Gadolinium-based cationic polymer contrast agent and preparation method thereof

Abstract

The gadolinium-based cationic polymer contrast agent is synthesized by chelating a cationic polymer and gadolinium ions, has high biological safety and can be used for cartilage imaging. The prepared contrast agent adopts an articular cavity direct injection method, so that the imaging effect is improved, and the contrast dosage is reduced. When the cartilage activity is normal, the content of glycosaminoglycan in the matrix is rich, and when the cartilage is inactivated or the activity is reduced, the content of glycosaminoglycan in the matrix is obviously reduced. The gadolinium-based cationic polymer contrast agent is combined with negatively charged glycosaminoglycan in a cartilage matrix through the charge effect. The gadolinium-based cationic polymer contrast agent can be used for noninvasive and quantitative evaluation of cartilage activity in nuclear magnetic resonance imaging, especially cartilage activity and stability in transplantation.

Description

TECHNICAL FIELD

[0001] The present application relates to a kind of polycation contrast agent and its preparation method, especially a kind of joint cartilage gadolinium-based contrast agent and its preparation method. BACKGROUND

[0002] Joint cartilage is mainly composed of extracellular matrix and a small amount of chondrocytes, and the extracellular matrix is mainly a network composed of collagen fibers and proteoglycans, with glycosaminoglycans (GAG) as the main component. GAG, also known as mucopolysaccharide, is a heteropolysaccharide with at least one monosaccharide residue with a negative charge carboxyl or sulfate group in its disaccharide unit. In the early stage of cartilage necrosis, the microstructure of cartilage is destroyed and loses function due to the loosening of collagen fibers and the loss of GAG. Therefore, the decrease in GAG content is the gold standard for judging cartilage inactivation.

[0003] The evaluation of the activity of cartilage after transplantation is the bottleneck of clinical imaging diagnosis, and existing imaging methods cannot accurately evaluate the biological activity of cartilage. Nuclear magnetic resonance imaging (MRI) has good soft tissue resolution and can observe joint cartilage from multiple directions. The present application aims to change the GAG content of postoperative cartilage and invent a cationic polymer contrast agent that adsorbs to cartilage through charge interaction with GAG. Gadolinium-based chelate contrast agents belong to paramagnetic T1 contrast agents and are the most widely used in clinical practice. In the present application, branched PEI is reacted with diethylenetriamine pentaacetic acid (DTPA), and then gadolinium ions are chelated to prepare a new type of cationic contrast agent. The gadolinium-based cationic polymer is a small-sized flexible segment with the characteristics of gadolinium ion contrast agent stability, good biocompatibility, and high-density positive charge on the surface, which is used for MRI imaging to evaluate cartilage activity. SUMMARY

[0004] The present application aims to provide a gadolinium-based cationic polymer contrast agent for nuclear magnetic resonance imaging, which can be used to evaluate the activity of joint cartilage transplantation in vivo.

[0005] Another object of the present application is to provide a preparation method of a gadolinium-based cationic polymer contrast agent.

[0006] The gadolinium-based cationic polymer contrast agent described in the present application is composed of polyetherimide (PEI) as a cationic polymer and chelated gadolinium ions, and its chemical structure is shown below: wherein m and n are the degree of polymerization, and the relative molecular weight of PEI is 300-70000.

[0007] The preparation method of the gadolinium-based cationic polymer contrast agent comprises the following steps: Step 1: a proper amount of PEI is dissolved in a 0.1 M sodium bicarbonate (NaHCO3) solution, and diethylene triamine pentaacetic acid dianhydride (DTPAA) is dissolved in anhydrous dimethyl sulfoxide, and then added dropwise into the above PEI solution. A 1 M NaOH solution is used to adjust the pH value to 8.5. Stir at room temperature for 2 h, and then perform dialysis treatment with deionized water as the dialysis liquid; Step 2: gadolinium chloride hexahydrate (GdCl3 6H2O) is dissolved in a 0.1 M sodium acetate buffer solution, and then added dropwise into the solution of step 1, and then stirred at room temperature for 24 h; Step 3: the reaction product of step 2 is subjected to dialysis treatment, and the dialysis liquid is sequentially 0.1 M citric acid buffer solution and deionized water. After dialysis, freeze-drying is performed to obtain the gadolinium-based cationic polymer contrast agent.

[0008] In step 1, the concentration of PEI is preferably 0.2 M; the mass ratio of GdCl3 6H2O:DTPAA:PEI is 1:1-2:2-10.

[0009] In step 2, the pH value of the sodium acetate buffer solution is preferably 6.0, and the pH value of the citric acid buffer solution is preferably 6.5.

[0010] The beneficial effects brought by the technical scheme of the present application Compared with the prior art, the present application has the following outstanding advantages: The gadolinium-based cationic polymer contrast agent prepared according to the present application has mild preparation conditions. After intra-articular injection, due to the potential difference between the large number of surface positive charges of the contrast agent and the negatively charged GAG in the cartilage matrix, the contrast agent is quickly delivered to the cartilage matrix by the electrostatic interaction between the charges, and the correlation between the GAG content and the nuclear magnetic signal intensity is established by MRI to accurately evaluate the cartilage activity. The present application overcomes the problem that the existing diagnosis can only observe the morphological changes of the cartilage, and has outstanding advantages in non-invasive diagnosis of the internal microstructure of the transplanted cartilage.

[0011] The prepared contrast agent is directly injected into the joint cavity to improve the imaging effect and reduce the amount of contrast agent. When the cartilage activity is normal, the content of glycosaminoglycan in the matrix is rich, and when the cartilage is inactivated or the activity decreases, the content of glycosaminoglycan in the matrix decreases significantly. The gadolinium-based cationic polymer contrast agent of the present application binds to the negatively charged glycosaminoglycan in the cartilage matrix through charge interaction. The gadolinium-based cationic polymer contrast agent can be used for non-invasive and quantitative evaluation of cartilage activity, especially the cartilage activity and stability during transplantation. BRIEF DESCRIPTION OF DRAWINGS

[0012] Figure 1 The T1 nuclear magnetic signal of the gadolinium-based cationic polymer contrast agent prepared in Examples 1-4 of the present application. DETAILED DESCRIPTION

[0013] The preparation method of the gadolinium-based cationic polymer contrast agent according to the present application is further illustrated below with examples.

[0014] Example 1 A gadolinium-based cationic polymer contrast agent was prepared according to the following steps: Step 1: 5 g of PEI was dissolved in 0.1 M NaHCO3 solution to obtain a PEI solution; 5 g of DTPAA was dissolved in anhydrous dimethyl sulfoxide, and then added dropwise to the above PEI solution; 1 M NaOH solution was used to adjust the pH value to 8.5; the reaction was stirred at room temperature for 2 h, and then dialysis treatment was performed using deionized water as the dialysis liquid; Step 2: 2.5 g of GdCl3 6H2O was dissolved in 0.1 M NaAc buffer solution (pH 6.0), and then added dropwise to the solution of step 1; the reaction was stirred at room temperature for 24 h; Step 3: the reaction product of step 2 was subjected to dialysis treatment, and the dialysis liquid was sequentially 0.1 M citric acid buffer solution (pH 6.5) for dialysis 3 times, and then deionized water for dialysis until the pH value was neutral; after dialysis, freeze-drying was performed to obtain the gadolinium-based cationic polymer contrast agent.

[0015] Example 2 A gadolinium-based cationic polymer contrast agent was prepared according to the following steps: Step 1: 3 g of PEI was dissolved in 0.1 M NaHCO3 solution to obtain a PEI solution; 1 g of DTPAA was dissolved in anhydrous dimethyl sulfoxide, and then added dropwise to the above PEI solution. 1 M NaOH solution was used to adjust the pH value to 8.5; the reaction was stirred at room temperature for 2 h, and then dialysis treatment was performed using deionized water as the dialysis liquid; Step 2: 1 g of GdCl3 6H2O was dissolved in 0.1 M NaAc buffer solution (pH 6.2), and then added dropwise to the solution of step 1; the reaction was stirred at room temperature for 24 h; Step 3: the reaction product of step 2 was subjected to dialysis treatment, and the dialysis liquid was 0.1 M citric acid buffer solution (pH 6.5); dialysis was performed 3 times, and then deionized water was used for dialysis until the pH value was neutral; after dialysis, freeze-drying was performed to obtain the gadolinium-based cationic polymer contrast agent.

[0016] Example 3 A gadolinium-based cationic polymer contrast agent was prepared according to the following steps: Step 1: 5 g PEI was dissolved in 0.1 M NaHCO3solution, 2 g DTPAA was dissolved in anhydrous dimethyl sulfoxide, and then added dropwise to the above PEI solution. The pH was adjusted to 8.5 using 1 M NaOH solution; the reaction was stirred at room temperature for 2 h, and then dialyzed with deionized water as the dialysis liquid; Step 2: 1 g GdCl3 6H2O was dissolved in 0.1 M NaAc buffer (pH 6.0), and then added dropwise to the solution of step 1, and the reaction was stirred at room temperature for 24 h; Step 3: The reaction product of step 2 was dialyzed with 0.1 M citric acid buffer (pH 6.8) as the dialysis liquid for 3 times, and then dialyzed with deionized water until the pH was neutral; after dialysis, it was freeze-dried to obtain the gadolinium-based cationic polymer contrast agent.

[0017] Example 4 A gadolinium-based cationic polymer contrast agent was prepared according to the following steps: Step 1: 5 g PEI was dissolved in 0.1 M NaHCO3solution, 2 g DTPAA was dissolved in anhydrous dimethyl sulfoxide, and then added dropwise to the above PEI solution. The pH was adjusted to 8.5 using 1 M NaOH solution; the reaction was stirred at room temperature for 2 h, and then dialyzed with deionized water as the dialysis liquid; Step 2: 1 g GdCl3 6H2O was dissolved in 0.1 M NaAc buffer (pH 6.0), and then added dropwise to the solution of step 1, and the reaction was stirred at room temperature for 24 h; Step 3: The reaction product of step 2 was dialyzed with 0.1 M citric acid buffer (pH 6.8) as the dialysis liquid for 3 times, and then dialyzed with deionized water until the pH was neutral; after dialysis, it was freeze-dried to obtain the gadolinium-based cationic polymer contrast agent.

Claims

1. A gadolinium-based cationic polymer contrast agent, which is composed of polyethyleneimine (PEI) as a cationic polymer and chelated gadolinium ions, and has the following chemical structure: wherein m and n are the polymerization degrees, and the relative molecular weight of PEI is 300-70,000. ， wherein 2.A preparation method of a gadolinium-based cationic polymer contrast agent, comprising the following steps: Step 1: a proper amount of PEI is dissolved in a 0.1 M sodium bicarbonate (NaHCO3) solution, and diethylene triamine pentaacetic dianhydride (DTPAA) is dissolved in anhydrous dimethyl sulfoxide, and then added dropwise into the above PEI solution; a 1 M NaOH solution is used to adjust the pH value to 8.5; the reaction is stirred at room temperature for 2 h, and then dialysis treatment is performed with deionized water as a dialysis liquid; Step 2: gadolinium chloride hexahydrate (GdCl3 6H2O) is dissolved in a 0.1 M sodium acetate buffer solution, and then added dropwise into the solution in step 1; the reaction is stirred at room temperature for 24 h; Step 3: the reaction product in step 2 is subjected to dialysis treatment with 0.1 M citric acid buffer solution and deionized water as dialysis liquids in sequence; and then freeze-drying is performed after dialysis to obtain the gadolinium-based cationic polymer contrast agent. In step 1, the concentration of PEI is preferably 0.2 M; and the mass ratio of GdCl3 6H2O:DTPAA:PEI is 1:1-2:2-10.

3. The method for preparing a gadolinium-based cationic polymer contrast agent according to claim 2, characterized in that... In step 2, the pH value of the sodium acetate buffer solution is preferably 6.0, and the pH value of the citric acid buffer solution is preferably 6.

5.

4. The method for preparing a gadolinium-based cationic polymer contrast agent according to claim 3, characterized in that... Preparation is performed according to the following steps:

5. A method for preparing a gadolinium-based cationic polymer contrast agent according to any one of claims 2 to 4, characterized in that, Step 1: 5 g of PEI is dissolved in a 0.1 M sodium bicarbonate (NaHCO3) solution to obtain a PEI solution; 5 g of diethylene triamine pentaacetic dianhydride (DTPAA) is dissolved in anhydrous dimethyl sulfoxide, and then added dropwise into the above PEI solution; a 1 M NaOH solution is used to adjust the pH value to 8.5; the reaction is stirred at room temperature for 2 h, and then dialysis treatment is performed with deionized water as a dialysis liquid; Step 2: 2.5 g of gadolinium chloride hexahydrate (GdCl3 6H2O) is dissolved in a 0.1 M sodium acetate buffer solution (pH 6.0), and then added dropwise into the solution in step 1; the reaction is stirred at room temperature for 24 h; Step 3: the reaction product in step 2 is subjected to dialysis treatment with 0.1 M citric acid buffer solution (pH 6.5) as a dialysis liquid, dialysis is performed for 3 times, and then deionized water is used for dialysis until the pH value is neutral; and then freeze-drying is performed after dialysis to obtain the gadolinium-based cationic polymer contrast agent. Preparation is performed according to the following steps:

6. The method for preparing a gadolinium-based cationic polymer contrast agent according to any one of claims 2 to 4, characterized in that, Step 1: 3 g of PEI is dissolved in a 0.1 M NaHCO3 solution to obtain a PEI solution; 1 g of DTPAA is dissolved in anhydrous dimethyl sulfoxide, and then added dropwise into the above PEI solution; a 1 M NaOH solution is used to adjust the pH value to 8.5; the reaction is stirred at room temperature for 2 h, and then dialysis treatment is performed with deionized water as a dialysis liquid; Step 2: 1 g of GdCl3 6H2O is dissolved in a 0.1 M sodium acetate buffer solution (pH 6.2), and then added dropwise into the solution in step 1; the reaction is stirred at room temperature for 24 h; ​ Step 3: The reaction product of step 2 was dialyzed against 0.1 M citric acid buffer (pH 6.5) for 3 times, and then against deionized water until the pH was neutral; after dialysis, the product was lyophilized to obtain the gadolinium-based cationic polymeric contrast agent.

7. The method for preparing a gadolinium-based cationic polymer contrast agent according to any one of claims 2 to 4, characterized in that, Preparation was carried out according to the following steps: Step 1: 5 g of PEI was dissolved in 0.1 M NaHCO3 solution, 2 g of DTPAA was dissolved in anhydrous dimethyl sulfoxide, and then added dropwise to the above PEI solution. The pH value was adjusted to 8.5 using 1 M NaOH solution; the reaction was stirred at room temperature for 2 h, and then dialyzed against deionized water; Step 2: 1 g of GdCl3 6H2O was dissolved in 0.1 M sodium acetate buffer (pH 6.0), and then added dropwise to the solution of step 1; the reaction was stirred at room temperature for 24 h; Step 3: The reaction product of step 2 was dialyzed against 0.1 M citric acid buffer (pH 6.8) for 3 times, and then against deionized water until the pH was neutral; after dialysis, the product was lyophilized to obtain the gadolinium-based cationic polymeric contrast agent.

8. A method for preparing a gadolinium-based cationic polymer contrast agent according to any one of claims 2 to 4, characterized in that, Preparation was carried out according to the following steps: Step 1: 5 g of PEI was dissolved in 0.1 M NaHCO3 solution, 2 g of DTPAA was dissolved in anhydrous dimethyl sulfoxide, and then added dropwise to the above PEI solution. The pH value was adjusted to 8.5 using 1 M NaOH solution; the reaction was stirred at room temperature for 2 h, and then dialyzed against deionized water; Step 2: 1 g of GdCl3 6H2O was dissolved in 0.1 M sodium acetate buffer (pH 6.0), and then added dropwise to the solution of step 1; the reaction was stirred at room temperature for 24 h; Step 3: The reaction product of step 2 was dialyzed against 0.1 M citric acid buffer (pH 6.8) for 3 times, and then against deionized water until the pH was neutral; after dialysis, the product was lyophilized to obtain the gadolinium-based cationic polymeric contrast agent.

9. The use of the gadolinium-based cationic polymeric contrast agent according to claim 1 for evaluating cartilage activity in nuclear magnetic resonance imaging, especially for evaluating cartilage activity after cartilage transplantation.

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