Chelating ligand compounds, targeted chelating ligands and their complexes and applications

Abstract

This invention relates to the field of chelating drug research, specifically to chelating ligand compounds, targeted chelating ligands and their complexes and applications. Compared with the prior art, the targeted molecule linking sites and linking methods disclosed in this invention have better targeting and in vivo stability. At the same time, the design of the targeted molecule linking sites in this invention can improve in vivo metabolism and distribution, thereby improving the druggability of preferred compounds and providing a better foundation for the clinical application of therapeutic radiopharmaceuticals.

Description

Technical Field

[0001] This invention 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. Technical Background

[0002] Compounds with chelating properties can undergo chelation reactions with metals and radioactive elements to form drugs with diagnostic and therapeutic functions, thus realizing their value in the clinical diagnosis and treatment of diseases. These compounds are called chelating ligand compounds.

[0003] For those skilled in the art, given the shortcomings of chelating ligand compounds in research, production, and clinical applications, the aim of designing and researching such molecules is to target structures capable of chelating a variety of metals and radioactive elements, with mild chelation reaction conditions, stable products, and good bio-metabolic properties.

[0004] Currently, chelating ligand compounds widely used in the pharmaceutical field include DOTA and DTPA. However, these compounds have several drawbacks: First, they can only chelate one or a few nuclides, making it impossible for a single molecular structure to achieve multiple functions such as diagnosis and treatment. Second, the chelation reaction conditions are demanding, often requiring heating to accelerate the reaction, thus limiting their use for modifying and developing heat-sensitive compounds and proteins. Third, their poor molecular stability leads to the decomposition of the chelated compound itself. While this can be mitigated through formulation development, it doesn't fundamentally solve the problem, increasing the difficulty of drug development and the risks associated with its use. Fourth, the poor biological stability and metabolic properties of chelating ligand compounds make them prone to dechelation, resulting in high in vivo toxicity and significant side effects.

[0005] Based on this, the inventors of this invention hope to design and synthesize chelating ligand compounds with better chelating and targeting properties, thereby providing more and better selectivity for clinical targeted drugs. Summary of the Invention

[0006] One of the objectives of this invention is to provide a novel chelating ligand compound. By constructing a flexible macrocyclic ligand framework adapted to a broad spectrum of atomic radius nuclides, the coordination ability and compatibility range are greatly improved, forming a stable metal complex that makes it difficult for radioactive metal nuclides to escape. Simultaneously, because it provides a large flexible chelating space, heating is not required during the chelation process, making it more compatible with various target probes.

[0007] The second objective of this invention is to provide multiple sites for the binding of target molecules, thereby enabling drugs to have better targeting and providing a good platform for multi-target drugs.

[0008] The third objective of this invention is that the complexes prepared based on the chelating ligand compound and the targeted chelating ligand compound formed therefrom, when used as radiopharmaceuticals as contrast agents for diagnosis or when utilizing the α and β rays and Auger electrons generated by the radionuclide for therapeutic purposes, exhibit better in vivo stability.

[0009] The fourth objective of this invention is that the drugs formed based on the chelating ligand compound and the targeted chelating ligand compound it forms have high stability, strong targeting, and better biological properties such as in vivo distribution and metabolism, thus greatly improving drug-likeness.

[0010] To achieve the above-mentioned objective, the present invention provides a chelating ligand compound, wherein the chelating ligand compound is a compound of general formula I or a pharmaceutically acceptable salt thereof:

[0011]

[0012] At least one of A1 and A2 contains an active group AG that can be linked to the target molecule; when both A1 and A2 contain an active group AG that can be linked to the target molecule, the active groups can be the same or different.

[0013] When A1 contains an active group AG that can connect with the target molecule, A1 is the target molecule linker arm;

[0014] When A1 does not contain an active group AG that can connect with the target molecule, A1 = R1;

[0015] When A2 contains an active group AG that can connect with the target molecule, A2 is the target molecule linker arm;

[0016] When A2 does not contain an active group AG that can connect with the target molecule, A2 = R2;

[0017] The target molecule linker arm connects a chelating compound at one end and a target molecule at the other end, forming a complex such as "chelating compound-linker-target molecule". R1 and R2 refer to groups that do not have the ability to link to the target molecule. They can be completely different from the target molecule linker arm, or they can be formed by replacing the active groups on the target molecule linker arm with inactive groups. R1 and R2 do not have the ability to further link to the target molecule, but they can be groups with other functionalities, or of course, they can be non-functional.

[0018] Preferably, when A1 is a target molecule linker arm, A1 is L1-AG1, and when A2 is a target molecule linker arm, A2 is L2-AG2; wherein AG1 and AG2 are 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,

[0019] The L1 and L2 are 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.

[0020] In some specific technical solutions, L1 and L2 are independently and arbitrarily selected.

[0021] Preferably, when A1 and A2 are non-targeted connecting arms, A1 and A2 are L N -NAG, L N Similar to the definitions of L1 and L2 mentioned above, NAG is an alkyl group or H;

[0022] K1 and K2 are heteroalkyl, heterocyclic alkyl, or heteroaryl groups containing heteroatoms O, S, P, or N; K1 and K2 can be the same or different; where K1 and K2 are heteroalkyl means that K1 and K2 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 and K2 are heterocyclic groups, meaning that K1 and K2 are 4-7 membered substituted or unsubstituted heterocyclic alkyl groups containing heteroatoms O, S, P, or N, preferably tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydropyrrolyl, or piperazineyl; K1 and K2 are heteroaryl groups, meaning that K1 and K2 are 4-7 membered heteroaryl groups containing heteroatoms O, S, P, or N, preferably furanyl, thiophenyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl, pyridinyl, pyridinyl, pyrazine, isoxazolyl, diazolyl, pyrazolyl, triazolyl, tetrazolyl, isothiazolyl, or thiadiazolyl.

[0023] In general formula I, the N atom is connected to H or a substituent. When a substituent is connected, the substituent can be a chelating arm of a heteroatom group Q with a lone pair of electrons or a target molecule linking arm L3 with an active group AG3 that can link to the target molecule. The N atom in general formula I includes both the N in the main ring structure and the N atom in K1 or K2.

[0024] The chelating arm has the structural formula JQ, where J is a substituted or unsubstituted, saturated or unsaturated ether, ester, ketone, amide or C0-C5 alkyl chain;

[0025] The structural formula of the target molecule linker arm L3 is L3-AG3. AG3 has the same definition as AG1 and AG2 mentioned above. AG1, AG2 and AG3 can be the same or different. L3 has the same definition as L1 and L2. L3, L1 and L2 can be the same or different.

[0026] The active group AG on the target molecule linker arm L is a crucial part for connection with the target molecule. This invention provides multiple target molecule linking sites, which lays the foundation for dual-antibody and triple-antibody formulations. Depending on the specific target molecule or target group requiring connection, different active groups AG can be linked to different linking sites to achieve connection with the target molecule. Of course, connection with the target molecule can be achieved through covalent bonds, coordinate bonds, or through hydrophobic bonds and electrostatic interactions.

[0027] Preferably, the group J further includes an active group AG4 that can be linked to the target molecule. AG4 has the same definition as AG1, AG2, and AG3 mentioned above. AG1, AG2, AG3, and AG4 can be the same or different.

[0028] Group J is a component of the chelating arm. When it has group AG4 on it (mainly on the side chain), these active groups can connect with the target molecule.

[0029] Therefore, under this technical solution, the present invention provides more target site connection points. As mentioned above, it can be understood that the target sites connected to these active groups can be the same or different.

[0030] In a preferred embodiment of this invention, the chelating ligand compound is selected from the following structures:

[0031]

[0032] Wherein, B1, B2, B3, and B4 are H or substituents. When a substituent is attached, the substituent can be a chelating arm of a heteroatom group Q with a lone pair of electrons, or a target molecule linking arm L3 with an active group AG3 that can link with the target molecule, or other substituents. The definitions of Q and L3 are the same as those mentioned above. X is a halogen (Br, Cl, I, F).

[0033] Furthermore, the preferred structures of the chelating ligand compounds are provided in this invention as follows:

[0034] (1) K1 and K2 are independently and arbitrarily selected from:

[0035]

[0036]

[0037] (2) Group Q is selected from:

[0038]

[0039]

[0040]

[0041] Wherein R, R', and R" are H, alkyl, hydroxyl, fatty acid group, amino, acyl, heteroalkyl, or heteroaryl; (3) the chelating ligand compound is arbitrarily selected from:

[0042]

[0043]

[0044]

[0045]

[0046]

[0047]

[0048]

[0049]

[0050]

[0051]

[0052] Further preferably, this invention also discloses a targeted chelating ligand compound formed by coupling a chelating ligand compound C with one or more target molecules TM. The active group in the chelating ligand compound is connected to the target molecule via covalent bonds, coordinate bonds, hydrophobic bonds, electrostatic interactions, or other means. Different target molecules can be selected for different target proteins; these target molecules, by binding to the active group, endow the chelating ligand compound with specific targeting properties.

[0053] For example, in this invention, preferably, 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 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.

[0054] More preferably, the TM is selected from any one or more of the following target molecules;

[0055]

[0056] Based on this, we further disclose the preferred targeted chelating ligand compound as follows:

[0057]

[0058]

[0059]

[0060]

[0061] Furthermore, the present invention also discloses the salt formed by the chelating ligand compound and an inorganic or organic acid. It should be understood that the salt referred to here is a pharmaceutically acceptable salt.

[0062] Furthermore, the present invention also discloses a method for preparing the chelating ligand compound, wherein the method comprises 2-hydroxy-isophthalaldehyde with or without 5-substituted or unsubstituted 5-position hydroxy-isophthalaldehyde and NH2—K1—NH 2， It is prepared from NH2—K2—NH2, where K1 and K2 are defined as described above.

[0063] The present invention also discloses metal complexes, which are formed by complexing the aforementioned chelating ligand compounds or couplings with metal elements.

[0064] Furthermore, the present invention also discloses the use of the aforementioned chelating ligand compounds, the derivatives, and the coupling compounds in the preparation of radiopharmaceuticals, iron removal agents, and drugs for treating heavy metal poisoning, as well as the use of metal complexes in the preparation of radiopharmaceuticals.

[0065] This invention 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., wherein the chelating ligand compounds disclosed in this invention have particularly significant advantages in chelating with lanthanides and actinides, such as those used in the embodiments of this invention. 68 Ga、 177 Lu、 225 Ac、 223 Ra、 89 Zr、 227 Th、 212 Pb.

[0066] This invention provides a class of chelating ligand compounds with a ligand framework adapted to a broad spectrum of atomic radius nuclides. This allows for chelation with radioactive metal nuclides without heating, and the strong coordination during chelation results in highly stable metal complexes that are difficult for radioactive metal nuclides to escape. Furthermore, this invention utilizes different functional groups with target molecule binding capabilities to replace different numbers and positions of chelating groups, forming chelating ligand compounds with varying numbers and positions. This enhances the binding affinity between the target molecule and the binding site. In summary, the chelating ligand compounds disclosed in this invention, as well as the radiopharmaceuticals prepared from compounds formed by binding these chelating ligand compounds to target molecules and / or complexes formed by complexing with metal nuclides, exhibit better targeting and stability when used as contrast agents for diagnosis or for therapeutic purposes utilizing α and β rays and Auger electrons generated by nuclides. We have found that the drugs formed from these chelating ligand compounds and their targeted chelating ligand compounds exhibit high stability, strong targeting, and better biological characteristics such as in vivo distribution and metabolism, thus possessing better drug-like properties. Attached Figure Description

[0067] Figure 1 H is the compound R1 in Example 1-1 1 NMR spectrum.

[0068] Figure 2 H is the compound R2-2 in Example 2-1 1 NMR spectrum.

[0069] Figure 3 H is the compound 28 in Examples 1-13 1 NMR spectrum.

[0070] Figure 4 H is the compound 36 in Examples 1-15 1 NMR spectrum.

[0071] Figure 5 H is the compound R22-1 in Examples 2-8 1 NMR spectrum.

[0072] Figure 6 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R1 and the metal nuclide Ga-68 in Example 3-1. The Ga ion should be located at the origin in the diagram.

[0073] Figure 7 The results are HPLC analysis of the chelation reaction product of compound R1 and metal nuclide Ga-68 in Example 3-1.

[0074] Figure 8The image shows the iTLC analysis results of the chelation reaction product of compound R1 and the metal nuclide Zr-89 in Example 3-1. The Zr ion should be located at the origin in the image.

[0075] Figure 9 The image shows the iTLC analysis results of the chelation reaction product of compound R1 and the metal nuclide Lu-177 in Example 3-1. The Lu ion should be located at the leading edge in the image.

[0076] Figure 10 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R2-2 and the metal nuclide Ga-68 in Example 3-2. The Ga ion should be located at the origin in the figure.

[0077] Figure 11 The image shows the HPLC analysis results of the chelation reaction between compound R2-2 and the metal nuclide Ga-68 in Example 3-2.

[0078] Figure 12 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R3-2 and the metal nuclide Ga-68 in Example 3-3. The Ga ion should be located at the origin in the figure.

[0079] Figure 13 The image shows the HPLC analysis results of the chelation reaction between compound R3-2 and the metal nuclide Ga-68 in Examples 3-3.

[0080] Figure 14 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R4-2 and the metal nuclide Ga-68 in Examples 3-4. The Ga ion should be located at the origin in the diagram.

[0081] Figure 15 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R5-4 and the metal nuclide Ga-68 in Examples 3-5. The Ga ion should be located at the origin in the figure.

[0082] Figure 16 The HPLC analysis results of the chelation reaction between compound R5-4 and the metal nuclide Ga-68 in Examples 3-5 are shown.

[0083] Figure 17 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R6-2 and the metal nuclide Ga-68 in Examples 3-6. The Ga ion should be located at the origin in the figure.

[0084] Figure 18 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R7-2 and the metal nuclide Ga-68 in Examples 3-7. The Ga ion should be located at the origin in the figure.

[0085] Figure 19This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R8-2 and the metal nuclide Ga-68 in Examples 3-8. The Ga ion should be located at the origin in the figure.

[0086] Figure 20 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R8 and the metal nuclide Ga-68 in Examples 3-9. The Ga ion should be located at the origin in the diagram.

[0087] Figure 21 The image shows the iTLC analysis results of the chelation reaction between compound R8 and the metal nuclide Zr-89 in Examples 3-9. The Zr ion should be located at the leading edge in the image.

[0088] Figure 22 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R9 and the metal nuclide Ga-68 in Examples 3-10. The Ga ion should be located at the origin in the figure.

[0089] Figure 23 The image shows the iTLC analysis results of the chelation reaction between compound R9 and the metal nuclide Zr-89 in Examples 3-10. The Zr ion should be located at the leading edge in the image.

[0090] Figure 24 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R10 and the metal nuclide Ga-68 in Examples 3-11. The Ga ion should be located at the origin in the diagram.

[0091] Figure 25 The image shows the iTLC analysis results of the chelation reaction between compound R10 and the metal nuclide Zr-89 in Examples 3-11. The Zr ions should be located at the leading edge in the image.

[0092] Figure 26 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R11 and the metal nuclide Ga-68 in Examples 3-12. The Ga ion should be located at the origin in the figure.

[0093] Figure 27 The image shows the iTLC analysis results of the chelation reaction between compound R11 and the metal nuclide Zr-89 in Examples 3-12. The Zr ion should be located at the leading edge in the image.

[0094] Figure 28 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R12 and the metal nuclide Ga-68 in Examples 3-13. The Ga ion should be located at the origin in the figure.

[0095] Figure 29 The image shows the iTLC analysis results of the chelation reaction between compound R12 and the metal nuclide Zr-89 in Examples 3-13. The Zr ion should be located at the leading edge in the image.

[0096] Figure 30 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R13 and the metal nuclide Ga-68 in Examples 3-14. The Ga ion should be located at the origin in the diagram.

[0097] Figure 31 The image shows the iTLC analysis results of the chelation reaction between compound R13 and the metal nuclide Zr-89 in Examples 3-14. The Zr ions should be located at the leading edge in the image.

[0098] Figure 32 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R15 and the metal nuclide Ga-68 in Examples 3-15. The Ga ion should be located at the origin in the figure.

[0099] Figure 33 The image shows the iTLC analysis results of the chelation reaction between compound R15 and the metal nuclide Zr-89 in Examples 3-15. The Zr ions should be located at the leading edge in the image.

[0100] Figure 34 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R16 and the metal nuclide Ga-68 in Examples 3-16. The Ga ion should be located at the origin in the figure.

[0101] Figure 35 The image shows the iTLC analysis results of the chelation reaction between compound R16 and the metal nuclide Lu-177 in Examples 3-16. The Lu ion should be located at the leading edge in the image.

[0102] Figure 36 The image shows the iTLC analysis results of the chelation reaction between compound R16 and the metal nuclide Zr-89 in Examples 3-16. The Zr ions should be located at the leading edge in the image.

[0103] Figure 37 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R17 and the metal nuclide Ga-68 in Examples 3-17. The Ga ion should be located at the origin in the diagram.

[0104] Figure 38 The image shows the iTLC analysis results of the chelation reaction between compound R17 and the metal nuclide Lu-177 in Examples 3-17. The Lu ion should be located at the leading edge in the image.

[0105] Figure 39 The image shows the iTLC analysis results of the chelation reaction between compound R17 and the metal nuclide Zr-89 in Examples 3-17. The Zr ion should be located at the leading edge in the image.

[0106] Figure 40 This is a schematic diagram of the iTLC analysis results of the chelation reaction between compound R18 and the metal nuclide Ga-68 in Examples 3-18. The Ga ion should be located at the origin in the figure.

[0107] Figure 41 The image shows the iTLC analysis results of the chelation reaction between compound R18 and the metal nuclide Zr-89 in Examples 3-18. The Zr ion should be located at the leading edge in the image.

[0108] Figure 42 The image shows the iTLC analysis results of the chelation reaction between compound R22-1 and the metal nuclide Zr-89 in Examples 3-19. The Zr ion should be located at the leading edge in the image.

[0109] Figure 43 The image shows the iTLC analysis results of the chelation reaction between the coupling protein R21-KT1 and the metal nuclide Zr-89 in Examples 3-20. The Zr ion should be located at the leading edge in the image.

[0110] Figure 44 The image shows the iTLC analysis results of the chelation reaction between the coupling protein R21-KT2 and the metal nuclide Zr-89 in Examples 3-21. The Zr ion should be located at the leading edge in the image.

[0111] Figure 45 The image shows the iTLC analysis results of the chelation reaction between the coupling protein R21-KT3 and the metal nuclide Zr-89 in Examples 3-22. The Zr ion should be located at the leading edge in the image.

[0112] Figure 46 The image shows the iTLC analysis results of the chelation reaction between the coupling protein R23-KT and the metal nuclide Zr-89 in Examples 3-23. The Zr ion should be located at the leading edge in the image.

[0113] Figure 47 The HPLC analysis results of the chelation reaction between the coupling protein R2-KT and the metal nuclide Ga-68 in Examples 3-24 are shown.

[0114] Figure 48 In Example 4-1 68 PET imaging results of Ga-R2-2 in a mouse model bearing U87-MG tumors.

[0115] Figure 49 In Example 4-2 68 PET imaging results of Ga-R3-2 in a mouse model bearing U87-MG tumor.

[0116] Figure 50 In Example 4-3 68 PET imaging results of Ga-R5-4 in a mouse model bearing U87-MG tumor.

[0117] Figure 51 In Example 4-4 89PET imaging results of Zr-R22-1 in a mouse model bearing U87-MG tumor.

[0118] Figure 52 For Examples 4-5 89 PET imaging results of Zr-R21-KT1 in a mouse model bearing U87-MG tumor.

[0119] Figure 53 For Examples 4-6 89 PET imaging results of Zr-R21-KT2 in a mouse model bearing U87-MG tumor.

[0120] Figure 54 For Examples 4-7 89 PET imaging results of Zr-R21-KT3 in a mouse model bearing U87-MG tumor. Detailed Implementation

[0121] To better understand the present invention, we will further elaborate on the present invention below with reference to specific embodiments.

[0122] 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.

[0123] Example Group 1

[0124] This invention is prepared from 2-hydroxy-1,3,5-benzenetrialdehyde and NH2-K (K can be different K1 and K2)-NH2 as raw materials. Specifically, it involves reacting 2-hydroxy-1,3,5-benzenetrialdehyde with an equivalent amount of NH2-K (K1 = K2)-NH2, or reacting 2 equivalents of 2-hydroxy-1,3,5-benzenetrialdehyde with 1 equivalent of NH2-K. 1- NH2 and 1 equivalent of NH2-K 2- The mixture of NH2 reacts to give a cyclic Schiff base, which is then reduced to a macrocyclic compound with sodium borohydride. The macrocyclic compound then reacts with a chelating arm intermediate containing a chelating group Q, and finally hydrolyzes to generate the target compound.

[0125] Below, we list the synthesis of some of the compounds of this invention.

[0126] Example 1-1

[0127]

[0128] 2-Hydroxybenzyl-1,3,5-tricarboxaldehyde (1 g, 5.61 mmol) was dissolved in 120 mL of anhydrous methanol solution and heated to 70 °C. At 70 °C, 40 mL of anhydrous methanol solution of 2,2′-(ethylenedioxy)bis(ethylamine) (0.832 g, 5.61 mmol) was added dropwise slowly, and the reaction was continued for 2 hours. No further treatment was performed to obtain a methanol solution of compound 1.

[0129] Under a nitrogen atmosphere, sodium borohydride (0.637 g, 16.8 mmol) was added to a methanol solution of compound 1 with stirring. The reaction was stirred for 1 hour, and the system color lightened. The solvent was removed under reduced pressure, and the residue was washed with dichloromethane / methanol / water extract and dried over anhydrous sodium sulfate. The solution was concentrated under reduced pressure to give 1.2 g of a pale yellow, foamy solid, compound 2.

[0130] LC-MS: 593.4(M+1), 297.2(M / 2+1)

[0131] Compound 2 (500 mg, 0.84 mmol), tert-butyl bromoacetate (658 mg, 3.4 mmol), DIEA (653 mg, 5.1 mmol), and acetonitrile (20 mL) were mixed and reacted at room temperature for 6 hours. The mixture was concentrated to dryness, extracted with ethyl acetate / water, dried and concentrated to obtain the crude product, and purified to obtain compound 3 (581 mg, yield 65.7%).

[0132] LC-MS: 1049.6(M+1), 525.3(M / 2+1).

[0133] Compound 3 (300 mg, 0.286 mmol) was dissolved in 4 M hydrochloric acid (3 mL) and reacted at 45 °C for 0.5 h. The solution was concentrated to dryness under reduced pressure to prepare and purify the target compound R1 (212 mg). See the detailed chromatogram for reference. Figure 1 .

[0134] LC-MS: 825.4(M+1), 413.2(M / 2+1)

[0135] 1 H-NMR (400M, D2O): δ3.49 (s, 8H), 3.68 (s, 8H), 3.87 (s, 16H), 4.45-4.50 (d, 12H), 7.40 (d, 4H)

[0136] Examples 1-2

[0137]

[0138] 2-Hydroxybenzaldehyde (0.3 g, 1.68 mmol) was dissolved in 45 mL of methanol, and a 15 mL methanol solution of bis(3-aminopropyl) ether (0.22 g, 1.68 mmol) was added dropwise. After the addition was complete, the reaction was continued for 1 hour. Then, sodium borohydride (382 mg, 10.1 mmol) was added in portions, and after reacting for 1 hour, the solution was concentrated and purified to give compound 4 (293 mg, 62%).

[0139] LC-MS: 561.4 (M+1)

[0140] Compound 4 (293 mg, 0.522 mmol), tert-butyl bromoacetate (458 mg, 2.35 mmol), DIEA (539 mg, 4.177 mmol), and 30 mL of acetonitrile were mixed and reacted at room temperature for 2 hours. The mixture was concentrated to dryness, extracted with ethyl acetate / water, dried and concentrated to obtain the crude product, and purified to obtain compound 5 (480 mg).

[0141] LC-MS: 509.3(M / 2+1), 339.9(M / 3+1)

[0142] Compound 5 (50 mg, 0.049 mmol) was dissolved in 4 M hydrochloric acid (1.5 mL) and reacted at 45 °C for 0.5 h. The solution was concentrated under reduced pressure to dryness, and the purified compound R8 (30 mg) was obtained.

[0143] LC-MS: 793.4(M+1), 397.2(M / 2+1)

[0144] H-NMR (300M, D2O): δ1.75-1.93 (br, 8H), 3.10 (s, 8H), 3.39-3.54 (br, 8H), 3.95-4.11 (m, 12H), 4.54 (br, 4H), 7.44 (s, 4H).

[0145] Examples 1-3

[0146] Following the method of Examples 1-2, only the bis(3-aminopropyl) ether was replaced with 2,2'-oxobis(ethylamine) to obtain compound R9.

[0147]

[0148] in:

[0149] Compound 6:

[0150] LC-MS: 505.3 (M+1)

[0151] Compound R9:

[0152] LC-MS: 737.3(M+1), 369.2(M / 2+1)

[0153] H-NMR (300M, D2O): δ3.25-3.28(m, 8H), 3.63-3.66(m, 8H), 3.84-3.87(m, 8H), 4.26-4.39(m, 12H), 7.44(s, 4H)

[0154] Examples 1-4

[0155] Following the method of Examples 1-2, only the bis(3-aminopropyl) ether was replaced with diethylenetriamine to obtain compound R10.

[0156]

[0157] in:

[0158] Compound 8:

[0159] LC-MS: 503.3 (M+1)

[0160] Compound R10:

[0161] LC-MS: 851.4(M+1), 426.2(M / 2+1)

[0162] H-NMR (300M, D2O): δ3.25-3.28(m, 8H), 3.63-3.66(m, 8H), 3.84-3.87(m, 8H), 4.26-4.39(m, 12H), 7.44(s, 4H)

[0163] Examples 1-5

[0164] Following the methods of Examples 1-3, only the bis(3-aminopropyl) ether was replaced with aminoethyl sulfide to obtain compound R11.

[0165]

[0166] in:

[0167] Compound R11:

[0168] LC-MS: 769.3(M+1), 385.2(M / 2+1)

[0169] H-NMR (300M, D2O): δ2.59 (br, 8H), 3.45 (br, 8H), 3.96 (br, 8H), 4.62 (br, 12H), 7.36 (s, 4H)

[0170] Examples 1-6

[0171]

[0172] 59.2 mg (0.1 mmol) of intermediate 2 was added to 2 ml of acetonitrile, followed by DIEA (129 mg, 1 mmol). Compound V 132 mg (0.45 mmol) was added with stirring. After reacting for 12 hours, the crude product of compound 12 was obtained and purified to 121 mg.

[0173] LC-MS: 708.3 (M / 2+1)

[0174] 50 mg of compound 12 was dissolved in 1 ml of a mixture of acetic acid and concentrated hydrochloric acid (1:1), reacted at 55 °C for 6 hours, and then concentrated to prepare purified compound R12.

[0175] Compound R12:

[0176] LC-MS: 543.2(M / 2+1), 362.5(M / 3+1)

[0177] H-NMR (300M, D2O): δ3.69(s, 8H), 3.79(s, 8H), 4.03(s, 8H), 4.58(s, 20H ), 6.61-6.63(d, 4H), 6.71-6.72(d, 4H), 7.10(s, 4H), 7.44-7.48(t, 4H). Tool

[0178] Examples 1-7

[0179]

[0180] Referring to Examples 1-5, compound V was replaced with compound VI to obtain compound R13.

[0181] Compound 13:

[0182] LC-MS: 651.3 (M / 2+1)

[0183] Compound R13:

[0184] LC-MS: 471.2 (M / 2+1)

[0185] H-NMR (300M, D2O): δ3.66-3.69 (m, 20H), 3.78 (s, 8H), 4.00 (s, 8H), 4.56 (s, 20H), 7.44-7.48 (t, 4H).

[0186] Examples 1-8

[0187]

[0188] Referring to Examples 1-5, compound 14 was prepared by replacing compound V with compound VII, and then further hydrolyzed with 4M hydrochloric acid to prepare lyophilized compound R14.

[0189] Compound R14:

[0190] LC-MS: 485.2 (M / 2+1)

[0191] Examples 1-9

[0192]

[0193] Referring to Examples 1-5, compound V was replaced with compound VIII to prepare compound 15, which was further hydrolyzed with 4M hydrochloric acid to prepare lyophilized compound R15.

[0194] Compound R15:

[0195] LC-MS: 715.2(M / 2+1), 477.2(M / 3+1)

[0196] Examples 1-10

[0197]

[0198] 118.5 mg (0.2 mmol) of intermediate 2 was added to 10 mL of dichloromethane, followed by TEA (258 g, 2 mmol). A mixed solution of 44 mg (0.18 mmol) of 2-(tert-butoxycarbonyloxyimino)-2-phenylacetonitrile and 5 mL of dichloromethane was added dropwise with stirring. After reacting for 12 hours, the crude product of compound 16 was obtained, and 69 mg of yellow oil was obtained after purification.

[0199] LC-MS: 693.4 (M+1)

[0200] 69 mg (0.1 mmol) of compound 16 was dissolved in 5 mL of acetonitrile, followed by the addition of DIEA (77 mg, 0.6 mmol). Compound V (147 mg, 0.5 mmol) was then added with stirring. After reacting for 12 hours, the crude product of compound 17 was obtained, which was purified to give 95 mg of a yellow solid. LC-MS: 616.8 (M / 2+1)

[0201] 95 mg (0.071 mmol) of compound 17 was dissolved in 2 ml of dichloromethane, followed by the addition of TFA (0.5 ml). The mixture was stirred at room temperature and reacted for 2 hours to obtain crude compound 18, which was then purified to give 80 mg of a yellow oil.

[0202] LC-MS: 666.8 (M / 2+1)

[0203] 80 mg (0.065 mmol) of compound 17 was dissolved in 2 ml of acetonitrile, followed by the addition of DIEA (25 mg, 0.195 mmol). Then, 25 mg (0.13 mmol) of tert-butyl bromoacetate was added with stirring. After reacting for 12 hours, the crude product of compound 19 was obtained and purified to give 70 mg of yellow oil.

[0204] LC-MS: 673.8 (M / 2+1)

[0205] 70 mg (0.052 mmol) of compound 19 was dissolved in 2 ml of 4 M hydrochloric acid, and the mixture was heated to 45 °C and reacted for 1 hour to prepare purified compound R16 (32 mg, 60.4%) as a white solid.

[0206] LC-MS: 510.7 (M / 2+1)

[0207] Examples 1-11

[0208]

[0209] 1,7-bis-BOC-1,4,7-triazaheptane (303 mg, 1 mmol) was dissolved in acetonitrile, and DIEA (129 mg, 1 mmol) and tert-butyl bromoacetate (195 mg, 1 mmol) were added. The mixture was stirred for 2 hours. After extraction and washing, the crude product was purified by column chromatography to give compound 20 (358 mg).

[0210] LC-MS: 474.2(M+23), 396.2(M+1-56)

[0211] Compound 20 (225 mg, 0.5 mmol) was dissolved in 20 mL of dichloromethane, and 1 M dioxane hydrochloride solution (1 mL) was added dropwise. After 30 minutes, the solution was diluted with 20 mL of dichloromethane and concentrated to give 163 mg of crude compound 21 hydrochloride. LC-MS: 252.2 (M+1)

[0212] Dissolve 2-hydroxybenzene-1,3,5-tricarboxaldehyde (80.2 mg, 0.45 mmol) in 30 mL of methanol and heat to 60 °C. Add dropwise 15 mL of methanol solution of compound 21 (163 mg, 0.5 mmol) and DIEA (64.5 mg, 0.5 mmol). After the addition is complete, continue the reaction for 2 hours.

[0213] After the temperature was raised to 45°C, sodium borohydride (76 mg, 2 mmol) was added to the reaction solution with stirring, and the reaction was stirred for 1 hour. The solvent was removed under reduced pressure, and the residue was extracted and washed to give crude compound 22 (101 mg).

[0214] LC-MS: 400.2 (M / 2+1)

[0215] Compound 22 crude product (101 mg, 0.13 mmol) was added to DMF, and DIEA (101 mg, 0.78 mmol) and compound V (172 mg, 0.59 mmol) were added while stirring. The mixture was stirred and reacted overnight. Ethyl acetate and water were added, and the mixture was extracted and washed to give 150 mg of crude product. After purification, 106 mg of compound 23 was obtained.

[0216] LC-MS: 826.4(M / 2+1), 551.3(M / 3+1)

[0217] Compound 23 (106 mg) was added to 2.5 ml of concentrated hydrochloric acid, reacted at 55 °C for 30 minutes, concentrated to dryness, and purified to obtain compound R17.

[0218] LC-MS: 556.2(M / 2+1), 371.2(M / 3+1)

[0219] Examples 1-12

[0220]

[0221] 2-Hydroxybenzaldehyde (89.1 mg, 0.5 mmol) was dissolved in 25 mL of methanol and heated to 45 °C. A 10 mL methanol solution of 2-(bis(2-aminoethyl)amino)ethylcarbamate (123 mg, 0.5 mmol) was added dropwise, and the reaction was continued for 1 hour. Compound 24 was obtained. Sodium borohydride (76 mg, 2 mmol) was added to the reaction system, and the reaction was continued for 2 hours, during which the color of the reaction system lightened. Dichloromethane was added and the mixture was extracted and concentrated with water to obtain compound 25 (197 mg, crude product). No further processing was performed, and the mixture was directly proceeded to the next reaction. LC-MS: 345.3 (M-100 / 2+1), 295.3 (M-200 / 2+1)

[0222] Compound 25 was dissolved in acetonitrile (5 ml), and tert-butyl bromoacetate (244 mg, 1.25 mmol) and DIEA (258 mg, 2 mmol) were added. The mixture was heated to 45 °C and reacted for 5 hours. After cooling, water and ethyl acetate were added, and the mixture was extracted and concentrated to obtain crude compound 26. Further purification yielded 189 mg of a yellow foamy solid. LC-MS: 572.4 (M-100 / 2+1), 522.4 (M-200 / 2+1)

[0223] Compound 25 (50 mg) was added to 2.5 ml of 4 M hydrochloric acid, reacted at 45 °C for 45 minutes, concentrated to dryness, and purified to obtain compound R18.

[0224] LC-MS: 821.4(M+1), 411.2(M / 2+1)

[0225] Examples 1-13

[0226]

[0227] 2-Hydroxybenzyl-1,3,5-tricarboxaldehyde (178 mg, 1 mmol) was dissolved in 45 mL of anhydrous methanol and heated to 45 °C. A mixture of 15 mL of methanol containing 2-(bis(2-aminoethyl)amino)ethylcarbamate (123 mg, 0.5 mmol) and diethylenetriamine (51.6 mg, 0.5 mmol) was added dropwise. After the addition was complete, the reaction was continued for 1 hour. Precursor 8, compounds 24, and 27 were obtained. Sodium borohydride (152 mg, 4 mmol) was added to the reaction system, and the reaction was continued for 2 hours, during which the color of the reaction system lightened. Filtration was used to prepare compounds 8, 25, and 28, respectively, with compound 28 being 93 mg. See the chromatogram for details. Figure 3 .

[0228] LC-MS: 646.4(M+1), 323.7(M / 2+1)

[0229] 1 H-NMR (400M, D2O): δ1.39 (s, 9H), 2.72 (br, 2H), 2.96 (br, 4H), 3.18 (br, 6H), 3.43-3.48 (m, 8H), 4.32-4.47 (m, 8H), 4.57 (s, 4H), 7.44-7.48 (m, 4H)

[0230] The compound 28 prepared in the previous step was dissolved in acetonitrile (5 ml), and DIEA (186 mg, 1.44 mmol) and tert-butyl bromoacetate (168 mg, 0.86 mmol) were added respectively. The mixture was heated to 45 °C and reacted for 5 hours. After cooling, water and ethyl acetate were added to extract and concentrate the product to obtain crude compound 29. Further purification yielded 112 mg of yellow foamy solid.

[0231] LC-MS: 608.9(M / 2+1), 406.3(M / 3+1)

[0232] Compound 29 obtained in the previous step was dissolved in a mixed solution of TFA:TIPS:H2O and reacted at room temperature for 3 hours. After preliminary treatment, compound R19 was prepared and purified.

[0233] LC-MS: 936.5(M+1), 468.7(M / 2+1)

[0234] Examples 1-14

[0235]

[0236] 2-Hydroxybenzaldehyde (178 mg, 1 mmol) was dissolved in 45 mL of anhydrous methanol and heated to 45 °C. A mixture of N-Boc,2,2-diaminoethylenediamine (102 mg, 0.5 mmol) and diethylenetriamine (51.6 mg, 0.5 mmol) in 15 mL of methanol was added dropwise. After the addition was complete, the reaction was continued for 1 hour. Compound 30 was obtained. Sodium borohydride (152 mg, 4 mmol) was added to the reaction system, and the reaction was continued for 2 hours. The color of the reaction system lightened. Filtering yielded 95 mg of compound 31.

[0237] LC-MS: 603.4(M+1), 302.2(M / 2+1)

[0238] 1 H NMR (400MHz, D2O) δ1.47 (d, 9H), 3.21-3.53 (m, 16H), 4.32-4.39 (d, 12H), 7.42 (s, 4H).

[0239] Compound 31, prepared in the previous step, was dissolved in acetonitrile (5 ml), and potassium carbonate (218 mg, 1.58 mmol) and 2-bromo-N-((4-methoxybenzyl)oxy)-N-methylacetamide (272 mg, 0.946 mmol) were added, respectively. The mixture was heated to 45 °C and reacted for 5 hours. After cooling, water and ethyl acetate were added, and the mixture was extracted and concentrated to obtain crude compound 32. Further purification yielded 175 mg of a yellow foamy solid.

[0240] LC-MS: 820.0(M / 2+1), 547.0(M / 3+1)

[0241] Compound 32 (100 mg, 0.061 mmol) from the previous step was dissolved in DCM (8 ml), and trifluoroacetic acid (2 ml) was slowly added dropwise at low temperature. The reaction was carried out at room temperature for 20 minutes. After preliminary purification, 80 mg of yellow solid compound 33 was further prepared.

[0242] LC-MS: 769.9(M / 2+1), 513.6(M / 3+1)

[0243] Compound 33 was dissolved in acetonitrile (3 ml), and DIEA and tert-butyl bromoacetate were added separately. The mixture was heated to 45 °C and reacted for 2 hours. After cooling, water and ethyl acetate were added to extract and concentrate the product to obtain crude compound 34. Further purification yielded 45 mg of yellow foamy solid.

[0244] LC-MS: 827.0(M / 2+1), 551.6(M / 3+1)

[0245] The compound 34 obtained in the previous step was dissolved in a mixed solution of (TFA:DCM = 2:1) and reacted at room temperature for 5 hours. After preliminary treatment, the compound R20 was prepared and purified.

[0246] LC-MS: 996.5(M / 2+1), 498.7(M / 3+1)

[0247] Examples 1-15

[0248]

[0249] 2-Hydroxybenzaldehyde (45 mg, 0.25 mmol) and 2,6-dicarboxy-4-methylphenol (41 mg, 0.25 mmol) were dissolved in 25 mL of methanol and heated to 45 °C. A mixture of 15 mL of diethylenetriamine (51.6 mg, 0.5 mmol) in methanol was added dropwise. After the addition was complete, the reaction was continued for 1 hour. Compound 35 was obtained. Sodium borohydride (152 mg, 4 mmol) was added to the reaction system, and the reaction was continued for 1 hour. The color of the reaction system lightened. Filtration was used to prepare 32 mg of compound 36. See the specific spectrum for reference. Figure 4 .

[0250] LC-MS: 487.3 (M+1)

[0251] 1 H NMR (400MHz, D2O) δ2.27 (s, 3H), 3.34-3.48 (m, 18H), 4.33-4.38 (m, 8H), 7.29 (s, 2H), 7.46 (s, 2H).

[0252] Compound 36 prepared in the previous step was dissolved in acetonitrile, and potassium carbonate and 2-bromo-N-((4-methoxybenzyl)oxy)-N-methylacetamide were added respectively. The mixture was heated to 45°C and reacted for 8 hours. After cooling, water and ethyl acetate were added to extract and concentrate the product to obtain crude compound 37. Further purification yielded 52 mg of yellow foamy solid.

[0253] LC-MS: 865.4(M / 2+1), 577.3(M / 3+1)

[0254] Compound 34 obtained in the previous step was dissolved in a mixed solution of TFA:DCM = 2:1 and reacted at room temperature for 3 hours. After preliminary treatment, compound R21 was prepared and purified.

[0255] LC-MS: 505.3(M / 2+1), 337.2(M / 3+1)

[0256] Example Group 2

[0257] In this set of embodiments, we designed different targeting molecules for different target proteins and combined these targeting molecules with the chelating ligand compounds obtained in Example 1 to obtain targeted chelating ligand compounds with targeting properties.

[0258] Example 2-1

[0259] In this embodiment, we prepared compound R2 by reacting compound 3 with thionyl chloride to obtain compound R2, and then coupled compound R2 with compound W to obtain compound R2-1, which was further hydrolyzed with hydrochloric acid to obtain compound R2-2.

[0260]

[0261]

[0262] Compound 3 (50 mg, 47.7 μmol) was dissolved in 2 mL of dry DCM solution. Under nitrogen purging, thionyl chloride (56.7 mg, 0.477 mmol) was added in one go. The tube was sealed and stirred for 3 h. The solvent was removed under reduced pressure to obtain compound R2. R2 was dissolved directly in 2 mL of acetonitrile and triethylamine (77 mg, 0.762 mmol) was added. Finally, compound W (46.3 mg, 95.3 μmol) was added. The mixture was purged with nitrogen three times. The system was heated to 60 °C and reacted for 16 h. The reaction solution was concentrated under reduced pressure and purified by preparative liquid chromatography to separate compound R2-1 (56.2 mg, yield 59.3%).

[0263] LC-MS: 993.5(M / 2+1), 662.7(M / 3+1), 497.3(M / 4+1).

[0264] 1 H-NMR (400M, D2O): δ1.62 (s, 36H), 2.32 (s, 4H), 2.81-2.98 (m, 4H), 3.31-3.75 (m, 40H), 3.93 (d, 16H), 4.24-4 .42(m,12H),4.52(s,8H),5.10(m,2H),7.57(d,4H),7.75-7.85(m,4H),8.06(s,2H),8.12(s,2H),8.97(s,2H)

[0265] Compound R2-1 (50 mg, 25.2 μmol) was dissolved in 4 M hydrochloric acid (1.5 mL), and the mixture was heated to 45 °C and reacted for 0.5 h. After concentration with water, the target compound R2-2 (28 mg, yield 63.1%) was obtained and purified. LC-MS: 881.4 (M / 2+1), 588 (M / 3+1), 441.2 (M / 4+1). See the detailed chromatograms for reference. Figure 2 .

[0266] 1 H-NMR (400M, D2O): δ2.31 (s, 4H), 2.81-2.98 (m, 4H), 3.33-3.77 (m, 40H), 3.92 (d, 16H), 4.23-4.42 (m, 13H), 4.50(s, 9H), 5.08(m, 2H), 7.56(d, 4H), 7.72-7.83(m, 4H), 8.05(s, 2H), 8.15(s, 2H), 8.96(s, 2H)

[0267] Example 2-2

[0268] In this embodiment, we prepared compound R3-1 by reacting compound 3 with an azide compound to obtain compound R3 and then by click coupling of compound N, and further hydrolyzed it with hydrochloric acid to obtain compound R3-2.

[0269]

[0270] Compound 3 (600 mg, 0.572 mmol) was dissolved in 30 mL of anhydrous tetrahydrofuran solution containing triphenylphosphine (600 mg, 2.288 mmol) and 2-acetyl-1-azido-1,2-dihydro-3H-1L3-benzo[D][1,2]iodazole-3-one (754 mg, 2.288 mmol), and the solution was purged with nitrogen. The mixture was heated to 60 °C, sealed, and stirred for 6 h. The solvent was removed under reduced pressure, and pre-TLC yielded a yellow foamy compound R3 (521 mg, yield 82.8%).

[0271] LC-MS: 550.3 (M / 2+1).

[0272] Compound R3 (50 mg, 45.5 μmol) and compound N (40 mg, 0.1 mmol) were dissolved sequentially in tetrahydrofuran / water (2 mL, 1:1) solution. Then, DIEA (29.3 mg, 0.227 mmol) and CuSO4 (0.36 mg, 2.27 μmol) were added sequentially. Under nitrogen bubbling, sodium vitamin C (0.45 mg, 2.27 μmol) was added, and the reaction was carried out at room temperature for 3 hours. 70 mg of a light red solid, R3-1, was obtained.

[0273] LC-MS: 948.4(M / 2+1), 632.6(M / 3+1), 474.7(M / 4+1).

[0274] Compound R3-1 (40 mg) was dissolved in 4 M hydrochloric acid aqueous solution (2 mL), heated to 45 °C and reacted for 1 hour. After dilution and concentration with an appropriate amount of water, lyophilized 15 mg of pale yellow solid R3-2 was obtained.

[0275] LC-MS: 836.3(M / 2+1), 557.9(M / 3+1), 418.7(M / 4+1).

[0276] Example 2-3

[0277] In this embodiment, we prepared compound R4-1 by click coupling of compound R3 and compound M, and further hydrolyzed it with hydrochloric acid to obtain compound R4-2.

[0278]

[0279]

[0280] Compound R3 (13 mg, 11.8 μmol) and compound M (32 mg, 26 μmol) were dissolved sequentially in tetrahydrofuran / water (2 mL, 1:1) solution. Then, DIEA (7.6 mg, 59 μmol) and CuSO4 (94.2 μg, 0.59 μmol) were added sequentially. Under nitrogen bubbling, sodium vitamin C (0.117 mg, 0.59 μmol) was added, and the reaction was carried out at room temperature for 2 hours. 22 mg of pale yellow solid R4-1 was obtained by lyophilization.

[0281] LC-MS: 883.4(M / 4+1), 707(M / 5+1), 589.3(M / 6+1).

[0282] Compound R4-1 (22 mg) was dissolved in 4M hydrochloric acid aqueous solution (1 mL), heated to 45 °C and reacted for 1 hour. After dilution and concentration with an appropriate amount of water, 12 mg of white cotton-like solid R4-2 was prepared by freeze-drying.

[0283] LC-MS: 827.8(M / 4+1), 662.5(M / 5+1).

[0284] Examples 2-4

[0285] In this embodiment, we reacted compound R3 with palladium on carbon to obtain compound R5-1, reacted it with thiocarbonyl diimidazole to obtain R5-2, and coupled it with compound X to prepare compound R5-3, which was further hydrolyzed with hydrochloric acid to obtain compound R5-4.

[0286]

[0287]

[0288] Compound R3 (400 mg, 0.364 mmol) was dissolved in methanol, and then 20% w / w palladium on carbon was added. The mixture was purged three times with hydrogen, stirred at room temperature for 16 h, and then filtered and concentrated to obtain a yellow oily substance R5-1 (362 mg, 95% yield).

[0289] LC-MS: 524.3 (M / 2+1), 349.9 (M / 3+1), 262.7 (M / 4+1). No post-processing required, proceed directly to the next step.

[0290] Compound R5-1 (360 mg, 0.344 mmol) was dissolved in DMF (5 mL), followed by the sequential addition of N,N-thiocarbonyldiimidazole (153.2 mg, 0.86 mmol) and triethylamine (69.6 mg, 0.688 mmol). The mixture was then heated to 50 °C and reacted for 1 h. Further purification by column chromatography yielded a pale yellow oil, R5-2 (220 mg, 51%).

[0291] Compound R5-2 (70 mg, 66.8 μmol) was dissolved in DCM (4 ml), and triethylamine (13.5 mg, 133.6 μmol) and compound X (111 mg, 133.6 μmol) were added sequentially. The mixture was reacted at room temperature for 8 h. The resulting pale yellow compound R5-3 (96 mg, 51.4%) was prepared and purified.

[0292] LC-MS: 932.8(M / 3+1), 699.8(M / 4+1), 560.1(M / 5+1).

[0293] Compound R5-3 was dissolved in 4M HCl (2 ml), heated to 45°C and reacted for 1 hour. The purified and lyophilized compound R5-4 (40 mg, 45.3%) was obtained.

[0294] LC-MS: 858.0(M / 3+1), 643.8(M / 4+1), 515.2(M / 5+1).

[0295] Examples 2-5

[0296] In this embodiment, we prepared compound R6-1 by coupling compound R5-2 with compound Y, and further hydrolyzed it with hydrochloric acid to obtain compound R6-2.

[0297]

[0298]

[0299] Compound R5-2 (70 mg, 66.8 μmol) was dissolved in DCM (3 mL), and triethylamine (13.5 mg, 133.6 μmol) and compound Y (111 mg, 133.6 μmol) were added sequentially. The reaction was carried out at room temperature for 8 h. The purified pale yellow compound R6-1 (80 mg, 43.1%) was prepared. LC-MS: 926.9 (M / 3+1), 695.4 (M / 4+1), 556.5 (M / 5+1).

[0300] Compound R6-1 was dissolved in 4M HCl (2 ml), and the mixture was heated to 45°C and reacted for 1 hour. The purified compound R6-2 (35 mg, 54.8%) was obtained by lyophilization. LC-MS: 740.5 (M / 3+1), 555.6 (M / 4+1), 444.7 (M / 5+1). Examples 2-6

[0301] In this embodiment, we prepared compound R7-1 by coupling compound R5-2 with compound H, and further hydrolyzed it with hydrochloric acid to obtain compound R7-2.

[0302]

[0303] Compound R5-2 (70 mg, 66.8 μmol) was dissolved in DCM (3 ml), and triethylamine (13.5 mg, 133.6 μmol) and compound H (97 mg, 133.6 μmol) were added sequentially. The mixture was reacted at room temperature for 8 h. The purified yellow oily compound R7-1 (92 mg, 57.8%) was prepared.

[0304] LC-MS: 860.7(M / 3+1), 645.8(M / 4+1), 516.8(M / 5+1).

[0305] Compound R7-1 was dissolved in 4M HCl (2.5 ml), heated to 45 °C and reacted for 1 hour. The purified and lyophilized compound R6-2 (51 mg, 63.7%) was obtained.

[0306] LC-MS: 748.6(M / 3+1), 561.7(M / 4+1), 449.6(M / 5+1).

[0307] Examples 2-7

[0308] In this embodiment, we prepared compound R8-1 by coupling compound R3 with compound R, and further hydrolyzed it with hydrochloric acid to obtain compound R8-2.

[0309]

[0310]

[0311] Compounds R3 (11 mg, 0.01 mmol) and R (35 mg, 0.02 mmol) were dissolved in ACN (2 ml) and reacted at room temperature for 1 hour. The reaction was confirmed to be complete by LC monitoring. The solution was concentrated to dryness to obtain R8-1. R8-1 was then cleaved in 1 ml of lysis buffer (TFA / TIS / H2O = 95:2.5:2.5) to obtain white solid R8-2 (3.6 mg).

[0312] LC-MS: 904.9(M / 4+1), 724.1(M / 5+1), 603.6(M / 6+1).

[0313] Examples 2-8

[0314]

[0315] Compound 8 (100 mg, 0.199 mmol) was dissolved in acetonitrile (5 ml), and 2-bromo-N-((4-methoxybenzyl)oxy)-N-methylacetamide (401 mg, 1.39 mmol) and DIEA (205 mg, 1.59 mmol) were added. The mixture was heated to 45 °C and reacted for 12 hours to prepare 183 mg of compound 38.

[0316] LC-MS: 873.4(M / 2+1), 582.6(M / 3+1)

[0317] Compound 38 was purified by hydrolysis to give compound R22.

[0318] LC-MS: 513.2 (M / 2+1).

[0319] The synthesis of compound R2-1 was carried out using compound 38 as a starting material to synthesize compound 39, yielding 94 mg of a yellow solid.

[0320] LC-MS: 894.8(M / 3+1), 671.3(M / 4+1), 537.3(M / 5+1)

[0321] Compound 39 (94 mg, 0.035 mmol) was dissolved in dichloromethane (4 ml), and then TFA (4.5 ml) was added. The mixture was stirred at room temperature for 6 hours, concentrated at low temperature, and then lyophilized to obtain compound R22-1 as a yellow solid (37 mg, 53.8%). See the detailed spectral reference. Figure 5 .

[0322] LC-MS: 981.5(M / 2+1), 654.3(M / 3+1), 491.2(M / 4+1)

[0323] 1H NMR (400MHz, D2O) δ8.94 (d, J=5.4Hz, 2H), 8.14 (d, J=9.3Hz, 2H), 7.99 (d, J=5.4Hz, 2H), 7.8 3-7.68 (m, 4H), 7.34 (s, 4H), 5.10 (dd, J = 8.8, 4.0Hz, 4H), 4.53-2.72 (m, 94H), 2.27 (s, 6H).

[0324] Examples 2-9

[0325]

[0326] Compound 40 was synthesized using the method for compound R2-1, and 493 mg was obtained after purification.

[0327] LC-MS: 756.6 (M / 3+1)

[0328] Compound 40 (454 mg, 0.2 mmol) was dissolved in TFA (3 ml), the PMB protecting group was removed at 45 °C, and then the solution was concentrated to dryness. 3 ml of 4-methylpiperidine was added to remove the Fmoc protecting group, and 113 mg of compound 41 was obtained after purification.

[0329] LC-MS: 662.9 (M / 2+1)

[0330] Compound 41 (26.5 mg, 0.02 mmol) was dissolved in DMF and added dropwise to a DMF solution of p-phenylisothiocyanate (19.2 mg, 0.1 mmol). After the reaction was complete, the solution was concentrated. Compound 42 (6.6 mg) was prepared and purified.

[0331] LC-MS: 759.4 (M / 2+1)

[0332] Add 3 mg of antibody KT1 (150 kDa) to PBS to a final volume of 1.5 mL, and adjust the pH to 8.9 ± 0.2 with NaHCO3. Dissolve compound 42 (0.6 mg) in DMSO (50 μL) and mix well. Incubate the mixture at 37 °C for 30 minutes at 200 rpm using a shaker.

[0333] 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 R21-KT1.

[0334] Using the same method, compound 42 was combined with antibody KT2 (75 kDa) to prepare the antibody-conjugated small molecule compound R21-KT2.

[0335] Using the same method, compound 42 was combined with antibody KT3 (149 kDa) to prepare the antibody-conjugated small molecule compound R21-KT3.

[0336] Example 2-10

[0337]

[0338] Referring to Examples 2-9, the antibody-conjugated small molecule compound R23-KT was prepared.

[0339] Example 2-11

[0340]

[0341] Referring to Examples 2-9, antibody-conjugated small molecule compound R2-KT was prepared.

[0342] Example Group 3

[0343] In this set of embodiments, we describe the chelating ligand compounds disclosed in this invention, the methods for chelating proteins with different metal nuclides by linking targeted small molecule chelating ligand compounds, and the methods for chelating proteins with coupled chelating ligand compounds.

[0344] Example 3-1

[0345] 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 added to 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 reacted 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). The reaction solution was injected into high-performance liquid chromatography (HPLC) to analyze the radiochemical purity of the product (chromatographic conditions: Phenomenex). Luna C18(2) column (5μm, 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%).

[0346] 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).

[0347] The chelation method of compound R1 with the metal nuclide Lu-177 is as follows: commercially available [product name] 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. Detect the radiochemical purity of the product using iTLC (stationary phase iTLC-SG plate, mobile phase: 0.1 M sodium citrate buffer at pH 5).

[0348] iTLC analysis results of the chelation reaction solution of compound R1 with the metal nuclide Ga-68 are as follows: Figure 6 As shown, the chelation reaction yield was 89%, indicating that R1 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0349] HPLC analysis results of the chelation reaction product of compound R1 and the metal nuclide Ga-68 are as follows: Figure 7 As shown, the radiochemical purity of the chelate product was 98%, indicating that R1 could undergo a highly efficient chelation reaction with Ga-68 ions.

[0350] The iTLC analysis results of the chelation reaction product of compound R1 and the metal nuclide Zr-89 are as follows: Figure 8 As shown, the radiochemical purity of the chelate product was greater than 95%, indicating that R1 could undergo a highly efficient chelation reaction with Zr-89 ions.

[0351] The iTLC analysis results of the chelation reaction product of compound R1 and the metal nuclide Lu-177 are as follows: Figure 9 As shown, the radiochemical purity of the chelate product was greater than 95%, indicating that R1 could undergo a highly efficient chelation reaction with Lu-177 ions.

[0352] Example 3-2

[0353] The chelation reaction conditions for the coupling chelating ligand compound R2-2 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 10 As shown, the chelation reaction yield was 92%, indicating that R2-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0354] HPLC analysis results of the chelation reaction product of compound R2-2 and the metal nuclide Ga-68 are as follows: Figure 11 As shown, the radiochemical purity of the chelate product was 99%, indicating that R2-2 and Ga-68 ions underwent a highly efficient chelation reaction.

[0355] Example 3-3

[0356] The chelation reaction conditions for the coupling chelating ligand compound R3-2 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 12 As shown, the chelation reaction yield was 90%, indicating that R3-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0357] HPLC analysis results of the chelation reaction product of compound R3-2 and the metal nuclide Ga-68 are as follows: Figure 13 As shown, the radiochemical purity of the chelate product was 99%, indicating that R3-2 and Ga-68 ions underwent a highly efficient chelation reaction.

[0358] Examples 3-4

[0359] The chelation reaction conditions for the coupling chelating ligand compound R4-2 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 14 As shown, the chelation reaction yield was 94%, indicating that R4-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0360] Examples 3-5

[0361] The chelation reaction conditions for the coupling chelating ligand compound R5-4 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 15 As shown, the chelation reaction yield was 94%, indicating that R5-4 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0362] HPLC analysis results of the chelation reaction product of compound R5-4 and the metal nuclide Ga-68 are as follows: Figure 16 As shown, the radiochemical purity of the chelate product was 94%, indicating that R3-2 and Ga-68 ions underwent a highly efficient chelation reaction.

[0363] Examples 3-6

[0364] The chelation reaction conditions for the coupling chelating ligand compound R6-2 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 17 As shown, the chelation reaction yield was 93%, indicating that R6-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0365] Examples 3-7

[0366] The chelation reaction conditions for the coupling chelating ligand compound R7-2 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 18 As shown, the chelation reaction yield was 91%, indicating that R7-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0367] Examples 3-8

[0368] The chelation reaction conditions of the coupling chelating ligand compound R8 with Ga-68 were the same as in Example 3-1. The iTLC analysis results of the reaction solution are shown in the figure below. The chelation reaction yield was 91%, indicating that R8 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0369] Examples 3-9

[0370] The chelation reaction conditions for the coupling chelating ligand compound R8-2 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 20 As shown, the chelation reaction yield was 90%, indicating that R8-2 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0371] Meanwhile, we used the same chelation reaction as in Example 3-1 to couple compound R8-2 with Zr-89. The iTLC analysis results of the reaction solution are as follows: Figure 21 As shown, the chelation reaction yield was greater than 95%, indicating that R8-2 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0372] Examples 3-10

[0373] The chelation reaction conditions for the coupling chelating ligand compound R9 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 22 As shown, the chelation reaction yield was 86%, indicating that R9 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0374] The chelation reaction conditions for the coupling chelating ligand compound R9 with Za-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 23As shown, the chelation reaction yield was greater than 95%, indicating that R9 can undergo a highly efficient chelation reaction with Za-89 ions.

[0375] Example 3-11

[0376] The chelation reaction conditions for the coupling chelating ligand compound R10 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 24 As shown, the chelation reaction yield was 87%, indicating that R10 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0377] The chelation reaction conditions for the coupling chelating ligand compound R10 with Za-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 25 As shown, the chelation reaction yield was greater than 95%, indicating that R10 can undergo a highly efficient chelation reaction with Za-89 ions.

[0378] Example 3-12

[0379] The chelation reaction conditions for the coupling chelating ligand compound R11 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 26 As shown, the chelation reaction yield was 91%, indicating that R11 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0380] The chelation reaction conditions for the coupling chelating ligand compound R11 with Zr-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 27 As shown, the chelation reaction yield was greater than 95%, indicating that R11 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0381] Example 3-13

[0382] The chelation reaction conditions for the coupling chelating ligand compound R12 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 28 As shown, the chelation reaction yield was 91%, indicating that R12 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0383] The chelation reaction conditions for the coupling chelating ligand compound R12 with Zr-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 29 As shown, the chelation reaction yield was greater than 95%, indicating that R12 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0384] Example 3-14

[0385] The chelation reaction conditions for the coupling chelating ligand compound R13 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 30 As shown, the chelation reaction yield was 92%, indicating that R13 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0386] The chelation reaction conditions for the coupling chelating ligand compound R13 with Zr-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 31 As shown, the chelation reaction yield was greater than 95%, indicating that R13 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0387] Example 3-15

[0388] The chelation reaction conditions for the coupling chelating ligand compound R15 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 32 As shown, the chelation reaction yield was 88%, indicating that R15 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0389] The chelation reaction conditions for the coupling chelating ligand compound R15 with Za-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 33 As shown, the chelation reaction yield was greater than 95%, indicating that R15 can undergo a highly efficient chelation reaction with Za-89 ions.

[0390] Example 3-16

[0391] The chelation reaction conditions for the coupling chelating ligand compound R16 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 34 As shown, the chelation reaction yield was 85%, indicating that R16 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0392] The chelation reaction conditions for the coupling chelating ligand compound R16 and Lu-177 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 35 As shown, the chelation reaction yield was greater than 95%, indicating that R16 can undergo a highly efficient chelation reaction with Lu-177 ions.

[0393] The chelation reaction conditions for the coupling chelating ligand compound R16 with Zr-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 36 As shown, the chelation reaction yield was greater than 95%, indicating that R16 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0394] Example 3-17

[0395] The chelation reaction conditions for the coupling chelating ligand compound R17 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 37 As shown, the chelation reaction yield was 89%, indicating that R17 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0396] The chelation reaction conditions for the coupling chelating ligand compound R17 and Lu-177 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 38 As shown, the chelation reaction yield was greater than 95%, indicating that R17 can undergo a highly efficient chelation reaction with Lu-177 ions.

[0397] The chelation reaction conditions for the coupling chelating ligand compound R17 with Zr-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 39 As shown, the chelation reaction yield was greater than 95%, indicating that R17 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0398] Example 3-18

[0399] The chelation reaction conditions for the coupling chelating ligand compound R18 with Ga-68 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 40 As shown, the chelation reaction yield was 89%, indicating that R18 can undergo a highly efficient chelation reaction with Ga-68 ions.

[0400] The chelation reaction conditions for the coupling chelating ligand compound R18 and Zr-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 41 As shown, the chelation reaction yield was greater than 95%, indicating that R18 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0401] Example 3-19

[0402] The chelation reaction conditions for the coupling chelating ligand compound R22-1 with Za-89 were consistent with those in Example 3-1, and the iTLC analysis results of the reaction solution are as follows: Figure 42 As shown, the chelation reaction yield was greater than 95%, indicating that R22-1 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0403] Example 3-20

[0404] The Zr-89 labeling method for the protein R21-KT1 of the conjugated chelate ligand compound is as follows: Place 100 μL of 0.25 M HEPES solution in a centrifuge tube, and add... 89 Zr 4+The solution was prepared by adjusting the pH to 6.0-6.5 with 0.25M Na₂CO₃ solution. Then, 0.1-1 mg of protein R21-KT1 was added, mixed thoroughly, and the reaction solution was placed at room temperature. After 60 min, the reaction 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 as follows: Figure 43 As shown, the chelation reaction yield was greater than 95%, indicating that the coupling protein R21-KT1 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0405] Example 3-21

[0406] The Zr-89 labeling method for the protein R21-KT2 of the conjugated chelate ligand compound was the same as in Examples 3-20, and the iTLC analysis results of the reaction solution are as follows: Figure 44 As shown, the chelation reaction yield was greater than 95%, indicating that the coupling protein R21-KT2 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0407] Example 3-22

[0408] The Zr-89 labeling method for the protein R21-KT3 of the conjugated chelate ligand compound was the same as in Examples 3-20, and the iTLC analysis results of the reaction solution are as follows: Figure 45 As shown, the chelation reaction yield was greater than 95%, indicating that the coupling protein R21-KT3 can undergo a highly efficient chelation reaction with Zr-89 ions.

[0409] Example 3-23

[0410] The Zr-89 labeling method for the protein R23-KT of the conjugated chelate ligand compound was the same as in Examples 3-20, and the iTLC analysis results of the reaction solution are as follows: Figure 46 As shown, the chelation reaction yield was greater than 95%, indicating that the coupling protein R23-KT can undergo a highly efficient chelation reaction with Za-89 ions.

[0411] Example 3-24

[0412] Ga-68 labeling method for protein R2-KT containing conjugated chelate ligands: Take a sample containing... 68 Ga 3+The eluent from the Ge-Ga generator was adjusted to pH 5.5-6.5 with 1M sodium acetate solution. The protein solution to be labeled was added at a concentration of 1-10 mg. The centrifuge tube was sealed and incubated at room temperature. After 30 min, the product solution was injected for HPLC analysis (HPLC analysis conditions: column: TOSOH G3000SWXL 7.8*300mm, 5μm; mobile phase: PBS buffer containing 10% acetonitrile; flow rate: 1.0 ml / min; detection wavelength: 280 nm). The HPLC analysis results of the reaction solution are as follows: Figure 47 As shown, the product has a radiochemical purity greater than 97%, indicating that the coupling protein R2-KT can undergo a highly efficient chelation reaction with Ga-68 ions.

[0413] Example Group 4

[0414] The targeting and metabolic properties of the chelating radioactive metal nuclides, linking targeted small molecule chelating ligand compounds and coupled chelating ligand compound proteins prepared in the above embodiments were evaluated in vivo using PET imaging in a mouse tumor model.

[0415] Example 4-1

[0416] R2-2, a Ga-68 chelate prepared in Example 3-2 with a radioactivity of 100 μCi, was injected via the tail vein into a nude mouse model bearing U87-MG tumors. PET imaging data were acquired 60 minutes after injection, and the results are as follows. Figure 48 As shown, the compound can be seen 68 Ga-R2-2 was significantly uptaken in U87-MG tumor tissue (as indicated by the arrow in the figure), but showed no significant retention in other organs besides the bladder.

[0417] Example 4-2

[0418] The PET imaging method for R3-2 chelating Ga-68 in a nude mouse model bearing U87-MG tumors is as described in Example 4-1, yielding... 68 PET imaging of Ga-R3-2 is as follows Figure 49 You can see 68 Ga-R3-2 was significantly taken up in U87-MG tumor tissue, but also significantly retained in the kidneys and liver.

[0419] Example 4-3

[0420] The PET imaging method for R5-4 chelating Ga-68 in a nude mouse model bearing U87-MG tumors is as described in Example 4-1, yielding... 68 PET imaging of Ga-R5-4 is as follows Figure 50 You can see 68Ga-R5-4 was significantly uptaken in U87-MG tumor tissue.

[0421] Example 4-4

[0422] The chelating compound with a radioactivity of 70 μCi prepared in Examples 3-19 was used. 89 Zr-R22-1 was injected via the tail vein into a nude mouse model bearing U87-MG tumors. PET imaging data were acquired 24 hours after injection. The results are as follows: Figure 51 As shown, you can see 89 Zr-R22-1 is taken up in U87-MG tumor tissue.

[0423] Examples 4-5

[0424] The products prepared in Examples 3-20 89 The PET imaging method for Zr-R21-KT1 was the same as in Examples 4-4, and PET images were obtained. Figure 52 You can see 89 Zr-R21-KT1 was significantly uptaken in U87-MG tumor tissue.

[0425] Examples 4-6

[0426] The preparations in Examples 3-21 89 The PET imaging method for Zr-R21-KT2 was the same as in Examples 4-4, and the PET images are shown below. Figure 53 You can see 89 Zr-R21-KT2 showed significant uptake in U87-MG tumor tissue.

[0427] Examples 4-7

[0428] The preparations in Examples 3-22 89 The PET imaging method for Zr-R21-KT3 was the same as in Examples 4-4, and the results were as follows: Figure 54 As shown, you can see 89 Zr-R21-KT3 was significantly uptaken in U87-MG tumor tissue.

[0429] The above describes specific embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of the present invention, and these improvements and modifications are also considered to be within the scope of protection of the present invention.

Claims

1. A chelating ligand compound, characterized in that, The chelating ligand compound is a compound of general formula I or a pharmaceutically acceptable salt thereof: At least one of A1 and A2 contains an active group AG that can be linked to the target molecule; when both A1 and A2 contain an active group AG that can be linked to the target molecule, the active groups can be the same or different. When A1 contains an active group AG that can connect with the target molecule, A1 is the target molecule linker arm; When A1 does not contain an active group AG that can connect with the target molecule, A1 = R1; When A2 contains an active group AG that can connect with the target molecule, A2 is the target molecule linker arm; When A2 does not contain an active group AG that can connect with the target molecule, A2 = R2; Preferably, when A1 is a target molecule linker arm, A1 is L1-AG1, and when A2 is a target molecule linker arm, A2 is L2-AG2. Preferably, AG1 and AG2 are 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. The L1 and L2 are 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, L1 and L2 are independently and arbitrarily selected. Preferably, when A1 and A2 are non-targeted connecting arms, A1 and A2 are L N -NAG, L N Similar to the definitions of L1 and L2 mentioned above, NAG is an alkyl group or H; K1 and K2 are heteroalkyl, heterocyclic alkyl, or heteroaryl groups containing heteroatoms O, S, P, or N; K1 and K2 can be the same or different; where K1 and K2 are heteroalkyl means that K1 and K2 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 and K2 are heterocyclic groups, meaning that K1 and K2 are 4-7 membered substituted or unsubstituted heterocyclic alkyl groups containing heteroatoms O, S, P, or N, preferably tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydropyrrolyl, or piperazineyl; K1 and K2 are heteroaryl groups, meaning that K1 and K2 are 4-7 membered heteroaryl groups containing heteroatoms O, S, P, or N, preferably furanyl, thiophenyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl, pyridinyl, pyridinyl, pyrazine, isoxazolyl, diazolyl, pyrazolyl, triazolyl, tetrazolyl, isothiazolyl, or thiadiazolyl. In general formula I, the N atom is connected to H or a substituent. When a substituent is connected, the substituent can be a chelating arm of a heteroatom group Q with a lone pair of electrons or a target molecule linking arm L3 with an active group AG3 that can link to the target molecule. The chelating arm has the structural formula JQ, where J is a substituted or unsubstituted, saturated or unsaturated ether, ester, ketone, amide or C0-C5 alkyl chain; The structural formula of the target molecule linker arm L3 is L3-AG3. AG3 has the same definition as AG1 and AG2 mentioned above. AG1, AG2 and AG3 can be the same or different. L3 has the same definition as L1 and L2. L3, L1 and L2 can be the same or different.

2. The chelating ligand compound according to claim 1, characterized in that, The J also includes a group AG4 that can be linked to the target molecule. AG4 has the same definition as AG1, AG2, and AG3 mentioned above. AG1, AG2, AG3, and AG4 can be the same or different.

3. The chelating ligand compound according to claim 1, characterized in that, The chelating ligand compound has the following structure: Wherein, B1, B2, B3, and B4 are H or substituents. When a substituent is attached, the substituent may be a chelating arm of a heteroatom group Q with a lone pair of electrons or a target molecule linking arm L3 with an active group AG3 that can link with the target molecule. Q and L3 are defined in the same way as L1 and L2 in claim 1. X is a halogen (Br, Cl, I, F).

4. The chelating ligand compound according to claim 1, characterized in that, The chelating ligand compound has any of the following preferred characteristics. (1) K1 and K2 are independently and arbitrarily selected from: (2) Group Q is selected from: -H2PO3、-H2PO4、-H2PO2、 Wherein R, R', and R" are H, alkyl, hydroxyl, fatty acid group, amino, acyl, heteroalkyl, or heteroaryl; (3) the chelating ligand compound is arbitrarily selected from:

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

5. The targeted chelating ligand compound 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, 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 chelating ligand compound is:

6. The salt formed by the chelating ligand compound of claim 1 and an inorganic or organic acid.

7. The method for preparing the chelating ligand compound according to claim 1, characterized in that, This method is prepared from 5-substituted or unsubstituted 2-hydroxy-isophthalaldehyde and NH2—K1—NH2, NH2—K2—NH2 as raw materials, wherein K1 and K2 are defined as described in any one of claims 1-4.

8. A metal complex formed from the chelating ligand compound of any one of claims 1 to 4, or the targeted chelating ligand compound of claim 5 or 6.

9. 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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