nissle1917 (EcN), attenuated transgenic Salmonella Typhimurium strain VNP20009 (VNP), Bifidobacteria, and Lactobacillus. Preferably, the bacteria are Escherichia coli Nissle1917 (EcN), which is known for its antibacterial properties, gut microbiota regulation, and facultative anaerobic activity.

[0066] In addition, the present invention also provides a method for preparing a drug delivery system, comprising the steps of:

[0067] Step S100: Mix the aggregation-induced light-emitting material with an organic solvent to obtain a mother liquor;

[0068] Step S200: The mother liquor is mixed with the bacterial solution and incubated to obtain a drug delivery system.

[0069] In this embodiment, the drug delivery system prepared by this method is combined through physical mixing or covalent bonding to obtain a bacterial drug delivery system for bacterial aggregation-induced emission materials. The bacterial-mediated bacterial drug delivery system can actively target hypoxic tumor tissue, achieve specific killing of tumor tissue, and significantly enhance the multimodal tumor imaging and anti-tumor efficacy of aggregation-induced emission materials.

[0070] In some embodiments, before step S100, the method further includes: reviving the cryopreserved bacterial strain, culturing it in a culture medium required by the bacteria, collecting colonies during the growth period, and obtaining a bacterial solution.

[0071] In some embodiments, step S100 includes: fully dissolving the aggregation-induced luminescence material INX-PF6 in DMSO to obtain a 1-1.5 mmol INX-PF6 mother solution.

[0072] In some embodiments, step S200 includes: taking 8-12 μL of INX-PF6 stock solution and adding it to 1 mL of a bacterial concentration of 5×10⁻⁶. 8 In a bacterial solution containing CFU, the bacteria are incubated with shaking at 37°C for 5-30 minutes to load the aggregation-induced luminescence material INX-PF6 onto the bacterial surface. The bacteria are then washed and centrifuged with PBS solution at pH 7.4 or ultrapure water, and the lower precipitate is collected to obtain the drug delivery system.

[0073] On the other hand, the present invention also provides the application of a drug delivery system in the preparation of antitumor drugs.

[0074] In this embodiment, the bacterial-mediated bacterial drug delivery system can actively target hypoxic tumor tissue, achieving specific killing of tumor tissue and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of aggregation-induced emission materials.

[0075] The following examples further illustrate the present invention in detail. It should also be understood that the following examples are only for further explanation of the present invention and should not be construed as limiting the scope of protection of the present invention. Any non-essential improvements and adjustments made by those skilled in the art based on the above description of the present invention are within the scope of protection of the present invention.

[0076] Example 1

[0077] This embodiment provides an aggregation-induced emission material INX-PF6 and a drug delivery system, specifically including the following:

[0078] The synthetic route of aggregation-induced emission material INX-PF6 is as follows: Figure 1 As shown, the specific steps are as follows:

[0079] 1) 6-Bromo-2,3-dihydro-1H-flavone-4-carboxaldehyde (200 mg, 0.68 mmol), p-methoxyaniline (229 mg, 1 mmol), sodium dithiosulfate (325 mg, 1 mmol), and palladium diacetate (20 mg, 0.6 mmol) were mixed in toluene under argon atmosphere. The reaction mixture was stirred at 120 °C for 48 hours. After cooling to room temperature, the solvent was removed by rotary evaporation under vacuum. The residue was reconstituted with dichloromethane and washed three times with saturated brine. The organic layer was dried on anhydrous Na₂SO₄ and then dried under reduced pressure. The crude product was purified by silica gel chromatography using n-hexane / dichloromethane (1:1, v / v) as the eluent to give compound 1, an orange powder, which is (6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavone-4-carboxaldehyde).

[0080] 2) A solution of 6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonen-4-carboxaldehyde (88 mg, 0.2 mmol) and 1-ethyl-2,3,3-trimethyl-3H-indole-1-onium iodide (63 mg, 0.2 mmol) was added dropwise to anhydrous ethanol for catalysis with piperidine. The mixture was refluxed under nitrogen for 24 hours, cooled to room temperature, and the solvent was evaporated under reduced pressure. The residue was purified by silica gel chromatography and eluted with dichloromethane / methanol (20:1, v / v) to give compound (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonen-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indole-1-onium iodide (INX-I).

[0081] 3) INX-I (74 mg, 0.1 mmol) was dissolved in anhydrous ethanol (10 mL) and mixed with saturated potassium hexafluorophosphate solution (10 mL). After stirring at room temperature for 1 hour, the anhydrous ethanol was evaporated with compressed air. The dark red precipitate was filtered again, washed with water, and dried under reduced pressure. The residue was purified by silica gel chromatography and eluted with dichloromethane / methanol (20:1, v / v) to give compound (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonen-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indole-1-onium hexafluorophosphate (V) salt (INX-PF6).

[0082] The structure of the INX-PF6 molecule is obtained through 1 H nuclear magnetic resonance (H nuclear magnetic resonance) Figure 2 ), 13 C nuclear magnetic resonance (C Figure 3 ), 19 F nuclear magnetic resonance (F Figure 4 ) and high-resolution mass spectrometer ( Figure 5 Characterization and verification were performed.

[0083] 1 The results of H nuclear magnetic resonance are 1H NMR(500MHz,DMSO-d6)δ8.42(d,J=14.6Hz,1H),7.72(dd,J=7.5,1.2Hz,1H),7.64(d,J=8.0 Hz,1H),7.60-7.52(m,2H),7.50-7.41(m,2H),7.33-7.24(m,4H),7.13-7.05(m,4H),6.72( dd,J=8.7,2.3Hz,1H),6.58(d,J=2.3Hz,1H),6.48(d,J=14.8Hz,1H),4.39(q,J=7.2Hz,2H) ,3.86(s,6H),2.76(t,J=6.0Hz,2H),2.71(t,J=6.1Hz,2H),1.95-1.80(m,2H),1.63(s,6H).

[0084] 13 The results of the C NMR are 13 C NMR (126MHz, DMSO) δ175.88,161.75,157.82,154.58,152.72,141.96,141.74,138 .36,135.25,129.36,129.31,128.50,126.87,125.84,123.12,115.92,114.93,114 .34,112.85,101.86,55.93,50.29,34.66,31.43,28.95,28.70,27.62,27.56,25.2 5,20.56,14.44,12.84,11.72.HRMS(ESI)calculated:609.3112;Found:609.3088.

[0085] The specific steps for preparing a bacterial aggregation-induced luminescent material drug delivery system are as follows:

[0086] Take 10 μL of INX-PF6 stock solution (dissolved in DMSO, 1 mmol) and add it to 1 mL of a bacterial concentration of 5 × 10⁻⁶. 8 CFUs were placed in Escherichia coli EcN (Escherichia coli Nissle 1917, EcN) and shaken at 37°C (180 rpm) for 20 minutes. The mixture was then washed with PBS and centrifuged twice at 6000 rpm for 3 minutes to obtain the bacterial aggregation-induced luminescent material drug delivery system EcN@INX-PF6.

[0087] The applications of bacterial aggregation-induced luminescent material drug delivery systems are as follows:

[0088] Add EcN bacterial suspension (5×10⁻⁶) to 1 mL of PBS containing 10 μM INX-PF6. 8 CFUs mL -1 After culturing the bacteria in a shaker (37°C, 200 rpm) for 30 minutes, the supernatant was removed by centrifugation, and a small amount of PBS was added to resuspend the bacteria in PBS. 1 μL of the stained bacterial solution was transferred to a glass slide and covered with a coverslip. Images were acquired using a confocal laser scanning microscope, as shown below. Figure 6 As shown. The molecular load on the bacterial surface was detected by flow cytometry, such as... Figure 7 As shown, after 24 hours of incubation, INX-PF6 was still able to adhere stably to the bacteria.

[0089] Three-dimensional tumor spheroids of CT26 tumor cells were constructed by dissolving agarose in DMEM and sterilizing at high temperature to obtain a 2% solution. This solution (60 μL) was then spread onto the bottom of a 96-well plate. CT26 cells (2 × 10⁻⁶) were then added. 3 (Cells / well) were inoculated into agarose-coated plates and cultured for 5 days. After the formation of three-dimensional tumor spheroids of approximately 180-200 μm, the tumor spheroids were transferred to agarose-coated confocal dishes and treated with INX-PF6 or EcN@INX-PF6 for 24 h. Images were acquired using a confocal laser scanning microscope to verify the penetration depth of the drug-loaded bacteria. Figure 8 As shown in the figure, the results indicate that EcN@INX-PF6 has excellent penetration performance.

[0090] A CT26 tumor-bearing mouse model was constructed, with a tumor volume of approximately 150 mm. 3 At that time, tumor-bearing mice were randomly divided into 7 groups (n=5 per group) and treated differently. Different treatment agents were injected into the tail vein of the tumor-bearing mice. Changes in tumor growth volume were observed, and the results are as follows: Figure 9 and Figure 10 As shown, the results indicate that EcN@INX-PF6 has a strong anti-tumor killing ability.

[0091] In summary, this invention provides an aggregation-induced emission (AIE) material and its preparation method, a drug delivery system and its preparation method, and its applications. The AIE material possesses unique characteristics; compared to traditional photosensitizers with planar conformations, it can utilize a larger proportion of absorbed energy to emit fluorescence and generate reactive oxygen species (ROS). Therefore, the novel AIE provided by this invention can efficiently generate ROS in a cellular environment; and it possesses photodynamic, photothermal therapeutic, and fluorescence, photoacoustic, and photothermal imaging properties. Furthermore, the drug delivery system includes the AIE material and bacteria. The tumor-targeting ability of bacteria is unaffected by the tumor genome composition and can stimulate innate and adaptive anti-tumor immune responses in the hypoxic, nutrient-deficient, and immunosuppressive environment of tumor tissue. Utilizing bacteria as a delivery carrier can expand the accumulation and penetration of drugs within tumor tissue, thereby optimizing therapeutic efficacy, achieving specific killing of tumor tissue, and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of the AIE material.

[0092] It should be understood that the application of the present invention is not limited to the examples above. Those skilled in the art can make improvements or modifications based on the above description, and all such improvements and modifications should fall within the protection scope of the appended claims.

Claims

1. A drug delivery system, characterized in that, It includes bacteria and aggregation-induced emission material; the aggregation-induced emission material is loaded on the surface of the bacteria; The bacteria are selected from one or more of Escherichia coli Nissle1917, transgenic Salmonella Typhimurium attenuated strain VNP20009, Bifidobacterium, and Lactobacillus. The general molecular structural formula of the aggregation-induced emission material is: Wherein, R1 is methoxy; R2 is ethyl.

2. A method for preparing the drug delivery system as described in claim 1, characterized in that, Including the following steps: The aggregation-induced emission material was mixed with an organic solvent to obtain a mother liquor; The mother liquor is mixed with a bacterial solution and incubated to obtain a drug delivery system.

3. The use of the drug delivery system as described in claim 1 in the preparation of a colorectal cancer drug.

Images