FAP-targeted polymeric compound as well as preparation method and application thereof

Abstract

The invention relates to the technical field of biological medicines, and discloses a FAP-targeted polymeric compound as well as a preparation method and application thereof. The chemical structure of the FAP-targeting polymeric compound is shown as any one of formulas (1)-(3), and the FAP-targeting polymeric compound comprises 3-6 targeting parts of fibroblast activating protein alpha (FAP-alpha). Compared with the existing compound, the FAP-targeted polymeric compound provided by the invention has higher tumor uptake and longer tumor residence time; in the diagnosis and treatment of FAP positive tumors, the FAP-targeted polymeric compound can clearly display the position and range of the tumors, discover the distant metastasis of the tumors in time, and provide more comprehensive disease assessment.

Description

Technical Field

[0001] This application relates to the field of biomedical technology, specifically to a polymeric compound targeting FAP, its preparation method, and its application. Background Technology

[0002] Tumor-associated fibroblasts (TAFs) are a crucial component of the tumor microenvironment, and the fibroblast activating protein (FAP) is widely recognized as a marker of TAFs, promoting tumor development and invasion. FAP belongs to the proline oligopeptidase family and is a type II transmembrane serine protein. Uniquely, it possesses both endopeptidase and exopeptidase activities and is highly expressed in the stroma of epithelial tumors (and other diseases involving tissue remodeling), while being expressed at low levels or not at all in normal tissues. This makes it a valuable target for tumor imaging and therapy.

[0003] FAP inhibitors (FAPIs) exhibit extremely high affinity and specificity for FAP and are used in the diagnosis of various solid tumors. Most existing FAPI small molecule compounds have a core structure of (4-quinolineyl)-glycyl-2-cyanopyrrolidine and have made significant progress in targeted tumor imaging. A research group in Heidelberg, Germany, developed the first batch of FAP-targeting drugs, FAPI-03 to FAPI-15, using this core structure. Among them, 68Ga-FAPI-04 showed the best efficacy and excellent tumor visualization capabilities. Kratochwil et al. applied 68Ga-FAPI-04 to 80 patients with 28 different types of solid tumors and found that 68Ga-FAPI-04 performed well in the vast majority of lesions. The subsequent development of 68Ga-FAPI-46 further improved drug uptake and retention time in tumors, and compared to 68Ga-FAPI-04, it had a better tumor-to-background ratio. Case reports on the application of 68Ga-FAPI in tumor diagnosis are numerous, indicating its great development potential.

[0004] The application of FAPIs in tumors and other diseases characterized by fibroblast activator protein overexpression is closely related to their structure. Structural analysis and design of FAPIs are crucial for producing FAPIs with excellent metabolic properties and for their application in disease diagnosis and treatment. Therefore, designing novel FAPI inhibitors with superior pharmacokinetic properties is of great significance. Summary of the Invention

[0005] This application provides a polymeric compound targeting FAP, its preparation method, and its application, aiming to develop a novel FAP inhibitor with excellent pharmacokinetic properties, and to use it as a drug for the diagnosis and treatment of various diseases caused by the overexpression of fibroblast activator protein (FAP).

[0006] To achieve the above objectives, the present application adopts the following technical solution.

[0007] In a first aspect, this application provides a polymeric compound targeting FAP, the chemical structure of which is shown in any one of formulas (1) to (3):

[0008] (1)

[0009] (2)

[0010] (3)

[0011] Where n and m are selected from any integers from 0 to 12;

[0012] Structural unit B is the targeting portion of fibroblast activation protein α (FAP-α);

[0013] Structural units D and Y are linking groups, one of which may be absent or both may be absent, and are selected from any of the following structures:

[0014]

[0015] Where n1, n2, and n3 are any integers from 1 to 12, X1 and Y1 are selected from C, N, O, and S, and R1 and R2 are each independently selected from hydrogen or any of the following structures:

[0016]

[0017] Where X is a halogen;

[0018] Structural unit E is selected from any of the following structures:

[0019]

[0020] Structural unit E2 is selected from any of the following structures:

[0021]

[0022] Structural unit E3 is selected from any of the following structures:

[0023]

[0024] The structural unit Z is selected from at least one of radioactive components, chelating agents, fluorescent dyes, contrast agents, or cytotoxic substances.

[0025] Preferably, the general structural formula of structural unit B is as follows:

[0026]

[0027] Among them, R a R b Each is independently selected from hydrogen, hydroxyl, and C. 1-6 Hydrocarbon group, mercapto group or halogen;

[0028] R c Selected from H, -CN, -B(OH)2, chloroacetyl, -C(O)alkyl, -C(O)aryl, -C=CC(O)aryl, -C=CS(O)2aryl, -CO2H, -NH2, urea, -SO3H, -PO3H2, ketoamide, trifluoromethyl, trifluoroacetyl, and tetrazolyl;

[0029] R d Selected from H, C 1-8 Straight-chain or branched hydrocarbon groups or hydroxyl groups;

[0030] R e Selected from aromatic monocyclic or bicyclic compounds priced between 5 and 10 yuan, non-aromatic monocyclic or bicyclic compounds priced between 5 and 10 yuan, aromatic monocyclic heterocyclic or bicyclic heterocyclic compounds priced between 5 and 10 yuan, and non-aromatic monocyclic heterocyclic or bicyclic heterocyclic compounds priced between 5 and 10 yuan with R e Excluding the pyridine ring;

[0031] The heterocycle contains 1 to 3 heteroatoms selected from O, N, or S, R e Hydrogen can be converted into halogens, -NHC 1-8 Alkyl, -OH substitution; when R e For a time At that time, it does not contain amino substituents;

[0032] R f The general structural formula is , where R f1 R f2 R f3 R f4 Each exists or does not exist;

[0033] R f1 For O, S, NR g R g It can be hydrogen, halogen, hydroxyl, mercapto or C 1-8 alkyl;

[0034] R f2 C 1-8 alkyl or n4 = any integer from 0 to 8;

[0035] R f3 It is an aromatic or non-aromatic monocyclic nitrogen-containing heterocycle; selected from any of the following structural species:

[0036]

[0037] R f4 It does not exist or is selected from any of the following structures:

[0038]

[0039] Where n1, n2, and n3 are any integers from 1 to 12.

[0040] Further preferred, R e Selected from any of the following structures, where the ring and any site of the substituents on the ring can be extended to connect R. f :

[0041] .

[0042] More preferably, structural unit B includes any one of the following chemical structures:

[0043]

[0044]

[0045] .

[0046] Preferably, the chelating agent comprises any one of DOTA, DOTAGA, NOTA, NOTAGA, TETA, DAPA, TRAP, HBED-CC, DFO, PCTA, DTPA, HYNIC, ATSM, EDTA, CB-TE2A, NOPO, NOTAM, MAG3, Deferoxamine, Cyclen, H3RESCA, Octaaminocryptands, diaza-18-crown ether-6, NTA, CB-cyclam, or isocyanate;

[0047] The radioactive component is selected from isotopes that emit alpha rays, beta rays, gamma rays, Auger electrons, X-rays, or fluorescence;

[0048] The fluorescent dyes include at least one of xanthine, acridine, azine, cyanine, styryl dyes, coumarin, porphyrin, metal ligand complexes, fluorescent proteins, nanocrystals, perylene, borodipyrrole methylene or phthalocyanine;

[0049] The cytotoxic substance includes at least one of MMAE, MMAF, DM1, PNU-159682, Cryptophycin-55, PBD, or Epothilone B.

[0050] A second aspect of this application provides a pharmaceutical composition comprising the aforementioned polymeric compound targeting FAP.

[0051] A third aspect of this application provides an FAP inhibitor comprising the aforementioned FAP-targeting polymeric compound, a salt or isomer of the aforementioned FAP-targeting polymeric compound.

[0052] A fourth aspect of this application provides the use of the above-described pharmaceutical composition or FAP inhibitor in the preparation of a medicament for treating or detecting a disease characterized by overexpression of the fibroblast activating protein FAP.

[0053] Preferably, the disease characterized by overexpression of fibroblast activator protein FAP includes any one of cancer, chronic inflammation, atherosclerosis, fibrosis, tissue remodeling, or scarring.

[0054] Compared with the prior art, the beneficial effects of this application are as follows:

[0055] The FAP-targeting polymeric compound of this application exhibits better FAP affinity than existing compounds, and in imaging experiments, 177 Lu-labeled radiocomplexes showed significant enrichment in the tumor region 1 hour after intravenous injection, and the retention time could extend up to 144 hours, compared with existing... 177 Compared to Lu-FAP-2286, the FAP-targeting polymeric compound of this application exhibits higher tumor uptake and longer tumor retention time. In the diagnosis and treatment of FAP-positive tumors, the FAP-targeting polymeric compound can clearly show the location and extent of the tumor, promptly detect distant metastases, and provide a more comprehensive assessment of the disease. Attached Figure Description

[0056] To more clearly illustrate the technical solutions of the embodiments of this application, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are only some embodiments recorded in this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0057] Figure 1 LC-MS spectrum of FAP-targeting polymer compound 3(FAPI-04)-DOTA;

[0058] Figure 2 UV spectrum of polymer compound 3 (FAPI-04)-DOTA targeting FAP;

[0059] Figure 3 for 68 Ga-3(FAPI-04)-DOTA and 177Radiometric high-performance liquid chromatography (HPLC) analysis chromatogram of Lu-3(FAPI-04)-DOTA;

[0060] Figure 4 for 68 Representative PET / CT images of Ga-3(FAPI-04)-DOTA in U87MG tumor-bearing mice;

[0061] Figure 5 for 177 Representative SPECT images of Lu-3(FAPI-04)-DOTA in U87MG tumor-bearing mice;

[0062] Figure 6 for 177 Representative SPECT images of Lu-FAP-2286 in U87MG tumor-bearing mice. Detailed Implementation

[0063] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of this application. All other embodiments obtained by those skilled in the art based on the embodiments of this application without creative effort are within the scope of protection of this application.

[0064] In the following description of this embodiment, the terms "including", "comprising", "having", and "containing" are all open-ended terms, meaning that they include but are not limited to.

[0065] In the following description of this embodiment, the term "and / or" is used to describe the association relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent: A existing alone, B existing alone, and A and B existing simultaneously. A and B can be singular or plural. The character " / " generally indicates that the preceding and following related objects have an "or" relationship.

[0066] In the following description of this embodiment, the term "at least one" means one or more, and "more than one" means two or more. "At least one of the following" or similar expressions refer to any combination of these items, including any combination of single or multiple items. For example, "at least one of a, b, or c", or "at least one of a, b, and c", can both mean: a, b, c, ab (i.e., a and b), ac, bc, or abc, where a, b, and c can be single or multiple.

[0067] The terminology used in the embodiments of this application is for the purpose of describing particular embodiments only and is not intended to be limiting of this application. The singular forms "a" and "the" as used in the embodiments of this application and the appended claims are also intended to include the plural forms, unless the context clearly indicates otherwise.

[0068] Those skilled in the art should understand that, in the following description of the embodiments of this application, the sequence of numbers does not imply the order of execution. Some or all steps may be executed in parallel or sequentially. The execution order of each process should be determined by its function and internal logic, and should not constitute any limitation on the implementation process of the embodiments of this application.

[0069] Those skilled in the art will understand that the numerical ranges in the embodiments of this application should be understood as each intermediate value between the upper and lower limits of the specifically disclosed range. Each smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this application. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0070] Unless otherwise stated, the technical / scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains. While this application describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this application. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0071] In a first aspect, this application provides a polymeric compound targeting FAP, the chemical structure of which is shown in any one of formulas (1) to (3):

[0072] (1)

[0073] (2)

[0074] (3)

[0075] Where n and m are selected from any integers from 0 to 12;

[0076] Structural unit B is the targeting portion of fibroblast activation protein α (FAP-α);

[0077] Structural units D and Y are linking groups, one of which may be absent or both may be absent, and are selected from any of the following structures:

[0078]

[0079] Where n1, n2, and n3 are any integers from 1 to 12, X1 and Y1 are selected from C, N, O, and S, and R1 and R2 are each independently selected from hydrogen or any of the following structures:

[0080]

[0081] Where X is a halogen;

[0082] Structural unit E is selected from any of the following structures:

[0083]

[0084] Structural unit E2 is selected from any of the following structures:

[0085]

[0086] Structural unit E3 is selected from any of the following structures:

[0087]

[0088] The structural unit Z is selected from at least one of radioactive components, chelating agents, fluorescent dyes, contrast agents, or cytotoxic substances.

[0089] In this application, structural unit B is the targeting portion of fibroblast activation protein α (FAP-α), and its general structural formula is shown below:

[0090]

[0091] Among them, R a R b Each is independently selected from hydrogen, hydroxyl, and C. 1-6 Hydrocarbon group, mercapto group or halogen;

[0092] R c Selected from H, -CN, -B(OH)2, chloroacetyl, -C(O)alkyl, -C(O)aryl, -C=CC(O)aryl, -C=CS(O)2aryl, -CO2H, -NH2, urea, -SO3H, -PO3H2, ketoamide, trifluoromethyl, trifluoroacetyl, and tetrazolyl;

[0093] R d Selected from H, C 1-8 Straight-chain or branched hydrocarbon groups or hydroxyl groups;

[0094] R e Selected from aromatic monocyclic or bicyclic compounds priced between 5 and 10 yuan, non-aromatic monocyclic or bicyclic compounds priced between 5 and 10 yuan, aromatic monocyclic heterocyclic or bicyclic heterocyclic compounds priced between 5 and 10 yuan, and non-aromatic monocyclic heterocyclic or bicyclic heterocyclic compounds priced between 5 and 10 yuan with Re Excluding the pyridine ring;

[0095] The heterocycle contains 1 to 3 heteroatoms selected from O, N, or S, R e Hydrogen can be converted into halogens, -NHC 1-8 Alkyl, -OH substitution; when R e For a time When it does not contain amino substituents.

[0096] Among them, R e Preferably, any of the following compounds, wherein the ring and any site of the substituents on the ring can be extended to connect R. f :

[0097]

[0098] R f The general structural formula is , where R f1 R f2 R f3 R f4 Each exists or does not exist. Specifically,

[0099] R f1 For O, S, NR g R g It can be hydrogen, halogen, hydroxyl, mercapto or C 1-8 alkyl;

[0100] R f2 C 1-8 alkyl or n4 = any integer from 0 to 8;

[0101] R f3 It is an aromatic or non-aromatic monocyclic nitrogen-containing heterocycle; selected from any of the following structural species:

[0102]

[0103] R f4 It does not exist or is selected from any of the following structures:

[0104]

[0105] Where n1, n2, and n3 are any integers from 1 to 12.

[0106] It should be noted that in this application, structural unit B may also be selected from self-cleaving or custom-made units, or non-self-cleaving units that can be selectively degraded by in vivo pH or enzymes, such as Val-Cit which can be cleaved by cathepsins.

[0107] In this application, the structural unit Z is selected from at least one of radioactive components, chelating agents, fluorescent dyes, contrast agents, or cytotoxic substances.

[0108] Wherein, when Z is a chelating agent, it is a chelating agent that binds to divalent or trivalent metal cations, preferably any one of DOTA, DOTAGA, NOTA, NOTAGA, TETA, DAPA, TRAP, HBED-CC, DFO, PCTA, DTPA, HYNIC, ATSM, EDTA, CB-TE2A, NOPO, NOTAM, MAG3, Deferoxamine, Cyclen, H3RESCA, Octaaminocryptands, diaza-18-crown ether-6, NTA, CB-cyclam, or isocyanate; more preferably, the chelating agent is any one of the following compounds:

[0109]

[0110] Wherein, when structural unit Z is a radioactive component, it is selected from radioactive isotopes, fluorescent isotopes, or combinations thereof, such as isotopes emitting alpha rays, beta rays, gamma rays, Auger electrons, X-rays, and fluorescence. Specifically, the radioactive component can be selected from 161Tb, 225Ac, 18F, 67Ga, 68Ga, 111In, 99mTc, 186Re, 188Re, 140La, 153Sm, 90Y, 165Dy, 169Er, 177Lu, 47Sc, 142Pr, 159Gd, 212Bi, 213Bi, 72As, 72Se, 97Ru, 109Pd, 105Rh, 119Sb, 123I, 12 The following are all of the following: 4I, 131I, 197Hg, 211At, 151Eu, 153Eu, 169Eu, 201Tl, 203Pb, 212Pb, 64Cu, 67Cu, 188Re, 186Re, 198Au, 225Ac, 227Th or 199Ag; more preferably, the following are all of the following: 68Ga, 177Lu, 18F, 99mTc, 188Re, 212Pb, 64Cu, 211At or 225Ac.

[0111] Wherein, when the structural unit Z is a fluorescent dye, it is selected from at least one of xanthine, acridine, azine, cyanine, styryl dye, coumarin, porphyrin, metal ligand-complex, fluorescent protein, nanocrystal, perylene, borodipyrrole methylene or phthalocyanine.

[0112] Wherein, when structural unit Z is a cytotoxic substance, it is selected from at least one of MMAE, MMAF, DM1, PNU-159682, Cryptophycin-55, PBD or Epothilone B.

[0113] When the polymeric compound targeting FAP contains three structural units B, structural unit E is absent, and structural unit Z contains multiple carboxyl groups, The structure of the polymeric compound that is directly linked to Z and targets FAP is as follows:

[0114]

[0115] Wherein, the structural unit Z is preferably:

[0116]

[0117] As a preferred embodiment of this application, the polymeric compound targeting FAP contains four structural units B, structural unit E is absent, and structural unit Z has multiple carboxyl groups. Connecting directly to Z yields the following compound structure:

[0118]

[0119] Wherein, the structural unit Z is preferably:

[0120]

[0121] This application also provides a pharmaceutical composition comprising the above-mentioned polymeric compound targeting FAP, and other ingredients.

[0122] This application also provides an FAP inhibitor, comprising any one of the above-mentioned FAP-targeting polymeric compounds, salts of the above-mentioned FAP-targeting polymeric compounds, or isomers of the above-mentioned FAP-targeting polymeric compounds, or a mixture of two or more thereof. Compared with existing compounds, the FAP-targeting polymeric compound exhibits higher tumor uptake and longer tumor retention time. In the diagnosis and treatment of FAP-positive tumors, the FAP-targeting polymeric compound can clearly show the location and extent of the tumor, promptly detect distant metastases, and provide a more comprehensive assessment of the disease.

[0123] The pharmaceutical composition and FAP inhibitor of this application can be used to prepare drugs for treating / detecting diseases characterized by overexpression of the fibroblast activating protein FAP. The diseases characterized by overexpression of the fibroblast activating protein FAP include any one of cancer, chronic inflammation, atherosclerosis, fibrosis, tissue remodeling, or scarring.

[0124] In this application, structural unit B is preferably any one of the following chemical structures:

[0125]

[0126]

[0127] .

[0128] When structural unit B is selected from the above compounds, the specific preparation method of the polymeric compound targeting FAP is as follows:

[0129] (1) When structural unit B is (denoted as R) 1-1 When the structural unit Z is DOTAGA, the synthesis process is as follows:

[0130]

[0131] Its preparation methods include:

[0132] S1, Compound 1 is tert-butyl esterified to give Compound 2; the molar ratio of Compound 1 to tert-butanol is 1:10-1:20;

[0133] S2, Compound 2 reacts with 3-methylaminopropanol to introduce a nitrogen-methyl group, yielding Compound 3; preferably, the molar ratio of Compound 2 to 3-methylaminopropanol is 1:4~8;

[0134] S3, Compound 3 is given by introducing a -Boc-protected piperazine group to obtain Compound 4; preferably, the molar ratio of Compound 3 to 1-Boc-piperazine is 1:15~25;

[0135] S4, compound 4, after removing the tert-butyl protecting group, yields compound 5, which undergoes an amide condensation reaction with (S)-1-(2-aminoacetyl)-4,4-difluoropyrrolidine-2-carboxynitrile to give compound 6, i.e., R. 1-1 -BOC; the preferred molar ratio of compound 4 to (S)-1-(2-aminoacetyl)-4,4-difluoropyrrolidine-2-carboxynitrile is 1:1.1~1.5;

[0136] S5, Compound 6, after removing the -Boc protecting group, undergoes an amide condensation reaction with 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid to give Compound 7; preferably, the molar ratio of Compound 6 to 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy)) is 4~6:1;

[0137] S6, compound 7 undergoes a de-Boc protecting group removal and an amide condensation reaction with DOTAGA-NHS to yield compound 8, the polymeric compound targeting FAP. The preferred molar ratio of compound 7 to DOTAGA-NHS is 1:1.2~1.5.

[0138] (2) When structural unit B is (denoted as R) 1-1 When the structural unit Z is DOTAGA, the synthesis process is as follows:

[0139]

[0140] Its preparation methods include:

[0141] S1, compound 1 is demethylated to give compound 2;

[0142] S2, Compound 2 reacts with 1-bromo-3-chloropropane to introduce a chloropropyl group, yielding Compound 3; preferably, the molar ratio of Compound 2 to 1-bromo-3-chloropropane is 1:3~5.

[0143] S3, compound 3 is given by introducing a Boc-protected piperazine group to obtain compound 4; preferably, the molar ratio of compound 3 to 1-Boc-piperazine is 1:15~25.

[0144] S4, compound 4, by introducing (S)-1-(2-aminoacetyl)-4,4-difluoropyrrolidine-2-carboxynitrile, yields compound 5, namely R. 1-1 -BOC; preferably, the molar ratio of compound 4 to (S)-1-(2-aminoacetyl)-4,4-difluoropyrrolidine-2-carboxynitrile is 1:1.1~1.5.

[0145] S5, compound 5, after removing the -Boc protecting group, undergoes an amide condensation reaction with 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid to give compound 6; preferably, the molar ratio of compound 5 to 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy)) is 4~6:1.

[0146] S6, compound 6, after removing the -Boc protecting group, undergoes an amide condensation reaction with DOTAGA-NHS to yield compound 7, which is the polymeric compound targeting FAP. The preferred molar ratio of compound 6 to DOTAGA-NHS is 1:1.2~1.5.

[0147] (5) When structural unit B is (denoted as R) 1-1 The structural unit Z is DOTAGA, and the synthesis process is as follows:

[0148]

[0149] Its preparation methods include:

[0150] S1, The preparation method of compound 1 is the same as steps S1-S4 in Example 2;

[0151] S2, compound 1 undergoes a condensation reaction with N-α-fluorenemethoxycarbonyl-N-ε-tert-butyloxycarbonyl-L-lysine after the -Boc protecting group is removed, to generate compound 2; preferably, the molar ratio of compound 1 to N-α-fluorenemethoxycarbonyl-N-ε-tert-butyloxycarbonyl-L-lysine is 1:2~3.

[0152] S3, compound 2, after removing the -Boc protecting group, undergoes a condensation reaction with 3-((fluorosulfonyl)oxy)benzoic acid to generate compound 3, i.e., R. 1-1 -BOC; the preferred molar ratio of compound 3 to 3-((fluorosulfonyl)oxy)benzoic acid is 1:1.1~1.3.

[0153] S4, compound 3, after removing the -Boc protecting group, undergoes an amide condensation reaction with 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid to give compound 4; preferably, the molar ratio of compound 3 to 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy)) is 4~6:1.

[0154] Step 5: Compound 4 undergoes an amide condensation reaction after the -Boc protecting group is removed, yielding compound 5, the FAP-targeting polymer. The preferred molar ratio of compound 4 to DOTAGA-NHS is 1:1.2~1.5.

[0155] The present application will be further described below through specific embodiments.

[0156] Example 1

[0157] This embodiment provides a method for preparing a polymeric compound targeting FAP, the chemical structure of which is shown below:

[0158]

[0159] Its synthesis process is as follows:

[0160]

[0161] The preparation methods specifically include:

[0162] 6-Methoxyquinoline-4-carboxylic acid was demethylated with hydrobromic acid to obtain 6-hydroxyquinoline-4-carboxylic acid. 38 mg (100 μmol) of 6-hydroxyquinoline-4-carboxylic acid and 325.8 mg (1 mmol) of cesium carbonate were added to 500 μL of DMF and stirred until homogeneous. 60 mg (350 μmol) of 1-bromo-4-chloropropane was added while stirring, and the mixture was heated to 60 °C and reacted for 8 h. After the reaction was complete, the reaction system was diluted with 500 μL of water and 1 mL of acetonitrile, and 200 μL of 6 M NaOH was added. The reaction was then directly purified by HPLC to obtain the product 7-(3-chloropropoxy)quinoline-4-carboxylic acid, with a yield of 65%.

[0163] 20 mg (75.4 μmol) of 7-(3-chloropropoxy)quinoline-4-carboxylic acid, 70 mg (377 μmol) of N-tert-butoxycarbonyl-piperazine, and 37.4 mg (226 μmol) of [a specific ingredient] were dissolved in 1 mL of DMF and stirred overnight at 60 °C. After the reaction was complete, the reaction mixture was diluted with an equal volume of water, and then purified by HPLC to give the product 7-(3-(4-tert-butoxycarbonylpiperazine-1-yl)propoxy)quinoline-4-carboxylic acid, in 90% yield.

[0164] 7-(3-(4-tert-butoxycarbonylpiperazin-1-yl)propoxy)quinoline-4-carboxylic acid (31.3 mg, 117.9 μmol) and HATU (53.8 mg, 141.4 μmol) were dissolved in 2 ml of DMF, and N,N-diisopropylethylamine (15 mg, 117.9 μmol) was added dropwise while stirring. After stirring for ten minutes, (S)-1-(2-aminoacetyl)-4,4-difluoropyrrolidine-2-carboxynitrile (24.5 mg, 129.7 μmol) and N,N-diisopropylethylamine (15 mg, 117.9 μmol) were added. After reacting overnight, the reaction was diluted with an equal volume of water and purified by HPLC to give the product (S)-N-(-)-phenylalanine-2-(2-cyano-4,4-difluoropyrrolidone-1-yl)-2-oxoethyl-6-(3-(4-tert-butoxycarbonylpiperazin-1-yl)propoxy)quinoline-4-carboxamide, with a yield of 80%.

[0165] (S)-N-(-)-phenylalanine-2-(2-cyano-4,4-difluoropyrrolidine-1-yl)-2-oxoethyl-6-(3-(4-tert-butoxycarbonylpiperazin-1-yl)propoxy)quinoline-4-carboxamide (35.4 mg, 60 μmol) was deprotected with trifluoroacetic acid. The reaction was monitored by TLC until completion. The solvent was removed under vacuum, and the product was dissolved in 1.5 ml of DMF. 3,3'-((2-((tert-butoxycarbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (8.6 mg, 20 μmol) and HATU (27.4 mg, 72 μmol) were added. DIPEA (11.6 mg, 90 μmol) was added dropwise while stirring. The reaction was allowed to proceed overnight. The reaction was monitored by TLC until completion. The solvent was removed under vacuum, and the product 6 was purified by HPLC with a yield of 50%.

[0166] Product 6 was deprotected with trifluoroacetic acid to remove the Boc protecting group and dissolved in 1 ml DMF. DOTAGA-tetratert-butyl ester (8.4 mg, 12 μmol), HATU (5.7 mg, 15 μmol), and DIPEA (2.6 mg, 20 μmol) were added. The reaction was carried out overnight. The reaction was monitored by TLC until it was complete. The solvent was removed under vacuum, and trifluoroacetic acid was added and stirred for 16 h. The product was purified by HPLC to obtain the final product compound 7 (11 mg, 50%), which is the FAP-targeting polymer compound, denoted as 3(FAPI-04)-DOTA.

[0167] The HPLC conditions were as follows: radioactive high-performance liquid chromatography with an Agilent analytical column (250 × 4.6 mm). The mobile phase consisted of an aqueous solution of 0.1% trifluoroacetic acid (A) and acetonitrile (B). The elution program was to increase the acetonitrile concentration from 10% to 90% over 0 to 15 minutes, with a flow rate of 1.0 mL / min and a UV detector wavelength of 254 nm.

[0168] The FAP-targeting polymer compound 3 (FAPI-04)-DOTA prepared in Example 1 was tested and evaluated, and its LC-MS spectrum is shown below. Figure 1 As shown, this indicates the successful synthesis of the target product. Its UV spectrum is as follows. Figure 2 As shown. From Figure 2 It can be seen that the purity of 3(FAPI-04)-DOTA is greater than 95%.

[0169] Animal experiments were conducted on the FAP-targeting polymer compound 3 (FAPI-04)-DOTA prepared in Example 1, as follows:

[0170] 1. Mark and quality control 3(FAPI-04)-DOTA:

[0171] 1.1. Take 1 mL 68 Add an appropriate amount of sodium acetate buffer (pH=4.45) to GaCl3 solution (2 mCi / mL), then add 10 nmol / 10 μL of 3(FAPI-04)-DOTA, heat to 95 °C for 15 min to carry out the reaction, and obtain... 68 Ga-3(FAPI-04)-DOTA.

[0172] After the reaction was completed and cooled to room temperature, the labeled precursor was purified by passing it through a pre-activated HLB column (10 mg specification). 68 Ga-3(FAPI-04)-DOTA was analyzed using Radio-HPLC to determine the retention time and radiochemical purity of the marker. The resulting chromatogram is shown below. Figure 3 As shown. By Figure 3 It can be seen that its radiochemical purity is greater than 95%.

[0173] 1.2. Take 0.1 mL 177 Add an appropriate amount of sodium acetate buffer (pH=4.45) to LuCl3 solution (1 mCi / 0.1 mL), then add 10 nmol / 10 μL of 3(FAPI-04)-DOTA, heat to 95 °C for 15 min to carry out the reaction, and obtain... 177 Lu-3(FAPI-04)-DOTA.

[0174] After the reaction was completed and cooled to room temperature, the labeled precursor was purified by passing it through a pre-activated HLB column (10 mg specification). 177 Lu-3(FAPI-04)-DOTA was analyzed using Radio-HPLC to determine the retention time and radiochemical purity of the marker. The resulting chromatogram is shown below. Figure 3 As shown. By Figure 3 It can be seen that its radiochemical purity is greater than 95%.

[0175] 1.3. Using FAP-2286 as a comparative example, 0.1 mL 177 Add an appropriate amount of sodium acetate buffer (pH=4.45) to LuCl3 solution (1 mci / 0.1 mL), then add 10 nmol / 10 μL of FAP-2286, heat to 95 ℃ for 15 min to carry out the reaction, and obtain... 177 Lu-FAP-2286 was used as a control sample.

[0176] 2. Establish an animal model

[0177] Purchase 4-6 week old male nude mice and inject them under the armpits. After inoculation, U87MG cells were cultured for 2-4 weeks, during which time tumor growth was observed. Tumors were monitored until they reached 100-300 mm in size. 3 Imaging or biological distribution studies are conducted at that time.

[0178] 3. Imaging Experiment

[0179] 1.85 MBq 68 Ga-3(FAPI-04)-DOTA (specific activity 37 MBq / mL) was injected into U87MG tumor-bearing nude mice. Small animal PET / CT imaging was performed at 0.5 hours, 1 hour, 2 hours, and 4 hours while the mice were in a prone position. Mice were anesthetized with isoflurane during imaging, and the experiment was repeated at least three times. The PET / CT images are shown below. Figure 4 As shown. From Figure 4 It can be known 68 Ga-3(FAPI-04)-DOTA exhibits high tumor uptake, and no decrease in tumor uptake was observed after 4 hours.

[0180] 6.4 MBq 177 Lu-3(FAPI-04)-DOTA (specific activity 64 MBq / mL) was injected into U87MG tumor-bearing nude mice, and SPECT / CT imaging was performed at 1 hour, 4 hours, 8 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 120 hours, and 144 hours. Mice were anesthetized with isoflurane during imaging, and the experiment was repeated at least three times. The SPECT images are shown below. Figure 5 As shown. From Figure 5 It can be known 177 144 hours after Lu-3(FAPI-04)-DOTA was injected into mice, the tumors still took up the drug.

[0181] 6.4 MBq 177 Lu-FAP-2286 was injected into U87MG tumor-bearing nude mice, and SPECT / CT imaging was performed at 1 hour, 4 hours, 8 hours, and 24 hours. Mice were anesthetized with isoflurane during imaging, and the experiment was repeated at least three times. The SPECT images are shown below. Figure 6 As shown. By Figure 6 It can be seen that, 177 Lu-FAP-2286 showed reduced tumor uptake after 24 hours.

[0182] from Figure 5 and Figure 6 It can be seen that, 177 Lu-3(FAPI-04)-DOTA had a significantly longer retention time in U87MG tumors than 177 Lu-FAP-2286.

[0183] Although this application has been described in detail in this specification with general descriptions and specific embodiments, some modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, such modifications or improvements made without departing from the spirit of this application are all within the scope of protection claimed in this application.

Claims

1. A polymeric compound targeting FAP, characterized in that, Its chemical structure is shown in any one of formulas (1) to (3): (1) (2) (3) Where n and m are selected from any integers from 0 to 12; Structural unit B is the targeting portion of fibroblast activation protein α (FAP-α); Structural units D and Y are linking groups, one of which may be absent or both may be absent, and are selected from any of the following structures: ； Where n1, n2, and n3 are any integers from 1 to 12, X1 and Y1 are selected from C, N, O, and S, and R1 and R2 are each independently selected from hydrogen or any of the following structures: ； Where X is a halogen; Structural unit E is selected from any of the following structures: ； Structural unit E2 is selected from any of the following structures: ； Structural unit E3 is selected from any of the following structures: ； The structural unit Z is selected from at least one of radioactive components, chelating agents, fluorescent dyes, contrast agents, or cytotoxic substances.

2. The compound according to claim 1, wherein the general structural formula of B is as follows: ； in, R a R b Each is independently selected from hydrogen, hydroxyl, and C. 1-6 Hydrocarbon group, mercapto group or halogen; R c Selected from H, -CN, -B(OH)2, chloroacetyl, -C(O) hydrocarbon, -C(O) aryl, -C=CC(O) aryl, -C=CS(O)2 aryl, -CO2H, -NH2, urea, -SO3H, -PO3H2, ketoamide group, trifluoromethyl, trifluoroacetyl and tetrazolyl; R d Selected from H, C 1-8 Straight-chain or branched hydrocarbon groups or hydroxyl groups; R e Selected from aromatic monocyclic or bicyclic compounds priced between 5 and 10 yuan, non-aromatic monocyclic or bicyclic compounds priced between 5 and 10 yuan, aromatic monocyclic heterocyclic or bicyclic heterocyclic compounds priced between 5 and 10 yuan, and non-aromatic monocyclic heterocyclic or bicyclic heterocyclic compounds priced between 5 and 10 yuan with R e Excluding the pyridine ring; The heterocycle contains 1 to 3 heteroatoms selected from O, N, or S, R e Hydrogen can be converted into halogens, -NHC 1-8 Alkyl, -OH substitution; when R e For a time At that time, it does not contain amino substituents; R f The general structural formula is , where R f1 R f2 R f3 R f4 Each exists or does not exist; R f1 For O, S, NR g R g It can be hydrogen, halogen, hydroxyl, mercapto or C 1-8 alkyl; R f2 C 1-8 alkyl or n4 = any integer from 0 to 8; R f3 It is an aromatic or non-aromatic monocyclic nitrogen-containing heterocycle; selected from any of the following structural species: ； R f4 It does not exist or is selected from any of the following structures: ； Where n1, n2, and n3 are any integers from 1 to 12.

3. The polymeric compound targeting FAP according to claim 2, characterized in that, R e Selected from any of the following structures, where the ring and any site of the substituents on the ring can be extended to connect R. f : 。 4. The polymeric compound targeting FAP according to claim 2, characterized in that, Structural unit B includes any one of the following chemical structures: ； ； 。 5. The polymeric compound targeting FAP according to claim 1, characterized in that: The chelating agent includes any one of DOTA, DOTAGA, NOTA, NOTAGA, TETA, DAPA, TRAP, HBED-CC, DFO, PCTA, DTPA, HYNIC, ATSM, EDTA, CB-TE2A, NOPO, NOTAM, MAG3, Deferoxamine, Cyclen, H3RESCA, Octaaminocryptands, diaza-18-crown ether-6, NTA, CB-cyclam, or isocyanate; The radioactive component is selected from isotopes that emit alpha rays, beta rays, gamma rays, Auger electrons, X-rays, or fluorescence; The fluorescent dyes include at least one of xanthine, acridine, azine, cyanine, styryl dyes, coumarin, porphyrin, metal ligand complexes, fluorescent proteins, nanocrystals, perylene, borodipyrrole methylene or phthalocyanine; The cytotoxic substance includes at least one of MMAE, MMAF, DM1, PNU-159682, Cryptophycin-55, PBD, or Epothilone B.

6. A pharmaceutical composition, characterized in that, Including the polymeric compound targeting FAP as described in any one of claims 1-5.

7. An FAP inhibitor, characterized in that, Includes the FAP-targeting polymeric compound according to any one of claims 1-5, a salt or isomer of the FAP-targeting polymeric compound.

8. Use of the pharmaceutical composition of claim 6 or the FAP inhibitor of claim 7 in the preparation of a medicament for treating or detecting a disease characterized by overexpression of the fibroblast activating protein FAP.

9. The application according to claim 8, characterized in that, The diseases characterized by overexpression of the fibroblast activator protein FAP include any one of cancer, chronic inflammation, atherosclerosis, fibrosis, tissue remodeling, or scarring.

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