Preparation method and application of five deuterium-substituted glucose

Abstract

The application belongs to the technical field of medicine production, and particularly relates to a preparation method and application of five deuterium-substituted glucose. The method comprises deuterium substitution of D-methyl glucoside, and demethylation of the deuterium-substituted D-methyl glucoside to obtain a target product, namely [2,3,4,6,6'-d5]-D-glucose. The preparation method of the five deuterium-substituted glucose is simple in process route, convenient to operate, mild in experimental conditions, and the synthesized five deuterium-substituted glucose is successfully applied to the description of glycolysis metabolism in a brain glioma model, which is helpful to the clinical conversion application of metabolic imaging technology.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and in particular to a method for preparing and applying deuterated glucose. Background Technology

[0002] Positron emission tomography (PET), currently the most advanced medical imaging technology, can achieve high-resolution imaging of cellular metabolism and function, enabling non-invasive, three-dimensional, and dynamic studies of human physiological and biochemical processes at the molecular level. PET is applied to tumors, including tumor diagnosis, differentiation between benign and malignant tumors, staging and classification of malignant tumors, early diagnosis and differentiation of recurrence and metastasis, selection of treatment plans and monitoring of chemotherapy efficacy, observation of tumor changes, and prognosis assessment. Unlike other examinations, PET examinations rely on positron emission tomography (PET drugs), which specifically concentrate in target organs to achieve diagnostic and evaluation purposes. Currently, the most commonly used positron emission tomography drug in PET examinations is... 18 F-fluorodeoxyglucose ( 18 F-FDG), 18 F-FDG, as a tumor metabolism drug, plays a significant role in the diagnosis and differential diagnosis of malignant tumors, but... 18 The fluorine used in F-FDG is fluorine-18, a positron-emitting radioactive isotope. While it provides high-resolution images of glucose uptake in tissues, it does not reflect information about glucose metabolism. Therefore, it often provides blurry results for tissues with inherently high glucose uptake, such as the brain. The radioactivity of FDG limits the possibility of repeated assessments of disease progression, and multiple tests on patients can easily cause secondary harm. Furthermore, although its half-life is significantly improved (109.8 min) compared to other imaging markers such as oxygen-15 (O-15), nitrogen-13 (N-13), and carbon-11 (C-11), it still cannot be used to track the entire metabolic process.

[0003] A new, non-invasive, and simple imaging technique has been developed for magnetic resonance imaging (MRI) that can capture information about cellular metabolism and create three-dimensional images. This helps radiologists make more accurate diagnoses of many diseases and track the progress of treatment, such as evaluating the effectiveness of treatments for tumors and other diseases. This technique, called "deuterium metabolism imaging" or DMI for short, uses MRI to scan the brain and other organs as they metabolize energy-producing substances, such as deuterium-labeled glucose, creating three-dimensional metabolic images using the spectra generated by the magnetic resonance of deuterium atoms.

[0004] To verify the effectiveness of this technology, researchers applied DMI to the brains and livers of rats and human subjects who ingested deuterium-labeled glucose (deuterium is a non-radioactive or stable hydrogen isotope). Researchers observed significant differences in glucose metabolism between glioma cells and normal brain cells in some mice during anti-cancer treatment, with high-contrast tomographic images showing a unique "Warburg effect." Similar metabolic patterns were obtained in brain scans of two glioma patients after oral administration of deuterium-labeled glucose (see [link to relevant documentation]). Figure 2 DMI was applied to the livers of mice and humans, and scans showed that glucose was converted into deuterated glycogen after oral administration of deuterium-labeled glucose.

[0005] Deuterated electron microscopy (DMI) can reveal the metabolic responsiveness of drug treatments before drugs affect tumor size, providing a new tool for drug evaluation. Theoretically, DMI technology is very powerful and can be applied to any disease involving metabolism, such as multiple sclerosis and other neurodegenerative diseases, and is not limited to the brain; it can also image other organs. DMI can also scan other deuterium-labeled substances to create three-dimensional images. Therefore, existing positron emission tomography (PET) techniques still need improvement and development. Summary of the Invention

[0006] In view of the shortcomings of the prior art, the purpose of this invention is to provide a method for preparing deuterated glucose that is simple to operate, has mild conditions, and is inexpensive, as well as its application, aiming to solve the problems of the prior art. 18 F-fluorodeoxyglucose, when used as a contrast agent, can easily cause secondary harm to patients with repeated testing, and it cannot be used to track the entire metabolic process. The deuterated glucose described herein has the following structure:

[0007] ,

[0008] Includes the following steps:

[0009] Step 1, Synthesis of [2,3,4,6,6'-d5]-D-methylglucoside: D-methylglucoside was added to heavy water, a catalyst was added, and the reaction was carried out at a suitable temperature under a hydrogen atmosphere. After the reaction was completed, [2,3,4,6,6'-d5]-D-methylglucoside was obtained.

[0010] Step 2, Synthesis of [2,3,4,6,6'-d5]-D-glucose: The [2,3,4,6,6'-d5]-D-methylglucosinolate prepared in the previous step was added to a suitable solvent and a suitable demethylating agent. The reaction was carried out at a suitable temperature. After the reaction was completed, the crude product was obtained and then recrystallized to obtain the target compound, namely [2,3,4,6,6'-d5]-D-glucose.

[0011] This invention has the following significant advantages: (1) It discloses for the first time a method for synthesizing [2,3,4,6,6'-d5]-D-glucose; (2) Compared with [6,6'-d2]-D-glucose, it has a simpler synthetic route, relatively milder reaction conditions, simpler operation, higher yield, and more efficient deuterium labeling; (3) Animal experiments have confirmed that this substrate, through... 2 H-MRS has been successfully applied to describe glycolytic metabolism in a glioma model under ultra-high field conditions. Attached Figure Description

[0012] Figure 1 A synthetic route diagram for [2,3,4,6,6'-d5]-D-glucose;

[0013] Figure 2 Left MRI shows a tumor in the right brain; right DMI shows a tumor metabolic map.

[0014] Figure 3 The NMR (H1N) spectrum of [2,3,4,6,6'-d5]-α-D-methylglucoside is shown.

[0015] Figure 4 The NMR (carbon) spectrum of [2,3,4,6,6'-d5]-α-D-methylglucoside is shown.

[0016] Figure 5 The NMR (H1N) spectrum of [2,3,4,6,6'-d5]-D-glucose;

[0017] Figure 6 The NMR (carbon spectrum) of [2,3,4,6,6'-d5]-D-glucose;

[0018] Figure 7 High-resolution 1H anatomical T2w images (CD) of (AB) glioma rats and control rats at different time points after infusion of [2,3,4,6,6'-d5]-D-glucose in glioma mice and control groups. 2 H spectrum;

[0019] Figure 8The results of infusions of [2,3,4,6,6'-d5]-D-glucose and [6,6'-d2]-D-glucose in glioma rats at (A) 0 min, (B) 60 min, and (C) 150 min after infusion. 2 H spectrum;

[0020] Figure 9 The signal intensities of glucose (A) and water (B) at 50 points in rats with gliomas after infusion of [2,3,4,6,6'-d5]-D-glucose (solid lines represent the fitted curves of equation (1-2), and the signal intensities are normalized to the water signal measured before infusion).

[0021] Figure 10 Normalized signal intensity of glutamate / glutamine (A), lactate (B), and lactate / Glx ratio (C) at different time points after infusion in four groups (error bars represent the standard deviation within the group);

[0022] Figure 11 To estimate the kinetic model parameters of glucose and water for each group based on formula (1-2) [(A) normalized G0 and Ws; (B) time constants for glucose consumption and water production]. Detailed Implementation

[0023] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0024] Example 1: A method for synthesizing [2,3,4,6,6'-d5]-D-glucose, comprising the following steps:

[0025] S1. Heavy water (400 mL) and 10% Ru / C (5.05 g, 5 mol%) were added to α-D-methyl glucoside. The system was purged with hydrogen and a hydrogen atmosphere was maintained. The mixture was heated to 80 °C and reacted for 24 h. After filtration, the filtrate was concentrated to obtain 19.5 g of [2,3,4,6,6'-d5]-α-D-methyl glucoside, with a yield of 98%. The detection data are as follows: [α] +50.51 (c 10, H2O); 1 H NMR (400 MHz, D2O) δ 4.829 (s, 1H), 3.657 (s, 1H), 3.444 (s, 3H); 13C NMR (100 MHz,D2O) δ 99.19, 71.33, 54.96; HR-ESIMS m / z: calculated for C7H9D5O6 - [MH] - :198.1104, found 198.0461.

[0026] S2. [2,3,4,6,6'-d5]-α-D-methylglucoside (19.5 g, 98 mmol) was dissolved in sulfuric acid (1 M, 200 mL). The system was purged with nitrogen and a nitrogen atmosphere was maintained. The mixture was heated to 100 °C and held for 11 h. The reaction solution was then cooled to room temperature and exchanged with D301 resin. The eluent was concentrated to obtain the crude product, which was recrystallized to give 13.6 g of [2,3,4,6,6'-d5]-α-D-glucose, with a yield of 75%. The detection data are as follows: [α] +134.42 (c10, H2O); 1 H NMR (400 MHz, D2O) δ5.243 (s, 1H), 4.657 (s, 2H), 3.838 (s, 1H), 3.469 (s, 2H); 13 C NMR (100 MHz,D2O) δ 95.82, 92.00, 72.69, 71.18; HR-ESIMS m / z: calculated for C6H7D5O6 - [MH] - : 184.0948, found 184.0896.

[0027] Figure 7 The images show glioma rats and normal rats at different time points. 2 H-spectrum. In both groups of rats, glucose was rapidly consumed after infusion of [2,3,4,6,6'-d5]-D-glucose. 2 H-labeled water gradually accumulated. In glioma rats, a significant lactate peak was observed 30 minutes after injection. In contrast, a weaker Glx peak with a lactate signal was clearly identifiable in the control group.

[0028] In addition to obvious water and glucose peaks, glioma rats also showed lactate peaks (~3.4 ppm to water peak), while control rats showed Glx peaks (~2.5 ppm to water peak).

[0029] Figure 8 Comparison [ 2 H5]-Tumor and [ 2 H2]-Tumor rats at three time points2 H spectrum. Due to 2 H is highly enriched. 2 The total signal-to-noise ratio (SNR) of the H5-glucose spectrum is higher than that of [ 2 [H2]-glucose profile. Due to [ 2 The additional labeling of H5-glucose at the C2-C4 positions is more pronounced during infusion, leading to increased water production. Lactate signal levels increase with […]. 2 H5]-glucose levels rise with infusion, especially at the end of collection.

[0030] All 50 sampling sites of glioma rats after infusion of [2,3,4,6,6'-d5]-D-glucose 2 The signal intensity of H-labeled glucose and water is as follows: Figure 9 As shown. The results indicate that the signal changes of glucose and water fit well with equation (1-2). Normalized G0 and W s The values ​​were 8.99 and 2.14, respectively. The time constants for glucose consumption and water production were 51.15 min and 26.35 min, respectively.

[0031] Figure 10 Quantitative parameters of four metabolites were compared. The Warburg effect could be observed by increasing the Lac / Glx ratio in the tumor group through infusion of [2,3,4,6,6'-d5]-D-glucose or [6,6'-d2]-D-glucose. 2 H5]-Tumor group and [ 2 The average Lac / Glx ratios for the H5-Control group at 60–90 min were 1.336 ± 0.149 and 0.584 ± 0.115, respectively. p <0.001), while [ 2 H2]-Tumor group and [ 2 The Lac / Glx ratios in the H2-Control group were 0.839±0.251 and 0.479±0.173, respectively (p=0.017). 60 minutes after infusion... 2 The Lac / Glx ratio in the H5-Tumor group was significantly higher than that in the [H5]-Tumor group. 2 H2]-Tumor group. 2 H5]-Tumor group and [ 2 The total Lac / Glx ratios for the H2-Tumor group were 1.489±0.237 and 0.893±0.262, respectively (p=0.002). Although throughout all time periods, [ 2 The signal strengths of Glx and Lac in the H5-Control group were both higher than those in […]. 2 H2]-Control group, but Lac / Glx ratio [ 2H5]-Control group and [ 2 There was no significant difference between the H2-Control groups (0.625±0.083 vs. 0.513±0.132). p =0.14).

[0032] Figure 11 Compare the dynamic model parameters of the four groups. 2 H2]-Control and [ 2 The normalized G0 values ​​for the H2-Tumor group were 5.77±0.52 and 6.49±0.57, respectively. 2 H2]-Control and [ 2 The normalized G0 values ​​for the H2-Tumor group were 2.39 ± 0.60 and 3.18 ± 0.42, respectively. 2 H5]-Control and [ 2 The ratio of H2]-Control was 2.41. Normalized Ws was consistently higher in the control group. 2 H5]-Control and [ 2 The H5-Tumor values ​​were 3.21±0.38 and 2.13±0.17, respectively. 2 H2]-Control and [ 2 The H2-Tumor values ​​were 1.63±0.29 and 1.08±0.14, respectively. Both groups of glioma rats had relatively large glucose consumption time constants, but [ 2 The difference in the H5 group was not significant, indicating that the glucose uptake level of glioma rats was high.

[0033] This invention leverages the universality and ease of implementation of 1H proton magnetic resonance spectroscopy (1H MR spectroscopy), along with its excellent resolution, to track metabolites transferred from deuterated glucose. It offers higher detection resolution and sensitivity, enabling the detection of dynamic exchanges of individual metabolites. By measuring changes in the 1H MR spectroscopy spectrum, metabolites undetectable by 2H MR spectroscopy can be detected, thus revealing the rate of metabolic circulation in the body. A single acquisition can provide steady-state information and metabolic rates for several metabolites. Furthermore, the deuterated glucose used in this invention is ingestible, and multiple tests will not cause harm to the human body. 1H MR spectroscopy can also be performed using a standard nuclear magnetic resonance (NMR) spectrometer, eliminating the need for specialized equipment and reducing costs. By using standard 1H MR spectroscopy acquisition hardware and signal processing, the conversion of deuterated labeling can be directly monitored. The method is simple, practical, highly accurate, and reliable, allowing for quantitative and localized analysis of metabolic status.

[0034] It should be understood that the application of the present invention is not limited to the examples above. Those skilled in the art can make improvements or modifications based on the above description, and all such improvements and modifications should fall within the protection scope of the appended claims.

Claims

1. The application of a stable pentadeuterated glucose, namely [2,3,4,6,6'-d5]-D-glucose, characterized in that, It was used as a contrast agent for deuterium metabolism imaging. The chemical structural formula of the [2,3,4,6,6'-d5]-D-glucose is: 。 2. The application according to claim 1, characterized in that, The contrast agent also includes at least one pharmaceutically acceptable excipient.

3. The application according to claim 2, characterized in that, The excipient is one of a carrier, a filler, or a solvent.

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