Cage-like chelate, targeted cage-like chelating ligand, and complex and use thereof

Abstract

The present application relates to a cage-like chelate, a targeted cage-like chelating ligand, and a complex and the use thereof. Provided in the present application is a cage-like chelate, which has a skeleton structure with more diverse modifiable sites and a higher flexibility, and has broad-spectrum adaptability. The cage-like chelate and the targeted cage-like ligand formed thereby can form a compound with a higher targeting ability and stability after complexation with a nuclide, thereby mitigating the daughter nuclide recoil effect of alpha nuclide-labeled drugs in organisms, and reducing toxic and side effects of alpha decay on normal tissues.

Description

Cage-like chelates, targeted cage-like chelating ligands and their complexes and applications

[0001] Related applications

[0002] This application claims priority to Chinese patent application No. 2024119910604, filed on December 31, 2024, entitled "Cage-like chelates, targeted cage-like chelating ligands and their complexes and applications", the entire contents of which are incorporated herein by reference. Technical Field

[0003] This application relates to the field of drug research with chelating structures, specifically to compounds and molecular fragments with chelating functions, compounds containing chelating structures and having targeting properties, their complexes and applications, particularly cage-like chelates, targeted cage-like chelating ligands and their complexes and applications. Background Technology

[0004] Radiopharmaceuticals (or "nuclear drugs") are a class of special drugs containing radioactive isotopes for medical diagnosis and treatment. They typically consist of a radioactive isotope paired with molecular reagents that specifically target particular organs and tissues. Radiopharmaceuticals utilize their targeted biological properties to reflect the state of diseased genes, molecules, metabolism, and function, enabling earlier and more specific insights into the molecular level of diseases. Furthermore, the radiation energy of the radioactive isotope can accurately kill tumors, making them a powerful tool for early disease diagnosis and precision treatment.

[0005] Radiopharmaceuticals consist of a chelating ligand linked to a targeted functional structure via a linker, which then chelates a radionuclide. This targeted functional structure can be an antibody, antibody fragment, small molecule, or peptide. These structures mediate the specific targeting of the entire drug molecule, delivering the radionuclide to the target site. Constructing therapeutic drugs by chelating different radionuclides with the same molecule is currently a hot topic in the industry. Therapeutic drugs allow the same targeting ligand and linker to be combined with radionuclides used for treatment or diagnostic imaging, respectively. Each therapeutic drug can have a corresponding diagnostic drug. After a patient is diagnosed with a disease, the appropriate drug can be used in conjunction with the treatment, saving time and improving efficiency.

[0006] However, several important technical problems still exist in the field of radiopharmaceuticals. First, the stability of radionuclide chelation is insufficient, resulting in instability of labeled radiopharmaceuticals and the deposition of free nuclides in bone, hematopoietic tissue, or other organs, causing radiation damage. Second, the regulation of metabolic properties during drug molecule development lacks systematic adjustment, leading to excessive randomness in the drug development process and significantly increasing development costs. Third, the reaction regulation of nuclide chelation is too stringent. Currently, commonly used DOTA-type chelating agents require heating during nuclide chelation, which affects the activity of heat-sensitive targeted functional structures (such as antibodies and antibody fragments), making it difficult to develop these highly active structures into radiopharmaceuticals. Fourth, secondary decay of nuclides can cause decomposition of the chelated structure or delabeling of decay daughter nuclei, leading to secondary distribution and toxic side effects. Summary of the Invention

[0007] To address the aforementioned technical problems, the inventors of this application believe that a superior chelating structure can solve the problems in the development and application of radiopharmaceuticals. Therefore, this application aims to provide a novel chelating structure, namely a cage-like structure, which resembles a net or clam-like molecular structure. This structure is highly suitable for capturing metal elements and further improves the thermodynamic and kinetic in vivo stability of the metal complex, reducing organ toxicity caused by demetallization, particularly slowing down the recoil nuclear effect of alpha nuclide decay. Simultaneously, it possesses more connection sites, providing a basis for connecting more target molecules and functional fragments, thereby enabling the drug to have better PKPD in vivo and further improving the druggability of therapeutic radiopharmaceuticals.

[0008] Based on this, this application discloses a cage-like chelate, wherein the chelating ligand compound is a compound of formula I or a pharmaceutically acceptable salt thereof:

[0009] in,

[0010] K1, K2, and K3 contain heteroalkyl, heterocycloalkyl, or heteroaryl atoms of O, S, P, or N; K1, K2, and K3 may be the same or different; where K1, K2, and K3 are heteroalkyl means that K1, K2, and K3 are composed of several R atoms. k It is formed by linking heteroatoms or heteroatom-containing groups, wherein R k Independently and arbitrarily selected from substituted or unsubstituted alkyl groups; preferably, R kSelected from substituted or unsubstituted methyl, substituted or unsubstituted ethyl, substituted or unsubstituted propyl; wherein K1, K2, and K3 are heteroalkylcycloalkyl groups, meaning that K1, K2, and K3 are 4-7 member substituted or unsubstituted heterocycloalkyl groups containing heteroatoms O, S, P, or N, preferably tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydropyrrolyl, or piperazineyl; K1, K2, and K3 are heteroaryl groups, meaning that K1, K2, and K3 are substituted or unsubstituted 4-7 member heteroaryl groups containing heteroatoms O, S, P, or N, preferably furanyl, thiophenyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl, pyridinyl, pyridinyl, isoxazolyl, diazolyl, pyrazolyl, triazolyl, tetrazolyl, isothiazolyl, or thiadiazolyl.

[0011] The N atom in Formula I is connected to H or a substituent.

[0012] The term "substituent" as used herein refers to the substitution or partial substitution of the hydrogen atom (H) on the nitrogen atom by a substituent or functional group. Here, "substitution" is optional, meaning that the substituents at each position in the structure can be the same or different. The term "substituent" encompasses all permissible substituents in organic compounds. In a broad sense, permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substitutions in organic compounds.

[0013] In one specific embodiment, this application further discloses that the substitution of the N atom can be a chelating arm of a heteroatom group with a lone pair of electrons, a target molecule linking arm with an active group AG that can link with a target molecule, or a non-target linking arm with a certain structure.

[0014] The chelating arm has the structural formula JQ, where Q is a heteroatom group; preferably, J is a substituted or unsubstituted, saturated or unsaturated ether, ester, ketone, amide, or C0-C5 alkyl chain; the target molecule linker has the structural formula L-AG, where AG is an active group capable of linking to the target molecule; in some technical solutions, AG is independently and arbitrarily selected from -X (Br, Cl, I, F), -OH, -COOH, -SH, -CO-N(CH3)OH, -H2PO3, H2PO4, -H2PO2, sulfonic acid, sulfinic acid, sulfone, sulfoxide, -OCH2COOH, -NH2, -NHR, -OCH2CH2NH2; -N3, -acetylene, -piperazine, piperidine, tetrahydropyrrole, -NCS, -NHS, -boronic acid (ester), -CHO, -COR.

[0015] Preferably, L is independently and arbitrarily selected from linear or branched, saturated or unsaturated alkyl chains, PEG, amino acids, polypeptides, piperazine, triazole, esters, sugars, ethers, ureas, guanidines, sulfonic acids, sulfinic acids, nucleic acids, amides, sulfones, sulfoxides, and their derivatives.

[0016] In some specific technical solutions, L is independently and arbitrarily selected. (n = 1 ~ 50), (n = 1 ~ 50),

[0017] When it is a non-targeted linker arm, it can be completely different from the target molecule linker arm, or it can be formed by replacing the active group on the target molecule linker arm with an inactive group. The non-targeted linker arm does not have the ability to further link to the target molecule, but it can be a group with other functions, or of course it can be non-functional.

[0018] The non-targeted connecting arm can be L N -NAG,L N Similar to the definition of L above, NAG is an alkyl group or H.

[0019] The active group AG on the target molecule linker arm L is a crucial part for linking with the target molecule. This application provides multiple target molecule linking sites, which lays the foundation for linking multiple target molecules or other functional molecules. Depending on the specific target molecule or target group requiring linking, different active groups AG can be linked to different linking sites to achieve connection with the target molecule. Of course, linking with the target molecule can be achieved through covalent bonds, coordinate bonds, or through hydrophobic bonds and electrostatic interactions.

[0020] In some technical solutions, J also includes a group AG1 that can be linked to the target molecule. AG1 has the same definition as AG, and AG1 and AG can be the same or different.

[0021] Group J is a component of the chelating arm. When it has group AG1 (mainly on the side chain), these active groups can connect to the target molecule. Therefore, this application provides more target connection sites under this technical solution. As mentioned above, it can be understood that the target sites connected by these active groups can be the same or different.

[0022] In some specific technical solutions, the cage-like chelate has any of the following preferred features:

[0023] (1) K1, K2, and K3 are independently and arbitrarily selected from:

[0024] (2) Group Q is selected from:

[0025] -H2PO3, -H2PO4, -H2PO2

[0026] Among them, R, R ’ "R" represents H, alkyl, hydroxyl, fatty acid group, amino, acyl, heteroalkyl, or heteroaryl.

[0027] (3) The cage-like chelate is arbitrarily selected from:

[0028] This application also discloses a targeted cage-like chelating ligand formed by coupling cage-like chelate C with one or more targeted molecules TM.

[0029] In some specific implementations, TM is independently and arbitrarily selected from biological macromolecules, or from drugs or small molecule compounds. The target molecules include, but are not limited to, one or more of antibodies, bispecific antibodies, triple antibodies, nanobodies, proteins, peptides, polymers, carbohydrates, nucleotides, oligonucleotides, oligosaccharides, vitamins, liposomes, cells, viruses, nanomaterials, small molecule drugs or fragments or derivatives. In particular, in some technical solutions, TM is selected from any one or more of the following target molecules.

[0030] Furthermore, this application also discloses that the targeted cage-like chelating ligand is:

[0031] Furthermore, this application also discloses the salt formed by the described cage-like chelate and an inorganic or organic acid. It should be understood that the salt referred to here means a pharmaceutically acceptable salt.

[0032] Furthermore, this application also discloses a method for preparing the cage-like chelate, which is prepared from 2-hydroxy-1,3,5-benzenetriformaldehyde and NH2—K1—NH2, NH2—K2—NH2, and NH2—K3—NH2 as raw materials, wherein K1, K2, and K3 are defined as described above.

[0033] This application also discloses metal complexes, which are metal complexes formed by the complexation of a cage-like chelate or a targeted cage-like chelating ligand with a metal.

[0034] This application also discloses the use of the aforementioned chelating ligand compounds, targeted chelating ligand compounds, organic or inorganic salt derivatives, and metal complexes in the preparation of radiopharmaceuticals, iron removal agents, and drugs for treating heavy metal poisoning.

[0035] This application discloses the use of metal complexes in radiopharmaceuticals. In this application, the metal is a metal nuclide, wherein the nuclide includes, but is not limited to, metal nuclides. 89 Zr、 47 Sc、 55 Co、 60 Cu、 61 Cu、 62 Cu、 64 Cu、 67 Cu、 66 Ga、 67 Ga、 68 Ga、 82 Rb、 86 Y、 87 Y、 90 Y、 97 Ru、 105 Rh、 109 Pd, 111 In、 117m Sn、 149 Pm, 52 Mn, 149 Tb, 152 Tb, 161 Tb, 99m Tc, 153 Sm、 177 Lu、 186 Re、 188 Re、 199 Au、 201 Tl、 203 Pb, 210 Pb, 212 Pb, 212 Bi、 213 Bi、 225 Ac、 223 Ra and 227 Th et al., among which the chelating ligand compounds disclosed in this application have particularly significant advantages in chelating with lanthanides and actinides, such as those used in the embodiments of this application. 68 Ga、 177 Lu、 225 Ac、 223 Ra、 89 Zr、 227 Th、 212 Pb.

[0036] This application breaks through the existing cyclic structure of chelates, boldly proposing and obtaining a cage-like chelate. This chelate provides a more diverse range of modifiable sites and a more flexible framework structure, which we call a "nuclide compartment," thus exhibiting broad adaptability and the ability to complex with various compounds, including... 68 Ga、177 Lu、 225 Ac、 223 Ra、 89 Zr、 227 Th、 212 Numerous nuclides, including Pb, were involved. Experiments showed that the cage-like chelate and its resulting targeting cage-like ligand exhibit excellent stability after complexing with the nuclides, effectively addressing the toxic side effects caused by poor stability. Simultaneously, the chelate provides more binding sites for the targeting molecule, thereby improving drug targeting and regulating biological properties such as in vivo distribution and metabolism. When the resulting radiopharmaceutical is used as a contrast agent for diagnosis or to exert therapeutic effects using alpha and beta rays and Auger electrons generated by the nuclide, it exhibits higher targeting and stability, significantly improving the daughter nucleus recoil effect of alpha-labeled drugs in vivo and reducing the toxic side effects of alpha decay in normal tissues. Attached Figure Description

[0037] To more clearly illustrate the technical solutions in the embodiments of this application or the conventional technology, the drawings used in the description of the embodiments or the conventional technology will be briefly introduced below. Obviously, the drawings described below are only embodiments of this application. For those skilled in the art, other drawings can be obtained based on the disclosed drawings without creative effort.

[0038] Figure 1 shows compound 2 in Example 1. 1 HNMR spectrum.

[0039] Figure 2 shows compound R1 in Example 1. 1 HNMR spectrum.

[0040] Figure 3 shows compound R2 in Example 5. 1 HNMR spectrum.

[0041] Figure 4 shows compound 8 in Example 6. 1 HNMR spectrum.

[0042] Figure 5 shows compound R3 in Example 6. 1 HNMR spectrum.

[0043] Figure 6 shows compound R4 in Example 7. 1 HNMR spectrum.

[0044] Figure 7 shows compound 26 in Example 13. 1 HNMR spectrum.

[0045] Figure 8 is a schematic diagram of the iTLC analysis results of the chelation reaction solution of compound R1 and metal nuclide Ga-68 in Example 15. In the figure, the Ga ion should be located at the origin.

[0046] Figure 9 shows the iTLC analysis results of the chelation reaction between compound R1 and the metal nuclide Zr-89 in Example 15. In the figure, the Zr ion should be located at the leading edge.

[0047] Figure 10 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1-1 and the metal nuclide Ga-68 in Example 16. In the figure, the Ga ion should be located at the origin.

[0048] Figure 11 shows the HPLC analysis results of the chelation reaction between compound R1-1 and the metal nuclide Ga-68 in Example 16.

[0049] Figure 12 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1-2 and the metal nuclide Ga-68 in Example 17. In the figure, the Ga ion should be located at the origin.

[0050] Figure 13 shows the HPLC analysis results of the chelation reaction between compound R1-2 and the metal nuclide Ga-68 in Example 17.

[0051] Figure 14 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1-5 and the metal nuclide Ga-68 in Example 18. In the figure, the Ga ion should be located at the origin.

[0052] Figure 15 shows the HPLC analysis results of the chelation reaction between compound R1-5 and the metal nuclide Ga-68 in Example 18.

[0053] Figure 16 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1-6 and the metal nuclide Ga-68 in Example 19. In the figure, the Ga ion should be located at the origin.

[0054] Figure 17 shows the HPLC analysis results of the chelation reaction between compound R1-6 and the metal nuclide Ga-68 in Example 19.

[0055] Figure 18 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1-9 and the metal nuclide Ga-68 in Example 20. In the figure, the Ga ion should be located at the origin.

[0056] Figure 19 shows the HPLC analysis results of the chelation reaction between compound R1-9 and the metal nuclide Ga-68 in Example 20.

[0057] Figure 20 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1-10 and the metal nuclide Ga-68 in Example 21. In the figure, the Ga ion should be located at the origin.

[0058] Figure 21 shows the iTLC analysis results of the chelation reaction between compound R1-10 and the metal nuclide Ga-68 in Example 21. The Ga ion should be located at the origin in the figure.

[0059] Figure 22 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R2 and the metal nuclide Ga-68 in Example 22. In the figure, the Ga ion should be located at the origin.

[0060] Figure 23 shows the iTLC analysis results of the chelation reaction between compound R2 and the metal nuclide Zr-89 in Example 22. The Zr ions should be located at the leading edge in the figure.

[0061] Figure 24 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R3 and the metal nuclide Ga-68 in Example 23. In the figure, the Ga ion should be located at the origin.

[0062] Figure 25 shows the iTLC analysis results of the chelation reaction between compound R3 and the metal nuclide Zr-89 in Example 23. In the figure, the Zr ion should be located at the leading edge.

[0063] Figure 26 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R4 and the metal nuclide Ga-68 in Example 24. In the figure, the Ga ion should be located at the origin.

[0064] Figure 27 shows the iTLC analysis results of the chelation reaction between compound R4 and the metal nuclide Zr-89 in Example 24. In the figure, the Zr ion should be located at the leading edge.

[0065] Figure 28 shows the iTLC analysis results of the chelation reaction between compound R4 and the metal nuclide Lu-177 in Example 24. The Lu ion should be located at the leading edge in the figure.

[0066] Figure 29 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R5 and the metal nuclide Ga-68 in Example 25. In the figure, the Ga ion should be located at the origin.

[0067] Figure 30 shows the iTLC analysis results of the chelation reaction between compound R5 and the metal nuclide Zr-89 in Example 25. In the figure, the Zr ion should be located at the leading edge.

[0068] Figure 31 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R7 and the metal nuclide Ga-68 in Example 26. In the figure, the Ga ion should be located at the origin.

[0069] Figure 32 shows the iTLC analysis results of the chelation reaction between compound R7 and the metal nuclide Zr-89 in Example 26. In the figure, the Zr ion should be located at the leading edge.

[0070] Figure 33 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R9 and the metal nuclide Ga-68 in Example 27. In the figure, the Ga ion should be located at the origin.

[0071] Figure 34 shows the iTLC analysis results of the chelation reaction between compound R9 and the metal nuclide Zr-89 in Example 27. In the figure, the Zr ion should be located at the leading edge.

[0072] Figure 35 is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R10-1 and the metal nuclide Ga-68 in Example 28. In the figure, the Ga ion should be located at the origin.

[0073] Figure 36 shows the iTLC analysis results of the chelation reaction between compound R10-1 and the metal nuclide Ga-68 in Example 28. In the figure, the Ga ion should be located at the origin.

[0074] Figure 37 shows the iTLC analysis results of the chelation reaction between compound R10-1 and the metal nuclide Zr-89 in Example 28. In the figure, the Zr ion should be located at the leading edge.

[0075] Figure 38 shows the iTLC analysis results of the chelation reaction between the coupling protein R10-KT and the metal nuclide Zr-89 in Example 29. In the figure, the Zr ion should be located at the leading edge.

[0076] Figure 39 shows the example 30. 68 PET imaging results of Ga-R1-1 in a mouse model bearing U87-MG tumors.

[0077] Figure 40 shows the example 31. 68 PET imaging results of Ga-R1-2 in a mouse model bearing U87-MG tumors.

[0078] Figure 41 shows the example 32. 68 PET imaging results of Ga-R1-5 in a mouse model bearing U87-MG tumor.

[0079] Figure 42 shows the example 33. 68 PET imaging results of Ga-R1-6 in a mouse model bearing U87-MG tumor.

[0080] Figure 43 shows the example 34. 68 PET imaging results of Ga-R1-9 in a mouse model bearing U87-MG tumor.

[0081] Figure 44 shows the example 35. 68 PET imaging results of Ga-R1-10 in a mouse model bearing U87-MG tumors.

[0082] Figure 45 shows the example 36. 68 PET imaging results of Ga-R10-1 in a mouse model bearing U87-MG tumor. Detailed Implementation

[0083] 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 embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0084] To better understand this application, we will further elaborate on it below with reference to specific embodiments.

[0085] Unless otherwise specified or noted in the examples, all reagents and instruments are commercially available conventional products that can be purchased. Unless otherwise specified or noted, all conditions can be carried out in accordance with conventional conditions in the field or the instructions of the relevant product seller.

[0086] This application uses substituted or unsubstituted 2-hydroxy-1,3,5-benzenetrialdehyde and NH2—K—NH2 as raw materials to prepare the target compound. Specifically, substituted or unsubstituted 2-hydroxy-1,3,5-benzenetrialdehyde is mixed and reacted with NH2—K—NH2 to obtain a macrocyclic compound (Schiff base), which is then reduced with sodium borohydride and reacted with tert-butyl bromoacetate (or hydroxylamine intermediate or phosphate intermediate). The obtained intermediate is hydrolyzed to generate the target compound.

[0087] Example 1

[0088] 2-Hydroxy-1,3,5-benzyltricarboxaldehyde (534 mg, 3 mmol) was dissolved in 30 mL of DMF solution and heated to 90 °C. At 90 °C, 30 mL of 2,2′-(ethylenedioxy)bis(ethylamine) (674.8 mg, 4.5 mmol) solution was added dropwise. After the addition was complete, the reaction was continued for 2 hours. The reaction solution was then concentrated to give compound 1.

[0089] Sodium borohydride (610 mg, 16 mmol) was added to a methanol solution of compound 1 with stirring, and the reaction was stirred for 1 hour. The solvent was removed under reduced pressure, and the residue was washed with ethyl acetate / water and dried over anhydrous sodium sulfate. The mixture was filtered, and 10 mL of dioxane (4 M) solution of hydrogen chloride was added to the filtrate. The precipitated solid was filtered and dried to give the hydrochloride salt of compound 2 (746 mg).

[0090] Compound 2:

[0091] LC-MS: 705.2 (M+1);

[0092] H-NMR (300M, D2O): δ 3.27-3.33 (m, 12H), 3.72-3.85 (m, 28H), 4.19-4.21 (d, 8H), 7.49-7.50 (d, 4H). See Figure 1 for the specific spectrum.

[0093] The hydrochloride salt of compound 2 (326 mg, 0.46 mmol), tert-butyl bromoacetate (632.8 mg, 3.24 mmol), DIEA (597 mg, 4.6 mmol), and 30 mL of acetonitrile were mixed and reacted at room temperature for 2 hours. The mixture was concentrated and dried, extracted with ethyl acetate / water, dried and concentrated to obtain the crude product, and purified to obtain compound 3 (323 mg).

[0094] Compound 3:

[0095] LC-MS: 695.4 (M / 2+1).

[0096] Compound 3 (200 mg, 0.144 mmol) was dissolved in 4 M hydrochloric acid (3 ml) and reacted at 45 °C for 0.5 h. The solution was concentrated and dried under reduced pressure to prepare the purified target compound R1 (10.1 mg). The specific spectrum is shown in Figure 2.

[0097] Compound R1:

[0098] LC-MS: 1053.1(M+1), 527.2(M / 2+1), 351.9(M / 3+1);

[0099] H-NMR (300M, D2O): δ3.9-3.64 (m, 12H), 3.79-4.05 (m, 36H), 4.40-4.63 (m, 12H), 7.59-7.63 (dd, 4H).

[0100] Example 2

[0101] In this embodiment, we prepared compounds R1-1, R1-2, R1-3, and R1-4 by coupling compound R1 with compound W.

[0102] Compound R1 (105.2 mg, 0.1 mmol), N,N-diisopropylethylamine (116.1 mg, 0.9 mmol), and HATU (114 mg, 0.3 mmol) were dissolved in 2 mL of DMF solution. After stirring for 10 min, compound W (145.8 mg, 0.3 mmol) was added. After reacting for 6 h, the reaction solution was concentrated under reduced pressure, and the compounds were purified by preparative liquid chromatography.

[0103] R1-1 (33mg), LC-MS: 761.3 (M / 2+1);

[0104] R1-2 (13 mg), LC-MS: 663.3 (M / 3+1);

[0105] R1-3 (9mg), LC-MS: 663.3 (M / 3+1);

[0106] R1-4 (15 mg), LC-MS: 663.3 (M / 3+1).

[0107] Example 3

[0108] In this embodiment, we prepared compounds R1-5, R1-6, R1-7, and R1-8 by coupling compound R1 with compound X.

[0109] Compound R1 (105.2 mg, 0.1 mmol), N,N-diisopropylethylamine (116.1 mg, 0.9 mmol), and HATU (114 mg, 0.3 mmol) were dissolved in 2 mL of DMF solution. After stirring for 10 min, compound X (249.6 mg, 0.3 mmol) was added. After reacting for 6 h, the reaction solution was concentrated under reduced pressure, and the compounds were purified by preparative liquid chromatography.

[0110] R1-5 (35mg), LC-MS: 622.2 (M / 3+1);

[0111] R1-6 (7mg), LC-MS: 670.5 (M / 4+1);

[0112] R1-7 (15mg), LC-MS: 670.5 (M / 4+1);

[0113] R1-8 (13 mg), LC-MS: 670.5 (M / 4+1).

[0114] Example 4

[0115] In this embodiment, we prepared compounds R1-9, R1-10, R1-11, and R1-12 by coupling compound R1 with compound Y.

[0116] Compound R1 (105.2 mg, 0.1 mmol), N,N-diisopropylethylamine (116.1 mg, 0.9 mmol), and HATU (114 mg, 0.3 mmol) were dissolved in 2 ml of DMF solution and stirred for 10 min. Then, compound Y (196.5 mg, 0.3 mmol) was added and reacted for 6 h. The reaction solution was concentrated under reduced pressure and the compounds were purified by preparative liquid chromatography.

[0117] R1-9 (19.6 mg), LC-MS: 845.9 (M / 2+1), 564.3 (M / 3+1);

[0118] R1-10 (13.2mg), LC-MS: 776.6 (M / 3+1), 582.9 (M / 4+1);

[0119] R1-11 (10.2mg), LC-MS: 776.6 (M / 3+1), 582.9 (M / 4+1);

[0120] R1-12 (7.5 mg), LC-MS: 776.6 (M / 3+1), 582.9 (M / 4+1).

[0121] Example 5

[0122] The hydrochloride salt of compound 2 (22 mg, 0.031 mmol), 1-benzyloxy-6-(bromomethyl)pyridin-2-one (64 mg, 0.22 mmol), DIEA (48 mg, 0.375 mmol), and 5 mL of acetonitrile were mixed and reacted at room temperature for 2 hours. The mixture was concentrated and dried, extracted with ethyl acetate / water, dried and concentrated again to obtain the crude product, and purified to obtain compound 5. Compound 5-A (9.5 mg, 0.005 mmol) was dissolved in 4 M hydrochloric acid (1 mL) and reacted at 45 °C for 0.5 hours. The mixture was concentrated and dried under reduced pressure, and purified to obtain the target compound R2 (3.34 mg). See Figure 3 for the specific chromatogram.

[0123] Compound R2:

[0124] LC-MS: 482.1(M / 3+1), 722.5(M / 2+1);

[0125] H-NMR (300M, D2O): δ3.57-4.77 (m, 60H), 6.58-6.75 (m, 10H), 7.38-7.44 (m, 12H).

[0126] Example 6

[0127] 2-Hydroxy-1,3,5-benzyltricarboxaldehyde (178 mg, 1 mmol) was dissolved in 30 mL of DMF solution and heated to 90 °C. At 90 °C, a 30 mL solution of diethylenetriamine (159 mg, 1.5 mmol) was added dropwise. After the addition was complete, the reaction was continued for 2 hours. The reaction solution was then concentrated to obtain compound 7.

[0128] Sodium borohydride (304 mg, 8 mmol) was added to a methanol solution of compound 7 with stirring, and the reaction was stirred for 1 hour. The solvent was removed under reduced pressure, and the residue was washed with ethyl acetate / water and dried over anhydrous sodium sulfate. The mixture was filtered, and 10 mL of dioxane (4 M) solution of hydrogen chloride was added to the filtrate. The precipitated solid was filtered and dried to give the hydrochloride salt of compound 8 (241 mg).

[0129] Compound 8:

[0130] LC-MS: 570.4 (M+1);

[0131] H-NMR (300M, D2O): δ 3.33-3.48 (m, 24H), 4.10-4.14 (d, 4H), 4.27 (s, 4H), 4.59-4.65 (t, 4H), 7.48 (s, 4H). See Figure 4 for the specific spectrum.

[0132] The hydrochloride salt of compound 8 (78 mg, 0.14 mmol), tert-butyl bromoacetate (267 mg, 1.4 mmol), DIEA (336 mg, 2.66 mmol), and 5 mL of acetonitrile were mixed and reacted at room temperature for 2 hours. The mixture was concentrated and dried, extracted with ethyl acetate / water, dried and concentrated to obtain the crude product, and purified to obtain compound 9 (70 mg).

[0133] Compound 9 (70 mg, 0.043 mmol) was dissolved in 4 M hydrochloric acid (1 ml) and reacted at 45 °C for 0.5 h. The solution was concentrated and dried under reduced pressure to obtain the target compound R3 (15.69 mg).

[0134] Compound R3:

[0135] LC-MS: 1091.6(M+1), 546.8(M / 2+1);

[0136] H-NMR (300M, D2O): δ3.16-4.59 (m, 54H), 7.73 (s, 4H), see Figure 5 for the specific spectrum.

[0137] Example 7

[0138] Following the method of Example 5, only tert-butyl bromoacetate was replaced with 1-benzyloxy-6-(bromomethyl)pyridin-2-one to obtain compound R4.

[0139] Compound R4:

[0140] LC-MS: 839.6(M / 2+1), 560.1(M / 3+1);

[0141] ¹H-NMR (300M, D₂O): δ 2.94–4.26 (m, 54H), 6.61–7.51 (m, 31H). See Figure 6 for the detailed spectrum.

[0142] Example 8

[0143] Following the method of Example 6, only diethylenetriamine was replaced with 2,2'-oxobis(ethylamine) to obtain compound R5.

[0144] in:

[0145] Compound 14:

[0146] LC-MS: 573.2 (M+1);

[0147] H-NMR (300M, D2O): δ3.04-3.16 (m, 12H), 3.91-4.04 (m, 24H), 7.61 (s, 4H).

[0148] Compound R5:

[0149] LC-MS: 921.4(M+1), 461.2(M / 2+1);

[0150] H-NMR (300M, D2O): δ4.51-5.09 (m, 48H), 7.51 (dd, 4H).

[0151] Example 9

[0152] Following the method of Example 6, only diethylenetriamine was replaced with aminoethyl sulfide to obtain compound R6.

[0153] in:

[0154] Compound R6:

[0155] LC-MS: 968.9(M+1), 485.0(M / 2+1);

[0156] H-NMR (300M, D2O): δ2.52-4.46 (m, 48H), 7.49-7.51 (d, 4H).

[0157] Example 10

[0158] Following the method of Example 5, only tert-butyl bromoacetate was replaced with 1-benzyloxy-6-(bromomethyl)pyridin-2-one to obtain compound R7.

[0159] Compound R7:

[0160] LC-MS: 656.1 (M / 2+1);

[0161] H-NMR (300M, D2O): δ4.31-5.22 (m, 48H), 6.41-8.02 (m, 22H).

[0162] Example 11

[0163] Following the method of Example 5, only tert-butyl bromoacetate was replaced with 1-benzyloxy-6-(bromomethyl)pyridin-2-one to obtain compound R8.

[0164] Compound R8:

[0165] LC-MS: 453.9(M / 3+1), 680.1(M / 2+1);

[0166] H-NMR (300M, D2O): δ2.91-4.31 (m, 48H), 6.11-8.09 (m, 22H).

[0167] Example 12

[0168] Following the method of Example 5, only tert-butyl bromoacetate was replaced with 2-bromo-N-((4-methoxybenzyl)oxy)-N-methylacetamide to obtain compounds R9-A and R9-B.

[0169] Compound R9-A:

[0170] LC-MS: 339.2(M / 4+1), 452.0(M / 3+1), 677.4(M / 2+1).

[0171] Compound R9-B:

[0172] LC-MS: 339.2(M / 4+1), 452.0(M / 3+1), 677.4(M / 2+1).

[0173] Example 13

[0174] Dissolve 123 mg (0.75 mmol) of 2-hydroxy-1,3,5-benzyltrialdehyde in 30 mL of DMF solution and heat to 90 °C. At 90 °C, add dropwise a mixture of 39 mg (0.375 mmol) of diethylenetriamine and 100 mg (0.375 mmol) of tert-butyl (2-(2-aminoethyl)amino)ethyl (100 mg, 0.375 mmol) carbamate in 30 mL of DMF solution. After the addition is complete, continue the reaction for 2 hours. Then concentrate the reaction solution.

[0175] Sodium borohydride (128 mg, 3.3 mmol) was added, and the mixture was stirred for 1 hour. The solvent was removed under reduced pressure, and the residue was washed with ethyl acetate / water and dried over anhydrous sodium sulfate. Preparative liquid chromatography purification gave compound 26 (60 mg).

[0176] Compound 26

[0177] LC-MS: 713.5 (M+1)

[0178] 1H-NMR (300M, D2O): δ1.47 (s, 9H), δ2.94-2.96 (m, 4H), 3.20-3.55 (m, 24H), δ4.04-4.13 (m, 4H), δ4.22-4.26 (m, 4H), δ4.53-4.62 (m, 4H), δ7.45-7.46 (d, 4H). See Figure 7 for the detailed spectrum.

[0179] Compound 26 (41 mg, 0.057 mmol), 2-bromo-N-((4-methoxybenzyl)oxy)-N-methylacetamide (55 mg, 0.191 mmol), DIEA (55 mg, 0.426 mmol), and 5 mL of acetonitrile were mixed and reacted at room temperature for 2 hours. The mixture was concentrated and dried, extracted with ethyl acetate / water, dried and concentrated to obtain the crude product, and purified to obtain compound 27 (35 mg).

[0180] Compound 27:

[0181] LC-MS: 791.0 (M / 2+1).

[0182] Compound 27 (30 mg, 0.0126 mmol) was dissolved in 4 M hydrochloric acid (3 ml) and reacted at 45 °C for 0.5 h. The solution was concentrated and dried under reduced pressure to obtain the target compound R10 (10.1 mg).

[0183] Compound R10:

[0184] LC-MS: 655.3 (M / 2+1).

[0185] Compound R10 (10 mg, 0.0076 mmol), tert-butyl bromoacetate (1.65 mg, 0.0084 mmol), DIEA (9.8 mg, 0.076 mmol), and 2 mL of acetonitrile were mixed and reacted at room temperature for 2 hours. The mixture was concentrated and dried, extracted with ethyl acetate / water, dried and concentrated to obtain the crude product, and purified to obtain compound 29 (9.8 mg).

[0186] Compound 29 (9.8 mg, 0.0072 mmol), N,N-diisopropylethylamine (9.8 mg, 0.076 mmol), and HATU (2.74 mg, 0.0072 mmol) were dissolved in 1 mL of DMF solution and stirred for 10 min. Then, compound W (3.55 mg, 0.0073 mmol) was added and reacted for 1 h. The reaction solution was concentrated under reduced pressure and compound R10-1 was purified by preparative liquid chromatography.

[0187] Compound R10-1:

[0188] LC-MS: 613 (M / 3+1).

[0189] Example 14

[0190] Compound 27 (114 mg, 0.05 mmol) was dissolved in 1 ml of dichloromethane, and 0.2 ml of trifluoroacetic acid was added to remove Boc and purify to give compound 30 (67 mg).

[0191] Compound 30:

[0192] LCMS: 757.8 (M / 3+1)

[0193] Compound 30 (66 mg, 0.03 mmol), Boc-5-amino-3-oxavaranic acid (7.2 mg, 0.033 mmol), HATU (12.5 mg, 0.033 mmol), and DIEA (39 mg, 0.3 mmol) were added sequentially to 0.5 ml of DMF. After 1 hour, the mixture was directly purified to obtain compound 31 (40 mg).

[0194] Compound 31:

[0195] LCMS: 724.8 (M / 3+1)

[0196] Compound 31 (39 mg, 0.016 mmol) was dissolved in 0.5 mL of trifluoroacetic acid. All protecting groups were removed, and the trifluoroacetic acid was concentrated to obtain 15.1 mg (0.011 mmol) of purified compound. The intermediate was dissolved in DMF and added to a DMF solution of p-phenylisothiocyanate (9.6 mg, 0.05 mmol). After the reaction was complete, the solution was concentrated. Compound 32 (3.7 mg) was obtained through purification.

[0197] Compound 32:

[0198] LC-MS: 801.9 (M / 2+1)

[0199] Add 2 mg of antibody KT (150 kDa) to PBS to a final volume of 1 mL, and adjust the pH to 8.9 ± 0.2 with NaHCO3. Dissolve compound 32 (0.4 mg) in DMSO (30 μL) and mix well. Incubate the reaction solution on a shaker at 200 rpm and 37 °C for 30 minutes.

[0200] The coupling reaction mixture was transferred to an ultrafiltration tube using a pipette, and the reaction tube was washed with ammonium citrate. The mixture was then washed five times with sodium ammonium citrate and recovered. The concentration was measured using a micro spectrophotometer to obtain the antibody-conjugated small molecule compound R10-KT.

[0201] Example 15

[0202] The chelation method of compound R1 with the metal nuclide Ga-68 is as follows: Take a sample containing... 68 Ga 3+ 1 mL of hydrochloric acid solution was placed in a centrifuge tube, and the pH was adjusted to 4.5-5.0 with 1 M sodium acetate solution. Then, an aqueous solution containing 20 nmol R1 was added, the centrifuge tube was sealed, and the reaction was carried out at room temperature for 10 min. The reaction yield was then analyzed by instantaneous thin-layer chromatography (iTLC) (iTLC analysis conditions: stationary phase iTLC-SG rapid chromatography; mobile phase: methanol / 1 M ammonium acetate = 1 / 1).

[0203] The chelation method of compound R1 with the metal nuclide Zr-89 is as follows: 100 μL of 0.25 M HEPES buffer solution is placed in a centrifuge tube, an aqueous solution containing 20 nmol R1 is added, and then 10 μL of a solution containing... 89 Zr 4+ The solution was mixed, and the pH of the reaction system was adjusted to between 6.0 and 6.5 with 0.25M Na2CO3 solution. The mixture was then reacted at room temperature for 1 hour. The radiochemical purity of the product was determined by iTLC (stationary phase iTLC-SG plate, mobile phase was 0.1M sodium citrate buffer at pH=5).

[0204] Figure 8 shows the iTLC analysis results of the chelation reaction solution of compound R1 with the metal nuclide Ga-68. The chelation reaction yield was 90%, indicating that R1 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0205] Figure 9 shows the iTLC analysis results of the chelation reaction solution of compound R1 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R1 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0206] Example 16

[0207] The chelation reaction conditions of compound R1-1 with Ga-68 were the same as in Example 15. The reaction solution was analyzed by iTLC and HPLC (chromatographic conditions: Phenomenex Luna C18(2) column (5μ, 150×4.60mm); flow rate 1mL / min; gradient elution, 0 to 3 minutes acetonitrile 10%, trifluoroacetic acid solution (0.1%) 90%, 3 to 10 minutes acetonitrile from 10% to 70%, trifluoroacetic acid solution from 90% to 30%, 10 to 12 minutes acetonitrile from 70% to 10%, trifluoroacetic acid solution from 30% to 90%, 12 to 15 minutes acetonitrile maintained at 10%, trifluoroacetic acid solution maintained at 90%). The iTLC analysis results of the reaction solution are shown in Figure 10. The chelation reaction yield was 91%, indicating that R1-1 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0208] The HPLC analysis results of the reaction solution are shown in Figure 11. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R1-1 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0209] Example 17

[0210] The chelation reaction conditions for compound R1-2 with Ga-68 were the same as in Example 15. The reaction solution was analyzed by iTLC and HPLC. The iTLC results are shown in Figure 12. The chelation reaction yield was 91%, indicating that R1-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0211] The HPLC analysis results of the reaction solution are shown in Figure 13. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R1-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0212] Example 18

[0213] The chelation reaction conditions for compound R1-5 with Ga-68 were the same as in Example 15. The reaction solution was analyzed by iTLC and HPLC. The iTLC results are shown in Figure 14. The results showed that the chelation reaction yield was 90%, indicating that R1-5 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0214] The HPLC analysis results of the reaction solution are shown in Figure 15. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R1-5 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0215] Example 19

[0216] The chelation reaction conditions for compound R1-6 with Ga-68 were the same as in Example 15. The reaction solution was analyzed by iTLC and HPLC. The iTLC results are shown in Figure 16. The results showed that the chelation reaction yield was 90%, indicating that R1-6 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0217] The HPLC analysis results of the reaction solution are shown in Figure 17. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R1-6 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0218] Example 20

[0219] The chelation reaction conditions for compound R1-9 with Ga-68 were the same as in Example 15. The reaction solution was analyzed by iTLC and HPLC. Compound R1-9 was labeled using a common method. The iTLC analysis results are shown in Figure 18. The chelation reaction yield was 91%, indicating that R1-9 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0220] The HPLC analysis results of the reaction solution are shown in Figure 19. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R1-9 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0221] Example 21

[0222] The chelation reaction conditions for compound R1-10 with Ga-68 were the same as in Example 15. The reaction solution was analyzed by iTLC and HPLC. The iTLC results are shown in Figure 20. The results showed that the chelation reaction yield was 92%, indicating that R1-10 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0223] The HPLC analysis results of the reaction solution are shown in Figure 21. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R1-10 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0224] Example 22

[0225] The chelation reaction conditions of compound R2 with Ga-68 and Zr-89 were the same as in Example 15, and the reaction solutions were analyzed by iTLC. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 22. The results showed that the chelation reaction yield was 91%, indicating that R2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0226] Figure 23 shows the iTLC analysis results of the chelation reaction solution of compound R2 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R2 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0227] Example 23

[0228] The chelation reaction conditions of compound R3 with Ga-68 and Zr-89 were the same as in Example 15, and the reaction solutions were analyzed by iTLC. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 24. The results showed that the chelation reaction yield was 92%, indicating that R3 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0229] Figure 25 shows the iTLC analysis results of the chelation reaction solution of compound R3 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R3 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0230] Example 24

[0231] The chelation reaction conditions of compound R4 with Ga-68 and Zr-89 were the same as in Example 15, and the reaction solutions were analyzed by iTLC. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 26. The results showed that the chelation reaction yield was 94%, indicating that R4 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0232] Figure 27 shows the iTLC analysis results of the chelation reaction solution of compound R4 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R4 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0233] The chelation method of compound R4 with the metal nuclide Lu-177 is as follows: Commercially available compound R4 is diluted with 0.04M ultrapure hydrochloric acid. 177 Adjust the LuCl3 solution to a suitable radiochemical concentration, take 0.5 mL and place it in a centrifuge tube. Adjust the pH to 5.0-5.5 with 0.25 M sodium acetate solution, then add an aqueous solution containing 20 nmol R1, mix well, and react at room temperature for 1 hour. The radiochemical purity of the product was determined by iTLC (stationary phase iTLC-SG plate, mobile phase: 0.1 M sodium citrate buffer at pH 5). The analytical results are shown in Figure 28. The results show that the radiochemical purity of the chelated product is greater than 95%, indicating that R1 can undergo a highly efficient chelation reaction with Lu-177 ions.

[0234] Example 25

[0235] The chelation reaction conditions of compound R5 with Ga-68 and Zr-89 were the same as in Example 15, and the reaction solutions were analyzed by iTLC. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 29. The results showed that the chelation reaction yield was 91%, indicating that R5 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0236] Figure 30 shows the iTLC analysis results of the chelation reaction solution of compound R5 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R5 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0237] Example 26

[0238] The chelation reaction conditions of compound R7 with Ga-68 and Zr-89 were the same as in Example 15, and the reaction solutions were analyzed by iTLC. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 31. The results showed that the chelation reaction yield was 91%, indicating that R7 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0239] Figure 32 shows the iTLC analysis results of the chelation reaction solution of compound R7 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R7 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0240] Example 27

[0241] The chelation reaction conditions of compound R9 with Ga-68 and Zr-89 were the same as in Example 15, and the reaction solutions were analyzed by iTLC. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 33. The results showed that the chelation reaction yield was 91%, indicating that R9 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0242] Figure 34 shows the iTLC analysis results of the chelation reaction solution of compound R9 with the metal nuclide Zr-89. The results show that the chelation reaction yield is greater than 95%, indicating that R9 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0243] Example 28

[0244] The chelation reaction conditions for compound R10-1 and Ga-68 were the same as in Example 15. The reaction solutions were analyzed by iTLC and HPLC, respectively. The iTLC analysis results of the Ga-68 labeled reaction solution are shown in Figure 35. The results showed that the chelation reaction yield was 90%, indicating that R10-1 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0245] The HPLC analysis results of the reaction solution are shown in Figure 36. The results show that the radiochemical purity of the product in the reaction solution is greater than 95%, further proving that R10-1 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0246] The chelation reaction conditions of compound R10-1 with Zr-89 were the same as in Example 15. The reaction solution was analyzed by iTLC, and the results are shown in Figure 37. The chelation reaction yield was greater than 95%, indicating that R10-1 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0247] Example 29

[0248] The Zr-89 labeling method for protein R10-KT modified by conjugated chelate ligand compounds is as follows: 100 μL of 0.25 M HEPES solution is placed in a centrifuge tube, and then... 89 Zr 4+ The pH of the solution was adjusted to 6.0-6.5 using 0.25M Na₂CO₃ solution. Then, 0.1-1 mg of protein R10-KT was added, mixed well, and the reaction solution was placed at room temperature. After 60 min, the reaction effect was monitored by iTLC (iTLC analysis conditions: stationary phase iTLC-SG rapid chromatography; mobile phase: 0.1M sodium citrate buffer, pH=5). The iTLC analysis results of the reaction solution are shown in Figure 38. The results show that the chelation reaction yield is greater than 95%, indicating that the coupled protein R10-KT can undergo a highly efficient chelation reaction with Zr-89 ions.

[0249] Example 30

[0250] R1-1, a Ga-68 chelate prepared in Example 16 with a radioactivity of 100 μCi, was injected via the tail vein into a nude mouse model bearing U87-MG tumors. PET imaging data was acquired 60 minutes after injection, and the results are shown in Figure 39. The compound can be seen... 68 Ga-R1-1 showed significant uptake in U87-MG tumor tissue (indicated by the arrow in the figure), but lower uptake in other organs besides the bladder.

[0251] Example 31

[0252] The R1-2 chelated Ga-68 prepared in Example 17 was used in a nude mouse model bearing U87-MG tumors for PET imaging studies, using the same method as in Example 28. The results are shown in Figure 40. The compound... 68 Ga-R1-2 was significantly uptaken in U87-MG tumor tissue (as indicated by the arrow in the figure) and also had high retention in the liver.

[0253] Example 32

[0254] The R1-5 chelated Ga-68 prepared in Example 18 was used in a nude mouse model bearing U87-MG tumors for PET imaging studies, using the same method as in Example 28. The results are shown in Figure 41. The compound... 68 Ga-R1-5 was significantly uptaken in U87-MG tumor tissue (as indicated by the arrow in the figure).

[0255] Example 33

[0256] The R1-6 chelate of Ga-68 prepared in Example 19 was used in a nude mouse model bearing U87-MG tumors for PET imaging studies, using the same method as in Example 28, as shown in Figure 42. The compound... 68 Ga-R1-6 was significantly uptaken in U87-MG tumor tissue (as indicated by the arrow in the figure), but there was also high uptake in the liver.

[0257] Example 34

[0258] The R1-9 chelated Ga-68 prepared in Example 20 was used in a nude mouse model bearing U87-MG tumors for PET imaging studies, using the same method as in Example 28, as shown in Figure 43. The compound... 68 Ga-R1-9 is uptaken in small amounts in U87-MG tumor tissue (as indicated by the arrow in the figure), and in higher amounts in the kidneys.

[0259] Example 35

[0260] The Ga-68 chelate R1-10 prepared in Example 21 was used in a nude mouse model bearing U87-MG tumors for PET imaging studies, using the same method as in Example 28, as shown in Figure 44. The compound... 68 Ga-R1-10 is uptaken in U87-MG tumor tissue (as indicated by the arrow in the figure), with higher uptake in the kidneys.

[0261] Example 36

[0262] The R10-1 chelated Ga-68 prepared in Example 28 was used in a nude mouse model bearing U87-MG tumors for PET imaging studies, using the same method as in Example 28, as shown in Figure 45. The compound... 68 Ga-R10-1 is uptaken in U87-MG tumor tissue (as indicated by the arrow in the figure), but the highest uptake is found in the liver.

[0263] The above describes specific embodiments of this application. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of this application, and these improvements and modifications are also considered to be within the scope of protection of this application.

[0264] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0265] The embodiments described above are merely illustrative of several implementation methods of this application, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the patent application. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of this application, and these all fall within the protection scope of this application. Therefore, the protection scope of this patent application should be determined by the appended claims.

Claims

1. A cage-like chelate, characterized in that, The chelating ligand compound is a compound of Formula I or a pharmaceutically acceptable salt thereof: in, K1, K2, and K3 are heteroalkyl, heterocycloalkyl, or heteroaryl groups containing heteroatoms O, S, P, or N; K1, K2, and K3 can be the same or different; where K1, K2, and K3 are heteroalkyl means that K1, K2, and K3 are composed of several R groups. k It is formed by linking heteroatoms or heteroatom-containing groups, wherein R k Independently and arbitrarily selected from substituted or unsubstituted alkyl groups; preferably, R k Selected from substituted or unsubstituted methyl, substituted or unsubstituted ethyl, substituted or unsubstituted propyl; wherein K1, K2, and K3 are heteroalkylcycloalkyl groups, meaning that K1, K2, and K3 are 4-7 member substituted or unsubstituted heterocycloalkyl groups containing heteroatoms O, S, P, or N, preferably tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydropyrrolyl, or piperazineyl; K1, K2, and K3 are heteroaryl groups, meaning that K1, K2, and K3 are substituted or unsubstituted 4-7 member heteroaryl groups containing heteroatoms O, S, P, or N, preferably furanyl, thiophenyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl, pyridinyl, pyridinyl, isoxazolyl, diazolyl, pyrazolyl, triazolyl, tetrazolyl, isothiazolyl, or thiadiazolyl. The N atom in Formula I is connected to H or a substituent.

2. The cage-like chelate according to claim 1, characterized in that, When the substituent of the N atom is a chelating arm of a heteroatom group with a lone pair of electrons; the chelating arm has the structural formula JQ, where Q is a heteroatom group; preferably, J is a substituted or unsubstituted, saturated or unsaturated ether, ester, ketone, amide or C0-C5 alkyl chain. When the substituent of the N atom is a target molecule linker arm with an active group AG capable of linking to the target molecule, the structural formula of the target molecule linker arm is L-AG, where AG is an active group capable of linking to the target molecule; preferably, AG is independently and arbitrarily selected from -X(Br, Cl, I, F), -OH, -COOH, -SH, -CO-N(CH3)OH, -H2PO3, H2PO4, -H2PO2, sulfonic acid, sulfinic acid, sulfone, sulfoxide, -OCH2COOH, -NH2, -NHR, -OCH2CH2NH2; -N3, -acetylene, -piperazine, piperidine, tetrahydropyrrole, -NCS, -NHS, -boronic acid (ester), -CHO, -COR. Preferably, L is independently and arbitrarily selected from linear or branched, saturated or unsaturated alkyl chains, PEG, amino acids, polypeptides, piperazine, triazole, esters, sugars, ethers, ureas, guanidines, sulfonic acids, sulfinic acids, nucleic acids, amides, sulfones, sulfoxides, and their derivatives. In some specific technical solutions, L is independently and arbitrarily selected. Or the substituent structure of the N atom is L. N -NAG,L N Similar to the definition of L above, NAG is an alkyl group or H.

3. The cage-like chelate according to claim 1, characterized in that, J also includes a group AG1 that can be linked to the target molecule. AG1 has the same definition as AG in claim 1. AG1 and AG can be the same or different.

4. The cage-like chelate according to claim 1, characterized in that, The cage-like chelate has any of the following preferred characteristics. (1) K1, K2, and K3 are independently and arbitrarily selected from: (2) Group Q is selected from: -H2PO3、-H2PO4、-H2PO2、 Where R, R', and R” represent H, alkyl, hydroxyl, fatty acid group, amino, acyl, heteroalkyl, and heteroaryl groups; (3) The cage-like chelate is arbitrarily selected from:

5. The targeted cage chelating ligand formed by coupling the cage chelate C according to any one of claims 1 to 4 with one or more targeting molecules TM.

6. The targeted cage-like chelating ligand according to claim 5, characterized in that, TM is independently and arbitrarily selected from biological macromolecules, and may also be selected from drugs or small molecule compounds. The target molecules include, but are not limited to, one or more of the following: antibodies, bispecific antibodies, triple antibodies, nanobodies, proteins, peptides, polymers, carbohydrates, nucleotides, oligonucleotides, oligosaccharides, vitamins, liposomes, cells, viruses, nanomaterials, small molecule drugs or fragments or derivatives. More preferably, the TM is selected from any one or more of the following target molecules; More preferably, the targeted cage-like chelating ligand is:

7. The salt formed by the cage-like chelate of claim 1 and an inorganic or organic acid.

8. The method for preparing the cage-like chelate according to claim 1, characterized in that, This method is prepared using 2-hydroxy-1,3,5-benzenetrialdehyde and NH2—K1—NH2, NH2—K2—NH2, and NH2—K3—NH2 as raw materials, wherein K1, K2, and K3 are defined as described in any one of claims 1-4.

9. A metal complex formed from the cage-like chelate according to any one of claims 1 to 4, or the targeted cage-like chelating ligand according to claim 5 or 6.

10. The use of the chelating ligand compound according to any one of claims 1 to 4, the targeted chelating ligand compound according to claim 5 or 6, the organic or inorganic salt derivative according to claim 7, and the metal complex according to claim 9 in the preparation of radiopharmaceuticals, iron removal agents, and drugs for treating poisoning caused by heavy metals.

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