A dual-mode imaging probe for Aβ patches, its preparation method and application

Abstract

This invention provides a dual-mode imaging probe for Aβ patches, its preparation method, and its application, belonging to the field of fine chemical technology. This invention uses quinoline-malononitrile as the fluorescent dye parent material, dimethylaminobenzene and other units as Aβ patch binding groups, introduces thiophene and other units as π-bridges to extend the probe's emission wavelength, and uses... 18 F was used as a PET imaging nuclide to prepare a dual-mode imaging probe for Aβ plaques. This probe has the advantages of long fluorescence emission wavelength, high affinity, good selectivity, strong stability and excellent biocompatibility. It can realize the staining and labeling of Aβ plaques and accurately reflect the distribution, morphology and size of the plaques.

Description

Technical Field

[0001] This invention relates to the field of fine chemical technology, and in particular to an Aβ patch dual-mode imaging probe, its preparation method, and its application. Background Technology

[0002] Alzheimer's disease (AD) is a progressive neurodegenerative disease, one of its main pathological features being the abnormal deposition of β-amyloid (Aβ) plaques in the brain. Currently, the primary technique used for imaging Aβ plaques is positron emission tomography (PET). PET imaging utilizes radioactive tracers (such as...) 18 Compounds labeled with [F] can bind to Aβ plaques, enabling three-dimensional quantitative imaging with excellent tissue penetration. However, PET imaging has low spatial resolution, making precise localization of Aβ plaques difficult. Fluorescence imaging technology, with its high sensitivity and high resolution, has unique advantages in revealing the microstructure of Aβ plaques. However, its application is limited by the attenuation and scattering of fluorescence signals in deep tissues. In summary, current Aβ imaging reagents developed based on single-modality methods cannot simultaneously meet the multiple requirements of spatial resolution, sensitivity, quantitative analysis, and tissue penetration.

[0003] Therefore, developing a dual-mode imaging probe for Aβ plaques that integrates the advantages of real-time dynamic monitoring by fluorescence imaging and quantitative detection by PET imaging, enabling it to simultaneously provide high-resolution local microscopic information and quantitative imaging data, will help provide more comprehensive imaging evidence for pathological research and treatment efficacy evaluation of Alzheimer's disease. Summary of the Invention

[0004] The purpose of this invention is to provide an Aβ patch dual-mode imaging probe, its preparation method, and its application, in order to solve the problem that existing Aβ imaging reagents cannot simultaneously meet the multiple requirements of spatial resolution, sensitivity, quantitative analysis, and tissue penetration.

[0005] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0006] This invention provides a dual-mode imaging probe for Aβ patches, the dual-mode imaging probe for Aβ patches having the structure shown in Formula I:

[0007]

[0008] In Formula I, R1 is selected from cyano, carboxyl, or... R2 is selected from C1 to C4 hydrocarbon groups. Where n is an independent integer between 0 and 10, and F is selected from...18 F or 19 F;R3 is selected from methyl, Where m is independently 0 or 1, and R5 is independently selected from -H, -CH3, -CH2OH, -CH2OTs, -CH2 18 F,-CH2 19 F, -CH2CH3, -CH2CH2OH, -CH2 CH2OTs, -CH2CH2 18 F or -CH2CH2 19 F;R4 is selected from -H, - at any position of the 5th, 6th, 7th, or 8th position of the quinoline group. 18 F or - 19 F.

[0009] Preferably, R1 is selected from cyano.

[0010] Preferably, R2 is selected from Where n is an independent integer between 0 and 10, and F is selected from... 18 F or 19 F.

[0011] Preferably, R3 is selected from Where m is independently 0 or 1, and R5 is independently selected from -H, -CH3, -CH2OH, -CH2OTs, -CH2 18 F,-CH2 19 F, -CH2CH3, -CH2CH2OH, -CH2 CH2OTs, -CH2CH2 18 F or -CH2CH2 19 F.

[0012] Preferably, when R2 is selected from When n is an independent integer between 0 and 10, R3 is selected from... Where m is independently 0 or 1, and R5 is independently selected from -CH2OH, -CH2OTs, and -CH2 18 F,-CH2 19 F, -CH2CH2OH, -CH2CH2OTs, -CH2CH2 18 F or -CH2CH2 19 F;

[0013] When R2 is selected When n is an independent integer between 0 and 10, F is selected from... 18 F or 19 F, R3 are selected from Where m is independently 0 or 1, and R5 is independently selected from -H, -CH3 or -CH2CH3.

[0014] Preferably, the Aβ patch dual-mode imaging probe has a structure as shown in Formula I-1, I-2, II-1, or II-2:

[0015]

[0016]

[0017] This invention provides a method for preparing the above-described Aβ patch dual-mode imaging probe, comprising the following steps:

[0018] (1) Under argon protection, N,N-dimethylformamide was used as a solvent to react compound A, compound X and sodium ethoxide to prepare compound B;

[0019] (2) Compound B and compound Y reacted under argon protection conditions with toluene as solvent to prepare compound C;

[0020] (3) Under argon protection, compound C and diethylaminotrifluoride were reacted with dichloromethane as solvent to prepare Aβ patch dual-mode imaging probe.

[0021] The compound A is

[0022] The compound X comprises 14-bromo-3,6,9,12-tetraoxatetradecane-1-ol, 2-(2-bromoethoxy)ethanol, 13-bromo-2,5,8,11-tetraoxatridecane, or 2-bromoethylmethyl ether.

[0023] The compound Y contains 5-(4-dimethylaminophenyl)thiophene-2-carboxaldehyde or 5-(4-(2-hydroxyethylmethylamino)phenyl)thiophene-2-carboxaldehyde.

[0024] Preferably, in step (1), the molar ratio of compound A, compound X and sodium ethoxide is 1:1 to 2:0.8 to 1.2, the reaction is carried out at room temperature, and the reaction time is 1 to 3 hours.

[0025] In step (2), the molar ratio of compound B to compound Y is 1:0.8 to 1.2, the reaction temperature is 100 to 120°C, and the reaction time is 2 to 4 hours.

[0026] Preferably, in step (3), the molar ratio of compound C to diethylaminotrifluoride is 1:0.8 to 1.2, the reaction temperature is -80 to -75°C, and the reaction time is 1 to 3 hours.

[0027] The present invention also provides an application of the above-described Aβ patch dual-mode imaging probe in the preparation of Aβ patch imaging reagents.

[0028] The beneficial effects of this invention are:

[0029] This invention uses quinoline-malononitrile as the fluorescent dye parent material, dimethylaminobenzene and other units as Aβ patch-binding groups, and introduces thiophene and other units as π-bridges to extend the probe emission wavelength, and uses... 18 F was used as a PET imaging nuclide to prepare a dual-mode imaging probe for Aβ plaques. This probe has the advantages of long fluorescence emission wavelength, high affinity, good selectivity, strong stability and excellent biocompatibility. It can realize the staining and labeling of Aβ plaques and accurately reflect the distribution, morphology and size of the plaques.

[0030] In fluorescence imaging, the probe exhibits weak fluorescence emission in phosphate buffer; however, the fluorescence signal is significantly enhanced to 40-fold when Aβ protein is present in the phosphate buffer. Furthermore, the probe displays a "spot-light" response and excellent blood-brain barrier penetration, enabling it to significantly identify Aβ plaques in 5xFAD mice. In PET imaging, the stable signal of the probe allows for effective imaging over a relatively long time window. In summary, this type of probe enables long-term tracking of Aβ plaques with a high signal-to-noise ratio through dual-mode fluorescence / PET imaging. Attached Figure Description

[0031] Figure 1 The fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Formula I-1 prepared in Example 1 in different proportions of glycerol / water mixed solvents.

[0032] Figure 2 The dual-mode imaging probe for Aβ patches with the structure shown in Formula I-1, prepared in Example 1, and different concentrations of Aβ were used. 42 Fluorescence spectra of protein aggregates after binding;

[0033] Figure 3 The fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Formula I-2 prepared in Example 2 in different proportions of glycerol / water mixed solvents;

[0034] Figure 4 The dual-mode imaging probe for Aβ patches with the structure shown in Formula I-2, prepared in Example 2, and different concentrations of Aβ were used. 42 Fluorescence spectra of protein aggregates after binding;

[0035] Figure 5 The fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Formula II-1 prepared in Example 3 in different proportions of glycerol / water mixed solvents.

[0036] Figure 6 The dual-mode imaging probe for Aβ patches with the structure shown in Formula II-1, prepared in Example 3, and different concentrations of Aβ were used. 42 Fluorescence spectra of protein aggregates after binding;

[0037] Figure 7 The fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Formula II-2 prepared in Example 4 in different proportions of glycerol / water mixed solvents.

[0038] Figure 8 The dual-mode imaging probe for Aβ patches with the structure shown in Formula II-2, prepared in Example 4, and different concentrations of Aβ were used. 42 Fluorescence spectra of protein aggregates after binding;

[0039] Figure 9 This is a comparison of the cytotoxicity of the Aβ plaque dual-mode imaging probe with the structure shown in Formula I-1 prepared in Example 1 to HepG2 cells at different concentrations.

[0040] Figure 10 The dual-mode imaging probe for Aβ plaques with the structure shown in Formula I-1, prepared in Example 1, is used to stain Aβ plaques on brain tissue sections of 5xFAD transgenic mice, wherein adjacent sections are stained with thiosulfate S to confirm the distribution of plaques.

[0041] Figure 11 The image shows a fluorescence imaging pattern of an Aβ patch using a dual-mode imaging probe of Formula I-1 prepared in Example 1.

[0042] Figure 12 The dual-mode imaging probe for Aβ patches with the structure shown in Formula I-1, prepared in Example 1, was used for positron emission tomography imaging of Aβ patches. Detailed Implementation

[0043] This invention provides a dual-mode imaging probe for Aβ patches, the dual-mode imaging probe for Aβ patches having the structure shown in Formula I:

[0044]

[0045] In Formula I, R1 is selected from cyano, carboxyl, or... R2 is selected from C1 to C4 hydrocarbon groups. Where n is an independent integer between 0 and 10, and F is selected from... 18 F or 19 F;R3 is selected from methyl, Where m is independently 0 or 1, and R5 is independently selected from -H, -CH3, -CH2OH, -CH2OTs, -CH218 F,-CH2 19 F, -CH2CH3, -CH2CH2OH, -CH2 CH2OTs, -CH2CH2 18 F or -CH2CH2 19 F;R4 is selected from -H, - at any position of the 5th, 6th, 7th, or 8th position of the quinoline group. 18 F or - 19 F. In this invention, the curve markings in each group indicate substitution sites.

[0046] In this invention, R1 is selected from cyano.

[0047] In this invention, R2 is selected from... Where n is an independent integer between 0 and 10, and F is selected from... 18 F or 19 F.

[0048] In this invention, R3 is selected from... Where m is independently 0 or 1, and R5 is independently selected from -H, -CH3, -CH2OH, -CH2OTs, -CH2 18 F,-CH2 19 F, -CH2CH3, -CH2CH2OH, -CH2 CH2OTs, -CH2CH2 18 F or -CH2CH2 19 F.

[0049] In this invention, when R2 is selected from... When n is an independent integer between 0 and 10, R3 is selected from... Where m is independently 0 or 1, and R5 is independently selected from -CH2OH, -CH2OTs, and -CH2 18 F、-CH2 19 F, -CH2CH2OH, -CH2CH2OTs, -CH2CH2 18 F or -CH2CH2 19 F.

[0050] In this invention, when R2 is selected from... When n is an independent integer between 0 and 10, F is selected from... 18 F or 19 F, R3 are selected from Where m is independently 0 or 1, and R5 is independently selected from -H, -CH3 or -CH2CH3.

[0051] In this invention, the Aβ patch dual-mode imaging probe has a structure as shown in Formula I-1, I-2, II-1, or II-2:

[0052]

[0053] This invention provides a method for preparing the above-described Aβ patch dual-mode imaging probe, comprising the following steps:

[0054] (1) Under argon protection, N,N-dimethylformamide was used as a solvent to react compound A, compound X and sodium ethoxide to prepare compound B;

[0055] (2) Compound B and compound Y reacted under argon protection conditions with toluene as solvent to prepare compound C;

[0056] (3) Under argon protection, compound C and diethylaminotrifluoride were reacted with dichloromethane as solvent to prepare Aβ patch dual-mode imaging probe.

[0057] The compound A is

[0058] The compound X comprises 14-bromo-3,6,9,12-tetraoxatetradecane-1-ol, 2-(2-bromoethoxy)ethanol, 13-bromo-2,5,8,11-tetraoxatridecane, or 2-bromoethylmethyl ether.

[0059] The compound Y contains 5-(4-dimethylaminophenyl)thiophene-2-carboxaldehyde or 5-(4-(2-hydroxyethylmethylamino)phenyl)thiophene-2-carboxaldehyde.

[0060] In this invention, in step (1), the molar ratio of compound A, compound X and sodium ethoxide is 1:1 to 2:0.8 to 1.2, preferably 1:1:1 or 1:2:1; the reaction is carried out at room temperature for 1 to 3 hours, preferably 2 hours.

[0061] In step (2), the molar ratio of compound B to compound Y is 1:0.8 to 1.2, preferably 1:1, the reaction temperature is 100 to 120°C, preferably 110°C, and the reaction time is 2 to 4 hours, preferably 3 hours.

[0062] In this invention, in step (3), the molar ratio of compound C to diethylaminotrifluoride is 1:0.8 to 1.2, preferably 1:1; preferably, compound C is added to dichloromethane solvent, and then diethylaminotrifluoride is added dropwise to carry out the reaction, the reaction temperature is -80 to -75°C, preferably -78°C, and the reaction time is 1 to 3 hours, preferably 2 hours.

[0063] The present invention also provides an application of the above-described Aβ patch dual-mode imaging probe in the preparation of Aβ patch imaging reagents.

[0064] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0065] Example 1

[0066] Compound A (100 mg, 0.40 mmol), 14-bromo-3,6,9,12-tetraoxatetradecane-1-ol (120 mg, 0.40 mmol), and sodium ethoxide (27 mg, 0.40 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of DMF as solvent, the mixture was stirred at 25 °C for 2 h under argon protection. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 74 mg of a yellow oily substance (compound B-1), with a yield of 39%.

[0067]

[0068] 1 H NMR (400MHz, CDCl3, ppm) δ: 9.10 (d, J=8.4Hz, 1H, Ph-H), 7.73 (t, J=8.0Hz, 1H, P h-H),7.63(d,J=8.8Hz,1H,Ph-H),7.44(t,J=7.7Hz,1H,Ph-H),6.82(s,1H,Ph-H ),4.54(t,J=4.8Hz,2H,-NCH2CH2O-),3.89(t,J=4.8Hz,2H,-NCH2CH2O-),3.56-3.71(m,20H,PEG-H),2.65(s,3H,-CH3); Massspectrometry(NSI-MS,m / z):[MH] - calcd.for[C 25 H 33 O6N3-H] - 470.2286; found, 470.2290.

[0069]

[0070] Compound B-1 (72 mg, 0.15 mmol) and 5-(4-dimethylaminophenyl)thiophene-2-carboxaldehyde (35 mg, 0.15 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of toluene as solvent, the mixture was heated to 110 °C under argon protection and refluxed with stirring for 3 h. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 42 mg of a dark red solid (compound C-1), with a yield of 41%.

[0071] 1 H NMR (400MHz, CDCl3, ppm) δ: 9.11 (d, J = 8.4Hz, 1H, Ph-H), 7.73 (t, J = 7.9Hz, 1H, Ph-H), 7.64 (d, J = 8.8Hz, 1H, Ph-H), 7.50 (d ,J=8.6Hz,2H,Ph-H),7.44(t,J=7.8Hz,1H,Ph-H),7.38(d,J=15.4Hz,1H,alkene-H),7.19(d,J=3.8Hz,1H,thioph-H),7. 14(s,1H,Ph-H),7.12(d,J=15.4Hz,1H,alkene-H),7.12(d,J=3.8Hz,1H,thioph-H),6.71(d,J=8.6Hz,2H,Ph-H),4.55(t Mass spectrometry(ESI-MS,m / z):[M+H] + calcd.for[C 38 H 44 O6N4S+H] + 685.3060; found, 685.3058.

[0072]

[0073] Compound C-1 (27 mg, 0.04 mmol) was placed in a dry 50 mL round-bottom flask. Diethylaminosulfur trifluoride (6 mg, 0.04 mmol) was added dropwise under argon protection at -78 °C using 10 mL dichloromethane as solvent. The reaction was continued at -78 °C for 2 h. After the reaction was complete, the mixture was quenched with saturated sodium bicarbonate solution, extracted with dichloromethane, dried, and the solvent removed. The mixture was then purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to obtain 13 mg of a dark red solid, which is the Aβ patch dual-mode imaging probe with the structure shown in Formula I-1, with a yield of 48%.

[0074] 1 H NMR (400MHz, CDCl3, ppm) δ: 9.13 (d, J = 8.4Hz, 1H, Ph-H), 7.73 (t, J = 7.9Hz, 1H, Ph-H), 7.60 (d, J = 8.7Hz, 1H, Ph-H), 7.51 (d, J = 8.5Hz, 2H, Ph-H),7.45(t,J=7.7Hz,1H,Ph-H),7.40(d,J=15.5Hz,1H,alkene-H),7.20(d,J=3.6Hz,1H,thioph-H),7.16(s,1H,Ph-H),7.13(d,J=3. 6Hz,1H,thioph-H),7.11(d,J=15.5Hz,1H,alkene-H),6.73(d,J=7.6Hz,2H,Ph-H),4.54(t,J=4.8Hz,2H,-NCH2CH2O-),4.53(dt,J1=3.7 Mass spectrometry(ESI-MS,m / z):[M+H] + calcd.for[C 38 H 43 FN4O5S+H] + 687.3016; found, 687.3014.

[0075] Example 2

[0076] Compound A (100 mg, 0.40 mmol), 2-(2-bromoethoxy)ethanol (68 mg, 0.40 mmol), and sodium ethoxide (27 mg, 0.40 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of DMF as solvent, the mixture was stirred at room temperature (25 °C) for 2 h under argon protection. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 62 mg of a yellow oily substance (compound B-2), with a yield of 38%.

[0077]

[0078] 1 H NMR (400MHz, CDCl3, ppm) δ: 9.09 (d, J=8.4Hz, 1H, Ph-H), 7.74 (t, J=7.8Hz, 1H, Ph-H), 7.59 (d,J=8.8Hz,1H,Ph-H),7.44(t,J=7.8Hz,1H,Ph-H),6.81(s,1H,Ph-H),4.52(t,J=5.3Hz,2 H,-NCH2CH2O-),3.89(t,J=5.3Hz,2H,-NCH2CH2O-),3.70(t,J=4.2Hz,2H,-OCH2CH2OH),3. 58(m,4H,-CH2OCH2CH2OCH2-),3.53(t,J=4.2Hz,2H,-OCH2CH2OH),2.64(s,3H,-CH3); Mass spectrometry(ESI-MS,m / z):[MH] - calcd.for[C 19 H 21 O3N3-H] - 404.1581; found, 404.1582.

[0079]

[0080] Compound B-2 (40 mg, 0.12 mmol) and 5-(4-dimethylaminophenyl)thiophene-2-carboxaldehyde (28 mg, 0.12 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of toluene as solvent, the mixture was heated to 110 °C under argon protection and refluxed with stirring for 3 h. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 43 mg of a dark red solid (compound C-2), with a yield of 66%.

[0081] 1H NMR (400MHz, CDCl3, ppm) δ: 9.07 (d, J = 8.4Hz, 1H, Ph-H), 7.72 (t, J = 7.6Hz, 1H, Ph-H), 7.59 (d, J = 8.6Hz, 1H, Ph-H), 7.49 ( d,J=8.4Hz,2H,Ph-H),7.42(t,J=7.8Hz,1H,Ph-H),7.36(d,J=15.4Hz,1H,alkene-H),7.17(d,J=3.6Hz,1H,thioph-H),7 .10(s,1H,Ph-H),7.09(d,J=3.6Hz,1H,thioph-H),7.07(d,J=15.4Hz,1H,alkene-H),6.70(d,J=8.2Hz,2H,Ph-H),4.53( Mass spectrometry(ESI-MS,m / z):[M+H] + calcd.for[C 32 H 32 O3N4S+H] + 553.2268; found, 553.2265.

[0082]

[0083] Compound C-2 (30 mg, 0.04 mmol) was placed in a dry 50 mL round-bottom flask. Diethylaminosulfur trifluoride (6 mg, 0.04 mmol) was added dropwise under argon protection at -78 °C using 10 mL dichloromethane as solvent. The reaction was continued at -78 °C for 2 h. After the reaction was complete, the mixture was quenched with saturated sodium bicarbonate solution, extracted with dichloromethane, dried, and the solvent removed. The mixture was then purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to obtain 6 mg of a dark red solid, which is the Aβ patch dual-mode imaging probe with the structure shown in Formula I-2, with a yield of 25%.

[0084] 1H NMR (400MHz, CDCl3, ppm) δ: 9.12 (dd, J1=0.8Hz, J2=8.4Hz, 1H, Ph-H), 7.73 (t, J=7.4Hz, 1H, Ph-H), 7.59 (d, J=8.7Hz, 1H, Ph-H), 7.50 (d, J=8.7Hz,2H,Ph-H),7.44(t,J=7.6Hz,1H,Ph-H),7.39(d,J=15.4Hz,1H,alkene-H),7.19(d,J=3.8Hz,1H,thioph-H),7.15(s,1H,Ph-H), 7.12(d,J=3.8Hz,1H,thioph-H),7.10(d,J=15.4Hz,1H,alkene-H),6.72(d,J=8.6Hz,2H,Ph-H),4.54(t,J=5.2Hz,2H,-NCH2CH2O-),4.4 Mass spectrometry(ESI-MS,m / z):[M+H] + calcd.for[C 32 H 31 FN4O2S+H] + 555.2225; found, 555.2228.

[0085] Example 3

[0086] Compound A (100 mg, 0.40 mmol), 13-bromo-2,5,8,11-tetraoxatridecane (216 mg, 0.80 mmol), and sodium ethoxide (27 mg, 0.40 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of DMF as solvent, the mixture was stirred at 25 °C for 2 h under argon protection. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 143 mg of a yellow oily substance (compound B-3), with a yield of 81%.

[0087]

[0088] 1H NMR (400MHz, CDCl3, ppm) δ: 9.04 (dd, J1=1.3Hz, J2=8.5Hz, 1H, Ph-H), 7.74 (m, 1H, Ph-H), 7.62 (d, J=8.6Hz, 1H, Ph-H), 7.42 (m, 1H, Ph-H), 6.77 (s, 1H, Ph-H),4.54(t,J=5.4Hz,2H,-NCH2CH2-),3.90(t,J=5.4Hz,2H,-NCH2CH2-),3.52-3.66(m,16H,PEG-H),3.37(s,3H,-OCH3),2.65(s,3H,-CH3); Mass spectrometry(NSI-MS,m / z):[M+H] + calcd.for[C 24 H 31 N3O5+H] + 442.2336; found, 442.2329.

[0089]

[0090] Compound B-3 (285 mg, 0.65 mmol) and 5-(4-(2-hydroxyethylmethylamino)phenyl)thiophene-2-carboxaldehyde (169 mg, 0.65 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of toluene as solvent, the mixture was heated to 110 °C under argon protection and refluxed with stirring for 3 h. After removing the solvent, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 300 mg of a dark red solid (compound C-3), with a yield of 67%.

[0091] 1H NMR (400MHz, DMSO-d6, ppm) δ: 9.04 (dd, J1=1.2Hz, J2=8.4Hz, 1H, Ph-H), 8.05 (d, J=9.0Hz, 1H, Ph-H), 7.89 (m, 1H, Ph-H), 7.60 (t, J=7.8Hz, 1H ,Ph-H),7.57(d,J=15.7Hz,1H,alkene-H),7.51(d,J=8.9Hz,2H,Ph-H),7.46(d,J=3.9Hz,1H,thioph-H),7.33(d,J=3.9Hz,1H,thioph-H),7 .31(d,J=15.7Hz,1H,alkene-H),7.00(s,1H,Ph-H),6.73(d,J=9.0Hz,2H,Ph-H),4.73(t,J=5.4Hz,1H,-OH),4.71(t,J=6.0Hz,2H,-NCH2CH2 Mass spectrometry(ESI-MS,m / z):[M+Na] + calcd.for[C 38 H 44 N4O6S+Na] + 707.2874; found, 707.2849.

[0092]

[0093] Compound C-3 (300 mg, 0.42 mmol) was placed in a dry 50 mL round-bottom flask. Diethylaminosulfur trifluoride (63 mg, 0.42 mmol) was added dropwise under argon protection at -78 °C using 10 mL dichloromethane as solvent. The reaction was continued at -78 °C for 2 h. After the reaction was complete, the mixture was quenched with saturated sodium bicarbonate solution, extracted with dichloromethane, dried, and the solvent removed. The mixture was then purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to obtain 169 mg of a dark red solid, which is the Aβ patch dual-mode imaging probe with the structure shown in Formula II-1, with a yield of 59%.

[0094] 1H NMR (400MHz, CDCl3, ppm) δ: 9.12 (d, J = 8.4Hz, 1H, Ph-H), 7.73 (t, J = 8.0Hz, 1H, Ph-H), 7.60 (d, J = 8.8Hz, 1H, Ph-H), 7.50 (d, J = 8.5Hz, 2H, Ph-H), 7.44 ( t,J=7.7Hz,1H,Ph-H),7.38(d,J=15.4Hz,1H,alkene-H),7.19(d,J=4.0Hz,1H,thioph-H),7.14(d,J=4.0Hz,1H,thioph-H),7.13(s,1H,Ph-H),7.11( d,J=15.4Hz,1H,alkene-H),6.72(d,J=8.6Hz,2H,Ph-H),4.63(dt,J1=5.0Hz,J2=47.1Hz,2H,-NCH2CH2F),4.54(t,J=4.8Hz,2H,-NCH2CH2O-),4.00(t ,J=4.8Hz,2H,-NCH2CH2O-),3.49-3.75(m,18H,PEG-H,-NCH2CH2F),3.34(s,3H,-OCH3),3.08(s,3H,-NCH3); Massspectrometry(ESI-MS,m / z):[M+H] + calcd.for[C 38 H 43 FN4O5S+H] + 687.3016; found, 687.3014.

[0095] Example 4

[0096] Compound A (100 mg, 0.40 mmol), 2-bromoethyl methyl ether (111 mg, 0.80 mmol), and sodium ethoxide (27 mg, 0.40 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of DMF as solvent, the mixture was stirred at room temperature (25 °C) for 2 h under argon protection. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 65 mg of a yellow oily substance (compound B-4), with a yield of 53%.

[0097]

[0098] 1H NMR (400MHz, DMSO-d6, ppm) δ: 8.90 (d, J=8.4Hz, 1H, Ph-H), 8.06 (d, J=8.9Hz, 1H, Ph-H), 7.88 (t, J= 8.1Hz,1H,Ph-H),7.59(t,J=7.8Hz,1H,Ph-H),6.80(s,1H,Ph-H),4.66(t,J=5.1Hz,2H,-NCH2CH2O -),3.82(t,J=5.1Hz,2H,-NCH2CH2O-),3.46(t,J=4.2Hz,2H,-OCH2CH2OCH3),3.32(t,J=4.2Hz,2H ,-OCH2CH2OCH3),3.12(s,3H,-OCH3),2.70(s,3H,-CH3); Massspectrometry(NSI-MS,m / z):[M+H] + calcd.for[C 18 H 19 N3O2+H] + 310.1550; found, 310.1543.

[0099]

[0100] Compound B-4 (115 mg, 0.37 mmol) and 5-(4-(2-hydroxyethylmethylamino)phenyl)thiophene-2-carboxaldehyde (96 mg, 0.37 mmol) were placed in a dry 50 mL round-bottom flask. Using 10 mL of toluene as solvent, the mixture was heated to 110 °C under argon protection and refluxed with stirring for 3 h. After solvent removal, the mixture was purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to give 116 mg of a dark red solid (compound C-4), with a yield of 57%.

[0101]

[0102] Compound C-4 (116 mg, 0.21 mmol) was placed in a dry 50 mL round-bottom flask. Diethylaminosulfur trifluoride (31 mg, 0.21 mmol) was added dropwise under argon protection at -78 °C using 10 mL dichloromethane as solvent. The reaction was continued at -78 °C for 2 h. After the reaction was complete, the mixture was quenched with saturated sodium bicarbonate solution, extracted with dichloromethane, dried, and the solvent removed. The mixture was then purified by silica gel column chromatography (developing solvent: dichloromethane:methanol = 25:1) to obtain 72 mg of a dark red solid, which is the Aβ patch dual-mode imaging probe with the structure shown in Formula II-2, with a yield of 59%.

[0103] 1H NMR (400MHz, CDCl3, ppm) δ: 9.09 (d, J = 8.2Hz, 1H, Ph-H), 7.72 (t, J = 7.6Hz, 1H, Ph-H), 7.58 (d, J = 8.8Hz, 1H, Ph-H), 7.50 (d, J = 8.8Hz, 2H, Ph-H), 7.43 (t, J = 7 .8Hz,1H,Ph-H),7.37(d,J=15.4Hz,1H,alkene-H),7.18(d,J=3.8Hz,1H,thioph-H),7.13(d,J=15.4Hz,1H,alkene-H),7.12(s,1H,Ph-H),7.11(d,J=3.8H z,1H,thioph-H),4.63(dt,J1=5.1Hz,J2=47.1Hz,2H,-NCH2CH2F),4.54(t,J=5.2Hz,2H,-NCH2CH2O-),4.00(t,J=5.2Hz,2H,-NCH2CH2O-),3.70(dt,J1=5. Mass spectrometry(NSI-MS,m / z):[M+H] + calcd.for[C 21 H 31 FN4O2S+H] + 555.2225; found, 555.2236.

[0104] Experimental Example 1

[0105] The fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Formula I-1 were tested in different viscosity environments: Formula I-1 was dissolved in analytical grade dimethyl sulfoxide to prepare a 1.0 × 10⁻⁶ m² / m² solution. -3 A stock solution of 1 mol / L was prepared. Then, 2 mL of glycerol / water mixed solvents with glycerol contents of 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, and 90% were prepared. 20 μL of the I-1 stock solution was added to each of the prepared glycerol / water mixed solvents, mixed thoroughly, and then transferred to quartz cuvettes (10 mm × 10 mm) to measure their fluorescence spectra. The results are shown below. Figure 1As shown, with an excitation wavelength of 470 nm, the maximum emission wavelength of the Aβ patch dual-mode imaging probe with the structure shown in Formula I-1 is approximately 720 nm, located in the near-infrared region. Furthermore, the fluorescence intensity significantly increases with increasing glycerol content in the mixed solvent, i.e., increasing viscosity.

[0106] Experiment Example 2

[0107] Test Aβ patch dual-mode imaging probe with structure shown in Equation I-1 and different concentrations of Aβ 42 Fluorescence spectra of protein aggregate responses: A concentration of 1.0 × 10⁻⁶ -3 The stock solution of I-1 was diluted to obtain a concentration of 1.0 × 10⁻⁶ mol / L. -4 A stock solution of mol / L. Then prepare Aβ. 42 The protein aggregate concentration was 2.0 × 10⁻⁶. -4 A mol / L stock solution was prepared. Then, 10 μL, 20 μL, 30 μL, 40 μL, 50 μL, 60 μL, 70 μL, 80 μL, 90 μL, and 100 μL of the above Aβ solution were added to 1.99 mL, 1.98 mL, 1.97 mL, 1.96 mL, 1.95 mL, 1.94 mL, 1.93 mL, 1.92 mL, 1.91 mL, and 1.90 mL of 10 mM phosphate buffer (pH 7.4), respectively. 42 Add 20 μL of a 1.0 × 10⁻⁶ solution to the stock solution of the protein aggregates. -4 The stock solution of mol / L I-1 was incubated with gentle shaking at 37℃ for 10 min, and its fluorescence spectrum was measured. The test results are as follows: Figure 2 As shown, with an excitation wavelength of 470 nm, I-1 and Aβ 42 The protein aggregate shows almost no fluorescence before binding, but when I-1 binds to Aβ... 42 After binding to the protein aggregates, there was a significant fluorescence enhancement at 657 nm, and this enhancement increased with the increase of Aβ. 42 As the concentration of protein aggregates increases, the fluorescence intensity of I-1 increases.

[0108] Experimental Example 3

[0109] Using the same method as in Experimental Example 1, the fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Equation I-2 were tested in different viscosity environments. The test results are as follows: Figure 3 As shown, with an excitation wavelength of 470 nm, the maximum emission wavelength of I-2 is approximately 720 nm, located in the near-infrared region. Furthermore, the fluorescence intensity significantly increases with increasing glycerol content in the mixed solvent, i.e., increasing viscosity.

[0110] Experiment Example 4

[0111] Using the same method as in Experimental Example 2, the dual-mode imaging probe for Aβ patches with the structure shown in Equation I-2 was tested in relation to different concentrations of Aβ. 42 The fluorescence spectrum of the protein aggregate response, the test results are as follows: Figure 4 As shown, with an excitation wavelength of 470 nm, I-2 and Aβ 42 The protein aggregate shows almost no fluorescence before binding, but when I-2 binds to Aβ... 42 After binding to the protein aggregates, there was a significant fluorescence enhancement at 657 nm, and this enhancement increased with the increase of Aβ. 42 As the concentration of protein aggregates increases, the fluorescence intensity of I-2 increases.

[0112] Experimental Example 5

[0113] Using the same method as in Experimental Example 1, the fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Equation II-1 were tested in different viscosity environments. The test results are as follows: Figure 5 As shown, with an excitation wavelength of 470 nm, the maximum emission wavelength of II-1 is approximately 720 nm, located in the near-infrared region. Furthermore, the fluorescence intensity significantly increases with increasing glycerol content in the mixed solvent, i.e., increasing viscosity.

[0114] Experimental Example 6

[0115] Using the same method as in Experimental Example 2, the dual-mode imaging probe for Aβ patches with the structure shown in Formula II-1 was tested in relation to different concentrations of Aβ. 42 The fluorescence spectrum of the protein aggregate response, the test results are as follows: Figure 6 As shown, with an excitation wavelength of 470 nm, Ⅱ-1 and Aβ 42 The protein aggregate shows almost no fluorescence before binding, but when II-1 binds to Aβ... 42 After binding to the protein aggregates, there was a significant fluorescence enhancement at 657 nm, and this enhancement increased with the increase of Aβ. 42 As the concentration of protein aggregates increases, the fluorescence intensity of II-1 increases.

[0116] Experimental Example 7

[0117] Using the same method as in Experimental Example 1, the fluorescence spectra of the Aβ patch dual-mode imaging probe with the structure shown in Equation II-2 were tested in different viscosity environments. The test results are as follows: Figure 7 As shown, with an excitation wavelength of 470 nm, the maximum emission wavelength of II-2 is approximately 720 nm, located in the near-infrared region. Furthermore, the fluorescence intensity significantly increases with increasing glycerol content in the mixed solvent, i.e., increasing viscosity.

[0118] Experimental Example 8

[0119] Using the same method as in Experimental Example 2, the dual-mode imaging probe for Aβ patches with the structure shown in Formula II-2 was tested in relation to different concentrations of Aβ.42 The fluorescence spectrum of the protein aggregate response, the test results are as follows: Figure 8 As shown, with an excitation wavelength of 470 nm, Ⅱ-2 and Aβ 42 The protein aggregate shows almost no fluorescence before binding, but when II-2 binds to Aβ... 42 After binding to the protein aggregates, there was a significant fluorescence enhancement at 657 nm, and this enhancement increased with the increase of Aβ. 42 Increased concentration of protein aggregates leads to enhanced fluorescence intensity of II-2.

[0120] Experimental Example 9

[0121] The cytotoxicity of the Aβ plaque dual-mode imaging probe with the structure shown in Formula I-1 was tested: HepG2 cells were seeded in 96-well plates at a density of 1.0 × 10⁶ cells / well. 4 Cells / wells were cultured in an incubator for 8 hours. Culture medium containing I-1 (concentrations of 0 μM, 1 μM, 2 μM, 4 μM, 8 μM, 16 μM, 32 μM, 50 μM, 75 μM, 100 μM, and 100 μL) was added to different wells, and the cells were cultured for another 24 hours. The culture medium was then aspirated from each well with a needle, and 100 μL of 0.5 mg / mL MTT solution was added. After incubation for 4 hours, the culture medium was aspirated, and 100 μL of dimethyl sulfoxide was added to each well. The absorbance at 590 nm was measured using a microplate reader. Cell viability was calculated using the formula: (OD...) treated / OD control )×100%. The result is as follows: Figure 9 As shown, when the concentration of I-1 reaches 100 μM, the cell survival rate remains above 80%, indicating that the Aβ plaque dual-mode imaging probe with the structure shown in Formula I-1 has low cytotoxicity.

[0122] Experimental Example 10

[0123] The Aβ plaque dual-mode imaging probe with the structure shown in Formula I-1 was used for fluorescence staining of brain tissue sections from 5xFAD transgenic mice: Brain tissue sections from 5xFAD transgenic mice were dewaxed and hydrated, then 100 μL of a 100 μM aqueous solution of Formula I-1 was added to the tissue sections. After incubation at room temperature for 10 min, the solution was carefully blotted away from the edges with clean paper, and the sections were then observed under a microscope. Adjacent sections were stained with the gold standard dye thiosulfinate S to determine the plaque distribution. The results are as follows: Figure 10 As shown, due to the inherent aggregation and quenching properties of thiosulfate S, some Aβ plaques exhibit insufficient fluorescence signal and inadequate contrast with the background, easily leading to unavoidable false negative results. In contrast, I-1 possesses excellent anti-quenching properties, enabling high-fidelity display of Aβ plaque information on brain tissue slices from 5xFAD transgenic mice, significantly improving signal quality and detection accuracy.

[0124] Experimental Example 11

[0125] The dual-mode imaging probe for Aβ patches with the structure shown in Formula I-1 was tested for Aβ patch fluorescence imaging. The 10-month-old 5xFAD transgenic mice and the control group normal mice used in this experiment were purchased from Nanjing Junke Biotechnology Co., Ltd. The mice were housed in sterile cages in a laminar flow fume hood in a sterile room and were fed with food and water that had been treated with high pressure steam.

[0126] Ten-month-old 5xFAD transgenic mice and control mice were injected intravenously with 2.3 mg / kg of I-1, followed by imaging using a PerkinElmer IVIS Spectrum CT small animal in vivo imaging system. During imaging, the mice were anesthetized with an oxygen flow of 2.0 L / min containing 2.0% isoflurane. The excitation and reception wavelengths were set to 500 nm and 700 nm, respectively. The results are as follows: Figure 11 As shown, before injection of I-1, the fluorescence signals in 5xFAD transgenic mice and control normal mice were weak. Thirteen hours after injection of I-1, the fluorescence signal in the brains of 5xFAD transgenic mice was significantly higher than that in control normal mice. This indicates that I-1 possesses excellent blood-brain barrier penetration ability and a "spot-lighting" response characteristic, enabling the imaging of Aβ plaques through fluorescence patterns.

[0127] Experimental Example 12

[0128] The dual-mode imaging probe for Aβ plaques with the structure shown in Formula I-1 was tested for positron emission tomography imaging of Aβ plaques: 10-month-old 5xFAD transgenic mice and control normal mice were injected intravenously with 10 MBq. 18 F-labeled I-1 was then imaged using the Inviscan IRIS PET-CT small animal in vivo imaging system. During the imaging process, the mice were anesthetized with an oxygen flow of 2.0 L / min containing 2.0% isoflurane. Results are as follows... Figure 12 As shown, during injection 18 After 12 minutes of F-labeled I-1, the signal intensity in the hippocampus, cortex and other regions of the brain of 5xFAD transgenic mice was significantly higher than that of normal control mice, indicating that I-1 can image Aβ plaques by positron emission tomography.

[0129] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A dual-mode imaging probe for Aβ patches, characterized in that, The Aβ patch dual-mode imaging probe has the structure shown in Formula I: Ⅰ Where n is an integer between 2 and 10, and F is... 18 F.

2. The Aβ patch dual-mode imaging probe according to claim 1, characterized in that, The Aβ patch dual-mode imaging probe has a structure as shown in Formula I-1 or I-2: 。 3. The method for preparing the Aβ patch dual-mode imaging probe according to claim 2, characterized in that, Includes the following steps: (1) Compound B was prepared by reacting compound A, compound X and sodium ethoxide under argon protection with N,N-dimethylformamide as solvent. (2) Compound B and compound Y reacted under argon protection conditions with toluene as solvent to prepare compound C; (3) Under argon protection, compound C and diethylaminotrifluoride were reacted with dichloromethane as solvent to prepare Aβ patch dual-mode imaging probe. The compound A is ; The compound X is 14-bromo-3,6,9,12-tetraoxatetradecane-1-ol or 2-(2-bromoethoxy)ethanol; The compound Y is 5-(4-dimethylaminophenyl)thiophene-2-carboxaldehyde.

4. The method for preparing the Aβ patch dual-mode imaging probe according to claim 3, characterized in that, In step (1), the molar ratio of compound A, compound X and sodium ethoxide is 1:1~2:0.8~1.2, the reaction is carried out at room temperature, and the reaction time is 1~3h; In step (2), the molar ratio of compound B to compound Y is 1:0.8~1.2, the reaction temperature is 100~120℃, and the reaction time is 2~4h.

5. The method for preparing the Aβ patch dual-mode imaging probe according to claim 3 or 4, characterized in that, In step (3), the molar ratio of compound C to diethylaminotrifluoride is 1:0.8~1.2, the reaction temperature is -80~-75℃, and the reaction time is 1~3h.

6. The use of the Aβ patch dual-mode imaging probe according to claim 1 or 2 in the preparation of Aβ patch imaging reagents.

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