A tumor stroma-targeted radiopharmaceutical and preparation method and application thereof

Abstract

The present application relates to a tumor stroma-targeted radiopharmaceutical and its preparation method and application. Specifically, the present application provides a novel tumor stroma-targeted probe compound, the structure of which is shown in formula I, and the substituents and groups thereof are described in the specification and claims. The probe described in the present application has the characteristics of anti-enzymolysis and high water solubility with glutathione as a connecting agent. The present application also provides a preparation method of the probe compound and its use in treating and / or diagnosing diseases related to tumor stroma fibroblast activation protein and neovascular integrin alpha v beta 3 receptor.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a FAP-targeted radiopharmaceutical, its preparation method, and its application. Background Technology

[0002] Malignant tumors are among the most deadly diseases threatening human health. Furthermore, the incidence of cancer continues to rise, with new cases projected to reach 28.4 million by 2040. The core of cancer treatment is early diagnosis, early detection, and early treatment. Due to its high sensitivity, nuclear medicine imaging, especially PET / CT, has irreplaceable advantages in tumor diagnosis and the detection of metastases compared to other diagnostic methods.

[0003] Tumors consist of tumor cells and tumor stroma, with the stromal cells primarily comprising vascular cells, fibroblasts, and immune cells. The tumor stroma is a crucial component of tumors, sometimes accounting for over 90% of the solid mass of a malignant tumor. Tumor-associated fibroblasts (CAFs) are a major component of the tumor stroma, secreting numerous growth factors, pro-inflammatory cytokines, and chemokines. They can reduce T-cell responses and recruit immunosuppressive cells to aid in tumor immune escape. Therefore, CAFs play a critical role in tumor development, metastasis, treatment resistance, and immune evasion, making them important target cells for tumor diagnosis and treatment research. Tumor fibroblast activating protein (FAP) is highly expressed in over 90% of epithelial cancers, such as head and neck cancer, breast cancer, lung cancer, pancreatic cancer, esophageal cancer, colorectal cancer, ovarian cancer, gastric cancer, and liver cancer, while it is not expressed or is expressed at very low levels in normal tissues. Furthermore, high FAP expression in CAFs has been proven to be a marker of tumor invasiveness and poor prognosis. Therefore, FAP is an important target for CAFs and also provides a reliable target for nuclear medicine tumor diagnosis and treatment.

[0004] Developing small-molecule radionuclide imaging and therapeutic drugs targeting FAP (fiber-dependent phosphatidylcholine) has become a hot topic in the field of nuclear medicine radiopharmaceuticals in recent years, representing the next frontier in radiopharmaceutical research and development. Currently, a series of radiolabeled FAP-specific inhibitors (FAPIs) have been developed for tumor diagnosis and treatment. 68 Ga-FAPI PET / CT imaging exhibits good tumor specificity in nearly 30 different types of tumors. Compared with [18F]FDG imaging, FAPI imaging has a lower background in the brain, liver, and oropharyngeal mucosa, and a higher detection rate for tumor lesions.

[0005] Neovascularization in the tumor stroma is fundamental to tumor development, progression, and metastasis. The αvβ3 receptor, a type of integrin receptor, is highly expressed in tumor neovascular endothelial cells but not in normal cells, making it a stable and reliable target for tumor diagnosis and treatment. Small peptides with the arginine-glycine-aspartic acid (RGD) amino acid sequence can specifically target the αvβ3 receptor. Based on this, several drugs for tumor diagnosis and treatment have been developed. Among them, technetium-99m (99mTc)-labeled RGD cyclic peptides can be used for tumor single-photon emission computed tomography (SPECT) diagnosis. To enhance tumor targeting, two RGD cyclic peptides are coupled into a dimer and then labeled with technetium-99m. The resulting [99mTc]Tc-3PRGD2 has entered Phase III clinical trials and can be used for broad tumor diagnosis. Therefore, targeting the αvβ3 receptor in tumor neovascularization can achieve tumor receptor-targeted, universally applicable diagnosis and treatment.

[0006] Compared to targeting tumor cells, targeting the tumor stroma offers unique advantages for tumor diagnosis and treatment. Diagnostic or therapeutic drugs must first pass through the bloodstream and then diffuse into the tumor cells via the tumor stroma. Due to the tumor microenvironment, vascular malformations or abnormalities, and cancer cell proliferation, intratumoral pressure is high. Therefore, drug delivery to tumor cells must overcome multiple physiological barriers. In contrast, stroma-targeted drugs do not need to penetrate the tumor stroma, resulting in higher targeting efficiency and thus higher diagnostic and therapeutic efficacy.

[0007] Nuclear medicine demands drastically different pharmacokinetic behaviors from tumor diagnostic and therapeutic drugs. Diagnostic drugs require rapid in-and-out action, meaning they must quickly target the tumor and be rapidly eliminated from the body to reduce blood background and facilitate diagnosis. Therapeutic drugs, on the other hand, require a relatively long blood half-life to promote high tumor uptake. Currently, most reported nuclear medicine tumor diagnostic and therapeutic drugs are used either for imaging in tumor diagnosis or for tumor treatment, making it difficult to achieve both simultaneously. Furthermore, for detecting small metastatic lesions, an appropriately prolonged blood half-life is more conducive to probe uptake, providing more reliable and clear imaging data. In terms of treatment, rapid metabolism and elution lead to lower effective doses and shorter retention times at the tumor site, necessitating higher doses or more frequent dosing to meet treatment needs and increasing the likelihood of adverse reactions.

[0008] Therefore, there is an urgent need in this field to develop a novel strategy for preparing tumor-targeting probes whose blood half-life can be modularly customized and controlled according to the half-life of the radionuclide and the requirements of diagnosis and treatment. At the same time, it not only has the characteristics of high uptake by tumors, but also can remain in the tumor for a long time, so as to overcome the shortcomings of existing technologies. Summary of the Invention

[0009] The main objective of this invention is to provide a novel tumor matrix-targeting nuclear medicine probe, its preparation method, and its applications. The probe uses glutathione (GSH) as a linker and can optionally be combined with tumor matrix (fibroblasts and neovascular endothelial cells) targeting molecules (FAPI, RGD small peptides), radionuclide-labeled chelating agents, albumin-binding small molecules, and covalently coupled small molecules. This allows the blood half-life of the compound to be customizable according to the radionuclide lifetime, enabling irreversible binding to the tumor and thus achieving efficient tumor diagnosis or treatment.

[0010] In a first aspect, the present invention provides a compound of formula I, or a pharmaceutically acceptable salt thereof.

[0011]

[0012] in,

[0013] R1 is a nuclide labeling group selected from the following group:

[0014]

[0015] R4 is a FAP inhibitor targeting tumor stromal fibroblasts or a RGD cyclic peptide of integrin αvβ3 receptor for neovascular endothelial cells selected from the following groups:

[0016]

[0017]

[0018] Indicates the connection point with other parts of the compound;

[0019] The nuclide is selected from the following group: 177 Lu、 90 Y、 18 F, 64 Cu、 68 Ga、 89 Zr、 225 Ac、 111 In、 99m Tc, 188 Re、 161 Tb, 212 Pb, 125 I, 131 I or 124 I.

[0020] In another preferred embodiment, R1 is

[0021] A second aspect of the present invention provides a compound of formula II, or a pharmaceutically acceptable salt thereof, having the following structure:

[0022]

[0023] Wherein, R1 and R4 are as defined in the first aspect of the present invention;

[0024] R2 is a small albumin-binding molecule selected from the following group:

[0025]

[0026] R3 is a covalently bonded group selected from the following group:

[0027]

[0028] In another preferred embodiment, R1 is In another preferred embodiment, R2 is In another preferred embodiment, R3 is In another preferred embodiment, the compound is selected from the group consisting of:

[0029]

[0030]

[0031] A third aspect of the invention provides a matrix-targeting probe comprising a radiolabeled compound as described in the first aspect of the invention, wherein the radiolabeled compound is selected from the group consisting of: 177 Lu、 90 Y、 18 F, 64 Cu、 68 Ga、 89 Zr、 225 Ac、 111 In、 99m Tc, 188 Re、 161 Tb, 212 Pb, 125 I, 131 I, 124 I.

[0032] A fourth aspect of the present invention provides a method for preparing a probe as described in the third aspect of the present invention, comprising the following steps:

[0033] (a) Providing a compound or pharmaceutically acceptable salt as described in the first or second aspect of the present invention, and a radionuclide;

[0034] (b) Prepare probes by wet labeling or lyophilization labeling methods;

[0035] The radionuclides are as defined above.

[0036] A fifth aspect of the present invention provides a composition comprising:

[0037] (i) a compound as described in the first or second aspect of the present invention, or a pharmaceutically acceptable salt thereof; and

[0038] (ii) Pharmaceutically acceptable carriers.

[0039] A sixth aspect of the invention provides the use of a compound as described in the first or second aspect of the invention or a composition as described in the fifth aspect of the invention for the preparation of a medicament for the treatment and / or diagnosis of diseases associated with fibroblast activation proteins.

[0040] In another preferred embodiment, the disease is selected from the group consisting of: head and neck cancer, breast cancer, lung cancer, pancreatic cancer, esophageal cancer, colorectal cancer, ovarian cancer, gastric cancer, liver cancer, or combinations thereof.

[0041] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here. Attached Figure Description

[0042] Figure 1 The high-resolution mass spectrum of compound GSH-FAPI prepared in Example 1 of this invention is shown.

[0043] Figure 2 The HPLC spectrum of compound GSH-FAPI prepared in Example 1 of this invention is shown.

[0044] Figure 3 The in vitro binding affinity test of the compound GSH-FAPI of the present invention with FAP protein is shown.

[0045] Figure 4 Radiolabeled ligands were shown 68 PET imaging of Ga-GSH-FAPI in tumor models.

[0046] Figure 5 Radiolabeled ligands were shown 161 Biodistribution and plasma half-life of Tb-GSH-FAPI.

[0047] Figure 6 Radiolabeled ligands were shown 161 Comparison of in vivo antitumor efficacy of Tb-GSH-FAPI and changes in mouse body weight.

[0048] Figure 7 Radiolabeled ligands were shown 68Ga-GSH-FAPI in PET imaging of gastric cancer patients. Detailed Implementation

[0049] Through extensive and in-depth research, the inventors unexpectedly developed a novel class of tumor-targeting nuclear medicine probe compounds. These compounds utilize glutathione as a linker to connect four components: a modified targeting molecule (R4), a radionuclide-labeled chelating agent (R1), an albumin-binding small molecule (R2), and a covalently coupled small molecule (R3). R2 and R3 are optional groups that can be adjusted according to the radionuclide lifetime. Based on this, the present invention was completed.

[0050] This invention provides a multifunctional complex constructed with GSH as the core linker, tumor-targeting ligands, radionuclide chelating groups, albumin-binding groups, and covalently bound groups. These complexes include, but are not limited to, multi-module combinations of GSH-FAPI (or RGD cyclic peptide)-radionuclide chelating agents and GSH-FAPI (or RGD cyclic peptide)-albumin-binding groups-covalently bound groups-radionuclide chelating agents, as well as their synthesis methods and specific applications in radiopharmaceutical diagnosis and treatment.

[0051] the term

[0052] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0053] As used herein, the terms “comprising,” “including,” and “containing” are used interchangeably and include not only closed definitions but also semi-closed and open definitions. In other words, the terms include “consisting of” and “substantially consisting of”.

[0054] As used herein, the abbreviations are explained as follows:

[0055] DMF: N,N-dimethylformamide

[0056] HOBT: 1-Hydroxybenzotriazole

[0057] EDCI: 1-Ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride

[0058] TFA: Trifluoroacetic acid

[0059] CAN: Acetonitrile

[0060] HATU: N,N,N',N'-Tetramethyl-O-(7-azabenzotriazol-1-yl)hexafluorophosphate urea

[0061] DIPEA: N,N-Diisopropylethylamine

[0062] NMM: N-methylmorpholine

[0063] DCC: 1,3-Dicyclohexylcarbodiimide

[0064] HOSU: N-hydroxysuccinimide

[0065] DCM: Dichloromethane

[0066] HBTU: Benzotriazole-N,N,N',N'-Tetramethylurea hexafluorophosphate

[0067] Tumor stroma

[0068] Tumor fibroblasts and neovascularized tumor stroma are the main components of tumor matrix. Tumor fibroblast activation protein (FAP) and αvβ3 receptor are highly expressed in tumor fibroblasts and tumor neovascularized endothelial cells, respectively, and are stable and reliable targets for tumor matrix-targeted diagnosis and treatment.

[0069] Fibroblast activating protein (FAP): Also known as seprase protein or melanoma membrane-bound gelatinase, FAP is a 170 kDa protein encoded by the FAP gene (2q23) in the human body. It is a homodimer belonging to the membrane gelatinase family of serine proteases. It is selectively expressed in the reaction matrix of epithelial carcinoma fibroblasts, granulation tissue of wound healing, and malignant cells of bone and soft tissue sarcomas. This protein is believed to be involved in fibroblast growth during development, the control of epithelial-mesenchymal interactions, tissue repair, and epithelial carcinogenesis. FAP belongs to the SC protease family, specifically the S9B proline oligopeptidase subfamily. Other members of the S9B subfamily include DPPIV, DPP8, and DPP9. FAP shows a high correlation with DPPIV, sharing approximately 50% of their amino acids.

[0070] Integrin αvβ3 receptor: Angiogenesis is fundamental to tumor development, progression, and metastasis. The αvβ3 receptor is a type of integrin receptor that is highly expressed in tumor neovascular endothelial cells but not in normal cells. It is a stable and reliable target for tumor diagnosis and treatment, and small peptides with the arginine-glycine-aspartic acid (RGD) amino acid sequence can specifically target the αvβ3 receptor.

[0071] This invention relates to a tumor matrix-targeting probe and its preparation method.

[0072] This invention provides a tumor matrix targeting probe of Formula I, which is based on a quinolinic acid-derived tumor matrix targeting compound.

[0073]

[0074] in,

[0075] R1 is a nuclide labeling group selected from the following group:

[0076]

[0077] R4 is a FAP-targeting inhibitor selected from the following group:

[0078]

[0079] Alternatively, the integrin αvβ3 receptor of newly formed vascular endothelial cells targets the RGD cyclic peptide:

[0080]

[0081] Indicates the connection point with other parts of the compound;

[0082] The nuclide is selected from the following group: 77 Lu、 90 Y、 18 F, 64 Cu、 68 Ga、 89 Zr、 225 Ac、 111 In、 99m Tc, 188 Re、 161 Tb, 212 Pb, 125 I, 131 I, 124 I.

[0083] The present invention also provides a compound represented by Formula II.

[0084]

[0085] R1 and R4 are defined as above;

[0086] R2 is an albumin-binding small molecule selected from the group consisting of amide bonds grafted onto glutathione.

[0087]

[0088] R3 is a covalently bonded group selected from the following group:

[0089]

[0090] In another preferred embodiment, R1 is

[0091] In another preferred embodiment, R2 is

[0092] In another preferred embodiment, R3 is

[0093] In another preferred embodiment, R4 is:

[0094] or

[0095]

[0096] In another preferred embodiment, in the compound, any one of R1, R2, R3 and R4 is independently a group corresponding to the specific compound of the present invention.

[0097] As used herein, the term "pharmaceutically acceptable salt" refers to a salt formed by a positively charged group on a compound of formula (I) and an anion, or a salt formed by a negatively charged group on a compound of formula (I) and a cation. Suitable anions include, but are not limited to, chloride, bromide, iodide, sulfate, nitrate, phosphate, citrate, methanesulfonate, trifluoroacetate, acetate, malate, toluenesulfonate, tartrate, fumarate, glutamate, glucuronide, lactate, glutarate, or maleate. Suitable cations include, but are not limited to, sodium, potassium, magnesium, calcium, and ammonium ions.

[0098] In another preferred embodiment, the pharmaceutically acceptable salt of the present invention refers to a salt formed by a compound of general formula (I) with an acid from the group below, such as, but not limited to: hydrofluoric acid, hydrochloric acid, hydrobromic acid, phosphoric acid, acetic acid, oxalic acid, sulfuric acid, nitric acid, methanesulfonic acid, aminosulfonic acid, salicylic acid, trifluoromethanesulfonic acid, naphthalenesulfonic acid, maleic acid, citric acid, acetic acid, lactic acid, tartaric acid, succinic acid, oxalic acid, pyruvic acid, malic acid, glutamic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, ethanesulfonic acid, naphthalenedisulfonic acid, malonic acid, and fumaric acid. Propionic acid, oxalic acid, trifluoroacetic acid, stearic acid, acetic acid, hydroxymaleic acid, phenylacetic acid, benzoic acid, glutamic acid, ascorbic acid, p-aminobenzenesulfonic acid, 2-acetoxybenzoic acid, and hydroxyethanesulfonic acid, etc.; or salts formed by compounds of general formula (I) with inorganic bases, such as, but not limited to, sodium salts, potassium salts, calcium salts, aluminum salts, or ammonium salts; or salts formed by compounds of general formula (I) with organic bases, such as, but not limited to, methylamine salts, ethylamine salts, ethanolamine salts, hydroxymethylaminomethane (TRIS) ammonium salts, etc.

[0099] The embodiments of this invention specifically describe the preparation method of the compound with structure (I) of this invention, but these specific methods do not constitute any limitation on this invention. The compounds of this invention can also be conveniently prepared by optionally combining various synthetic methods described in this specification or known in the art, such combinations can be easily performed by those skilled in the art.

[0100] Typically, the raw materials and reagents used in the preparation process of the compounds of the present invention can be purchased commercially unless otherwise specified.

[0101] Preparation of radionuclide-labeled compounds: The FAPI compound is dissolved in a buffer solution or deionized water, and then a radionuclide (such as...) is added. 68 A solution of Ga was prepared, the pH was adjusted, and the mixture was reacted at 95°C for 15 minutes. After cooling to room temperature, an HLB separation column was taken and activated by rinsing with 10 mL of anhydrous ethanol and then 10 mL of water. The reaction solution was diluted to 5 mL with water and then loaded onto the separation column. The separation column was rinsed with water to remove unreacted ions, and then rinsed with ethanol solution to obtain the labeled complex.

[0102] Pharmaceutical Compositions and Administration

[0103] The pharmaceutical compositions of the present invention comprise, within a safe and effective range, the compound of the present invention or a pharmacologically acceptable salt thereof, and a pharmacologically acceptable excipient or carrier. "Safe and effective range" refers to an amount of the compound sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical composition contains 1-2000 mg of the compound of the present invention per dose, more preferably, 5-1000 mg of the compound of the present invention per dose. Preferably, "one dose" is one capsule or tablet.

[0104] "Pharmaceutically acceptable carriers" refers to one or more compatible solid or liquid fillers or gelling substances that are suitable for human use and must have sufficient purity and sufficiently low toxicity. "Compatibility" here means that the components in the composition can be mixed with and with the compounds of the present invention without significantly reducing the efficacy of the compounds. Examples of pharmaceutically acceptable carriers include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), and emulsifiers (such as Tween). Wetting agents (such as sodium dodecyl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0105] The pharmaceutical composition is an injection, capsule, tablet, pill, powder, or granule.

[0106] There are no particular limitations on the administration of the compounds or pharmaceutical compositions of the present invention. Representative administration methods include (but are not limited to): oral, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous), and local administration.

[0107] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with: (a) fillers or compatibilizers, such as starch, lactose, sucrose, glucose, mannitol, and silica; (b) binders, such as hydroxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, and gum arabic; (c) humectants, such as glycerin; (d) disintegrants, such as agar, calcium carbonate, potato starch or cassava starch, alginate, certain complex silicates, and sodium carbonate; (e) slowing agents, such as paraffin; (f) absorption accelerators, such as quaternary ammonium compounds; (g) wetting agents, such as cetyl alcohol and glyceryl monostearate; (h) adsorbents, such as kaolin; and (i) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium dodecyl sulfate, or mixtures thereof. Buffers may also be included in the dosage forms of capsules, tablets, and pills.

[0108] Solid dosage forms such as tablets, sugar pills, capsules, pellets, and granules can be prepared using coatings and shells, such as casings and other materials known in the art. They may contain opacifying agents, and the release of the active compound or compound from such compositions can be delayed in a portion of the digestive tract. Examples of encapsulating components that can be used are polymeric substances and waxes. If necessary, the active compound may also be formed into microcapsules with one or more of the excipients described above.

[0109] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, or tinctures. In addition to the active compound, liquid dosage forms may contain inert diluents conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, e.g., ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butanediol, dimethylformamide, and oils, particularly cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, or mixtures of these substances.

[0110] In addition to these inert diluents, the composition may also contain auxiliaries such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents and fragrances.

[0111] In addition to the active compound, the suspension may contain suspending agents such as ethoxylated isooctadecyl alcohol, polyoxyethylene sorbitol and dehydrated sorbitol esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances.

[0112] Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions, or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents, or excipients include water, ethanol, polyols, and suitable mixtures thereof.

[0113] Dosage forms of the compounds of the present invention for topical administration include ointments, powders, patches, sprays, and inhalers. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants that may be necessary.

[0114] The compounds of this invention can be administered alone or in combination with other pharmaceutically acceptable compounds (such as oncology drugs).

[0115] The treatment method of the present invention can be used alone or in combination with other treatment methods or drugs.

[0116] When using the pharmaceutical composition, a safe and effective amount of the compound of the present invention is applied to the mammal (such as a human) requiring treatment. The dosage administered is the pharmaceutically considered effective dose. For a person weighing 60 kg, the daily dose is typically 1–2000 mg, preferably 5–1000 mg. Of course, the specific dosage should also take into account factors such as the route of administration and the patient's health condition, which are all within the scope of the skill of a skilled physician.

[0117] Compared with the prior art, the main advantages of the present invention are:

[0118] 1. Advantages of Glutathione (GSH) as a Linker: By adjusting the type of linker, the affinity of the tracer and its biodistribution characteristics in vivo can be modified. Pharmacokinetics, initial tumor uptake, and tumor retention can all be improved by modifying the linker without altering the target ligand. Endogenous GSH as a linker can reduce immune recognition, enhance tumor-targeted uptake, and prolong blood circulation. The γ-glutamyl bond of GSH can resist degradation by common proteases, improving its stability in vivo. As a zwitterionic peptide, GSH possesses both positive and negative charges, enhancing molecular hydrophilicity. It contains free -NH3. + / -COO- can form hydrogen bonds or electrostatic interactions with Site II of albumin, prolonging blood circulation time and improving its pharmacokinetic behavior by enhancing the molecule's hydrophilicity and "stealth" properties.

[0119] 2. Blood half-life can be adjusted according to different needs: According to the specific requirements of diagnosis and treatment, this invention can couple or not couple albumin-bound small molecules to give them appropriate in vivo metabolic kinetics, higher tumor uptake dose and longer tumor retention time, so as to meet the needs of radionuclide therapy or diagnosis.

[0120] 3. Irreversible tumor uptake: The molecular probe of this invention can covalently bind to the tumor site, achieving irreversible tumor targeting.

[0121] 4. Enhanced tumor targeting and retention: The molecular probe of this invention can simultaneously achieve receptor-ligand and covalent binding. The probe taken up by the tumor through receptor-ligand action is fixed inside the tumor through covalent binding and does not efflux. The absolute value of uptake is higher, which is not available in other tumor matrix targeting probes. This achieves high uptake and long-term retention.

[0122] 5. Excellent in vivo pharmacokinetic behavior: It is rapidly cleared and metabolized by normal organs, and is not taken up by the liver and spleen. It can be excreted through the kidneys, resulting in a higher tumor / normal organ uptake ratio of the probe, which is more conducive to nuclear medicine imaging of tumors.

[0123] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions as described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or as recommended by the manufacturer. Unless otherwise stated, percentages and parts are weight percentages and parts by weight.

[0124] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as are familiar to those skilled in the art. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to the methods of this invention. The preferred embodiments and materials described herein are for illustrative purposes only.

[0125] Unless otherwise specified in the examples, the reaction temperature is room temperature (20℃~35℃).

[0126] Compound preparation examples

[0127] Example 1: Synthesis of GSH-FAPI

[0128]

[0129] (1) Dissolve Compd 2 (1 eq) in 10 mL DMF, cool to 0 °C, add HOBT (1.2 eq) and EDCI (1.2 eq), react at 0 °C for 10 min, add Compd 1 (1.2 eq) and NMM (3.3 eq), react at room temperature for 2 h. LC-MS monitor the reaction until complete, evaporate the solvent, add 10 mL acetonitrile aqueous solution, place at 0 °C, add lithium hydroxide, adjust pH to alkaline (pH paper test), react at room temperature for 30 min. LC-MS monitor the hydrolysis reaction until complete, adjust pH to neutral, evaporate the solvent, add 10 mL TFA solution, react at room temperature for 2 h, add 100 mL diethyl ether, a large amount of solid precipitates, centrifuge and dry, and purify by reverse-phase preparative HPLC to obtain Compd 3, yield 37%.

[0130] (2) Compd 4 (1eq) and Compd 3 (1eq) were dissolved in 5 mL ACN and 5 mL H2O, and then 2.5 mL of buffer solution with pH = 7.2 was added. The reaction was carried out at room temperature for 1 h. The reaction was monitored by LC-MS to ensure complete reaction. After purification by reverse-phase preparative liquid chromatography, Compd 5 was obtained with a yield of 42.2%.

[0131] (3) Compd 7 (1 eq) was dissolved in 6 mL DMF, and then HATU (1.5 eq) and DIPEA (3 eq) were added. The mixture was reacted at 0 °C for 5 minutes. Then Compd 6 (1 eq) was added and the mixture was reacted at room temperature overnight. The reaction was monitored by LC-MS until it was complete. The mixture was then evaporated to dryness, and 5 mL TFA was added. The mixture was reacted at room temperature for 2 hours. Then 100 mL diethyl ether was added. A large amount of solid precipitated out. The solid was centrifuged and dried. The solid was purified by reverse-phase preparative liquid chromatography to obtain Compd 8 with a yield of 76.2%.

[0132] (4) Compd 9 (1 eq) was dissolved in 4 mL DMF, cooled to 0 °C, and HOBT (1.5 eq) and EDCI (1.5 eq) were added. The mixture was reacted at 0 °C for 10 minutes, then Compd 8 (1 eq) and NMM (3 eq) were added, and the mixture was reacted at room temperature for 2 hours. The reaction was monitored by LC-MS until complete. The mixture was evaporated to dryness, and 4.75 mL TFA and 0.25 mL H2O were added. The mixture was reacted at room temperature for 2 hours, and then 100 mL diethyl ether was added. A large amount of solid precipitated out. After centrifugation and drying, the crude product was obtained. Compd 10 was obtained by reverse-phase preparative HPLC purification with a yield of 45.9%.

[0133] (5) Compd 10 (1 eq) was dissolved in 10 ml DMF, then DCC (1.2 eq) and HOSu (1.2 eq) were added, and the mixture was reacted at room temperature for 6 hours. Then Compd 5 (1.2 eq) and DIPEA (3 eq) were added, and the mixture was reacted at room temperature for 1 hour. The reaction was monitored by LC-MS until it was complete. The mixture was then evaporated to dryness, and 10 ml of 20% piperidine solution was added. The mixture was reacted at room temperature for 10 minutes, and then evaporated to dryness. After purification by reverse-phase preparative liquid chromatography, GSH-FAPI was obtained with a yield of 30.5%.

[0134] The synthetic route is shown above. Mass spectrometry analysis revealed the target compound based on its molecular weight; HPLC purity is >95%. High-resolution mass spectrometry and HPLC spectra are shown below. Figure 1 and Figure 2 As shown.

[0135] Example 2: Synthesis of Alb-GSH-FAPI

[0136]

[0137] (1) The first step is the same as the synthesis route of GSH-FAPI. Compd 2 (1 eq) was dissolved in 10 ml DMF, cooled to 0 °C, HOBT (1.2 eq) and EDCI (1.2 eq) were added, and the reaction was carried out at 0 °C for 10 min. Then, Compd 1 (1.2 eq) and NMM (3.3 eq) were added, and the reaction was carried out at room temperature for 2 h. Compd 3 was obtained by reverse-phase preparative liquid chromatography.

[0138] (2) Compd 4 (1eq) and Compd 3 (1eq) were dissolved in DMF, and HATU and DIPEA were added. The reaction was carried out at room temperature for 12 h. The reaction was monitored by LC-MS until it was complete. 30% TFA and DCM were added, and the reaction was carried out at room temperature for 30 min. Compd 5 was obtained after reverse-phase preparative liquid chromatography purification.

[0139] (3) Dissolve Compd 5 (1 eq) in 6 mL of DMF, then add HBTU (1.5 eq) and DIPEA (3 eq), react at 0 °C for 5 minutes, add Compd 6 (1 eq), react at room temperature overnight, monitor the reaction to ensure complete reaction by LC-MS, evaporate to dryness, add 95% TFA, react at room temperature for 2 hours, centrifuge to dryness, and purify by reverse-phase preparative liquid chromatography to obtain Compd 7.

[0140] (4) Compd 9 (1 eq) was dissolved in 6 mL DMF, and then HATU (1.5 eq) and DIPEA (3 eq) were added. The mixture was reacted at 0 °C for 5 minutes. Then Compd 8 (1 eq) was added and the mixture was reacted at room temperature overnight. The reaction was monitored by LC-MS until it was complete. The mixture was then evaporated to dryness, and 5 mL TFA was added. The mixture was reacted at room temperature for 2 hours. Then 100 mL diethyl ether was added. A large amount of solid precipitated out. The solid was centrifuged and dried. The solid was purified by reverse-phase preparative liquid chromatography to obtain Compd 10.

[0141] (5) Compd 11 (1 eq) was dissolved in 4 mL DMF, cooled to 0 °C, and HOBT (1.5 eq) and EDCI (1.5 eq) were added. The mixture was reacted at 0 °C for 10 minutes, then Compd 10 (1 eq) and NMM (3 eq) were added. The mixture was reacted at room temperature for 2 hours. The reaction was monitored by LC-MS until complete. The mixture was evaporated to dryness, and 4.75 mL TFA and 0.25 mL H2O were added. The mixture was reacted at room temperature for 2 hours, and then 100 mL diethyl ether was added. A large amount of solid precipitated out. The solid was centrifuged and dried to obtain the crude product, which was purified by reverse-phase preparative HPLC to obtain Compd 12.

[0142] (6) Compd 12 (1 eq) was dissolved in 10 ml DMF, then DCC (1.2 eq) and HOSu (1.2 eq) were added, and the mixture was reacted at room temperature for 6 hours. Then Compd 7 (1.2 eq) and DIPEA (3 eq) were added, and the mixture was reacted at room temperature for 1 hour. The reaction was monitored by LC-MS until it was complete. The mixture was then evaporated to dryness, and 10 mL of 20% piperidine solution was added. The mixture was reacted at room temperature for 10 minutes, and then evaporated to dryness. After purification by reverse-phase preparative liquid chromatography, Alb-GSH-FAPI was obtained.

[0143] Example 3: Synthesis of Alb-SF-GSH-FAPI

[0144]

[0145] (1) The first step is the same as the synthesis route of GSH-FAPI. Compd 2 (1 eq) was dissolved in 10 ml DMF, cooled to 0 °C, HOBT (1.2 eq) and EDCI (1.2 eq) were added, and the reaction was carried out at 0 °C for 10 min. Then, Compd 1 (1.2 eq) and NMM (3.3 eq) were added, and the reaction was carried out at room temperature for 2 h. Compd 3 was obtained by reverse-phase preparative liquid chromatography.

[0146] (2) Compd 4 (1eq) and Compd 3 (1eq) were dissolved in DMF, and HATU and DIPEA were added. The reaction was carried out at room temperature for 12 h. The reaction was monitored by LC-MS until it was complete. 30% TFA and DCM were added, and the reaction was carried out at room temperature for 30 min. Compd 5 was obtained after reverse-phase preparative liquid chromatography purification.

[0147] (3) Dissolve Compd 5 (1 eq) in 6 mL of DMF, then add HBTU (1.5 eq) and DIPEA (3 eq), react at 0 °C for 5 minutes, add Compd 6 (1 eq), react at room temperature overnight, monitor the reaction to ensure complete reaction by LC-MS, evaporate to dryness, add 95% TFA, react at room temperature for 2 hours, centrifuge to dryness, and purify by reverse-phase preparative liquid chromatography to obtain Compd 7.

[0148] (4) Compd 9 (1 eq) was dissolved in 6 mL DMF, and then HATU (1.5 eq) and DIPEA (3 eq) were added. The mixture was reacted at 0 °C for 5 minutes. Then Compd 8 (1 eq) was added and the mixture was reacted at room temperature overnight. The reaction was monitored by LC-MS until it was complete. The mixture was then evaporated to dryness, and 5 mL TFA was added. The mixture was reacted at room temperature for 2 hours. Then 100 mL diethyl ether was added. A large amount of solid precipitated out. The solid was centrifuged and dried. The solid was purified by reverse-phase preparative liquid chromatography to obtain Compd 10.

[0149] (5) Compd 11 (1 eq) was dissolved in 4 mL DMF, cooled to 0 °C, and HOBT (1.5 eq) and EDCI (1.5 eq) were added. The mixture was reacted at 0 °C for 10 minutes, then Compd 10 (1 eq) and NMM (3 eq) were added. The mixture was reacted at room temperature for 2 hours. The reaction was monitored by LC-MS until complete. The mixture was evaporated to dryness, and 4.75 mL TFA and 0.25 mL H2O were added. The mixture was reacted at room temperature for 2 hours, and then 100 mL diethyl ether was added. A large amount of solid precipitated out. The solid was centrifuged and dried to obtain the crude product, which was purified by reverse-phase preparative HPLC to obtain Compd 12.

[0150] (6) Compd 12 (1 eq) was dissolved in 10 ml DMF, then DCC (1.2 eq) and HOSu (1.2 eq) were added, and the mixture was reacted at room temperature for 6 hours. Then Compd 7 (1.2 eq) and DIPEA (3 eq) were added, and the mixture was reacted at room temperature for 1 hour. The reaction was monitored by LC-MS until it was complete. The mixture was then evaporated to dryness, and 10 mL of 20% piperidine solution was added. The mixture was reacted at room temperature for 10 minutes, and then evaporated to dryness. After purification by reverse-phase preparative liquid chromatography, Compd 13 was obtained.

[0151] (7) Compd 13 (1 eq) was dissolved in 6 mL of DMF, and then HBTU (1.5 eq) and DIPEA (3 eq) were added. The mixture was reacted at 0 °C for 5 minutes, and then Compd 14 (1 eq) was added. The mixture was reacted overnight at room temperature. The reaction was monitored by LC-MS until complete. The mixture was then evaporated to dryness, and 95% TFA was added. The mixture was reacted at room temperature for 2 hours, centrifuged, and dried. The product was purified by reverse-phase preparative HPLC to obtain Alb-SF-GSH-FAPI. The synthetic route is shown above.

[0152] Example 4: Preparation of radioactive ligands

[0153] 68 The Ga radioactive labeling process is as follows: ... 68 GaCl3 hydrochloric acid solution (0.1M HCl, 370-555 MBq) was added to a centrifuge tube. The pH was adjusted to 4.0 with 1 mol / L sodium acetate solution. 50 nmol GSH-FAPI or FAPI-04 was added, and the mixture was incubated at 95°C for 15 minutes. After cooling to room temperature, an HLB separation column was activated and rinsed successively with 10 mL of anhydrous ethanol and 10 mL of water. The reaction solution was diluted to 5 mL with water and purified using an HLB solid-phase extraction column (Waters Corporation, Milford, Massachusetts, USA). The column was rinsed with water to remove unbound compounds. 68 Ga 3+ Ions were obtained by rinsing with a 50% ethanol solution. 68 Ga-labeled radioligands. The labeled compound is obtained by diluting with physiological saline or water for injection and then filtering under sterile conditions.

[0154] 161 The Tb radiolabeling process is as follows: Take GSH-FAPI or FAPI-04 (50 nmol) dissolved in 0.1 mL of sodium acetate buffer (0.5 M, pH 5.5), and add... 161 TbCl3 solution (0.04M HCl, 185-370 MBq) was reacted at 95℃ for 30 minutes, followed by reaction with... 68 The Ga radiolabeled ligands were purified using the same purification method. The labeled radioligands required no further purification and could be used directly in subsequent in vitro and in vivo experiments.

[0155] Biological test cases

[0156] Test Example 1: In vitro label stability test

[0157] Experimental methods:

[0158] Determination of radiochemical purity: 1 μL of the product was dropped onto instant thin-layer chromatography paper and subjected to radiochemical thin-layer chromatography analysis to obtain the radiochemical purity.

[0159] 50 μL of the labeled compound was co-incubated with 950 μL of 10% FBS (fetal bovine serum) or PBS buffer at room temperature. At 30 min, 60 min, 90 min, and 120 min after mixing, 1 μL of the mixture was added to instant thin-layer chromatography paper for radiometric analysis, detected by a gamma detector. The labeling rate was determined according to the above method, and the change in radionuclide labeling stability was obtained. Table 1 shows... 68 Ga-GSH-FAPI and 161Labeling stability of Tb-GSH-FAPI in 10% FBS or PBS.

[0160] Table 1. In vitro stability evaluation of radiolabeled ligands

[0161]

[0162] Experimental results:

[0163] As can be seen from Table 1, 68 Ga-GSH-FAPI and 161 The Tb-GSH-FAPI nucleolabeling rate remained around 95% after two hours, and no significant degradation was observed in either 10% FBS or PBS. It exhibited high stability in solution and can be used for further experiments.

[0164] Test Example 2: In vitro FAP competitive binding assay of GSH-FAPI

[0165] To determine the binding affinity of GSH-FAPI to FAP, we performed a competitive binding assay in FAP-positive U87MG cells. U87MG cells were seeded in 6-well plates and cultured until the cell count reached approximately 1 × 10⁻⁶ cells per well. 6 One. Then the cells were combined with 68 Ga-FAPI-04 (370 KBq per well) and different concentrations (10 -12 Up to 10 -5 Cells were co-incubated with unlabeled GSH-FAPI or FAPI-04 (M) for 1 hour. After discarding the culture medium, the cells were washed twice with PBS (0.5 mL) and then lysed with 0.25% trypsin. The lysates were collected in test tubes and radioactivity was counted using a gamma counter. All experiments were performed in triplicate. 68 Ga-FAPI-04 is used as a radioactive tracer for the quantitative detection of FAP binding.

[0166] The results are as follows Figure 3 As shown, GSH-FAPI can significantly inhibit cellular response to... 68 Ga-FAPI-04 uptake. GSH-FAPI and FAPI-04 inhibition. 68 Ga-FAPI-04 absorbs IC 50 The values ​​were 0.54 nM and 1.442 nM, respectively, indicating that GSH-FAPI has a stronger affinity for FAP than FAPI-04 and has good FAP protein-specific binding ability.

[0167] Test Example 3: 68 Ga-GSH-FAPI and 68PET-CT imaging of Ga-FAPI-04 in tumor-bearing mouse models

[0168] Experimental animals and administration methods: HT1080-FAP tumor-bearing mice and 4T1 orthotopic breast cancer mice were administered via tail vein injection.

[0169] Experimental grouping and drug dosage:

[0170] 68 Ga-GSH-FAPI group: 200 μCi injected via tail vein 68 Ga-GSH-FAPI;

[0171] 68 Ga-FAPI-04 group: 200 μCi injected via tail vein 68 Ga-FAPI-04;

[0172] Blank control (Blocking) group: Tail vein injection 68 Thirty minutes before administering Ga-GSH-FAPI (200 μCi), administer unlabeled FAPI-04 (100 μg / animal) via tail vein injection.

[0173] Validation in HT1080-FAP tumor model and 4T1 orthotopic breast cancer model mice 68 The targeting ability of Ga-GSH-FAPI to tumors. Prepared according to Example 4. 68 Ga-labeled complex solution (radiochemical purity greater than 97%), when the tumor volume reaches approximately 200-300 mm. 3 In this study, a small animal PET / CT imaging system was used to perform PET / CT imaging on tumor-bearing mice. After anesthesia, mice were injected with a tracer via the tail vein and then placed on an imaging bed for 120 minutes of dynamic PET scanning. In the blockade experiment, a 500-fold overdose (100 μg / mouse) of FAPI-04 was injected beforehand as a competing ligand. 68 PET imaging was performed 1 hour after Ga-GSH-FAPI.

[0174] exist Figure 4 A and Figure 4 In B, the circled area indicates the location of the tumor. 68 The uptake of Ga-FAPI-04 at the tumor site is rapid, with clearance occurring 0.5 hours after injection. This invention... 68 Ga-labeled probes showed significant uptake and retention at tumor sites within 2 hours, and were rapidly cleared from normal tissues such as muscle, liver, and kidneys. A high radioactive signal was observed in the bladder. The probes are metabolized and excreted in urine. (See the graph below.) 68 Compared to Ga-FAPI-04, at each imaging time point, 68Ga-GSH-FAPI significantly increased absolute uptake in tumors, improved the tumor-to-muscle ratio, and significantly increased tumor retention time.

[0175] Therefore, the labeled compounds of the present invention exhibit superior tumor uptake and tumor background ratio, giving the labelers of the present invention significant advantages in practical applications. On the one hand, this facilitates longer imaging time, and with the guarantee of high imaging contrast, the diagnosis of primary micro-lesions will be more accurate, which is beneficial for target delineation; on the other hand, the high uptake and long retention of the radionuclide-labeled probe at the tumor site will lay the foundation for radionuclide targeted therapy.

[0176] Test Example 4: 161 Tb-GSH-FAPI and 161 Blood clearance and biodistribution of Tb-FAPI-04 in tumor-bearing mouse models

[0177] To further evaluate the pharmacokinetic behavior and in vivo tumor targeting of GSH-FAPI, blood clearance and biodistribution studies were conducted. When the tumor volume in 4T1 tumor-bearing mice reached 200-300 mm... 3 Mice were then randomly assigned to groups. In the blood clearance experiment, each group of mice was injected with 1.85 MBq via the tail vein. 161 Tb-GSH-FAPI or 161 Tb-FAPI-04 (n=5). Micro-blood samples were collected via the orbital venous plexus at 1, 3, 5, 10, 15, 30, 45, 60, 90, 120, and 180 minutes post-injection. Radioactivity counts were determined using a gamma counter after weighing, and data are expressed as percentage of the injected dose per unit tissue (%ID / g). In the in vitro biodistribution study, mice in each group were injected with 1.85 MBq of the tracer (n=3 at each time point). Animals were sacrificed at 1, 4, and 24 hours post-injection, and tissue samples were immediately dissected. Tumors and major organs were collected, and radioactivity was measured using a gamma counter after weighing. Data were standardized to %ID / g.

[0178] 61Tb-GSH-FAPI and 161 The biodistribution and blood clearance results of Tb-FAPI-04 in tumor-bearing mice are as follows: Figure 5 As shown. 161 The distribution half-life of Tb-GSH-FAPI is 1.472 min, and the elimination half-life is 90.72 min. 161The distribution half-life of Tb-FAPI-04 is 0.742 min, and the clearance half-life is 23.8 min. These results indicate that both GSH-FAPI and FAPI-04 are rapidly distributed in vivo. FAPI-04 is rapidly cleared, which can severely affect imaging quality and treatment efficacy. In contrast, the clearance half-life of GSH-FAPI in this invention is significantly prolonged, indicating that GSH, as a linker, can significantly affect the blood half-life of oncoFAP molecules, prolonging blood circulation time and thus enhancing tumor uptake and retention.

[0179] Results of biological distribution experiments as follows Figure 5 As shown, 161 Tb-GSH-FAPI and 161 Tb-FAPI-04 showed significant uptake at tumor sites in model mice after intravenous administration. 161 Tumor uptake at 1, 4, and 24 hours after Tb-GSH-FAPI was 1.57±0.35% ID / g, 1.17±0.13% ID / g, and 0.57±0.09% ID / g, respectively. 161 The tumor uptake in the Tb-FAPI-04 group at 1, 4, and 24 hours was 1.34±0.07 ID / g, 0.75±0.23 ID / g, and 0.38±0.12 ID / g, respectively. Comparison revealed that at each time point... 161 The tumor-specific uptake of Tb-GSH-FAPI was stronger than that of... 161 Tb-FAPI-04 was inhibited, and tumor clearance was significantly slowed. These results indicate that GSH, as a linker-optimized probe, can significantly prolong its blood circulation time, increasing both tumor uptake and retention time.

[0180] Test Example 5: 161 Tb-GSH-FAPI and 161 In vivo pharmacodynamic study of Tb-FAPI-04 against tumors

[0181] Experimental animals and administration method: 4T1 orthotopic breast cancer mice, administered via tail vein injection.

[0182] Experimental grouping and drug dosage:

[0183] 161 Tb-GSH-FAPI group: 0.5 mCi / 1 mCi administered via tail vein. 161 Tb-GSH-FAPI;

[0184] 161 Tb-FAPI-04 group: 0.5 mCi / 1 mCi injected via tail vein. 161 Tb-FAPI-04;

[0185] In order to investigate161 The anti-tumor potential of Tb-GSH-FAPI was demonstrated in a targeted radiotherapy experiment in a 4T1 orthotopic breast cancer mouse model, and... 161 Tb-FAPI-04 and physiological saline were used as controls. 161 The antitumor efficacy of Tb-labeled radioligands was evaluated using the following protocol: when the tumor volume reached approximately 100-120 mm. 3 Mice were randomly divided into five groups (n=5) to begin the treatment experiment. The corresponding dose was administered via a single intravenous injection. 161 Tb-GSH-FAPI or 161 Tb-FAPI-04. Mouse body weight and tumor volume were measured and recorded every other day. The major and minor axes of the tumor were measured using calipers, and the volume was calculated according to the formula: Volume (mm²) 3 The tumor volume is calculated as (major diameter (mm) × minor diameter (mm) × minor diameter (mm) × 0.5), and the relative tumor volume (the ratio of the volume on the day of treatment to the initial volume) is used as the evaluation index, and monitoring continues until 14 days after drug administration.

[0186] Treatment results as follows Figure 6 As shown, in Figure 6 In (a), compared with the NS group, after 14 days of treatment 161 The tumor inhibition effect in the Tb-FAPI-04 group was not significant (0.5 mCivs NS, P>0.05; 1 mCivs NS, P=0.0174), while... 161 Medium and high concentrations of Tb-GSH-FAPI both showed significant therapeutic effects on tumors (P<0.0001). Furthermore, at the same therapeutic dose, 161 The therapeutic effect of Tb-GSH-FAPI is significantly better than that of Tb-GSH-FAPI. 161 Tb-FAPI-04 describes the new type 161 Tb-GSH-FAPI probe ratio 161 Tb-FAPI-04 has superior therapeutic effects. For example... Figure 6 As shown in (b), no significant weight loss was observed in any of the treatment groups during the treatment process.

[0187] Test Example 6: 68 Ga-GSH-FAPI and 68 Ga-FAPI-04 in PET imaging of gastric cancer patients

[0188] Patient and administration method: A 46-year-old male patient with gastric cancer was administered intravenously every other day. 68 Ga-GSH-FAPI and 68 Ga-FAPI-04, with an injection dose of 1 mCi, was used for PET / CT imaging.

[0189] Compare 68 Ga-GSH-FAPI compared to 68 Diagnostic efficacy of Ga-FAPI-04 PET / CT imaging in gastric cancer.

[0190] Experimental results: PET / CT imaging results are as follows Figure 7 As shown, 68 Ga-GSH-FAPI can significantly accumulate at the tumor (gastric cancer) site. Compared to 68 In addition to showing gastric cancer lesions, Ga-FAPI-04... 68 Ga-GSH-FAPI can detect lymph node metastases. This indicates that... 68 Ga-GSH-FAPI PET / CT imaging is superior to other imaging methods in terms of diagnostic sensitivity, specificity, and accuracy in gastric cancer. 68 Ga-FAPI-04 has a higher detection rate in early-stage gastric cancer primary lesions and metastatic lesions.

[0191] In summary, the FAP-targeting radiolabeled complex provided by this invention can significantly prolong its circulating half-life, enhance tumor uptake and retention time. This novel property is not found in other FAPI imaging agents and is expected to be used for radionuclide therapy and imaging of tumors with high FAP expression.

[0192] discuss:

[0193] Using GSH as a multifunctional linker, modular modifications of FAPI targeting molecules, radionuclide chelators, albumin-binding groups, and covalently coupled small molecules were achieved to construct tunable drug carriers. By controlling the number and type of albumin-binding groups linked to GSH, the half-life of the radionuclide (e.g., ...) can be matched. 68 Ga or 177 Lu), to optimize pharmacokinetics.

[0194] All documents mentioned in this invention are incorporated herein by reference as if each document were individually incorporated by reference. Furthermore, it should be understood that after reading the foregoing teachings of this invention, those skilled in the art can make various alterations or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims.

Claims

1. A compound of Formula I, or a pharmaceutically acceptable salt thereof, in, R1 is a nuclide labeling group selected from the following group: R4 is a FAP inhibitor targeting tumor stromal fibroblasts selected from the following group: Alternatively, the integrin αvβ3 receptor of newly formed vascular endothelial cells targets the RGD cyclic peptide: Indicates the connection point with other parts of the compound.

2. The compound according to claim 1, characterized in that, R1 is 3. A compound of formula II, or a pharmaceutically acceptable salt thereof, characterized in that, The structure is as follows: Wherein, R1 and R4 are as defined in claim 1; R2 is a small albumin-binding molecule selected from the following group: R3 is a covalently bonded group selected from the following group:

4. The compound according to claim 3, characterized in that, R3 is 5. The compound according to claim 1 or 3, characterized in that, The compounds are selected from the group consisting of:

6. A FAP targeting probe, characterized in that, The probe comprises a compound as described in claim 1 or 3 labeled with a radionuclide, wherein the radionuclide is selected from the group consisting of: 177 Lu、 90 Y、 18 F, 64 Cu、 68 Ga、 89 Zr、 225 Ac、 111 In、 99m Tc, 188 Re、 161 Tb, 212 Pb, 125 I, 131 I, 124 I.

7. A method for preparing the probe as described in claim 6, characterized in that, Includes the following steps: (a) Provide the compound or pharmaceutically acceptable salt as described in claim 1 or 3, and the radionuclide; (b) Prepare probes by wet labeling or lyophilization labeling methods; The radionuclide is as defined in claim 6.

8. A composition, characterized in that, Include: (i) the compound of any one of claims 1-5, or a pharmaceutically acceptable salt thereof, or the probe of claim 6; and (ii) Pharmaceutically acceptable carriers.

9. Use of a compound as claimed in any one of claims 1-5, or a pharmaceutically acceptable salt thereof, or a probe as claimed in claim 6, or a composition as claimed in claim 8, characterized in that, Used to prepare a drug for the treatment and / or diagnosis of diseases associated with fibroblast activation proteins.

10. The use as described in claim 9, characterized in that, The diseases mentioned are selected from the following group: head and neck cancer, breast cancer, lung cancer, pancreatic cancer, esophageal cancer, colorectal cancer, ovarian cancer, stomach cancer, liver cancer, or combinations thereof.

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