A targeted near-infrared fluorescent compound and its preparation method and application

Abstract

The present invention provides a targeted near-infrared fluorescent compound and a preparation method and application thereof, and relates to the technical field of organic fluorescent molecules. The targeted near-infrared fluorescent compound has a structure as shown in the following formula I. The present invention uses an organic total synthesis method to modify the S0456 near-infrared small molecule on the CXCR4 inhibitor to obtain a targeted near-infrared fluorescent compound as shown in formula I. The targeted near-infrared fluorescent compound has a good active targeting effect when identifying CXCR4-overexpressing tumors such as triple-negative breast cancer and head and neck squamous cell carcinoma; and the targeted near-infrared fluorescent compound retains the water solubility of the dye and the specificity for tumor cells, and has the advantages of good water solubility and high fluorescence quantum yield. #imgabs0#

Description

Technical Field

[0001] The invention relates to the technical field of near-infrared fluorescent molecules, and in particular to a targeted near-infrared fluorescent compound and a preparation method and application thereof. Background Art

[0002] In recent years, near-infrared imaging technology has provided an effective tool for precision tumor surgery. Currently, the only near-infrared contrast agent approved for clinical use in China is indocyanine green (ICG), which provided the ability to visualize tumors for clinical surgery in its early applications. However, as research deepened, it was found that the tumor imaging effect of ICG was greatly affected by individual differences among patients and surgical time, making it difficult to formulate standardized surgical plans.

[0003] In addition, ICG has the following problems: First, it has a low fluorescence quantum yield of <1% in aqueous solution, which greatly reduces the detection limit of the equipment. According to the Chinese expert consensus on the application of indocyanine green near-infrared imaging in laparoscopic radical gastrectomy (2019 edition), the maximum injection dose of ICG in humans should be less than 2 mg / kg. At this dose, the literature reports that the concentration of ICG in human tumors is 10~1000 nM (reference: Wang Z, Ni K, Zhang X, Ai S, Guan W, Cai H, Wang Y, Lu Q, Lane LA (2018) Method for real-time tissue quantification of indocyanine green revealing optimal conditions for near infrared fluorescence guided surgery. Anal Chem 90:7922–7929.), which places extremely high demands on the detection equipment. Second, in a short period of time, the metabolic rate of ICG in tumors and normal tissues is not much different. Only after more than 12 hours can sufficient fluorescence contrast (tumor: normal tissue) be generated, which increases the burden on hospitals and patients. Third, ICG is prone to photobleaching after aggregation, which greatly reduces its stability during imaging. Fourth, ICG is enriched in tumors through the EPR effect, lacks active targeting of tumor cells, and greatly reduces the accuracy of fluorescent tracer imaging. These factors greatly limit the clinical application of ICG. Therefore, it is necessary to further search for tumor-specific near-infrared fluorescent tracers suitable for clinical surgical imaging.

[0004] In cancer, the CXCL12 / CXCR4 biological axis can stimulate the proliferation and metastasis of tumor cells, regulate the inflammatory state of tumors, and activate the immune response within tumors through autocrine and paracrine factors. CXCR4 is expressed at high levels in many solid tumors and hematological tumors and is an effective target for tumor diagnosis and treatment. Small molecule compounds that act as CXCL12 / CXCR4 antagonists have great development and commercialization potential. So far, dozens of small molecule compounds have been studied and entered clinical trials.

[0005] In view of this, the present invention is proposed. Summary of the invention

[0006] The purpose of the present invention is to provide a targeted near-infrared fluorescent compound and its preparation method and application. The targeted near-infrared fluorescent compound of the present invention is a novel NY-CXCR4 near-infrared fluorescent molecule targeting chemokine receptor type 4 receptor obtained by connecting the near-infrared fluorescent molecule S0456 to the CXCR4 inhibitor, which can specifically target triple-negative breast cancer and head and neck squamous cell carcinoma, and has the advantages of good water solubility and high fluorescence quantum yield.

[0007] In order to achieve the above-mentioned purpose of the present invention, the following technical solutions are particularly adopted:

[0008] In a first aspect, the present invention provides a targeted near-infrared fluorescent compound, wherein the targeted near-infrared fluorescent compound has a structure as shown in the following formula I:

[0009]

[0010] Formula 1.

[0011] In a second aspect, the present invention provides a method for preparing the targeted near-infrared fluorescent compound as described in the first aspect, the preparation method comprising the following steps:

[0012] (1) Compound S0456 reacts with p-hydroxyphenylpropionate to obtain compound A;

[0013] The structural formula of the compound A is shown below:

[0014] ;

[0015] (2) Compound A reacts with dipyrrolidino (N-succinimidyloxy) carbonium hexafluorophosphate to obtain compound B;

[0016] The structural formula of the compound B is shown below:

[0017] ;

[0018] (3) Compound B reacts with a CXCR4 inhibitor to obtain a targeted near-infrared fluorescent compound as shown in Formula I;

[0019] The structure of the compound CXCR4 inhibitor is: .

[0020] Furthermore, in step (1), the molar ratio of the compound S0456 to p-hydroxyphenylpropionate is 1:(2-4).

[0021] Furthermore, in step (1), the p-hydroxyphenylpropionate is selected from sodium p-hydroxyphenylpropionate.

[0022] Furthermore, in step (1), the reaction is carried out in a solvent, and the solvent is selected from dimethyl sulfoxide.

[0023] Furthermore, in step (1), the reaction is carried out under a protective gas atmosphere, and the protective gas is selected from nitrogen.

[0024] Furthermore, in step (1), the reaction temperature is 60-70° C., and the reaction time is 5-7 h.

[0025] Furthermore, in step (1), after the reaction is completed, a post-treatment step is also included:

[0026] The reaction solution obtained after the reaction in step (1) is added dropwise to a mixture of ethyl acetate and ethanol, and the mixture is filtered and dried to obtain the compound A.

[0027] Furthermore, in the mixed solution, the volume ratio of ethyl acetate to ethanol is 1:((0.5~2).

[0028] Furthermore, in step (2), the compound A and the dipyrrolidino group ( N -succinimidyloxy)carbonium hexafluorophosphate has a molar ratio of 1:(1.2~2).

[0029] Furthermore, in step (2), the reaction is carried out in the presence of a base selected from N,N -Diisopropylethylamine.

[0030] Furthermore, in step (2), the molar ratio of compound A to base is 1:(0.5~5).

[0031] Furthermore, in step (2), the reaction is carried out in a solvent, and the solvent is selected from dimethyl sulfoxide.

[0032] Furthermore, in step (2), the reaction temperature is 30-40° C., and the reaction time is 1-3 h.

[0033] Furthermore, in step (2), after the reaction is completed, a post-treatment step is also included:

[0034] The reaction solution obtained after the reaction in step (2) is added dropwise to an acetone solution, and the mixture is filtered and dried to obtain the compound B.

[0035] Furthermore, in step (3), the molar ratio of compound B to the CXCR4 inhibitor is 1:(0.5-2).

[0036] Furthermore, in step (3), the reaction is carried out in the presence of a base, and the base is selected from N,N -Diisopropylethylamine.

[0037] Furthermore, in step (3), the molar ratio of compound B to base is 1:(0.5~5).

[0038] Furthermore, in step (3), the reaction is carried out in a solvent, and the solvent is selected from dimethyl sulfoxide.

[0039] Furthermore, in step (3), the reaction temperature is 30-40° C., and the reaction time is 18-30 h.

[0040] Furthermore, in step (3), after the reaction is completed, a post-treatment step is also included:

[0041] The reaction solution obtained after the reaction in step (3) is added dropwise to an acetone solution, and the mixture is filtered and dried to obtain a targeted near-infrared fluorescent compound represented by formula I.

[0042] In a third aspect, the present invention provides a use of the targeted near-infrared fluorescent compound or its racemate, stereoisomer, or pharmaceutically acceptable salt as described in the first aspect in the preparation of a tumor imaging agent.

[0043] Furthermore, the tumor includes a triple-negative breast cancer tumor and / or a head and neck squamous cell carcinoma tumor.

[0044] In a fourth aspect, the present invention provides a use of the targeted near-infrared fluorescent compound or its racemate, stereoisomer, or pharmaceutically acceptable salt as described in the first aspect in the preparation of a reagent for identifying CXCR4-overexpressing tumors.

[0045] Compared with the prior art, the present invention has the following beneficial effects:

[0046] (1) The present invention uses an organic total synthesis method to synthesize S0456 modified with a CXCR4 inhibitor to obtain the targeted near-infrared fluorescent compound NY-CXCR4. The probe can actively target chemokine receptor type 4 (CXCR4), can be rapidly cleared in normal tissues, and can remain in the tumor site for a long time. It has a good active targeting effect in identifying triple-negative breast cancer and head and neck squamous cell carcinoma.

[0047] (2) The targeted near-infrared fluorescent compound NY-CXCR4 of the present invention has the advantages of water solubility of the dye and high fluorescence quantum yield, etc. It has great development potential in the fields of near-infrared fluorescence navigation tumor surgery and medical cell labeling. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] In order to more clearly illustrate the specific implementation methods of the present invention or the technical solutions in the prior art, the drawings required for use in the specific implementation methods or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some implementation methods of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0049] Figure 1 This is a high performance liquid chromatography analysis diagram of NY-CXCR4 fluorescent molecules.

[0050] Figure 2 This is the NY-CXCR4 fluorescence molecular mass spectrum.

[0051] Figure 3 This is a diagram of the fluorescence performance of the NY-CXCR4 fluorescent molecule.

[0052] Figure 4 It is the specific targeting effect of NY-CXCR4 on triple-negative breast cancer.

[0053] Figure 5 It is the specific targeting effect of NY-CXCR4 on head and neck squamous cell carcinoma. DETAILED DESCRIPTION

[0054] Unless otherwise defined herein, scientific and technical terms used in conjunction with the present invention shall have the meanings commonly understood by those of ordinary skill in the art. The meaning and scope of the terms should be clear, however, in the case of any potential ambiguity, the definitions provided herein take precedence over any dictionary or external definitions. In this application, unless otherwise stated, the use of "or" means "and / or". In addition, the use of the term "including" and other forms is non-limiting.

[0055] It should be noted that specific details are described in the following description to facilitate a full understanding of the present invention. However, the present invention can be implemented in a variety of other ways different from those described herein, and those skilled in the art can make similar generalizations without violating the connotation of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0056] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0057] In a first aspect, the present invention provides a targeted near-infrared fluorescent compound, wherein the targeted near-infrared fluorescent compound has a structure as shown in the following formula I:

[0058]

[0059] Formula 1.

[0060] In the present invention, the S0456 near-infrared small molecule is modified on the CXCR4 inhibitor to obtain the targeted near-infrared fluorescent compound shown in the above formula I, hereinafter referred to as NY-CXCR4. Among them, the CXCR4 inhibitor of the present invention selects the structural analogue of plerixafor As a targeting group connected with a fluorescent group, the obtained NY-CXCR4 can be quickly cleared in normal tissues, but can be retained for a long time in the tumor site. The connection with the fluorescent group further expands the application value of the drug, and is used for surgical navigation and tumor removal. The results show that the NY-CXCR4 obtained by the present invention has a good active targeting effect in identifying triple-negative breast cancer and head and neck squamous cell carcinoma; and NY-CXCR4 also has the advantages of water solubility of the dye and high fluorescence quantum yield.

[0061] Among them, Plerixafor is a small non-peptide molecule that specifically antagonizes the CXCR4 receptor. The primary structure of Plerixafor contains two cyclopolysaccharide rings connected by a phenyl linker. Under physiological conditions, the protonated nitrogen atom on the ring interacts with the carboxylic acid group on CXCR4. This can inhibit the binding of CXCL12 to CXCR4, block downstream signal transduction and regulate various physiological activities. In the process of developing tumor-specific fluorescent probes, anti-tumor drugs with tumor-specific killing effects have certain research value and can be used for the development of specific targeted fluorescent probes.

[0062] In a second aspect, the present invention provides a method for preparing the targeted near-infrared fluorescent compound according to the first aspect, the preparation method comprising the following steps:

[0063] (1) Compound S0456 reacts with p-hydroxyphenylpropionate to obtain compound A;

[0064] The structural formula of the compound A is shown below:

[0065] ;

[0066] (2) Compound A reacts with dipyrrolidino (N-succinimidyloxy) carbonium hexafluorophosphate to obtain compound B;

[0067] The structural formula of the compound A is shown below:

[0068] ;

[0069] (3) Compound B reacts with a CXCR4 inhibitor to obtain a targeted near-infrared fluorescent compound as shown in Formula I;

[0070] The structure of the compound CXCR4 inhibitor is: .

[0071] More specifically, the preparation method of the targeted near-infrared fluorescent compound is as follows:

[0072] (1) Compound S0456 reacts with p-hydroxyphenylpropionate to obtain compound A; wherein the reaction formula is as follows:

[0073]

[0074] (2) Compound A and dipyrrolidine ( N -succinimidyloxy)carbonium hexafluorophosphate to react to obtain compound B; wherein the reaction formula is as follows:

[0075]

[0076] (3) Compound B reacts with a CXCR4 inhibitor to obtain a targeted near-infrared fluorescent compound as shown in Formula I; wherein the reaction formula is as follows:

[0077] .

[0078] In the present invention, a CXCR4 inhibitor ( ) modified S0456 near-infrared small molecule formula I shown as a targeted near-infrared fluorescent compound, namely NY-CXCR4, which has a good active targeting effect in identifying CXCR4-overexpressing tumors such as triple-negative breast cancer and head and neck squamous cell carcinoma. The targeted near-infrared fluorescent compound shown in formula I retains the water solubility of the dye and its specificity for tumor cells. As a tracer, it has the advantages of good water solubility and high fluorescence quantum yield, and has great development potential in the fields of fluorescence navigation tumor surgery and medical cell labeling.

[0079] In an optional embodiment, in step (1), the molar ratio of the compound S0456 to p-hydroxyphenylpropionate is 1:(2-4), for example, it can be 1:2, 1:2.2, 1:2.4, 1:2.6, 1:2.8, 1:3, 1:3.2, 1:3.4, 1:3.6, 1:3.8, 1:4, etc.

[0080] In an optional embodiment, in step (1), the salt of p-hydroxyphenylpropionate is selected from sodium p-hydroxyphenylpropionate.

[0081] In an optional embodiment, in step (1), the reaction is carried out in a solvent.

[0082] In an optional embodiment, in step (1), the solvent is selected from dimethyl sulfoxide.

[0083] In an optional embodiment, in step (1), the reaction is carried out under a protective gas atmosphere.

[0084] In an optional embodiment, in step (1), the protective gas is selected from nitrogen;

[0085] In an optional embodiment, in step (1), the reaction temperature is 60-70°C, for example, it can be 60°C, 61°C, 62°C, 63°C, 64°C, 65°C, 66°C, 67°C, 68°C, 69°C, 70°C, etc.

[0086] In an optional embodiment, in step (1), the reaction time is 5 to 7 h, for example, 5 h, 5.2 h, 5.4 h, 5.6 h, 5.8 h, 6 h, 6.2 h, 6.4 h, 6.6 h, 6.8 h, 7 h, etc.

[0087] In an optional embodiment, in step (1), after the reaction is completed, a post-treatment step is further included:

[0088] The reaction solution obtained after the reaction in step (1) is added dropwise to a mixture of ethyl acetate and ethanol, and the mixture is filtered and dried to obtain the compound A.

[0089] In an optional embodiment, in the mixed solution, the volume ratio of ethyl acetate to ethanol is 1:(0.5~2), for example, it can be 1:0.5, 1:0.6, 1:0.7, 1:0.8, 1:0.85, 1:0.9, 1:0.95, 1:1, 1:1.05, 1:1.1, 1:1.15, 1:1.2, 1:1.4, 1:1.6, 1:1.8, 1:2, etc.

[0090] In an optional embodiment, in step (2), the molar ratio of compound A to dipyrrolidino (N-succinimidyloxy) carbonium hexafluorophosphate is 1:(1.2~2), for example, it can be 1:1, 1:1.2, 1:1.25, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1:1.9, 1:2, etc.

[0091] In an optional embodiment, in step (2), the reaction is carried out in the presence of a base selected from N,N -Diisopropylethylamine.

[0092] In an optional embodiment, in step (2), the molar ratio of compound A to base is 1:(0.5~5), for example, it can be 1:0.5, 1:0.6, 1:0.7, 1:0.8, 1:0.9, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1:1.9, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, 1:5, etc.

[0093] In an optional embodiment, in step (2), the reaction is carried out in a solvent.

[0094] In an optional embodiment, in step (2), the solvent is selected from dimethyl sulfoxide.

[0095] In an optional embodiment, in step (2), the reaction temperature is 30-40°C, for example, 30°C, 31°C, 32°C, 33°C, 34°C, 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, etc.

[0096] In an optional embodiment, in step (2), the reaction time is 1 to 3 h, for example, 1 h, 1.2 h, 1.4 h, 1.6 h, 1.8 h, 2 h, 2.2 h, 2.4 h, 2.6 h, 2.8 h, 3 h, etc.

[0097] In an optional embodiment, in step (2), after the reaction is completed, a post-treatment step is also included:

[0098] The reaction solution obtained after the reaction in step (2) is added dropwise to an acetone solution, and the mixture is filtered and dried to obtain the compound B.

[0099] In an optional embodiment, in step (3), the molar ratio of compound B to the CXCR4 inhibitor is 1:(0.5-2), for example, it can be 1:0.5, 1:0.6, 1:0.7, 1:0.8, 1:0.85, 1:0.9, 1:0.95, 1:1, 1:1.05, 1:1.1, 1:1.15, 1:1.2, 1:1.4, 1:1.6, 1:1.8, 1:2, etc.

[0100] In an optional embodiment, in step (3), the reaction is carried out in the presence of a base.

[0101] In an optional embodiment, in step (3), the base is selected from N,N -Diisopropylethylamine.

[0102] In an optional embodiment, in step (3), the molar ratio of compound B to base is 1:(0.5~2), for example, it can be 1:0.5, 1:0.6, 1:0.7, 1:0.8, 1:0.9, 1:1, 1:1.2, 1:1.4, 1:1.6, 1:1.8, 1:2, 1:2.2, 1:2.4, 1:2.6, 1:2.8, 1:3, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, 1:5, etc.

[0103] In an optional embodiment, in step (3), the reaction is carried out in a solvent, and the solvent is selected from dimethyl sulfoxide.

[0104] In an optional embodiment, in step (3), the reaction temperature is 30-40°C, for example, 30°C, 31°C, 32°C, 33°C, 34°C, 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, etc.

[0105] In an optional embodiment, in step (3), the reaction time is 18 to 30 h, for example, 18 h, 19 h, 20 h, 21 h, 22 h, 23 h, 24 h, 25 h, 26 h, 27 h, 28 h, 29 h, 30 h, etc.

[0106] In an optional embodiment, in step (3), after the reaction is completed, a post-treatment step is also included:

[0107] The reaction solution obtained after the reaction in step (3) is added dropwise to an acetone solution, and the mixture is filtered and dried to obtain a targeted near-infrared fluorescent compound represented by formula I.

[0108] In a third aspect, the present invention provides a use of the targeted near-infrared fluorescent compound or its racemate, stereoisomer, or pharmaceutically acceptable salt as described in the first aspect in the preparation of a tumor imaging agent.

[0109] In an alternative embodiment, the tumor comprises a triple-negative breast cancer tumor and / or a head and neck squamous cell carcinoma tumor.

[0110] In a fourth aspect, the present invention provides a use of the targeted near-infrared fluorescent compound or its racemate, stereoisomer, or pharmaceutically acceptable salt in the preparation of a reagent for identifying CXCR4-overexpressing tumors.

[0111] In an optional embodiment, the application is specifically: using the targeted near-infrared fluorescent compound or its racemate, stereoisomer, pharmaceutically acceptable salt as described in the first aspect to establish a tumor imaging model, and using intraoperative fluorescence navigation equipment to diagnose the tumor effect.

[0112] In an optional embodiment, the present invention provides a triple-negative breast cancer tumor development model and a method for establishing the same, which specifically comprises the following steps: inoculating a CXCR4-high-expressing cell line of human triple-negative breast cancer on the back of a mouse, and injecting the targeted near-infrared fluorescent compound (NY-CXCR4) of the present invention via tail vein injection.

[0113] More specifically, the present invention provides a triple-negative breast cancer tumor development model and an establishment method thereof, which specifically comprises the following steps: using a CXCR4 high-expressing cell line, human triple-negative breast cancer MDA-MB-231, to be inoculated on the back of Balb / c mice, and injecting the targeted near-infrared fluorescent compound (NY-CXCR4) of the present invention through tail vein injection.

[0114] It should be noted that after the above-mentioned triple-negative breast cancer tumor imaging model is established, the intraoperative fluorescent navigation equipment is used to diagnose the effect of small molecules on triple-negative breast cancer tumors; the triple-negative breast cancer can be removed during surgery, and pathological sections can be used to determine whether NY-CXCR4 can specifically target triple-negative breast cancer tumors.

[0115] In an optional embodiment, the present invention provides a head and neck squamous cell carcinoma development model and a method for establishing the same, which specifically comprises the following steps: inoculating a CXCR4 high-expressing cell line human head and neck squamous cell carcinoma into the back of a mouse, and injecting the targeted near-infrared fluorescent compound (NY-CXCR4) of the present invention via tail vein injection.

[0116] More specifically, the present invention provides a head and neck squamous cell carcinoma imaging model and an establishment method thereof, which specifically comprises the following steps: inoculating the CXCR4 high-expressing cell line human head and neck squamous cell carcinoma FaDu on the back of Balb / c mice, and injecting the targeted near-infrared fluorescent compound (NY-CXCR4) of the present invention via tail vein injection.

[0117] It should be noted that after the above-mentioned head and neck squamous cell carcinoma imaging model is established, intraoperative fluorescence navigation equipment can be used to diagnose the small molecule effect of head and neck squamous cell carcinoma tumors. The head and neck squamous cell carcinoma is removed during surgery, and pathological sections are used to determine whether NY-CXCR4 can specifically target head and neck squamous cell carcinoma tumors.

[0118] As mentioned in the present invention, the term "pharmaceutically acceptable salt" means that the salt is not only physiologically acceptable to the subject, but also refers to a synthetic substance with pharmaceutical use value, such as a salt formed as an intermediate when performing chiral separation. Although the salt of this intermediate cannot be directly administered to the subject, the salt can play a role in obtaining the final product of the present invention.

[0119] As mentioned in the present invention, the pharmaceutically acceptable salt of the compound of formula I is a salt formed with a base or an alkali metal. The acid that forms a pharmaceutically acceptable salt with the compound of formula I includes an inorganic acid and an organic acid. More specifically, the alkali metal that forms a pharmaceutically acceptable salt with the compound of formula I includes but is not limited to lithium, sodium, potassium, magnesium, calcium, aluminum, zinc, etc.; the base that forms a pharmaceutically acceptable salt with the compound of formula I includes but is not limited to choline, diethanolamine, morpholine, etc.

[0120] The present invention is further described below by way of examples. Unless otherwise specified, the materials in the examples are prepared according to existing methods or directly purchased from the market.

[0121] Example 1

[0122] This embodiment provides a targeted near-infrared fluorescent compound NY-CXCR4, wherein the targeted near-infrared fluorescent compound NY-CXCR4 has a structure as shown in the following formula I:

[0123]

[0124] Formula 1.

[0125] The synthesis route of the targeted near-infrared fluorescent compound NY-CXCR4 is as follows:

[0126]

[0127] The preparation method of the targeted near-infrared fluorescent compound NY-CXCR4 specifically comprises the following steps:

[0128] (1) Dissolve S0456 (888 mg, 1 mmol) in 10 ml DMSO and stir until completely dissolved. Add sodium p-hydroxyphenylpropionate (424 mg, 2 mmol) into the flask. React at 65 °C for 6 h under nitrogen protection. Add the resulting solution dropwise into a mixed solution of 200 mL ethyl acetate and ethanol (V 乙酸乙酯 :V 乙醇 =1:1), a dark green precipitate was produced, which was filtered and dried in a vacuum to obtain 1.05 g of solid (i.e., compound A), with a yield of 95%.

[0129] (2) Compound A (1 g, 0.98 mmol) was placed in a round-bottom flask and dissolved in 20 mL of DMSO. Dipyrrolidino (N-succinimidyloxy) carbonium hexafluorophosphate (483 mg, 1.176 mmol) and DIEA (151 mg, 1.176 mmol) were added in sequence. The mixture was stirred at 37 °C for 2 h and precipitated in acetone solution. The precipitate was filtered and dried under vacuum to obtain 1.499 g (i.e., compound B).

[0130] (3) Compound B (1.15 g, 1 mmol) was placed in a round-bottom flask, dissolved in 20 mL DMSO, and then added (0.431 g, 1 mmol), DIEA (258 mg, 2 mmol), stirred at 37 ° C for 24 h, precipitated in acetone solution, filtered and dried in vacuum to obtain 1.6 g. The product was purified by preparative liquid chromatography to obtain the targeted near-infrared fluorescent compound NY-CXCR4 shown in formula I.

[0131] The structure was characterized by HPLC and mass spectrometry, and the results of structure determination are as follows:

[0132] like Figure 1 As shown, the purity of the targeted near-infrared fluorescent compound NY-CXCR4 is 96.1%.

[0133] like Figure 2 As shown, LCMS (ESI): m / z: C 74 H 89 N7O 14 S4, [M-3H] found 465.15, [M-2H] found 698.65.

[0134] like Figure 3 As shown, the targeted near-infrared fluorescent compound NY-CXCR4 has a maximum excitation wavelength Ex max = 773 nm and a maximum fluorescence emission wavelength Em max = 795 nm, indicating that the molecule is a near-infrared fluorescent probe.

[0135] Application Example 1

[0136] This application example provides an application of a NY-CXCR4 near-infrared fluorescent molecule in the specific identification of triple-negative breast cancer.

[0137] The triple-negative breast cancer model was constructed using the MDA-MB-231 cell line: The CXCR4-high-expressing human triple-negative breast cancer MDA-MB-231 cell line was inoculated on the back of Balb / c mice, and 10 nmol of NY-CXCR4 molecules were injected into the tail vein of each mouse. The intraoperative fluorescence navigation device was used to diagnose the tumor effect of small molecules on triple-negative breast cancer. The triple-negative breast cancer was removed during surgery, and the pathological sections were used to determine whether NY-CXCR4 could specifically target triple-negative breast cancer tumors.

[0138] Test results such as Figure 4 As shown:

[0139] The left side shows the effect of using intraoperative fluorescence navigation equipment to diagnose triple-negative breast cancer tumors with small molecules. When 10 nmol NY-CXCR4 was injected 8 h, 12 h, and 24 h later, the back of the mouse showed high fluorescence signals only in the tumor area, while the surrounding normal tissues showed low fluorescence. This shows that NY-CXCR4 has good specificity in identifying triple-negative breast cancer with overexpression of CXCR4 receptors, and can achieve rapid clearance in normal tissues, while staying in the breast cancer tumor site for a long time, thereby obtaining a higher tumor-to-background signal ratio.

[0140] The right side shows the distribution of fluorescence signals of isolated organs at corresponding time points. After the injection of 10 nmol NY-CXCR4, it can be seen that the fluorescence intensity of tumor tissue is significantly higher than that of muscle tissue at each time point. This shows that NY-CXCR4 can accumulate in tumors and achieve specific intraoperative identification of breast cancer tumors.

[0141] Application Example 2

[0142] This application example provides an application of a NY-CXCR4 near-infrared fluorescent molecule in the specific identification of head and neck squamous cell carcinoma.

[0143] The CXCR4-high-expressing cell line human head and neck squamous cell carcinoma FaDu was inoculated on the back of Balb / c mice, and 10nmol of NY-CXCR4 molecules were injected into the tail vein of each mouse. The intraoperative fluorescence navigation device was used to diagnose the small molecule effect of head and neck squamous cell carcinoma tumors. The head and neck squamous cell carcinoma was removed during surgery, and the pathological sections were used to determine whether NY-CXCR4 could specifically target head and neck squamous cell carcinoma tumors.

[0144] Test results such as Figure 5 As shown:

[0145] The left side shows the effect of using intraoperative fluorescence navigation equipment to diagnose head and neck squamous cell carcinoma tumors with small molecules. When 10 nmol NY-CXCR4 was injected 8 h, 12 h, and 24 h later, the back of the mouse showed high fluorescence signals only in the tumor area, while the surrounding normal tissues showed low fluorescence. This shows that NY-CXCR4 has good specificity in identifying head and neck squamous cell carcinoma with overexpression of CXCR4 receptors, and can achieve rapid clearance in normal tissues and long-term retention in the head and neck squamous cell carcinoma tumor site, thereby achieving the effect of in vivo diagnosis.

[0146] The right side shows the distribution of fluorescence signals of ex vivo organs at the corresponding time. After the injection of 10 nmol NY-CXCR4, it can be seen that the fluorescence intensity of tumor tissue is significantly higher than that of muscle tissue at each time point. This shows that NY-CXCR4 can accumulate in tumors and achieve intraoperative specific identification of head and neck squamous cell carcinoma tumors.

[0147] In summary, the present invention aims to provide a near-infrared fluorescent probe and its preparation method and application, which can actively target chemokine receptor type 4 (CXCR4), can be quickly cleared in normal tissues, and stay in the tumor site for a long time, so as to achieve the function of in vivo diagnosis. The modified S0456 near-infrared small molecule, NY-CXCR4, has good active targeting effects in identifying triple-negative breast cancer and head and neck squamous cell carcinoma. This near-infrared fluorescent dye retains the water solubility of the dye and its specificity for tumor cells. The tracer has the advantages of good water solubility and high fluorescence quantum yield, and has great development potential in the fields of near-infrared fluorescence navigation tumor surgery and medical cell labeling.

[0148] The novel near-infrared fluorescent tracer targeting CXCR4 receptor of the present invention is used for the specific targeting of triple-negative breast cancer and head and neck squamous cell carcinoma. The S0456 is used as the parent, and S0456 and are connected together by p-hydroxyphenylpropionic acid to finally condense into NY-CXCR4 active targeting near-infrared fluorescent molecule. Animal experiments have confirmed that NY-CXCR4 can target tumors.

[0149] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A targeted near-infrared fluorescent compound, characterized in that: The targeted near-infrared fluorescent compound has a structure as shown in the following formula I: Formula 1.

2. A method for preparing a targeted near-infrared fluorescent compound according to claim 1, characterized in that: The preparation method comprises the following steps: (1) Compound S0456 reacts with p-hydroxyphenylpropionate to obtain compound A; wherein the reaction formula is as follows: ； (2) Compound A reacts with dipyrrolidino (N-succinimidyloxy) carbonium hexafluorophosphate to obtain compound B; The structural formula of the compound B is shown below: ； (3) Compound B reacts with a CXCR4 inhibitor to obtain a targeted near-infrared fluorescent compound as shown in Formula I; The structure of the compound CXCR4 inhibitor is: .

3. The method for preparing the targeted near-infrared fluorescent compound according to claim 2, characterized in that: In step (1), the molar ratio of the compound S0456 to p-hydroxyphenylpropionate is 1:(2-4); And / or, in step (1), the p-hydroxyphenylpropionate is selected from sodium p-hydroxyphenylpropionate; And / or, in step (1), the reaction is carried out in a solvent selected from dimethyl sulfoxide; And / or, in step (1), the reaction is carried out under a protective gas atmosphere, and the protective gas is selected from nitrogen; And / or, in step (1), the reaction temperature is 60-70° C., and the reaction time is 5-7 h.

4. The method for preparing the targeted near-infrared fluorescent compound according to claim 2 or 3, characterized in that: In step (1), after the reaction is completed, a post-treatment step is also included: The reaction solution obtained after the reaction in step (1) is added dropwise to a mixture of ethyl acetate and ethanol, and the mixture is filtered and dried to obtain the compound A; And / or, in the mixed solution, the volume ratio of ethyl acetate to ethanol is 1:(0.5~2).

5. The method for preparing the targeted near-infrared fluorescent compound according to claim 2, characterized in that: In step (2), the compound A and the dipyrrolidone ( N -succinimidyloxy)carbonium hexafluorophosphate has a molar ratio of 1:(1.2-2); And / or, in step (2), the reaction is carried out in the presence of a base selected from N,N -diisopropylethylamine; And / or, in step (2), the molar ratio of compound A to base is 1:(0.5-5); And / or, in step (2), the reaction is carried out in a solvent selected from dimethyl sulfoxide; And / or, in step (2), the reaction temperature is 30-40° C., and the reaction time is 1-3 h.

6. The method for preparing a targeted near-infrared fluorescent compound according to claim 2 or 5, characterized in that: In step (2), after the reaction is completed, a post-treatment step is also included: The reaction solution obtained after the reaction in step (2) is added dropwise to an acetone solution, and the mixture is filtered and dried to obtain the compound B.

7. The method for preparing the targeted near-infrared fluorescent compound according to claim 2, characterized in that: In step (3), the molar ratio of compound B to CXCR4 inhibitor is 1:(0.5-2); And / or, in step (3), the reaction is carried out in the presence of a base selected from N,N -diisopropylethylamine; And / or, in step (3), the molar ratio of compound B to base is 1:(0.5-5); And / or, in step (3), the reaction is carried out in a solvent selected from dimethyl sulfoxide; And / or, in step (3), the reaction temperature is 30-40° C., and the reaction time is 18-30 h.

8. The method for preparing a targeted near-infrared fluorescent compound according to claim 2 or 7, characterized in that: In step (3), after the reaction is completed, a post-treatment step is also included: The reaction solution obtained after the reaction in step (3) is added dropwise to an acetone solution, and the mixture is filtered and dried to obtain a targeted near-infrared fluorescent compound represented by formula I.

9. Use of the targeted near-infrared fluorescent compound or a pharmaceutically acceptable salt thereof as claimed in claim 1 in preparing a reagent for tumor imaging; in, The tumor is a triple-negative breast cancer tumor and / or a head and neck squamous cell carcinoma tumor.

10. Use of the targeted near-infrared fluorescent compound and pharmaceutically acceptable salt according to claim 1 in the preparation of a reagent for identifying CXCR4-overexpressing tumors.

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