Application of levothyroxine sodium in the preparation of drugs for the prevention or treatment of liver injury

Abstract

This invention relates to novel applications of levothyroxine sodium, specifically its use in the preparation of drugs for the prevention or treatment of liver injury. This invention reveals that levothyroxine sodium improves liver morphology and liver weight ratio in mice with acetaminophen (APAP)-induced acute liver injury (ALI), reduces serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin (TbiL) levels, and increases serum glutathione (GSH) levels. This suggests a novel pharmaceutical use for levothyroxine sodium in the prevention, relief, and / or treatment of acute and chronic liver injury.

Description

Technical Field

[0001] This invention relates to the field of compound drugs, and more specifically to the use of levothyroxine sodium in the treatment of liver injury. Background Technology

[0002] Liver injury (LI), classified as acute or chronic, has become an increasingly serious public health problem, with its prevalence and importance growing daily. Clinically, various factors can lead to liver injury, including drugs, herbal compounds, chemicals, and toxins. Acetaminophen (APAP, commonly known as paracetamol) is the most common drug causing toxic liver injury. With the increasing incidence of drug-induced liver injury, the incidence of acute liver failure is also rising, accounting for approximately 80% of deaths and making it the third leading cause of death among liver disease patients in my country, after liver cancer and cirrhosis, seriously threatening human health. Therefore, the search for drugs to prevent and treat liver injury is of profound significance.

[0003] Levothyroxine sodium is indicated for long-term replacement therapy of congenital hypothyroidism (cretinism) and hypothyroidism of various causes in children and adults. It can also be used for simple goiter, chronic lymphocytic thyroiditis, and suppression (and replacement) therapy after thyroid cancer surgery. Sometimes it can be used as adjunctive therapy for hyperthyroidism and can also be used for suppression tests to diagnose hyperthyroidism.

[0004]

[0005] Levothyroxine sodium (MF:C) 15 H 11 l4NO4, CAS: 51-48-9)

[0006] Currently, there are no reports of levothyroxine sodium being used to treat liver injury. This patent is the first to demonstrate that levothyroxine sodium can be used to prevent, alleviate, and / or treat acute and chronic liver injury. Summary of the Invention

[0007] The technical problem to be solved by this invention is to provide a new pharmaceutical use of levothyroxine sodium in the prevention and treatment of liver damage.

[0008] Technical solution

[0009] The use of levothyroxine sodium in the preparation of drugs for the prevention or treatment of liver injury.

[0010] The liver injury described is either acute or chronic.

[0011] Specifically:

[0012] Levothyroxine sodium has a novel application in the prevention and treatment of acute liver injury by improving the liver weight ratio in mice with acetaminophen-induced (APAP) acute liver injury (ALI), reducing serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin (TbiL) levels, and increasing serum glutathione (GSH) levels.

[0013] Beneficial effects

[0014] 1. Levothyroxine sodium can be used to treat acute and chronic liver injury and related diseases, and has a hepatoprotective effect. In Examples 1-4 of this invention, after pre-administration of levothyroxine sodium for 7 days, an acute liver injury model in mice was induced by acetaminophen (APAP). By measuring indicators such as liver coefficient ratio, serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) levels, total bilirubin, and glutathione, it was confirmed that levothyroxine sodium has a significant effect in improving acetaminophen-induced acute liver injury, demonstrating that levothyroxine sodium can be used to prevent and treat acute and chronic liver injury, and has broad application prospects.

[0015] 2. Levothyroxine sodium falls under the category of "repurposing an existing drug," and its new drug application is classified as a Class 2 modified drug, 2.4, for a new indication containing a known active ingredient. This invention found that levothyroxine sodium at 100 μg / kg is superior to silymarin meglumine tablets (10 mg / kg). This invention also found that levothyroxine sodium may improve liver damage through other mechanisms rather than by increasing glutathione (GSH) levels, i.e., its mechanism of action differs from that used for anti-hepatic damage. Levothyroxine sodium is a marketed drug with proven safety and quality; therefore, this invention has significant market development and clinical application potential. Attached Figure Description

[0016] Figure 1 Effects of different doses of levothyroxine sodium on liver coefficient of liver injury (SM: silymarin meglumine tablets, the same below; n=8, expressed as mean±SD, #P<0.05 compared with the blank group, *P<0.05, **P<0.01 compared with the model group, $P<0.05 compared with the positive drug SM group);

[0017] Figure 2 Effects of different doses of levothyroxine sodium on transaminases in liver injury, where A is alanine aminotransferase (ALT) and B is aspartate aminotransferase (AST); (n=8, expressed as mean±SD, ##P<0.01 compared with the blank group, *P<0.05, **P<0.01 compared with the model group, $P<0.05 compared with the positive drug SM group);

[0018] Figure 3Effects of different doses of levothyroxine sodium on total bilirubin in liver injury (n=8, expressed as mean±SD, #P<0.05 compared with the control group, *P<0.05, **P<0.01 compared with the model group);

[0019] Figure 4 Effects of different doses of levothyroxine sodium on glutathione levels in liver injury (n=8, expressed as mean±SD, ##P<0.01 compared with the control group, **P<0.01 compared with the model group, $P<0.05 compared with the positive drug SM group). Detailed Implementation

[0020] The pharmacological effects of levothyroxine sodium in improving liver injury are further illustrated below with reference to embodiments, but this does not limit the scope of protection of this invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention. In this invention, experimental materials or experimental methods not described in detail or particularly emphasized are all conventional experimental materials or experimental methods in the art, and those skilled in the art can obtain such experimental materials or have the ability to conduct such experiments.

[0021] Example 1: Levothyroxine sodium improves the liver coefficient ratio in acetaminophen-induced acute liver injury. I. Experimental materials

[0022] 1. Instruments and reagents

[0023] Constant flow syringe pump LSP04-1A (Baoding Lange Constant Flow Pump Co., Ltd.), Infinite200Pro microplate reader (Tecan, Switzerland), electronic analytical balance (Mettler-Toledo, Shanghai), Milli-Q ultrapure water system (Millipore, USA), miniature vortex apparatus (Dalong Xingchuang Experimental Instruments Co., Ltd.), intelligent adjustable peristaltic pump (Kammer Fluid Technology (Shanghai) Co., Ltd.), tissue scissors and surgical forceps (Shanghai Surgical Instrument Factory).

[0024] Levothyroxine sodium was purchased from Merck Pharmaceuticals (Jiangsu) Co., Ltd., acetaminophen from Dalian Meilun Biotechnology Co., Ltd., silymarin meglumine lyophilized powder for injection from Nanjing Chenxiang Pharmaceutical Research Co., Ltd., silymarin meglumine tablets from Jiangsu Zhongxing Pharmaceutical Co., Ltd., sodium chloride injection from Anhui Shuanghe Pharmaceutical Co., Ltd., and reagent kits for alanine aminotransferase, aspartate aminotransferase, total bilirubin, and glutathione from Nanjing Jiancheng Bioengineering Institute.

[0025] 2. Laboratory animals

[0026] Seventy-two male C57BL / 6J mice were purchased from Jiangsu Huachuang Xinno Pharmaceutical Technology Co., Ltd. (Experimental Animal Production License No. SYXK(Su)2022-0013).

[0027] II. Experimental Methods

[0028] 1. Animal grouping, model establishment, and drug administration

[0029] Male C57BL / 6J mice were randomly divided into 6 groups (n=12 per group): a control group, a model group, low-, medium-, and high-dose levothyroxine (LEV) groups (10, 30, and 100 μg / kg), and a positive control group (silymarin meglumine tablets, 10 mg / kg). The treatment groups were pre-treated with LEV by gavage for one week, while the control and model groups were treated with saline by gavage for one week. On day 8, an acute liver injury model was induced in mice by intraperitoneal injection of 300 mg / kg acetaminophen (APAP) solution. Blood and liver tissue were collected 6 hours after APAP modeling for biochemical and molecular analysis.

[0030] 2. Detection of liver coefficient ratio

[0031] Six hours after modeling, blood was collected from the eyeballs of mice in each group and placed in 1.5 mL EP tubes. After standing for 30 min, the tubes were centrifuged at 3,500 rpm for 15 min at 4°C. The supernatant was collected, and the mice's abdomens were opened with surgical scissors to observe the morphology of the liver. The liver was then isolated. The mice's hearts were perfused with 1×PBS until the blood was completely washed away. A specimen of the left lobe of the liver was then fixed in 4% paraformaldehyde solution.

[0032] 3. Data Processing

[0033] All data were analyzed using GraphpadPrism 8 software and expressed as mean ± SD. Students' t test was used for comparisons between two groups. One-way ANOVA was used for comparisons of three or more groups. Dunnett's test was used for the test, and P < 0.05 was considered statistically significant.

[0034] III. Experimental Results

[0035] like Figure 1 As shown, compared with the blank group, the liver weight ratio in the model group increased after APAP stimulation ( # P<0.01 indicates that the liver is congested, edematous, or hyperplastic and hypertrophic, indicating that the liver is severely damaged after APAP stimulation. Pretreatment with different doses of levothyroxine sodium followed by APAP stimulation significantly improved the liver compared with the model group. Statistical results show that the improvement effect of the medium and high dose LEV group is better than that of SM, and compared with the SM group, the high dose LEV group has a significant difference in reducing the liver coefficient.

[0036] Example 2: The effect of levothyroxine sodium on improving serum transaminase levels in mice with acetaminophen-induced acute liver injury.

[0037] I. Experimental Materials

[0038] 1. The instruments and reagents are the same as in Example 1.

[0039] 2. The experimental animals are the same as in Example 1.

[0040] II. Experimental Methods

[0041] 1. Animal grouping, modeling, and drug administration are the same as in Example 1.

[0042] 2. Detection of alanine aminotransferase (ALT) and aspartate aminotransferase (AST)

[0043] Six hours after modeling, blood was collected from the eyeballs of mice in each group and placed in 1.5 mL EP tubes. After standing for 30 min, the tubes were centrifuged at 3500 rpm for 10 min, and the supernatant was collected as serum. The level of transaminase in the serum was detected according to the ALT and AST kits.

[0044] 3. Data processing is the same as in Example 1.

[0045] III. Experimental Results

[0046] The results are shown in Figure 2. Compared with the control group, the ALT and AST levels in the model group mice were significantly increased after APAP stimulation. ## P<0.01, # (P<0.05) indicates severe liver damage after APAP stimulation. Pretreatment with different doses of LEV followed by APAP stimulation significantly improved transaminase levels in the treatment group. Statistical results showed that, compared with the SM group, 100 μg / kg LEV significantly improved aspartate aminotransferase levels, further demonstrating that LEV was significantly more effective than SM in improving liver function. * P<0.05, ** P<0.01).

[0047] Example 3: The effect of levothyroxine sodium on improving serum total bilirubin in mice with acetaminophen-induced acute liver injury.

[0048] I. Experimental Materials

[0049] 1. The instruments and reagents are the same as in Example 1.

[0050] 2. The experimental animals are the same as in Example 1.

[0051] II. Experimental Methods

[0052] 1. Animal grouping, modeling, and drug administration are the same as in Example 1.

[0053] 2. Detection of total bilirubin (TbiL)

[0054] Blood was collected from the eyeballs of mice in each group 6 hours after modeling, placed in 1.5 mL EP tubes, allowed to stand for 30 min, centrifuged at 3500 rpm for 10 min, and the supernatant was collected as serum. The TbiL content in the serum was detected according to the kit.

[0055] 3. Data processing is the same as in Example 1.

[0056] III. Experimental Results

[0057] The results are as follows Figure 3 As shown, compared with the blank group, the TBiL level in the model group mice was significantly increased after APAP stimulation. # (P<0.05) indicates severe liver damage after APAP stimulation. Pretreatment with different doses of LEV followed by APAP stimulation resulted in a dose-dependent reduction in TBiL levels by LEV. Statistical results showed that the high-dose LEV group did not significantly differ from the positive control drug SM in reducing TBiL. * P<0.05, ** P<0.01).

[0058] Example 4: The effect of levothyroxine sodium on improving serum glutathione levels in mice with acetaminophen-induced acute liver injury.

[0059] I. Experimental Materials

[0060] 1. The instruments and reagents are the same as in Example 1.

[0061] 2. The experimental animals are the same as in Example 1.

[0062] II. Experimental Methods

[0063] 1. Animal grouping, modeling, and drug administration are the same as in Example 1.

[0064] 2. Detection of glutathione (GSH)

[0065] Blood was collected from the eyeballs of mice in each group 6 hours after modeling, placed in 1.5 mL EP tubes, allowed to stand for 30 min, centrifuged at 3500 rpm for 10 min, and the supernatant was collected as serum. The GSH content in the serum was detected according to the kit.

[0066] 3. Data processing is the same as in Example 1.

[0067] III. Experimental Results

[0068] As shown in Figure 4, compared with the control group, the GSH level in the model group mice was significantly reduced after APAP stimulation. ##(P<0.01) After pretreatment with different doses of LEV followed by APAP stimulation, GSH was improved to varying degrees. Statistical results showed that the effect of LEV in raising GSH was not as good as that of SM in any dose group, indicating that LEV may improve liver damage through other means rather than by increasing GSH levels.

Claims

1. Application of levothyroxine sodium in the preparation of drugs for the prevention or treatment of acetaminophen-induced acute liver injury.

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