CN118791488B

CN118791488B - Compounds, chelates and their use in magnetic resonance imaging

Info

Publication number: CN118791488B
Application number: CN202411290102.1A
Authority: CN
Inventors: 岑国栋; 王晓宇; 牟霞
Original assignee: Chengdu Shibeikang Biological Medicine Technology Co ltd
Current assignee: Chengdu Shibeikang Biological Medicine Technology Co ltd
Filing date: 2024-09-14
Publication date: 2024-11-15
Anticipated expiration: 2044-09-14

Abstract

The invention discloses a compound, a chelate and application thereof in magnetic resonance imaging, and belongs to the technical field of medical imaging. The chelate formed by chelating the ligand compound and the metal ion Gd ³⁺ has high relaxation rate, higher chelation stability and solubility, solves the problem that the conventional gadolinium contrast agent releases free Gd ³⁺, has good safety, and has potential clinical application prospect as a magnetic resonance imaging contrast agent.

Description

Compounds, chelates and their use in magnetic resonance imaging

Technical Field

The invention belongs to the technical field of medical imaging, and particularly relates to a compound, a metal chelate of the compound and application of the compound in a magnetic resonance imaging contrast agent.

Background

Magnetic resonance imaging (Magnetic Resonance Imaging, MRI) is based on the fact that nuclei with magnetic moment undergo energy transitions under the influence of a magnetic field, and the difference in hydrogen nuclear density between different tissues of the human body is imaged by using the principle of magnetic resonance. Because of the low MRI sensitivity, contrast agents are required to improve the imaging contrast between the focal site and healthy tissue. The magnetic resonance imaging contrast agent containing paramagnetic and superparamagnetic metals can obtain stronger signals and higher spatial ban rate due to the fact that the relaxation time of protons is changed, and is the most commonly used magnetic resonance imaging contrast agent at present.

The first paramagnetic magnetic resonance imaging contrast agent put into clinical use in 1983 was a chelate of diethylenetriamine pentaacetic acid (DTPA) with Gd (iii) (Gd-DTPA) developed by Schering corporation, germany, which is effective in shortening the spin-lattice relaxation time (T ₁) of tissues, and is the most widely used MRI positive contrast agent at present. However, in practical application, it is found that Gd-DTPA belongs to a small molecular ionic contrast agent, has high in vivo osmotic pressure, rapidly enters a cell gap after entering blood, is easy to rapidly excrete after being metabolized by kidneys, has short in vivo residence time, poor tissue or organ selectivity and high price. In view of this, there is a need to develop a magnetic resonance imaging contrast agent with better performance, such as high relaxation rate, good solubility, and high stability, to improve diagnostic imaging efficiency.

Among the patent documents known at present, patent document EP 438206 discloses a bicyclic polyazamacrocyclic carboxylic acid (bicyclopolyazamacrocyclocarboxylic acid) chelate of the formula:

wherein X represents a carboxylate or phosphate group, R ₀ represents an alkyl or phenyl group, or R ₀ is a group bonded to a biomolecule, wherein PCTA is a bicyclic polyazamacrocyclic carboxylic acid chelate known to those skilled in the art, and the relaxation rate of PCTA is about 4-6mM ^-1∙S^-1∙Gd^-1.

Still further, WO93/11800 discloses that the R ₀ group in the above formula is selected from H, OH or C ₁-C₃ alkyl. Patent document US5403572 discloses that the R ₀ group may be an alcohol compound, the synthesis of which comprises first synthesizing the alcohol chain and then coupling the chain to the macrocyclic nitrogen atom by alkylation. Patent document US6450956 discloses that the R ₀ group is-CH ₂-CH₂ -CO-NY-Y, wherein Y is a heavy amino alcohol chain, such as the amino alcohol chain having a molecular weight of 500-1500, which, although being designed to give compounds with higher relaxation rates, also presents problems of expensive synthesis and excessive viscosity of the compounds.

On this basis, patent document CN 103224495A discloses a bicyclic polyazamacrocyclic carboxylic chelate of the formula:

Wherein X represents C ₁-C₃ alkyl, Y represents-CONH ₂、-CO-NR₇R₈ or-NR ₇-CO-R₈, R7 and R8 are each independently selected from H, C ₁-C₆ alkyl, C ₁-C₆ hydroxyalkyl. The chelate disclosed in this patent document has a relaxation rate of 9 to 15 mM ∙ S ^-1∙Gd^-1 at 0.5T in water.

With the development of magnetic resonance imaging technology, the use of gadolinium-based magnetic resonance imaging contrast agents has increased year by year, and although gadolinium contrast agents are currently widely considered clinically safe, adverse reactions such as headache, nausea, taste change or urticaria occur in some patients, and anaphylactic shock or renal-induced systemic fibrosis may occur in very few patients. A part of preclinical studies show that free Gd ³⁺ can be released after intravenous injection of gadolinium-based contrast agent, and free Gd ³⁺ can rapidly distribute to bone and liver with small amount of deposition. Thus, the chelation stability of gadolinium-based contrast agents is a critical factor to be considered in the drug development process.

The technical personnel of the invention find that the prior gadolinium bicyclo polyaza macrocyclic carboxylic acid chelate has room for continuous improvement in the aspects of chelation stability, water solubility and relaxation rate in long-term research. Based on the method, the ligand compound and the chelate formed by chelating the ligand compound with metal ions are developed, and the chelate has the advantages of high relaxation and high stability, and has potential clinical application prospect as a magnetic resonance imaging contrast agent.

Disclosure of Invention

The invention aims to provide a dicyclic polyazamacrocyclic carboxylic acid ligand compound, which is chelated with metal ions to form a chelate, and the chelate has good stability and high relaxation rate and has potential to be developed into a magnetic resonance imaging contrast agent for clinical application. It is also an object of the present invention to provide a process for the preparation of said chelate and a pharmaceutical composition comprising said chelate.

The aim of the invention is realized by the following technical scheme:

in a first aspect of the invention, the present invention provides a ligand compound of formula i, or an enantiomer thereof, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof:

；

Wherein G is selected from OR _a、NR_bR_c、SR_d, wherein R _a、R_b、R_c、R_d are independently selected from hydrogen, deuterium, hydroxyalkyl, mercaptoalkyl, alkoxyalkyl, thioalkyll, aminoalkyl, substituted aminoalkyl, carboxyalkyl, esteralkyl, amidoalkyl, phenyl, phenylalkyl, aromatic heterocyclic alkyl, heterocyclic alkyl OR a group selected from the group consisting of:

C ₁-C₆ alkyl group, C ₃-C₆ cycloalkyl group,

Wherein the C ₁-C₆ alkyl is optionally substituted identically or differently with a phenyl substituent optionally substituted identically or differently 1,2 or 3 times with a halogen atom or a group selected from:

Hydroxy, C ₁-C₃ alkyl, C ₁-C₃ haloalkyl and C ₁-C₃ alkoxy.

Preferably, G is-OH or-OCH ₃.

R ₁、R₂、R₃ is independently selected from-COOH, carboxylate, -P (O) (OH) ₂、R_e-P(O)(OH)₂, phosphate, R _e is selected from H or C ₁-C₃ alkylene.

X is selected from C ₁-C₆ alkyl.

Y represents-CO-NR _fR_g or-NR _f-CO-R_g, wherein R _f、R_g independently of one another represents H or C ₁-C₆ alkyl or C _l-C₆ hydroxyalkyl and at least one of R _f or R _g represents C _l-C₆ hydroxyalkyl.

Preferably, Y is-A-B-R ₄, wherein A is selected from-CO-; b is selected from-N (R ₅)-,R₅ is selected from H, C ₁-C₃ alkyl, C ₁-C₃ hydroxyalkyl; R ₄ is selected from C ₁-C₆ hydroxyalkyl).

K ₁-K₈ is each independently selected from H, - (CH ₂)_j-CH₃ or (CH ₂)_i -OH) wherein j is 0-8,i and 1-8.

Preferably, K ₁-K₈ is each independently selected from-H, C ₁-C₆ alkyl, C ₁-C₆ hydroxyalkyl.

The C ₁-C₆ alkyl groups described herein are selected from any of the linear or branched alkyl groups of C ₁、C₂、C₃、C₄、C₅、C₆ known to those skilled in the art.

Preferably, the C ₁-C₆ alkyl group includes, but is not limited to -CH₂-、-CH₂-CH₂-、-CH₂-CH₂-CH₂-、-CH₂-CH₂-CH₂-CH₂-、-CH₂-CH₂-CH₂-CH₂-CH₂-、-CH₂-CH₂-CH₂-CH₂-CH₂-CH₂-.

The C ₁-C₆ hydroxyalkyl groups described herein are selected from the group consisting of straight or branched alkyl groups of C ₁、C₂、C₃、C₄、C₅、C₆ containing one or more hydroxyl groups. In particular embodiments of the invention, the C _l-C₆ hydroxyalkyl groups include, but are not limited to -CH₂-CH₂OH、-CHOH-CH₂OH、-CH-(CH₂OH)₂、-(CH₂)_p-(CHOH)_q-CH₂OH, where p=1-3, q=1-4, and p+q=2-5, or-C- (CH ₂OH)₃).

The definition of the C ₁-C₃ alkyl or the C ₁-C₃ hydroxyalkyl is the same as that of the C ₁-C₃ alkyl or the C ₁-C₃ hydroxyalkyl, and the description is omitted.

Further, the present invention provides ligand compounds represented by the formula II-1 and the formula II-2:

；

in the above formula, n is selected from integers between 1 and 6, specifically 1,2, 3, 4, 5 and 6.

R ₅ is selected from H, C ₁-C₂ alkyl, C ₁-C₂ hydroxyalkyl;

Specifically, the C ₁-C₂ alkyl is selected from-CH ₃、-CH₂-CH₃; the C ₁-C₂ hydroxyalkyl is selected from the group consisting of-CH ₂OH、-CH₂-CH₂ OH.

R ₄ isWherein R ₆ and R ₇ are selected from H or-CH ₂ OH, m is selected from integers between 0 and 4, in particular 0,1, 2, 3, 4, when m is 0, it means that the-CHOH-group is absent.

In one embodiment of the invention, the ligand compound is represented by formula III-1 a, formula III-1 b:

；

in the above formula, n is selected from 1,2 or 3; m is selected from 0, 1,2,3 or 4, when m is 0, it means that the-CHOH-group is absent.

R ₅ is selected from H, -CH ₃ or-CH ₂-CH₂ OH.

R ₈、R₉、R₁₀ are each independently selected from H or-CH ₂ OH, and at least one of R ₈、R₉、R₁₀ is-CH ₂ OH; preferably, two of R ₈、R₉、R₁₀ are-CH ₂ OH; more preferably, R ₈、R₉、R₁₀ is-CH ₂ OH.

In another embodiment of the present invention, the ligand compound is represented by formula III-2 a, formula III-2 b:

（Ⅲ-2a）、

（Ⅲ-2b）；

R ₅ is selected from H, -CH ₃ or-CH ₂-CH₂ OH.

R ₈、R₉、R₁₀ are each independently selected from H or-CH ₂ OH, and at least one of R ₈、R₉、R₁₀ is-CH ₂ OH; preferably, two of R ₈、R₉、R₁₀ are-CH ₂ OH; more preferably, R ₈、R₉、R₁₀ are each-CH ₂ OH.

In a most preferred embodiment of the invention, the ligand compound is selected from the following structures:

、、

、。

The ligand compounds of the present invention also include stereoisomers, tautomers, N-oxides, hydrates, solvates, or mixtures thereof.

The ligand compounds of the present invention also include multimers, preferably dimers, trimers or tetramers, formed from the ligand compounds described above.

The ligand compound also comprises a conjugate obtained by coupling the ligand compound with any peptide, protein or monoclonal antibody through a bond forming group.

The bond-forming groups include amino, hydroxyl, carboxyl, carbonyl, thiol, thioether, and the like.

Peptides commonly known in the art that can be conjugated to the ligand compounds include, but are not limited to ingr polypeptides, NGR polypeptides, pasireotide pasireotide, triptorelin triptorelin polypeptides, octreotide octreotide.

In a second aspect of the present invention, there is provided a chelate complex of the ligand compound according to the first aspect of the present invention with a metal ion selected from paramagnetic metal ions or radionuclide ions.

The paramagnetic metal ion is selected from Gd ³⁺、Mn²⁺、Yb³⁺ or Fe ³⁺.

The radionuclide ions are selected from ⁹⁹Tc、¹¹⁷Sn、¹¹¹In、⁹⁷Ru、⁶⁷Ga、⁸⁹Zr、¹⁷⁷Lu、⁴⁷Sc、¹⁰⁵Rh、¹⁸⁸Re、⁶⁰Cu、⁶²Cu、⁶⁴Cu、⁶⁷Cu、⁹⁰Y、¹⁵⁹Gd、¹⁴⁹Pr or ¹⁶⁶ Ho.

Preferably, the chelate is a chelate of a ligand compound according to the first aspect of the invention with Gd ³⁺.

Specifically, the chelate has the structure shown in formula AO-I:

（AO-Ⅰ）

wherein G, X, Y, K ₁-K₈、R₁、R₂、R₃ is as described in the first aspect of the invention.

Further, the chelate has a structure represented by the formula AO-II-1, formula AO-II-2:

Wherein n and R ₄、R₅ are as described in the first aspect of the invention.

Further, the chelate has a structure represented by the formula AO-III-1 a, the formula AO-III-1 b, the formula AO-III-2 a, and the formula AO-III-2 b:

Wherein n, m, R ₅、R₈、R₉、R₁₀ are as described in the first aspect of the invention.

In a preferred embodiment of the invention, the chelate has the following formula:

、、

、。

In a third aspect of the invention, the invention provides a method of preparing a chelate comprising the steps of:

wherein G, n, R ₄、R₅ are as described in the first aspect of the invention.

As shown in the formula, the compound M2 is dissolved in water, the pH is regulated to 6, the compound M1 and dioxane are added, HOBT and EDCI are added under stirring, the pH of the reaction solution is regulated to 6, the reaction is carried out for 24 hours, the reduced pressure concentration is carried out, the equal volume of ethanol and diethyl ether are added for dispersing solid, the solid is filtered, and the chelate M is obtained through reversed phase preparation chromatography separation.

In a fourth aspect of the invention, the invention provides a pharmaceutical composition comprising a ligand compound according to the first aspect of the invention or a chelate according to the second aspect of the invention, together with pharmaceutically acceptable adjuvants or auxiliaries.

The pharmaceutically acceptable excipients or auxiliaries are used depending on the particular composition and the particular mode of administration. The pharmaceutical compositions of the present invention exist in a variety of suitable formulations including, but not limited to, lipid nanoparticles prepared by the ligand compounds or chelates.

In a fifth aspect of the invention, the present invention provides the use of a ligand compound according to the first aspect of the invention, a chelate according to the second aspect of the invention or a pharmaceutical composition according to the fourth aspect of the invention for the preparation of a contrast agent for magnetic resonance imaging.

The chelate formed by chelating the ligand compound provided by the invention with Gd ³⁺ shows high relaxation and good pharmacokinetic characteristics. The technical staff of the invention unexpectedly find that the chelate prepared by the invention has higher chelation stability and solubility, particularly shows excellent chelation stability in plasma, overcomes the problem that the conventional gadolinium contrast agent releases free Gd ³⁺ after intravenous injection, and avoids adverse reaction of patients caused by free Gd ³⁺. The Gd ³⁺ chelate prepared by the invention has potential clinical application prospect as a magnetic resonance imaging contrast agent.

Detailed Description

The technical solutions of the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only some embodiments of the present invention, but not all. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.

Interpretation of the terms

The term "C ₁-C₆ alkyl" refers to any straight or branched chain group of C ₁、C₂、C₃、C₄、C₅、C₆ known to those skilled in the art.

The term "C ₁-C₆ hydroxyalkyl" refers to a straight or branched chain group of C ₁、C₂、C₃、C₄、C₅、C₆ containing one or more hydroxyl groups.

The term "pharmaceutically acceptable" means that the substance or composition must be chemically and/or toxicologically compatible with the other ingredients comprising the formulation and/or the mammal being treated therewith.

The term "pharmaceutically acceptable excipients or auxiliaries" may include any solvents, solid excipients, diluents or other liquid excipients, etc., suitable for the particular dosage form of interest.

The following is further illustrated by specific examples, it being understood that these examples are for the purpose of illustration only and are not to be construed as limiting the disclosure in any way.

The names of the compounds corresponding to the English abbreviations are as follows:

HCl: hydrochloric acid

EDCI:1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride

HOBT: 1-hydroxybenzotriazoles

Gd ₂O₃: gadolinium oxide

The structures of the compounds of the examples of the present invention were characterized by nuclear magnetic resonance (¹ H-NMR) or liquid chromatography-mass spectrometry (LC-MS). As used herein, "room temperature" means 10 to 25 ℃.

Preparation of Gd ³⁺ chelate

EXAMPLE 1 Synthesis of chelate 1

0.88G (9.7 mmol,3.6 eq) of 3-amino-1, 2-propanediol was dissolved in 50ml of water, the pH of the reaction solution was adjusted to 6 with 2N HCl, 2.11g (2.7 mmol,1 eq) of Compound X and 40ml of dioxane were added to the reaction solution, 1.57g of HOBT (11.6 mmol,4.3 eq) and 2.22g of EDCI (11.6 mmol,4.3 eq) were continuously added with stirring, the pH of the reaction solution was adjusted to 6, after 24 hours of reaction, the reaction solution was concentrated to about 20ml under reduced pressure, 100ml ethanol and 100ml of diethyl ether were added to the reaction flask to disperse solids, and filtration was carried out to obtain 0.86g (32%, 0.86 mmol) of chelate 1 of example 1 by reverse phase preparative chromatography with a purity of 97.8%.

LC-MS(ESI+)：m/z＝1001.2(M+H)⁺。

EXAMPLE 2 Synthesis of chelate 2

The preparation method is the same as that of example 1, and the 3-amino-1, 2-propanediol in example 1 is replaced by 2-aminoethanol, so that chelate 2 of example 2 can be prepared with purity of 98.2%.

LC-MS(ESI+)：m/z＝911.1(M+H)⁺。

EXAMPLE 3 Synthesis of chelate 3

The preparation method is the same as that of example 1, and the chelate 3 of example 3 can be prepared by replacing 3-amino-1, 2-propanediol in example 1 with 2-amino-1, 3-propanediol, and the purity is 97.3%.

LC-MS(ESI+)：m/z＝1001.3(M+H)⁺。

EXAMPLE 4 Synthesis of chelate 4

The preparation method is the same as that of example 1, and the 3-amino-1, 2-propanediol in example 1 is replaced by 3- (methylamino) -1, 2-propanediol, so that chelate 4 of example 4 can be prepared with a purity of 97.2%.

LC-MS(ESI+)：m/z＝1043.3(M+H)⁺。

EXAMPLE 5 Synthesis of chelate 5

The preparation method is the same as that of example 1, and the 3-amino-1, 2-propanediol in example 1 is replaced by 2-amino-2-hydroxymethyl-1, 3-propanediol, so that chelate 5 of example 5 can be prepared with purity of 98.4%.

LC-MS(ESI+)：m/z＝1091.3(M+H)⁺。

EXAMPLE 6 Synthesis of chelate 8

The preparation method is the same as that of example 1, and the 3-amino-1, 2-propanediol in example 1 is replaced by glucosamine, so that chelate 8 of example 6 can be prepared with purity of 97.4%.

LC-MS(ESI+)：m/z＝1271.5(M+H)⁺。

EXAMPLE 7 Synthesis of chelate complex X

Synthesis of compound int.3:

4.73g (20.0 mmol,1 eq) of int.1 (cf. J. Am. chem. Soc. 2018, 140, 11, 3916-3928), 17.10g (64.0 mmol,3.2 eq) of diethyl 2-bromoglutarate and 11.06g (80.0 mmol,4 eq) of anhydrous K ₂CO₃ are added to 500ml of acetonitrile and stirred under reflux with heating overnight. After the completion of the reaction, the reaction mixture was filtered while it was still hot, the solid was washed with acetonitrile and the filtrate was concentrated, and the concentrate was subjected to silica gel column chromatography to obtain 11.61g (73%, 14.6 mmol) of the objective compound int.3 with a purity of 95.2%.

LC-MS(ESI+)：m/z＝795.8(M+H)⁺。

¹H NMR (400 MHz, CDCl₃) δ 6.72 (s, 2H), 4.18 – 4.12 (m, 12H), 3.88 – 3.62 (m, 10H), 2.89 – 2.81 (m, 2H), 2.80 – 2.72 (m, 2H), 2.65 – 2.55 (m, 4H), 2.51 – 2.32 (m, 6H), 2.08 – 1.81 (m, 6H), 1.26 – 1.23 (m, 18H).

Synthesis of compound Int 4:

10.00g (12.6 mmol) of Int.3 were dissolved in a mixed solution of 60ml of 5N sodium hydroxide and 60ml of methanol. The reaction solution is heated, refluxed and stirred for 5 to 8 hours, after the reaction is finished, the reaction solution is concentrated, water is used for dissolving, the aqueous solution is subjected to ion exchange through Amberlite IRC50 resin, the obtained solution is concentrated, and the obtained solution is solidified and crystallized by ethanol, so that 6.77g (86 percent, 10.8 mmol) of target compound int.4 with the purity of 95.8 percent is obtained.

LC-MS(ESI+)：m/z＝627.7(M+H)⁺,LC-MS(ESI-)：m/z＝625.4(M-H)⁺。

¹H NMR (400 MHz, CD₃OD) δ 6.70 (s, 2H), 3.84 – 3.76 (m, 5H), 3.70 – 3.60 (m, 5H), 2.86 – 2.73 (m, 4H), 2.62 – 2.54 (m, 4H), 2.48 – 2.29 (m, 6H), 2.06 – 1.73 (m, 6H).

Synthesis of chelate X:

6.27g (10.0 mmol) of int.4 was dissolved in 150ml of water, the pH was adjusted to 6, 1.99g (5.5 mmol) of Gd ₂O₃ was added to the solution, and the reaction solution was heated to 60℃to react for 8 hours. The reaction solution was concentrated and solidified and crystallized with ethanol to obtain 6.09g (78%, 7.8 mmol) of the target chelate X with a purity of 96.7%.

LC-MS(ESI+)：m/z＝782.3(M+H)⁺。

EXAMPLE 8 Synthesis of chelate 10

The preparation method is the same as that of the example 1, and the chelate X in the example 1 is replaced by the chelate Y, so that the chelate 10 of the example 8 can be prepared with the purity of 96.8%.

LC-MS(ESI+)：m/z＝987.4(M+H)⁺。

EXAMPLE 9 Synthesis of chelate 11

The preparation method is the same as that of example 2, and chelate X in example 2 is replaced by chelate Y, so that chelate 11 of example 9 can be prepared with purity of 96.2%.

LC-MS(ESI+)：m/z＝897.3(M+H)⁺。

EXAMPLE 10 Synthesis of chelate 12

The preparation method is the same as that of example 3, and chelate X in example 3 is replaced by chelate Y, so that chelate 12 of example 9 can be prepared with purity of 98.3%.

LC-MS(ESI+)：m/z＝987.2(M+H)⁺。

EXAMPLE 11 Synthesis of chelate 13

The preparation method is the same as that of example 4, and chelate X in example 4 is replaced by chelate Y, so that chelate 13 of example 11 can be prepared with purity of 98.4%.

LC-MS(ESI+)：m/z＝1029.3(M+H)⁺。

EXAMPLE 12 Synthesis of chelate 14

The preparation method is the same as that of example 5, and chelate 14 of example 12 can be prepared by replacing chelate X in example 5 with chelate Y, and the purity is 97.7%.

LC-MS(ESI+)：m/z＝1077.5(M+H)⁺。

EXAMPLE 13 Synthesis of chelate 17

The preparation method is the same as that of example 6, and chelate X in example 6 is replaced by chelate Y, so that chelate 17 of example 13 can be prepared with purity of 97.4%.

LC-MS(ESI+)：m/z＝1257.4(M+H)⁺。

EXAMPLE 14 Synthesis of chelate complex Y

Synthesis of compound int.6:

the preparation method is the same as that of int.3 in example 7, and int.1 in the preparation method is replaced by int.5 (refer to chem. Commun., 2013, 49, 2712-2714 for synthesis), so that compound int.6 can be prepared with purity of 96.1%.

LC-MS(ESI+)：m/z＝871.6(M+H)⁺。

¹H NMR (400 MHz, CDCl₃) δ 7.46 – 7.41 (m, 2H), 7.37 – 7.33 (m, 2H), 7.32 – 7.25 (m, 1H), 6.70 (s, 2H), 4.99 (s, 2H), 4.18 – 4.09 (m, 12H), 3.87 – 3.61 (m, 7H), 2.88 – 2.80 (m, 2H), 2.78– 2.70 (m, 2H), 2.63 – 2.52 (m, 4H), 2.51 – 2.32 (m, 6H), 2.08 – 1.81 (m, 6H), 1.26 – 1.23 (m, 18H).

Synthesis of compound Int 7:

8.70g (10.0 mmol) of int.7 was added to 100ml of methanol solution, 1.22g (10.0 mmol,1 eq) of palladium oxide was added to the reaction system, benzyl groups were removed by catalytic hydrogenation, after the reaction was completed, the reaction solution was filtered, the filtrate was concentrated under reduced pressure, and the concentrate was subjected to silica gel column chromatography to give 5.08g (65%, 6.5 mmol) of the objective compound int.7 with a purity of 97.3%.

LC-MS(ESI+)：m/z＝781.7(M+H)⁺。

¹H NMR (400 MHz, CDCl₃) δ 6.56 (s, 2H), 4.15– 4.11 (m, 12H), 3.86 – 3.70 (m, 5H), 3.61 – 3.58 (m, 2H), 2.90 – 2.81 (m, 2H), 2.80 – 2.71 (m, 2H), 2.63 – 2.53 (m, 4H), 2.52 – 2.31 (m, 6H), 2.07 – 1.81 (m, 6H), 1.26 – 1.23 (m, 18H).

Synthesis of compound Int 8:

the preparation method is the same as that of Int.4 in example 7, and Int.3 in the preparation method is replaced by Int.7, so that the compound Int.8 can be prepared with the purity of 95.2%.

LC-MS(ESI+)：m/z＝613.4(M+H)⁺；

¹H NMR (400 MHz, CD₃OD) δ 6.60 (s, 2H), 3.82 – 3.80 (m, 2H), 3.68 – 3.64 (m, 5H), 2.85 – 2.74 (m, 4H), 2.62 – 2.54 (m, 4H), 2.48 – 2.29 (m, 6H), 2.05 – 1.81 (m, 6H).

Synthesis of chelate Y:

The preparation method is the same as that of the chelate X in the example 7, and the chelate Y can be prepared by replacing Int.4 in the preparation method with Int.8, and the purity is 95.6%.

LC-MS(ESI+)：m/z＝768.2(M+H)⁺。

Comparative example 1 Synthesis of chelate complexes 1-Gadopiclenol

Chelate complexes 1-Gadopiclenol were prepared as comparative example 1 of the present invention according to the preparation method of example 2 in patent CN101305006a, purified by reverse phase preparative chromatography, and dried to give the compound of target comparative example 1 with a purity of 97.5%. Structural identification was consistent, LC-MS (esi+): m/z= 971.1 (m+h) ⁺, i.e. the molecular weight of the main product is 970.1. The chemical structure of the product was as shown in comparative example 1 above.

Effect test example 1 relaxation measurement at 3.0T

Relaxation measurements at 3.0T were made by a whole-body 3.0T MRI scanner using knee coils. The sample tubes were placed in 3 columns of 4 and 5 tubes in a plastic seat in a water filled tank and the temperature was adjusted to 37 ℃.

For MRI sequences, the shortest possible echo Time (TE) of 7.46 milliseconds is used. The inversion time is selected to optimize the sequence to measure T1 values corresponding to the estimated T1 range for all relaxation times of the contrast agent-containing solution. The following inversion Times (TI) were used: 50. 100, 150, 200, 300, 500, 700, 1000, 1400, 2100, 3200, and 4500 milliseconds. After registering the last echo, the sequence runs at a constant relaxation delay of 3.4 seconds (the range of variable TR is 3450-7900 milliseconds).

Relaxation was assessed using three different concentrations of each compound (3 concentrations ranging from 0.05 to 2 mM).

The T1 relaxation times of the compounds of the examples were measured in water and human plasma. Human plasma preparation: for each experiment, fresh blood was removed from volunteers using a 10mL citrate tube. The 10mL citrate tube was carefully inverted ten times to mix the blood and anticoagulant and centrifuged at room temperature for 15 minutes.

The relaxation r ₁ is calculated based on the relaxation time T1 measured in water and plasma:

where "T1 intrinsic" is the relaxation time of the corresponding blank solvent, "[ contrast agent ]" represents the concentration of the compound normalized to gadolinium, and the results are shown in table 1.

TABLE 1 relaxivity (normalized to Gd) in water and human plasma at 3.0T,37 ℃ [ L/mmol/s ]

As can be seen from the results of the table, the relaxation of the chelates 1-5, 8, 10-14 and 17 prepared by the invention is obviously stronger than that of 1-Gadopiclenol, and the contrast effect of the chelates 1-5, 8, 10-14 and 17 is better; in particular, the chelate compounds 5, 8, 14, 17 have a more pronounced relaxation and a more advantageous contrast effect.

Effect test example 2 Tris measurement of chelate stability in HCl buffer

The chelate complex prepared in the example of the present invention and the chelate complex prepared in the comparative example 1 were dissolved in 10mM Tris-HCl buffer (pH 7.4) respectively, and the final concentration was 5mmolGd/L. An aliquot was removed and the remaining portion of the clear and colorless solution was autoclaved at 121 ℃ for 20min; after autoclaving, the solution is still clear and colorless. The amount of released Gd ³⁺ of the chelate before and after autoclaving was determined by HPLC-ICP-MS, in particular the change in the chromatogram of the intensity of Gd ³⁺ before and after autoclaving was monitored by ICP-MS. The results are shown in the following table:

TABLE 2 release of Gd ³⁺ from solution

Note that: the quantitative limit was 0.1% of the total gadolinium.

As can be seen from the results of the table, the stability of the chelate 1, 5, 8, 14 and 17 prepared by the invention in Tris-HCl buffer solution is better than that of the chelate 1-Gadopiclenol, the change of the chromatograph before and after autoclaving is not detected in the detection system of the chelate 1, 5, 8, 14 and 17, which indicates that the chelate prepared by the invention is stable in the autoclaving operation process, the release amount of Gd ³⁺ is less than 0.1 percent, the quantitative limit is lower, and the stability is better than that of 1-Gadopiclenol.

Effect test example 3 measurement of chelate stability in plasma

The chelate prepared in the example of the present invention and the chelate prepared in comparative example 1 were dissolved in human plasma at a concentration of 1mmol Gd/L, respectively. As a reference for released Gd ³⁺, 0.1mmol/L gadolinium chloride (GdCl ₃) was dissolved in human plasma. The plasma samples were incubated at 37℃for 15 days in a 5% CO ₂ atmosphere, maintaining a pH of 7.4. Aliquots were removed at the beginning and end of the incubation, and the amount of released Gd ³⁺ from the chelate was determined by HPLC-ICP-MS, in particular by ICP-MS monitoring, and the change in the chromatogram of the monitored Gd intensity was assessed by peak area analysis. The results are shown in the following table:

TABLE 3 release of Gd ³⁺ in plasma

Note that: the quantitative limit is 0.1% of the total amount of gadolinium

From the above table results, it can be seen that no chromatographic change before and after measurement is detected in the chelate 1, 5, 8, 14 and 17 detection system prepared by the invention, which indicates that no free Gd ³⁺ ion exists in the system, and that chelate 1, 5, 8, 14 and 17 is stable in plasma, gd ³⁺ release amount is less than 0.1%, and the stability of chelate in plasma is better than 1-Gadopiclenol.

Effect test example 4 Water solubility measurement

The water solubility of the compounds was determined in a microcentrifuge tube in 0.5mL of buffer (10 mM Tris-HCl) at room temperature (20 ℃). The measuring method comprises the following steps: the solid compound was gradually added to the buffer solution, the suspension was mixed using a shaker, and treated in an ultrasonic bath for 5min until insoluble substances were present. The solubility of the compound, which is the mass of solute dissolved when a solid substance reaches a saturated state in 100g of solvent at a certain temperature, was calculated. The measurement results are shown in the following table:

TABLE 4 Water solubility

As can be seen from the results of the table, the solubility of each of the compounds 1, 5, 8, 14 and 17 prepared by the invention is more than 1000mg/100ml, and the compounds can be used as contrast agents for intravenous injection administration.

Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some or all of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit of the invention.

Claims

1. A chelate compound, characterized in that the chelate compound has a structure represented by the formula (AO-ii-1), formula (AO-ii-2):

Wherein n is selected from integers between 1 and 6;

R ₅ is selected from H, C ₁-C₂ alkyl;

R ₄ is Wherein R ₆ and R ₇ are selected from H or-CH ₂ OH, m is selected from integers between 0 and 4, and when m is 0, it means that the-CHOH-group is absent.

2. The chelate of claim 1, wherein the chelate has a structure represented by the formula (AO-iii-1 a), formula (AO-iii-2 a):

Wherein n is selected from 1, 2 or 3; m is selected from 0, 1, 2, 3 or 4, when m is 0, it means that a-CHOH-group is absent; r ₅ is selected from H, -CH ₃.

3. The chelate of claim 1, wherein the chelate has a structure represented by the formula (AO-III-1 b) or the formula (AO-III-2 b)

Wherein n is selected from 1,2 or 3; r ₅ is selected from H, -CH ₃;R₈、R₉、R₁₀ are each independently selected from H or-CH ₂ OH, and at least one of R ₈、R₉、R₁₀ is-CH ₂ OH.

4. Chelate according to claim 1, characterized in that it has the following formula:

、、

、。

5. A method for preparing chelate, comprising the following steps:

；

wherein G is-OH or-OCH ₃;

n is selected from integers between 1 and 6;

R ₅ is selected from H, C ₁-C₂ alkyl;

R ₄ is Wherein R ₆ and R ₇ are selected from H or-CH ₂ OH, m is selected from integers between 0 and 4, when m is 0, it means that the-CHOH-group is absent;

6. A pharmaceutical composition comprising the chelate of any one of claims 1-4, and a pharmaceutically acceptable adjuvant.

7. Use of a chelate according to any one of claims 1-4 or a pharmaceutical composition according to claim 6 for the preparation of a contrast agent for magnetic resonance imaging.