Chelating ligand compound, derivative and preparation method therefor, conjugate formed thereby and use thereof

Abstract

Provided are a chelating ligand compound, a derivative and a preparation method therefor, a conjugate formed thereby and a use thereof. The chelating ligand compound has larger chelating space and a larger chelating capability, can form a stable structure with various metal ions, especially metal ions having a large atomic radius and much charge, thereby possessing higher in-vitro stability, and providing more possibilities for radiopharmaceutical therapy. Moreover, due to the large framework of the chelating ligand compound, high-temperature heating is not needed during metal nuclide labeling, thereby preventing the structure of the framework from being damaged in the heating process; additionally, the metal nuclide labeling process is simplified, thereby shortening the time, and increasing the radiochemical purity of a final target radiopharmaceutical.

Description

Chelated ligand compound, derivative, preparation method thereof, conjugate formed therefrom and application thereof Technical Field

[0001] The present invention relates to the research field of chelate ligand compounds capable of chelating with metal ions, especially nuclides, and in particular to a chelate ligand compound, a derivative thereof, a preparation method thereof, a conjugate formed therefrom and applications thereof. Background Art

[0002] Radiochelates or radiolabeled targeting compounds have been widely used in diagnostics and usually include at least a chelating ligand with a complexed radionuclide, a linker, and a targeting molecule or a cross-linking group.

[0003] Currently, an increasing number of metal nuclides are being developed as new therapeutic and diagnostic metals. However, due to their large ionic radius and high charge, the chelating ligands currently known in the prior art are unable to stably complex with them, resulting in the inability of the formed metal chelates to maintain sufficient stability in the body. This, in turn, is expected to cause a series of side effects, thus limiting the application of these metal nuclides in medicine, especially in diagnostic medicine.

[0004] At the same time, existing relatively mature chelating ligand compounds, such as traditional DOTA and DTPA ligands, require high-temperature conditions in the labeling process of some large molecular weight nuclides. Such high-temperature conditions may destroy the structural integrity of the antibody targeting part, which also makes traditional ligands unable to adapt to the chelation needs of more metal nuclides.

[0005] Therefore, for those skilled in the art, developing new chelating ligand compounds that can meet the chelation needs of more metal nuclides, especially those with larger ionic radius and more free charge, has become a hot spot and difficulty in the research and development of these technicians.

[0006] Summary of the Invention

[0007] One of the objectives of the present invention is to provide a novel chelating ligand compound whose chelating groups are capable of chelating metal nuclides with larger ionic radii and higher free charges. This serves as a foundation for further chelating the metal nuclides after attachment to a targeting molecule, resulting in a conjugate with enhanced targeting and biological properties.

[0008] The present invention also provides a method for preparing the above-mentioned chelated ligand compound and its derivatives, and their use in preparing radiopharmaceuticals. The radiopharmaceuticals can be used for the diagnosis and treatment of related diseases.

[0009] In order to achieve the above-mentioned object of the invention, the present invention provides a new chelate ligand compound, which comprises a chelate group containing multiple nitrogen atoms and at least one linker connected to the nitrogen atom on the chelate group:

[0010] in:

[0011] Z is a chelating group containing multiple nitrogen atoms and has the following structure:

[0012] R1 and R2 are substituents on the benzene ring, and the substituents do not contain residues that react with the target molecule; more preferably, R1=R2; more preferably, R1 and R2 are both methyl groups;

[0013] K is an alkyl chain containing heteroatoms O, S, P or N, and is composed of several R k Connected by heteroatoms or groups containing heteroatoms, where R k independently and arbitrarily selected from substituted or unsubstituted alkyl;

[0014] L is a linker portion connected to the N atom in the chelating group Z, and the linker portion L has a terminal group X connected to the target molecule or targeting group.

[0015] The terminal group X on the linker portion L is an important part for connecting to the target molecule or targeting group. Different terminal groups X can be connected according to the connection requirements of different target molecules or targeting groups, so that the connection with the target molecule can be achieved through covalent bonding, ligand bonding, or hydrophobic bonding and electrostatic interaction.

[0016] The other end of the linker portion L is connected to the nitrogen atom of the chelating group Z, thereby forming a chelating ligand compound that can be coupled to different target molecules or targeting groups.

[0017] Sometimes we may need several identical or different terminal groups X. These terminal groups X can be designed on one linker portion L or on different linker portions L. The specific design ideas and methods can refer to the connection method between the targeting molecule or targeting group and the linker portion L in the prior art.

[0018] When K is an alkyl chain containing heteroatoms O, S, or P, the linker portion L is connected to one, two, three, or four of the four nitrogen atoms on the chelating group Z; when K is an alkyl chain containing heteroatoms N, the N atoms on the K chain can also be connected to the linker portion L, thereby providing more connection sites for the linker portion L. Of course, this does not mean that all nitrogen atoms need to be connected to the linker portion L, as long as at least one nitrogen atom is connected to the linker portion L.

[0019] When R1 and R2 are both located in the para position of the hydroxyl group, except for the nitrogen atom that may exist on the alkyl chain K, the environments of the other four nitrogen atoms are the same. When K is an alkyl chain containing a heteroatom N, the nitrogen atom on the alkyl chain K is not completely equivalent to the other nitrogen atoms due to their positions, and thus two nitrogen atoms in different environments are formed.

[0020] As a further preferred technical solution, the linker portion L is selected from a group having the following formula: L m ——X,

[0021] in:

[0022] X is -COOH, -OH, -CO, -SH, -CO-N(CH3)OH, -NH2,

[0023] —H2PO3, —H2PO4, —H2PO2 or halogen;

[0024] L m is a substituted or unsubstituted alkyl group, preferably, L m It is a substituted or unsubstituted C0~C3 alkyl group.

[0025] Further preferably, the chelating ligand compound further comprises a group Q, wherein the group Q is a heteroatom group with a lone pair of electrons;

[0026] There are two ways to connect the group Q and the chelate group Z.

[0027] One is that the group Q is connected between the chelating group Z and the linker arm portion L,

[0028] The second is that the group Q is directly connected to the N atoms in the chelate group Z that are not connected to the linker part L,

[0029] It should be noted that, although two connection modes of the group Q and the chelating group Z are provided herein, this does not mean that in a specific embodiment, the group Q can only adopt one connection mode, not the other. On the contrary, in a technical solution, for the group Q, there may be two different connection modes with the chelating group Z at the same time, and these groups Q may be the same or different.

[0030] Further preferably, the group Q is selected from the group having the following formula, Q m -X Q ;

[0031] in:

[0032] Q mis a substituted or unsubstituted alkyl group, preferably, Q m is a substituted or unsubstituted C0~C3 alkyl group;

[0033] X Q A heteroatom group with a lone pair of electrons, including but not limited to —COOH, —H2PO3, —H2PO4 or —H2PO2.

[0034] Furthermore, the present invention also discloses R k Selected from substituted or unsubstituted methyl, substituted or unsubstituted ethyl, substituted or unsubstituted propyl, these Rk are connected through heteroatoms to form a group containing two or three R k The alkyl chain K.

[0035] Further preferably, the present invention further discloses that the alkyl chain K containing heteroatoms O, S, P or N in the chelating ligand compound is selected from:

[0036] Any one of .

[0037] At the same time, the present invention further discloses that the chelating ligand compound is selected from: Any one of .

[0038] Furthermore, the present invention also discloses a derivative of the chelating ligand, which is a compound having a structure shown in the following formula or a salt thereof formed with an inorganic acid or an organic acid,

[0039] In this formula, K is defined as above, i.e., K is an alkyl chain containing heteroatoms O, S, P or N, and is composed of several R k Connected by heteroatoms or groups containing heteroatoms, where R k Independently and arbitrarily selected from substituted or unsubstituted alkyl; and preferably R k Selected from substituted or unsubstituted methyl, substituted or unsubstituted ethyl, substituted or unsubstituted propyl, these Rk are connected through heteroatoms to form a group containing two or three R k At the same time, K is further preferably any one of the following formulae:

[0040] Furthermore, the present invention also discloses a conjugate formed by coupling the chelating ligand compound with the target molecule or targeting group. As mentioned above, the target molecule or targeting group is coupled with the terminal group X of the linker portion L.

[0041] The target molecule can be selected from biological macromolecules, or can be selected from drugs or small molecule compounds. The target molecule and targeting group include but are not limited to antibodies, proteins, peptides, carbohydrates, nucleotides, oligonucleotides, oligosaccharides, vitamins, liposomes, small molecule drugs or fragments or derivatives, etc.

[0042] Furthermore, the present invention also provides a method for preparing the chelating ligand compound, which is prepared by dissolving substituted or unsubstituted 2,6-diformylphenol and NH2—K—NH2 as raw materials, wherein K is defined as described above.

[0043] The present invention also discloses a metal complex, which is formed by complexing the aforementioned chelating ligand compound or coupling compound with a metal element.

[0044] Furthermore, the present invention discloses the use of the aforementioned chelated ligand compounds, derivatives, conjugates, and metal complexes in the preparation of radiopharmaceuticals. Radiopharmaceuticals prepared by combining different target molecules with different radionuclides to form compounds can be used for targeted diagnosis and treatment of various diseases.

[0045] The present invention discloses the use of metal complexes in radiopharmaceuticals. In this application, the metal is a metal nuclide, wherein the nuclide can be 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 Such as 105 Rh, 109 Pd, 111 In, 117m Sn, 149 Pm, 52 Mn, 149 Tb, 152 Tb, 153 Sm, 177 Lu, 186 Re、 188 Re、 199 Au, 201 Tl, 203 Pb, 212 Pb,212 Bi, 213 Bi, 225 Ac, 223 Ra and 227 Th, etc., wherein the chelating ligand compound disclosed in the present invention has a particularly significant advantage in chelating with lanthanides and actinides, such as the one used in the embodiment of the present invention. 68 Ga.

[0046] The chelating ligand compounds disclosed in the present invention possess a larger chelating space and greater chelating capacity, and can form stable structures with a variety of metal ions, particularly those with large atomic radii and high charges. This allows for greater in vitro stability and expands the potential for radiopharmaceutical therapy. Furthermore, because the chelating ligand compounds disclosed in the present invention have a large skeleton, they do not require high-temperature heating when reacting with metal nuclides, thus avoiding the possibility of the skeleton structure being destroyed during heating. This also simplifies the metal nuclide labeling process, shortens the time, and improves the radiochemical purity of the final target radiopharmaceutical. BRIEF DESCRIPTION OF THE DRAWINGS

[0047] FIG1 is a schematic diagram of the single crystal structure of compound 2 obtained in Example 1.

[0048] FIG2 is a schematic diagram of the single crystal structure of the target chelating ligand compound R1 obtained in Example 1.

[0049] Figure 3 is the compound 2 obtained in Example 1 1 HNMR spectrum.

[0050] Figure 4 shows the compound 3 obtained in Example 1 1 HNMR spectrum.

[0051] Figure 5 shows the structure of compound 11 obtained in Example 7. 1 HNMR spectrum.

[0052] Figure 6 shows the compound R4 obtained in Example 7. 1 HNMR spectrum.

[0053] Figure 7 shows the compound R6 obtained in Example 9 1 HNMR spectrum.

[0054] Figure 8 shows the compound in Example 14 68 Schematic diagram of thin layer chromatography detection results of Ga-R1.

[0055] Figure 9 shows the compound in Example 14 68 Schematic diagram of thin layer chromatography detection results of Ga-R2.

[0056] Figure 10 shows the compound in Example 14 68Schematic diagram of thin layer chromatography detection results of Ga-R3.

[0057] Figure 11 shows the compound in Example 14 68 Schematic diagram of thin layer chromatography detection results of Ga-R4.

[0058] Figure 12 shows the compound in Example 14 68 Schematic diagram of thin layer chromatography detection results of Ga-R5.

[0059] Figure 13 shows the compound in Example 14 68 Schematic diagram of thin layer chromatography detection results of Ga-R6.

[0060] Figure 14 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-4.

[0061] Figure 15 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-5.

[0062] Figure 16 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-6.

[0063] Figure 17 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-7.

[0064] Figure 18 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-10.

[0065] Figure 19 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-11.

[0066] Figure 20 shows the compound in Example 15 68 Schematic diagram of the HPLC analysis results of Ga-R1-12.

[0067] FIG. 21 shows the injection of compounds in Example 16. 68 PET scan of Ga-R1-4 in nude mice.

[0068] FIG. 22 shows the injection of compounds in Example 16. 68 PET scan of Ga-R1-6 in nude mice.

[0069] FIG. 23 shows the injection of compounds in Example 16. 68 PET scan of Ga-R1-12 in nude mice.

[0070] FIG. 24 is a diagram of Example 17.68 Schematic diagram of the distribution results of Ga-R1-4 in tumor-bearing mice. DETAILED DESCRIPTION

[0071] In order to better understand the present invention, the present invention is further described below with reference to specific embodiments.

[0072] The reagents and instruments not otherwise specified or indicated in the examples are all conventional commercial products and can be purchased. Any conditions not otherwise specified or indicated can be carried out in accordance with conventional conditions in the art or the instructions of the relevant product sellers.

[0073] The invention uses substituted or unsubstituted 2,6-diformylphenol and NH2—K—NH2 as raw materials for preparation. Specifically, the substituted or unsubstituted 2,6-diformylphenol and NH2—K—NH2 are mixed to react to obtain a macrocyclic compound (Schiff base), which is then reduced with sodium borohydride and reacted with tert-butyl bromoacetate (or a hydroxylamine intermediate or a phosphate intermediate). The obtained intermediate is hydrolyzed to generate the target compound.

[0074] Example 1

[0075] Dissolve 2,6-diformyl-4-methylphenol (1.225 g, 7.4 mmol) in 30 ml of DMF and heat to 90°C. Add 2,2′-(ethylenedioxy)bis(ethylamine) (1.10 g, 7.4 mmol) in 30 ml of DMF dropwise at 90°C. Continue the reaction for 2 hours. Concentrate the reaction mixture to obtain Compound 1.

[0076] Sodium borohydride (0.562 g, 14.8 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 extracted and washed with ethyl acetate / water, then dried over anhydrous sodium sulfate. Filtered, 10 ml of a 4 M solution of hydrogen chloride in dioxane was added to the filtrate, and the precipitated solid was filtered and dried to obtain the hydrochloride salt of compound 2 (2.6 g).

[0077] LC-MS: 561.4(M+1), 281.2(M / 2+1)

[0078] H-NMR (300M, D2O): δ 2.28 (s, 6H), 3.31-3.33 (t, 8H), 3.77 (s, 8H), 3.85-3.87 (t, 8H), 4.29 (s, 8H), 7.28 (s, 4H). See Figure 3 for the specific spectrum.

[0079] 100 mg of compound 2 was weighed and dissolved in 2 ml of methanol. The mixture was filtered, and 4 ml of ether was added to the filtrate. The mixture was covered and allowed to stand for 3 weeks. Single crystals grew, as shown in FIG1 .

[0080] The hydrochloride of compound 2 (600 mg, 0.85 mmol), tert-butyl bromoacetate (623 mg, 3.2 mmol), DIEA (1.4 g, 10.8 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 a crude product, which was then purified to obtain compound 3 (788 mg).

[0081] LC-MS: 509.3 (M / 2+1).

[0082] Compound 3 (788 mg, 0.775 mmol) was dissolved in 4M hydrochloric acid (3 ml) and reacted at 45°C for 0.5 hour. The mixture was concentrated to dryness under reduced pressure to obtain the target compound R1 (594 mg). The specific spectrum is shown in Figure 4.

[0083] LC-MS: 793.4(M+1), 397.2(M / 2+1)

[0084] H-NMR (300M, D2O): δ2.26 (s, 6H), 3.56-3.58 (t, 8H), 3.75 (s, 8H), 3.94-3.96 (t, 8H), 4.03 (s, 8H), 4.52 (br, 8H), 7.32 (s, 4H)

[0085] Weigh 10 mg of compound R1 and dissolve it in 1 ml of ultra-dried, filtered MeOH. Add 232 μl of 0.024 M Cu(NO₃)₂ / MeOH solution. Add 840 μl of 0.1 M NaOH / MeOH solution, and the solution turns dark green. Filter, add 4 ml of ether, cover, and let stand for 2 weeks. Single crystals will grow, with the structure shown in Figure 2.

[0086] Example 2

[0087] In this example, we used compound R1 to couple with compound W to prepare compounds R1-1, R1-2, R1-3, R1-4, R1-5, and R1-6.

[0088] Compound 4 (79.2 mg, 0.1 mmol), N,N-diisopropylethylamine (64.5 mg, 0.5 mmol) and HATU (152 mg, 0.4 mmol) were dissolved in 2 ml of DMF solution and stirred for 10 min. Compound W (146 mg, 0.3 mmol) was added and reacted for 6 h. The reaction solution was concentrated under reduced pressure and purified using preparative liquid phase to isolate the following compound:

[0089] R1-1(21.3mg), LC-MS: 631.3(M / 2+1)

[0090] R1-2(0.2mg), LC-MS: 577.3(M / 3+1)

[0091] R1-3(0.3mg), LC-MS: 577.3(M / 3+1)

[0092] R1-4(29.1mg), LC-MS: 577.3(M / 3+1)

[0093] R1-5(39.4mg), LC-MS: 733.6(M / 3+1), 550.5(M / 4+1)

[0094] R1-6(6.9mg), LC-MS: 667.5(M / 4+1), 534.2(M / 5+1)

[0095] Example 3

[0096] In this example, we used compound R1 to couple with compound X to prepare compounds R1-7, R1-8, R1-9, R1-10, R1-11, and R1-12.

[0097] Compound R1 (79.2 mg, 0.1 mmol), N,N-diisopropylethylamine (64.5 mg, 0.5 mmol) and HATU (152 mg, 4 mmol) were dissolved in 2 ml of DMF solution and stirred for 10 min. Compound X (250 mg, 3 mmol) was added and reacted for 2 h. The reaction solution was concentrated under reduced pressure and purified using preparative liquid phase to isolate the compound:

[0098] R1-7(19.6mg), LC-MS: 804.4(M / 2+1), 536.6(M / 3+1)

[0099] R1-8(0.1mg), LC-MS: 808.4(M / 3+1), 606.5(M / 4+1)

[0100] R1-9(0.1mg), LC-MS: 808.4(M / 3+1), 606.5(M / 4+1)

[0101] R1-10(29.6mg), LC-MS: 808.4(M / 3+1), 606.5(M / 4+1)

[0102] R1-11(34.4mg), LC-MS: 810.1(M / 4+1), 648.3(M / 5+1)

[0103] R1-12(39.1mg), LC-MS: 676.3(M / 6+1)

[0104] Example 4

[0105] In this example, we used compound R1 to couple with compound Y to prepare compounds R1-13, R1-14, R1-15, R1-16, R1-17, and R1-18.

[0106] Compound 4 (79.2 mg, 0.1 mmol), N,N-diisopropylethylamine (64.5 mg, 0.5 mmol) and HATU (114 mg, 0.3 mmol) were dissolved in 2 ml of DMF solution and stirred for 10 min. Compound Y (196.5 mg, 0.3 mmol) was added and reacted for 2 h. The reaction solution was concentrated under reduced pressure and purified using preparative liquid phase to isolate compound 4.

[0107] R1-13(13.9mg), LC-MS: 715.8(M / 2+1), 477.6(M / 3+1)

[0108] R1-14(0.1mg), LC-MS: 690.3(M / 3+1), 517.9(M / 4+1)

[0109] R1-15(0.2mg), LC-MS: 690.3(M / 3+1), 517.9(M / 4+1)

[0110] R1-16(19.8mg), LC-MS: 690.3(M / 3+1), 517.9(M / 4+1)

[0111] R1-17(22.8mg), LC-MS: 677.3(M / 4+1)

[0112] R1-18(21.5mg), LC-MS: 836.7(M / 4+1), 669.6(M / 5+1)

[0113] Example 5

[0114] Dissolve 2,6-diformyl-4-methylphenol (0.82 g, 5 mmol) in 300 ml of methanol, and add a solution of bis(3-aminopropyl) ether (0.66 g, 5 mmol) in 300 ml of methanol dropwise. Continue the reaction for 2 hours. Then, add sodium borohydride (760 mg, 20 mmol) in portions. After 1 hour of reaction, concentrate and purify to obtain compound 5 (1.01 g). LC-MS: 529.4 (M+1).

[0115] H-NMR (300M, D2O): δ1.95-2.01 (m, 8H), 2.28 (s, 6H), 3.14-3.17 (t, 8H), 3.60-3.63 (t, 8H), 4.24 (s, 8H), 7.24 (s, 4H)

[0116] Compound 5 (528 mg, 1 mmol), tert-butyl bromoacetate (1.17 g, 6 mmol), DIEA (1.03 g, 8 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 a crude product, which was then purified to obtain compound 6 (804 mg).

[0117] Compound 6 (328 mg, 0.33 mmol) was dissolved in 4M hydrochloric acid (3 ml) and reacted at 45°C for 0.5 hours. The mixture was concentrated to dryness under reduced pressure to obtain the target compound R2 (124 mg).

[0118] LC-MS: 761.4(M+1), 381.3(M / 2+1)

[0119] H-NMR (300M, D2O): δ2.11 (br, 8H), 2.28 (s, 6H), 3.45 (br, 8H), 3.55 (br, 8H), 3.90 (br, 8H), 4.52 (br, 8H), 7.26 (s, 4H)

[0120] Example 6

[0121] According to the method of Example 5, only bis(3-aminopropyl)ether was replaced by 2,2'-oxybis(ethylamine) to obtain compound R3.

[0122] in:

[0123] Compound 8:

[0124] LC-MS: 473.3 (M+1)

[0125] H-NMR (300M, D2O): δ2.28 (s, 6H), 3.25-3.28 (t, 8H), 3.70-3.73 (t, 8H), 4.25 (s, 8H), 7.24 (s, 4H).

[0126] Compound R3:

[0127] LC-MS: 705.3(M+1), 353.2(M / 2+1)

[0128] H-NMR (300M, D2O): δ2.28 (s, 6H), 3.49 (br, 8H), 3.56 (br, 8H), 3.90 (br, 8H), 4.52 (br, 8H), 7.26 (s, 4H).

[0129] Example 7

[0130] According to the method of Example 5, only bis(3-aminopropyl)ether was replaced by diethylenetriamine to obtain compound R4.

[0131] in:

[0132] Compound 11:

[0133] LC-MS: 471.3 (M+1)

[0134] H-NMR (300M, D2O): δ 2.28 (s, 6H), 3.33-3.36 (m, 8H), 3.42-3.46 (m, 8H), 4.34 (s, 8H), 7.30 (s, 4H), see Figure 5 for details.

[0135] Compound R4:

[0136] LC-MS: 819.4(M+1), 352.2(M / 2+1)

[0137] H-NMR (300M, D2O): δ 2.28 (s, 6H), 3.14 (m, 8H), 3.30 (m, 4H), 3.43 (m, 8H), 3.90 (s, 8H), 4.46 (s, 8H), 7.33 (s, 4H), see Figure 6 for details.

[0138] Example 8

[0139] According to the method of Example 5, only bis(3-aminopropyl)ether was replaced by aminoethyl sulfide to obtain compound R5.

[0140] in:

[0141] Compound R5:

[0142] LC-MS: 737.3(M+1), 369.1(M / 2+1)

[0143] H-NMR (300M, D2O): δ2.28 (s, 6H), 2.58 (br, 8H), 3.41 (br, 8H), 3.90 (br, 8H), 4.52 (br, 8H), 7.26 (s, 4H)

[0144] Example 9

[0145] 70.6 mg (0.1 mmol) of intermediate 2 was added to 2 ml of acetonitrile, followed by DIEA (1.29 g, 10 mmol), and 176 mg (0.6 mmol) of compound V was added under stirring. After reacting for 6 hours, the crude product of compound 17 was obtained through purification to obtain 119 mg.

[0146] LC-MS: 707.3 (M / 2+1)

[0147] 50 mg of compound 17 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 concentrated to obtain compound R6.

[0148] Compound R6:

[0149] LC-MS: 527.4 (M / 2+1)

[0150] H-NMR (300M, D2O): δ 2.14 (s, 6H), 3.66 (s, 6H), 3.78 (s, 6H), 3.99 (s, 6H), 4.56 (s, 6H), 6.60-6.62 (d, 4H), 6.70-6.71 (d, 4H), 7.10 (s, 4H), 7.43-7.47 (t, 4H), see Figure 7 for details.

[0151] Example 10

[0152] Referring to Example 9, Compound V was replaced by Compound VI to obtain Compound R7

[0153] Compound 19:

[0154] LC-MS: 635.3 (M / 2+1)

[0155] H-NMR (300M, D2O): δ 2.33 (s, 6H), 3.11-3.19 (m, 20H), 3.66-3.78 (m, 24H), 4.24 (s, 8H), 4.91 (s, 8H), 7.25 (s, 4H), 7.41-7.45 (m, 20H) Compound R7:

[0156] LC-MS: 455.2 (M / 2+1)

[0157] Example 11

[0158] Referring to Example 9, Compound V was replaced with Compound VII to obtain Compound R8

[0159] Compound R8:

[0160] LC-MS: 469.2 (M / 2+1)

[0161] Example 12

[0162] Referring to Example 9, compound V was replaced by compound VIII to obtain compound R9

[0163] Compound R9:

[0164] LC-MS: 699.2 (M / 2+1)

[0165] Example 13

[0166] 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 obtain compound 25 (358 mg).

[0167] LC-MS: 474.2(M+23), 396.2(M+1-56)

[0168] Compound 25 (225 mg, 0.5 mmol) was dissolved in 20 ml of dichloromethane, and a 1 M solution of hydrogen chloride in dioxane (1 ml) was added dropwise. After 30 minutes, the mixture was diluted with 20 ml of dichloromethane and concentrated to give a crude product of 163 mg of the hydrochloride salt of compound 26. LC-MS: 252.2 (M+1)

[0169] 2,6-diformyl-4-methylphenol (74 mg, 0.45 mmol) was dissolved in 30 ml of methanol and heated to 60° C. A 20 ml methanol solution of compound 26 (163 mg, 0.5 mmol) was added dropwise and the reaction was continued for 2 hours.

[0170] After the temperature was lowered to 45°C, sodium borohydride (76 mg, 2 mmol) was added to the reaction solution with stirring, and the mixture was stirred for 1 hour. The solvent was removed under reduced pressure, and the residue was extracted and washed to obtain a crude product of compound 27 (79 mg).

[0171] LC-MS: 384.2 (M / 2+1)

[0172] The crude product of compound 27 (79 mg, 0.1 mmol) was added to DMF, and DIEA (88 mg, 0.68 mmol) and compound V (199 mg, 0.68 mmol) were added under stirring. The mixture was stirred and reacted overnight. Ethyl acetate and water were added, and the mixture was extracted and washed to obtain 126 mg of the crude product. 81 mg of compound 28 was obtained after purification.

[0173] LC-MS: 540.8 (M / 3+1)

[0174] Compound 28 (81 mg) was added with 2 ml of concentrated hydrochloric acid and reacted at 55°C for 30 minutes. The mixture was concentrated to dryness to prepare and purify compound R10.

[0175] LC-MS: 540.2 (M / 2+1)

[0176] The present invention provides a chelating ligand compound having a ligand skeleton that is friendly to nuclides with large atomic radius, so that heating is not required when chelating with radioactive metal nuclides, and the coordination effect in the chelation is strong, the metal complex formed is highly stable, and the radioactive metal nuclides are not easy to escape; at the same time, the present invention utilizes different functional groups capable of binding to target molecules to replace carboxyl groups of different numbers and positions, thereby forming chelating ligand compounds with different numbers and positions, which can improve the sites and abilities of binding to target molecules when binding to target molecules. In summary, the chelating ligand compound disclosed in the present invention, and the radiopharmaceutical prepared by using the compound formed by combining the chelating ligand compound with the target molecule and / or the complex formed by complexing with the metal nuclide, have better targeting and stability when used as a diagnostic imaging agent or when using the α and β rays generated by the nuclide to exert therapeutic effects.

[0177] Next, we will further illustrate the method by combining the example of preparing markers and the example of in vivo PET imaging experiments.

[0178] Example 14

[0179] The Ge-Ga generator was washed with 0.1M hydrochloric acid solution to obtain a solution containing radioactive nuclides. 68 Ga 3+eluent; add 1M sodium acetate solution to the eluent to adjust the pH value of the eluent to 4.0; add 20nM chelating ligand compound R1 to the eluent, shake well and react at room temperature for 10 minutes to obtain a radioactive nuclide Ga-68 labeled compound 68 Ga-R1.

[0180] Detection of compounds by instant thin layer chromatography 68 The radiochemical purity of Ga-R1 was determined by instant thin layer chromatography (TLC) under the following conditions: iTLC-SG chromatography plate; methanol / 1M ammonium acetate = 1 / 1 (v / v) as the developing agent. The results are shown in FIG8 . 68 The Ga-R1 product accounted for 95.9% of the total activity of the reaction solution, indicating that the chelating ligand compound R1 can be efficiently labeled at room temperature.

[0181] According to the above method, the chelating ligand compound R2, compound R3, and compound 28 are respectively 68 Ga 3+ Radiolabeling was performed in a Ge-Ga generator to obtain a compound labeled with the radionuclide Ga-68. 68 Ga-R2, compounds 68 Ga-R3.

[0182] Then, the compounds were determined by instant thin layer chromatography. 68 Ga-R2, compounds 68 The radiochemical purity of Ga-R3 is shown in Figures 9 and 10. According to the results shown in the figures, it can be seen that the chelating ligand compounds R2, R3 and 28 can all be efficiently labeled at room temperature.

[0183] According to the above method, the chelating ligand compound R4, compound R5, and compound R6 are respectively 68 Ga 3+ Radiolabeling was performed in a Ge-Ga generator to obtain a compound labeled with the radionuclide Ga-68. 68 Ga-R4, compound 68 Ga-R5 and compounds 68 Ga-R6.

[0184] Then, the compounds were determined by instant thin layer chromatography. 68 Ga-R4, compound 68 Ga-R5 and compounds 68 The radiochemical purity of Ga-R6 is shown in Figures 11 to 13. According to the results shown in the figures, it can be seen that the chelating ligand compounds R4, R5 and R6 can all be efficiently labeled at room temperature.

[0185] Each compound disclosed in the present invention can react with 68 Ga and other radionuclides are highly efficient markers and are not listed here one by one.

[0186] Example 15

[0187] Next, we take the coupling of compound R1 and compound W to prepare compound R1-4 as an example to illustrate the labeling of radionuclides in compounds with macrocyclic coupling targeting structures.

[0188] The Ge-Ga generator was washed with 0.1M hydrochloric acid solution to obtain a solution containing radioactive nuclides. 68 Ga 3+ eluent; add 1M sodium acetate solution to the eluent to adjust the pH value of the eluent to 4.2; add 20μM compound R1-4 to the eluent, shake well, and react at room temperature for 10 minutes, add 5mL water to dilute the reaction solution, and then purify it using Oasis HLB solid phase extraction cartridge, load the sample, rinse with water, and elute with 50% ethanol solution to obtain the product, the labeled compound 68 Ga-R1-4.

[0189] The product was analyzed by high performance liquid chromatography with a radioactive detector. The analysis conditions of the high performance liquid chromatography were as follows: Phenomenex Luna C18(2) column (5μ, 150×4.60mm); flow rate 1mL / min; gradient elution, acetonitrile 10% and trifluoroacetic acid solution (0.1%) 90% from 0 to 3 minutes, acetonitrile from 10% to 70% and trifluoroacetic acid solution from 90% to 30% from 3 to 10 minutes, acetonitrile from 70% to 10% and trifluoroacetic acid solution from 30% to 90% from 10 to 12 minutes, and acetonitrile maintained at 10% and trifluoroacetic acid solution maintained at 90% from 12 to 15 minutes. The results are shown in FIG14 , as can be seen. 68 The retention time of Ga-R1-4 was 7.88 minutes. The results showed that the radiochemical purity of the product was 97%.

[0190] According to the above method, use 68 Ga 3+ Label compound R1-5, compound R1-6, compound R1-7, compound R1-10, compound R1-11, and compound R1-12 to obtain labeled compounds 68 Ga-R1-5, 68 Ga-R1-6, 68 Ga-R1-7, 68 Ga-R1-10, 68 Ga-R1-11, 68 Ga-R1-12.

[0191] The labeled compounds were analyzed using high performance liquid chromatography with a radioactive detector, and the experimental results are shown in Figures 15-20.

[0192] It can be seen that the radiochemical purity of the labeled product is high.

[0193] It can be seen from the above radionuclide labeling examples that the macrocyclic coupling targeting compound structure disclosed in the present invention can be efficiently labeled at room temperature, and the obtained product has high radiochemical purity.

[0194] Each compound disclosed in the present invention can react with 68 Ga and other radionuclides are highly efficient markers and are not listed here one by one.

[0195] Example 16

[0196] Take the labeled macrocyclic coupling targeting compound 68 Ga-R1-4, 68 Ga-R1-6, 68 Ga-R1-12 was used to conduct PET phenomenon experiments.

[0197] 200 μC of radiochemically labeled compound 68 Ga-R1-4 was injected into the nude mouse model bearing U87MG tumor via tail vein. PET imaging data were collected for 10 minutes 60 minutes after injection. The results are shown in Figure 21, indicating that the compound 68 Ga-9 was obviously taken up and retained in U87MG tumor tissue (the position indicated by the arrow in the figure).

[0198] According to the above method, 200 μC of radiochemically labeled compound 68 Ga-R1-6, compound 68 Ga-R1-12 was injected into the nude mouse model bearing U87MG tumor through the tail vein, and the PET images shown in Figures 22 and 23 were obtained. It can be seen that the radiochemically labeled compound 68 Ga-R1-6, compound 68 Ga-R1-12 was obviously taken up and retained in U87MG tumor tissue (the position indicated by the arrow in the figure).

[0199] Example 17

[0200] Next we will 68 Ga-R1-4 is taken as an example to illustrate the pharmacokinetic data of the compound obtained in the present invention in animals.

[0201] Nine nude mice with U87gm tumors were randomly divided into three groups, with three mice in each group. 68Ga-R1-4 was injected into tumor-bearing mice via the tail vein, and then the nude mice were killed 30, 60, and 120 minutes after injection. The organs were separated, weighed, and the radioactivity count in each organ was measured to calculate the radioactivity uptake value per unit mass of tissue. 68 The distribution results of Ga-R1-4 in tumor-bearing mice are shown in Figure 24, indicating that the marker has a higher uptake and retention in the tumor, while the uptake in non-tumor tissues is lower, and the uptake of the drug in the tumor continues to increase, while in other non-tumor tissues it is gradually metabolized and shows a decreasing trend.

[0202] The above is a specific embodiment of the present invention. It should be noted that those skilled in the art can make several 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. Chelating ligand compound, characterized in that, The chelating ligand compound includes a chelating group containing multiple nitrogen atoms and at least one linking arm connected to the nitrogen atoms on the chelating group: Wherein: Z is a chelating group containing multiple nitrogen atoms and has the following structure: R1 and R2 are substituents on the benzene ring, and the substituent does not contain residues reactive with the target molecule; further preferably, R1 = R2; more preferably, both R1 and R2 are methyl groups; K is an alkyl chain containing a heteroatom O, S, P or N, composed of several R k connected by a heteroatom or a heteroatom-containing group, where R k is independently and optionally selected from substituted or unsubstituted alkyl; L is a linker moiety connected to the N atom in the chelating group Z, and the linker moiety L has a terminal group X connected to the target molecule or the targeting group.

2. The chelating ligand compound according to claim 1, wherein: The chelating ligand compound further includes a group Q, and the group Q is a heteroatom group with a lone pair of electrons; There are two ways of connecting the group Q to the chelating group Z, One is that the group Q is connected between the chelating group Z and the linking arm part L, Second, the group Q is directly connected to those N atoms in the chelating group Z that are not connected to the linker arm portion L.

3. The chelating ligand compound according to claim 1, characterized in that: The connecting arm portion L is selected from groups having the following formula: L m ——X, Wherein: X is —COOH, —OH, —CO, —SH, —CO—N(CH3)OH, —NH2, —H2PO3, —H2PO4, —H2PO2 or a halogen; L m is a substituted or unsubstituted alkyl group, preferably, L m is a substituted or unsubstituted C0-C3 alkyl group.

4. The chelating ligand compound according to claim 1, wherein R k selected from substituted or unsubstituted methyl, substituted or unsubstituted ethyl, and substituted or unsubstituted propyl.

5. The chelating ligand compound according to claim 1, characterized in that, The chelating ligand compound further has any of the following preferences, (1) The alkyl chain K containing a heteroatom O, S, P or N is selected from: any one of; (2) The group Q is selected from groups having the following formula, Q m -X Q ; Wherein: Q m is a substituted or unsubstituted alkyl group, preferably, Q m is a substituted or unsubstituted C0-C3 alkyl group; X Q is a heteroatom group with lone pair electrons, including but not limited to —COOH, —H2PO3, —H2PO4 or —H2PO2; (3) The chelating ligand compound is optionally selected from:

6. The derivative of the chelating ligand as described in claim 1, characterized in that, The derivative is a compound having the structure shown by the following formula or a salt formed by the compound and an inorganic acid or an organic acid, 7. The conjugate formed by coupling the chelating ligand compound according to any one of claims 1 to 5 with a target molecule or a targeting group.

8. The method for preparing the chelating ligand compound according to claim 1, characterized in that, This method is prepared from substituted or unsubstituted 2,6-diformylphenol dissolved in NH2—K—NH2, wherein the definition of K is as described in any one of claims 1-5.

9. The metal complex formed by the chelating ligand compound according to any one of claims 1 to 5 or the conjugate according to claim 7.

10. Use of the chelating ligand compound according to any one of claims 1 to 5, the derivative according to claim 6, the conjugate according to claim 7, and the metal complex according to claim 9 in the preparation of radiopharmaceuticals.

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