A near-infrared-emitting aggregation-induced emission organic photothermal molecule, its preparation method and applications

Abstract

The present invention discloses a near-infrared-emitting aggregation-induced emission organic photothermal molecule, its preparation method and application, belonging to the technical field of biomedicine, and the structure is shown in formula I: #imgabs0# (I); wherein, R1, R2, R3, R4, R5, and R6 are independently selected from groups containing alkyl chains. The present invention also provides a preparation method of the above-mentioned organic photothermal molecule and its application in the preparation of near-infrared aggregation-induced emission multifunctional protein nanoparticles. The near-infrared-emitting aggregation-induced emission organic photothermal molecule provided by the present invention exhibits excellent near-infrared imaging effects and photothermal capabilities under laser irradiation. The obtained near-infrared aggregation-induced emission multifunctional protein nanoparticles have excellent photothermal effects and imaging resolutions, have excellent targeting to brain tumors, have practical medical significance in crossing the blood-brain barrier, tumor killing, and combined photothermal and ferroptosis therapy, and are expected to be widely used in intelligent drug delivery and nanomedicine.

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 ( Figure 11 ) or Angiopep-2 ( Figure 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. Figure 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, said R 1 , R 2 , R 3 , R 4 , R 5 , R 6 are independently selected from groups containing an alkyl chain.

[0011] Preferably, said alkyl chain is a straight chain of C 1 -C 20 , a branched chain of C 1 -C 20 , a cyclic alkyl chain of C 1 -C 20 , an alkyl chain in which carbon atoms are substituted by one or more of oxygen atom, alkenyl, alkynyl, aryl, carbonyl, hydroxyl, amino, carboxyl, cyano, nitro and ester group, or an alkyl chain in which hydrogen atoms are substituted by one or more of fluorine atom, chlorine atom, bromine atom and iodine atom.

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

[0013]

[0014] Beneficial effects: The present invention provides a novel near-infrared-emitting aggregation-induced emission organic photothermal molecule with an electron donor-electron acceptor structure, using thieno[3,4-b]thiophene-3,6-dicarboximide with a long alkyl chain as an electron acceptor group and triphenylamine as a molecular rotor and electron donor unit. Under laser irradiation, it can not only emit light in the second near-infrared region but also generate heat, and has good photostability. In addition, a long alkyl chain is introduced into triphenylamine to regulate the stacking mode of molecules in the aggregated state. Specifically, the large conjugated main chain of the electron acceptor group has a quinone-like characteristic to allow large electron delocalization, forming a lower bandgap. Its large planar structure is conducive to improving the long-wavelength absorption ability and high molar absorption rate. The long alkyl chain on the electron acceptor avoids excessive intermolecular stacking, avoiding disadvantages such as poor imaging effect, high signal-to-noise ratio, and poor photostability caused by aggregation-induced quenching. The electron donor is electron-rich triphenylamine, and 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 ability of the material. The long alkyl chain in triphenylamine gives the molecule greater rotational freedom of the rotor, 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 emission 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 preparation method of an organic photothermal molecule with near-infrared emission and aggregation-induced emission, comprising the following steps:

[0016] (1) React sodium hydride and 4-nitrobenzonitrile under ice bath conditions, then add carbon disulfide and continue the reaction at room temperature, and then add 4,5,9,10-tetrabromo-2,7-bis(2-octyldodecyl)benzo[lmn][3,8]phenanthroline-1,3,6,8(2H,7H)-tetrone and continue stirring and reacting. After the reaction is completed, perform post-treatment to obtain Compound 1;

[0017] (2) Dissolve Compound 1 and ammonium chloride in an organic solvent and carry out a reflux reaction, then add iron powder and continue the reaction. After the reaction is completed, perform post-treatment to obtain Compound 2;

[0018] (3) Dissolve Compound 2, a haloaromatic hydrocarbon or a haloaromatic ether compound or its derivative, a deprotonating agent, a ligand, and a catalyst in an organic solvent, mix them and carry out a reflux reaction. After the reaction is completed, perform post-treatment to obtain the organic photothermal molecule with near-infrared emission and aggregation-induced emission.

[0019] Preferably, the haloaromatic hydrocarbon or the haloaromatic 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, 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-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl or 2-dicyclohexylphosphino-2',6'-diisopropoxy-1,1'-biphenyl; and / or,

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

[0023] An application of an organic photothermal molecule with near-infrared emission and aggregation-induced emission in the preparation of near-infrared aggregation-induced emission multifunctional protein nanoparticles.

[0024] A preparation method of near-infrared aggregation-induced emission multifunctional protein nanoparticles, comprising the following steps:

[0025] Mix the above-mentioned organic photothermal molecule solution with near-infrared emission and aggregation-induced emission and transferrin solution to carry out a self-assembly reaction. After the reaction is completed, wash, centrifuge, filter and collect to obtain the near-infrared aggregation-induced emission multifunctional protein nanoparticles.

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

[0027] Preferably, the concentration of the near-infrared-emitting aggregation-induced emission organic photothermal molecular 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 emission organic photothermal molecular solution to the transferrin solution is 1:3 (w / w).

[0030] Beneficial effects: Holotransferrin is a protein rich in iron, mainly used to transport iron absorbed by the digestive tract and iron released by the degradation of red blood cells. The embedded nanoparticles have excellent properties such as high stability, biocompatibility, and biodegradability, and have the ability to highly bind to organic small molecules. Transferrin is essential for ferroptosis. It can transport Fe 3+ into the nuclear endosome, where Fe 3+ is further reduced to Fe 2+ . Finally, mediated by divalent metal transporter 1, Fe 2+ is released from the endosome into the cytosolic labile iron pool. 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, holotransferrin, as a potential nanocarrier, can effectively cross the blood-brain barrier and simultaneously induce ferroptosis of brain tumor cells, ultimately leading to the process of ferroptosis in cells. The near-infrared-emitting aggregation-induced emission organic photothermal molecule provided by the present invention has the characteristics of near-infrared second-region luminescence, heat generation, and good photostability. Subsequently, holotransferrin is used as a biological functional carrier,

[0031] The present invention mixes an organic photothermal molecule with near-infrared emission and aggregation-induced emission and holotransferrin in a fixed ratio, uniformly loads the above organic photothermal molecule on the protein to form a near-infrared molecule-holotransferrin composite nanoparticle. There are strong hydrogen bond and van der Waals force interactions between the organic photothermal molecule and transferrin. This ingenious and unique interaction makes the performance of the protein nanoparticle stable. The presence of holotransferrin enables the protein to efficiently cross the blood-brain barrier and promote the occurrence of ferroptosis in the tumor site. Moreover, the organic photothermal molecule with near-infrared emission and aggregation-induced emission exhibits excellent near-infrared imaging effects and photothermal capabilities under laser irradiation. The obtained multifunctional protein nanoparticles with near-infrared aggregation-induced emission have excellent photothermal effects and imaging resolutions, have excellent targeting for brain tumors, and have practical medical significance in crossing the blood-brain barrier, tumor killing, and combined photothermal and ferroptosis therapy, and are expected to be widely used in intelligent drug delivery and nanomedicine.

[0032] The multifunctional protein nanoparticles with near-infrared aggregation-induced emission prepared by the preparation method as described above.

[0033] Beneficial effects: The product structure provided by the present invention includes a large planar benzene ring structure with high absorption and a twisted backbone 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 emission materials. The introduction of an electron-donating terminal alkyl chain balances the luminescence and heating capabilities of the materials, enabling this new material to have good luminescence imaging performance. The present invention is a near-infrared nanodiagnosis and treatment system integrating 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 in a block.

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

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

[0037] Use of a multifunctional protein nanoparticle with near-infrared aggregation-induced emission in the preparation of a drug for diagnosing or treating brain tumors.

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

[0039] The synthesis method of the near-infrared-emitting aggregation-induced emission organic photothermal molecule provided by the present invention has a simple and efficient synthesis process. The obtained novel organic molecule has luminescence performance in the second near-infrared region and excellent photothermal performance. At the same time, the preparation process of the near-infrared aggregation-induced emission multifunctional protein nanoparticles in the present invention is simple and has excellent performance, which can effectively cross the blood-brain barrier and accumulate in the brain tumor site, realizing visualization and photothermal therapy killing of brain tumors. There are strong hydrogen bond and van der Waals force interactions between the molecular structure of the organic material and holotransferrin. 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 this application are used to provide a further understanding of this application. The schematic embodiments of this application and their descriptions are used to explain this application and do not constitute an improper limitation of this application. In the drawings:

[0041] Figure 1 It is the fluorescence spectrum diagram of the near-infrared-emitting aggregation-induced emission 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 It is the molecular docking and molecular dynamics simulation results of the interaction between the near-infrared organic molecule and holotransferrin in the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4;

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

[0044] Figure 4 It is the transmission scanning electron microscope image of the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4;

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

[0046] Figure 6 It is the in vitro simulation of crossing the blood-brain barrier of the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4;

[0047] Figure 7 It is the induction of ferroptosis in U87 cells by the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4;

[0048] Figure 8 It is the tumor-targeted imaging using the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4;

[0049] Figure 9 Photothermal therapy / imaging effect of multifunctional protein nanoparticles with near-infrared aggregation-induced emission obtained in Example 4 on brain tumors;

[0050] Figure 10 Combined treatment effect of multifunctional protein nanoparticles with near-infrared aggregation-induced emission obtained in Example 4 on brain tumors;

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

[0052] Figure 12 Modification of nanoparticles with transduction peptide Angiopep-2 for crossing the blood-brain barrier;

[0053] Figure 13 Imaging effect of traditional near-infrared photodiagnostic materials after binding to proteins. Specific embodiments

[0054] The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0055] To make the above objects, features, and advantages of the present invention more obvious and understandable, the present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0056] Unless otherwise specified, 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 embodiments of the present invention are not obtained by purchasing through commercial channels;

[0058] Among them, re-evaporated N,N-dimethylformamide is purchased from Aladdin Chemistry Co., Ltd.

[0059] Example 1

[0060] Preparation method of Compound 1, the synthesis route is as follows:

[0061]

[0062] Specifically, it includes the following steps:

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

[0064] 1 H NMR (500 MHz, CDCl 3 ), 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, CDCl 3 ) 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 N 6 O 8 S 4 , 1295.5737; found, 1295.5784.

[0065] Example 2

[0066] Preparation method of Compound 2, and the synthetic route is as follows:

[0067]

[0068] Specifically, it includes the following steps:

[0069] Under nitrogen protection, 200 mg of 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 continued for 40 minutes. After the reaction was completed, it was cooled to room temperature, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography, using a dichloromethane / methanol mixed solution (volume ratio 10:1) as the eluent to obtain a green powder product, Compound 2, with a yield of 76%.

[0070] 1 H NMR (500 MHz, CDCl 3 ) 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, CDCl 3 ) 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 N 6 O 4 S 4 , 1235.6253; found, 1235.6283.

[0071] Example 3

[0072] Preparation method of an organic photothermal molecule NDA-18 with near-infrared emission and aggregation-induced emission, and the synthetic route is as follows:

[0073]

[0074] Specifically, the following steps are included:

[0075] Under nitrogen protection, the compound 2 (100 mg, 0.08 mmol) obtained in Example 2, 1-bromo-4-(octadecyloxy)benzene (170 mg, 0.40 mmol), sodium tert-butoxide (53.6 mg, 0.56 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (18.2 mg, 0.2 mmol), and 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, it was cooled to room temperature, and the mixture was 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 the eluent to obtain the near-infrared-emitting aggregation-induced emission organic photothermal molecule NDA-18 as a green powder product with a yield of 75%.

[0076] 1 H NMR (500 MHz, CDCl 3 ) 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,CDCl 3 ) 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 H254 N 6 O 8 S 4 , 2613.8603; found, 2613.7075.

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

[0078] Example 4

[0079] A method for preparing multifunctional protein nanoparticles with near-infrared aggregation-induced emission, comprising the following steps:

[0080] Dissolve 12 g of holotransferrin in 4 mL of ultrapure water to obtain a holotransferrin solution, and dissolve 4 g of the organic photothermal molecule NDA-18 with near-infrared emission and aggregation-induced emission obtained in Example 3 in 4 mL of tetrahydrofuran to form a solution. Then mix the above holotransferrin solution and the organic photothermal molecule solution with near-infrared emission and aggregation-induced emission evenly at room temperature and react for 3 hours. After the reaction, wash, centrifuge, filter and collect to obtain multifunctional protein nanoparticles with near-infrared aggregation-induced emission.

[0081] Figure 2 These are the results of molecular docking and molecular dynamics simulation of the interaction between the near-infrared organic molecule and holotransferrin in the obtained multifunctional protein nanoparticles with near-infrared aggregation-induced emission. It can be seen that holotransferrin can bind to the organic photothermal molecule with near-infrared emission and aggregation-induced emission at specific sites, and the binding energy is -5.56 kcal / mol. It is proved that there are strong hydrogen bond and van der Waals force interactions between the two, thus making the structure and performance of the multifunctional protein nanoparticles with near-infrared aggregation-induced emission stable.

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

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

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

[0085] Example 5

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

[0087]

[0088] 1 H NMR (500 MHz, CDCl 3 ) 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 preparation method of an organic photothermal molecule NDA-6 with near-infrared emission and aggregation-induced emission, which is different from Example 3 in that 1-bromo-4-(octadecyloxy)benzene is replaced with 4-n-hexyloxybromobenzene of equal molar mass. 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, CDCl 3 ) 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,CDCl 3) 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 for C 120 H 158 N 6 O 8 S 4 , 1940.1058; found, 1940.1410.

[0093] Example 7

[0094] A preparation method of an organic photothermal molecule NDA-6A with near-infrared emission and aggregation-induced emission, which is different from Example 3 in that 1-bromo-4-(octadecyloxy)benzene is replaced with 1-bromo-4-hexylbenzene of equal molar mass. Other process steps and parameters are the same as those in Example 3. The yield is 61%. The synthetic route is as follows:

[0095]

[0096] 1 H NMR (500 MHz, CDCl 3 ) 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, CDCl 3) 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 N 6 O 4 S 4 , 1876.1261; found, 1876.1603.

[0097] Example 8

[0098] A preparation method of an organic photothermal molecule NDA-6F with near-infrared emission and aggregation-induced emission, 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. Other process steps and parameters are the same as those in Example 3. The yield is 49%. The synthetic route is as follows:

[0099]

[0100] 1 H NMR (500 MHz, CDCl 3 ) 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, CDCl 3 ) 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, CDCl 3) 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 N 6 O 4 S 4 , 2811.6362; found, 2811.3159.

[0101] Examples 9 - 12

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

[0103] Technical effects:

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

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

[0106] Figure 6Part B in this is the in vitro simulation of the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4. It can be seen that the FITC-labeled near-infrared aggregation-induced emission multifunctional protein nanoparticles (N18Tf NPs in the figure) show strong signals in the cytoplasm of U87 cells in the lower chamber, demonstrating their good in vitro blood-brain barrier penetration and cell uptake ability.

[0107] 2. The near-infrared aggregation-induced emission multifunctional protein nanoparticles induce ferroptosis in U87 cells.

[0108] 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. Then, it was incubated with a specific primary antibody against the ferroptosis-related marker protein GPX. Detection was carried out with a secondary antibody (HRP-labeled), and developed with a chemiluminescent substrate. The expression differences of ferroptosis-related proteins between the experimental group and the control group were analyzed. The decrease in GPX expression in the experimental group indicated that particle treatment could induce ferroptosis in U87 cells.

[0110] Figure 7 For the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4 to induce ferroptosis in U87 cells, it can be seen that after adding the near-infrared aggregation-induced emission multifunctional protein nanoparticles (corresponding to the two right columns of bands), the GPX4 protein significantly decreased, indicating that the cells may be in an oxidative stress and iron-related death pathway.

[0111] 3. Tumor-targeted imaging of the near-infrared aggregation-induced emission multifunctional protein nanoparticles, and the results are as Figure 8 shown. It can be seen that the fluorescence signal of the nanoparticles gradually increases at the tumor site and reaches a peak at 12 hours, indicating effective accumulation and targeting of the tumor site.

[0112] Figure 9 For the photothermal therapy / imaging effect of the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4 on brain tumors. It can be seen that after adding the near-infrared aggregation-induced emission multifunctional protein nanoparticles to the brain tumor area, the highest temperature reached 42 degrees Celsius within 5 minutes, and this temperature showed an obvious difference from the surrounding environmental temperature, demonstrating the precise and efficient photothermal killing ability of the brain tumor area.

[0113] Figure 10 For the combined treatment effect of the near-infrared aggregation-induced emission multifunctional protein nanoparticles obtained in Example 4 on brain tumors, it specifically includes the following steps:

[0114] The multifunctional protein nanoparticles with near-infrared aggregation-induced emission not only have high efficiency in inducing tumor ferroptosis. After turning on the 808 laser, the high-efficiency photothermal ability ensures the smooth progress of photothermal therapy, thus realizing the combined treatment mode of ferroptosis / photothermal killing for brain tumors. According to Figure 10 As can be seen, during the treatment period, the nanoparticle treatment group showed a longer survival rate, indicating the potential of the nanoparticles for the combined treatment of brain tumors.

[0115] The above are only the preferred specific embodiments of the present application, but the protection scope of the present application is not limited thereto. Any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present application should be covered by the protection scope of the present application. Therefore, the protection scope of the present application should be subject to 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: ； Wherein, R1, R2, R3, R4, R5, and R6 are independently selected from the following groups: C1-C 20 Straight chain alkyl, C1-C 20 In the branched alkyl group, the carbon atoms may be replaced by oxygen atoms, and the hydrogen atoms may be replaced by one or more of fluorine atoms, chlorine atoms, bromine atoms and iodine atoms.

2. An organic photothermal molecule that emits near-infrared aggregation-induced luminescence, characterized in that: Selected from the following structures: 。 3. The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 2, characterized in that: The following steps are involved: (1) Sodium hydride and 4-nitrophenylacetonitrile are reacted in an ice bath, then carbon disulfide is added to continue the reaction at room temperature, and then After the reaction is completed, the mixture is stirred and reacted 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) dissolving the compound 2, halogenated aromatic hydrocarbon or halogenated aromatic ether compound or its derivative, deprotonating agent, ligand and catalyst in an organic solvent, mixing and performing reflux reaction, and performing post-treatment after the reaction to obtain the near-infrared emitting aggregation-induced luminescence organic photothermal molecule; The halogenated aromatic hydrocarbon or halogenated aromatic ether compound or its derivative in step (3) is selected from one or more of p-bromoanisole, 4-n-hexyloxybromobenzene, 1-bromo-4-(octadecyloxy)benzene, 1-bromo-4-hexylbenzene, and 1-bromo-4-(perfluorohexyl)benzene.

4. 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.

5. The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 3, characterized in that: The ligand is 2-dicyclohexylphospho-2',4',6'-triisopropylbiphenyl or 2-dicyclohexylphospho-2',6'-diisopropoxy-1,1'-biphenyl.

6. The method for preparing a near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 3, characterized in that: The catalyst is tris(dibenzylideneacetone)dipalladium(0).

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

8. 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 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.

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

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

11. Use of the near-infrared emitting aggregation-induced luminescence organic photothermal molecule according to claim 1 or the near-infrared aggregation-induced luminescence multifunctional protein nanoparticle according to claim 10 in the preparation of drugs for diagnosing or treating brain tumors.

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