Near-infrared emission aggregation-induced emission organic photo-thermal molecule as well as preparation method and application thereof

Abstract

The invention discloses a near-infrared emission aggregation-induced emission organic photo-thermal molecule and a preparation method and application thereof, and belongs to the technical field of biomedicine, and the structure is shown as a formula I: # imgabs0 # (I), wherein R1, R2, R3, R4, R5 and R6 are independently selected from groups containing alkyl chains. The invention also provides a preparation method of the organic photo-thermal molecule and application of the organic photo-thermal molecule in preparation of multifunctional protein nanoparticles with near-infrared aggregation-induced emission. The near-infrared emission aggregation-induced emission organic photo-thermal molecule provided by the invention shows excellent near-infrared imaging effect and photo-thermal capability under laser irradiation, and the obtained near-infrared aggregation-induced emission multifunctional protein nano particle has excellent photo-thermal effect and imaging resolution, has excellent targeting to brain tumor, and has good application prospects in blood brain barrier crossing, brain tumor targeting and brain tumor targeting. And tumor killing, photothermal and ferroptosis combined treatment has practical medical significance, and is expected to be widely applied to the aspects of intelligent drug delivery and nano medical treatment.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical technology, and in particular relates to an organic photothermal molecule that emits near-infrared aggregation-induced luminescence, and a preparation method and application thereof. Background Art

[0002] Glioblastoma multiforme is a common and malignant brain tumor that is difficult to treat. Although surgery, chemotherapy, and radiotherapy are conventional treatments, they are usually not completely effective, and the patient's prognosis is poor. In recent years, near-infrared light diagnosis and treatment nanomedicines have attracted widespread attention in the early diagnosis and precision treatment of tumors, and have made significant research progress. The advantage of near-infrared light diagnosis and treatment lies in its excellent near-infrared imaging effect and efficient photothermal killing ability, which makes it an important technology for non-invasive tumor treatment. However, for brain tumors, the existence of the blood-brain barrier significantly limits the effective delivery and application of traditional light diagnosis and treatment nanomedicines. The current common blood-brain barrier crossing scheme is to modify the material, such as adding apolipoprotein E ( Fig.11 ) or Angiopep-2 ( Fig.12 ) etc. However, such modifications are often difficult to operate, with a low modification ratio and cumbersome steps.

[0003] Studies have shown that transferrin has good cross-over ability due to its high expression of receptors on the blood-brain barrier, thus providing a new solution for the targeted treatment of brain tumors. Combining transferrin with near-infrared light therapy not only avoids tedious modifications, but also enables accurate diagnosis and treatment of brain tumors. However, traditional near-infrared light therapy materials, such as indocyanine green (ICG), have poor imaging effects and high signal-to-noise ratios (such as fluorescence quenching caused by aggregation effects) after binding to proteins. Fig.13 As shown in the figure, it is difficult to use it as a reliable imaging material. In addition, the light / thermal stability of near-infrared light diagnostic and therapeutic materials is also a challenge.

[0004] It can be seen that how to provide a near-infrared second-zone optical diagnostic and therapeutic material with excellent performance and be able to be effectively applied in the transferrin nanodrug system has become an urgent need to improve the accuracy of brain tumor diagnosis and treatment. Summary of the invention

[0005] In order to solve the above technical problems, the present invention proposes an organic photothermal molecule that emits near-infrared aggregation-induced luminescence, and a preparation method and application thereof.

[0006] To achieve the above object, the present invention provides the following technical solutions:

[0007] A near-infrared emitting aggregation-induced luminescence organic photothermal molecule, the structure of which is shown in Formula I:

[0008]

[0009] (I);

[0010] Wherein, R1, R2, R3, R4, R5 and R6 are independently selected from groups containing alkyl chains.

[0011] Preferably, the alkyl chain is C1-C 20 Straight chain, C1-C 20 The branched chain, C1-C 20 A cyclic alkyl chain, an alkyl chain in which the carbon atoms are replaced by one or more of oxygen atoms, alkenyl groups, alkynyl groups, aryl groups, carbonyl groups, hydroxyl groups, amino groups, carboxyl groups, cyano groups, nitro groups and ester groups, and an alkyl chain in which the hydrogen atoms are replaced by one or more of fluorine atoms, chlorine atoms, bromine atoms and iodine atoms.

[0012] More preferably, the structure of the near-infrared emitting aggregation-induced luminescence organic photothermal molecule is as follows:

[0013]

[0014] Beneficial effects: The present invention provides a novel near-infrared emitting aggregation-induced luminescence organic photothermal molecule with an electron donor-electron acceptor structure, with thionaphthalene tetracarboxylic acid diimide having a long alkyl chain as the electron acceptor group, and triphenylamine as the molecular rotor and electron donor unit. Under laser irradiation, it can emit near-infrared zone II light and generate heat, and has good photostability. In addition, a long alkyl chain is introduced into triphenylamine to regulate the stacking mode of the molecule in the aggregated state. Specifically, the large conjugated main chain of the electron acceptor group has quinone-like characteristics to allow large electron delocalization to exist, forming a lower band gap, and its large planar structure is conducive to improving long-wavelength absorption capacity and high molar absorptivity. The long alkyl chain on the electron acceptor avoids excessive stacking between molecules, avoids the shortcomings of poor imaging effect, high signal-to-noise ratio, poor photostability, etc. caused by aggregation-induced quenching. The electron donor is electron-rich triphenylamine. This design helps to form an efficient twisted intramolecular charge transfer effect, which helps to improve the near-infrared luminescence performance and enhance the photothermal capacity of the material. The long alkyl chain in triphenylamine brings greater freedom of rotor rotation to the molecule, and the intermolecular interaction is greatly suppressed in the aggregated state, ensuring its efficient and stable photothermal performance. Therefore, the near-infrared emitting aggregation-induced luminescence organic photothermal molecule provided by the present invention can achieve efficient near-infrared luminescence performance and excellent photothermal performance, providing material support for the subsequent construction of functional protein nanoparticles.

[0015] A method for preparing near-infrared emitting aggregation-induced luminescence organic photothermal molecules comprises the following steps:

[0016] (1) reacting sodium hydride and 4-nitrophenylacetonitrile in an ice bath, then adding carbon disulfide and continuing the reaction at room temperature, then adding 4,5,9,10-tetrabromo-2,7-bis(2-octyldodecyl)benzo[lmn][3,8]phenanthrole-1,3,6,8(2H,7H)-tetraone and continuing the reaction by stirring, and performing post-treatment after the reaction to obtain compound 1;

[0017] (2) dissolving the compound 1 and ammonium chloride in an organic solvent for reflux reaction, then adding iron powder to continue the reaction, and performing post-treatment after the reaction to obtain compound 2;

[0018] (3) The compound 2, halogenated aromatic hydrocarbon or halogenated aromatic ether compound or its derivative, depolymerizing agent, ligand and catalyst are dissolved in an organic solvent and mixed to carry out reflux reaction. After the reaction is completed, post-treatment is carried out to obtain the near-infrared emitting aggregation-induced luminescence organic photothermal molecule.

[0019] Preferably, the halogenated aromatic hydrocarbon or halogenated aromatic ether compound or its derivative in step (3) includes one or more of p-bromoanisole, 4-n-hexyloxybromobenzene, 1-bromo-4-(octadecyloxy)benzene, 1-bromo-4-hexylbenzene, and 1-bromo-4-(perfluorohexyl)benzene.

[0020] Preferably, the deprotonating agent is sodium tert-butoxide or potassium tert-butoxide; and / or,

[0021] The ligand is 2-dicyclohexylphosphine-2,4,6-triisopropylbiphenyl or 2-dicyclohexylphosphine-2',6'-diisopropoxy-1,1'-biphenyl; and / or,

[0022] The catalyst is tris(dibenzylideneacetone)dipalladium(0).

[0023] Application of a near-infrared emitting aggregation-induced luminescence organic photothermal molecule in the preparation of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles.

[0024] A method for preparing near-infrared aggregation-induced luminescence multifunctional protein nanoparticles comprises the following steps:

[0025] The above-mentioned near-infrared emitting aggregation-induced luminescence organic photothermal molecule solution is mixed with the transferrin solution to carry out a self-assembly reaction, and after the reaction is completed, the solution is washed, centrifuged, filtered and collected to obtain the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles.

[0026] Preferably, the transferrin is one or more of holo-transferrin, functionally modified transferrin and controllably modified transferrin.

[0027] Preferably, the concentration of the near-infrared emitting aggregation-induced luminescence organic photothermal molecule solution is 1 mg / mL; and / or,

[0028] The concentration of the transferrin solution is 3 mg / mL; and / or,

[0029] The volume ratio / mass ratio of the near-infrared emitting aggregation-induced luminescence organic photothermal molecule solution to the transferrin solution is 1:3 (w / w).

[0030] Beneficial effects: All-iron transferrin is a protein rich in iron, which is mainly used to carry iron absorbed by the digestive tract and iron released by red blood cell degradation. Its embedded nanoparticles have excellent properties such as high stability, biocompatibility, and biodegradability, and have the ability to highly bind to small organic molecules. Transferrin is necessary for ferroptosis, which can transfer Fe 3+ Transported to the endosome, where Fe 3+ Further reduced to Fe 2+ Finally, under the mediation of divalent metal transporter 1, Fe 2+ Released from endosomes into the unstable iron pool in the cytoplasm. Abnormal expression or dysfunction of these iron-related proteins will increase the intracellular iron ion concentration due to metabolic imbalance, thereby triggering the Fenton reaction, leading to lipid peroxidation (LPO) and ferroptosis of brain tumor cells. Therefore, full transferrin, as a potential nanocarrier, can effectively cross the blood-brain barrier and simultaneously induce ferroptosis of brain tumor cells, ultimately leading to ferroptosis of cells. The near-infrared emitting aggregation-induced luminescence organic photothermal molecules provided by the present invention have the characteristics of near-infrared second zone luminescence, heat generation, and good photostability. Subsequently, full iron transferrin is used as a biological functional carrier,

[0031] The present invention mixes near-infrared emitting aggregation-induced luminescence organic photothermal molecules with all-iron transferrin in a fixed ratio, and uniformly loads the above-mentioned organic photothermal molecules on the protein to form near-infrared molecule-all-iron transferrin composite nanoparticles. There are strong hydrogen bonds and van der Waals interactions between the organic photothermal molecules and transferrin. This ingenious and unique interaction makes the performance of the protein nanoparticles stable. The presence of all-iron transferrin allows the protein to efficiently cross the blood-brain barrier and promote the occurrence of ferroptosis in the tumor site. The near-infrared emitting aggregation-induced luminescence organic photothermal molecules show excellent near-infrared imaging effect and photothermal ability under laser irradiation. The obtained near-infrared aggregation-induced luminescence multifunctional protein nanoparticles have excellent photothermal effect and imaging resolution, have excellent targeting for brain tumors, and have practical medical significance in crossing the blood-brain barrier, tumor killing, and combined photothermal and ferroptosis treatment, and are expected to be widely used in smart drug delivery and nanomedicine.

[0032] The multifunctional protein nanoparticles with near-infrared aggregation-induced luminescence are prepared by the above-mentioned preparation method.

[0033] Beneficial effects: The product structure provided by the present invention comprises a large planar benzene ring structure with high absorption and a twisted skeleton for enhancing red-shifted emission. The structural modification of D-π-A and the introduction of phenyl groups improve the brightness of such aggregation-induced luminescent materials, and the introduction of the terminal alkyl chain of the electron donor balances the luminescence and heat generation capabilities of the material, so that this new material has good luminescence imaging performance. The present invention is a near-infrared nano-diagnosis and treatment system that integrates crossing the blood-brain barrier, targeting, inducing ferroptosis, fluorescence imaging, and photothermal therapy, and has important scientific research and clinical application prospects in the fields of biotechnology and medical technology.

[0034] Preferably, the protein carrier is spherical, polyhedral or aggregated.

[0035] The size of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles is 80-200 nm.

[0036] Beneficial effects: Protein particles with a particle size of less than 200 nanometers can enhance the permeability and retention (EPR) effect of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles, thereby improving the effect of targeted enrichment of tumors.

[0037] Application of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles in the preparation of drugs for diagnosing or treating brain tumors.

[0038] Compared with the prior art, the present invention has the following advantages and technical effects:

[0039] The method for synthesizing near-infrared emitting aggregation-induced luminescence organic photothermal molecules provided by the present invention has a simple and efficient synthesis process, and the obtained new organic molecules have luminescence performance in the near-infrared second region and excellent photothermal performance. At the same time, the preparation process of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles in the present invention is simple and has excellent performance. It can effectively cross the blood-brain barrier and be enriched in the brain tumor site, realizing the visualization of brain tumors and photothermal therapy and killing. There are strong hydrogen bonds and van der Waals interactions between the molecular structure of the organic material and the full-iron transferrin. This ingenious and unique interaction makes the performance of the protein nanoparticles stable. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] The drawings constituting a part of the present application are used to provide a further understanding of the present application. The illustrative embodiments and descriptions of the present application are used to explain the present application and do not constitute an improper limitation on the present application. In the drawings:

[0041] Figure 1This is the fluorescence spectrum of the near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-18 obtained in Example 3 tested in a mixed solvent of water and tetrahydrofuran, and the molecule emits in the near-infrared region;

[0042] Figure 2 The molecular docking and molecular dynamics simulation results of the interaction between the near-infrared organic molecule and the full-iron transferrin in the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4;

[0043] Figure 3 This is a scanning electron microscope image of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4;

[0044] Figure 4 This is a transmission scanning electron micrograph of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4;

[0045] Figure 5 The particle size distribution of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4;

[0046] Figure 6 The near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4 were simulated to cross the blood-brain barrier in vitro;

[0047] Figure 7 The multifunctional protein nanoparticles with near-infrared aggregation-induced luminescence obtained in Example 4 induce ferroptosis in U87 cells;

[0048] Figure 8 The multifunctional protein nanoparticles with near-infrared aggregation-induced luminescence obtained in Example 4 are used for tumor targeting imaging;

[0049] Fig. 9 The photothermal treatment / imaging effect of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4 on brain tumors;

[0050] Fig.10 The combined therapeutic effect of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4 on brain tumors;

[0051] Fig.11 Modification of nanoparticles with apolipoprotein E for crossing the blood-brain barrier;

[0052] Fig.12 Modify nanoparticles with the delivery peptide Angiopep-2 for crossing the blood-brain barrier;

[0053] Fig.13 This is the imaging effect after traditional near-infrared light diagnostic materials are combined with proteins. DETAILED DESCRIPTION

[0054] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0055] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the present invention is further described in detail below with reference to the accompanying drawings and specific embodiments.

[0056] Unless otherwise specified, the room temperature or normal temperature in the embodiments of the present invention refers to 25±3°C.

[0057] Unless otherwise specified, the raw materials in the examples of the present invention were not purchased from commercial sources;

[0058] The re-evaporated N,N-dimethylformamide was purchased from Anage Chemical Company.

[0059] Example 1

[0060] The preparation method of compound 1, the synthetic route is as follows:

[0061]

[0062] The specific steps include:

[0063] Under nitrogen protection, sodium hydride (210 mg, 5.24 mmol) was added to 10 mL of re-evaporated N,N-dimethylformamide, and placed in an ice bath. 4-Nitrophenylacetonitrile (425 mg, 2.62 mmol) was added and stirred for 30 minutes. Then carbon disulfide (300 mg, 3.93 mmol) was added and the reaction temperature was raised to room temperature for 2 hours. When the reaction solution changed from colorless to light green and gradually to brown, 4,5,9,10-tetrabromo-2,7-bis(2-octyldodecyl)benzo[lmn][3,8]phenanthrolene-1,3,6,8(2H,7H)-tetraone (300 mg, 0.262 mmol) was added at once and stirred at room temperature for 1 hour until the reaction solution turned dark purple. Then, 20 mL of physiological saline was added to terminate the reaction, and then ethyl acetate (EA) was used for extraction. The organic phase was collected, dried over anhydrous sodium sulfate, and evaporated to remove the solvent. The residue obtained after evaporation was separated by column chromatography using dichloromethane and petroleum ether (4:1) as eluent to obtain dark green solid compound 1 with a yield of 52%.

[0064] 1H NMR (500 MHz, CDCl3), δ (ppm): 8.41 (d, J = 5.0 Hz, 4H), 7.92 (d, J= 10.0 Hz, 4H), 4.20 – 4.10 (m, 4H), 1.99 (s, 2H), 1.27 (t, 64H), 0.85 (m,12H). 13 C NMR (126 MHz, CDCl3) δ [ppm]: 162.07, 147.84, 146.50, 145.96,139.18, 128.27, 124.79, 116.03, 99.68, 46.22, 36.42, 31.94, 31.90, 30.08,29.70, 29.66, 29.62, 29.57, 29.38, 29.35, 22.70, 22.69, 14.14. MS (MALDI-TOF)[m / z]: calcd for C 72 H 90 N6O8S4, 1295.5737; found, 1295.5784.

[0065] Example 2

[0066] The preparation method of compound 2, the synthetic route is as follows:

[0067]

[0068] The specific steps include:

[0069] Under nitrogen protection, 200 mg of the compound 1 obtained in Example 1 and 70 mg of ammonium chloride were dissolved in a mixed solution of 15 mL of tetrahydrofuran and 20 mL of ethanol, and refluxed at 80°C for 10 minutes. Subsequently, 50 mg of iron powder was added and the reaction was continued for 40 minutes. After the reaction was completed, the mixture was cooled to room temperature, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography using a mixed solution of dichloromethane / methanol (volume ratio 10:1) as an eluent to obtain a green powder product compound 2 with a yield of 76%.

[0070] 1 H NMR (500 MHz, CDCl3) δ [ppm]: 7.54 – 7.39 (d, 4H), 6.86 – 6.69 (d,4H), 4.23 – 4.10 (d, 4H), 3.99 (s, 2H), 2.00 (s, 2H), 1.27 (m, 64H), 0.85(dt, 12H).13 C NMR (126 MHz, CDCl3) δ [ppm]: 13 C NMR (126 MHz, THF) δ 162.29,150.86, 150.04, 147.36, 144.89, 127.99, 125.22, 120.88, 117.34, 115.16,114.12, 101.80, 45.68, 36.45, 32.09, 30.47, 29.90, 26.41, 22.00, 13.54. MS(MALDI-TOF) [m / z]: calcd for C 72 H 94 N6O4S4, 1235.6253; found, 1235.6283.

[0071] Example 3

[0072] A method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-18, the synthesis route is as follows:

[0073]

[0074] The specific steps include:

[0075] Under nitrogen protection, compound 2 obtained in Example 2 (100 mg, 0.08 mmol), 1-bromo-4-(octadecyloxy)benzene (170 mg, 0.40 mmol), sodium tert-butoxide (53.6 mg, 0.56 mmol), 2-dicyclohexylphosphine-2,4,6-triisopropylbiphenyl (18.2 mg, 0.2 mmol), tris(dibenzylideneacetone)dipalladium (0) (9.2 mg, 0.1 mmol) were dissolved in toluene and refluxed at 120°C for 12 hours. After the reaction was completed, the mixture was cooled to room temperature, extracted with dichloromethane, and washed three times with brine. Subsequently, the mixture was dried over anhydrous sodium sulfate, the solvent was removed under reduced pressure, and the residue was purified by silica gel column chromatography using dichloromethane and petroleum ether (2:1) as eluents to obtain a green powder product, near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-18, with a yield of 75%.

[0076] 1H NMR (500 MHz, CDCl3) δ [ppm]: 7.49 – 7.44 (d, 4H), 7.18 – 7.12 (d,8H), 6.99 – 6.94 (d, 4H), 6.94 – 6.86 (d, 8H), 4.24 – 4.16 (d, 4H), 4.00 –3.96 (m, 8H), 2.05 (s, 2H), 1.27 (m, 192H), 0.85 (dt, 24H). 13 C NMR (126 MHz, CDCl3) δ [ppm]: 162.33, 156.40, 149.46, 139.37, 133.98, 128.05, 127.61,124.01, 118.49, 115.50, 115.19, 115.10, 77.29, 77.04, 76.78, 68.45, 68.31,34.89, 31.96, 31.90, 31.46, 30.21, 30.13, 29.74, 29.69, 29.66, 29.50, 29.40,29.35, 29.00, 26.44, 26.15, 25.95, 22.72, 14.15. MS (MALDI-TOF) [m / z]: calcdfor C 168 H 254 N6O8S4, 2613.8603; found, 2613.7075.

[0077] Figure 1 This is the fluorescence spectrum of the organic photothermal molecule NDA-18, which emits near-infrared aggregation-induced emission, tested in a mixed solvent of water and tetrahydrofuran. The molecule emits in the near-infrared region. It can be seen that with the increase of water content, the fluorescence intensity decreases and then increases, showing an obvious TICT-aggregation-induced emission phenomenon.

[0078] Example 4

[0079] A method for preparing near-infrared aggregation-induced luminescence multifunctional protein nanoparticles comprises the following steps:

[0080] 12 g of all-iron transferrin was dissolved in 4 mL of ultrapure water to obtain an all-iron transferrin solution, and 4 g of the near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-18 obtained in Example 3 was dissolved in 4 mL of tetrahydrofuran to form a solution. Then the above all-iron transferrin solution and the near-infrared emitting aggregation-induced luminescence organic photothermal molecule solution were mixed evenly at room temperature and reacted for 3 hours. After the reaction was completed, the mixture was washed, centrifuged, filtered and collected to obtain near-infrared aggregation-induced luminescence multifunctional protein nanoparticles.

[0081] Figure 2 The molecular docking and molecular dynamics simulation results of the interaction between near-infrared organic molecules and all-iron transferrin in the obtained infrared aggregation-induced luminescence multifunctional protein nanoparticles show that all-iron transferrin can bind to the near-infrared emitting aggregation-induced luminescence organic photothermal molecules at characteristic sites with a binding energy of -5.56 kcal / mol. This proves that there is a strong hydrogen bond and van der Waals force interaction between the two, which makes the structure and performance of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles stable.

[0082] Figure 3 This is a scanning electron microscope image of the obtained near-infrared aggregation-induced luminescence multifunctional protein nanoparticles. It can be seen that the particle size and morphology of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles are uniform.

[0083] Figure 4 This is a transmission scanning electron micrograph of the obtained near-infrared aggregation-induced luminescence multifunctional protein nanoparticles. It can be seen that the particle size of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles is about 140 nanometers and the morphology is uniform.

[0084] Figure 5 : is the particle size distribution of the obtained near-infrared aggregation-induced luminescence multifunctional protein nanoparticles. It can be seen that the particle size of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles is about 140 nanometers, and the size distribution is uniform.

[0085] Example 5

[0086] A method for preparing an organic photothermal molecule NDA-1 that emits near-infrared aggregation-induced luminescence is different from that in Example 3 in that 1-bromo-4-(octadecyloxy)benzene is replaced with p-bromoanisole of equal molar mass. The other process steps and parameters are the same as those in Example 3. The yield is 46%. The synthetic route is as follows:

[0087]

[0088] 1H NMR (500 MHz, CDCl3) δ [ppm]: 7.54 – 7.39 (d, 4H), 6.86 – 6.69 (d,4H), 4.23 – 4.10 (d, 4H), 3.99 (s, 2H), 2.00 (s, 2H), 1.27 (m, 64H), 0.85(dt, 12H).

[0089] Example 6

[0090] A method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-6, which is different from Example 3 in that 1-bromo-4-(octadecyloxy)benzene is replaced with 4-n-hexyloxybromobenzene of equal molar mass. The other process steps and parameters are the same as those in Example 3. The yield is 52%. The synthesis route is as follows:

[0091]

[0092] 1 H NMR (500 MHz, CDCl3) δ [ppm]: 7.49 – 7.44 (d, 4H), 7.18 – 7.12 (d,8H), 6.99 – 6.94 (d, 4H), 6.94 – 6.86 (d, 8H), 4.24 – 4.16 (d, 4H), 4.00 –3.96 (m, 8H), 2.05 (s, 2H), 1.27 (m, 64H), 0.85 (dt, 12H). 13 C NMR (126 MHz, CDCl3) δ [ppm]: 162.19, 156.44, 149.39, 139.32, 129.46, 127.95, 127.68,127.34, 118.37, 115.52, 115.31, 114.99, 77.32, 77.07, 76.82, 68.55, 68.30,31.96, 31.94, 31.67, 31.51, 31.40, 30.20, 29.75, 29.69, 29.40, 26.46, 25.83,25.79, 22.73, 22.68, 22.66, 14.16, 14.09. MS (MALDI-TOF) [m / z]: calcd forC 120 H 158 N6O8S4, 1940.1058; found, 1940.1410.

[0093] Example 7

[0094] The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-6A is different from that in Example 3 in that 1-bromo-4-(octadecyloxy)benzene is replaced with 1-bromo-4-hexylbenzene of equal molar mass. The other process steps and parameters are the same as those in Example 3. The yield is 61%. The synthesis route is as follows:

[0095]

[0096] 1 H NMR (500 MHz, CDCl3) δ [ppm]: 7.53 – 7.02 (m, 24H), 4.19 (s, 4H), 2.60 (s, 8H), 2.03 (s, 2H), 1.27 (m, 64H), 0.84 (d, 12H). 13 C NMR (126 MHz, CDCl3) δ [ppm]: 144.24, 143.38, 139.19, 130.29, 129.49, 129.02, 128.04,126.56, 125.80, 125.16, 124.39, 121.26, 120.23, 35.53, 34.05, 31.78, 31.52,31.10, 30.15, 29.68, 29.38, 29.15, 25.52, 22.71, 22.67, 22.27, 14.15. MS(MALDI-TOF) [m / z]: calcd for C 120 H 158 N6O4S4, 1876.1261; found, 1876.1603.

[0097] Example 8

[0098] A method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-6F, which is different from Example 3 in that 1-bromo-4-(octadecyloxy)benzene is replaced with 1-bromo-4-(perfluorohexyl)benzene of equal molar mass. The other process steps and parameters are the same as those in Example 3. The yield is 49%. The synthesis route is as follows:

[0099]

[0100] 1H NMR (500 MHz, CDCl3) δ [ppm]: 7.70-7.50 (m, 12H), 7.35 – 7.22 (m, 12H), 4.21 (s, 4H), 2.07 (s, 2H), 1.22 (m, 64H), 0.85 (s, 12H). 19 F NMR (500MHz, CDCl3) δ [ppm]: -81.30 (s, 3F), -110.70 (s, 2F), -122.02 (s, 2F), -122.13 (S, 2F), -123.35 (S, 2F), -126.67 (S, 2F). 13 C NMR (126 MHz, CDCl3) δ[ppm]: 162.23, 162.07, 149.51, 146.85, 144.85, 139.34, 128.74, 128.56,125.16, 124.89, 124.13, 116.95, 115.76, 114.10, 101.01, 46.25, 36.37, 34.90,33.87, 31.97, 31.92, 31.87, 31.82, 31.65, 31.54, 31.47, 30.33, 30.21, 30.14,29.74, 29.71, 29.64, 29.58, 29.41, 29.38, 29.34, 29.30, 29.21, 28.99, 26.42,24.77, 22.74, 22.71, 22.69, 22.67, 22.62, 14.17, 14.13, 14.08, 14.04, 14.00,11.19. MS (MALDI-TOF) [m / z]: calcd for C 120 H 106 F 52 N6O4S4, 2811.6362; found, 2811.3159.

[0101] Examples 9-12

[0102] A method for preparing near-infrared aggregation-induced luminescence multifunctional protein nanoparticles, which is different from Example 4 only in that the near-infrared emitting aggregation-induced luminescence organic photothermal molecule NDA-18 obtained in Example 3 is replaced with an equal amount of near-infrared emitting aggregation-induced luminescence organic photothermal molecules obtained in Example 5, Example 6, Example 7 or Example 8. The other process steps and parameters are the same as those in Example 4.

[0103] Technical effect:

[0104] 1. In vitro simulation experiment of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles crossing the blood-brain barrier.

[0105] BBB permeability was assessed in an in vitro dialysis chamber model. The upper chamber was seeded with a layer of endothelial cells, bEnd.3, to simulate the blood-brain barrier, while the lower chamber was seeded with U87 cells to simulate glioblastoma tumors ( Figure 6 (Part A).

[0106] Figure 6 Part B is a simulation of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4 crossing the blood-brain barrier in vitro. It can be seen that the FITC-labeled near-infrared aggregation-induced luminescence multifunctional protein nanoparticles (N18Tf NPs in the figure) show a strong signal in the cytoplasm of U87 cells in the lower chamber, proving that they have good in vitro blood-brain barrier penetrability and cellular uptake ability.

[0107] 2. Near-infrared aggregation-induced luminescence multifunctional protein nanoparticles induce ferroptosis in U87 cells.

[0108] In order to verify that the particles induce ferroptosis in U87 cells, the following Western blot experiment was designed:

[0109] First, U87 cells were divided into two groups. The experimental group was treated with the multifunctional protein nanoparticles prepared in Example 4, and the control group was not treated. After that, a specific primary antibody against the ferroptosis-related marker protein GPX was used for incubation. The cells were detected by a secondary antibody (HRP labeled) and developed with a chemiluminescent substrate. The expression differences of ferroptosis-related proteins in the experimental group and the control group were analyzed. The expression of GPX in the experimental group decreased, indicating that particle treatment can induce ferroptosis in U87 cells.

[0110] Figure 7 The near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4 induced ferroptosis in U87 cells. It can be seen that after the addition of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles (corresponding to the two columns of strips on the right), the GPX4 protein was significantly reduced, indicating that the cells may be in oxidative stress and iron-related death pathways.

[0111] 3. Near-infrared aggregation-induced luminescence multifunctional protein nanoparticles tumor-targeted imaging, the results are as follows Figure 8 As shown, it can be seen that the fluorescence signal of the nanoparticles at the tumor site gradually increased and reached a peak at 12 hours, indicating effective accumulation and targeting of the tumor site.

[0112] Fig. 9The photothermal treatment / imaging effect of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles on brain tumors obtained in Example 4. It can be seen that after adding near-infrared aggregation-induced luminescence multifunctional protein nanoparticles, the maximum temperature of the brain tumor area reached 42 degrees Celsius within 5 minutes, which is significantly different from the ambient temperature, demonstrating the precise and efficient photothermal killing ability of the brain tumor area.

[0113] Fig.10 The combined therapeutic effect of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles obtained in Example 4 on brain tumors specifically includes the following steps:

[0114] The multifunctional protein nanoparticles with near-infrared aggregation-induced luminescence not only have the ability to induce tumor ferroptosis efficiently. After turning on the 808 laser, the efficient photothermal capacity ensures the smooth progress of photothermal therapy, thus realizing the combined treatment mode of ferroptosis / photothermal killing of brain tumors. Fig.10 It can be seen that the nanoparticle treatment group exhibited a longer survival rate during the treatment period, indicating the potential of nanoparticles for combined treatment of brain tumors.

[0115] The above are only preferred specific implementations of the present application, but the protection scope of the present application is not limited thereto. Any changes or substitutions that can be easily thought of by a person skilled in the art within the technical scope disclosed in the present application should be included in the protection scope of the present application. Therefore, the protection scope of the present application should be based on the protection scope of the claims.

Claims

1. An organic photothermal molecule that emits near-infrared aggregation-induced luminescence, characterized in that: The structure is shown in Formula I: （I）； Wherein, R1, R2, R3, R4, R5 and R6 are independently selected from groups containing alkyl chains.

2. The near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 1, characterized in that: The alkyl chain is C1-C 20 Straight chain, C1-C 20 The branched chain, C1-C 20 A cyclic alkyl chain, an alkyl chain in which the carbon atoms are replaced by one or more of oxygen atoms, alkenyl groups, alkynyl groups, aryl groups, carbonyl groups, hydroxyl groups, amino groups, carboxyl groups, cyano groups, nitro groups and ester groups, and an alkyl chain in which the hydrogen atoms are replaced by one or more of fluorine atoms, chlorine atoms, bromine atoms and iodine atoms.

3. The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 1 or 2, characterized in that: The following steps are involved: (1) reacting sodium hydride and 4-nitrophenylacetonitrile in an ice bath, then adding carbon disulfide and continuing the reaction at room temperature, then adding 4,5,9,10-tetrabromo-2,7-bis(2-octyldodecyl)benzo[lmn][3,8]phenanthrole-1,3,6,8(2H,7H)-tetraone and continuing the reaction by stirring, and performing post-treatment after the reaction to obtain compound 1; (2) dissolving the compound 1 and ammonium chloride in an organic solvent for reflux reaction, then adding iron powder to continue the reaction, and performing post-treatment after the reaction to obtain compound 2; (3) The compound 2, halogenated aromatic hydrocarbon or halogenated aromatic ether compound or its derivative, deprotonating agent, ligand and catalyst are dissolved in an organic solvent, mixed and subjected to reflux reaction, and after the reaction is completed, post-treatment is performed to obtain the near-infrared emitting aggregation-induced luminescence organic photothermal molecule.

4. The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 3, characterized in that: The halogenated aromatic hydrocarbon or halogenated aromatic ether compound or its derivative in step (3) includes one or more of p-bromoanisole, 4-n-hexyloxybromobenzene, 1-bromo-4-(octadecyloxy)benzene, 1-bromo-4-hexylbenzene, and 1-bromo-4-(perfluorohexyl)benzene.

5. The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 3, characterized in that: The deprotonating agent is sodium tert-butoxide or potassium tert-butoxide; and / or, The ligand is 2-dicyclohexylphosphine-2,4,6-triisopropylbiphenyl or 2-dicyclohexylphosphine-2',6'-diisopropoxy-1,1'-biphenyl; and / or, The catalyst is tris(dibenzylideneacetone)dipalladium(0).

6. Use of a near-infrared emitting aggregation-induced luminescence organic photothermal molecule as claimed in claim 1 or 2 in the preparation of near-infrared aggregation-induced luminescence multifunctional protein nanoparticles.

7. A method for preparing near-infrared aggregation-induced luminescence multifunctional protein nanoparticles, characterized in that: The following steps are involved: The near-infrared emitting aggregation-induced luminescence organic photothermal molecule solution of claim 1 or 2 is mixed with a transferrin solution to carry out a self-assembly reaction, and after the reaction is completed, the solution is washed, centrifuged, filtered and collected to obtain the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles.

8. The method for preparing a near-infrared aggregation-induced luminescence multifunctional protein nanoparticle according to claim 7, characterized in that: The transferrin is one or more of holo-transferrin, functionally modified transferrin and controllably modified transferrin.

9. The multifunctional protein nanoparticles with near-infrared aggregation-induced luminescence prepared by the preparation method according to claim 8.

10. Use of the near-infrared aggregation-induced luminescence multifunctional protein nanoparticles according to claim 9 in the preparation of drugs for diagnosing or treating brain tumors.

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