A photosensitizer targeting beta amyloid and its preparation method and application

Abstract

The application discloses a photosensitizer targeting beta amyloid, and a preparation method and application thereof. The photosensitizer is a photosensitizer with A-D-A configuration, and a structural formula thereof is shown as formula I. The photosensitizer synthesized in the application has a longer emission wavelength, and after being combined with beta amyloid, the fluorescence signal is obviously enhanced, and can effectively generate singlet oxygen, that is, the photosensitizer can target mark A beta protein, and is used for fluorescence imaging of the A beta protein; and after the photosensitizer synthesized in the application is combined with beta amyloid in the brain of an Alzheimer's disease (AD) model mouse, the cognitive ability of the AD mouse can be effectively improved, A beta-mediated neurotoxicity can be reduced, and clearance of A beta plaques can be promoted, that is, the photosensitizer can be used for early diagnosis and treatment of Alzheimer's disease.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine, and specifically relates to a photosensitizer targeting β-amyloid protein, its preparation method, and its application. Background Technology

[0002] Alzheimer's disease (AD), an irreversible neurodegenerative disease, is mainly characterized by cognitive impairment and progressive memory loss. While AD research has garnered widespread attention, its specific pathogenic mechanisms remain unclear. Currently, the most widely accepted theories include: the abnormal deposition of β-amyloid protein, the Tau protein hyperphosphorylation theory, the immune-inflammatory response theory, the mitochondrial dysfunction theory, and the oxidative stress theory. The amyloid β-protein (Aβ) theory plays a crucial role in the early diagnosis and treatment of AD, and Leqembi, a monoclonal antibody drug targeting β-amyloid protein, received accelerated approval from the FDA in 2023. This theory posits that amyloid precursor protein (APP) is cleaved at the β site (Asp1) by β-secretase to produce a 99-amino acid C-terminal fragment (CTF). This CTF is then further cleaved by γ-secretase to form Aβ fragments of varying lengths. Under normal physiological conditions, the formation and degradation of Aβ are in a state of equilibrium. However, in the brains of AD patients, the production and clearance of Aβ protein are uncoordinated. Excessive Aβ in the cerebral cortex or hippocampus misfolds to form neurotoxic oligomers and insoluble fibers, leading to cognitive decline.

[0003] In recent years, photooxidation of Aβ protein has emerged as a novel drug intervention strategy for Alzheimer's disease (AD). Many researchers believe that photooxidation of the Aβ protein side chains reduces its tendency to self-assemble into β-sheet-rich aggregates. Met residues are most susceptible to oxidative stress; in Met35, the sulfur electron pair is occupied by oxygen atoms, producing sulfoxides or sulfones, which reduces the hydrophobicity of the β chain and increases the flexibility of the C-terminal domain of the Aβ protein, thereby inhibiting Aβ protein aggregation. His13 and His14 on the Aβ protein side chains can also be photooxidized to dehydro-2-imidazoline, thus hindering conformational changes in the peptide and intermolecular assembly between Aβ monomers. In this case, photosensitization of Met and His may lead to a reduction in metal-catalyzed ROS (a key player in Aβ-induced cytotoxicity), significantly reducing its cytotoxicity. Furthermore, there are reports that photooxidized β-amyloid protein is more easily cleared by microglia.

[0004] Currently reported photosensitizers for the photooxidation of Aβ protein mainly include small organic molecule photosensitizers (thioflavones, curcumin derivatives, rose red, methylene blue, etc.), metal complexes, and ligand photosensitizers. Since the 1930s, quinoline compounds have primarily been used as antimalarial drugs. With the development of photosensitizers, quinoline compounds have shown great potential value in the field due to their excellent drug-like properties and their near-infrared emission wavelengths and effective generation of reactive oxygen species. Summary of the Invention

[0005] The primary objective of this invention is to overcome the shortcomings and deficiencies of the prior art and provide a photosensitizer that targets β-amyloid protein.

[0006] Another object of the present invention is to provide a method for preparing the photosensitizer that targets β-amyloid protein.

[0007] Another object of the present invention is to provide the application of the photosensitizer that targets β-amyloid protein.

[0008] The objective of this invention is achieved through the following technical solution:

[0009] A photosensitizer targeting β-amyloid protein, being an ADA-configured photosensitizer, comprises an electron-withdrawing group A, a π-conjugated moiety B, an electron-donating group C, a π-conjugated moiety D, and an electron-withdrawing group E; wherein moiety A is primarily 1-methyl-6-dimethylaminoquinoline, moiety B is an ethylene-linking bridge, moiety C is a derivative group such as triphenylamine and tetraphenylethylene, moiety D is an ethylene-linking bridge, and moiety E is primarily 1-methyl-6-dimethylaminoquinoline; preferably, it has the following structure:

[0010]

[0011] In the formula, R is selected from any of the following groups:

[0012]

[0013] The preparation method of the photosensitizer targeting β-amyloid protein includes the following steps:

[0014] (1) N,N-dimethyl-1,4-p-phenylenediamine (p-dimethylaminoaniline), crotonaldehyde and toluene were added to hydrochloric acid solution and refluxed under a protective gas atmosphere at 115±5℃. After the reaction was completed, compound 1 was obtained by separation and purification, and its structural formula is shown in Formula II:

[0015]

[0016] (2) Compound 1 was added to an organic solvent, followed by the addition of iodomethane. The mixture was refluxed under a protective atmosphere at 80±5℃. After the reaction was completed, compound 2 was obtained by separation and purification. Its structural formula is shown in Formula III.

[0017]

[0018] (3) Compound 2 and aldehyde ligand compound were added to an organic solvent, acetic acid was added, and the mixture was refluxed (condensation reaction) at 80±5℃. After the reaction was completed, the photosensitizer targeting β amyloid protein was obtained by purification.

[0019] The protective gas mentioned in steps (1) and (2) is at least one of nitrogen and argon.

[0020] The molar ratio of N,N-dimethyl-1,4-p-phenylenediamine and crotonaldehyde in step (1) is 1:1 to 2; preferably 1:1.5 to 2.

[0021] The amount of toluene used in step (1) is calculated as 8 to 10 ml of toluene per gram of N,N-dimethyl-1,4-p-phenylenediamine; preferably, it is calculated as 8 ml of toluene per gram of N,N-dimethyl-1,4-p-phenylenediamine.

[0022] The concentration of the hydrochloric acid solution mentioned in step (1) is 6 mol / L.

[0023] The reaction described in step (1) is carried out under stirring conditions; the reaction time is 4 to 6 hours.

[0024] The separation and purification described in step (1) is preferably achieved by the following steps: after the reaction is completed, the reaction solution is extracted to remove toluene, then the pH is adjusted to neutral under ice bath conditions, and finally purified by silica gel column chromatography after extraction with dichloromethane to obtain compound 1.

[0025] The conditions for silica gel column chromatography are as follows: the volume ratio of petroleum ether to ethyl acetate is 20:1, and the silica gel is 200-300 mesh.

[0026] The organic solvent mentioned in steps (2) and (3) is preferably ethanol.

[0027] The molar ratio of compound 1 to iodomethane in step (2) is 1:1 to 5; preferably 1:2 to 3.

[0028] The reaction described in step (2) is carried out under stirring conditions; the reaction time is 8 to 12 hours (preferably 10 hours).

[0029] The separation and purification described in step (2) is preferably achieved by the following steps: after the reaction is completed, the solvent is removed by rotary evaporation of the reaction solution, and then purified by silica gel column chromatography to obtain compound 1.

[0030] The conditions for silica gel column chromatography are as follows: the volume ratio of dichloromethane to methanol is 40:1, and the silica gel is 200-300 mesh.

[0031] The aldehyde ligand compound mentioned in step (3) is any one of the following formulas:

[0032]

[0033] The molar ratio of compound 2 to the aldehyde ligand compound in step (3) is 4 to 5:1; preferably 4:1.

[0034] The reaction described in step (3) is carried out under stirring conditions; the reaction time is 3 to 5 hours (preferably 5 hours).

[0035] The amount of acetic acid used in step (3) is calculated based on its final concentration in the system being 0.75% by volume (final concentration being 7.5 μL / mL).

[0036] The purification described in step (3) is carried out by thin-layer chromatography. The conditions for thin-layer chromatography are as follows: the silica gel is 200-300 mesh, and the developing solvent is dichloromethane and methanol mixed in a volume ratio of 100:1.

[0037] The application of the photosensitizer targeting β-amyloid protein in the preparation of Aβ fluorescent imaging agents.

[0038] The use of the photosensitizer targeting β-amyloid protein in the preparation of products for the diagnosis and / or treatment of Alzheimer's disease.

[0039] The products mentioned include photosensitizers, detection or diagnostic reagents, detection or diagnostic kits, therapeutic drugs, etc.

[0040] The therapeutic agents mentioned include Aβ photo-oxidative therapy agents.

[0041] The preferred method of photo-oxidation therapy is to use a 520nm, 1000mW laser for 10±2 minutes.

[0042] The β-amyloid protein is preferably Aβ. 1-42 Aggregates.

[0043] The present invention has the following advantages and effects compared with the prior art:

[0044] 1. This invention synthesizes a novel A-π-D-π-A quinoline onion compound, using 6-dimethylamino-1-methylquinoline as the parent compound. This compound has not been reported in existing photosensitizers for labeling Aβ protein. It can be applied to fluorescence imaging and photo-oxidative therapy for labeling Aβ protein, and can be used for the diagnosis and treatment of early Alzheimer's disease.

[0045] 2. The β-amyloid protein targeting photosensitizer synthesized in this invention has a long emission wavelength, which can specifically detect β-amyloid protein. After binding with β-amyloid protein, the fluorescence signal is significantly enhanced, and it can effectively generate singlet oxygen to photo-oxidize the Aβ protein side chain, which reduces its tendency to self-assemble into aggregates rich in β-sheets. The photo-oxidized Aβ protein is more easily cleared by microglia, thereby significantly reducing its neurotoxicity.

[0046] 3. The 2-substituted arylvinylquinoline derivatives with the A-π-D-π-A configuration synthesized in this invention, using quinoline vinyl derivatives as fluorescent groups, can target and label Aβ protein. After binding to β-amyloid protein in the brain of Alzheimer's disease (AD) model mice, they can effectively improve the cognitive ability of AD mice. This has important guiding significance for the development of Alzheimer's disease diagnostic and therapeutic reagents. Attached Figure Description

[0047] Figure 1 The synthetic route of the photosensitizer in this invention is shown in the figure (a: crotonaldehyde, HCl:H2O = 1:1, 115℃; b: CH3I, EtOH, 80℃; c: different aldehyde ligands, CH3COOK, 80℃).

[0048] Figure 2 This is the structural formula of the photosensitizer synthesized in this invention.

[0049] Figure 3 This is the fluorescence spectrum of the photosensitizer synthesized in this invention and β-amyloid protein.

[0050] Figure 4 This is a diagram showing the ability of the photosensitizer synthesized in this invention to generate singlet oxygen.

[0051] Figure 5 The experiment on AD model mice with the photosensitizer (QM-TA-2) synthesized in this invention in the water maze test; where (A) is the latency period; (B) is the number of times the virtual platform is crossed; (C) is the time spent on the target platform; (D) is the distance spent on the target platform; and (E) is the representative swimming path of different groups of mice.

[0052] Figure 6The image shows HE staining of AD model mice with the photosensitizer (QM-TA-2) synthesized in this invention; where A is the HE staining result of the mouse brain; and B is the statistical result of HE staining of degenerated cells in the mouse brain.

[0053] Figure 7 The images show Nissl staining of AD model mice with the photosensitizer (QM-TA-2) synthesized in this invention; where A is Nissl staining images of the brains of mice in different groups; and B is the quantitative statistical results of surviving brain cells in mice in different groups.

[0054] Figure 8 Immunohistochemical images of AD model mice with the photosensitizer (QM-TA-2) synthesized by the inventors; wherein, A is an immunohistochemical image of Aβ plaques in the mouse brain; B is the quantitative statistical results of Aβ plaques in the mouse brain. Detailed Implementation

[0055] The present invention will be further described in detail below with reference to embodiments, but the implementation of the present invention is not limited thereto. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in this technical field. Test methods in the following embodiments that do not specify specific experimental conditions are generally performed according to conventional experimental conditions or experimental conditions recommended by the manufacturer. Unless otherwise specified, the reagents and raw materials used in the present invention are commercially available.

[0056] Example 1

[0057] 1. Preparation of the compound (see synthetic route diagram) Figure 1 ):

[0058] (1) Synthesis steps of compound 1:

[0059] Weigh 5.8 g of N,N-dimethyl-1,4-p-phenylenediamine (42.60 mmol), dissolve it thoroughly in 250 mL of 6 M hydrochloric acid solution, add 6.5 mL of crotonaldehyde, and add 40 mL of pure toluene solution under argon protection. The reaction is refluxed at 115 °C, and the reaction progress is monitored every 1 hour until the reaction is complete. The reaction solution is directly poured into a separatory funnel for extraction to remove the upper toluene layer. The pH of the aqueous phase is adjusted to neutral under ice bath conditions, and finally extracted with dichloromethane. Purification is achieved by silica gel column chromatography (petroleum ether:ethyl acetate = 20:1, v / v; silica gel 200–300 mesh) to obtain a yellow solid compound 1.

[0060]

[0061] Characterization by proton nuclear magnetic resonance (NMR):

[0062] N,N,2-trimethylquinolin-6-amine: Yellow solid; Yield: 32.21%; 1 H NMR (400MHz, Chloroform-d): δ=7.91 (dd, J=14.3, 8.9Hz, 2H), 7.36 (dd, J=9.3, 2. 8Hz,1H),7.18(d,J=8.4Hz,1H),6.81(d,J=2.9Hz,1H),3.07(s,6H),2.70(s,3H).

[0063] (2) Synthesis steps of compound 2:

[0064] 2.6 g (13.96 mmol) of compound 1 was dissolved in 40 mL of ethanol. Under argon protection, 4.86 g (34.24 mmol) of iodomethane was added dropwise, and the mixture was refluxed at 80 °C for 10 h. After the reaction was complete, the solvent was removed by rotary evaporation, and the mixture was further purified by silica gel column chromatography (dichloromethane:methanol = 40:1, v / v; silica gel 200–300 mesh) to give an orange-yellow solid, compound 2, in 25.06% yield.

[0065]

[0066] 6-(dimethylamino)-1,2-dimethylquinolin-1-ium: Yellow solid; Yield: 25.06%; 1 H NMR (400MHz, DMSO-d6): δ = 8.73 (d, J = 8.6Hz, 1H), 8.35 (d, J = 9.7Hz, 1H), 7.86 (d, J = 8.6Hz, 1H), 7. 77(dd,J=9.9,3.0Hz,1H),7.27(d,J=3.0Hz,1H),4.36(s,3H),3.13(d,J=1.2Hz,6H),2.95(s,3H).

[0067] (3) Synthetic steps of ADA-type compounds:

[0068] Compound 2 and different aldehyde ligands (molar ratio 4:1) were weighed and dissolved in ethanol. An appropriate amount of acetic acid (150 μL, final concentration 7.5 μL / mL) was added, and the mixture was refluxed at 80 °C for 5 h. The mixture was purified by prepared thin-layer chromatography. A 1 mm silica gel (200-300 mesh) coated on a support plate was used as the stationary phase, and a dichloromethane:methanol (100:1, v / v) mixture was used as the developing solvent. A deep blue solid was obtained at room temperature.

[0069]

[0070] 2,2'-((1E,1'E)-((phenylazanediyl)bis(4,1-phenylene))bis(ethene-2,1-diyl))bis(6-(dimethylamino)-1-methylquinolin-1-ium):Blue solid;Yield:54.21%; 1 H NMR(400MHz,DMSO-d6)δ8.70(d,J=9.1Hz,2H),8.35(dd,J=9.5,5.5Hz,4H),8.02(d,J=15.8Hz,2H),7.89(d,J=8.5Hz,4H),7.80–7.63(m,4H),7.47(t,J=7.7Hz,2H),7.37–7.02(m,8H),4.48(s,6H),3.35(s,12H). 13 C NMR(101MHz,DMSO-d6)δ150.73,149.69,149.10,143.65,141.51,132.09,130.88,130.61,130.34,126.64,123.39,122.98,121.14,120.40,117.88,106.23,56.51,19.01.

[0071]

[0072] 2,2'-((1E,1'E)-((2,2-diphenylethene-1,1-diyl)bis(4,1-phenylene))bis(ethene-2,1-diyl))bis(6-(dimethylamino)-1-methylquinolin-1-ium):Blue solid;Yield:46.32%; 1 H NMR(400MHz,DMSO-d6)δ8.72(d,J=9.1Hz,2H),8.34(dd,J=14.3,9.5Hz,4H),7.92(d,J=15.9Hz,2H),7.83–7.64(m,8H),7.37–6.89(m,16H),4.48(s,6H),3.15(s,12H).

[0073]

[0074] 2,2'-((1E,1'E)-(((4-methoxyphenyl)azanediyl)bis(4,1-phenylene))bis(ethene-2,1-diyl))bis(6-(dimethylamino)-1-methylquinolin-1-ium): Blue solid; Yield: 52.45%; 1 H NMR (400MHz, DMSO-d6) δ8.69(d,J=9.1Hz,2H),8.48–8.28(m,4H),8.07(d,J=15.6Hz,2H),7.89(d,J=8.5Hz,4H),7.72(dd,J=9.7,2.9Hz,2H),7. 63(d,J=15.8Hz,2H),7.29(d,J=2.9Hz,2H),7.19(d,J=8.9Hz,2H),7.14 (m,4H),7.06(d,J=9.0Hz,2H),5.08(s,3H),3.82(s,3H),3.15(s,12H).

[0075] The excitation and emission wavelengths of the synthesized compounds QM-TA-2, QM-TB-2, and QM-OTA-2 are shown in Table 1.

[0076] Table 1 Excitation and emission wavelengths of photosensitizers

[0077] name λex(nm) λem(nm) QM-TA-2 542 720 QM-TB-2 494 678 QM-OTA-2 545 674

[0078] λex: Excitation wavelength of the probe;

[0079] λem: The maximum emission wavelength of the probe.

[0080] 2. Fluorescence response of photosensitizer to Aβ aggregates:

[0081] Photosensitizers (QM-TA-2, QM-TB-2, and QM-OTA-2) and Aβ were prepared separately. 1-42 The final concentrations of the aggregates (β-Amyloid(1-42), Shanghai Qiangyao Biotechnology Co., Ltd.) were 1 μM and 10 μM, respectively. After thorough mixing, the mixture was incubated at room temperature for 10 minutes, and the fluorescence spectra were detected by a fluorescence spectrophotometer. PBS buffer was used as a control, and the assay was repeated three times. The results are as follows: Figure 3 As shown.

[0082] 3. Measurement of singlet oxygen produced by the photosensitizer:

[0083] The generation of singlet oxygen was detected using 9,10-anthratridiyl-bis(methylene)dicarboxylic acid (ABDA) as an indicator, with the commercial photosensitizer rose red (RB) as a control. First, mixed solutions of ABDA (final concentration 100 μM) and different photosensitizers (final concentration 10 μM) were prepared. Then, the absorbance at the characteristic peak of ABDA at 378 nm was recorded after irradiation with white light (10 mW / cm²) for different times (0–500 s). The results are as follows: Figure 4 As shown.

[0084] 4. Evaluation of the efficacy of anti-AD drugs;

[0085] (1) The mice were divided into the following groups (n=6 / group): (1) Wt: Wt mice (C57BL6, 11 months old, male) injected intravenously with 0.9% saline; (2) AD: Tg mice (C57BL6, APPsw / PSEN1, 11 months old, male, Huafukang Biotechnology Co., Ltd.) injected intravenously with 0.9% saline; (3) AD+Light: Tg mice were injected intravenously with 0.9% saline and irradiated with laser (520nm, 1000mW) for 10 min 1 h after injection; (4) AD+QM-TA-2: QM-TA-2 was injected intravenously; (5) AD+QM-TA-2+Light: QM-TA-2 was injected intravenously and irradiated with laser (520nm, 1000mW) for 10 min 1 h after injection; QM-TA-2 was dissolved in 0.9% saline for 15 consecutive days and then injected intravenously at 5 mg / kg / day. Before and after irradiation, the local temperature of the head was monitored using a thermal infrared imaging camera.

[0086] (2) Behavioral studies were conducted on different groups of mice (n=6 / group) using the water maze (MWM) test. The experimental setup consisted of a circular water tank (120 cm in diameter, 45 cm in height, water temperature 20±1℃, depth 32 cm, the water was made opaque by adding non-toxic white TiO2). A platform (9 cm in diameter, 30 cm in height) was hidden 2.0 cm below the water surface. Mice in different groups were trained for 5 consecutive days. Mice had 90 seconds to find the hidden platform; if not, they were guided to the platform. The escape latency was recorded as 90 seconds. A computerized digital video tracking system was used to record the escape latency each time. 24 hours after the 5th day of training, the platform was removed from the tank. The time taken to traverse the platform (original platform position), average swimming speed, time taken, swimming path length in the target quadrant, and the virtual platform were recorded using video tracking software to analyze the mice's memory consolidation.

[0087] (3) HE staining: Six mice in each group were euthanized by misalignment. The brains were removed and bonded in 4% formaldehyde for 24 hours, then embedded in paraffin. Paraffin sections were dewaxed and rehydrated according to routine procedures, stained with hematoxylin for 5 minutes, eosin (Wuhan Sewell Biotechnology Co., Ltd.) for 1 minute, and then dehydrated with graded series of alcohols (70%, 95%, and 100%), cleared with xylene, and sealed. Computer image analysis was used to study the histopathological abnormalities of the hippocampus. The degenerated cell index was calculated using ImageJ software in three representative mouse images.

[0088] (4) Nissl staining: 4 μm slides were rinsed twice in 10 mM PBS buffer with 0.5% (v / v) toluene violet solution (containing a small amount of glacial acetic acid) for 15 minutes each time, followed by rinsing with distilled water. The slides were then dehydrated in fractionated ethanol (70%, 95%, and 100%, v / v) and soaked in xylene and a precipitate solution. Cells in the hippocampal Cortex and CA1, CA3, and DG regions were counted using computer image analysis. The percentage of surviving neurons was calculated using ImageJ software in three representative images from each mouse.

[0089] (5) Immunohistochemistry: Antigen retrieval was performed in a microwave oven (medium heat for 8 minutes, stop heating for 8 minutes, medium-low heat for 7 minutes) (P70D20TL-P4, Galanz Microwave Oven Appliances Co., Ltd.). After cooling at room temperature, it was washed twice with PBS (pH 7.4). Then, it was quenched in the dark with 3% (v / v) hydrogen solution at room temperature for 25 minutes, washed twice with PBS (pH 7.4) for 5 minutes each time, and further incubated in bovine serum albumin (BSA) blocking solution for 30 minutes. After blocking, β-amyloid 1-16 (6E10) monoclonal antibody (Wuhan Sewell Biotechnology Co., Ltd.) was diluted 1:100 (v / v) and incubated overnight, followed by secondary antibody (Wuhan Sewell Biotechnology Co., Ltd.) diluted 1:500 (v / v) and incubated for 50 minutes. Washed twice with PBS, incubated in 3,3′-diaminobenzidine tetrahydrochloride (DAB), washed with distilled water, dehydrated in fractionated ethanol (70%, 80%, 90%, 95%, and 100%, v / v), placed in xylene, and then covered. Using ImageJ software, the percentage of Aβ plaque deposition area relative to the total brain area in three representative images for each mouse was calculated.

[0090] The results are as follows Figures 5-8 As shown:

[0091] Results of the Morris water maze experiment ( Figure 5 The results showed that AD mice treated with the drug and exposed to light exhibited a similar tendency-based search strategy to wild-type mice. Figure 5 (E) I and V), while other groups used an edge search strategy, indicating that mice treated with photooxidation and wild-type mice had better spatial learning and memory. This was achieved by comparing the latency, number of times the virtual platform was crossed, time spent in the target quadrant, and time distance between the virtual platform and the different groups of mice. Figure 5 (II, III, IV in (E)) further demonstrate that photo-oxidative therapy can significantly improve cognitive impairment in AD mice and effectively enhance their spatial memory ability.

[0092] Following behavioral tests, we collected brain slices from six mice in each group for HE staining, Nissl staining, and immunohistochemical staining to investigate the effects of photooxidative therapy on neurons and Aβ plaques. I first used HE staining to evaluate the degree of neuronal degeneration in the cerebral cortex and hippocampus of each group of mice. The HE staining results are shown below. Figure 6 As shown, compared with the AD group, the drug-treated and light-treated group showed a significant reduction in the number of apoptotic and necrotic cells in the cortex and hippocampal CA1, CA3, and DG regions, exhibiting reduced cell division, nuclear pyknosis, irregular morphology, and darker HE staining. Statistical analysis revealed that after drug-treated and light-treated mice, the degeneration cell index was comparable to the wild-type group and significantly lower than that of the AD group, as well as AD mice treated with 520nm laser alone or QM-TA-2 alone. Nissl staining was used to further evaluate the protective effect of QM-TA-2-mediated photo-oxidative therapy against neuronal damage. Figure 7 Nissl staining and statistical analysis showed that the drug plus light irradiation group contained more Nissl body neurons, similar to wild-type mice, while the AD group, as well as the drug-only or light-only groups, showed a significant reduction in Nissl bodies. HE staining and Nissl staining results both indicated that QM-TA-2-mediated photooxidation of Aβ could reduce Aβ-mediated neurotoxicity and protect neurons. We further investigated the effect of QM-TA-2-mediated photooxidation on Aβ content using immunohistochemical staining. Figure 8 The number of Aβ plaques in AD mice was significantly reduced after drug administration and light treatment, and was significantly lower than that in the AD group, as well as in the drug administration alone or light treatment alone groups, indicating that QM-TA-2-mediated photo-oxidation can promote the clearance of Aβ plaques.

[0093] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. A photosensitizer targeting β-amyloid protein, characterized in that: The photosensitizer has the following structure: Equation I; In the formula, R is selected from any of the following groups: 、 、 。 2. The method for preparing the photosensitizer targeting β-amyloid protein according to claim 1, characterized in that, Includes the following steps: (1) N,N-dimethyl-1,4-p-phenylenediamine, crotonaldehyde, and toluene were added to a hydrochloric acid solution and refluxed under a protective atmosphere at 115 ± 5 °C. After the reaction was completed, compound 1 was obtained by separation and purification, and its structural formula is shown in Formula II: Formula II; (2) Compound 1 was added to an organic solvent, followed by the addition of iodomethane. The mixture was refluxed under a protective atmosphere at 80 ± 5 °C. After the reaction was completed, compound 2 was obtained by separation and purification. Its structural formula is shown in Formula III. Formula III; (3) Compound 2 and aldehyde ligand compound were added to an organic solvent, acetic acid was added, and the mixture was refluxed at 80 ± 5 °C. After the reaction was completed, the photosensitizer targeting β amyloid protein was obtained by purification. The aldehyde ligand compound mentioned in step (3) is any one of the following formulas: 、 、 。 3. The method for preparing the photosensitizer targeting β-amyloid protein according to claim 2, characterized in that: The molar ratio of N,N-dimethyl-1,4-p-phenylenediamine and crotonaldehyde in step (1) is 1:1 to 2; The molar ratio of compound 1 to iodomethane in step (2) is 1:1 to 5; The molar ratio of compound 2 to the aldehyde ligand compound in step (3) is 4 to 5:

1.

4. The method for preparing the photosensitizer targeting β-amyloid protein according to claim 2, characterized in that: The separation and purification described in step (1) are achieved through the following steps: after the reaction is completed, the reaction solution is extracted to remove toluene, then the pH is adjusted to neutral under ice bath conditions, and finally purified by silica gel column chromatography after extraction with dichloromethane to obtain compound 1; wherein the silica gel column chromatography conditions are: the volume ratio of petroleum ether to ethyl acetate is 20:1, and the silica gel is 200-300 mesh. The separation and purification described in step (2) are achieved by the following steps: the reaction solution after the reaction is completed is removed by rotary evaporation to remove the solvent, and then purified by silica gel column chromatography to obtain compound 1; wherein the conditions for silica gel column chromatography are: the volume ratio of dichloromethane to methanol is 40:1, and the silica gel is 200-300 mesh. The purification described in step (3) is carried out by thin-layer chromatography. The conditions for thin-layer chromatography are as follows: the silica gel is 200-300 mesh, and the developing solvent is dichloromethane and methanol mixed in a volume ratio of 100:

1.

5. The method for preparing the photosensitizer targeting β-amyloid protein according to claim 2, characterized in that: The protective gas mentioned in steps (1) and (2) is at least one of nitrogen and argon; The amount of toluene used in step (1) is calculated as 8 to 10 ml of toluene per gram of N,N-dimethyl-1,4-p-phenylenediamine; The concentration of the hydrochloric acid solution mentioned in step (1) is 6 mol / L; The reaction described in step (1) is carried out under stirring conditions; the reaction time is 4 to 6 hours. The organic solvent mentioned in steps (2) and (3) is ethanol; The reaction described in step (2) is carried out under stirring conditions; the reaction time is 8 to 12 hours. The reaction is carried out under the stirring conditions described in step (3); the reaction time is 3 to 5 hours. The amount of acetic acid used in step (3) is calculated based on its final concentration in the system being 0.75% by volume.

6. The use of the photosensitizer targeting β-amyloid protein as described in claim 1 in the preparation of Aβ fluorescent imaging agents.

7. The use of the photosensitizer targeting β-amyloid protein as described in claim 1 in the preparation of products for diagnosing Alzheimer's disease.

8. The use of the photosensitizer targeting β-amyloid protein according to claim 1 in the preparation of products for treating Alzheimer's disease, characterized in that: R is selected from the following groups: 。

Images