Aggregation-induced emission material, preparation method of aggregation-induced emission material, drug delivery system and preparation method and application of drug delivery system

Abstract

The invention relates to the technical field of fluorescent materials, in particular 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 of the drug delivery system. The AIE photosensitizer can utilize a larger proportion of absorption energy to emit fluorescence, generating reactive oxygen species (ROS) or light and heat. Therefore, the AIE material provided by the invention can efficiently generate ROS and light and heat in a cell environment; the material has the characteristics of photodynamic and photo-thermal therapy and fluorescence, photoacoustic and photo-thermal imaging; besides, the drug delivery system comprises the AIE material and bacteria, and accumulation and permeation of the drug in the tumor tissue can be enhanced by using the bacteria as a delivery carrier, so that the curative effect is optimized, specific killing of the tumor tissue is realized, and the multi-modal tumor imaging and anti-tumor effects of the AIE material are remarkably enhanced.

Description

Technical Field

[0001] The present invention relates to the technical field of fluorescent materials, and in particular to an aggregation-induced emission material and a preparation method thereof, a drug delivery system and a preparation method and application thereof. Background Art

[0002] Malignant tumors have always been a huge threat to human health. Traditional treatments, such as surgery, chemotherapy, and radiotherapy, often face the 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 attracted 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 interactions between traditional photosensitizer molecules limit the generation of reactive oxygen species (ROS). In addition, the limited diffusion range and short lifespan of ROS in solid tumors limit the therapeutic effect.

[0003] In this context, aggregation-induced emission (AIE) materials provide a new perspective for cancer treatment due to their unique optical properties and wide application potential. The luminescence of AIE materials is enhanced when the molecules aggregate, which is in sharp contrast to the "aggregation-induced luminescence quenching" (ACQ) phenomenon of traditional organic light-emitting materials. This characteristic of AIE materials makes them show great application potential in bioimaging, integrated disease diagnosis and treatment, etc. Especially in the field of cancer treatment, AIE materials can enhance the therapeutic effect by increasing the efficiency of ROS generation and improving its diffusion in tumor tissues. However, how AIE materials can efficiently target tumor sites, induce lasting anti-tumor effects in the body, and reduce accumulation in organs such as the liver and spleen are still bottleneck problems that need to be solved. The efficiency of existing technologies in targeting tumors needs to be improved and developed.

[0004] Bacteria have advantages such as targeted colonization at tumor sites. As a unique treatment method, bacterial therapy has become a new direction for cancer treatment. However, the mechanism of action of bacterial therapy is still unclear, and problems such as poor controllability, poor safety, and low efficacy when used alone limit its application development. Therefore, it is of great significance to develop NIR fluorescent molecules and new strategies that can be used for bacteria-mediated tumor diagnosis and treatment, and to integrate the advantages of AIE optical diagnosis and treatment and bacterial targeting to improve the accuracy, safety, and efficiency of cancer diagnosis and treatment. Summary of the invention

[0005] In view of the above-mentioned deficiencies in the prior art, the object of the present invention is to provide an aggregation-induced emission material and a preparation method thereof, a drug delivery system and a preparation method and application thereof, 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, wherein the molecular structure of the aggregation-induced emission material is shown in formula (1):

[0008]

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

[0010] In the aggregation-induced emission material, R1 is a methoxy group and R2 is an ethyl group.

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

[0012] 6-bromo-2,3-dihydro-1H-flavone-4-carbaldehyde, Sodium dithiosulfate carbonate, palladium diacetate and an organic solvent are heated and stirred to obtain a first product;

[0013] Will The catalyst is mixed with the first product and subjected to reflux treatment to obtain a second product;

[0014] The second product is 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 the sodium carbonate dithiosulfate to the 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 emission material, wherein the temperature of the heating and stirring treatment is 110°C-140°C, the time of the heating and stirring treatment is 36h-54h; and / or the time of the reflux treatment is 20h-30h.

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

[0019] In the drug delivery system, the bacteria include one or more of Escherichia coli Nissle1917, transgenic attenuated Salmonella typhimurium strain VNP20009, bifidobacteria, and lactic acid bacteria.

[0020] A method for preparing a drug delivery system comprises the steps of:

[0021] mixing the aggregation-induced emission material with an organic solvent to obtain a mother solution;

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

[0023] The invention discloses an application of a drug delivery system in the preparation of anti-tumor drugs.

[0024] Beneficial effects: The present invention provides an aggregation-induced emission (AIE) material and a preparation method thereof, a drug delivery system and a preparation method and application thereof. AIE has unique characteristics as a photosensitizer. Compared with traditional photosensitizers with a planar conformation, AIE photosensitizers can use a larger proportion of absorbed energy to emit fluorescence and generate ROS. Therefore, the AIE provided by the present invention can efficiently generate ROS in a cellular environment; and it has photodynamic, photothermal therapy and fluorescence, photoacoustic, and photothermal imaging characteristics; in addition, the drug delivery system includes AIE materials and bacteria, and the tumor targeting ability of bacteria is not affected by the composition of the tumor genome, and can stimulate innate and adaptive anti-tumor immune responses in an environment of hypoxia, nutrient deficiency, and immunosuppression in tumor tissue. Using bacteria as a delivery carrier can expand the accumulation and penetration of drugs in tumor tissues, thereby optimizing the therapeutic effect, achieving specific killing of tumor tissues, and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of AIE materials. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The synthetic route for preparing the aggregation-induced emission material INX-PF6 in Example 1;

[0026] Figure 2 is the aggregation-induced emission material INX-PF6 in Example 1 1 H NMR spectrum;

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

[0028] Figure 4 is the aggregation-induced emission material INX-PF6 in Example 1 19 F NMR spectrum;

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

[0030] Figure 6 Confocal images of EcN after co-incubation with INX-PF6;

[0031] Figure 7 This is a data graph showing the change in fluorescence intensity after EcN loaded with INX-PF6 detected by flow cytometry;

[0032] Figure 8 Confocal imaging of EcN@INX-PF6 penetrating into CT26 tumor spheres;

[0033] Fig. 9 This is a data graph showing the changes in tumor volume of mice in each group after in vivo treatment;

[0034] Fig.10 These are images of in vitro tumor tissues of mice in each group after in vivo treatment. DETAILED DESCRIPTION

[0035] The present invention provides an aggregation-induced luminescence material and a preparation method thereof, a drug delivery system and a preparation method and application thereof. To make the purpose, technical solution and effect of the present invention clearer and more specific, the present invention is further described in detail below. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0036] It will be understood by those skilled in the art that, unless otherwise defined, all terms (including technical and scientific terms) used herein have the same meaning as generally understood by those skilled in the art in the art to which the present invention belongs. It should also be understood that terms such as those defined in general dictionaries should be understood to have meanings consistent with the meanings in the context of the prior art, and will not be interpreted with idealized or overly formal meanings unless specifically defined as herein.

[0037] Strong intermolecular interactions in traditional photosensitizers hinder the generation of ROS, thereby weakening the efficacy of photodynamic therapy. In addition, the limited diffusion range and short lifespan of ROS in solid tumors also affect the therapeutic effect. Therefore, it is imperative to innovate new photosensitizers with enhanced ROS generation capabilities to enhance photodynamic therapy of deep solid tumors.

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

[0039]

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

[0041] In this embodiment, aggregation-induced emission (AIE) materials have unique characteristics as photosensitizers. Compared with traditional photosensitizers with planar conformations, AIE photosensitizers can use a larger proportion of absorbed energy to emit fluorescence and generate ROS. Therefore, the AIE materials provided by the present invention can efficiently generate ROS in a cellular environment; and they have photodynamic, photothermal therapy, fluorescence, photoacoustic, and photothermal imaging characteristics; in addition, the drug delivery system includes AIE materials and bacteria, and the tumor targeting ability of bacteria is not affected by the composition of the tumor genome, and can stimulate innate and adaptive anti-tumor immune responses in an environment of hypoxia, nutrient deficiency, and immunosuppression in tumor tissue. Using bacteria as a delivery carrier can expand the accumulation and penetration of drugs in tumor tissues, thereby optimizing the therapeutic effect, achieving specific killing of tumor tissues, and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of AIE materials.

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

[0043] Specifically, when R1 is a methoxy group and R2 is an ethyl group, the chemical structure of the aggregation-induced emission material is The aggregation-induced luminescence material has photodynamic therapy, photothermal therapy, fluorescence, photoacoustic and photothermal imaging properties; the aggregation-induced luminescence material is then combined by physical mixing or covalent connection to obtain a bacterial-aggregation-induced luminescence material bacterial drug delivery system, and the bacteria-mediated bacterial drug delivery system can actively target hypoxic tumor tissues and achieve specific killing of tumor tissues, significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of the aggregation-induced luminescence material.

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

[0045] Step S10: 6-bromo-2,3-dihydro-1H-flavone-4-carbaldehyde, Sodium dithiosulfate carbonate, palladium diacetate and an organic solvent are heated and stirred to obtain a first product; the chemical structure of the first product is

[0046] Step S20: The catalyst is mixed with the first product and refluxed to obtain a second product; the chemical structure of the second product is

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

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

[0049] In some embodiments, the 6-bromo-2,3-dihydro-1H-flavone-4-carbaldehyde, The molar ratio of the sodium carbonate dithiosulfate and the 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 can obtain a first product with high purity and complete reaction after heating and stirring, which is beneficial to the next step of synthesis.

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

[0051] Specifically, the step S10 includes: 6-bromo-2,3-dihydro-1H-flavonoid-4-carboxaldehyde, Sodium carbonate dithiosulfate, palladium diacetate and an organic solvent are mixed under an inert atmosphere and then heated and stirred; then cooled to room temperature, the solvent is removed by vacuum rotary evaporation, the residue is redissolved with dichloromethane, and washed three times with saturated brine; the organic layer is dried over anhydrous Na2SO4, and then dried under reduced pressure; the crude product is purified by silica gel chromatography using n-hexane / dichloromethane as eluent to obtain an orange powdery solid compound, namely 6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavone-4-carbaldehyde.

[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 is The molar ratio of the two is (1-2):(1-2). After mixing according to the above molar ratio and subjecting to reflux treatment, a second product with higher purity and more complete reaction can be obtained.

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

[0056] Specifically, the step S20 includes: 6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavonoid-4-carboxaldehyde and A solution of the product is added in anhydrous ethanol, 50-100 μL of piperidine is added dropwise as a catalyst, and the mixture is refluxed for 24 hours under an inert atmosphere. After cooling to room temperature, the solvent is evaporated under reduced pressure. The residue is purified by silica gel chromatography and eluted with dichloromethane / methanol to obtain the second product, i.e., (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavon-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indol-1-ium iodide (INX-I).

[0057] In some embodiments, the temperature of the heating and stirring treatment is 110°C-140°C, the time of the heating and stirring treatment is 36h-54h; and / or the time of the reflux treatment is 20h-30h. Under this condition, the heating and stirring treatment can make the raw materials react quickly and shorten the reaction time.

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

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

[0060] In some embodiments, the step S30 comprises: 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 the dark red precipitate again, washing with water, and drying under reduced pressure. The residue is purified by silica gel chromatography and eluted with dichloromethane / methanol to obtain an aggregation-induced emission material, namely (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavon-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indol-1-ium hexafluorophosphate (V) salt (INX-PF6)

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

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

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

[0064] Specifically, a fluorescent material with aggregation-induced luminescence properties is first synthesized, and then combined with engineered bacteria under physical or chemical effects to prepare a bacterial drug delivery system of bacteria-aggregation-induced luminescence material. After tail vein injection, 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), transgenic attenuated Salmonella typhimurium strain VNP20009 (VNP), bifidobacteria, and lactic acid bacteria. Preferably, the bacteria is Escherichia coli Nissle1917 (EcN), which is well known for its antibacterial properties, intestinal microbiota regulation, and facultative anaerobicity.

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

[0067] Step S100: mixing the aggregation-induced emission material with an organic solvent to obtain a mother solution;

[0068] Step S200: mixing the mother solution with the bacterial solution, and incubating the mixture to obtain a drug delivery system.

[0069] In this embodiment, the drug delivery system prepared by the preparation method is combined by physical mixing or covalent connection to obtain a bacterial drug delivery system of bacteria-aggregation-induced luminescence material. The bacteria-mediated bacterial drug delivery system can actively target hypoxic tumor tissue and achieve specific killing of tumor tissue, which significantly enhances the multimodal tumor imaging and anti-tumor efficacy of aggregation-induced luminescence material.

[0070] In some embodiments, before step S100, the method further includes: reviving the low-temperature stored bacterial strains, culturing them in a culture medium required by the bacteria, collecting colonies in the growth phase, and obtaining a bacterial solution.

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

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

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

[0074] In this embodiment, the bacteria-mediated bacterial drug delivery system can actively target hypoxic tumor tissues, achieve specific killing of tumor tissues, and significantly enhance the multimodal tumor imaging and anti-tumor efficacy of aggregation-induced luminescence materials.

[0075] The following examples are further given to illustrate the present invention in detail. It should also be understood that the following examples are only used to further illustrate the present invention and cannot be understood as limiting the scope of protection of the present invention. Some non-essential improvements and adjustments made by those skilled in the art based on the above content of the present invention belong to 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, which specifically include the following:

[0078] The synthetic route of the 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 carbonate dithiosulfate (325 mg, 1 mmol), and palladium diacetate (20 mg, 0.6 mmol) were mixed in toluene under argon, and the reaction mixture was stirred at 120°C for 48 hours. After cooling to room temperature, the solvent was removed by vacuum rotary evaporation, and the residue was redissolved in dichloromethane and washed three times with saturated brine. The organic layer was dried over 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 obtain compound 1 as an orange powder, namely (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-flavon-4-carboxaldehyde (88 mg, 0.2 mmol) and 1-ethyl-2,3,3-trimethyl-3H-indol-1-ium iodide (63 mg, 0.2 mmol) was added dropwise with 50-100 μL of piperidine as catalyst in anhydrous ethanol, and the mixture was refluxed for 24 hours under nitrogen. After cooling to room temperature, 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 obtain compound (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavon-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indol-1-ium iodide (INX-I).

[0081] 3) INX-I (74 mg, 0.1 mmol) was dissolved in anhydrous ethanol (10 mL) and mixed with a saturated potassium hexafluorophosphate solution (10 mL). After stirring at room temperature for 1 hour, the anhydrous ethanol was evaporated with compressed air, and 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 obtain compound (E)-2-(2-(6-bis(4-methoxyphenyl)amino-2,3-dihydro-1H-flavon-4-yl)vinyl)-1-ethyl-3,3-dimethyl-3H-indol-1-ium hexafluorophosphate (V) salt (INX-PF6).

[0082] The structure of INX-PF6 molecule is 1 H NMR ( Figure 2 ), 13 C NMR ( Figure 3 ), 19 F NMR ( Figure 4 ) and high-resolution mass spectrometer ( Figure 5 ) for characterization verification.

[0083] 1 The results of H NMR 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 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] Preparation of bacteria-aggregation-induced luminescence material drug delivery system, the specific steps are as follows:

[0086] Take 10 μL of INX-PF6 stock solution (dissolved in DMSO, 1 mmol) and add it to 1 mL of bacterial concentration of 5×10 8 The CFUs of Escherichia coli EcN (Escherichia coli Nissle1917, EcN) were placed in a 37°C constant temperature shaker (180 rpm) for 20 minutes, then washed with PBS, and centrifuged twice at 6000 rpm for 3 minutes to obtain the bacteria-aggregation-induced luminescence material drug delivery system EcN@INX-PF6.

[0087] The applications of bacteria-aggregation-induced luminescence material drug delivery system are as follows:

[0088] Add the bacterial suspension of EcN (5 × 10 8 CFUs mL -1 ). After the bacteria were cultured in a shaker (37°C, 200 rpm) for 30 minutes, the supernatant was removed by centrifugation, a small amount of PBS was added and the cells were resuspended in PBS. 1 μL of the stained bacterial solution was transferred to a glass slide and then covered with a coverslip. Images were collected using a confocal laser scanning microscope, such as Figure 6 As shown. And the situation of bacterial surface loading molecules was detected by flow cytometry, such as Figure 7 As shown, INX-PF6 was still able to stably attach to bacteria after 24 h of incubation.

[0089] To construct a three-dimensional tumor sphere structure of CT26 tumor cells, agarose was dissolved in DMEM and sterilized at high temperature to obtain a 2% solution. The solution (60 μL) was used to coat the bottom of a 96-well plate. CT26 cells (2×10 3 After forming three-dimensional tumor spheroids of about 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 collected using a confocal laser scanning microscope to verify the penetration depth of drug-loaded bacteria, such as Figure 8 As shown, the results show that EcN@INX-PF6 has good penetration performance.

[0090] Establishment of CT26 tumor-bearing mouse model, tumor volume ≈ 150mm 3 The tumor-bearing mice were randomly divided into 7 groups for different treatments (5 mice in each group). Different treatment agents were injected into the tail vein of the tumor-bearing mice. The changes in the tumor growth volume of the mice were observed. The results are as follows Fig. 9 and Fig.10 As shown, the results showed that EcN@INX-PF6 had strong anti-tumor killing ability.

[0091] In summary, the present invention provides an aggregation-induced luminescence material and a preparation method thereof, a drug delivery system and a preparation method and application thereof. The aggregation-induced luminescence material has unique characteristics. Compared with traditional photosensitizers with a planar conformation, AIE photosensitizers can use a larger proportion of absorbed energy to emit fluorescence and generate ROS. Therefore, the new AIE provided by the present invention can efficiently generate ROS in a cellular environment; and it has photodynamic, photothermal therapy and fluorescence, photoacoustic, and photothermal imaging characteristics; in addition, the drug delivery system includes aggregation-induced luminescence materials and bacteria, and the tumor targeting ability of bacteria is not affected by the composition of the tumor genome, and can stimulate innate and adaptive anti-tumor immune responses in an environment of hypoxia, nutrient deficiency and immunosuppression in tumor tissue. Using bacteria as a delivery carrier can expand the accumulation and penetration of drugs in tumor tissues, thereby optimizing the therapeutic effect, achieving specific killing of tumor tissues, and significantly enhancing the multimodal tumor imaging and anti-tumor efficacy of aggregation-induced luminescence materials.

[0092] It should be understood that the application of the present invention is not limited to the above examples. For ordinary technicians in this field, improvements or changes can be made based on the above description. All these improvements and changes should fall within the scope of protection of the claims attached to the present invention.

Claims

1. An aggregation-induced emission material, characterized in that: The molecular structure of the aggregation-induced emission material is shown in formula (1): Wherein, R1 is selected from one of hydrogen, methyl and methoxy; R2 is selected from one of methyl and ethyl.

2. The aggregation-induced emission material according to claim 1, characterized in that: R1 is methoxy; R2 is ethyl.

3. A method for preparing an aggregation-induced emission material, characterized in that: Includes steps: 6-bromo-2,3-dihydro-1H-flavone-4-carbaldehyde, Sodium dithiosulfate carbonate, palladium diacetate and an organic solvent are heated and stirred to obtain a first product; Will The catalyst is mixed with the first product and subjected to reflux treatment to obtain a second product; The second product is mixed with a saturated potassium hexafluorophosphate solution to obtain an aggregation-induced emission material.

4. The method for preparing the aggregation-induced emission material according to claim 3, characterized in that: The 6-bromo-2,3-dihydro-1H-flavone-4-carboxaldehyde, The molar ratio of the sodium carbonate dithiosulfate to the palladium diacetate is (0.5-1.5):(0.8-1.5):(0.8-1.5):(0.4-1).

5. The method for preparing the aggregation-induced emission material according to claim 3, characterized in that: The first product The molar ratio is (1-2):(1-2).

6. The method for preparing the aggregation-induced emission material according to claim 3, characterized in that: The temperature of the heating and stirring treatment is 110° C.-140° C., the time of the heating and stirring treatment is 36 h-54 h; and / or the time of the reflux treatment is 20 h-30 h.

7. A drug delivery system, characterized in that It comprises bacteria and the aggregation-induced emission material according to any one of claims 1 to 2; the aggregation-induced emission material is loaded on the surface of the bacteria.

8. The drug delivery system according to claim 7, characterized in that The bacteria include one or more of Escherichia coli Nissle1917, transgenic attenuated Salmonella typhimurium strain VNP20009, bifidobacteria, and lactic acid bacteria.

9. A method for preparing a drug delivery system according to any one of claims 7 to 8, characterized in that: Includes steps: mixing the aggregation-induced emission material with an organic solvent to obtain a mother solution; The mother solution is mixed with a bacterial solution and incubated to obtain a drug delivery system.

10. Use of the drug delivery system according to any one of claims 7 to 8 in the preparation of anti-tumor drugs.

Images