Nuclide probe targeting COX-2 as well as preparation method and application of nuclide probe

Abstract

The invention relates to the technical field of nuclide probes, and discloses a COX-2 targeting nuclide probe as well as a preparation method and application thereof, and the COX-2 targeting nuclide probe comprises a radionuclide labeled compound as shown in a formula I; the targeting COX-2 nuclide probe provided by the invention can be used as a diagnosis and treatment reagent to be applied to tumors with high COX-2 protein expression in human or animal bodies, and is especially suitable for tumor imaging and radionuclide treatment; animal experiments show that the nuclide probe has a high tumor / muscle uptake ratio and shows good application potential. Formula I.

Description

Technical Field

[0001] This invention relates to the field of radionuclide probe technology, and more specifically, to a radionuclide probe targeting COX-2, its preparation method, and its application. Background Technology

[0002] COX-2 (cyclooxygenase-2) is a key enzyme mediating arachidonic acid metabolism. It mainly catalyzes the production of prostaglandins. In addition to regulating the body's inflammatory response, it can also enhance the proliferation of tumor cells, inhibit apoptosis, promote angiogenesis and invasion and metastasis. It is closely related to the occurrence, development, treatment response and prognosis of various solid tumors. At the same time, it is also an important potential target for tumor targeted therapy and molecular imaging. In triple-negative breast cancer (TNBC), approximately 45%-75% of patients exhibit COX-2 overexpression. COX-2 expression levels are significantly correlated with increased tumor histological grade and lymph node metastasis risk. Furthermore, it can enhance tumor cell resistance to anthracyclines and taxane chemotherapy drugs by activating the PI3K / Akt signaling pathway, making it an independent risk factor for poor prognosis in TNBC patients. In high-grade serous ovarian cancer (HGSOC), COX-2 overexpression rates reach 50%-70%, with a higher positivity rate in BRCA wild-type patients. In non-small cell lung cancer (NSCLC) patients, COX-2 expression is upregulated in approximately 35%-60% of cases, with adenocarcinoma subtypes showing a higher COX-2 positivity rate than squamous cell carcinoma subtypes. High expression can enhance tumor angiogenesis by promoting prostaglandin E2 production, and is also significantly associated with resistance to EGFR-TKIs (epidermal growth factor receptor tyrosine kinase inhibitors) and an increased risk of local recurrence after radiotherapy. In pancreatic cancer and glioblastoma (GBM), the overexpression rate of COX-2 in pancreatic cancer patients is 40%-65%, which is positively correlated with the depth of tumor invasion and the degree of nerve infiltration, and can reduce the response rate of patients to gemcitabine-based chemotherapy regimens. In GBM, the overexpression rate of COX-2 is as high as 65%-85%, which can promote tumor cells to cross the blood-brain barrier and is also associated with radiotherapy resistance. It has now become an attractive biomarker for targeted imaging (such as PET imaging) and personalized treatment of solid tumors.

[0003] Indomethacin is a classic nonsteroidal anti-inflammatory drug (NSAID), belonging to the non-selective cyclooxygenase (COX) inhibitors. Its main targets are COX-1 and COX-2, with significantly higher inhibitory activity against COX-2 than against COX-1. In the development of tumor-specific radionuclide probes, indomethacin, with its ability to specifically recognize COX-2-overexpressing tumors, holds significant research value. Based on its high affinity for COX-2, it can be used to develop specific targeting radionuclide probes, aiding in preoperative PET imaging of tumors and supporting precise tumor diagnosis.

[0004] In view of this, the present invention proposes a COX-2 targeting nuclide probe, its preparation method and application. Summary of the Invention

[0005] To address the problems in related technologies, this invention proposes a COX-2-targeting nuclide probe, its preparation method, and its application, thereby overcoming the aforementioned technical problems in existing related technologies.

[0006] The purpose of this invention is to provide a COX-2-targeting probe, its preparation method, and its application. By enhancing the target affinity of the chemical probe and utilizing the structure-selectivity relationship of indomethacin, the parent compound is optimized using a covalent method to obtain a new COX-2-targeting radioligand, which is then used for PET imaging. From its design, synthesis, and in vitro and in vivo evaluation, this radionuclide probe can specifically target COX-2-overexpressing tumors, enabling non-invasive tumor diagnosis and monitoring.

[0007] Therefore, the specific technical solution adopted by the present invention is as follows: According to a first aspect of the invention, a COX-2 targeting nuclide probe is provided, the COX-2 targeting nuclide probe comprising a radiolabeled compound of formula I:

[0008] Formula I; The Y group includes any one of group I and group II. Linkage sites of representative groups:

[0009] The radionuclides include 55 Co、 68 Ga、 64 Cu、 86 Y、 89 Zr、 90 Y、 111 In、 177 Lu、 225 Any one of Ac.

[0010] Furthermore, the radiolabeled compound of Formula I includes either NY-COX-2-NOTA or NY-COX-2-DOTA; The structural formula of NY-COX-2-NOTA is as follows:

[0011] The structural formula of NY-COX-2-DOTA is: .

[0012] According to a second aspect of the present invention, a method for preparing a COX-2-targeting nuclide probe is provided, comprising the following steps: S1. The resin type selected is 2-CL resin, and the synthesis method is FMOC solid-phase synthesis, starting from the C-terminus of the peptide. FMOC-LYS(DDE)-OH and the resin are condensed using condensation reagents HBTU / DIEA and DCM as solvents. The reaction formula is shown below: ; S2. FMOC is removed using a DMF solution containing 20% ​​piperidine for 20 minutes. After removal, the resin is washed with DMF four times, 1 minute each time. The resin peptide is then dehydrated and condensed using DIC+HOBT as a condensation reagent and DMF as a solvent. This step is repeated to sequentially condense FMOC-GLU-OTBU and the small molecule. The reaction formula is shown below: ; S3. DDE is removed by hydrazine hydrate, followed by solvent coupling of DOTA-NHS via the condensation reagent DIEA / DMF, as shown in the following reaction formula: ; S4. Using a lysis buffer with a ratio of 95% TFA, 1% H2O, 2% EDT, and 2% TIS, the lysis time was 2 hours. After lysis, the lysate was filtered into ice-cold diethyl ether and centrifuged to obtain the crude product. Then, HPLC separation was performed to obtain a purity of 95%, and the pure product was obtained by lyophilization. The reaction formula is shown below: ; S5. Combine sodium acetate solution, the compound solution shown in Formula I, and the radioactive nuclide [ 68 The GaCl3 solution was mixed and reacted to obtain the COX-2 targeting nuclide probe, wherein the reaction formula is shown below: .

[0013] Furthermore, in step S1, FMOC-LYS(DDE)-OH is formed by the condensation of DIEA and HBTU with the resin; In step S1, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours. In step S1, the condensation reaction is carried out in a solvent, and the solvent is dichloromethane; In step S1, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is O-benzotriazole-tetramethylurea hexafluorophosphate. In step S1, the molar ratio of FMOC-LYS(DDE)-OH to the condensing agent is 1:(1.2~2); In step S1, the condensation reaction is carried out in the presence of a base, wherein the base is N,N-diisopropylethylamine; In step S1, the molar ratio of FMOC-LYS(DDE)-OH to the base is 1:(0.5~5).

[0014] Furthermore, in step S2, FMOC-GLU-OTBU undergoes dehydration condensation with the resin peptide using DIC / HOBT / DMF as a solvent; In step S2, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours. In step S2, the condensation reaction is carried out in a solvent, and the solvent is N,N-dimethylformamide; In step S2, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is 1-hydroxybenzotriazole; In step S2, the molar ratio of FMOC-GLU-OTBU to the condensing agent is 1:(1.2~2); In step S2, the condensation reaction is carried out in the presence of a base, wherein the base is N,N'-diisopropylcarbodiimide; In step S2, the molar ratio of FMOC-GLU-OTBU to the base is 1:(0.5~5).

[0015] Furthermore, in step S3, the resin peptide is dehydrated and condensed with DOTA using DIC / HOBT / DMF as a solvent to obtain a resin peptide. The peptide and resin are separated using a DCM solution containing 20% ​​trifluoroethanol, rotary evaporated, and lyophilized to obtain a fully protected peptide. In step S3, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours. In step S3, the condensation reaction is carried out in a solvent, and the solvent is N,N-dimethylformamide; In step S3, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is 1-hydroxybenzotriazole; In step S3, the molar ratio of indomethacin to the condensing agent is 1:(1.2~2); In step S3, the condensation reaction is carried out in the presence of a base, wherein the base is N,N-diisopropylethylamine; In step S3, the molar ratio of indomethacin to the base is 1:(0.5~5); In step S3, the reagent used for separating the peptides and resin is a dichloromethane solution containing 20% ​​trifluoroethanol. In step S3, the temperature for deprotection treatment is 30~40℃, and the treatment time is 1~3 h.

[0016] Furthermore, in step S4, the peptide and DOTA(OtBu)3 are mixed in equimolar amounts, DMF is used as a solvent, DIEA / HBTU is used as a condensing agent to couple DOTA(OtBu)3, and then the mixture is lyophilized. In step S4, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours. In step S4, the condensation reaction is carried out in a solvent, and the solvent is N,N-dimethylformamide; In step S4, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is O-benzotriazole-tetramethylurea hexafluorophosphate. In step S4, the molar ratio of the polypeptide to the condensing agent is 1:(1.2~2); In step S4, the condensation reaction is carried out in the presence of a base, wherein the base is N,N-diisopropylethylamine; In step S4, the molar ratio of the polypeptide to the base is 1:(0.5~5); In step S4, the molar ratio of the polypeptide to DOTA(OtBu)3 is 1:(0.5~5).

[0017] Furthermore, in step S6, the radioactive labeling includes the following steps: The sodium acetate solution, the compound solution shown in Formula I, and the radionuclide solution are mixed and reacted to obtain the COX-2 targeting nuclide probe. The volume ratio of sodium acetate solution to radioactive nuclide solution is 1:1; The sodium acetate solution has a concentration of 1-3 M and a pH of 4; The concentration of the compound solution shown in Formula I is 5 × 10⁻⁶. -6 M; The radioactivity of the radionuclide solution is 1~10 mCi; The radioactive labeling temperature is 85~95℃, and the radioactive labeling time is 5~20 min.

[0018] According to a third aspect of the invention, there is provided the use of a COX-2 targeting nuclide probe or its racemic, stereoisomer, or pharmaceutically acceptable salt in the preparation of a reagent for tumor imaging; The tumors include any one of triple-negative breast cancer, ovarian cancer, non-small cell lung cancer, pancreatic cancer, and glioblastoma.

[0019] According to a fourth aspect of the invention, there is provided the use of a COX-2 targeting nuclide probe or its racemic, stereoisomer, or pharmaceutically acceptable salt in the preparation of a reagent for identifying COX-2 overexpressing tumors.

[0020] The beneficial effects of this invention are as follows: 1) This invention utilizes an organic total synthesis method to synthesize a series of radionuclide probes. By linking indomethacin with a chelating agent ligand of a specific structure using glutamic acid and lysine, COX-2 targeting probes are generated. These probes can be rapidly cleared in normal tissues, thereby quickly achieving the tumor background signal ratio that meets clinical needs.

[0021] 2) The series of COX-2-targeting radionuclide probes synthesized in this invention utilize polyglutamic acid linked to indomethacin and DOTA. This design further enhances the tumor uptake of the radionuclide probes, mainly due to the close relationship between the characteristics of the tumor microenvironment and the molecular properties of polyglutamic acid. The hydrophilicity and negative charge of polyglutamic acid can enhance the interaction between the molecule and tumor cell surface receptors, thereby increasing the uptake rate. Attached Figure Description

[0022] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0023] Figure 1 The mass spectrum of NY-COX-2-DOTA provided in Example 1; Figure 2 Provided for Example 1 68 High performance liquid chromatography of Ga-NY-COX-2-DOTA; Figure 3 The COX-2 targeting probe provided in Example 1 68 A diagram illustrating the specific targeting effect of Ga-NY-COX-2-DOTA on triple-negative breast cancer. Figure 4 The αvβ3-targeting nuclide probe provided in Example 1 68 Organ distribution histogram of Ga-NY-COX-2-DOTA over 2 hours. Detailed Implementation

[0024] To further illustrate the various embodiments, the present invention provides accompanying drawings, which are part of the disclosure of the present invention. These drawings are mainly used to illustrate the embodiments and can be used in conjunction with the relevant descriptions in the specification to explain the operating principles of the embodiments. With reference to these drawings, those skilled in the art should be able to understand other possible implementation methods and the advantages of the present invention. The components in the drawings are not drawn to scale, and similar component symbols are generally used to represent similar components.

[0025] According to a first aspect of the invention, a COX-2 targeting probe is provided, the COX-2 targeting probe comprising a radiolabeled compound of formula I:

[0026] Formula I; The Y group is selected from any one of the following groups. Linkage sites of representative groups: ; In this invention, a novel radionuclide probe with COX-2 targeting capability was designed and synthesized. Using indomethacin as a recognition group, glutamate and lysine were linked to a chelating agent Y (NOTA, DOTA) to construct a novel radionuclide probe with COX-2 targeting capability, which can specifically target and identify COX-2 overexpressing tumors. The radionuclide probe obtained in this invention can specifically recognize COX-2 overexpressing tumors and can be rapidly cleared in normal tissues, thereby quickly achieving a tumor background signal ratio (TBR>1.5) that meets clinical requirements for preoperative tumor PET imaging.

[0027] Indomethacin is a classic nonsteroidal anti-inflammatory drug (NSAID), belonging to the non-selective cyclooxygenase (COX) inhibitors. Its main targets are COX-1 and COX-2, with significantly higher inhibitory activity against COX-2 than against COX-1. In the development of tumor-specific radionuclide probes, indomethacin, with its specificity against COX-2-overexpressing tumors, holds significant research value. Based on its high affinity for COX-2, it can be used to develop specific targeted radionuclide probes, aiding in preoperative PET imaging of tumors and supporting precise tumor diagnosis.

[0028] As an optional implementation, the COX-2 targeting nuclide probe comprises a radiolabeled compound of Formula I, including any one of the following compounds:

[0029] NY-COX-2-NOTA

[0030] NY-COX-2-DOTA; As an optional implementation, the radionuclide includes 55 Co、 68 Ga、 64 Cu、 86 Y、 89 Zr、 90 Y、 111 In、 177 Lu、225 Any one of Ac, preferably 68 Ga.

[0031] According to a second aspect of the present invention, a method for preparing a COX-2-targeting nuclide probe is provided, comprising the following steps: S1. The resin type selected is 2-CL resin, and the synthesis method is FMOC solid-phase synthesis, starting from the C-terminus of the peptide. FMOC-LYS(DDE)-OH and the resin are condensed using condensation reagents HBTU / DIEA and DCM as solvents. The reaction formula is shown below: ; S2. FMOC is removed using a DMF solution containing 20% ​​piperidine for 20 minutes. After removal, the resin is washed with DMF four times, 1 minute each time. The resin peptide is then dehydrated and condensed using DIC+HOBT as a condensation reagent and DMF as a solvent. This step is repeated to sequentially condense FMOC-GLU-OTBU and the small molecule. The reaction formula is shown below: ; S3. DDE is removed by hydrazine hydrate, followed by solvent coupling of DOTA-NHS via the condensation reagent DIEA / DMF, as shown in the following reaction formula: ; S4. Using a lysis buffer with a ratio of 95% TFA, 1% H2O, 2% EDT, and 2% TIS, the lysis time was 2 hours. After lysis, the lysate was filtered into ice-cold diethyl ether and centrifuged to obtain the crude product. Then, HPLC separation was performed to obtain a purity of 95%, and the pure product was obtained by lyophilization. The reaction formula is shown below: ; S5. Combine sodium acetate solution, the compound solution shown in Formula I, and the radioactive nuclide [ 68 The GaCl3 solution was mixed and reacted to obtain the COX-2 targeting nuclide probe, wherein the reaction formula is shown below: .

[0032] As an optional implementation, in step S1, the molar ratio of FMOC-LYS(DDE)-OH to HBTU is 1:(1.2~2), for example, it can be 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.

[0033] As an optional implementation, in step S1, the temperature of the condensation reaction is 30~40℃, for example, it can be 30℃, 31℃, 32℃, 33℃, 34℃, 35℃, 36℃, 37℃, 38℃, 39℃, 40℃, etc., and the time of the condensation reaction is 2~4 h, for example, it can be 2 h, 3 h, 4 h, etc.

[0034] As an optional implementation, in step S1, the condensation reaction is carried out in a solvent, wherein the solvent is dichloromethane.

[0035] As an optional implementation, in step S1, the condensation reaction is carried out in the presence of a condensing agent selected from O-benzotriazole-tetramethylurea hexafluorophosphate.

[0036] As an optional implementation, in step S1, the condensation reaction is carried out in the presence of a base selected from N,N-diisopropylethylamine.

[0037] As an optional implementation, in step S1, the molar ratio of FMOC-LYS(DDE)-OH to the 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.

[0038] As an optional implementation, step S1 further includes a post-processing step after the condensation reaction is completed.

[0039] As an optional implementation, in step S2, the processing temperature is 30~40℃, for example, it can be 30℃, 31℃, 32℃, 33℃, 34℃, 35℃, 36℃, 37℃, 38℃, 39℃, 40℃, etc., and the deprotection processing time is 2~4h, for example, it can be 2h, 2.2h, 2.4h, 2.6h, 2.8h, 3h, 3.2h, 3.4h, 3.6h, 3.8h, 4h, etc.

[0040] As an optional implementation, in step S2, the molar ratio of FMOC-GLU-OTBU and HOBT is 1:(1.2~2), for example, it can be 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.

[0041] As an optional implementation, the condensation reaction in step S2 is carried out in the presence of a base, wherein the base is selected from N,N'-diisopropylcarbodiimide; As an optional implementation, in step S2, the molar ratio of FMOC-GLU-OTBU to the alkali 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.

[0042] As an optional implementation, in step S2, the molar ratio of the polypeptide to the indomethacin molecule 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.

[0043] As an optional implementation, in step S3, the temperature of the condensation reaction is 30~40℃, for example, it can be 30℃, 31℃, 32℃, 33℃, 34℃, 35℃, 36℃, 37℃, 38℃, 39℃, 40℃, etc., and the time of the condensation reaction is 2~4 h, for example, it can be 2 h, 3 h, 4 h, etc.

[0044] As an optional implementation, in step S3, the condensation reaction is carried out in a solvent, wherein the solvent is selected as N,N-dimethylformamide; As an optional implementation, in step S3, the molar ratio of DOTA(OtBu)3 to 1-hydroxybenzotriazole is 1:(1.2~2), for example, it can be 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.

[0045] As an optional implementation, in step S3, the molar ratio of DOTA(OtBu)3 to the 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.

[0046] As an optional implementation, in step S5, the radioactive labeling includes the following steps: Sodium acetate solution, compound solution of Formula I, and radioactive nuclide [ 68 The GaCl3 solution was mixed and reacted to obtain the COX-2 targeting nuclide probe.

[0047] As an optional implementation, the volume ratio of the sodium acetate solution to the radionuclide solution is 1:1.

[0048] As an optional implementation, the concentration of the sodium acetate solution is 1~3 M, for example, it can be 1 M, 1.2 M, 1.5 M, 1.8 M, 2 M, 2.2 M, 2.5 M, 2.8 M, 3 M, etc., preferably 2 M, and the pH is 4.

[0049] As an optional implementation, the concentration of the compound solution represented by Formula I is 5 × 10⁻⁶. -6 M.

[0050] As an optional implementation, the radioactivity of the radionuclide solution is 1~10 mCi, for example, it can be 1 mCi, 2 mCi, 3 mCi, 4 mCi, 5 mCi, 6 mCi, 7 mCi, 8 mCi, 9 mCi, 10 mCi, etc., preferably 5 mCi.

[0051] As an optional implementation, in step S5, the temperature of the radioactive labeling is 85~95℃, for example, it can be 85℃, 86℃, 87℃, 88℃, 89℃, 90℃, 91℃, 92℃, 93℃, 94℃, 95℃, etc., and the radioactive labeling time is 5~20 min, for example, it can be 5 min, 6 min, 8 min, 10 min, 12 min, 14 min, 15 min, 16 min, 18 min, 20 min, etc.

[0052] In a preferred embodiment, step S5 specifically includes the following steps: Using a 5 mL EP tube as the reaction vessel, 1 mL of 2M sodium acetate solution (pH=4), 10 μg of the compound solution shown in Formula I, and 1 mL of […] were added sequentially. 68 A GaCl3 radionuclide solution (5 mCi) was used to heat the mixed leaves to 90°C and react for 10 min to obtain the COX-2 targeting nuclide probe. 68 Ga-NY-COX-2-DOTA 68 Ga-NY-COX-2-NOTA.

[0053] According to a third aspect of the invention, there is provided the use of a COX-2 targeting nuclide probe or its racemic, stereoisomer, or pharmaceutically acceptable salt in the preparation of a reagent for tumor imaging.

[0054] This invention relates to the application of a COX-2-targeting nuclide probe, specifically a compound of Formula I, or its precursor compound, isotopic compound, salt, or hydrate. This compound, when labeled with a radionuclide, yields a class of COX-2-targeting nuclide probes, which can be used as diagnostic and therapeutic agents in human or animal lesions with high COX-2 protein expression, particularly as tumor imaging agents and radionuclide therapeutic drugs. In animal experiments, the radiocomplex of this invention exhibits a high tumor / muscle ratio, demonstrating promising application prospects.

[0055] As an optional implementation, the tumor includes any one of triple-negative breast cancer, ovarian cancer, non-small cell lung cancer, pancreatic cancer, and glioblastoma.

[0056] As an optional implementation, the present invention provides a liver cancer tumor imaging model and a method for establishing the same, specifically including the following steps: inoculating a COX-2 highly expressed triple-negative breast cancer cell line into the axilla of a mouse, and injecting the COX-2 nuclide probe described in the present invention via tail vein injection.

[0057] It is important to note that after the triple-negative breast cancer imaging model is established, small animal PET imaging equipment is used to visualize the small molecules in triple-negative breast cancer.

[0058] According to a fourth aspect of the invention, there is provided the use of a COX-2 targeting nuclide probe or its racemic, stereoisomer, or pharmaceutically acceptable salt in the preparation of a reagent for identifying COX-2 overexpressing tumors.

[0059] To facilitate understanding of the above technical solutions of the present invention, the present invention will be further described below through embodiments. Unless otherwise specified, the materials in the embodiments are prepared according to existing methods or purchased directly from the market.

[0060] As mentioned in this 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 that has pharmaceutical value, such as a salt formed as an intermediate during chiral resolution, although such intermediate salt cannot be directly given to the subject, but can play a role in obtaining the end product of this invention.

[0061] As mentioned in this invention, a pharmaceutically acceptable salt of the compound represented by Formula I is a salt formed with an alkali or alkali metal. Acids that form pharmaceutically acceptable salts with the compound represented by Formula I include inorganic acids and organic acids. More specifically, alkali metals that form pharmaceutically acceptable salts with the compound represented by Formula I include, but are not limited to, lithium, sodium, potassium, magnesium, calcium, aluminum, zinc, etc.; bases that form pharmaceutically acceptable salts with the compound represented by Formula I include, but are not limited to, choline, diethanolamine, morpholine, etc.

[0062] Example 1 This embodiment provides a COX-2 targeting probe, wherein the COX-2 targeting probe is a radioactive nuclide. 68 Ga-marked [ 68 The structural formula of Ga]Ga-NY-COX-2-DOTA is as follows: ; The radionuclide 68 NY-COX-2-DOTA marked with Ga (denoted as Ga) 68 The crafting route for Ga-NY-COX-2-DOTA is as follows: (1) The resin type selected is 2-CL resin, and the synthesis method is FMOC solid-phase synthesis. The synthesis starts from the C-terminus of the peptide. The FMOC-LYS(DDE)-OH and the resin are condensed using condensation reagents HBTU / DIEA and DCM as solvents. The reaction formula is shown below: ; (2) FMOC was removed using a DMF solution containing 20% ​​piperidine for 20 min. After removal, the resin was washed with DMF 4 times, 1 min each time. The resin peptide was dehydrated and condensed using DIC+HOBT as a condensation reagent and DMF as a solvent. This step was repeated to condense FMOC-GLU-OTBU and small molecules sequentially, as shown in the following reaction formula: ; (3) DDE is removed by hydrazine hydrate, and DOTA-NHS is solvent-coupled by condensation reagent DIEA / DMF, as shown in the following reaction formula: ; (4) A lysis buffer with a ratio of 95% TFA, 1% H2O, 2% EDT, and 2% TIS was used. The lysis time was 2 hours. After lysis, the lysate was filtered into ice-cold ether and centrifuged to obtain the crude product. Then, HPLC separation was performed to prepare a purity of 95%. The pure product was obtained by freeze-drying. The reaction formula is shown below: ; (5) Using a 5 mL EP tube as the reaction vessel, add 1 mL of 2 M pH=4 sodium acetate solution, 10 μg of the compound NY-COX-2-DOTA shown in Formula I, and 1 mL of 68Ga solution (5 mCi) sequentially. Heat the mixture to 90 °C and react for 10 min to obtain the COX-2 targeting nuclide probe 68Ga-NY-COX-2-DOTA. The reaction formula is shown below: .

[0063] The structure was characterized by high performance liquid chromatography and mass spectrometry, and the structural determination results are as follows: like Figure 1 Shown, LCMS (ESI): m / z: Chemical Formula: C51H68ClN9O18, [MH]-found 1128.25, [M-2H]2H-found 564.05, [M-3H]-found 375.6; like Figure 2 As shown, 68 The purity of Ga-NY-COX-2-DOTA is 95.39%. like Figure 3 As shown, the tumor region exhibited characteristics at different time points in mice carrying 4T1 tumors, as studied using miniature PET / MR imaging. 68 Ga-NY-COX-2-DOTA was significantly absorbed by the tumor in the left anterior axillary region, with a clean background and clear imaging. This confirms the role of 4T1 tumors in... 68 Specific uptake of Ga-NY-COX-2-DOTA.

[0064] Application Example 1 This application example demonstrates the use of a COX-2-targeting nuclide probe in the specific identification of triple-negative breast cancer. Triple-negative breast cancer cells with high COX-2 expression were seeded under the armpit of Balb / c nude cells, and 3.33 MBq of the COX-2-targeting nuclide probe provided in Example 1 was injected via the tail vein. 68 The efficacy of Ga-NY-COX-2-DOTA molecules in diagnosing breast cancer was evaluated using a small animal PET / MR in vivo imaging system.

[0065] Figure 3 The COX-2 targeting probe provided in Example 1 68 A diagram illustrating the specific targeting effect of Ga-NY-COX-2-DOTA on triple-negative breast cancer. (See diagram for reference.) Figure 3 As shown, at different time points after injection, triple-negative breast cancer-bearing mice exhibited high radioactive signals in the tumor area, indicating that... 68Ga-NY-COX-2-DOTA showed good specificity in identifying COX-2 overexpressing tumors. Based on in vivo data from female mice... 68 The biological distribution data of Ga-NY-COX-2-DOTA are shown below. Figure 4 The highest uptake was observed in the kidneys, with lower uptake in various non-target organs. The 2-hour ex vivo organ distribution showed a tumor-to-muscle ratio of 9.65.

[0066] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A nuclide probe targeting COX-2, characterized in that, The COX-2-targeting nuclide probe comprises a radiolabeled compound of Formula I: Formula I; Wherein, the Y group includes either group I or group II: The radionuclides include 55 Co、 68 Ga、 64 Cu、 86 Y、 89 Zr、 90 Y、 111 In、 177 Lu、 225 Any one of Ac.

2. The nuclide probe targeting COX-2 according to claim 1, characterized in that, The radionuclide-labeled compound of Formula I includes either NY-COX-2-NOTA or NY-COX-2-DOTA. The structural formula of NY-COX-2-NOTA is as follows: The structural formula of NY-COX-2-DOTA is: 。 3. A method for preparing a COX-2-targeting radionuclide probe according to any one of claims 1-2, characterized in that, Includes the following steps: S1. The resin type selected is 2-CL resin, and the synthesis method is FMOC solid-phase synthesis, starting from the C-terminus of the peptide. FMOC-LYS(DDE)-OH and the resin are condensed using condensation reagents HBTU / DIEA and DCM as solvents. The reaction formula is shown below: ； S2. FMOC is removed using a DMF solution containing 20% ​​piperidine for 20 minutes. After removal, the resin is washed with DMF four times, 1 minute each time. The resin peptide is then dehydrated and condensed using DIC+HOBT as a condensation reagent and DMF as a solvent. This step is repeated to sequentially condense FMOC-GLU-OTBU and the small molecule. The reaction formula is shown below: ； S3. DDE is removed by hydrazine hydrate, followed by solvent coupling of DOTA-NHS via the condensation reagent DIEA / DMF, as shown in the following reaction formula: ； S4. Using a lysis buffer with a ratio of 95% TFA, 1% H2O, 2% EDT, and 2% TIS, the lysis time was 2 hours. After lysis, the lysate was filtered into ice-cold diethyl ether and centrifuged to obtain the crude product. Then, HPLC separation was performed to obtain a purity of 95%, and the pure product was obtained by lyophilization. The reaction formula is shown below: ； S5. Combine sodium acetate solution, the compound solution shown in Formula I, and the radioactive nuclide [ 68 The GaCl3 solution was mixed and reacted to obtain the COX-2 targeting nuclide probe, wherein the reaction formula is shown below: 。 4. The method for preparing a COX-2-targeting radionuclide probe according to claim 3, characterized in that, In step S1, FMOC-LYS(DDE)-OH is formed by the condensation of DIEA and HBTU with the resin; And / or, in step S1, the temperature of the condensation reaction is 30~40℃ and the time is 2~4 h; And / or, in step S1, the condensation reaction is carried out in a solvent, and the solvent is dichloromethane; And / or, in step S1, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is O-benzotriazole-tetramethylurea hexafluorophosphate. And / or, in step S1, the molar ratio of FMOC-LYS(DDE)-OH to the condensing agent is 1:(1.2~2); And / or, in step S1, the condensation reaction is carried out in the presence of a base, wherein the base is N,N-diisopropylethylamine; And / or, in step S1, the molar ratio of FMOC-LYS(DDE)-OH to the base is 1:(0.5~5).

5. The method for preparing a COX-2-targeting radionuclide probe according to claim 3, characterized in that, In step S2, FMOC-GLU-OTBU undergoes dehydration condensation with the resin peptide using DIC / HOBT / DMF as a solvent; And / or, in step S2, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours. And / or, in step S2, the condensation reaction is carried out in a solvent, and the solvent is N,N-dimethylformamide; And / or, in step S2, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is 1-hydroxybenzotriazole; And / or, in step S2, the molar ratio of FMOC-GLU-OTBU to the condensing agent is 1:(1.2~2); And / or, in step S2, the condensation reaction is carried out in the presence of a base, wherein the base is N,N'-diisopropylcarbodiimide; And / or, in step S2, the molar ratio of FMOC-GLU-OTBU to the base is 1:(0.5~5).

6. The method for preparing a COX-2-targeting radionuclide probe according to claim 3, characterized in that, In step S3, the resin peptide is dehydrated and condensed with DOTA using DIC / HOBT / DMF as a solvent to obtain the resin peptide. The peptide and resin are separated using a DCM solution containing 20% ​​trifluoroethanol, rotary evaporated, and lyophilized to obtain the fully protected peptide. And / or, in step S3, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours; And / or, in step S3, the condensation reaction is carried out in a solvent, and the solvent is N,N-dimethylformamide; And / or, in step S3, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is 1-hydroxybenzotriazole; And / or, in step S3, the molar ratio of indomethacin to the condensing agent is 1:(1.2~2); And / or, in step S3, the condensation reaction is carried out in the presence of a base, wherein the base is N,N-diisopropylethylamine; And / or, in step S3, the molar ratio of indomethacin to the base is 1:(0.5~5); And / or, in step S3, the reagent used for separating the peptide and resin is a dichloromethane solution containing 20% ​​trifluoroethanol; And / or, in step S3, the temperature for deprotection treatment is 30~40℃, and the treatment time is 1~3 h.

7. The method for preparing a COX-2-targeting radionuclide probe according to claim 3, characterized in that, In step S4, the peptide and DOTA(OtBu)3 are mixed in equimolar amounts, DMF is used as the solvent, and DIEA / HBTU is used as the condensing agent to couple DOTA(OtBu)3, and then the mixture is lyophilized. And / or, in step S4, the condensation reaction is carried out at a temperature of 30-40°C for 2-4 hours; And / or, in step S4, the condensation reaction is carried out in a solvent, and the solvent is N,N-dimethylformamide; And / or, in step S4, the condensation reaction is carried out in the presence of a condensing agent, wherein the condensing agent is O-benzotriazole-tetramethylurea hexafluorophosphate. And / or, in step S4, the molar ratio of the peptide to the condensing agent is 1:(1.2~2); And / or, in step S4, the condensation reaction is carried out in the presence of a base, wherein the base is N,N-diisopropylethylamine; And / or, in step S4, the molar ratio of the polypeptide to the base is 1:(0.5~5); And / or, in step S4, the molar ratio of the peptide to DOTA(OtBu)3 is 1:(0.5~5).

8. The method for preparing a COX-2-targeting radionuclide probe according to claim 3, characterized in that, In step S6, the radioactive labeling includes the following steps: The sodium acetate solution, the compound solution shown in Formula I, and the radionuclide solution are mixed and reacted to obtain the COX-2 targeting nuclide probe. And / or, the volume ratio of sodium acetate solution to radioactive nuclide solution is 1:1; And / or, the concentration of the sodium acetate solution is 1~3 M, and the pH is 4; And / or, the concentration of the compound solution shown in Formula I is 5 × 10⁻⁶. -6 M; And / or, the radioactivity of the radionuclide solution is 1~10 mCi; And / or, the radiolabeling temperature is 85~95℃, and the radiolabeling time is 5~20 min.

9. The use of a COX-2 targeting probe or its racemic, stereoisomer, or pharmaceutically acceptable salt according to any one of claims 1-2 in the preparation of a reagent for tumor imaging; in, The tumors include any one of triple-negative breast cancer, ovarian cancer, non-small cell lung cancer, pancreatic cancer, and glioblastoma.

10. The use of a COX-2 targeting nuclide probe or its racemic, stereoisomer, or pharmaceutically acceptable salt thereof, according to any one of claims 1-2, in the preparation of a reagent for identifying COX-2 overexpressing tumors.

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