Compound, chelate and use thereof in magnetic resonance imaging

Abstract

A compound, a chelate and the use thereof in magnetic resonance imaging, belonging to the technical field of medical imaging. Specifically disclosed is a compound as represented by general formula (I) or a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a mixture thereof, which can significantly increase plasma exposure, has high bioavailability, can prolong an in vivo imaging time window, and has the advantages of long-acting and low-dose imaging. The present invention also has good clearance characteristics and low risk of tissue residues, thereby reducing the potential risk of release of free Gd3+, and providing a long-acting, low-dose and high-safety magnetic resonance contrast solution for clinical use.

Description

Compounds, chelates and their applications in magnetic resonance imaging

[0001] Related applications: This application claims priority to Chinese invention patent application No. CN202411973602.5 filed on December 30, 2024 and Chinese invention patent application No. CN202511423783.9 filed on September 30, 2025, which are hereby expressly incorporated herein by reference. Technical Field

[0002] This invention belongs to the field of medical imaging technology, specifically relating to a compound, particularly a metal chelate of the compound, its preparation method, and its application in magnetic resonance imaging contrast agents. Background Technology

[0003] Gadolinium (III)-based contrast agents (GBCAs) are widely used clinically in magnetic resonance imaging (MRI). Once in the body, gadolinium-containing chelates significantly shorten the relaxation time of protons in tissues, thereby enhancing the clarity and contrast of MRI images and improving lesion detection rates. They provide more diagnostically valuable information than plain scans for the localization and characterization of lesions in the brain, spinal cord, and central nervous system. They are also used for MRI of the abdomen, chest, pelvis, limbs, and other human tissues, as well as for renal function assessment. Compared to other contrast agents, gadolinium contrast agents offer high tissue resolution and good safety.

[0004] Currently available gadolinium contrast agents all contain a nine-coordinate gadolinium ion [Gd(III)], which can coordinate with an octardate polyaminocarboxylic acid ligand and a water molecule. Based on ligand structure, they can be divided into linear and macrocyclic types. There are six linear gadolinium contrast agents, including gadopentetate diglucamine, gadodiamine, and gadofusamide; and three macrocyclic gadolinium contrast agents, namely gadoteric acid dimeglumine, gadoteryl alcohol, and gadobutrol. Existing gadolinium contrast agents generally exhibit a strictly passive extracellular distribution of GBCA in vivo and are excreted only through the kidneys; some, in addition to extracellular distribution, can also be partially excreted through the liver after uptake. Besides their typical imaging potential, such as imaging or contrast of the central nervous system, blood vessels, limbs, heart, head / face / neck, abdomen, and breast, gadolinium contrast agents also provide liver imaging through enhanced liver parenchyma caused by GBCA uptake in hepatocytes.

[0005] While existing macrocyclic gadolinium contrast agents offer certain safety advantages, they still suffer from several limitations: First, their imaging time windows are limited, making it difficult to meet the continuous enhancement requirements of long-duration scans or complex sequences; second, clinical imaging signals are highly dose-dependent, often requiring doubled doses to obtain ideal signals, increasing the potential risk of free gadolinium release; third, existing contrast agents are mostly used for specific organ imaging, lacking clinical applications of "single injection—multi-site enhancement throughout the body." Furthermore, although patent CN107667096B describes attempts to improve pharmacokinetic properties through structural modification, it still suffers from problems such as rapid drug clearance and unsatisfactory tissue signal residue, making it difficult to balance maintaining imaging intensity and reducing tissue exudation.

[0006] Therefore, there is still a clinical need for a new type of gadolinium chelate that can provide higher exposure, longer imaging window, faster background elution, and is suitable for imaging multiple sites throughout the body at the same or lower gadolinium dose. Summary of the Invention

[0007] This invention describes a new class of highly relaxible extracellular gadolinium chelates, their preparation methods, and their use as MRI contrast agents.

[0008] On the one hand, the present invention provides a compound of general formula (I) or a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a mixture thereof:

[0009] R1 and R3 are independently selected from hydrogen, deuterium, alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted hydroxyalkyl, substituted or unsubstituted mercaptoalkyl, substituted or unsubstituted alkoxyalkyl, substituted or unsubstituted thioetheralkyl, substituted or unsubstituted aminoalkyl, COOH, and COO. - The group may be esterified, amide-based, substituted or unsubstituted carboxylalkyl, substituted or unsubstituted esterylalkyl, substituted or unsubstituted amideylalkyl, phenyl, aromatic heterocyclic, aromatic heterocyclic alkyl, heterocyclic, heterocyclic alkyl, substituted or unsubstituted phenylcycloalkyl, or substituted or unsubstituted phenylalkyl; wherein the substituents in the above-mentioned substituted or unsubstituted groups are optionally selected from 1 to 3 of the following groups: halogen, hydroxyl, C1-C3 alkyl, halo-C1-C3 alkyl, or C1-C3 alkoxy; wherein the alkyl group is substituted by a phenyl group that is substituted 1 to 3 times in any way: hydroxyl, C1-C3 alkyl, halo-C1-C3 alkyl, or C1-C3 alkoxy;

[0010] R2 is selected from hydrogen, substituted or unsubstituted C1-C6 alkyl, C3-C6 cycloalkyl, (C1-C2 alkoxy)-(C2-C3 alkyl)-, substituted or unsubstituted heterocyclic aryl or phenyl; wherein the substituent in the substituted or unsubstituted C1-C6 alkyl is selected from phenyl substituted 1 to 3 times by any of the following groups: halogen, C1-C3 alkyl, halo-C1-C3 alkyl or C1-C3 alkoxy; the substituent in the substituted or unsubstituted heterocyclic aryl is selected from 1 to 3 of the following groups: C1-C3 alkyl, halo-C1-C3 alkyl or C1-C3 alkoxy;

[0011] Alternatively, R2 and R3 may form a five- or six-membered heterocyclic alkyl group;

[0012] And exclude the following situations:

[0013] R1 and R3 are selected from hydrogen, while R2 is selected from methyl, ethyl, isopropyl, 2-methylpropyl, octyl, cyclopropyl, cyclopentyl, 2-methoxyethyl, 2-ethoxyethyl or phenyl; or R1 is selected from hydrogen or methyl, while R2 and R3 are selected from hydrogen.

[0014] Furthermore, the alkyl group is selected from C1-C6 alkyl groups, and the cycloalkyl group is selected from C3-C6 cycloalkyl groups.

[0015] Furthermore, the above-mentioned compounds are:

[0016] The present invention also provides a compound of formula (II) or a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a mixture thereof:

[0017] The definitions of R1, R2, and R3 are the same as any of the corresponding definitions mentioned above;

[0018] The following conditions are excluded: R1 and R3 are selected from hydrogen, while R2 is selected from methyl, ethyl, isopropyl, 2-methylpropyl, octyl, cyclopropyl, cyclopentyl, 2-methoxyethyl, 2-ethoxyethyl or phenyl; or R1 is selected from hydrogen or methyl, while R2 and R3 are selected from hydrogen.

[0019] Furthermore, the above-mentioned compound is:

[0020] Furthermore, the present invention also provides a compound of formula (VII) or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a mixture thereof:

[0021] The definitions of R1, R2, and R3 are the same as any of the corresponding definitions mentioned above.

[0022] Furthermore, the above compounds are selected from:

[0023] Furthermore, the hydrogen in any of the above compounds can be replaced by one or more deuterium atoms.

[0024] In a second aspect, the present invention provides a method for preparing a compound of general formula (I) or general formula (VII), wherein,

[0025] The preparation of compound of general formula (VII) includes the following steps: condensing compound of formula (III) or its salt with compound of formula (V) to obtain compound of formula (VI), and further deprotecting compound of formula (VI) to obtain compound of general formula (VII);

[0026] Wherein, R1, R2, and R3 are defined as in claim 6, and R4 is selected from methyl, tert-butyl, or benzyl;

[0027] Alternatively, the preparation of compounds of general formula (I) includes the following methods:

[0028] Method 1: The compound of general formula (VII) is chelated to obtain the compound of general formula (I);

[0029] Method 2: A substitution reaction is carried out between a compound of formula (III) or its salt and a compound of formula (IV) to obtain a compound of general formula (I);

[0030] Wherein, R1, R2, and R3 are defined as in the corresponding definitions of claims 1-4; and LG represents an activated leaving group, preferably phenol, p-chlorophenol, or p-nitrophenol.

[0031] Furthermore, the above-mentioned compound (IV) is synthesized by the following steps: esterification of compound (II) with LG;

[0032] The definitions of R1, R2, and R3 are the same as the corresponding definitions of any of the above items, and LG is selected from phenol, p-chlorophenol, or p-nitrophenol.

[0033] Thirdly, the present invention provides the use of any of the above-mentioned compounds or their stereoisomers, pharmaceutically acceptable salts, or mixtures thereof in the preparation of diagnostic agents; preferably, the present invention provides the use of any of the above-mentioned compounds or their stereoisomers, pharmaceutically acceptable salts, or mixtures thereof in the preparation of contrast agents for magnetic resonance imaging.

[0034] Fourthly, the present invention provides a pharmaceutical composition comprising any of the above-described compounds or their stereoisomers, pharmaceutically acceptable salts or mixtures thereof, and pharmaceutically acceptable carriers and / or excipients.

[0035] The compound names corresponding to the English abbreviations mentioned in this invention are:

[0036] EDCI: 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride;

[0037] HOBT: 1-Hydroxybenzotriazole;

[0038] HATU: 2-(7-Azobenzotriazole)-N,N,N',N'-Tetramethylurea hexafluorophosphate;

[0039] TFA: Trifluoroacetic acid;

[0040] DBU: 1,8-diazabicyclo[5.4.0]undec-7-ene;

[0041] THF: Tetrahydrofuran;

[0042] DIC: N,N'-Diisopropylcarbodiimide;

[0043] PPh3: Triphenylphosphine.

[0044] Compared with the prior art, the present invention has the following beneficial effects:

[0045] (1) The gadolinium chelate and its salts of the present invention have significantly higher relaxation (r1) in pure water and plasma than the comparative compounds in vitro, and have higher potential for MRI contrast enhancement, providing a basis for achieving stronger signal or reduced equivalent imaging at the same dose.

[0046] (2) The pharmacokinetics (AUC) and C0.05 of the gadolinium chelate and its salt in vivo. maxIt is higher, especially at equimolar doses, plasma exposure can be more than 10 times higher than the commercially available control, peak concentration can be more than 2 times higher, with higher bioavailability and stronger initial imaging enhancement; in addition, it can maintain a higher peak signal even when the dose is at least halved, achieving "reduced dose without reduced effect", and the signal persistence window is extended by 1 time, rapid elution within 2 hours, tissue residue is close to the background, and it has the advantages of long-lasting imaging and low exudation safety.

[0047] (3) The gadolinium chelate and its salt of the present invention also have good scavenging properties and low risk of tissue residue, thereby reducing free Gd 3+ This approach aims to eliminate potential risks and provide a long-lasting, low-dose, and highly safe magnetic resonance imaging (MRI) contrast solution for clinical use. Attached Figure Description

[0048] Figure 1 shows MRI images of cerebral blood vessels in mice before and after administration of compounds 1-2 and compound 21 of Example 21;

[0049] Figure 2 shows the Δ signal intensity-time curves of mouse cerebral blood vessels after administration of compounds 1-2 and 21. Detailed Implementation

[0050] The present invention will be further described in detail below with reference to embodiments and experimental examples. The embodiments and experimental examples of the present invention are only used to illustrate the technical solutions of the present invention and are not intended to limit the present invention. Any equivalent substitutions made in the art based on the content disclosed in the present invention shall fall within the protection scope of the present invention.

[0051] The structure of the compound in the embodiments of this invention is determined by nuclear magnetic resonance (NMR). 1 Determined by ¹H NMR or LC-MS.

[0052] In this invention, "room temperature" refers to 10–35°C.

[0053] Example 1: Preparation of Compound 1

[0054] [7,10-bis(carboxymethyl)-4-(1,17-dihydroxy-9,9-bis{8-hydroxy-3,6-dioxane-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazaoct-1-yl}-3,6,12,15-tetraoxane-16-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-4,7,11,14-tetraazaheptadecane-2-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 1)

[0055] Step 1: Synthesis of Compound 1a

[0056] 5.67 g (35.17 mmol, 1.00 q.) of O-tert-butyl-serine, 14.65 g (123.10 mmol, 3.50 q.) of KBr and 47.43 g (281.36 mmol, 8.00 q.) of HBr (48%) were added to a 250 ml three-necked flask containing 57 ml of drinking water. The reaction system was stirred and cooled to 0 °C.

[0057] 2.55 g (36.91 mmol, 1.05 eq.) of NaNO2 was dissolved in 26 ml of drinking water and added dropwise to the reaction system. The internal temperature was controlled at 0 ± 5 °C during the dropwise addition. After the dropwise addition was completed, the mixture was stirred at 0 ± 5 °C for 4 hours and monitored by TLC. After the reaction was completed, the mixture was extracted with 113 ml of ethyl acetate, washed with 113 ml of water, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain 6.81 g (86%, 30.24 mmol) of intermediate compound 1a.

[0058] LC-MS (ESI-): m / z = 223.1 (MH) - .

[0059] Step 2: Synthesis of Compound 1b

[0060] 4.51 g (22.37 mmol, 1.05 eq.) glycine benzyl ester hydrochloride, 4.79 g (21.30 mmol, 1.00 eq.) compound 1a, 6.12 g (31.95 mmol, 1.50 eq.) EDCI and 4.32 g (31.95 mmol, 1.5 eq.) HOBT were added to a 500 mL three-necked flask containing 100 mL dichloromethane solvent. The reaction system was stirred and cooled to 0 °C under nitrogen protection. Then, 8.25 g (63.90 mmol, 3.0 eq.) N,N-diisopropylethylamine was added dropwise to the reaction flask. The reaction mixture was then brought to room temperature and stirred overnight.

[0061] After the reaction was completed by TLC monitoring, the reaction system was washed twice with 1 L of water, the organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure and then subjected to silica gel column chromatography to obtain 6.58 g (83%, 17.68 mmol) of intermediate compound 1b.

[0062] LC-MS(ESI+): m / z=316.0(Mt-Bu+H) + .

[0063] Step 3: Synthesis of compound 1c

[0064] 5.63 g (15.13 mmol, 1.00 eq.) of compound 1b and 7.79 g (15.13 mmol, 1.00 eq.) of 2,2',2”-(1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid tritert-butyl ester were added to 389 mL of acetonitrile, followed by 6.27 g (45.39 mmol, 3.00 eq.) of potassium carbonate and 0.23 g (1.51 mmol, 0.10 eq.) of sodium iodide. The reaction system was stirred and heated to 60 °C under nitrogen protection overnight. After the reaction was completed by TLC monitoring, the mixture was filtered, and the filtrate was concentrated under reduced pressure and then purified by silica gel column chromatography to obtain 6.83 g (56%, 8.47 mmol) of intermediate compound 1c.

[0065] LC-MS(ESI+): m / z=806.8(M+H) + .

[0066] Step 4: Synthesis of Compound 1d

[0067] 5.76 g (7.15 mmol, 1.00 eq.) of compound 1c, 3.38 g (1.43 mmol, 0.20 eq.) of 10% Pd / C and 230 ml of isopropanol were added to a hydrogenation reactor. The reactor was purged with hydrogen three times, pressurized to P = 1.00 MPa, heated to 80 °C and reacted for 8 h. Heating was stopped, the temperature was lowered, the reaction solution was filtered, and the filtrate was concentrated under reduced pressure and then purified by silica gel column chromatography to obtain 4.25 g (83%, 5.93 mmol) of intermediate compound 1d.

[0068] LC-MS (ESI-): m / z = 714.5 (MH) - .

[0069] Step 5: Synthesis of compound 1e

[0070] 3.52 g (4.92 mmol, 12.00 eq.) of compound 1d, 2.18 g (5.74 mmol, 14.00 eq.) of HATU and 205 mL of N,N-dimethylacetamide were added to a 500 mL three-necked flask. The reaction system was stirred and cooled to 0 °C under nitrogen protection. Then, 1.59 g (12.30 mmol, 30.00 eq.) of N,N-diisopropylethylamine dissolved in 82 mL of N,N-dimethylacetamide was added. The resulting reaction mixture was stirred at 0-5°C for 30 min, and then 114.0 mg (0.41 mmol, 1.00 eq.) of 2,2-di(aminomethyl)prop-1,3-diamine hydrochloride was added. The reaction mixture was stirred at room temperature for 3 h. The reaction mixture was then added to 3 L of purified water and stirred. 2 L of dichloromethane was added for extraction, and the organic phase was collected. The organic phase was washed three times with 2 L of saturated sodium chloride solution and dried with anhydrous sodium sulfate. After concentration under reduced pressure, crude product 1e was obtained. This crude product did not require further characterization and was directly used in the next chemical step.

[0071] Step Six: Synthesis of Compound 1f

[0072] The crude product 1e obtained in the previous step was treated with TFA (105 mL) and stirred overnight at room temperature. The reaction mixture was concentrated under reduced pressure to obtain an oil, which was separated by reversed-phase preparative chromatography to give 263.4 mg (32% (overall yield of the two-step chemical reaction), 0.13 mmol) of intermediate compound 1f.

[0073] LC-MS(ESI+): m / z(z=2)=1013.5(M+2H) 2+ m / z (z = 3) = 676.0 (M + 3H) 3+ m / z (z = 4) = 507.3 (M + 4H) 4+ .

[0074] Step 7: Synthesis of Compound 1

[0075] 202.6 mg (0.10 mmol, 1.00 eq.) of compound 1f was dissolved in 265 mL of purified water. After adding 72.5 mg (0.20 mmol, 2.00 eq.) of gadolinium oxide, the reaction mixture was heated to 90-100 °C and stirred for 5 h. The reaction solution was then cooled to room temperature and lyophilized. The crude product was separated by reverse preparative chromatography to obtain 211.4 mg (80%, 0.08 mmol) of compound 1 with a purity of 98.4%.

[0076] LC-MS (ESI+): 1323.0 (M+2H) 2+ m / z (z = 3) = 882.0 (M + 3H) 3+ m / z (z = 4) = 661.5 (M + 4H)4+ .

[0077] Example 2: Preparation of Compound 2

[0078] [7,10-bis(carboxymethyl)-4-(1,15-dicarboxy-8,8-bis{7-carboxy-3,6-dioxane-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diahepta-1-yl}-2,5,11,14-tetraoxane-15-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-3,6,10,13-tetraazapentadecanane-1-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 2)

[0079] The preparation method is the same as that in Example 1, except that 1b in Example 1 is replaced with intermediate 2b and step 1a is omitted to obtain compound 2. The final synthesis yield is 76% and the purity is 98.9%.

[0080] LC-MS (ESI+): 1350.7 (M+2H) 2+ m / z (z = 3) = 900.9 (M + 3H) 3+ m / z (z = 4) = 676.1 (M + 4H) 4+ .

[0081] The synthetic route for compound 2b is as follows:

[0082] Step 1: Synthesis of compound 2a

[0083] 14.63 g (72.54 mmol, 1.05 eq.) glycine benzyl ester hydrochloride, 11.07 g (69.09 mmol, 1.00 eq.) mono-tert-butyl malonate, 19.87 g (103.64 mmol, 1.5 eq.) EDCI and 14.00 g (103.64 mmol, 1.5 eq.) HOBT were added to a 1 L three-necked flask containing 221 mL of dichloromethane solvent. The reaction system was stirred and cooled to 0 °C under nitrogen protection. Then, 26.75 g (207.27 mmol, 3.0 eq.) of N,N-diisopropylethylamine was added dropwise to the reaction flask. The reaction mixture was then brought to room temperature and stirred overnight.

[0084] After the reaction was completed by TLC monitoring, the reaction system was washed twice with 1 L of water, the organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure and then subjected to silica gel column chromatography to obtain 18.05 g (85%, 58.73 mmol) of intermediate compound 2a.

[0085] LC-MS(ESI+): m / z=252.1(Mt-Bu+H) + .

[0086] Step 2: Synthesis of compound 2b

[0087] 15.41 g (50.14 mmol, 1.00 eq.) of compound 2a, 8.40 g (55.15 mmol, 1.10 eq.) of 1,8-diazabicyclo[5.4.0]undec-7-ene and 154 mL of tetrahydrofuran were added to a 500 mL three-necked flask and stirred and cooled to -60 °C. 17.46 g (52.65 mmol, 1.05 eq.) of carbon tetrabromide was dissolved in 200 mL of tetrahydrofuran and added dropwise to the reaction system. The internal temperature was controlled at -60 ± 5 °C during the dropwise addition. After the addition was complete, the mixture was stirred for 2 hours and monitored by TLC. After the reaction was completed, 100 mL of saturated ammonium chloride solution was added to quench the reaction. The mixture was stirred and brought to room temperature. The reaction solution was extracted twice with dichloromethane, using 300 mL of dichloromethane each time. The organic phases were combined and washed once with saturated sodium chloride. The mixture was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and then purified by silica gel column chromatography to obtain 10.26 g (53%, 26.57 mmol) of intermediate compound 2b.

[0088] LC-MS(ESI+): m / z=330.1(Mt-Bu+H) + .

[0089] Example 3: Preparation of Compound 3

[0090] The preparation method is the same as that in Example 1. Compound 3 is obtained by replacing glycine benzyl ester hydrochloride in Example 1 with compound 3a and replacing 1a with 2-bromopropionic acid, thus omitting step 1a. The final synthesis yield is 79%, and the purity is 98.8%.

[0091] LC-MS(ESI+): m / z(z=2)=1403.5(M+2H) 2+ m / z (z = 3) = 936.0 (M + 3H) 3+ m / z (z = 4) = 702.2 (M + 4H) 4+ .

[0092] Synthesis of compound 3a

[0093] 56.25 g (769.02 mmol, 6.00 eq.) of isobutylamine and 250 mL of tetrahydrofuran were added to a 1 L three-necked flask and stirred until cooled to 0 ± 5 °C. 29.36 g (128.17 mmol, 1.00 eq.) of benzyl bromoacetate was dissolved in 200 mL of tetrahydrofuran and added dropwise to the reaction system, maintaining the internal temperature at 0 ± 5 °C during the addition. After the addition was complete, the mixture was stirred at 0 ± 5 °C for 1 hour, and monitored by TLC. After the reaction was completed, the mixture was concentrated under reduced pressure and purified by silica gel column chromatography to obtain 16.45 g (58%, 74.34 mmol) of intermediate compound 3a with a purity of 98.9%.

[0094] LC-MS(ESI+): m / z=222.1(M+H) + .

[0095] Example 4: Preparation of Compound 4

[0096] [4,7-bis(carboxylate-methyl)-10-{1-[(carboxymethyl)amino]-3-hydroxy-1-oxoylide-2-yl}-1,4,7,10-tetraazacyclododecane-1-yl]gadolinium acetate (compound 4)

[0097] Step 1: Synthesis of compound 4a

[0098] 694.5 mg (0.97 mmol, 1.00 eq.) of N-{3-[(2-methylprop-2-yl)oxy]-2-(4-{1-[(2-methylprop-2-yl)oxy]-1-oxylideneethyl-2-yl}-7,10-bis{2-[(2-methylprop-2-yl)oxy]-2-oxylideneethyl}-1,4,7,10-tetraazacyclododecane-1-yl)propionyl}glycine was added to 21 mL of TFA and stirred overnight at room temperature. The reaction mixture was concentrated under reduced pressure to give 4a, which was used directly for the next chemical step without further characterization.

[0099] Step 2: Synthesis of Compound 4

[0100] The 4a obtained in the previous step was dissolved in 100 ml of purified water. 177.6 mg (0.49 mmol, 0.50 eq.) of gadolinium oxide was added, and the reaction mixture was heated to 90-100 °C and stirred for 5 h. The reaction solution was then cooled to room temperature and lyophilized. The product was separated by reversed-phase preparative chromatography to obtain 348.7 mg (56% (overall yield of the two-step chemical reaction), 0.54 mmol) of compound 4 with a purity of 99.6%.

[0101] LC-MS(ESI+): m / z=647.4(M+H) + .

[0102] Example 5: Preparation of Compound 5

[0103] (10-{1-carboxy-2-[(carboxymethyl)amino]-2-oxoylideneethyl}-4,7-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl)gadolinium acetate (compound 5)

[0104] Step 1: Synthesis of compound 5a

[0105] 897.8 mg (1.23 mmol, 1.00 eq.) of N-{3-[(2-methylprop-2-yl)oxy]-2-(4-{1-[(2-methylprop-2-yl)oxy]-1-oxylideneethyl-2-yl}-7,10-bis{2-[(2-methylprop-2-yl)oxy]-2-oxylideneethyl}-1,4,7,10-tetraazacyclododecane-1-yl)-1,3-dioxylidenepropyl}glycine was added to 27 mL of TFA and stirred overnight at room temperature. The reaction mixture was concentrated under reduced pressure to give 5a, which was used directly for the next chemical step without further characterization.

[0106] Step 2: Synthesis of Compound 5

[0107] The 5a obtained in the previous step was dissolved in 100 ml of purified water. 224.8 mg (0.62 mmol, 0.50 eq.) of gadolinium oxide was added, and the reaction mixture was heated to 90-100 °C and stirred for 5 h. The reaction solution was then cooled to room temperature and lyophilized. The product was separated by reversed-phase preparative chromatography to obtain 382.6 mg (47% (overall yield of the two-step chemical reaction), 0.58 mmol) of compound 5 with a purity of 98.9%.

[0108] LC-MS(ESI+): m / z=661.1(M+H) + .

[0109] Example 6: Preparation of Compound 6

[0110] [4,7-bis(carboxymethyl)-10-{1-[(carboxymethyl)amino]-1-oxoylidenepropyl-2-yl}-1,4,7,10-tetraazacyclododecane-1-yl]gadolinium acetate (compound 6)

[0111] Step 1: Synthesis of compound 6a

[0112] 2.00 g (3.11 mmol, 1.00 eq.) of N-[2-(4-{1-[(2-methylprop-2-yl)oxy]-1-oxylidene eth-2-yl}-7,10-bis{2-[(2-methylprop-2-yl)oxy]-2-oxylidene ethyl}-1,4,7,10-tetraazacyclododecane-1-yl)propionyl]glycine was added to 60 mL of TFA and stirred overnight at room temperature. The reaction mixture was concentrated under reduced pressure to give 6a, which was used directly for the next chemical step without further characterization.

[0113] Step 2: Synthesis of Compound 6

[0114] The 6a obtained in the previous step was dissolved in 200 ml of purified water. 565.5 mg (1.56 mmol, 0.50 eq.) of gadolinium oxide was added, and the reaction mixture was heated to 90-100 °C and stirred for 5 h. The reaction solution was then cooled to room temperature and lyophilized. Separation by reversed-phase preparative chromatography yielded 743.1 mg (38% (overall yield of the two-step reaction), 1.18 mmol) of compound 6. Purity: 99.4%.

[0115] LC-MS(ESI+): m / z=631.3(M+H) + .

[0116] Example 7: Preparation of Compound 7

[0117] [4,7-bis(carboxymethyl)-10-(1-{2-[1,7-dioxylidene-7-(1-{2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}tetrahydro-1H-pyrrolo-2-yl)-4,4-bis({[(1-{2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}tetrahydro-1H-pyrrolo-2-yl)carbonyl]amino}methyl)-2,6-diazaheptane-1-yl]tetrahydro-1H-pyrrolo-1-yl}-1-oxylidenepropionyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 7)

[0118] The preparation method is the same as that in Example 3, except that 3a in Example 3 is replaced with benzyl proline to obtain compound 7. The final synthesis yield is 76% and the purity is 98.7%.

[0119] LC-MS(ESI+): m / z(z=2)=1371.4(M+2H) 2+ m / z (z = 3) = 914.6 (M + 3H) 3+m / z (z = 4) = 686.2 (M + 4H) 4+ .

[0120] Example 8: Preparation of Compound 8

[0121] [4,7-bis(carboxymethyl)-10-(1-{2-[1,7-dioxylidene-7-(1-{2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}hexahydropyridin-2-yl)-4,4-bis({[(1-{2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}hexahydropyridin-2-yl)carbonyl]amino}methyl)-2,6-diazaheptane-1-yl]hexahydropyridin-1-yl}-1-oxylidenepropyl-2-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 8)

[0122] The preparation method is the same as that in Example 3, except that 3a in Example 3 is replaced with benzyl 2-piperidinecarboxylate to obtain compound 8. The final synthesis yield is 72% and the purity is 98.1%.

[0123] LC-MS(ESI+): m / z(z=2)=1399.4(M+2H) 2+ m / z (z = 3) = 933.3 (M + 3H) 3+ m / z (z = 4) = 700.2(M + 4H) 4+ .

[0124] Example 9: Preparation of Compound 9

[0125] [4,7-bis(carboxymethyl)-10-(4,14-dicyclopropyl-9,9-bis{5-cyclopropyl-3,6-dioxonyl-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazaoctyl-1-yl}-3,6,12,15-tetraoxonyl-16-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-4,7,11,14-tetraazaheptadecane-2-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 9)

[0126] The preparation method is the same as that in Example 3, except that isobutylamine in Example 3 is replaced with cyclopropylamine to obtain compound 9. The final synthesis yield is 70% and the purity is 98.4%.

[0127] LC-MS(ESI+): m / z(z=2)=1371.4(M+2H) 2+ m / z (z = 3) = 914.6 (M + 3H) 3+ m / z (z = 4) = 686.2 (M + 4H) 4+ .

[0128] Example 10: Preparation of Compound 10

[0129] [4-(9,9-bis{3,6-dioxonyl-8-mercapto-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazaoct-1-yl}-3,6,12,15-tetraoxonyl-1,17-dimercapto-16-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-4,7,11,14-tetraazaheptadecane-2-yl)-7,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 10)

[0130] The preparation method is the same as that in Example 1, except that O-tert-butyl-serine in Example 1a is replaced with S-tert-butyl-DL-cysteine ​​hydrochloride to obtain compound 10. The final synthesis yield is 66% and the purity is 98.3%.

[0131] LC-MS(ESI+): m / z(z=2)=1355.3(M+2H) 2+ m / z (z = 3) = 903.8 (M + 3H) 3+ m / z (z = 4) = 678.1 (M + 4H) 4+ .

[0132] Example 11: Preparation of Compound 11

[0133] [4,7-bis(carboxymethyl)-10-(3-hydroxy-1-{2-[7-(1-{3-hydroxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}tetrahydro-1H-pyrrolo-2-yl)-4,4-bis({[(1-{3-hydroxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}tetrahydro-1H-pyrrolo-2-yl)-4,4-bis({[(1-{3-hydroxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}tetrahydro-1H-pyrrolo-2-yl) [1,4,7,10-tetraazacyclododecane-1-yl]propionyl]tetrahydro-1H-pyrrolo-2-yl]carbonyl]amino]methyl]1,7-dioxane-2,6-diazaheptane-1-yl]tetrahydro-1H-pyrrolo-1-yl]-1-oxanepropyl-2-yl]-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (Compound 11)

[0134] The preparation method is the same as that in Example 1, except that glycine benzyl ester hydrochloride in Example 1b is replaced with proline benzyl ester to obtain compound 11. The final synthesis yield is 70% and the purity is 98.0%.

[0135] LC-MS(ESI+): m / z(z=2)=1403.4(M+2H) 2+ m / z (z = 3) = 935.9 (M + 3H) 3+ m / z (z = 4) = 702.2 (M + 4H) 4+ .

[0136] Example 12: Preparation of Compound 12

[0137] [4,7-bis(carboxymethyl)-10-(3-hydroxy-1-{2-[7-(1-{3-hydroxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}hexahydropyridin-2-yl)-4,4-bis({[(1-{3-hydroxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]propionyl}hexahydropyridin-2-yl)carbonyl]amino}methyl)-1,7-dioxane-2,6-diazepeptide-1-yl]hexahydropyridin-1-yl}-1-oxanepropyl-2-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 12)

[0138] The preparation method is the same as that in Example 1, except that glycine benzyl ester hydrochloride in Example 1b is replaced with benzyl 2-piperidinecarboxylate to obtain compound 12. The final synthesis yield is 65% and the purity is 98.4%.

[0139] LC-MS(ESI+): m / z(z=2)=1431.4(M+2H) 2+ m / z (z = 3) = 954.6 (M + 3H) 3+ m / z (z = 4) = 716.2 (M + 4H) 4+ .

[0140] Example 13: Preparation of Compound 13

[0141] [10-(1-carboxyl-2-{2-[7-(1-{2-carboxyl-2-[4,7,10-tris(carboxylmethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}tetrahydro-1H-pyrrolo-2-yl)-4,4-bis({[(1-{2-carboxyl-2-[4,7,10-tris(carboxylmethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}tetrahydro-1H-pyrrolo-2-yl)-4,4-bis({[(1-{2-carboxyl-2-[4,7,10-tris(carboxylmethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl} [-tetraazacyclododecane-1-yl]acetyl]tetrahydro-1H-pyrrolo-2-yl]carbonyl]amino]methyl]-1,7-dioxane-2,6-diazepeptide-1-yl]tetrahydro-1H-pyrrolo-1-yl]-2-oxane-ethyl]-4,7-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 13)

[0142] The preparation method is the same as that in Example 2, except that glycine benzyl ester hydrochloride in Example 2a is replaced with proline benzyl ester to obtain compound 13. The final synthesis yield is 60% and the purity is 98.4%.

[0143] LC-MS(ESI+): m / z(z=2)=1431.3(M+2H) 2+ m / z (z = 3) = 954.5 (M + 3H) 3+ m / z (z = 4) = 716.2 (M + 4H) 4+ .

[0144] Example 14: Preparation of Compound 14

[0145] [10-(1-carboxy-2-{2-[7-(1-{2-carboxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}hexahydropyridin-2-yl)-4,4-bis({[(1-{2-carboxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}hexahydropyridin-2-yl)carbonyl]amino}methyl)-1,7-dioxane-2,6-diazepeptide-1-yl]hexahydropyridin-1-yl}-2-oxane-ethyl)-4,7-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 14)

[0146] The preparation method is the same as that in Example 2, except that glycine benzyl ester hydrochloride in Example 2a is replaced with benzyl 2-piperidinecarboxylate to obtain compound 14. The final synthesis yield is 62% and the purity is 98.3%.

[0147] LC-MS(ESI+): m / z(z=2)=1459.4(M+2H)2+ m / z (z = 3) = 973.2 (M + 3H) 3+ m / z (z = 4) = 730.2 (M + 4H) 4+ .

[0148] Example 15: Preparation of Compound 15

[0149] [4-(12,12-bis{6,9-dioxane-5-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-7,10-diaza-2-thia-undecane-11-yl}-6,9,15,18-tetraoxane-19-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,22-dithia-7,10,14,17-tetraaza-tetrazotriane-5-yl)-7,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 15)

[0150] The preparation method is the same as that in Example 1, except that O-tert-butyl-serine in Example 1a is replaced with DL-methionine to obtain compound 15. The final synthesis yield is 72% and the purity is 98.7%.

[0151] LC-MS(ESI+): m / z(z=2)=1411.3M+2H) 2+ m / z (z = 3) = 941.2 (M + 3H) 3+ m / z (z = 4) = 706.2 (M + 4H) 4+ .

[0152] Example 16: Preparation of Compound 16

[0153] [4,7-bis(carboxymethyl)-10-{1,17-dihydroxy-9,9-bis[8-hydroxy-5-(2-methylpropyl)-3,6-dioxonyl-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazaoctyl-1-yl]-4,14-bis(2-methylpropyl)-3,6,12,15-tetraoxonyl-16-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-4,7,11,14-tetraazaheptadecane-2-yl}-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 16)

[0154] The preparation method is the same as that in Example 1, except that glycine benzyl ester hydrochloride is replaced with 3a to obtain compound 16. The final synthesis yield is 72% and the purity is 98.1%.

[0155] LC-MS(ESI+): m / z(z=2)=1435.4(M+2H) 2+ m / z (z = 3) = 957.3 (M + 3H) 3+ m / z (z = 4) = 718.2 (M + 4H) 4+ .

[0156] Example 17: Preparation of Compound 17

[0157] [10-(1-carboxy-13-{2-carboxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}-8,8-bis(5-{2-carboxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}-7-methyl-3-oxylidene-2,5-diazaoctyl-1-yl)-15-methyl-3-(2-methylpropyl)-2,5,11-trioxylidene-3,6,10,13-tetraazahexahexadecane-1-yl)-4,7-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 17)

[0158] The preparation method was the same as in Example 2, except that glycine benzyl ester hydrochloride was replaced with 3a to obtain compound 17. The final synthesis yield was 62% and the purity was 98.2%.

[0159] LC-MS(ESI+): m / z(z=2)=1463.4(M+2H) 2+ m / z (z = 3) = 975.9 (M + 3H) 3+ m / z (z = 4) = 732.2 (M + 4H) 4+ .

[0160] Example 18: Preparation of Compound 18

[0161] [4,7-bis(carboxymethyl)-10-{1,17-dihydroxy-9,9-bis[8-hydroxy-4-methyl-5-(2-methylpropyl)-3,6-dioxonyl-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazaoct-1-yl]-5,13-dimethyl-4,14-bis(2-methylpropyl)-3,6,12,15-tetraoxonyl-16-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-4,7,11,14-tetraazaheptadecane-2-yl}-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 18)

[0162] The preparation method is the same as that in Example 16, except that benzyl bromoacetate in Example 16 is replaced with benzyl 2-bromopropionate to obtain compound 18. The final synthesis yield is 65% and the purity is 98.5%.

[0163] LC-MS(ESI+): m / z(z=2)=1463.5(M+2H) 2+ m / z (z = 3) = 976.0 (M + 3H) 3+ m / z (z = 4) = 732.2 (M + 4H) 4+ .

[0164] Example 19: Preparation of Compound 19

[0165] [10-(1-carboxy-13-{2-carboxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}-8,8-bis(5-{2-carboxy-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]acetyl}-4,7-dimethyl-3-oxylidene-2,5-diazaoctyl-1-yl)-4,12,15-trimethyl-3-(2-methylpropyl)-2,5,11-trioxylidene-3,6,10,13-tetraazahexahexadecane-1-yl)-4,7-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 19)

[0166] The preparation method is the same as that in Example 17, except that 2-bromoacetic acid benzyl ester in Example 17 is replaced with 2-bromopropionic acid benzyl ester to obtain compound 19. The final synthesis yield is 62% and the purity is 98.8%.

[0167] LC-MS(ESI+): m / z(z=2)=1491.4(M+2H)2+ m / z (z = 3) = 994.6 (M + 3H) 3+ m / z (z = 4) = 746.2 (M + 4H) 4+ .

[0168] Example 20: Preparation of Compound 20

[0169] [7,10-bis(carboxymethyl)-4-(4,14-dimethyl-9,9-bis{5-methyl-3,6-dioxane-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazaoctyl-1-yl}-3,6,12,15-tetraoxane-16-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-4,7,11,14-tetraazaheptadecane-2-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 20)

[0170] The preparation method is the same as that in Example 3, except that 3a in Example 3 is replaced with benzyl (methylamino)acetate to obtain compound 20. The final synthesis yield is 67% and the purity is 98.3%.

[0171] LC-MS(ESI+): m / z(z=2)=1319.4(M+2H) 2+ m / z (z = 3) = 879.9 (M + 3H) 3+ m / z (z = 4) = 660.2(M + 4H) 4+ .

[0172] Example 21: Preparation of Compound 21

[0173] [4,7-bis(carboxymethyl)-10-(1,19-dicarboxy-10,10-bis{9-carboxy-3,6-dioxane-7-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-2,5-diazanon-1-yl}-4,7,13,16-tetraoxane-17-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]-5,8,12,15-tetraazanonadecan-3-yl)-1,4,7,10-tetraazacyclododecane-1-yl]tetragadolinium acetate (compound 21)

[0174] The preparation method is the same as that in Example 1, except that O-tert-butyl-serine in Example 1a is replaced with glutamic acid-5-tert-butyl ester to obtain compound 21. The final synthesis yield is 64% and the purity is 98.2%.

[0175] LC-MS(ESI+): m / z(z=2)=1407.3(M+2H) 2+ m / z (z = 3) = 938.0 (M + 3H) 3+ m / z (z = 4) = 704.2 (M + 4H) 4+ .

[0176] The intermediates and their mass spectrometry data included in the above embodiments are shown in the table below:

[0177] Comparative Example 1: Gadovist (purchased from Anaiji Chemical, purity ≥ 98%)

[0178] Comparative Example 2: Synthesis of Gadoquatrane

[0179] The compound of Example 3 in patent CN107667096B was prepared according to the preparation method of Example 3 and is used as Comparative Example 2 of this application. Finally, it was purified by reversed-phase preparative chromatography and dried to obtain the target Comparative Example 2 compound with a purity of 98.5%. The structure was consistent with the analysis, and the LC-MS (ESI+) of the main product was: m / z (z=2) = 1290.5 (M+H). 2+ m / z (z = 3) = 860.9 (M + H) 3+ The chemical structure of the product is the same as that of Comparative Example 2 above.

[0180] Experimental Example 1: Relaxation Measurement

[0181] 1. Instruments

[0182] Meishi Medical 7.0T Small Animal Magnetic Resonance Imaging System.

[0183] 2. Preparation of test samples

[0184] At room temperature, using pure water and human blood plasma as matrices, the compounds of the examples and the comparative compounds were prepared into test samples with concentrations of 1.0, 0.75, 0.6, 0.5, 0.4, and 0.25 mmol Gd / L, respectively. The samples were sonicated for 2 minutes to remove air bubbles, centrifuged to ensure the liquid level was consistent, sealed to prevent evaporation, and placed in a honeycomb array sample holder in order of concentration for later use.

[0185] 3. Test methods

[0186] Before the formal scan, after the gradient and RF tests pass, the sample is placed at the center of the coil, and local shimming (based on the water peak) is performed until the linewidth is <30–50Hz. A rapid localization image is acquired to ensure that the single-layer slice covers all samples. After confirmation, the localization / tuning / matching / field modulation is performed, and the geometry of the coil and sample is recorded. T1 arrays of all concentration samples are acquired sequentially. The sequence parameters (IR-SE sequence) are as follows:

[0187] Field of view (FOV) 80×80mm 2 Matrix 128×128; Thickness 3mm; Repetition Time (TR) 3000ms; Echo Time (TE) 7.5ms; 8 TI arrays with times of 10, 300, 580, 860, 1150, 1400, 1700, and 2000ms respectively; Accumulation count 1; Number of layers 1.

[0188] 4. Data Processing and Result Analysis

[0189] Data processing: Take the average signal within each sample's circular ROI (avoiding the boundary by 1–2 pixels), fit it to obtain T1(m), and convert T1 to R1 = 1 / T1 (unit: s). -1 (T1 needs to be converted to seconds); Perform univariate linear regression on (R1, [C]):

[0190] R1 = r1[C] + R10, and the resulting slope r1 is the relaxation rate of the compound (unit: mM^-1s^-1).

[0191] The relaxation properties of each compound measured in pure water and human plasma are shown in the table below:

[0192] Table 1. Relaxability (mM^-1s^-1) in water and human plasma

[0193] The relaxation (r1) of compounds 1, 2, 10, 11, 13, and 21 in the examples was significantly higher than that of the comparative compounds in pure water and plasma in vitro. In contrast, compounds 3, 7, 8, and 9 in the examples showed a level that was basically equivalent to that of the comparative compounds. The overall results indicate that the compounds of the present invention have higher potential for MRI contrast enhancement, providing strong evidence for obtaining stronger signals at the same dose or maintaining equivalent imaging effects under reduced dose conditions.

[0194] Experimental Example 2: Pharmacokinetic Experiment in SD Rats

[0195] 1. Experimental animals

[0196] SPF-grade male SD rats, 6-8 weeks old, weighing 180-250g, with 6 rats in each group, were randomly assigned to groups according to their weight and had free access to food and water.

[0197] 2. Administration

[0198] Test samples: compounds from each example and comparative example;

[0199] Administration method: Weigh the patient before administration. Calculate the dosage based on body weight at 0.1 mmol Gd / kg. Prepare the injection solution using 0.9% sodium chloride as the solvent. Prepare and use immediately. Administer intravenously.

[0200] 3. Sample collection and processing

[0201] After blood was collected via the jugular vein, the blood samples were placed on ice. At least 150 μL of blood was collected per time point. The samples were temporarily placed on ice before centrifugation and the plasma was separated by centrifugation within one hour. The collected whole blood samples were placed in blood collection tubes containing anticoagulant (EDTA-K2) and centrifuged at 6800g and 2-8℃ for 6 min. At least 75 μL of plasma was collected and stored in a -80℃ freezer for testing.

[0202] 4. Data Processing and Result Analysis

[0203] Bioanalysis: An ICP-MS standard curve was established to detect the plasma concentration of gadolinium, with an upper limit of quantification of 20,000 ng / mL and a lower limit of quantification of 0.1 ng / mL. The accuracy of quality control samples was evaluated simultaneously with the analysis of the samples. The accuracy of more than 66% of the quality control samples was between 80-120%.

[0204] Data processing: Pharmacokinetic parameters, such as AUC, were calculated using WinNonlin based on blood drug concentration data at different time points. all AUC inf T 1 / 2 C max and T max When plotting plasma drug concentration-time curves, BLQ is always recorded as 0. When calculating pharmacokinetic parameters, the concentration before administration is calculated as 0; C max Previous BLQs (including "No peak") are calculated as 0; C max Subsequent BLQs (including "No peak") will not be included in the calculation.

[0205] The main pharmacokinetic parameters (Mean ± SD) of each compound in rats are shown in the table below:

[0206] Table 2. Pharmacokinetic parameters of gadolinium in plasma (Mean ± SD)

[0207] As can be seen from Table 2, at equimolar gadolinium dosages, the AUC of the compounds in the examples is... all All were significantly higher than the comparative compounds, with the AUCall of Examples 1 and 2 being approximately 5 times that of the comparative compounds, and Example 21 exceeding 10 times. This indicates that the total exposure level of the compounds of the present invention in vivo is significantly increased, which is beneficial for enhancing imaging signal intensity and extending the imaging time window. Meanwhile, the C of the compounds in the examples... max The value is approximately twice that of the comparative example, indicating that the compounds of the present invention can reach higher peak plasma concentrations more quickly, which is beneficial for rapidly obtaining excellent initial enhancement effects. Furthermore, the half-lives of the compounds in the examples are all at least half that of the comparative example compounds, demonstrating that the compounds of the present invention have good clearance properties and a low risk of tissue residue, thereby reducing free Gd. 3+ Eliminate potential hazards and enhance safety.

[0208] Experimental Example 3: Evaluation of Magnetic Resonance Imaging (MRI) Results

[0209] This study aims to evaluate the magnetic resonance enhancement effect and pharmacodynamic characteristics of the compound in the example and the comparative compound in mouse cerebral blood vessels through a single imaging experiment and pharmacodynamic experiment.

[0210] 1. Test materials

[0211] Instrument: MedSci 7.0T Small Animal Magnetic Resonance Imaging System.

[0212] Animals: Balb / c mice, 7-12 weeks old; single imaging test, males, 3 mice per group; time-dependent test, half males and half females, 2 mice per group.

[0213] Test samples: Take each compound and prepare injection solutions using 0.9% sodium chloride as solvent. Prepare and use immediately.

[0214] 2. Dosing regimen

[0215] Single imaging test: All test samples were administered at an equivalent dose of 0.1 mmol Gd / kg.

[0216] Pharmacokinetic studies (time-dependent): Compound 1 was administered at 0.1 mmol Gd / kg; Compound 2 and Compound 3 were administered at a low dose of 0.04 mmol Gd / kg.

[0217] 3. Test methods

[0218] Mice were fasted overnight before the examination and anesthetized with 1%-1.5% isoflurane gas at a ventilation rate of 0.8-1.0 L / min O2. After anesthesia, a tail vein access was established. The mice were fixed in a prone position in the magnetic resonance imaging (MRI) scanner, using a special head coil for mice. After tuning and shimming, cerebral vascular MRA data were first acquired before drug administration, followed by injection of the test substance via the tail vein, and data were acquired after drug administration.

[0219] Single imaging study: Images were acquired before administration (pre) and immediately after administration.

[0220] Pharmacokinetic study (time-dependent): Images were collected at multiple time points, including before administration (pre) and 0.5 min, 5 min, 15 min, and 30 min after administration.

[0221] 4. Data Processing

[0222] MRI angiography was performed using 3D CE-MRA sequences. The MRI images were analyzed and processed using ImageJ and MatLab software. The signal intensity (SI) of the target region (blood vessel) was measured, and the imaging effect was expressed as MRI Δ signal intensity. The calculation formula is as follows:

[0223] MRI signal intensity = (signal intensity after drug administration / signal intensity before drug administration) - 1.

[0224] 5. Results Analysis

[0225] 5.1 Single Imaging

[0226] Figure 1 shows MRI images of mouse cerebral blood vessels before and after administration of compounds 1-2 and compound 21. As can be seen from the figure, compound 21 showed the highest contrast enhancement, sharp blood vessel edges, and no signal overflow, which was significantly better than the comparative examples.

[0227] 5.2 Pharmacokinetics

[0228] Table 3. Δ signal intensity (Mean ± SD) of cerebral blood vessels on MRI

[0229] Table 3 shows the Δ signal intensity of mouse cerebral blood vessels on MRI at different administration times for compounds in Comparative Examples 1-2 and Compound 21. Comparative Example 1 was administered at a high dose of 0.1 mmol Gd / kg, while Comparative Example 2 and Compound 2 were administered at low doses of 0.04 mmol Gd / kg. Figure 2 shows the Δ signal intensity-time curves of mouse cerebral blood vessels on MRI after administration of compounds in Comparative Examples 1-2 and Compound 21. As can be seen from Figure 2 and Table 3, Compound 21, under low-dose conditions, not only achieved a peak signal enhancement of 9.22 times at 0.5 min, significantly better than the comparative groups, but also maintained a 3.27-fold enhancement level at 15 min, indicating that its vascular retention ability was superior to both comparative groups, effectively expanding the imaging window and demonstrating the dual advantages of "higher peak value and longer effective window." Furthermore, the experiment found that by 2 h, the signal of Compound 21 had fallen back to the baseline level basically consistent with that of Comparative Examples 1 and 2, showing rapid clearance without delayed accumulation, exhibiting both efficient imaging and rapid metabolism, demonstrating a safe profile.

[0230] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A compound of formula (I) or a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a mixture thereof: R1 and R3 are independently selected from hydrogen, deuterium, alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted hydroxyalkyl, substituted or unsubstituted mercaptoalkyl, substituted or unsubstituted alkoxyalkyl, substituted or unsubstituted thioetheralkyl, substituted or unsubstituted aminoalkyl, COOH, and COO. - The alkyl group may be esterified, amide-based, substituted or unsubstituted carboxylalkyl, substituted or unsubstituted esteralkyl, substituted or unsubstituted amidealkyl, phenyl, aromatic heterocyclic, aromatic heterocyclic alkyl, heterocyclic, heterocyclic alkyl, substituted or unsubstituted phenylcycloalkyl, or substituted or unsubstituted phenylalkyl; wherein the substituent in the substituted or unsubstituted group is optionally selected from 1 to 3 of the following groups: halogen, hydroxyl, C1-C3 alkyl, halo-C1-C3 alkyl, or C1-C3 alkoxy; wherein the alkyl group is substituted by a phenyl group that is substituted 1 to 3 times in any way: hydroxyl, C1-C3 alkyl, halo-C1-C3 alkyl, or C1-C3 alkoxy; R2 is selected from hydrogen, substituted or unsubstituted C1-C6 alkyl, C3-C6 cycloalkyl, (C1-C2 alkoxy)-(C2-C3 alkyl)-, substituted or unsubstituted heterocyclic aryl or phenyl; wherein the substituent in the substituted or unsubstituted C1-C6 alkyl is selected from phenyl substituted 1 to 3 times by any of the following groups: halogen, C1-C3 alkyl, halo-C1-C3 alkyl or C1-C3 alkoxy; the substituent in the substituted or unsubstituted heterocyclic aryl is selected from 1 to 3 of the following groups: C1-C3 alkyl, halo-C1-C3 alkyl or C1-C3 alkoxy; Alternatively, R2 and R3 may form a five- or six-membered heterocyclic alkyl group; And exclude the following situations: R1 and R3 are selected from hydrogen, while R2 is selected from methyl, ethyl, isopropyl, 2-methylpropyl, octyl, cyclopropyl, cyclopentyl, 2-methoxyethyl, 2-ethoxyethyl or phenyl; or R1 is selected from hydrogen or methyl, while R2 and R3 are selected from hydrogen.

2. The compound or its stereoisomer, pharmaceutically acceptable salt, or mixture thereof according to claim 1, characterized in that, The alkyl group is selected from C1-C6 alkyl groups, and the cycloalkyl group is selected from C3-C6 cycloalkyl groups.

3. The compound or its stereoisomer, pharmaceutically acceptable salt, or mixture thereof according to claim 1, characterized in that, The compound is:

4. A compound of formula (II) or a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a mixture thereof: The definitions of R1, R2, and R3 are the same as the corresponding definitions in claim 1; The following conditions are excluded: R1 and R3 are selected from hydrogen, while R2 is selected from methyl, ethyl, isopropyl, 2-methylpropyl, octyl, cyclopropyl, cyclopentyl, 2-methoxyethyl, 2-ethoxyethyl or phenyl; or R1 is selected from hydrogen or methyl, while R2 and R3 are selected from hydrogen.

5. The compound according to claim 4, or its stereoisomers, pharmaceutically acceptable salts, or mixtures thereof, characterized in that, The compound is selected from the following:

6. A compound of general formula (VII) or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a mixture thereof: The definitions of R1, R2, and R3 are the same as the corresponding definitions in claim 1.

7. The compound according to any one of claims 1-6, or its stereoisomer or pharmaceutically acceptable salt, or a mixture thereof, characterized in that, The hydrogen in the compound can be replaced by one or more deuterium atoms.

8. A method for preparing a compound of general formula (I) or general formula (VII), characterized in that, The preparation of compound of general formula (VII) includes the following steps: condensing compound of formula (III) or its salt with compound of formula (V) to obtain compound of formula (VI), and further deprotecting compound of formula (VI) to obtain compound of general formula (VII); Wherein, R1, R2, and R3 are defined as in claim 6, and R4 is selected from methyl, tert-butyl, or benzyl; Alternatively, the preparation of compounds of general formula (I) includes the following methods: Method 1: The compound of general formula (VII) is chelated to obtain the compound of general formula (I); Method 2: A substitution reaction is carried out between a compound of formula (III) or its salt and a compound of formula (IV) to obtain a compound of general formula (I); Wherein, R1, R2, and R3 are defined as in the corresponding definitions of claims 1-4; and LG represents an activated leaving group, preferably phenol, p-chlorophenol, or p-nitrophenol.

9. The synthesis method according to claim 8, characterized in that, The compound of formula (IV) is synthesized by the following steps: esterification of the compound of formula (II) with LG; The definitions of R1, R2, and R3 are the same as those in claim 6, and LG is selected from phenol, p-chlorophenol, or p-nitrophenol.

10. Use of the compound or its stereoisomer, pharmaceutically acceptable salt or mixture thereof as claimed in any one of claims 1-3, the compound or its stereoisomer, pharmaceutically acceptable salt or mixture thereof as claimed in any one of claims 4-5, or the compound or its stereoisomer, pharmaceutically acceptable salt or mixture thereof as claimed in claim 6 in the preparation of a diagnostic agent; preferably, use in the preparation of a contrast agent for magnetic resonance imaging.

11. A pharmaceutical composition comprising the compound or stereoisomer of any one of claims 1-3, a pharmaceutically acceptable salt or a mixture thereof, the compound or stereoisomer of any one of claims 4-5, a pharmaceutically acceptable salt or a mixture thereof, or the compound or stereoisomer of claim 6, a pharmaceutically acceptable salt or a mixture thereof, and a pharmaceutically acceptable carrier and / or excipient.

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