Zwitterionic cyanine dye as well as preparation method and application thereof

Abstract

The invention relates to zwitterionic cyanine dye as well as a preparation method and application thereof. According to the invention, a sulfonic acid group is introduced to a specific position of an existing cyanine dye, so that the cyanine dye can keep good hydrophilicity, charge distribution of cyanine dye molecules is more balanced, and the protein binding rate is lower, thereby improving the metabolic rate when the cyanine dye is used as a kidney clearance type probe, and further realizing ureter imaging. Meanwhile, the zwitterionic cyanine dye disclosed by the invention is high in kidney metabolism rate, and in-vivo metabolism can be completed within about 6 hours, so that the biological safety is higher.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technology, and more specifically to a zwitterionic cyanine dye and a preparation method and application thereof. Background Art

[0002] The urinary system is an important pathway for human metabolism and excretion, and is mainly composed of the kidneys, ureters, bladder, and urethra. During the excretion process, abnormal metabolic waste, foreign microorganisms, and drugs can easily cause infection and damage to the kidneys, ureters, and bladder, leading to a series of urinary system diseases. In recent years, the widespread use of ultrasound, computerized tomography (CT), magnetic resonance imaging (MRI), and other examination methods has enabled people to image the tissue structure of the urinary system and detect tiny tumors in the urinary system at an early stage, but it also poses challenges to clinicians while achieving early diagnosis. In addition, the surgeon mainly relies on visual information to perform clinical operations during the operation. Increasing the contrast between tissues and achieving precise intraoperative operations has become another problem that needs to be solved urgently.

[0003] As a new imaging method, near-infrared fluorescence imaging technology has been widely used in sentinel lymph node imaging, tumor localization, and vascular imaging. After near-infrared fluorescent dyes are injected intravenously or locally, they are gathered in specific parts. Under the irradiation of excitation light with a wavelength of 700-900nm, near-infrared fluorescent dyes will emit fluorescence that is longer than the wavelength of the incident light and invisible to the naked eye. This fluorescence can be captured by near-infrared fluorescence imaging equipment. After entering the camera through a filter, the light signal is converted into an electronic signal through data processing, and the strength of the signal is finally displayed on the screen as a grayscale value. Biological tissues have the weakest absorption of light quanta in this band. Compared with traditional imaging methods, near-infrared fluorescence imaging can achieve real-time imaging during surgery, providing intuitive visual guidance for the surgeon, and because tissues have little absorption and scattering effects on light within this wavelength, it has strong tissue penetration (1-2 cm) and no hazards such as ionizing radiation.

[0004] Hydrophilic fluorescent substances can be excreted from the body through the urinary system, and fluorescent imaging technology can be used to image organs to determine whether there are lesions in the organs. However, in near-infrared fluorescence imaging of the urinary system, unlike the kidneys and bladder, the ureters are mainly used for transportation and do not have storage functions, so it is difficult to capture the fluorescence information of the ureters. It is currently believed that if it is necessary to achieve ureteral imaging in vivo, the probe must have a high renal metabolic rate in addition to good hydrophilicity, because a high renal metabolic rate can allow the ureters to fully enrich the fluorescent signal, thereby achieving ureteral imaging.

[0005] In addition, the probes remain in the body for a long time and cannot be metabolized and cleared out of the body in time, which will cause certain toxicity to the circulatory system. Therefore, if the probes have a fast renal metabolism rate, that is, they remain in the body for a short time, their biosafety can be further improved.

[0006] Patent CN117186151A discloses a near-infrared zwitterionic cyanine dye, and points out that it can achieve dynamic monitoring of urinary system lesions and visual identification of lesion areas through real-time monitoring of fluorescence imaging, which is of great significance for the diagnosis of kidney damage, ureteral obstruction, etc. At the same time, with the help of real-time fluorescence imaging equipment, it can provide more accurate guidance for the precision treatment of ureteral surgery. However, from the experiment of its renal clearance efficiency test, it can be seen that it takes about 8 hours to be basically metabolized out of the body (that is, it takes about 8 hours to reach the highest urine recovery rate in the renal clearance efficiency test). The metabolic rate is still slow, making it difficult to fully enrich the signal in the ureter to achieve ureteral imaging. In addition, further shortening its metabolic time can improve its biological safety. Summary of the invention

[0007] The primary purpose of the present invention is to overcome the problem that the renal metabolic rate of the current renal-cleared fluorescent probe needs to be further improved and the existing renal-cleared fluorescent probe is difficult to achieve ureteral imaging, and provide a zwitterionic cyanine dye. The present invention introduces a sulfonic acid group at a specific position of the existing cyanine dye, which not only allows the cyanine dye to maintain good hydrophilicity, but also makes the charge distribution of the cyanine dye molecule more balanced and the protein binding rate lower, thereby improving its metabolic rate when used as a renal-cleared probe, and then can achieve ureteral imaging. At the same time, since the renal metabolic rate of the zwitterionic cyanine dye of the present invention is fast, the metabolism in the body is completed in about 6 hours, so the biological safety is higher.

[0008] A further object of the present invention is to provide a method for preparing the zwitterionic cyanine dye.

[0009] Another object of the present invention is to provide the use of the above-mentioned zwitterionic cyanine dye or its pharmaceutically acceptable salt, solvate, enantiomer, diastereomer, tautomer in the preparation of fluorescent probes.

[0010] Another object of the present invention is to provide a fluorescent probe comprising the above-mentioned zwitterionic cyanine dye and / or its pharmaceutically acceptable salt, solvate, enantiomer, diastereomer, tautomer.

[0011] The above-mentioned object of the present invention is achieved by the following technical solutions: The present invention provides a zwitterionic cyanine dye having a structure shown in any one of formulas (I) to (IV): , Wherein, Y is F, Cl, Br or I; X is NH, SH or O; n is an integer of 0 to 30; and n1 is an integer of 2 to 30.

[0012] The present invention is in existing cyanine dye The sulfonic acid group is introduced at a specific position of the cyanine dye, which not only allows the cyanine dye to maintain good hydrophilicity, but also makes the charge distribution of the cyanine dye molecule more balanced and the protein binding rate lower, thereby increasing its metabolic rate when used as a renal clearance probe, thereby enabling ureteral imaging. At the same time, since the zwitterionic cyanine dye of the present invention has a fast renal metabolism rate and completes metabolism in the body in about 6 hours, it has higher biological safety.

[0013] In addition, the zwitterionic cyanine dye of the present invention is a single molecule, label-free, zwitterion-neutral, has high specificity, high sensitivity, good biocompatibility and good optical stability, and can be rapidly metabolized by the kidneys without being nonspecifically taken up by other tissues or organs. It has good specificity and can play an important role in future medical optical examinations, and has excellent application prospects.

[0014] In addition, the present invention can also introduce a carboxyl group (Formula (II) or Formula (III)) or a PEG-carboxyl group (Formula (IV)) at a specific position of the six-membered ring of the cyanine dye mother core structure. The carboxyl group can undergo a condensation reaction with an amino acid, thereby providing more modifiable sites for the subsequent targeted modification of the fluorescent probe. By introducing different functional groups (such as oligopeptides or polypeptide sequences), the probe can have higher specificity and targeting, and can be further used for tracking and detecting cancer. The PEG chain can not only improve the solubility, stability and biocompatibility of the drug, but also improve the targeting of the drug by binding to a targeting ligand. In addition, the functionally modified PEG chain can be further used to construct a nanoscale drug delivery system.

[0015] Preferably, n is an integer from 1 to 30.

[0016] More preferably, n is an integer from 1 to 15.

[0017] More preferably, n is an integer of 1 to 3.

[0018] Preferably, n1 is an integer from 2 to 20.

[0019] Preferably, the zwitterionic cyanine dye has any of the following structures: , , , , or .

[0020] A method for preparing the above zwitterionic cyanine dye comprises the following steps: When the zwitterionic cyanine dye is of formula (I): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the zwitterionic cyanine dye; ; When the zwitterionic cyanine dye has the structure of formula (II): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the compound of formula (I); S2. The compound of formula (I) undergoes a condensation reaction with compound (2) to obtain the zwitterionic cyanine dye; ; When the zwitterionic cyanine dye is of formula (III): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the compound of formula (I); S3. The compound of formula (I) undergoes a condensation reaction with compound (5) to obtain the zwitterionic cyanine dye; ; When the zwitterionic cyanine dye is of formula (IV): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the compound of formula (I); S4. The compound of formula (I) undergoes a condensation reaction with compound (6) under anaerobic conditions to obtain the zwitterionic cyanine dye; .

[0021] The inventors of the present invention have found that the condensation reaction needs to be carried out under anaerobic conditions in order to successfully introduce the PEG flexible segment into the rigid six-membered ring of the cyanine dye core structure.

[0022] Preferably, in step S1, the reaction temperature of the condensation reaction is 80-100° C., and the reaction time is 12-24 h.

[0023] More preferably, in step S1, the solvent for the condensation reaction is methanol, and the catalyst is acetic anhydride and sodium acetate.

[0024] Preferably, in steps S2 to S4, the reaction temperature of the condensation reaction is 65 to 85° C., and the reaction time is 6 to 12 h.

[0025] Preferably, in steps S2 to S4, the condensation reaction is carried out in the presence of a solvent, and the solvent is at least one of water or N,N-dimethylformamide.

[0026] Preferably, in steps S2 to S4, the condensation reaction is carried out in the presence of an activator, and the activator is selected from one or more of potassium carbonate, cesium carbonate, sodium acetate, potassium acetate or triethylamine.

[0027] Preferably, the compound (14) is obtained by a substitution reaction between the compound (12) and the compound (13); .

[0028] The inventors of the present invention have found that in the process of preparing the zwitterionic cyanine dye of the present invention, due to the steric effect and the influence of charge distribution, it is necessary to modify the terminal of compound (12) to an iodine atom in order to successfully Introduced middle.

[0029] More preferably, the substitution reaction is carried out in the presence of sodium iodide, the reaction temperature is 110-130° C., and the solvent is one or both of o-dichlorobenzene and toluene.

[0030] More preferably, the compound (12) is obtained by a substitution reaction between compound (11) and sodium iodide.

[0031] More preferably, the solvent for the substitution reaction is acetonitrile, the reaction temperature is 60-70° C., and the reaction time is 8-12 h.

[0032] More preferably, the compound (11) is obtained by condensation reaction of compound (9) and compound (10).

[0033] Further preferably, the condensation reaction is carried out in the presence of sodium hydroxide at a reaction temperature of 70-90°C.

[0034] The structures of the compounds involved are as follows:

[0035] Preferably, compound (13) is obtained by condensation reaction of compound (16) and compound (17).

[0036] More preferably, the condensation reaction is carried out in the presence of acetic acid at a reaction temperature of 110-130°C.

[0037] The structures of the compounds involved are as follows:

[0038] The present invention also protects the use of the above zwitterionic cyanine dye or its pharmaceutically acceptable salt, solvate, enantiomer, diastereomer, tautomer in the preparation of fluorescent probes.

[0039] Preferably, the fluorescent probe is a fluorescent probe for organ imaging, and the organ is a kidney, a ureter or a bladder.

[0040] Preferably, the pharmaceutically acceptable salt is hydrochloride, hydrobromide, nitrate, methylnitrate, sulfate, bisulfate, aminosulfate, phosphate, acetate, glycolate, phenylacetate, propionate, butyrate, isobutyrate, valerate, maleate, hydroxymaleate, acrylate, fumarate, malate, tartrate, citrate, salicylate, para-aminosalicylate, glycolate, lactate, enanthate, phthalate, oxalate, succinate, benzoate, o-acetyl At least one of oxybenzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, mandelate, tannate, formate, stearate, ascorbate, palmitate, oleate, pyruvate, bishydroxynaphthoate, malonate, laurate, glutarate, glutamate, propionate lauryl sulfate, methanesulfonate, ethanesulfonate, 2-hydroxyethanesulfonate, benzenesulfonate, p-aminobenzenesulfonate, p-toluenesulfonate (toluenesulfonate) or naphthalene-2-sulfonate.

[0041] The present invention also protects a fluorescent probe, which contains the above-mentioned zwitterionic cyanine dye and / or its pharmaceutically acceptable salt, solvate, enantiomer, diastereomer, tautomer.

[0042] Compared with the prior art, the present invention has the following beneficial effects: The present invention introduces a sulfonic acid group at a specific position of an existing cyanine dye, which not only allows the cyanine dye to maintain good hydrophilicity, but also makes the charge distribution of the cyanine dye molecule more balanced and the protein binding rate lower, thereby increasing its metabolic rate when used as a kidney-cleared probe, thereby enabling ureteral imaging. At the same time, since the zwitterionic cyanine dye of the present invention has a fast renal metabolic rate and completes metabolism in the body in about 6 hours, it has higher biological safety.

[0043] In addition, the zwitterionic cyanine dye of the present invention is a single molecule, label-free, zwitterion-neutral, has high specificity, high sensitivity, good biocompatibility and good optical stability, and can be rapidly metabolized by the kidney without being nonspecifically taken up by other tissues or organs. It has good specificity and can more accurately perform early diagnosis of diseases, intraoperative navigation treatment, tissue and organ function evaluation, etc. It can play an important role in future medical optical examinations and has excellent application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 The ultraviolet absorption spectrum and fluorescence emission spectrum of the zwitterionic cyanine dye of Example 2 are shown.

[0045] Figure 2 The ultraviolet absorption spectrum and fluorescence emission spectrum of the zwitterionic cyanine dye of Example 3 are shown.

[0046] Figure 3 The figure is a statistical graph of the recovery rate of the zwitterionic cyanine dye in urine within 24 hours after injection of the zwitterionic cyanine dye in Example 2, wherein Figure 3 A in the figure is a curve showing the change of urine recovery rate over time. Figure 3 Figure B shows the results of three repeated experiments on urine recovery rate.

[0047] Figure 4 The figure is a statistical graph of the recovery rate of the zwitterionic cyanine dye in urine within 24 hours after injection of the zwitterionic cyanine dye in Example 3, wherein Figure 4 A in the figure is a curve showing the change of urine recovery rate over time. Figure 4 Figure B shows the results of three repeated experiments on urine recovery rate.

[0048] Figure 5 This is an organ imaging diagram of the fluorescent probe prepared by the zwitterionic cyanine dye of Example 2 in normal mice; wherein He represents the heart, Li represents the liver, Sp represents the spleen, Lu represents the lung, Ki represents the kidney, Bl represents the bladder, Sk represents the skin, Mu represents the muscle, and In represents the small intestine.

[0049] Figure 6 This is a quantitative diagram of organ imaging of the fluorescent probe prepared with the zwitterionic cyanine dye of Example 2 in normal mice; wherein He represents heart, Li represents liver, Sp represents spleen, Lu represents lung, Ki represents kidney, Bl represents bladder, Sk represents skin, Mu represents muscle, and In represents small intestine.

[0050] Figure 7This is an organ imaging diagram of the fluorescent probe prepared by the zwitterionic cyanine dye of Example 3 in normal mice; wherein He represents the heart, Li represents the liver, Sp represents the spleen, Lu represents the lung, Ki represents the kidney, Bl represents the bladder, Sk represents the skin, Mu represents the muscle, and In represents the small intestine.

[0051] Figure 8 This is a quantitative image of organ imaging of the fluorescent probe prepared with the zwitterionic cyanine dye of Example 3 in normal mice; wherein He represents the heart, Li represents the liver, Sp represents the spleen, Lu represents the lung, Ki represents the kidney, Bl represents the bladder, Sk represents the skin, Mu represents the muscle, and In represents the small intestine.

[0052] Fig. 9 A in the figure is the zwitterionic cyanine dye ureteral imaging result diagram of Example 2, Fig. 9 B in the figure is the zwitterionic cyanine dye ureteral imaging result of Example 3. DETAILED DESCRIPTION

[0053] In order to more clearly and completely describe the technical solution of the present invention, the present invention is further described in detail by specific examples below. It should be understood that the specific examples described herein are only used to explain the present invention and are not used to limit the present invention. Various changes can be made within the scope of the present invention. Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0054] Example 1 This embodiment provides a zwitterionic cyanine dye, and its synthesis route is as follows:

[0055] The specific steps include: 1) Take a 100 mL double-necked round-bottom flask, under argon atmosphere, add compound 9 (CAS: 5407-04-5, 10.3 g, 65 mmol), compound 10 (CAS: 1120-71-4, 8 mL, 97.5 mmol), solvent (1,2-dichloroethane, 6 mL) and sodium hydroxide (2.6 g, 65 mmol), heat to 70 ° C and stir overnight. After the reaction is completed, discard the supernatant and collect the white viscous liquid to obtain compound 11 (9.5 g, 60%). LRMS (ESI) m / z: [MH]- Calcd for C8H18ClNO3S243.07; Found 243.75.

[0056] 2) In a 100 mL double-necked round-bottom flask, add compound 11 (15 g, 65 mmol), sodium iodide (19.5 g, 130 mmol) and acetonitrile (20 mL) in sequence under argon atmosphere, heat to 65 °C and stir for 12 h. After the reaction is completed, spin dry, recrystallize from ethyl acetate, collect the red viscous solid, and obtain compound 12. LRMS (ESI) m / z: [MH]- Calcd forC8H18INO3S 355.01, Found 355.20.

[0057] 3) Take a 100 mL double-necked round-bottom flask, under an argon atmosphere, add acetic acid (30 mL), sodium acetate 3.2 g, phenylhydrazine-4-sulfonic acid (3.76 g, 20 mmol) and methyl isopropyl ketone (3.2 mL, 30 mmol) in sequence, heat to 110 ° C and stir overnight. After the reaction is completed, no treatment is required, and the sample is directly loaded on the column chromatography, and the target product is eluted with a volume ratio of dichloromethane: methanol = 15:1. Collect the yellow liquid, spin dry and dry to obtain pink solid compound 13 (3.8 g, 80%). 1H NMR (400 MHz, DMSO-d6): δ 7.63 (s, 1H), 7.55 (d, J = 7.9 Hz, 1H), 7.34 (d, J = 7.9 Hz, 1H), 2.21(s, 3H), 1.25 (s, 6H). LRMS (ESI) m / z: [M+H]+ Calcd for C11H14NO3S 240.07;Found 240.19.. 4) Take a 100mL round-bottom flask, add compound 12 (2 g, 6 mmol), compound 13 (2g, 8 mmol), sodium iodide (900 mg, 6 mmol) and o-dichlorobenzene (50 mmol), heat to 110°C in an oil bath, and react for 48 h. After the reaction, recrystallize with ethanol and collect the pink solid to obtain compound 14 (1.2 g, yield 46%). 1H NMR (500 MHz, Methanol-d4): δ 8.23–8.09 (m, 2H), 8.06 (s, 1H), 4.84–4.61 (m, 2H), 3.88–3.76 (m, 2H), 3.72–3.55 (m, 4H), 3.46–3.37 Found 445.22.

[0058] 5) Take a 10 mL round-bottom flask, add compound 14 (120 mg, 0.27 mmoL), compound 15 (18.6 mg, 0.11 mmoL), anhydrous ethanol (2 mL) and sodium acetate (54 mg, 0.65 mmoL) in sequence, heat to 80°C in an oil bath, and react for 12 h. After the reaction, HPLC separation and analysis were performed to obtain green compound 1, which is a zwitterionic cyanine dye (166 mg, 60%). LRMS (ESI) m / z: [MH]- Calcd for C46H68ClN4O12S43+ 1031.34 ; Found 1028.07.

[0059] Example 2 This embodiment provides a zwitterionic cyanine dye, and its synthesis route is as follows:

[0060] The specific steps include: Take a 10 mL round-bottom flask, add compound 1 (102.8 mg, 0.1 mmoL), compound 2 (CAS: 2393-17-1, 83 mg, 0.5 mmoL), water (2 mL) and sodium bicarbonate (25 mg, 0.3 mmoL) in sequence, heat to 80 ° C in an oil bath, and react for 6 h. After the reaction, high performance liquid phase separation and analysis were performed to obtain blue compound 3, which is a zwitterionic cyanine dye (53 mg, 50%). LRMS (ESI) m / z: [MH]- Calcd for C55H78N5O14S43+1160.44; Found 1156.07.

[0061] Example 3 This embodiment provides a zwitterionic cyanine dye, and the synthesis route is as follows:

[0062] The specific steps include: Take a 10 mL round-bottom flask, add compound 1 (102.8 mg, 0.1 mmoL), compound 6 (CAS: 791028-27-8, 83 mg, 0.5 mmoL), water (2 mL) and sodium bicarbonate (25 mg, 0.3 mmoL) in sequence, then introduce inert gas to replace the air, and heat to 80°C in an oil bath under an inert gas atmosphere for 6 h. After the reaction, high performance liquid phase separation and analysis were performed to obtain blue compound 4, which is a zwitterionic cyanine dye (53 mg, 50%). LRMS (ESI) m / z: [MH]- Calcd forC53H82N5O16S43+1172.46 ; Found 1168.03.

[0063] Performance Testing 1. Spectrum test The zwitterionic cyanine dyes of Examples 2 and 3 were prepared into PBS solutions of 0.05 mg / mL of a constant-brightness fluorescent probe. The absorption spectra of each sample at 400-900 nm were measured by an ultraviolet spectrometer, and the fluorescence spectra were measured by a fluorescence spectrometer. The excitation wavelengths used in Examples 2 and 3 were 700 nm and 600 nm, respectively. The results are as follows: Figure 1 and 2 shown. Figure 1 The ultraviolet absorption spectrum curve and the fluorescence emission spectrum curve of the zwitterionic cyanine dye of Example 2 are shown. Figure 2 The ultraviolet absorption spectrum curve and the fluorescence emission spectrum curve of the zwitterionic cyanine dye of Example 3 are shown in FIG. Figure 1It can be seen that the absorption wavelength of the ultraviolet absorption spectrum is between 650 and 750 nm, and the emission wavelength of the fluorescence emission spectrum is between 700 and 800 nm. Figure 2 It can be seen that the absorption wavelength of the ultraviolet absorption spectrum is between 550 and 650 nm, and the emission wavelength of the fluorescence emission spectrum is between 700 and 800 nm.

[0064] 2. Renal clearance efficiency test Take the zwitterionic cyanine dyes of Examples 2 and 3 and prepare a PBS solution of 2 mg / mL of the ever-bright fluorescent probe. Take 9 Balb / c mice and randomly divide them into three groups, A, B, and C, with 3 mice in each group. 150 μL of the probe solution was injected into the tail vein. After the injection, the mice were placed individually in a clean metabolic cage, and the urine was collected within 24 hours and the volume was recorded. The standard curve of probe concentration-ultraviolet absorption intensity was prepared by ultraviolet absorption spectroscopy, and the recovery rate of the probe in mouse urine was calculated. The results are shown in Figure 2. Figure 3 and 4 shown.

[0065] Figure 3 The figure is a statistical graph of the recovery rate of the zwitterionic cyanine dye in urine within 24 hours after injection of the zwitterionic cyanine dye in Example 2, wherein Figure 3 A in the figure is a curve showing the change of urine recovery rate over time. Figure 3 Figure B shows the results of three repeated experiments on urine recovery rate. Figure 4 The figure is a statistical graph of the recovery rate of the zwitterionic cyanine dye in urine within 24 hours after injection of the zwitterionic cyanine dye in Example 3, wherein Figure 4 A in the figure is a curve showing the change of urine recovery rate over time. Figure 4 B in the figure is the result of three repeated experiments on urine recovery rate. Figure 3 It can be seen from A that the zwitterionic cyanine dye of Example 2 was basically metabolized out of the body 5-6 hours after tail vein injection, and the urine recovery rate basically reached the highest value. Figure 4 A shows that the zwitterionic cyanine dye of Example 3 is basically metabolized out of the body about 5 hours after tail vein injection, and the urine recovery rate basically reaches the highest value, indicating that the zwitterionic cyanine dye of the present invention has a fast renal metabolism rate. Figure 3 B and Figure 4 B shows that the urine recovery rate of the probe is relatively high, about 97% on average, indicating that the zwitterionic cyanine dye provided by the present invention is mainly metabolized by the kidney.

[0066] 3. Probe tissue distribution study The zwitterionic cyanine dyes of Examples 2 and 3 were prepared into a PBS solution of 2 mg / mL of a constantly bright fluorescent probe. Fifteen healthy Balb / c mice were randomly divided into five groups (1h group, 2h group, 3h group, 24h group, and control group), and 150 μL of the probe solution was injected into the tail vein. The rats were killed at 1h, 2h, 3h, and 24h after injection, and the main organs (heart He, liver Li, spleen Sp, lung Lu, kidney Ki, bladder Bl, small intestine In, skin Sk, muscle Mu) were collected and fluorescent imaging was performed. The results are shown in Figure 2. Figures 5 to 8 shown.

[0067] Figure 5 and Figure 7 Organ imaging diagrams of the fluorescent probes prepared from zwitterionic cyanine dyes of Example 2 and Example 3 in normal mice; wherein He represents heart, Li represents liver, Sp represents spleen, Lu represents lung, Ki represents kidney, Bl represents bladder, Sk represents skin, Mu represents muscle, and In represents small intestine. Figure 6 and Figure 8 The organ imaging quantitative diagrams of the fluorescent probes prepared by zwitterionic cyanine dyes in Examples 2 and 3 in normal mice; wherein He represents the heart, Li represents the liver, Sp represents the spleen, Lu represents the lung, Ki represents the kidney, Bl represents the bladder, Sk represents the skin, Mu represents the muscle, and In represents the small intestine. Figures 5 to 8 It can be seen that the fluorescent probe prepared from the zwitterionic cyanine dye provided by the present invention is mainly metabolized by the urinary system, and is rapidly enriched in the kidney and / or bladder after the probe is injected, and no obvious fluorescent signal is observed in other organs, indicating that the nonspecific uptake of the probe by other tissues and organs is extremely low, and can be used for the detection of urinary system diseases.

[0068] 4. Ureteric imaging applications The zwitterionic cyanine dyes of Example 2 and Example 3 were prepared into a PBS solution of 2 mg / mL of a constantly bright fluorescent probe. The solution was injected into mice via the tail vein, and the laparotomy and in vivo imaging of the small animals were performed at 30 minutes. The results are as follows: Fig. 9 As shown. Among them, Fig. 9 A in the figure is the zwitterionic cyanine dye ureteral imaging result diagram of Example 2, Fig. 9 B in the figure is the zwitterionic cyanine dye ureteral imaging result of Example 3. Fig. 9 It can be seen that the kidneys, ureters and bladders in the urinary system are all fully displayed, indicating that the zwitterionic cyanine dye provided by the present invention has the ability to image not only the kidneys and bladder, but also the ureter, and can be applied to the diagnosis and treatment of ureter-related diseases.

[0069] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the embodiments here. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. A zwitterionic cyanine dye, characterized in that: It has a structure shown in any one of formula (I) to formula (IV): ， Wherein, Y is F, Cl, Br or I; X is NH, SH or O; n is an integer of 0 to 30; and n1 is an integer of 2 to 30.

2. The zwitterionic cyanine dye according to claim 1, characterized in that The n is an integer of 1 to 30.

3. The zwitterionic cyanine dye according to claim 1, characterized in that The zwitterionic cyanine dye has any of the following structures: 、 、 、 , or .

4. A method for preparing the zwitterionic cyanine dye according to any one of claims 1 to 3, characterized in that: The steps include: When the zwitterionic cyanine dye is of formula (I): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the zwitterionic cyanine dye; When the zwitterionic cyanine dye has the structure of formula (II): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the compound of formula (I); S2. The compound of formula (I) undergoes a condensation reaction with compound (2) to obtain the zwitterionic cyanine dye; When the zwitterionic cyanine dye is of formula (III): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the compound of formula (I); S3. The compound of formula (I) undergoes a condensation reaction with compound (5) to obtain the zwitterionic cyanine dye; When the zwitterionic cyanine dye is of formula (IV): S1. Compound (14) and compound (15) undergo condensation reaction to obtain the compound of formula (I); S4. The compound of formula (I) undergoes a condensation reaction with compound (6) under anaerobic conditions to obtain the zwitterionic cyanine dye; The specific compounds involved are as follows: 。 5. The preparation method according to claim 4, characterized in that: The compound (14) is obtained by a substitution reaction between the compound (12) and the compound (13); The specific compounds involved are as follows: 。 6. The preparation method according to claim 4, characterized in that: In step S1, the reaction temperature of the condensation reaction is 80-100° C., and the time is 12-24 h.

7. The preparation method according to claim 4, characterized in that: In steps S2-S4, the reaction temperature of the condensation reaction is 65-85°C and the time is 6-12 h.

8. Use of the zwitterionic cyanine dye according to any one of claims 1 to 3 or a pharmaceutically acceptable salt, solvate, enantiomer, diastereomer or tautomer thereof in the preparation of a fluorescent probe.

9. The use according to claim 8, characterized in that: The fluorescent probe is a fluorescent probe used for organ imaging, and the organ is a kidney, a ureter or a bladder.

10. A fluorescent probe, characterized in that Contains the zwitterionic cyanine dye according to any one of claims 1 to 3 and / or its pharmaceutically acceptable salt, solvate, enantiomer, diastereomer, tautomer.

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