Preparation of a PET imaging agent targeting ACE2 for differential diagnosis of COVID-19 infection and tumors

Abstract

The present invention belongs to the C07K technical field, and more specifically relates to the preparation and application of a PET imaging agent targeting ACE2 for the differential diagnosis of novel coronavirus infection and tumors. A PET imaging agent targeting ACE2 for the differential diagnosis of novel coronavirus infection and tumors is prepared, comprising the following steps: S1: 18 Purification of F; S2: Preparation of PET imaging agent. The present invention prepares PET imaging agent ( 18 The F-MLN-4760 method is simple, with mild reaction conditions and shortened reaction time compared to common substitution reactions. The structure was verified by LC-MS, and two chlorines were simultaneously replaced by fluorines.

Description

Technical Field

[0001] The present invention belongs to the field of C07K technology, and more specifically relates to the preparation of a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors. Background Art

[0002] Angiotensin-converting enzyme 2 (ACE2) has been used as a protease for more than 20 years. ACE2 is the functional receptor of the S glycoprotein of coronaviruses and their variants, such as SARS and the new coronavirus. The new coronavirus invades lung cells by binding to the S protein and ACE2, and studies have found that the affinity between ACE2 and the receptor binding domain of the new coronavirus is 10-20 times higher than that of the SARS virus. Therefore, selecting an effective and selective ACE2 inhibitor and analyzing the distribution of the inhibitor in vivo is a method to determine the tissues and organs susceptible to infection by the new coronavirus. At the same time, studies have found that genetic abnormalities such as abnormal amplification of ACE2 mutation copies can occur in tumors. It is known that ACE2 is highly expressed in tumors such as colorectal adenocarcinoma, renal papillary cell carcinoma, pancreatic cancer, gastric adenocarcinoma, and lung adenocarcinoma, and constitutes an important marker for the differential diagnosis of the corresponding tumors.

[0003] MLN-4760 is an effective and selective ACE2 inhibitor. When ordinary MLN-4760 enters the animal body, it cannot be detected in vivo by conventional biochemical methods. Therefore, how to conveniently and quickly detect the distribution of MLN-4760 in the body is a problem that needs to be solved at present.

[0004] pass 18 F-labeled ligands for receptor imaging are the main areas of research in molecular nuclear medicine today. Positron emission tomography (PET) can provide monitoring data with high sensitivity and high resolution. Among the large number of positron-emitting nuclides, 18 F has a small atomic radius, a half-life of 109.8 minutes, a maximum positron energy of 0.64 MeV, and an average range in tissue of 0.22 mm. It has excellent nuclear and chemical properties and is the preferred nuclide for labeling ligands such as peptides. The dichloro-substituted benzene ring structure in the structure of MLN-4760 makes it easy to replace chlorine with 18 F. This makes it easier to detect 18 The distribution of F-MLN-4760 in the body makes it an ACE2-targeted PET imaging agent.

[0005] Explore the possibility of replacing chlorine in MLN-4760 with 18 The synthesis method of F and its application in PET are the research objectives of the present invention. Summary of the invention

[0006] In order to solve the above technical problems, the first aspect of the present invention provides a preparation of a PET imaging agent targeting ACE2 for the differential diagnosis of novel coronavirus infection and tumors, comprising the following steps:

[0007] S1: 18 Purification of F.

[0008] S2: Preparation of PET imaging agent.

[0009] In some preferred embodiments, the step S1 18 The specific operation of purifying F includes the following steps:

[0010] 1) Enrichment of cyclotron produced by sep-pak QMA column 18 F ions;

[0011] 2) Elute the sep-pak QMA column with saline to recover 18 F, spare.

[0012] In some preferred embodiments, the specific operation of preparing the PET imaging agent described in S2 comprises the following steps:

[0013] S2-1: dissolving the ACE2 inhibitor MLN-4760 in the solution;

[0014] S2-2: Take the solution in step S1 again 18 F is heated for 0.5h-1h to azeotropically remove water, and mixed with the solution in step S2-1 to obtain solution A for standby use;

[0015] S2-3: Add phase reaction catalyst to solution A, seal in a high pressure reactor, and react at 170-220°C for 2-4h;

[0016] S2-4: After the reaction is completed, the mixture is cooled to room temperature, enriched using a C-18 column, and eluted with an ethanol-physiological saline solution to obtain a PET imaging agent.

[0017] It should be noted that the ACE2 inhibitor refers to MLN4760.

[0018] The PET imaging agent prepared by this application is 18 F-MLN-4760.

[0019] MLN4760 and 18 The structural formula of F-MLN-4760 is shown in formula (1) and formula (2):

[0020]

[0021]

[0022] In some preferred embodiments, the solutions described in S2-1 and S2-2 are aprotic polar solutions.

[0023] In some preferred embodiments, the aprotic polar solution includes at least one of dimethyl sulfoxide, N,N-dimethylformamide, 1,3-dimethyl-2-imidazolidinone, cyclopentane, hexamethylphosphoric triamide, and acetonitrile.

[0024] It should be noted that the type of aprotic polar solution added in the present application is the solvent provided in the present application, and the technology of the present application can be implemented in the preparation of the present application.

[0025] Preferably, the aprotic polar solution is dimethyl sulfoxide.

[0026] In some preferred embodiments, the phase reaction catalyst includes at least one of crown ethers, quaternary ammonium salts, quaternary phosphonium salts, and metal halide salts.

[0027] In some preferred embodiments, the phase reaction catalyst includes tetramethylammonium chloride, tetrabutylammonium bromide, tetraphenylphosphonium bromide, PEG with a molecular weight greater than 1000, diether derivatives of low molecular weight PEG, and the like.

[0028] In some preferred embodiments, the mass ratio of the phase reaction catalyst to solution A is 1:10-20.

[0029] In some preferred embodiments, the fraction of the ethanol-physiological saline solution is 30-100%.

[0030] In some preferred embodiments, the temperature in step S2-3 is 180°C.

[0031] The second aspect of the present invention provides an application of a PET imaging agent targeting ACE2 for the differential diagnosis of new coronavirus infection and tumors, which is used in PET imaging.

[0032] In some preferred embodiments, 18 The specific application process of F-MLN-4760 is as follows:

[0033] According to 37MBq / 10kg body weight 18 F-MLN-4760 was injected intravenously into mice. After the initial distribution and clearance in the body was completed for more than 15 minutes, the PET device was used for signal acquisition.

[0034] Organs and tissues with high ACE2 expression and organs that metabolize the imaging agent will show high uptake of the imaging agent. It can be used to monitor pathological changes of ACE2 in tissues and organs caused by COVID-19 infection, monitor pathological recovery of ACE2 after infection, diagnose diseases of the renin-angiotensin-aldosterone (RAAS) system, and screen tumor-related ACE2 targets and identify tumors.

[0035] Beneficial effects: The PET imaging agent prepared by the present invention has the following advantages over the prior art:

[0036] 1. Prepared by the present invention 18 The F-MLN-4760 method is simple, and compared with common substitution reactions, the reaction conditions are mild and the reaction time is shortened, and the closed reaction is convenient for radiation protection. The structure was verified by LC-MS, and two chlorines were simultaneously replaced by fluorines;

[0037] 2. The method prepared by the present invention 18 F-MLN-4760 can be used after intravenous injection to monitor pathological changes in ACE2 in tissues and organs caused by COVID-19 infection, monitor pathological recovery after infection, diagnose diseases of the renin-angiotensin-aldosterone (RAAS) system, and screen for tumor-related ACE2 targets and identify tumors. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 Prepared by Example 1 of the present application 18 LC-MS UV spectrum of F-MLN-4760;

[0039] Figure 2 Prepared by Example 1 of the present application 18 LC-MS mass spectrum of F-MLN-4760;

[0040] Figure 3 Prepared in Example 1 of this application 18 Transverse PET / CT image and PET MIP image of the mouse heart 30 minutes after tail vein injection of F-MLN-4760 (lower right), the arrow points to the heart position. Example

[0041] Example 1

[0042] A PET imaging agent targeting ACE2 for differential diagnosis of COVID-19 infection and tumors is prepared, comprising the following steps:

[0043] S1: 18 Purification of F:

[0044] 1) Enrichment of cyclotron produced by sep-pak QMA column 18 F ions;

[0045] 2) Elute the sep-pak QMA column with saline to recover 18 F, spare.

[0046] S2: Preparation of PET imaging agent:

[0047] S2-1: Dissolve 5 mg of MLN4760 in 2.5 mL of dimethyl sulfoxide to prepare a 2 mg / mL stock solution;

[0048] S2-2: Take the solution in step S1 again 18 F is heated for 1 h to azeotropize and remove water, and the mixture is mixed with the solution in step S2-1 to obtain solution A for standby use;

[0049] S2-3: Add phase reaction catalyst to solution A, seal in a high pressure reactor, and react at 180°C for 2h;

[0050] S2-4: After the reaction is completed, the mixture is cooled to room temperature, enriched with a C-18 column, and eluted with a 30% by volume ethanol-physiological saline solution to obtain a PET imaging agent.

[0051] The phase reaction catalyst is tetramethylammonium chloride, and the added amount is 20 mg.

[0052] The mass ratio of the phase reaction catalyst to solution A is 1:15;

[0053] Solution A contains 100 μL of DMSO-treated 18 F (370 MBq) and 10 μL of stock solution (containing 20 μg of MLN4760, the solution obtained in step S2-1).

[0054] The total reaction volume in step S2-3 was 500 μL, which was supplemented to the target volume with dimethyl sulfoxide.

[0055] 18 The specific application process of F-MLN-4760 is as follows:

[0056] According to 37MBq / 10kg body weight 18 F-MLN-4760 was injected intravenously into mice. After the initial distribution and clearance in the body was completed for more than 15 minutes, the PET device was used for signal acquisition.

[0057] Organs and tissues with high ACE2 expression and organs that metabolize the imaging agent will show high uptake of the imaging agent. It can be used to monitor pathological changes in ACE2 in tissues and organs caused by COVID-19 infection, monitor pathological recovery after infection, diagnose diseases of the renin-angiotensin-aldosterone (RAAS) system, and screen for tumor-related ACE2 targets and identify tumors.

[0058] Example 2

[0059] A PET imaging agent targeting ACE2 for the differential diagnosis of novel coronavirus infection and tumors is prepared. Its specific implementation method is the same as that in Example 1, except that the reaction catalyst is tetraphenylphosphonium bromide.

Claims

1. A method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of COVID-19 infection and tumors, characterized in that: The following steps are involved: S1: 18 Purification of F. S2: Preparation of PET imaging agent; S1 step 18 The specific operation of purifying F includes the following steps: 1) Enrichment of cyclotron-produced 18 F ions; 2) Elute the sep-pak QMA column with saline to recover 18 F, spare; The specific operation of preparing the PET imaging agent described in S2 comprises the following steps: S2-1: dissolving the ACE2 inhibitor MLN-4760 in the solution; S2-2: Take the solution again and compare it with the solution in step S1 18 F is heated to remove water, and mixed with the solution in step S2-1 to obtain solution A for standby use; S2-3: Add phase reaction catalyst to solution A, seal in a high pressure reactor, and react at 170-220°C for 2-4h; S2-4: After the reaction is completed, the reaction is cooled to room temperature, and a C-18 column is used for enrichment, and an ethanol-physiological saline solution is used for elution to obtain a PET imaging agent; The structural formula of ACE2 inhibitor MLN-4760 is shown in formula (1): Formula (1); The PET imaging agent is 18 F-MLN-4760, structural formula is shown in formula (2): Formula (2).

2. The method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors according to claim 1, characterized in that: The solutions described in S2-1 and S2-2 are both aprotic polar solutions.

3. The method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors according to claim 2, characterized in that: The aprotic polar solution is at least one of dimethyl sulfoxide, N,N-dimethylformamide, 1,3-dimethyl-2-imidazolidinone, cyclopentane sulfone, hexamethylphosphoric triamide and acetonitrile.

4. The method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors according to claim 3, characterized in that: The phase reaction catalyst is at least one of crown ethers, quaternary ammonium salts, quaternary phosphonium salts and metal halide salts.

5. The method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors according to claim 1 or 4, characterized in that: The mass ratio of the phase reaction catalyst to solution A is 1:10-20.

6. The method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors according to claim 1, characterized in that: The volume fraction of the ethanol-physiological saline solution is 30-100%.

7. The method for preparing a PET imaging agent targeting ACE2 for differential diagnosis of novel coronavirus infection and tumors according to claim 1, characterized in that: The temperature in step S2-3 is 180°C.

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