A fapi-46 derivative containing a hydrazine group nicotinamide group and its use

Abstract

The present application relates to the field of radiopharmaceutical chemistry and clinical nuclear medicine technology, and particularly relates to a FAPI-46 derivative containing a hydrazine-based nigroamide group and application thereof. A radioactive preparation obtained by labeling the FAPI-46 derivative containing the hydrazine-based nigroamide group with a radionuclide is simple to prepare, has high radiochemical purity, good stability, high uptake at a tumor site of a tumor-bearing mouse and a high target-to-non-target ratio, and has specific binding with fibroblast activation protein (FAP) in the tumor, and is a new type of tumor radioactive drug with promotional application value.

Description

Technical Field

[0001] This invention relates to the fields of radiopharmaceutical chemistry and clinical nuclear medicine, specifically to a FAPI-46 derivative containing a hydrazine-nicotinamide group and its applications. Background Technology

[0002] Fibroblast activating protein (FAP) is an important molecular marker on the surface of tumor-associated fibroblasts. Because it is selectively expressed on the cell membrane of fibroblasts in most solid tumor stroma, it has become an important target for tumor detection and treatment.

[0003] FAPI small molecule inhibitors based on (4-quinolineyl)-glycyl-2-cyanopyrrolidine exhibit high affinity and high selectivity. Through continuous structural modification and improvement, a series of... 68 Ga and 18 F-labeled FAPI complexes have been reported for tumor imaging in primary and metastatic lesions. However, these probes are all PET imaging agents, which are expensive and their clinical application is somewhat limited. 99m Tc, as the most common nuclide used in SPECT imaging, can be derived from... 99 Mo / 99m Tc generator rinsing and 99m Tc-labeled drugs can be prepared in kits, making them easy to promote and use in clinical practice. Therefore, the development of novel drugs targeting FAP is crucial. 99m Tc-based radiopharmaceuticals for tumors have significant practical implications.

[0004] at present 99m There are few reports on Tc-labeled molecular probes targeting FAP tumors, among which [ 99m Tc]Tc-FAPI-34 and [ 99m Tc][Tc-(CN-PEG4-FAPI)6] + It exhibits high affinity and high tumor uptake, but its high uptake in non-target organs in the abdomen somewhat limits its application. To develop superior SPECT-like tumor molecular probes targeting FAP, in 2021, we developed a... 99m Tc-labeled FAPI derivatives (patent number: ZL2021115355016) use D-proline-modified FAPI derivatives containing HYNIC groups (HYNIC-DP-FAPI) and synergistic ligands tris(hydroxymethylglycine) and sodium triphenylphosphine trisulfonate (TPPTS) with... 99m Tc coordination forms a stable 99mTc(HYNIC-DP-FAPI)(Tricine / TPPTS). This complex exhibits good in vitro and in vivo stability and FAP affinity. Biodistribution shows high uptake at the tumor site in tumor-bearing mice, and lower uptake in non-target organs such as the heart, liver, and muscle. However, due to high uptake in the blood, the tumor / blood ratio needs to be improved.

[0005] Linkers connect the targeting group and the chelating group attached to the radionuclide, playing a crucial role in regulating the affinity of radiopharmaceuticals and their biodistribution properties in vivo. To effectively address... 99m To address the issue of improving tumor uptake and tumor / blood ratio of Tc-labeled FAP-targeting tumor molecular probes, this invention modifies the structure of the pharmacophore of FAPI-46 to obtain ligands containing different linkers, which, along with the synergistic ligand Tricine / TPPTS, [further details needed]. 99m Tc coordination forms stable, highly stable, highly targeted, and well-balanced target-to-non-target ratio tumor molecular probes that target FAP. These probes are easy to deploy and can be used for early tumor diagnosis, staging, and efficacy evaluation, laying a solid foundation for the realization of translational medicine and precision medicine. Summary of the Invention

[0006] This invention provides a FAPI-46 derivative containing hydrazine-nicotinamide and its application. The derivative has good stability, is easy to prepare, and can be used for tumor diagnosis and treatment after radiolabeling. It exhibits high tumor uptake and a good target / non-target ratio, which has important scientific significance and application prospects in the field of tumor diagnosis and treatment.

[0007] Specifically, the present invention provides the following technical solutions:

[0008] A FAPI-46 derivative containing a hydrazine-nicotinamide group and its applications, wherein the structural formula is (I):

[0009]

[0010] In the formula, X is:

[0011] The corresponding derivative prepared from this derivative 99m Tc complexes specifically bind to FAP, exhibiting very low uptake in non-target organs, high tumor uptake values, and high tumor / blood and tumor / muscle ratios, thus achieving excellent results in tumor diagnosis and treatment.

[0012] The present invention also provides a radioactive preparation comprising the above-mentioned FAPI-46 derivative containing a hydrazine-nicotinamide group labeled with a radionuclide and its application.

[0013] Preferably, in the above-mentioned radioactive preparation, the radionuclide portion is a metallic radionuclide.

[0014] Preferably, in the above-mentioned radioactive agents, the metallic radionuclide is... 99m Tc, 99 Tc, 94m Tc, 94 Tc, 52 Mn, 186 Re or 188 Re.

[0015] Most preferably, in the above-mentioned radioactive agent, the radionuclide is... 99m Tc, the structural formula of the radioactive agent is (II):

[0016]

[0017] In the formula, X is:

[0018] The present invention also provides the use of the above-mentioned radioactive agents in the preparation of radiopharmaceuticals for tumors.

[0019] The beneficial effects of this invention are as follows: This invention provides a FAPI-46 derivative containing hydrazine nicotinamide and its application. The radioactive preparation obtained by labeling it with a radionuclide has high uptake in tumors and a good tumor / non-target ratio, which is a novel radiopharmaceutical with promotional significance. Detailed Implementation

[0020] This invention provides a FAPI-46 derivative containing a hydrazine-nicotinamide group and its applications. In a preferred embodiment, this invention provides a derivative with the general structural formula [ 99m The radioactive preparation of Tc-(HYNIC-X-F46)(Tricine / TPPTS):

[0021]

[0022] In this structural formula: the nitrogen atom on the hydrazine group in the HYNIC-X-F46 molecule, the phosphorus atom in the co-ligand TPPTS, and the oxygen and nitrogen atoms in Tricine are... 99m Tc coordination yields [ 99m [Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex. X represents different linkers, X =

[0023] The preparation steps are as follows:

[0024] a: Synthesis of ligand HYNIC-X-F46:

[0025] 6-Chloronicotinic acid (compound 1) was dissolved in 80% hydrazine hydrate and stirred at 100°C for 4 h. Part of the solvent was removed under reduced pressure. Deionized water was added to the reaction flask to adjust the pH to 5.0, and then the mixture was filtered. The filter cake was washed and dried under vacuum to obtain compound 2. Compound 2 and sodium 2-formylbenzenesulfonate were dissolved in N,N-dimethylformamide (DMF) and stirred at room temperature for 3 h. N-hydroxysuccinimide (NHS) and dicyclohexylcarbodiimide (DCC) were added to the mixture and stirred at room temperature for 18 h. The filtrate was then concentrated. Ethyl acetate (EA) was added, and the mixture was heated under reflux for 1 h and filtered. The filter cake was dried under vacuum to obtain compound 3. Compound 3 and compound 4 were added to a reaction flask, dissolved in DMF, and then triethylamine (TEA) was added. The mixture was reacted at 110°C for 6–15 h. Column chromatography purification yielded compound 5. An appropriate amount of compound 6 was weighed into a round-bottom flask, dissolved in DMF, and then compound 5, 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI), 1-hydroxybenzotriazole (HOBT), and N-ethyldiisopropylamine (DIPEA) were added sequentially. The reaction was carried out at room temperature for 6-8 hours. The solvent was evaporated, and the mixture was purified by column chromatography to obtain HYNIC-X-F46. The specific synthetic route is as follows:

[0026]

[0027] Among them, compound 4 is as follows: X represents different binders.

[0028] b:[ 99m Preparation of the Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex:

[0029] Weigh appropriate amounts of Tricine, TPPTS, and HYNIC-X-F46 ligand and dissolve them in physiological saline. Adjust the pH of the solution to 5.0, and add an appropriate amount of freshly rinsed Na+. 99m TcO4 can be obtained by heating in a boiling water bath for 30 minutes. 99m Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex.

[0030] The series prepared by the above method 99mThe [Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex has a radiochemical purity greater than 95%, is hydrophilic, and exhibits good in vitro stability. It shows high uptake and good retention at tumor sites in tumor-bearing mice. Inhibition by FAP inhibitors significantly reduces tumor uptake, indicating that its uptake in tumors is specific to FAP. Imaging results show significant concentration at tumor sites, low uptake in non-target tissues, and that tumor uptake can be significantly inhibited by FAP inhibitors. Therefore, it is a high-performance novel SPECT molecular probe for tumor imaging.

[0031] The following embodiments are used to illustrate the present invention, but are not intended to limit the scope of the invention. Where specific techniques or conditions are not specified in the embodiments, they should be performed in accordance with the techniques or conditions described in the literature in the field, or in accordance with the product manual.

[0032] Example 1

[0033] This embodiment provides a FAPI-46 derivative containing a hydrazine-nicotinamide group and its application, referred to as […]. 99m Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS), the structural formula is as follows:

[0034]

[0035] In this structural formula: the nitrogen atom on the hydrazine group in the HYNIC-X-F46 molecule, the phosphorus atom in the co-ligand TPPTS, and the oxygen and nitrogen atoms in Tricine are... 99m Tc coordination yields [ 99m [Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex. X represents different linkers, X =

[0036] The preparation methods are as follows, but are not limited to the exemplified complexes:

[0037] 1.[ 99m Preparation of Tc-(HYNIC-DPro-F46)(Tricine / TPPTS)

[0038] Synthesis of a.HYNIC-DPro-F46

[0039] Synthesis of Compound 2. Compound 1 (1.57 g, 10 mmol) was dissolved in 80% hydrazine hydrate (10 mL). The solution was then stirred at 100 °C for 4 h, cooled to room temperature, and partially removed under reduced pressure. 20 mL of deionized water was added to the reaction flask, and the pH was adjusted to 5.0 with concentrated hydrochloric acid to form a precipitate, which was then filtered. The filter cake was washed with 95% ethanol and diethyl ether, and dried under vacuum to give a yellow solid, which was Compound 2 (1.30 g, 85%). 1 H NMR (400MHz, DMSO-d6): δ8.50 (d, J = 2.3Hz, 1H), 7.83 (dd, J = 8.9, 2.3Hz, 1H), 6.68 (d, J = 9.0Hz, 1H).

[0040] Synthesis of Compound 3. Compound 2 (1.28 g, 8 mmol) and sodium 2-formylbenzenesulfonate (0.87 g, 4 mmol) were dissolved in DMF (10 mL) and stirred at room temperature for 3 h. Then, NHS (0.46 g, 4 mmol) and DCC (1.60 g, 8 mmol) were added to the mixture. After stirring at room temperature for 18 h, the reaction mixture was filtered to remove insoluble solids, and the filtrate was concentrated. EA (50 mL) was added to the concentrate, and the mixture was heated under reflux for 1 h, followed by filtration. The filter cake was dried under vacuum to give a pale yellow solid, which was Compound 3 (1.33 g, 76%). 1 H NMR (600MHz, DMSO-d6): δ11.86(s,1H),9.13(s,1H),8.76(d,J=2.3Hz,1H),8.14(dd,J=9.0,2.4 Hz,1H),8.00(dd,J=7.6,1.5Hz,1H),7.77(dd,J=7.6,1.6Hz,1H)7.36-7.31(m,3H),2.69(s,4H).

[0041] Synthesis of compound 5A. Compound 3 (1.611 g, 3.7 mmol) and compound 4A (0.421 g, 3.7 mmol) were added to a 50 mL reaction flask, dissolved in 10 mL of DMF, and then TEA (2.590 mL, 18.5 mmol) was added. The reaction was carried out at 110 °C for 9 h. After cooling to room temperature, the solvent was removed by rotary evaporation, and the mixture was purified by column chromatography [DCM / MeOH = 2 / 1 (v / v)] to give a yellow solid, which was compound 5A (0.971 g, 60%). 1H NMR (400MHz, D2O): δ8.99(d,J=10.2Hz,1H),8.36(s,1H),8.23(d,J=8.4Hz,1H),7.95(t,J=8.1Hz,2H),7.46(t,J=7.4Hz,1H),7.37(t ,J=7.6Hz,1H),7.26(t,J=7.8Hz,1H),4.55(t,J=6.8Hz,1H),3.71(q,J=7.4Hz,2H),2.36(h,J=11.8,11.1Hz,1H),2.16-1.80(m,3H).

[0042] Synthesis of compound HYNIC-DPro-F46. Compound 6 (50 mg, 0.10 mmol) was weighed into a round-bottom flask, dissolved in 2 mL of DMF, and then compound 5A (53 mg, 0.12 mmol), EDCI (38 mg, 0.20 mmol), HOBT (14 mg, 0.10 mmol), and DIPEA (0.17 mL, 1.00 mmol) were added sequentially. The reaction was carried out at room temperature for 8 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the mixture was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to obtain the ligand HYNIC-DPro-F46 (32 mg, 33%). 1 H NMR (400MHz, Methanol-d4) δ8.91 (d, J=6.5Hz, 1H), 8.49 (dd, J=12.8, 4.5Hz, 1H), 8.38 (s, 1H), 8.20 (t, J=6. 9Hz, 1H), 7.97-7.91 (m, 1H), 7.87 (d, J=8.9Hz, 2H), 7.71 (dd, J=59.1, 8.8Hz, 1H), 7.53 (d, J=9.5Hz, 1H), 7.48 -7.38(m,3H),7.39-7.26(m,3H),5.11(d,J=9.3Hz,1H),5.00(d,J=7.2Hz,1H),4.33-4.03(m,4H),3.66(d,J=40.3Hz, 8H),3.19(td,J=7.3,6.9,2.4Hz,2H),3.16-3.05(m,4H),2.59(s,3H),2.33(s,2H),2.02-1.81(m,6H); HR-MS(ESI)for C 43 H 46 F2N 11 O7S[M-Na] - :found 898.3270,calcd 898.3275.

[0043] The synthetic route is as follows:

[0044]

[0045] b.[ 99m Preparation of the Tc]Tc-(HYNIC-DPro-F46)(Tricine / TPPTS) complex

[0046] Weigh 1 mg Tricine and 2 mg TPPTS and dissolve them in 0.5 mL of physiological saline. Adjust the pH of the solution to 5.0 with succinate buffer (pH 5.0). Then, add 20 μg of HYNIC-DPro-F46 ligand and 0.5 mL of freshly rinsed […]. 99m The [Tc]NaTcO4 (approximately 370 MBq) can be obtained by heating in a boiling water bath for 30 minutes. 99m Tc]Tc-(HYNIC-DPro-F46)(Tricine / TPPTS) complex.

[0047] 2.[ 99m Preparation of Tc-(HYNIC-DAla-F46)(Tricine / TPPTS)

[0048] Synthesis of a.HYNIC-DAla-F46

[0049] The synthesis of compound 3 is consistent with the synthesis method of compound 3 in HYNIC-DPro-F46.

[0050] Synthesis of compound 5B. Compound 3 (0.881 g, 2 mmol) and compound 4B (0.178 g, 2 mmol) were added to a reaction flask, dissolved in 5 mL of DMF, and then TEA (1.37 mL, 10 mmol) was added. The reaction was carried out at 110 °C for 6 h. After cooling to room temperature, the solvent was removed by rotary evaporation, and the mixture was purified by column chromatography [DCM / MeOH = 5 / 1 (v / v)] to give a pale yellow solid, which was compound 5B (0.510 g, 62%). 1 H NMR(600MHz,D2O)δ8.61(s,1H),8.25(d,J=2.2Hz,1H),7.95-7.88(m,2H),7.65(d,J=7.8Hz,1H),7.37(t,J= 7.5Hz,1H),7.28(q,J=7.2,6.7Hz,1H),6.91(d,J=9.0Hz,1H),4.10(q,J=7.2Hz,1H),1.27(d,J=7.3Hz,3H).

[0051] Synthesis of compound HYNIC-DAla-F46. Compound 6 (50 mg, 0.10 mmol) was weighed into a round-bottom flask, dissolved in 2 mL of DMF, and then compound 5B (50 mg, 0.12 mmol), EDCI (38 mg, 0.20 mmol), HOBT (14 mg, 0.10 mmol), and DIPEA (0.17 mL, 1.00 mmol) were added sequentially. The reaction was carried out at room temperature for 7 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the ligand HYNIC-DAla-F46 (44 mg, 49%) was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to obtain HYNIC-DAla-F46. 1 H NMR (400MHz, Methanol-d4) δ8.91(s,1H),8.62-8.57(m,1H),8.51(d,J=4.5Hz,1H),8.22(d,J=8.0Hz,1H),8.07(d d,J=8.9,2.4Hz,1H),7.93(dd,J=7.9,1.3Hz,2H),7.87(d,J=9.4Hz,1H),7.52(dd,J=9.4,2.8Hz,1H),7.47-7.42(m ,3H),7.41-7.34(m,1H),5.12(d,J=9.0Hz,1H),4.98(d,J=6.9Hz,1H),4.27(s,2H),4.17(d,J=44.1Hz,2H),3.83( s,2H),3.63-3.53(m,4H),3.11(s,5H),2.78(d,J=12.2Hz,6H),2.28(t,J=6.5Hz,3H),1.95(s,2H).HR-MS(ESI)for C 41 H 45 F2N 11 NaO7S[M+H] + :found 896.3088,calcd 896.3084.

[0052] The synthetic route is as follows:

[0053]

[0054] b.[ 99m Preparation of the Tc]Tc-(HYNIC-DAla-F46)(Tricine / TPPTS) complex

[0055] Weigh 1 mg Tricine and 2 mg TPPTS and dissolve them in 0.5 mL of physiological saline. Adjust the pH of the solution to 5.0 with succinate buffer (pH 5.0). Then, add 20 μg of HYNIC-DAla-F46 ligand and 0.5 mL of freshly rinsed […]. 99m The [Tc]NaTcO4 (approximately 370 MBq) can be obtained by heating in a boiling water bath for 30 minutes. 99m Tc]Tc-(HYNIC-DAla-F46)(Tricine / TPPTS) complex.

[0056] 3.[ 99m Preparation of Tc]Tc-(HYNIC-Gly-F46)(Tricine / TPPTS

[0057] Synthesis of a.HYNIC-Gly-F46

[0058] The synthesis of compound 3 is consistent with the synthesis method of compound 3 in HYNIC-DPro-F46.

[0059] Synthesis of compound 5C. Compound 3 (0.881 g, 2 mmol) and compound 4C (0.150 g, 2 mmol) were added to a 50 mL reaction flask, dissolved in 5 mL of DMF, and then TEA (1.37 mL, 10 mmol) was added. The reaction was carried out at 110 °C for 6 h. After cooling to room temperature, the solvent was removed by rotary evaporation, and the mixture was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to give a brown solid, which was compound 5C (0.385 g, 48%). 1 H NMR(600MHz,D2O)δ8.64(s,1H),8.36(d,J=2.4Hz,1H),8.03(d,J=7.7Hz,1H),7.92(dd,J=8.9,2.4Hz,1H),7.73 (dd,J=7.9,1.3Hz,1H),7.44(t,J=7.5Hz,1H),7.34(td,J=7.6,1.3Hz,1H),7.03(d,J=8.9Hz,1H),3.75(s,2H).

[0060] Synthesis of compound HYNIC-Gly-F46. Compound 6 (50 mg, 0.10 mmol) was weighed into a round-bottom flask, dissolved in 2 mL of DMF, and then compound 5C (48 mg, 0.12 mmol), EDCI (38 mg, 0.20 mmol), HOBT (14 mg, 0.10 mmol), and DIPEA (0.17 mL, 1.00 mmol) were added sequentially. The reaction was carried out at room temperature for 8 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the ligand HYNIC-Gly-F46 (51 mg, 58%) was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to obtain HYNIC-Gly-F46. 1 H NMR (400MHz, Methanol-d4) δ8.92 (s, 1H), 8.69-8.61 (m, 1H), 8.50 (d, J = 4.3Hz, 1H), 8.22 (d ,J=7.7Hz,1H),8.08(dd,J=8.9,2.4Hz,1H),7.97-7.91(m,2H),7.86(d,J=4.5Hz,1H),7.56 -7.51(m,1H),7.44(d,J=3.7Hz,3H),7.36(d,J=7.7Hz,1H),5.11(d,J=9.8Hz,1H),4.26(d, J=4.5Hz, 4H), 3.59 (d, J=16.3Hz, 8H), 2.58 (s, 11H), 2.28 (t, J=6.4Hz, 2H); HR-MS (ESI) for C 40 H 43 F2N 11 NaO7S[M+H] + :found 882.2907,calcd 882.2927.

[0061] The synthetic route is as follows:

[0062]

[0063] b.[ 99m Preparation of the Tc]Tc-(HYNIC-Gly-F46)(Tricine / TPPTS) complex

[0064] Weigh 1 mg Tricine and 2 mg TPPTS and dissolve them in 0.5 mL of physiological saline. Adjust the pH of the solution to 5.0 with succinate buffer (pH 5.0). Then, add 20 μg of HYNIC-Gly-F46 ligand and 0.5 mL of freshly rinsed […]. 99m The [Tc]NaTcO4 (approximately 370 MBq) can be obtained by heating in a boiling water bath for 30 minutes. 99mTc]Tc-(HYNIC-Gly-F46)(Tricine / TPPTS) complex.

[0065] 4.[ 99m Preparation of Tc]Tc-(HYNIC-Gly2-F46)(Tricine / TPPTS

[0066] Synthesis of a.HYNIC-Gly2-F46

[0067] The synthesis of compound 3 is consistent with the synthesis method of compound 3 in HYNIC-DPro-F46.

[0068] Synthesis of compound 5D. Compound 3 (0.881 g, 2 mmol) and compound 4D (0.264 g, 2 mmol) were added to a 50 mL reaction flask, dissolved in 5 mL of DMF, and then TEA (1.37 mL, 10 mmol) was added. The reaction was carried out at 110 °C for 6 h. After cooling to room temperature, the solvent was removed by rotary evaporation, and the mixture was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to give a yellow solid, which was compound 5D (0.424 g, 46%). 1 H NMR (600MHz, Methanol-d4) δ8.94(d,J=14.4Hz,1H),8.66(dd,J=2.3,1.2Hz,1H),8.22(d,J=7.8Hz,1H),8.16-8.08(m,1 H),7.93(dt,J=7.8,1.2Hz,1H),7.45(t,J=7.2Hz,1H),7.38-7.26(m,2H),4.08(d,J=13.3Hz,2H),3.81(d,J=1.3Hz,2H).

[0069] Synthesis of compound HYNIC-Gly2-F46. Compound 6 (50 mg, 0.10 mmol) was weighed into a round-bottom flask and dissolved in 2 mL of DMF. Then, compound 5D (55 mg, 0.12 mmol), EDCI (38 mg, 0.20 mmol), HOBT (14 mg, 0.10 mmol), and DIPEA (0.17 mL, 1.00 mmol) were added sequentially, and the mixture was reacted at room temperature for 8 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the mixture was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to give ligand HYNIC-Gly2-F46 (44 mg, 47%). 1H NMR (600MHz, Methanol-d4) δ8.93(s,1H),8.66(d,J=2.4Hz,1H),8.48(dd,J=12.2,4.5Hz,1H),8.22(d,J=7.8Hz,1H),8.09(dd,J=9.1,2 .6Hz,1H),7.92(dd,J=14.5,7.8Hz,1H),7.83(dd,J=16.6,9.4Hz,1H),7.52(dd,J=9.4,2.9Hz,1H),7.47-7.39(m,3H),7.37-7.30(m,2H) ,5.11-5.08(m,1H),4.29-4.21(m,2H),4.08(d,J=14.8Hz,4H),3.90(d,J=11.6Hz,2H),3.48(s,2H),3.32(d,J=0.8Hz,2H),2.76(s,1H) ,2.49-2.39(m,4H),2.34(d,J=7.7Hz,2H),2.13(d,J=0.7Hz,3H),1.89-1.80(m,2H),1.64(p,J=7.0Hz,2H),1.26(s,1H); HR-MS(ESI)for C 42 H 47 F2N 12 O8S[M+2H-Na] + :found 917.3245,calcd 917.3323.

[0070] The synthetic route is as follows:

[0071]

[0072] b.[ 99m Preparation of the Tc]Tc-(HYNIC-Gly2-F46)(Tricine / TPPTS) complex

[0073] Weigh 1 mg Tricine and 2 mg TPPTS and dissolve them in 0.5 mL of physiological saline. Adjust the pH of the solution to 5.0 with succinate buffer (pH 5.0). Then, add 20 μg of HYNIC-Gly2-F46 ligand and 0.5 mL of freshly rinsed […]. 99m The [Tc]NaTcO4 (approximately 370 MBq) can be obtained by heating in a boiling water bath for 30 minutes. 99m Tc]Tc-(HYNIC-Gly2-F46)(Tricine / TPPTS) complex.

[0074] 5.[ 99mPreparation of Tc-(HYNIC-PG2-F46)(Tricine / TPPTS)

[0075] Synthesis of a.HYNIC-PG2-F46

[0076] The synthesis of compound 3 is consistent with the synthesis method of compound 3 in HYNIC-DPro-F46.

[0077] Synthesis of compound 5E. Compound 3 (0.645 g, 1.5 mmol) and compound 4E (0.500 g, 1.5 mmol) were added to a 50 mL reaction flask, dissolved in 8 mL of DMF, and then TEA (1 mL, 7.2 mmol) was added. The reaction was carried out at 110 °C for 15 h. After cooling to room temperature, the solvent was removed by rotary evaporation, and the mixture was purified by column chromatography [DCM / MeOH = 3 / 1 (v / v)] to give a yellow solid, which was compound 5E (0.490 g, 61%). 1 H NMR (600MHz, Methanol-d4): δ8.93(d,J=13.0Hz,1H),8.45(s,1H),8.20(t,J=10.4Hz ,1H),7.92(t,J=7.6Hz,1H),7.45(t,J=7.4Hz,2H),7.34(dd,J=16.4,9.0Hz,1H),7.30 (d,J=8.1Hz,1H),4.57(d,J=27.9Hz,1H),4.45(d,J=6.5Hz,1H),4.08(dd,J=43.6,16 .9Hz,2H),3.82(d,J=10.2Hz,2H),3.69(s,2H),3.60-3.51(m,2H),2.16-1.96(m,8H).

[0078] Synthesis of compound HYNIC-PG2-F46. Compound 6 (50 mg, 0.10 mmol) was weighed into a round-bottom flask, dissolved in 2 mL of DMF, and then compound 5E (65 mg, 0.12 mmol), EDCI (38 mg, 0.20 mmol), HOBT (14 mg, 0.10 mmol), and DIPEA (0.17 mL, 1.00 mmol) were added sequentially. The reaction was carried out at room temperature for 7 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the ligand HYNIC-PG2-F46 (33 mg, 29%) was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to obtain HYNIC-PG2-F46. 1H NMR (600MHz, Methanol-d4) δ8.92(s,1H),8.49-8.46(m,1H),8.19(d,J=7.9Hz,1H),7.91(d,J=7.9 Hz,2H),7.87-7.79(m,1H),7.47-7.40(m,3H),7.32(td,J=18.2,17.1,10.2Hz,3H),5.21-5.03(m, 2H),4.55(s,4H),4.24(dd,J=25.2,8.4Hz,3H),4.05-3.93(m,3H),3.56(d,J=14.3Hz,8H),3.00(s ,2H),2.92(d,J=7.5Hz,3H),2.48-2.29(m,8H),2.04(s,6H),1.81(p,J=6.8Hz,4H); HR-MS(ESI)for C 52 H 60 F2N 14 NaO 10 S[M+H] + :found 1133.4202,calcd1133.4197.

[0079] The synthetic route is as follows:

[0080]

[0081] b.[ 99m Preparation of the Tc]Tc-(HYNIC-PG2-F46)(Tricine / TPPTS) complex

[0082] Weigh 1 mg Tricine and 2 mg TPPTS and dissolve them in 0.5 mL of physiological saline. Adjust the pH of the solution to 5.0 with succinate buffer (pH 5.0). Then add 20 μg of HYNIC-PG2-F46 ligand and 0.5 mL of freshly rinsed […]. 99m The [Tc]NaTcO4 (approximately 370 MBq) can be obtained by heating in a boiling water bath for 30 minutes. 99m Tc]Tc-(HYNIC-PG2-F46)(Tricine / TPPTS) complex.

[0083] 6.[ 99m Preparation of Tc-(HYNIC-PEG-F46)(Tricine / TPPTS)

[0084] Synthesis of HYNIC-PEG-F46

[0085] The synthesis of compound 3 is consistent with the synthesis method of compound 3 in HYNIC-DPro-F46.

[0086] Synthesis of compound 5F. Compound 3 (0.881 g, 2 mmol) and compound 4F (0.530 g, 2 mmol) were added to a 50 mL reaction flask, dissolved in 10 mL of DMF, and then TEA (2.590 mL, 18.5 mmol) was added. The reaction was carried out at 110 °C for 6 h. After cooling to room temperature, the solvent was removed by rotary evaporation, and the mixture was purified by column chromatography [DCM / MeOH = 5 / 1 (v / v)] to give a pale yellow solid, which was compound 5F (0.660 g, 48%). 1 H NMR (400MHz, Methanol-d4) δ8.99-8.92(m,1H),8.64(t,J=3.1Hz,1H),8.23(dd,J=7.9,1.4Hz,1H),8.11-8.05(m,1H),7.93(dd ,J=7.8,1.4Hz,1H),7.46(td,J=7.7,1.4Hz,1H),7.41-7.30(m,2H),3.66-3.62(m,6H),3.62-3.56(m,12H),2.88-2.82(m,2H).

[0087] Synthesis of compound HYNIC-PEG-F46. Compound 6 (50 mg, 0.10 mmol) was weighed into a round-bottom flask, dissolved in 2 mL of DMF, and then compound 5F (53 mg, 0.12 mmol), EDCI (38 mg, 0.20 mmol), HOBT (14 mg, 0.10 mmol), and DIPEA (0.17 mL, 1.00 mmol) were added sequentially. The reaction was carried out at room temperature for 7 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the ligand HYNIC-PEG-F46 (56 mg, 52%) was purified by column chromatography [DCM / MeOH = 4 / 1 (v / v)] to obtain HYNIC-PEG-F46. 1H NMR (600MHz, Methanol-d4) δ8.93(d,J=6.9Hz,1H),8.62(s,1H),8.47(d,J=4.4Hz,1H),8.21( dd,J=16.4,7.9Hz,1H),8.06-8.01(m,1H),7.93(t,J=7.7Hz,1H),7.82(d,J=9.5Hz,1H),7.50- 7.40(m,3H),7.39-7.26(m,3H),5.10(d,J=9.1Hz,1H),4.56(s,2H),4.31-4.07(m,4H),3.63- 3.54(m,22H),3.47(s,2H),3.04(s,4H),2.48-2.32(m,8H),1.83-1.76(m,2H); HR-MS(ESI)for C 49 H 61 F2N 11 NaO 11 S[M+H] + :found 1072.4129,calcd 1072.4133.

[0088] The synthetic route is as follows:

[0089]

[0090] b.[ 99m Preparation of the Tc-(HYNIC-PEG-F46)(Tricine / TPPTS) complex

[0091] Weigh 1 mg Tricine and 2 mg TPPTS and dissolve them in 0.5 mL of physiological saline. Adjust the pH of the solution to 5.0 with succinate buffer (pH 5.0). Then, add 20 μg of HYNIC-PEG-F46 ligand and 0.5 mL of freshly rinsed […]. 99m The [Tc]NaTcO4 (approximately 370 MBq) can be obtained by heating in a boiling water bath for 30 minutes. 99m Tc]Tc-(HYNIC-PEG-F46)(Tricine / TPPTS) complex.

[0092] This invention [ 99m Performance determination of the Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex:

[0093] 1. Identification of coordination compounds

[0094] [ 99mTc-(HYNIC-X-F46) (Tricine / TPPTS) was identified using high-performance liquid chromatography (HPLC): a C18 reverse-phase column was used in an SCL-10AVP HPLC system. Phase A was water (containing 0.1% trifluoroacetic acid), and phase B was acetonitrile (containing 0.1% trifluoroacetic acid). The gradient was as follows: 0-2 min, phase B 10%; 2-10 min, phase B changed from 10% to 90%; 10-15 min, phase B 90%; 15-20 min, phase B changed from 90% to 10%; 20-30 min, phase B 10%. The injection volume was 20 μL, and the flow rate was 1 mL / min. The retention time (R) was determined. t )for:[ 99m Tc]Tc-(HYNIC-DPro-F46)(Tricine / TPPPTS):10.21min;[ 99m Tc]Tc-(HYNIC-DAla-F46)(Tricine / TPPPTS):10.83min;[ 99m Tc]Tc-(HYNIC-Gly-F46)(Tricine / TPPPTS):10.71min;[ 99m Tc]Tc-(HYNIC-Gly2-F46)(Tricine / TPPPTS):10.20min;[ 99m Tc]Tc-(HYNIC-PG2-F46)(Tricine / TPPPTS):10.43min;[ 99m Tc]Tc-(HYNIC-PEG-F46)(Tricine / TPPPTS):10.88min.

[0095] 2. Determination of the lipid-water partition coefficient of the complex

[0096] Take 0.9 mL of pH 7.4 phosphate buffer (0.025 mol / L) into a 5 mL centrifuge tube, add 1 mL of n-octanol and 0.1 mL of […]. 99m A Tc-(HYNIC-X-F46)(Tricine / TPPTS) solution was prepared, capped, vortexed, and centrifuged for 5 min (5000 r / min). Then, 3 × 0.1 mL samples were taken from both the organic and aqueous phases, and the radioactivity counts of the two phases were measured. The partition coefficient D (D = radioactivity of organic phase / radioactivity of aqueous phase) was calculated. This process was repeated five times. The measured […] 99m The lipid-water partition coefficient (logD) of the Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex is negative, indicating that it is a hydrophilic substance.

[0097] Table 1 [99m The lipid-water partition coefficient of Tc-(HYNIC-X-F46)(Tricine / TPPTS)

[0098]

[0099] 3. Stability determination of complexes

[0100] The radiochemical purity of the complexes was determined after 4 hours of incubation at room temperature and in mouse serum at 37°C. The results showed that six […]. 99m The Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) complex exhibited radiochemical purity greater than 90% after being placed in mouse serum at room temperature and 37°C for 4 hours, indicating good in vitro stability.

[0101] 4. Biodistribution experiment of the complex in tumor-bearing mice

[0102] 0.10 mL was injected via the tail vein into a Balb / c mouse model bearing U87MG tumor. 99m Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) labeled solution (approximately 3.7 × 10⁻⁶) 5 Bq), mice were anesthetized with isoflurane gas one hour after injection and then sacrificed. Additionally, FAPI was used to […]. 99m In vivo inhibition experiments were conducted on mice using [Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS], as follows: 0.10 mL of physiological saline containing 100 μg FAPI and 0.10 mL of […]. 99m Tc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) labeled solution (approximately 3.7 × 10⁻⁶) 5 Mice were injected via tail vein with Bq, and euthanized one hour later after being anesthetized with isoflurane gas. Relevant tissues and organs, including heart, liver, lungs, kidneys, spleen, bones, small intestine, large intestine, pancreas, stomach, muscles, blood, and tumors, were collected, cleaned, weighed, and their radioactivity counts were measured using a γ-counter. The percentage injection dose per gram (%ID / g) for each tissue was calculated. Four mice were used for each time item. The results are shown in Tables 2-7.

[0103] Table 2 [ 99m Biodistribution (%ID / g) of Tc-(HYNIC-DPro-F46)(Tricine / TPPTS) in Balb / c nude mice bearing U87MG tumors

[0104]

[0105] Table 3 [ 99mBiodistribution (%ID / g) of Tc-(HYNIC-DAla-F46)(Tricine / TPPTS) in Balb / c nude mice bearing U87MG tumors

[0106]

[0107]

[0108] Table 4 [ 99m Biodistribution (%ID / g) of Tc-(HYNIC-Gly-F46)(Tricine / TPPTS) in Balb / c nude mice bearing U87MG tumors

[0109]

[0110] Table 5 [ 99m Biodistribution (%ID / g) of Tc-(HYNIC-Gly2-F46)(Tricine / TPPTS) in Balb / c nude mice bearing U87MG tumors

[0111]

[0112] Table 6 [ 99m Biodistribution (%ID / g) of Tc-(HYNIC-PG2-F46)(Tricine / TPPTS) in Balb / c nude mice bearing U87MG tumors

[0113]

[0114]

[0115] Table 7 [ 99m Biodistribution (%ID / g) of Tc-(HYNIC-PEG-F46)(Tricine / TPPTS) in Balb / c nude mice bearing U87MG tumors

[0116]

[0117] 5. SPECT imaging of the complex in tumor-bearing mice

[0118] Injected via tail vein into a Balb / c mouse model of U87MG tumor. 99m 0.2 mL (approximately 37 MBq) of Tc-(HYNIC-X-F46)(Tricine / TPPTS) solution was administered, followed by isoflurane gas anesthesia 1 hour later. The inhibition group received 0.10 mL of physiological saline containing 100 μg FAPI and 0.10 mL of […]. 99mTc]Tc-(HYNIC-X-F46)(Tricine / TPPTS) labeled solution (approximately 3.7 × 10⁻⁶) 5 Bq) was injected via the tail vein, and one hour later, mice were anesthetized with isoflurane gas. The mice were then fixed in a prone position and SPECT / CT was used for imaging. SPECT imaging results showed that in the experimental group... 99m Tc-(HYNIC-X-F46)(Tricine / TPPTS) showed significant concentration in tumors, while tumor uptake was significantly reduced in the inhibition group, further demonstrating its specificity in tumor uptake and indicating that it can serve as a novel SPECT molecular probe with excellent tumor affinity.

[0119] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, these modifications or improvements made without departing from the spirit of the present invention involve changing different linking agents, such as amino acids, peptide chains, polyethylene glycol (PEG) chains, fatty acid chains, etc., or utilizing N-tris(hydroxymethyl)methylglycine (Tricine) and ethylenediamine-N,N'-diacetic acid (EDDA), N-tris(hydroxymethyl)methylglycine (Tricine) and sodium diphenylphosphine-3-sulfonate (TPPMS), N-tris(hydroxymethyl)methylglycine (Tricine) and disodium 3,3'-(phenylphosphinediyl)bis(phenyl-1-sulfonic acid) (TPPDS), N-tris(hydroxymethyl)methylglycine (Tricine) and nicotinic acid (NIC), N-tris(hydroxymethyl)methylglycine... Radioactive preparations obtained by radiolabeling coligands such as tricine and isonicotinic acid, N-tris(hydroxymethyl)methylglycine and 3,5-pyridinedicarboxylic acid (PDA), N-tris(hydroxymethyl)methylglycine and 3-pyridinesulfonic acid (PSA), N-tris(hydroxymethyl)methylglycine and glucohepanoate, N-tris(hydroxymethyl)methylglycine and glucosamine, N-tris(hydroxymethyl)methylglycine and mannitol, and N-tris(hydroxymethyl)methylglycine and diphenylphosphine benzoic acid with radioactive nuclides are all within the scope of protection claimed in this invention.

Claims

1. A FAPI-46 derivative containing a hydrazinonicotinamide group, characterized in that, The FAPI-46 derivative containing a hydrazine group nicotinamide group has the structural formula (I): wherein X is:

2. A radiopharmaceutical formulation, characterized in that, The radioactive preparation comprises the FAPI-46 derivative containing a hydrazine group nicotinamide group according to claim 1, which is labeled with a radionuclide.

3. The radiopharmaceutical of claim 2, wherein, The radionuclide is 99m Tc, 99 Tc, 94m Tc, 94 Tc, 52 Mn, 186 Re or 188 Re.

4. The radiopharmaceutical of claim 3, wherein, The radioactive preparation has the structural formula (II): In the formula, X is as defined in claim 1.

5. Use of the radioactive preparation according to any one of claims 2-4 in the preparation of a tumor imaging agent.

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