CN105524113A - 99mTcN nuclear-labeled triazole-ring-containing glucosine complex and its preparation method and application - Google Patents

Abstract

The invention discloses a ^99m TcN (TAGDTC) ₂ complex and its preparation method and application. The complex has a ^99m TcN core as the central core, and the organic ligand coordinated therewith is glucosine containing a triazole ring The ligand is obtained through the synthesis of the ligand TAGDTC and the preparation of the ^99m TcN (TAGDTC) ₂ complex. The complex has high radiochemical purity, good stability, high uptake and good retention in the tumor site of tumor-bearing mice, high tumor/muscle ratio, good SPECT tumor imaging effect, and can be used as a new type of tumor Imaging agent promotion and application.

Description

99mTcN nuclear-labeled triazole-ring-containing glucosine complex and its preparation method and application

Technical field

The invention relates to the technical fields of ^99m Tc-labeled radiopharmaceutical chemistry and clinical nuclear medicine, in particular to a ^99m TcN nuclear-labeled triazole ring-containing glucosine complex, a preparation method and application.

Background technique

At present, cancer has become the first cause of death, and the morbidity and mortality are still rising, posing a huge threat to public health. In order to reduce the mortality rate of malignant tumors and improve the cure rate, it is of great significance to carry out early diagnosis. Radionuclide tumor imaging can non-invasively detect the molecular biological behavior and pathophysiological changes of tumors, and can accurately detect the location of lesions with high specificity. Today, radionuclide tumor imaging technology has become a major category of cancer diagnosis. Methods, especially with the use of SPECT-CT and PET-CT, radionuclide tumor imaging has become one of the advantages of nuclear medicine diagnosis.

Among tumor imaging drugs, ¹⁸ F-fluorodeoxyglucose ( ¹⁸ F-FDG) is currently the most widely used tumor imaging agent in clinical practice, and its clinical value has been recognized. In the human body, ¹⁸ F-FDG can enter tumor cells through glucose transporters, and then be converted into 6-phosphate- ¹⁸ F-FDG by hexokinase, and 6-phosphate- ¹⁸ F-FDG cannot be further metabolized into tumor cells, while 6 -Phospho- ¹⁸ F-FDG is negatively charged and cannot freely pass through the cell membrane, and finally stays in tumor cells. However, ¹⁸ F-FDG is a positron imaging agent, and the positron nuclide [ ¹⁸ F] needs to be produced by an accelerator, which is expensive, which limits its popularization and application in clinical diagnosis to some extent. However, ^99m Tc has excellent nuclear properties, and the successful development of ^{99Mo- 99m Tc} generator makes ^99m Tc radiopharmaceuticals easy to prepare, can be packaged, cheap and easy to obtain, and reliable in quality. Therefore, ^99m Tc-labeled glucose tumors are significantly Imaging agents have become a research hotspot in radiopharmaceuticals.

So far, most of the ^99m Tc-labeled glucose derivatives are still in the stage of laboratory research, and only ^99m Tc-ethylenedicysteine-deoxyglucose (referred to as ^99m Tc-ECDG) has entered the phase II clinical research stage, but its tumor uptake value and tumor/blood Therefore, it is of great practical significance to further develop new ^99m Tc-labeled glucose tumor imaging agents with excellent performance.

Click chemistry is a newly developed modular synthesis method, because of its convenient and easy-to-obtain raw materials, simple and easy reaction, mild reaction conditions, insensitive to oxygen and water, good product selectivity, high yield, easy purification of products, The advantages of simple post-processing have attracted great attention from the majority of chemical researchers. In particular, the Cu(I)-catalyzed 1,3-dipolar cycloaddition reaction of azide and alkyne produces a triazole ring with high stability, good water solubility and certain biological activity. The bond between nuclides and biomolecules reduces the lipid solubility of complexes, making Click chemistry widely used in radiopharmaceutical synthesis in recent years.

Due to the good chemical stability of the ^99m TcN triple bond and the biodistribution properties of the corresponding complexes are quite different from [ ^99m TcO] ³⁺ core or [ ^99m TcO ₂ ] ⁺ core complexes, the ^99m TcN core complexes The research has drawn a lot of attention. Especially the method of successfully preparing [ ^99m TcN] ²⁺ intermediate at room temperature using SDH (succinyl hydrazide, H ₂ NNHCOCH ₂ CH ₂ CONHNH ₂ ) as N ^3- ion donor and SnCl ₂ as reducing agent The breakthrough has laid a good foundation for the research and development of new [ ^99m TcN ] ²⁺ radiopharmaceuticals.

在已成功制备^99m TcN(DGDTC) ₂配合物的基础上(JunboZhang,JialeiRen,XiaoLin,XuebinWang.Synthesisandbiologicalevaluationofanovel ^99m Tcnitridoradiopharmaceuticalwithdeoxyglucosedithiocarbamate,showingtumoruptake[J],Bioorganic&MedicinalChemistryLetters,2009,19:2752–2754)，为了改进其肿瘤摄取值In order to solve the problem of low tumor/muscle ratio, the present invention utilizes Click chemistry to skillfully convert glucose into triazole ring-containing glucosine ligand (abbreviated as: TAGDTC), and then use the The sulfur atom coordinates with the ^99m TcN nucleus to form a stable ^99m TcN nucleus glucosamine complex to explore new tumor molecular probes, which has important scientific significance and application development value.

Contents of the invention

The purpose of the present invention is to provide a ^99m TcN nuclear-labeled triazole ring-containing glucosine complex with high radiochemical purity and good stability, which is applied in the field of tumor imaging, and also provides a preparation method thereof.

In order to achieve the above object, the present invention adopts the following technical scheme: a ^99m TcN nuclear-labeled triazole ring-containing glucosine complex ( ^99m TcN(TAGDTC) ₂ ), the structural formula of which is:

In this structural formula: with the [ ^99m TcN] ²⁺ nucleus as the central core, the 4 sulfur atoms in the two TAGDTC ligand molecules coordinate with ^99m Tc to obtain a ^99m TcN(TAGDTC) ₂ complex.

The preparation method of ^99m TcN (TAGDTC) ₂ complex is as follows:

a: Synthesis of ligand TAGDTC:

Its synthetic route is:

Step 1: Put an appropriate amount of GPN3 and tert-butyl-propargyl amide in a 50mL two-necked flask, add tetrahydrofuran to dissolve, then add a small amount of aqueous solution dissolved in copper sulfate pentahydrate and vitamin C, stir at room temperature, and react under nitrogen overnight. After the reaction was over, a light yellow solid was obtained by distillation under reduced pressure, which was dissolved by adding a small amount of water, extracted three times with ethyl acetate, and the organic phases were combined and dried with anhydrous MgSO ₄ . The solvent was distilled off under reduced pressure, and purified on a silica gel column (petroleum ether-ethyl acetate) to obtain a light yellow solid.

The second step: Dissolve an appropriate amount of the product of the first step in methanol, add 1mol/L HCl to adjust the pH to 2, and heat the reaction at 60°C for 4h. After the reaction was completed, a saturated NaHCO ₃ solution was added to adjust to neutrality, extracted with dichloromethane, the organic phase was separated, dried, filtered, and the solvent was removed. The residue was dissolved in 50 mL of methanol, an appropriate amount of KOH was added, and stirred at room temperature for 3 h. After the reaction, the pH was adjusted to neutral with 1mol/L HCl, the solvent was distilled off under reduced pressure, ethanol was added, filtered, the filtrate was collected, and the solvent was removed to obtain a crude product. The crude product was dissolved in an appropriate amount of water, a certain amount of KOH was added, CS ₂ was added dropwise in an ice-water bath, and the reaction was continued for 1 h in an ice-water bath. After the reaction, the solvent was distilled off under reduced pressure, and recrystallized from ethanol to obtain TAGDTC as a light yellow solid.

b: Preparation of ^99m TcN(TAGDTC) ₂ complex:

The preparation of ^99m TcN(TAGDTC) ₂ adopts ligand exchange reaction, and the reaction scheme is as follows:

^99m TcO ₄ ^- +SDH+SnCl ₂ ·2H ₂ O+PDTA→[ ^99m TcN] _int ²⁺

[ ^99m TcN] _int ²⁺ +TAGDTC→ ^99m TcN(TAGDTC) ₂

The specific steps are: adding an appropriate amount of fresh ^99m TcO ₄ ^-eluent to the SDH freeze-dried kit containing succinyl dihydrazide (SDH), 1,2-propanediaminetetraacetic acid, and SnCl ₂ 2H ₂ O, After shaking well, react at room temperature for 15 minutes to obtain [ ^99m TcN] ²⁺ intermediate. Then 1 mL of TAGDTC aqueous solution with a concentration of 1 g/L was added to the [ ^99m TcN ] ²⁺ intermediate prepared above, mixed well and reacted in a boiling water bath for 20 minutes to obtain the ^99m TcN(TAGDTC) ₂ complex.

The chemical reagents used above are all commercially available products with a wide range of sources and are easy to obtain; the SDH freeze-dried kit is provided by Beijing Shihong Drug Research and Development Center.

The radiochemical purity of the ^99m TcN(TAGDTC) ₂ complex prepared by the above-mentioned method is greater than 90%, and it has good stability in vitro, high uptake and good retention in the tumor of tumor-bearing mice, high tumor/muscle ratio, SPECT Imaging shows that there is obvious concentration in the tumor site, and it can become a new type of tumor imaging agent.

The biodistribution data of ^99m TcN (TAGDTC) ₂ complex and ^99m TcN (DGDTC) were compared in S180 sarcoma-bearing mice 2 hours after injection, and the results are shown in Table 1.

Table 1 Biodistribution data of ^99m TcN(TAGDTC) ₂ and ^99m TcN(DGDTC) ₂ in S180 sarcoma-bearing mice 2 hours after injection (x±s,%ID/g)

The above results show that the tumor/blood ratio of ^99m TcN(TAGDTC) ₂ and ^99m TcN(DGDTC) ₂ complexes is not much different, but the tumor uptake and tumor/muscle ratio of the former is significantly higher than that of the latter, ^99m TcN(TAGDTC) The pro-tumor performance of ₂ is obviously better than that of ^99m TcN(DGDTC) ₂ , and it has broad application prospects as a new glucose tumor imaging agent.

Experiments show that the properties of the ^99m TcN(TAGDTC) ₂ complex are as follows:

1. Chromatographic identification of ^99m TcN(TAGDTC) ₂ complex

Thin-layer chromatography (TLC) identification: the two systems adopted are respectively, polyamide film as a support, acetonitrile as a developing agent; polyamide film as a support, physiological saline as a developing agent. The chromatography results of the determination are shown in Table 2.

R _f value of each component in table 2

The radiochemical purity of the marker was greater than 90% as determined by the above chromatographic identification.

2. Determination of lipid-water partition coefficient of ^99m TcN(TAGDTC) ₂ complex

Take 0.98mL pH7.4 phosphate buffer (0.025mol/L) in a 10mL centrifuge tube, add 1.0mL n-octanol and 0.02mL ^99m TcN (TAGDTC) ₂ complex solution to the centrifuge tube, cover the stopper, shake well Evenly, centrifuge for 5min (5000rpm), then take 0.1mL from the organic phase and the aqueous phase respectively, measure the radioactive counts of the two phases and calculate the logP value (P=activity of the organic phase/activity of the aqueous phase). The lipid-water partition coefficient of ^99m TcN(TAGDTC) ₂ logP=-0.97±0.01, indicating that it is a hydrophilic substance.

3. Stability determination of ^99m TcN(TAGDTC) ₂ complex

The radiochemical purity of the labeled ^99m TcN (TAGDTC) ₂ complex was measured at room temperature and in mouse serum at 37°C for different times (1, 2, 3, 4, 5, 6 hours). The experimental results It shows that the radiochemical purity of the complex is greater than 90% at room temperature and in mouse serum at 37°C for 6 hours, indicating that it has good stability in vitro.

4. Biodistribution experiment of ^99m TcN(TAGDTC) ₂ complex in S180 sarcoma-bearing mouse model

Inject 0.10 mL of complex solution (about 7.4×10 ⁵ Bq ) into the tail vein of Kunming mice (female, about 18-20 g) bearing the S-180 sarcoma model, and decapitate them at 0.5 h, 2 h, and 4 h after injection execute. The heart, liver, lung, kidney, spleen, intestine, muscle, bone, blood, tumor and other related tissues and organs were collected, cleaned and weighed, and their radioactive counts were measured with a technetium analyzer. The number of mice was 5. The percent injected dose per gram (%ID/g) of each tissue was calculated, and the results are shown in Table 3.

Table 3 Biodistribution of ^99m TcN(TAGDTC) ₂ complex in S180 sarcoma-bearing mouse model (x±s,%ID/g)

5. SPECT imaging experiment of ^99m TcN(TAGDTC) ₂ complex in mice bearing S-180 sarcoma

Inject 0.1 mL of labeled ^99m TcN (TAGDTC) ₂ (about 1.85×10 ⁷ Bq) into Kunming mice (female, weighing about 18-20 g) bearing S-180 sarcoma model from the tail vein, and perform SPECT to show picture. The results of SPECT imaging showed that ^99m TcN (TAGDTC) ₂ complexes were significantly concentrated in the tumor site, and the ratio of tumor to non-target in the region of interest (ROI) reached 8.92, which further confirmed that ^99m TcN (TAGDTC) ₂ complexes can be used as a A novel tumor imaging agent with excellent performance.

detailed description:

Describe the present invention in detail below by embodiment:

A ^99m TcN nuclear-labeled triazole ring-containing glucosine complex ( ^99m TcN(TAGDTC) ₂ ), the structural formula of which is:

In this structural formula: with the [ ^99m TcN] ²⁺ nucleus as the central core, the 4 sulfur atoms in the two TAGDTC ligand molecules coordinate with ^99m Tc to obtain a ^99m TcN(TAGDTC) ₂ complex.

The preparation method of ^99m TcN (TAGDTC) ₂ complex is as follows:

a. Synthesis of ligand TAGDTC:

Step 1: Put GPN3 (5mmol, 2.16g) and tert-butyl-propargyl amide (5mmol, 0.78g) in a 50mL two-necked flask, add 50mL tetrahydrofuran to dissolve, then add 90mg copper sulfate pentahydrate and 180mg A small amount of aqueous solution of vitamin C was stirred at room temperature and reacted overnight under nitrogen. After the reaction was over, distilled under reduced pressure to obtain a pale yellow solid, which was dissolved in 20 mL of water, extracted with ethyl acetate (3×10 mL), and the organic phases were combined and dried with anhydrous MgSO ₄ . The solvent was distilled off under reduced pressure, and purified on a silica gel column (petroleum ether-ethyl acetate) to obtain a light yellow solid. ¹ H-NMR (400MHz, CDCl ₃ ): δ (ppm) 7.57 (s, 1H), 5.19-5.24 (t, 1H), 5.06-5.11 (t, 1H), 4.98-5.03 (m, 1H), 4.43 -4.46(t,2H),4.38-4.39(d,4H),4.09-4.12(t,2H),3.84-3.88(m,1H),3.71-3.74(m,1H),3.47-3.53(m, 1H),2.22-2.29(m,2H),2.01-2.08(m,12H),1.44(s,9H). ¹³ CNMR(400MHz,CDCl ₃ ):δ(ppm)170.34,170.15,169.70,169.01,168.96 ,155.43,145.09,121.83,100.19,79.12,72.18,71.32,70.71,67.77,65.38,61.26,60.38,46.64,35.48,28.83,27.86,20.33,20.24,20.08.MS( ^ESI ):

Step 2: Dissolve an appropriate amount of the product from the first step in 50 mL of methanol, add 1 mol/L HCl to adjust the pH to 2, and heat at 60° C. for 4 h. After the reaction was completed, a saturated NaHCO ₃ solution was added to adjust to neutrality, extracted with dichloromethane, the organic phase was separated, dried, filtered, and the solvent was removed. The residue was dissolved in 50 mL of methanol, KOH (8 mmol, 0.45 g) was added, and stirred at room temperature for 3 h. After the reaction, the pH was adjusted to neutral with 1mol/L HCl, the solvent was distilled off under reduced pressure, ethanol was added, filtered, the filtrate was collected, and the solvent was removed to obtain a crude product. The crude product was dissolved in 30 mL of water, 113 mg of KOH was added, 0.2 mL of CS ₂ was added dropwise in an ice-water bath, and the reaction was continued in the ice-water bath for 1 h. After the reaction, the solvent was distilled off under reduced pressure, and recrystallized from ethanol to obtain TAGDTC as a light yellow solid. IR(KBr)/cm ^-1 : 3449(νOH), 1036(νC=S). ¹ H-NMR (400MHz, CDCl ₃ ): δ(ppm) 7.84-7.91(m, 1H), 4.34-4.47(m ,4H),3.81-3.91(d,2H),3.63-3.67(m,2H),3.19-3.52(m,8H). ¹³ C-NMR(400MHz,CDCl ₃ ):δ(ppm)215.143,184.044, 147.030, 126.623, 62.973, 60.808, 49.850, 45.122, 34.398, 26.363, 25.963. MS (ESI): 393.1 (MK ^- ).

b. Preparation of ^99m TcN(TAGDTC) ₂ complex

Add 0.5-1 mL ^of 37-370 MBq fresh ^99m TcO ₄ -eluent into the SDH freeze-dried kit, shake well, and react at room temperature for 15 minutes to obtain [ ^99m TcN] ²⁺ intermediate. Then 1 mL of TAGDTC aqueous solution with a concentration of 1 g/L was added to the [ ^99m TcN ] ²⁺ intermediate prepared above, mixed well and reacted in a boiling water bath for 20 minutes to obtain the ^99m TcN(TAGDTC) ₂ complex.

Claims (3)

1. a ^kind99mTcN (TAGDTC) ₂ complexes, its structural formula is as follows: In this structural formula: with the [ ^99m TcN] ²⁺ nucleus as the central core, the 4 sulfur atoms in the two TAGDTC ligand molecules coordinate with ^99m Tc to obtain a ^99m TcN(TAGDTC) ₂ complex. 2. as claimed in claim 1 ^99mTcN (TAGDTC) ₂ preparation methods of complexes, its processing steps are as follows: a: Synthesis of ligand TAGDTC: Step 1: Put an appropriate amount of GPN3 and tert-butyl-propargyl amide in a 50mL two-necked flask, add tetrahydrofuran to dissolve, then add a small amount of aqueous solution dissolved in copper sulfate pentahydrate and vitamin C, stir at room temperature, and react under nitrogen Overnight, after the reaction is over, distill under reduced pressure to obtain a light yellow solid, add a small amount of water to dissolve, extract 3 times with ethyl acetate, combine the organic phases and use anhydrous _MgSO4Dry , remove the solvent by distillation under reduced pressure, purify on a silica gel column to obtain a light yellow solid ; The second step: Dissolve an appropriate amount of the first step product in methanol, add 1mol/L HCl to adjust the pH to 2, heat the reaction at 60°C for 4h; after the reaction is completed, add saturated NaHCO ₃ solution to adjust to neutrality, and extract with dichloromethane , separate the organic phase, dry and filter to remove the solvent; dissolve the residue in 50 mL of methanol, add an appropriate amount of KOH, and stir at room temperature for 3 h; after the reaction, adjust the pH to neutral with 1mol/L HCl, remove the solvent by distillation under reduced pressure, and add ethanol , filter, collect the filtrate, and remove the solvent to obtain a crude product; dissolve the crude product in an appropriate amount of water, add a certain amount of KOH, add CS ₂ dropwise in an ice-water bath, and continue to react in an ice-water bath for 1 hour. After the reaction, reduce pressure The solvent was distilled off, and recrystallized from ethanol to obtain light yellow solid TAGDTC; b: Preparation of ^99m TcN(TAGDTC) ₂ complex: Add an appropriate amount of fresh ^99m TcO ₄ ^-eluent to the SDH freeze-dried kit containing succinyl dihydrazide, 1,2-propanediaminetetraacetic acid, SnCl ₂ 2H ₂ O, shake well, and React for 15 minutes to obtain the [ ^99m TcN] ²⁺ intermediate, then add 1 mL of TAGDTC aqueous solution with a concentration of 1g/L to the [ ^99m TcN] ²⁺ intermediate prepared above, mix well and react in a boiling water bath for 20 minutes to obtain The ^99m TcN (TAGDTC) ₂ complex. 3. The use of the ^99m TcN (TAGDTC) ₂ complex as claimed in claim 1 in the preparation of tumor imaging agents in the field of nuclear medicine imaging.